C H A P T E R

35

Tumor necrosis factor

Haichao Wang 1,2, Christopher]. Czura I a n d Keuin ]. Tracey 1
1North Shore-LIJ Research Institute, Manhasset, New York, USA
2North Shore University Hospital, Manhasset, New York, USA

Science is w o n d e r f u l l y e q u i p p e d to a n s w e r the q u e s t i o n 'How?'

b u t it gets terribly c o n f u s e d w h e n you ask the q u e s t i o n 'Why?'
Erwin Chargaff

INTRODUCTION STRUCTURE

Tumor necrosis factor (TNE TNF-a, cachectin) is a
pleiotropic pro-inflammatory cytokine that exerts
multiple biologic effects. TNF was originally identified
as an anti-tumor agent that induced necrotic cell
death in sarcomas and other tumor types. Localized,
low-level expression of TNF participates in beneficial
tissue remodeling and host defense responses. The
expression of TNF is tightly controlled, because sys-
temic overproduction of TNF activates inflammatory
responses to infection and injury, and mediates
hypotension, diffuse coagulation, and widespread
tissue damage. The diverse activities of TNF led to the
simultaneous and apparently paradoxical pursuit of
TNF as an anti-tumor strategy, and TNF inhibitors to
attenuate lethal systemic inflammation. This chapter
has been written as an introduction to the broad topic
of TNE its molecular biology, biochemistry, and
physiology; by necessity, only a portion of the primary
literature for each topic is reviewed.
Tumor necrosis factor is synthesized as a 26-kDa type
II transmembrane precursor that is displayed on the
plasma membrane, with the N-terminus in the cyto-
plasm and the C-terminus exposed to the extracellu-
lar space. The TNF precursor is proteolytically cleaved
between alanine (-1) and valine (+ 1) to yield a bio-
logically active 17-kDa mature TNF (Kriegler et al.,
1988) that forms a trimer in solution (Smith and
Baglioni, 1987). In solution, 17-kDa TNF is primarily
comprised of two anti-parallel [3 sheets; these TNF
monomers self-associate in a head-to-tail fashion
into non-covalently linked, symmetrical bell-shaped
homotrimers (Smith and Baglioni, 1987; Jones et al.,
1989). The C-terminus of each subunit is embedded
within the base of the trimer, while the N-terminus is
relatively free from the structure and is not critical for
TNF biologic activity (Creasey et al., 1987). Mutational
analyses of mature TNF indicate that each trimer
has three receptor interaction sites located in the

The Cytokine Handbook, 4th Edition, edited by Angus W. Thomson & Michael T Lotze Copyright 9 2003 Elsevier Science Ltd.
ISBN 0-12-689663-1, London All rights of reproduction in any form reserved.
838 TUMOUR NECROSIS FACTOR
intersubunit grooves near the base of the trimer
(Zhang et al., 1992). The trimeric structure of TNF is
important for its biologic activity, because mutations
that destabilize the trimer or attenuate monomer
association result in the loss of TNF biologic activity
(Lin, 1992). Recently identified TNF muteins have
been shown to possess decreased toxicity in mice
without loss of its antitumor effects (Van Ostade et al.,
1993; Barbara et al., 1994). For example, residues
within the inter-subunit grooves at the base of the
trimer dictate receptor specificity: the double mutant
R32W/S86T exhibited wild-type binding to the 55-
kDa receptor (TNFR1) with no measurable binding to
the 75-kDa TNF receptor (TNFR2). In contrast, the
D143N/A145R double mutant fails to bind TNFR1, but
does bind TNFR2 with five- to 10-fold lower affinity
than wild-type TNF (Loetscher et al., 1993). There
may be differences between species, because the
R32W/S86T double mutant, which was non-toxic in
mice, was significantly toxic to healthy baboons (Van
Zee et al., 1994). A recently identified triple mutant
($52I, Y56F, plus deletion of the seven N-terminal
residues) has 11- and 71-fold lower affinity for the 55-
kDa and 75-kDa receptors, respectively, and is 10-fold
less toxic than wild-type TNF (Cha et al., 1998). These
findings have raised hope for an engineered TNF
protein that possesses tumor cytotoxicity activity, but
with reduced systemic toxicity as compared with the
native cytokine.
Transmembrane pro-TNF is also biologically active,
but less is known about its structure-function rela-
tionship. Prior to transport to the cell surface, pro-
TNF is assembled into a bell-shaped trimer that is
similar to the structure of mature TNF (Jones et al.,
1989). A 20-residue linker sequence connects the
transmembrane domain with the N-terminus, which
facilitates but is not necessary for trimer formation.
During the interaction of effector and target cells,
membrane-bound pro-TNF in the effector cells binds
TNF receptors on the target cells, and induces TNF
responses via receptor aggregation (Kriegler et al.,
1988). Pro-TNF is a more potent activator of TNF
receptor 2 (TNFR2) than mature TNF (Grell et al.,
1994).
Highly acidic environments alter the tertiary struc-
ture of TNF such that the 13-sheet structure is aban-
doned in favor of ~ helices (Narhi et al., 1996). TNF is
unusual in that it has no ~ helix in its native structure
THE CYTOKINES AND CHEMOKINES
but forms an a-helical structure in the molten globule
(an intermediate structure produced by acid-induced
unfolding of proteins). With the resulting increase
in hydrophobicity at low pH, TNF becomes more
cytolytic, perhaps by insertion into membrane to
form sodium ion channels (Tracey et al., 1986;
Yoshimura and Sone, 1987; Baldwin et al., 1996). The
conformational change at low pH may be physiologi-
cally relevant, because a pH of about 3.6 has been
reported in the space between macrophages and their
substrates (Silver et al., 1988). Recent membrane con-
ductance studies have shown that the capacity of TNF
to increase membrane conductance does not corre-
late with its ability to interact with membranes. These
results suggest that TNF itself does not form a chan-
nel, but may increase sodium flux via interaction with
endogenous ion channels or with plasma membrane
proteins that are coupled to ion channels (van der
Goot et al., 1999).
BIOSYNTHESIS AND
REGULATION
TNF is produced by numerous immune cells in
response to an assortment of activating stimuli (Table
35.1). Among these stimuli, lipopolysaccharide (LPS)
(in monocytes), T-cell receptor activation (in T lym-
phocytes), crosslinking of surface immunoglobulin
(sIg) (in B lymphocytes), ultraviolet light (in epithelial
cells), and viral infections have been widely studied.
Suppressors of TNF expression in macrophages are
also highly diverse and include prostaglandin E 2
(PGE2), transforming growth factor 13 (TGFI3), IFN~,
IFNT, IL-4, IL-6, IL-10, G-CSE certain viral products
such as adenoviral proteins, dexamethasone, gluco-
corticoids, cyclosporin A, spermine, pentoxifylline
cAME and cholinergic agonists including acetyl-
choline and nicotine.
The biosynthesis of TNF is tightly controlled at
many different levels to ensure the production of van-
ishingly small quantities in quiescent cells, yet TNF is
capable of rapid and significant up-regulation in acti-
vated cells (Beutler et al., 1985). TNF is expressed as
an immediate early gene, and a variety of stimuli
induce high levels of TNF mRNA within 15-30 min-
utes with no requirement for de novo protein synthe-
sis, suggesting that the factors necessary for the
 BIOSYNTHESIS AND REGULATION
Table 35.1 Cell source a n d stimuli o f TNF production
Cell sources I m m u n e cells:
Macrophages/monocytes, natural
killer cells, Kupffer cells, B cells, T
cells, basophils, eosinophils, glial
cells, mast cells
N o n - i m m u n e cells:
Astrocytes, granulosa cells,
osteoblasts, cardiac myocytes,
fibroblasts, keratinocyte, neurons,
neutrophils, T cells, retinal pigment
epithelial cells, smooth muscle cells,
spermatogenic cells, tumor cells
Stimuli Bacterial endotoxin (LPS), antibodies
to LFA-3, calcium ionophores, C5a,
CD44, CD45, enterotoxin, GM-CSE
hypoxia, IL-1, leukotrienes, mellitin,
MIP- 1~, mycobacterial
lipoarabinomannan, nitric oxide,
oxygen radicals, parasites, phorbol
esters, synthetic lipid A, TNE toxic
shock toxin- 1, viruses
Irradiation
induction of TNF expression pre-exist in unstimu-
lated cells. The cell-type-specific regulation of TNF
synthesis is reflected by the differential use of regula-
tory elements on TNF biosynthesis in different cells.
Each one of the synthetic steps from sensing the pres-
ence of a stimulus to TNF secretion is controlled by
different molecular events regulating the TNF gene,
mRNA and protein.
Transcriptional regulation
TNF is regulated at the transcriptional level by both
activators and repressors. Besides a TATA box pro-
moter located 20 base pairs (bp) upstream from the
transcription start site, a number of regulatory
sequences are also found upstream of the TNF gene
(Figure 35.1), including three nuclear factor (NF)~:B
sites designated ~:1, ~:2 and ~:3 (Goldfeld et al., 1993);
three nuclear factor of activated T cells (NFAT) bind-
ing sites for NFATp, NFAT-149, NFAT-117 and NFAT-76
(Tsai et al., 1996); one 'Y' box; one cAMP-responsive
element (CRE) for activation transcription factor-2
(ATF-2)/Jun (Tsai et al., 1996); two SV40 promoter-1
(SP-1) sites (Kramer et al., 1994); one activating pro-
tein- 1 (AP-1) and one AP-2 binding site for Fos/Jun
(Leitman et al., 1992); one E26 transformation-
THE CYTOKINES AND CHEMOKINES
839
Y box SP1 NFAT NFAT ~:3 AP1 AP2
-232 -164 -149 -117 -89 -58 -27
I I , I, I ,I , I , I , I , [TNFGENE~"
~:1 ~:2 Egr-1 Ets-1 CRE NFAT SP1 TATA
-577 -201 -163 -118 -100 -76 -45 -20
FIGURE 35.1 Model of the TNF promoter. Putative
cis-acting consensus sequences are denoted by boxes
and numbers indicating the position in relation to the
transcription start site.
specific (Ets) binding site (Kramer et al., 1995); and an
early growth responsive- 1 (Egr-1) binding site
(Kramer et al., 1994). Although K3 matches the bind-
ing consensus sequence for the transcription factor
NFKB, K3 appears to bind to NFATp instead of NFKB.
The CRE element in TNF promoter binds mainly to
ATF-2/Jun. Genetic analyses suggest that additional
regulatory elements may be important in regulating
TNF transcription (Wong and Goeddel, 1988). TNF
mRNA expression is negatively regulated by factors
that are less well characterized. Several regions within
the TNF promoter, including b a s e s - 2 8 0 to -172,
-254 to -230, -125 to -82, and - 9 5 to -36, relative
to the transcription start site, seem to be critical for
inhibition ofTNF mRNA expression (Fong et al., 1995;
An et al., 1999). The transcriptional regulators that
bind these regions remain enigmatic. While AP-1 and
AP-2 consensus binding sequences have been noted
within these regions (An et al., 1999), evidence for
binding of these repressors to the TNF promoter is
lacking (Fong et al., 1995). More recently, Singh et al.
(2001) have found that heat-shock factor 1 (HSF1)
binds to regions - 1080 to -845, -533 to -196 and
-326 to - 3 9 of the TNF promoter relative to the
transcription start site, and represses TNF gene
transcription.
The regulation of TNF transcription is cell-type spe-
cific, partially because of differential use of these reg-
ulatory elements on the TNF promoter. In calcium- or
T-cell ligand-activated At-5 T lymphocytes, a func-
tional interaction between NFATp on the K3 site and
ATF-2/Jun on the CRE element, but not AP-1 or AP-2,
plays a crucial role in the induction of TNF expression
(Tsai et al., 1996). In calcium-activated A20 B lympho-
cytes, NFATp on the K3 element is not required, but
NFATp on NFAT-76 participates in the induction of
TNF synthesis (Tsai et al., 1996). In phorbol myristate
acetate (PMA)-activated T cells, the interaction
between Ets and ATF/Jun is essential for both basal
840 T U M O U R NECROSIS FACTOR
and induced TNF expression. Post-translational mod-
ification of the transcription factors that regulate TNF
transcription, such as NFATp, NF)cB and AP-1, has
been implicated in the regulation of TNF induction.
Protein kinases and phosphatases are also involved in
the induction of TNF (Lee et al., 1994), as are pro-
teases and phospholipase D (Balboa et al., 1992).
Some of these enzymes modify the activity of TNF
induction-related transcription factors such as NFKB
and AP- 1 (see below).
Translational regulation
TNF synthesis is highly inducible upon cell stimula-
tion, and is tightly regulated in quiescent cells; this
enforced inhibition of TNF expression protects
against the harmful effects of TNF overexpression. In
response to LPS stimulation, macrophages and
monocytic cells produce high levels of TNF protein,
an effect that is blocked by pretreatment with dexam-
ethasone, spermine or CNI-1493 (a tetravalent
guanylhydrazone) (Bianchi et al., 1996; Beutler et al.,
1986; Cohen et al., 1996; Zhang et al., 1997). Recent
studies have shown that cholinergic agonists such as
acetylcholine and nicotine inhibit TNF synthesis via
post-transcriptional mechanisms (Borovikova et al.,
2000). A key element for both negative and positive
translational regulation of TNF has been identified in
the 3' untranslated region (UTR) of TNF mRNA and
many other cytokines, growth factors and oncopro-
teins (Caput et al., 1986; Han et al., 1990). The mini-
mal motif UUAUUUAUU renders the TNF mRNA
unstable (Shaw and Kamen, 1986; Zubiaga et al.,
1995), and the presence of a single octanucleotide
(UUAUUUAU) in the 3' UTR is sufficient to signifi-
cantly inhibit translation (Kruys et al., 1989). This
sequence physically interacts with the poly(A) tail of
the TNF mRNA, preventing the mRNA from forming
large polysomes (Crawford et al., 1997). Regulation of
mRNA turnover is mediated through this region by
trans-acting proteins such as tristetraprolin (TTP)
(Lai et al., 1999), HuR (Dean et al., 2001), TIA- 1 (Piecyk
et al., 2000), and TIAR (Gueydan et al., 1999), which
bind AO-rich elements and stabilize or destabilize the
transcript. Mice deficient in tristetraprolin (TTP), the
prototype of a family of CCCH zinc-finger proteins,
develop an inflammatory syndrome mediated by
excess TNF (Lai et al., 1999). Other AO-rich-binding
THE CYTOKINES A N D C H E M O K I N E S
proteins, such as TIA-1, normally function as transla-
tional silencers (Piecyk et al., 2000), because
macrophages derived from TIA-1 knockout mice have
a significantly higher proportion of TNF transcript
associated with polysomes. TIA-1 knockout mice fail
to inhibit TNF production and are exquisitely sensi-
tive to endotoxemia (Piecyk et al., 2000). The 3' UTR of
TNF mRNA is equally important for stimulating TNF
synthesis, because a polymorphism (a GAU trinu-
cleotide insertional mutation) present in the regula-
tory 3' UTR ofTNF mRNA of NZW mice prevents the
interaction of RNA-binding proteins with target
sequences, and significantly reduces TNF production
(Di Marco et al., 2001).
Signal transduction cascades
regulating TNF production
Mitogen-activated protein (MAP) kinases such as the
extracellular signal-regulated kinase (ERIO, c-Jun N-
terminal kinase (JNIO, p38 and Big MAP kinase
(BMIO/ERK5 have a critical role in the regulation of
TNF production (Zhu et al., 2000). The roles of
ERK1/2- and p38 MAPK-dependent pathways in reg-
ulating cytokine production have been demonstrated
primarily through the use of specific inhibitors (such
as PD098059 for ERK1/2 activation, and SB203580 for
p38 MAP kinase) (Cuenda et al., 1995; Alessi et al.,
1995), which prevent cytokine production in LPS-
stimulated macrophage/monocytes (Feng et al., 1999;
Guha et al., 2001). p38 MAP kinase may be critical in
the translational control of TNF synthesis, because
activation of the MAP kinase cascade enhances the
translational efficiency of TNF mRNA (Lee et al.,
1994). Mitogen-activated protein (MAP) kinase/extra-
cellular signal-regulated kinase (ERIO kinase (MEKIO
activates TNF induction-associated transcription fac-
tors such NFKB, c-Jun and p38 (Cohen et al., 1996; Lee
S.Y. et al., 1996). An abridged listing of transcriptional
and translational control mechanisms in TNF synthe-
sis is shown in Figure 35.2. Multiple signaling path-
ways converge on NFKB, a ubiquitous transcription
factor involved in the transduction of activation sig-
nals from the cytoplasm to the nucleus. NFKB resides
in the cytosol as a trimer consisting of p50, p65
and I~:B subunits. Upon stimulation, IKB is released,
and the p50/p65 heterodimer migrates to the nucleus
where it interacts with target DNA sequences
 BIOSYNTHESIS AND REGULATION
I Inflammatory stimuli , I
I aa,c/ 0dc421
IMEKK,,21 I ASK1 .....I
IMKK3,61
I ".
I MAPKAP:K2 1
Y
j
TNF DNA ~ ,. mRNA ,. Protein
FIGURE 35.2 Schematic illustration of signal trans-
duction pathways regulating TNF transcription and
translation in macrophages.
resulting in the transcription of TNF and other pro-
inflammatory cytokines.
Post-translational regulation
TNF is synthesized as a 233-amino-acid membrane-
anchored prohormone that is subsequently processed
proteolytically to yield the mature 157 residue
cytokine. Analysis of the cleavage site sequences in
human TNF revealed significant homology with
sites in collagen, %-inhibitory proteins, and %-
macroglobulin cleaved by matrix metalloproteinases
(MMPs). The TNFcz-converting enzyme (TACE) is an
MMP-like enzyme (Black et al., 1997; Moss et al.,
1997), and metalloproteinase inhibitors block TNF
processing (Gearing et al., 1994). Gearing and col-
leagues (1995) have found that TACE activity is
expressed ubiquitously, even in HeLa cells that do not
produce TNE and in insect cells where pro-TNF is
expressed by means of a baculovirus transfectant. A
serine protease, proteinase 3 (PR-3), may also process
TNE and serve as an extracellular alternate processing
enzyme for TNF and IL-I[3 (Coeshott et al., 1999).
Endotoxin, as well as TNF and other cytokines, can
THE CYTOKINES AND CHEMOKINES
841
modulate MMP levels (Panagakos et al., 1996), and
plasminogen and plasmin increase secretion of
MMPs and induce cleavage to their active forms (Lee
et al., 1996).
Application
Uncontrolled synthesis of TNF has been implicated in
many human diseases of systemic inflammation,
including sepsis, arthritis and Crohn disease. Conse-
I quently, there has been a great deal of interest in
developing therapeutic strategies to inhibit TNF activ-
ity, and several inhibitors have been developed to
attenuate TNF activity, release, translation or tran-
scription. Corticosteroids, the most studied class of
compounds that suppress the production of TNF,
inhibit the transcription of TNF and other cytokines.
One immunosuppressive mechanism of these drugs
is through the stimulation of IKB~ production
(Auphan et al., 1995; Scheinman et al., 1995), which in
turn results in the inactivation of NFKB. Corticos-
teroids also inhibit gene transcription by preventing
the transcription factor, AP-1, from binding to its tar-
get promoter (Marx, 1995). Pentoxifylline and other
protein kinase C (PKC) inhibitors regulate the pro-
duction of TNF and other cytokines by blocking PKC-
or PKA-catalyzed activation of NF~:B (Biswas et al.,
1994). Cytokine-suppressing anti-inflammatory drugs
(CSAIDs) (Young et al., 1994) have no significant effect
on the transcription of TNF or other cytokines, but
rather suppress TNF translation by inhibiting the
activity of p38 MAP kinase (Lee et al., 1994). CNI- 1493,
a tetravalent guanylhydrazone that suppresses sys-
temic and central inflammation (Meistrell et al., 1997;
Tracey, 1998), inhibits TNF by preventing phosphory-
lation of p38 MAP kinase (Cohen et al., 1996; Denham
et al., 2000). At the post-translational level, thalido-
mide selectively suppresses the release of TNF but not
of other cytokines (Sampaio et al., 1991). IL-4 and
IL-10 suppress TNF activity by inhibiting the bio-
synthesis of matrix-destructive metalloproteinases
(Lacraz et al., 1992, 1995) and down-regulating the
proteolytic processing of pro-TNF to its mature form.
Furthermore, IL-4 inhibits the activation of the tran-
scriptional factors NFKB and AP-1, providing yet
another mechanism by which IL-4 can attenuate TNF
activity. Like IL-4, IL-10 also inhibits TNF as well as
other cytokines at multiple levels (Bogdan et al., 1992).
842 TUMOUR NECROSIS FACTOR
RECEPTORS
The pleiotropic effects of TNF are mediated through
two distinct but structurally homologous TNF recep-
tors, the 60-kDa receptor 1 (TNFR1, also known as
p55 or p60) and the 80-kDa receptor 2 (TNFR2, also
known as p75 or p80), which are both type I trans-
membrane glycoproteins and members of the TNF
receptor superfamily. Members of this receptor family
share sequence homologies within their extracellular
domains, consisting of multiple cystine-rich repeats
of about 40 amino acids in length (Mallett and
Barclay, 1991). TNFR1 and TNFR2 are present on
virtually all cell types except for red blood cells
(Hohmann et al., 1989). TNFR1 is more ubiquitous,
but TNFR2 is more abundant on endothelial cells and
cells of hematopoietic lineage (Hohmann et al., 1989;
Brockhaus et al., 1990). Both receptors can bind to
TNF with high affinity; the dissociation constants (Ka)
are 2-5 • 10-~~ 3-7 • 10 -11, respectively.
The precise roles of the TNF-induced receptor sig-
naling in mediating the effects of TNF continue to be
the focus of intensive study. TNFR1 is generally con-
sidered to be responsible for the majority of biologic
actions of soluble TNE Direct signaling through
TNFR2 occurs less extensively and appears to be con-
fined mainly to cells of the immune system, and may
be especially important during cell-cell contact
(Kriegler et al., 1988). Fibroblasts derived from
TNFRl-knockout mice are deficient in leukocyte
adhesion, and display reduced expression of inter-
cellular adhesion molecule- 1 (ICAM-1), vascular cell
adhesion molecule-1 (VCAM-1), and CD44. These
cells also fail to up-regulate major histocompatibility
complex (MHC) class I expression, cytokine secretion,
cell proliferation, and NFKB activation. These activi-
ties appear to be specific attributes of the TNFR1
receptor, because activation of cells expressing only
TNFR2 with exogenous TNF has no effect on these
functions (Mackay et al., 1994). Similar observations
were made with TNF mutants that selectively bind
TNFR1. TNFRl-selective TNF mutants that bind
TNFR2 poorly are equally cytotoxic and cytostatic to
carcinoma and leukemic cell lines, but are less pro-
inflammatory, than wild-type TNE TNFRl-selective
mutants also recapitulate the hypotensive and proin-
flammatory activities of wild-type TNF in vivo (Van
Zee et al., 1994), whereas TNFR2 by itself does not
THE CYTOKINES AND CHEMOKINES
seem to be sufficient to stimulate these functions
(Barbara et al., 1994). These observations suggest that
the role of TNFR2 may be to potentiate the effects of
TNFR1. A unique 'passing model' has been suggested
to explain the role of TNFR2 in mediating TNF
responses (Tartaglia et al., 1993b). The rapid rate of
dissociation of the TNF-TNFR2 complex may facili-
tate the interaction of TNF with TNFR1 by recruiting
TNF to the vicinity of the plasma membrane. In this
scenario, TNFR2 can be envisioned as passing TNF to
TNFR1, the main TNF signal transducer. In some
cases, however, TNFR2 is also believed to mediate
several TNF effects, such as proliferation of T and B
cells (Tartaglia et al., 1991; Barbara et al., 1994), induc-
tion of NF~:B, and cytotoxicity (Hohmann et al., 1990;
Grell et al., 1994). TNF receptor activities are also sub-
ject to regulation by a wide variety of agents such as
PKC and PKA modulators, LPS, retinoic acid, gluco-
corticoids and cytokines including TNF; IL-2, IL-4,
IL-6, IL-8; IFNct, 13,7; GM-CSF; and thyroid-stimulating
hormone. Therefore, expression of either TNFR1 or
TNFR2 is dependent upon cell type, as well as extra-
cellular stimuli.
Soluble receptors for TNF (sTNFR, also known as
TNF-BP for TNF-binding protein) are derived from
both TNFR1 and TNFR2 by proteolytic processing.
Both sTNFR1 and sTNFR2 are normally present in
blood and urine (Seckinger et al., 1989), and may be
produced by monocytes and macrophages (Gatanaga
et al., 1991; Leeuwenberg et al., 1994). Increased levels
of sTNFRs are found in many human diseases includ-
ing rheumatoid arthritis, systemic lupus erythemato-
sus, malignancy, sepsis, after surgery and chronic
infection (Chikanza et al., 1993; Aderka, 1996). A num-
ber of stimuli, including TNE IL-1, PMA and endo-
toxin, trigger proteolysis of the TNFRs, resulting in
increased levels of sTNFRs in the circulation. The pro-
teases responsible for this processing have not yet
been identified, but may involve the same metallo-
proteinases that process the TNF precursor to its
mature form, because metalloproteinase inhibitors
block the shedding of both TNF receptors (Crowe
et al., 1995; Mullberg et al., 1995). Recently, matrix
metalloproteinase processing of both precursor TNF
and the TNF receptors has been implicated in
humans in vivo; administration of an MMP inhibitor
prior to endotoxin infusion attenuated the release
of both TNF and TNFRs into the circulation. This
 SIGNAL TRANSDUCTION 843

attenuation was accompanied by an increase in
membrane-bound TNF and TNFRs in monocytes iso-
lated from these patients (Dekkers et al., 1999). Solu-
ble TNFRs bind and neutralize both circulating and
membrane-bound TNF (Seckinger et al., 1989; Kohno
et al., 1990). In vivo, administration of sTNFRs neu-
tralizes or modulates TNF activity when sTNFRs are
present in high concentrations; lower concentrations,
however, stabilize the TNF trimer, providing a TNF
reservoir (Mohler et al., 1993).
SIGNAL TRANSDUCTION
TNF mediates or initiates diverse biologic responses
by interaction with TNFR1 or TNFR2. Investiga-
tions into the signaling events activated by the lig-
and/receptor complex have been hampered by the
diversity of cell-specific TNF responses, and a lack of
similarities with other known cell surface receptor
signaling pathways. X-Ray data suggest that the TNF
homotrimer induces receptor trimerization, which
activates receptor signaling. Several downstream sig-
naling events initiated by the TNF receptor complex
have been characterized, and signaling molecules
that mediate the initial interaction with the ligand-
occupied receptor have been identified.
TNFRl-mediated signaling pathway
Multiple intracellular functional domains within
TNFR1 transduce intracellular signals to coordinate
the biological activity of TNF (Tartaglia et al., 1993a)
by interacting with intracellular adaptor proteins
(Figure 35.3). Three functional domains of TNFR1 -
the C-terminal death domain (Tartaglia et al.,
1993a), and the adjacent N-SMase (neutral sphingo-
myelinase) and A-SMase (acidic sphingomyelinase)

.SD I ASD Deathdomain TNFR1

I~N I I ~c-P,C I
I

N-terminus Death domain

N-SMase [ LA-sMase II P~C I

Ceramide
(membrane)
Ceramide
(endosome) I,~ I--IN'~ I L~'DD L ~-I P~D,P~-~ I
I MEKKI'2 II Caspase-2 II Caspase-8 I
I~ I
NF-KB /I-KB
complex IJNKI Protease cascade

~ er0r~
Cell growth Inflammation Anti-apoptosis Apoptosis

FIGURE 35.3 The TNF receptor 1 (TNFR1)-mediated signaling transduction pathways. TNF triggers different sig-
naling pathways via three functional domains of TNFR1 to mediate various activities.

THE CYTOKINES AND CHEMOKINES

844 TUMOUR NECROSIS FACTOR
activating domains (NSD and ASD) (Schutze et al.,
1992; Wiegmann et al., 1994) - transfer signals from
extracellular TNF to intracellular adaptor proteins
that can contain additional death domains and/or
caspase recruitment domains (CARD). TRADD
(TNFRl-associated death domain protein) binds to
the death domain ofTNFR1 (Hsu et al., 1995) to medi-
ate both apoptotic and antiapoptotic pathways.
Proapoptotic effects of TNF are also mediated
through activation of phosphatidylcholine-specific
phospholipase c (PC-PLC) by the ASD domain
(Wiegmann et al., 1994; Machleidt et al., 1996), which
shares receptor regions and function with the death
domain. The factor associated with N-SMase activa-
tion (FAN) binds the NSD domain of TNFR1 (Adam-
Klages et al., 1996) to mediate a number of
inflammatory and cell proliferation responses.
Together, it is apparent that TNFR1 signaling diverges
into three functional branches at the level of the
receptor itself." one that is mediated through the death
domain to positively and negatively regulate apop-
totic responses, and two that are processed through
sphingomyelinases to modulate apoptotic and
inflammatory pathways.
Death domain-mediated pathways
The TNFRl-associated death domain protein
(TRADD) mediates both apoptosis and antiapoptosis
pathways by activating the interleukin-1 converting
enzyme (ICE) pathway or the NFKB pathway, respec-
tively (Hsu et al., 1995). The TRADD protein contains
a death domain similar to the death domain of
TNFR1. While overexpression of an isolated TRADD
death domain can trigger cell death, overexpression of
the full-length protein does not, suggesting that
TRADD does not carry a caspase recruitment domain
(CARD). This has led to one currently accepted view of
TRADD as an adaptor protein that links TNFR1 to
other death effectors. Once bound to TNFR1, TRADD
recruits MORT1/FADD (Fas-associating protein with
death domain) or RIP (receptor-interacting protein)
through its C-terminal death domain to activate
divergent apoptotic pathways (Grimm et al., 1996),
and recruits TNF receptor-associated factor 2 (TRAF2)
through its N-terminal TRAF-interacting domain
(Hsu et al., 1996). Thus, TNFR1 death domain signal-
ing diverges at the level of TRADD into three distinct
THE CYTOKINES AND CHEMOKINES
pathways: one that is dependent upon TRAF2 and
activates inflammatory and antiapoptotic responses
via NFKB and c-Jun; a second pathway that activates
caspase 2 through the activity of RIP to induce apop-
tosis; and a caspase 8-dependent apoptotic pathway
that is dependent upon FADD activity.
The TNFR1-TRADD complex initiates apoptotic
signaling through pathways dependent upon either
FADD or RIP. Overexpression of FADD (Chinnaiyan
et al., 1995; Boldin et al., 1996) or RIP (Stanger et al.,
1995) results in apoptosis; however, a dominant-
negative mutant of FADD inhibits TNF-induced
apoptosis but not TNF-mediated NFKB activation
(Hsu et al., 1996), demonstrating that the apoptotic
and inflammatory cascades diverge at the level of the
TRADD-FADD complex. Activated FADD recruits the
FADD-homologous ICE/CED-3-1ike protease (FLICE),
also referred to as pro-caspase 8 or MACH, which
interacts with the caspase recruitment domains
(CARD) of FADD, where it oligomerizes and auto-
proteolytic activity generates catalytically active
capase-8 (Boldin et al., 1996; Muzio et al., 1996). The
FADD-FLICE interaction is negatively regulated by
PED/PEA- 15, possibly by the dissociation of the com-
plex (Condorelli et al., 1999). The death-associated
protein (DAP) kinase is another death domain-
containing protein that operates downstream of
FLICE but upstream of other caspases to activate
apoptosis (Cohen et al., 1999). The FADD-activated
caspase cascade induces apoptosis by proteolytically
activating protein precursors, such as lamin, actin
and polymerase, or by activating proteins that lead to
cell death (Figure 35.3).
TNFR1 can also induce apoptosis via the TRADD-
RIP pathway. All three RIP family members (RIP, RIP2
[also known as Rick or CARDIAK] or RIP3) possess
protein kinase and death domains, and transduce
intracellular apoptotic signals. The death domain of
RIP can mediate the death signal in the absence of the
kinase domain, suggesting that RIP does not possess a
CARD but rather induces apoptosis by recruiting
other downstream effector molecule(s) through the
RIP death domain. RIP binds preferentially to TRADD
via protein-protein interaction between the death
domains on both proteins (Hsu et al., 1996), but can
also bind weakly to TNFR1 (Stanger et al., 1995).
TNFR1-TRADD-RIP activate apoptosis via the inter-
mediate effector, RAIDD (for RIP-associated Ich-l/
 SIGNAL TRANSDUCTION
CED-3 homologous protein with death domain),
which binds RIP via a death domain (Duan and Dixit,
1997). The RAIDD/RIP complex recruits caspase-2
(Ich-1) to activate the protease cascade leading to
apoptosis.
RIP can also bind to TRAF2, which can be recruited
to TRADD simultaneously with RIP. The significance
of RIP binding to TRADD and TRAF2 as well as TNFR1
is still obscure, and it is possible that RIP may coordi-
nate these signaling pathways. Overexpression of
TRAF2 activates NF~:B (Rothe et al., 1995) but TRAF2
is not required for the induction of apoptosis (Hsu
et al., 1996). A serine-threonine kinase known as
NFKB-inducing kinase (NIK) binds specifically to
TRAF2 and mediates TRAF2-dependent activation of
NF~cB (Malinin et al., 1997). The binding between
TRAF2 and NIK is mainly dependent on the C-terminus
of TRAF2, but also involves the N-terminal of TRAF2.
Because NIK can be autophosphorylated and has
sequence similarity to several kinases that participate
in MAP kinase cascades, it may act in a kinase cascade
to activate NF~cB. The cascade downstream of NIK
that is responsible for NFKB activation, however,
is still unclear. TRAF2-dependent activation of
MEKK and c-Jun amino-terminal kinase (JNK) / stress-
activated protein kinase (SAPK) through TNFR1 has
been reported (Liu et al., 1996; Natoli et al., 1997). As
MEKK has been shown to induce the phosphorylation
and degradation of IKB~, resulting in the activation
of NFKB (Lee et al., 1997), MEKK may function as
a downstream factor of NIK and mediate TRADD-
TRAF2-directed activation of NFKB to prevent TNF-
induced apoptosis (Figure 35.3). It is plausible that
this may be one of the mechanisms of TNF self-
tolerance. Because NF~:B and JNK are major factors
regulating the inflammatory response, this pathway
may also play a role in the proinflammatory effect of
TNE
NSD- mediated pathway
NSD (neutral sphingomyelinase domain) occupies a
central role in mediating a number of TNF activities,
including inflammatory responses and cell pro-
liferation through the MAP kinase ERK and PIA2,
respectively (Figure 35.3). The novel WD repeat ({X6_94-
[GH-X23_41-WD]}N4_8) protein FAN couples TNFR1 to
N-SMase via protein-protein interactions between
THE CYTOKINES AND CHEMOKINES
845
the C-terminal WD repeats of FAN and a cytoplasmic,
nine-amino-acid binding motif (residues 310-318) of
TNFR1 (Adam-Klages et al., 1996). FAN specifically
activates N-SMase, and not A-SMase (see ASD-
mediated pathway below), which modulates the pro-
duction of ceramide, a product of the sphingomyelin
(SM) degradation. Ceramide is generated at several
distinct subcellular locations that influence its action
(Wiegmann et al., 1994). Activated N-SMase func-
tions at the outer leaflet of the plasma membrane to
trigger the degradation of sphingomyelin (SM) into
ceramide, which in turn mediates further down-
stream events in NSD-mediated TNF response.
Ceramide is also produced in endosomes by A-SMase,
where ceramide mediates a distinct signaling path-
way. On the plasma membrane, ceramide further
activates ceramide-activated protein kinase (CAPK)
(Winston and Riches, 1995), which phosphorylates
cytoplasmic raf-1. Activated raf-1 can activate the
MAP kinase cascade and activate MAPK/ERK (extra-
cellular signal-regulated kinase), which in turn
activates PI2K2 (Lin et al., 1993). PUK2 may be respon-
sible for the generation of the arachidonic acid
metabolites, leukotrienes and prostaglandins, which
contribute to TNF proinflammatory activities.
MAPK/ERK also induces the expression of c-myc and
other genes to regulate cell proliferation and differen-
tiation. Activation of ERK may act as a negative-
feedback loop to attenuate TNF-induced apoptosis,
because inhibition of ERK is required for induction of
apoptosis in some systems (Xia et al., 1995).
ASD-mediated pathway
N-SMase and A-SMase are activated independently
by different cytoplasmic domains ofTNFR1, and elicit
discrete signaling pathways (Wiegmann et al., 1994).
Like the death domain, the ASD can induce activation
of NFKB to protect against apoptosis (Van Antwerp
et al., 1996; Wang et al., 1996), but is also involved in
induction of apoptosis (Pena et al., 1997). The ASD
maps to the same stretch of residues within the death
domain, and may in fact be the same domain
(Tartaglia et al., 1993a; Wiegmann et al., 1994). The
ASD mediates activation of A-SMase via PC-PLC
(Schutze et al., 1992). The connection between the
ASD in TNFR1 and PC-PLC is still obscure, but
appears to involve the activity of protein kinase C
846 T U M O U R NECROSIS FACTOR
(PKC) (Mtiller et al., 1995). Activated PC-PCL pro-
duces 1,2-diacylglycerol (DAG), which activates A-
SMase and contributes to the activation of NFKB;
TNF-induced A-SMase and NFKB activity is blocked
by the specific PC-PLC inhibitor, D609, which effec-
tively inhibits DAG production (Schutze et al., 1992).
A-SMase produces ceramide in the acidic endoso-
mal/lysosomal compartment (Spence, 1993). How-
ever, the precise role of A-SMase-induced ceramide
is unclear, because cells that are deficient in
ceramidase, and therefore accumulate intracellular
ceramide, are equally responsive to apoptotic signals
from TNFR1. In addition, TNF- or CD95-induced
apoptosis does not induce the degradation of labeled
sphingomyelin (Segui et al., 2000). A-SMase activity in
TNF-induced NFKB may in fact be cell-type specific,
because a recent study by Kouba et al. (2001) revealed
that A-SMase activity can be used as a surrogate
mediator of NF~:B activation.
TNFR2-mediated signaling pathway
Direct signaling through TNFR2 is less well character-
ized, and appears to be confined primarily to cells of
the immune system. TNFR2 mediates specific TNF
effects including proliferation of T and B cells
(Tartaglia et al., 1991; Barbara et al., 1994), induction
of NFKB and cytotoxicity (Hohmann et al., 1990a,
1990b; Grell et al., 1994), and may potentiate the
effects of TNFR1. TNFR2 can down-regulate human
and murine activated T cells by inducing apoptosis
(Zheng et al., 1995). TNFR2 lacks an intracellular
death domain, suggesting that TNFR2 uses distinct
signaling pathway(s) to induce apoptosis. Down-
regulation of the anti-apoptotic protein Bcl-xL (Boise
and Thompson, 1997) has been correlated with
TNFR2-induced apoptosis (Lin et al., 1997), but the
precise mechanisms triggering TNFR2-induced apop-
tosis remain to be elucidated. Whether TNFR2 alone
can mediate the activation of NFKB is controversial.
Although TRAF2 interacts with TNFR2, and TRAF2-
TRAF1 (Rothe et al., 1995) and TRAF2-TANK (Cheng
and Baltimore, 1996) heterocomplexes are able to
mediate the activation of NFKB, the results of a recent
study (Chainy et al., 1996) indicate that TNFR2 does
not participate in the activation of NFKB in many cell
types, including immune cells.
THE CYTOKINES A N D C H E M O K I N E S
BIOLOGIC EFFECTS
TNF was originally characterized as a protein induc-
ing necrosis of methylcholanthrene (Meth A)-induced
sarcomas in vivo (Carswell et al., 1975) and believed to
be a growth modulatory agent that exhibits primarily
antitumor activity. The discovery of the systemic
inflammatory role of TNF in non-malignant disease
(Tracey et al., 1986b) led to a focus on the pathogenic
role of TNF in many immune-mediated diseases. The
net effects of TNF are influenced by a complex array of
cell- and tissue-specific factors. The diverse role of
TNF in mediating cellular responses is summarized in
Table 35.2.
Central nervous system
Systemic TNF produces fever and anorexia via hypo-
thalamic centers that regulate body temperature and
appetite (Tracey et al., 1988; Plata-Salaman, 1991).
The anorectic effect of TNF is attenuated by insulin
and appears to be independent of serotonergic sig-
nals (Tracey and Cerami, 1990). Cells of the central
nervous system (CNS), including microglia, astrocytes
and neurons (Cheng et al., 1994), express TNF and
high-affinity TNF receptors (Smith and Baglioni,
1992). Furthermore, elevated TNF levels have been
detected in brain injury and certain CNS diseases
including stroke (Liu et al., 1994), meningococcal
meningitis (Leist et al., 1988), human immuno-
deficiency virus infection (Grimaldi et al., 1991),
Alzheimer's disease (Fillit et al., 1991) and multiple
sclerosis (Hofman et al., 1989). These findings
strongly suggest that TNF is involved in various
biologic processes within the CNS.
The inflammatory effects of TNF are pivotal to the
development of cerebral inflammation and edema
during meningitis. High levels of TNF in cerebrospinal
fluid during meningitis are predictive of poor out-
come (Leist et al., 1988; Waage et al., 1989; Saukkonen
et al., 1990). The in vivo inflammatory effects of TNF
have been implicated in the development of the
plaques characteristic of multiple sclerosis, and in
cerebral ischemia in experimental models of stroke,
in which immunologic or pharmacologic inhibition of
TNF activity can reduce the severity of brain damage
(Saukkonen et al., 1990; Meistrell et al., 1997).
 BIOLOGIC EFFECTS 847

TABLE 35.2 Selected list of biological effects of TNF

Immune cells
Mo nocytes/macro p hages
Autocrine induction of TNF
Induction of IL- 1, IL-8,
GM-CSF, M-CSE IFNT,
NGF, TGF[3, DGF and E G 2
Chemotaxis
Stimulation of metabolism
Inhibition of differentiation
Suppression of proliferation
Internalization of
complement-coated virus
Polymorphonuclear leukocytes
Priming of integrin response
Release of granule
components
Increasing phagocytic
capacity
Enhanced production of
superoxide
Increased adherence to
extracellular matrix
Suppression of chemotaxis to
N- formyl- 1-1eucyl- 1-phenylalanine
Suppression of cell surface
expression of sialophlorin CD43
Lymphocytes
Induction ofT-cell colony
formation
Induction of superoxide in B cells
Activation of cytotoxic T-cell
invasiveness
Induction of apoptosis in mature
T cells
Non-immune cells In vivo
Vascular endothelial cells Central nervous system
Modulation of angiogenesis Fever
Increasing permeability Anorexia
Enhanced expression of MHC 1 Altered pituitary hormone
Procoagulant and antifibrinolytic secretion
Increasing permeability of
albumin Cardiovascular
Suppression of proliferation Shock
Rearrangement of cytoskeleton ARDS
Induction of NO synthase, IL-1, Capillary leakage syndrome
IL-3 receptor, G-CSE GM-CSE
ICAM- 1, VCAM- 1, P- and E-selectin, Gastrointestinal
surface antigen, platelet-activating Ischemia
factor, prostacyclin Colitis
Inhibition of integrin B30, Hepatic necrosis
thromomodulin, glutathione, Inhibition of albumin
protein S expression
Decreased catalase activity
Fibroblasts in liver
Induction of proliferation Suppression of HBV gene
Induction of IL-1, 6, IFN~2, expression
leukemia inhibitory factor,
metalloproteinases (MMTs) Metabolic
Suppression of respiratory activity LPL suppression
Inhibition of collagen synthesis, Net protein catabolism
MMT inhibitor Net lipid catabolism
Stress hormone release
Adipocytes Insulin resistance
Enhanced release of free fatty acids
and glycerol Inflammatory
Suppression of lipoprotein lipase Activation of cell cytotoxicity
(LPL) Enhanced NK cell function
Mediation of IL-2 tumor
Endocrine system toxicity
Stimulation of adrenocorticotrophic Protective role in cutaneous
hormone and prolactin leishmaniasis
Inhibition of thyroid-stimulating
hormone
Follicle-stimulating hormone and
growth hormone
Enhancing IL- 1 inhibition of
steroidogenesis

Although high levels of TNF play a pivotal role in
tissue injury, accumulating evidence also indicates
that low levels of TNF play critical roles in the repair
and regeneration of damaged tissue in the CNS
(Tracey et al., 1989). TNF prevents neuronal death
following metabolic excitotoxic insults (Cheng et al.,
1994), and activates CNS tissue repair by inducing
proliferation of astrocytes (Selmaj et al., 1990) and
neuronal progenitor cells (Mehler et al., 1993). The
proliferative effects of TNF may be dependent upon
the production of nerve growth factor (NGF) (Gadient
et al., 1990), which is protective in tissue damage
(Mattson et al., 1993). Mice genetically deficient for
both TNFR1 and TNFR2 are more sensitive to central
ischemic insult and have more extensive cortical
damage (Bruce et al., 1996), suggesting that TNF can
be protective in ischemic brains. This contrasts with
the overexpression of TNF during cerebral ischemia

THE CYTOKINES A N D C H E M O K I N E S
848 T U M O U R NECROSIS FACTOR
in normal rats subjected to focal infarction, where
TNF significantly increases brain damage (Meistrell
et al., 1997).
Cardiovascular system
Endothelial cells serve as a semipermeable barrier in
blood vessels, and mediate coagulation. During sys-
temic inflammation, sepsis, multiple organ failure and
other acute TNF-mediated diseases, TNF enhances
tissue factor expression and suppresses co-factor
activity for the anticoagulant protein C on endothelial
cells (Bevilacqua et al., 1986; Nawroth et al., 1986). TNF
also directly induces actin filament rearrangement,
leading to endothelial cell damage and loss of tight
junctions; this, in turn, causes capillary leakage syn-
drome as plasma proteins and water leak into tissues
(Sato et al., 1986; Stephens et al., 1988; Brett et al.,
1989). TNF exerts a negative inotropic effect on
myocardial contractility through a mechanism that
remains largely unknown. Evidence suggests that TNF
induces the release of platelet-activating factor (PAF)
(Alloatti et al., 1999), as well as stimulation of nitric
oxide (NO) production (Finkel et al., 1992; Habib et al.,
1996; Alloatti et al., 1999). Both epithelial cells and car-
diac myocytes express TNF receptors and are capable
of synthesizing TNF under certain forms of stress
(Giroir et al., 1994; Kapadia et al., 1995).
Elevated levels of TNF have been identified in
patients with advanced heart failure (Moriarty et al.,
1990; Katz et al., 1994). TNF may play a central role in
the pathogenesis of heart failure, and the known bio-
logic effects of TNF may contribute to the develop-
ment of many of the clinical hallmarks of heart failure,
including left ventricular dysfunction, cardiomyopa-
thy, cachexia and pulmonary edema (Millar et al.,
1989; Natanson et al., 1989; Suffredini et al., 1989;
Hegewisch et al., 1990). The negative inotropic effects
of TNF on isolated cardiac myocytes require TNF
concentrations as low as 0.7-1.4 x 10-9 M (Yokoyama
et al., 1993; Kapadia et al., 1995), which have been
reported in congestive heart failure (Levine et al.,
1990).
Direct injection of TNF produces hypotension,
metabolic acidosis, hemaconcentration and death
within minutes, mimicking the cardiovascular
response in endotoxin-induced septic shock (Tracey
et al., 1986a). This is caused by several TNF-mediated
THE CYTOKINES A N D C H E M O K I N E S
effects including a decrease in peripheral vascular
resistance, falling cardiac output and loss of intravas-
cular volume through capillary leakage (Tracey et al.,
1986a, 1987b; Natanson et al., 1989). TNF-induced
NO production in endothelium has been implicated
in decreases of both peripheral vascular tone and
cardiac function (Finkel et al., 1992), whereas
TNF-induced rearrangement of actin filaments and
loss of tight junctions are responsible for the capillary
leakage syndrome (Sato et al., 1986; Stephens et al.,
1988; Brett et al., 1989). TNF is extremely toxic to the
lungs and is pivotal in the development of adult respi-
ratory distress syndrome (ARDS), characterized by
pulmonary edema, hypoxia and a high mortality rate.
ARDS develops because of TNF-induced activation of
pulmonary endothelium, margination of leukocytes
with degranulation of granulocytes and capillary
leakage, which precipitates the collection of edema-
tous fluid in alveoli and prevents adequate perfusion
(Stephens et al., 1988).
Human diseases
Although TNF can preserve cellular and biochemical
homeostasis thereby participating in beneficial tissue
remodeling and host-defense responses, overproduc-
tion of TNF is harmful, even lethal, to the host (Tracey
et al., 1989). Therefore, either insufficient or excessive
production of TNF can be deleterious, and TNF has
been directly implicated in the pathogenesis of many
disease states (Table 35.3).
Septic shock
TNF plays a pivotal role in septic shock, because it ful-
fils Koch's postulates, a logical chain of evidence that
can be modified here for considering cytokine biol-
ogy. First, TNF is released systemically during over-
whelming sepsis, and in some cases leads to lethality
(Girardin et al., 1988; Waage et al., 1989). Persistent
increases in TNF are associated with the development
of multiple organ failure and mortality (Girardin et al.,
1988; Calandra et al., 1990; van der Poll and Lowry,
1995). Second, administration of exogenous TNF
causes shock and tissue injury that is physiologi-
cally, metabolically, hematologically and pathologi-
cally indistinguishable from septic shock syndrome
(Tracey et al., 1986a, 1987a). Third, neutralization of
 BIOLOGIC EFFECTS 849

TABLE 35.3 Selected TNF-associated diseases

Infectious diseases Autoimmune diseases Cancer and others

Bacterial Glomerulonephritis Hairy cell leukemia
Meningococcal disease Guillain-Barr6 syndrome T-cell leukemia
Leprosy Inflammatory bowel disease Chronic lymphocytic leukemia
Tuberculosis Systemic lupus erythematosus Malignant lymphoma
Septic shock syndrome Insulin-dependent diabetes Colorectal cancer
Nocardia brasiliensis mellitus Lung cancer
Cryptogenic fibrosing Rheumatoid arthritis Cervical cancer
alveolitis Dermatitis
Pneurnocystis carinii Celiac disease Euthyroid sick syndrome
pneumonia Myasthenia gravis Hemorrhagic shock
Systemic sclerosis Disseminated intravascular
Viral coagulopathy
AIDS Anemia
HIV-related lung disease Myocardial ischemia
Measles Obesity
Hepatitis Shock
Viral meningitis Stroke
Murine retrovirus infection Rejection of transplanted organs
Pulmonary fibrosis
Parasitic Chronic osteomyelitis
Cerebral malaria Graves' disease
Toxoplasma gondii Asthma
Dysentery Injury
Shigella dysenteriae Inflammatory hyperalgesia
Children with pertussis Down's syndrome
Cachexia

TNF with antibodies prevents septic shock syndrome
during lethal bacteremia (Tracey et al., 1987b). In sup-
port of this view, TNFRl-deficient knockout mice are
resistant to endotoxemic shock (Pfeffer et al., 1993).
TNF induces other mediators of sepsis such as IL-1,
and infusion of anti-TNF antibody attenuates the
activity of these mediators (Tracey et al., 1987b; Fong
et al., 1989).
TNF and soluble receptors ofTNF (sTNFRs) are lib-
erated during sepsis (Girardin et al., 1992). As men-
tioned above, sTNFRs are inhibitors of TNF biologic
activities but may also serve as a reservoir of TNF, and
increased levels of sTNFRs are found in many TNF-
associated diseases. In fact, increased levels of sTN-
FRs correlate with TNF levels and outcome in severe
meningococcemia (Girardin et al., 1992). In general,
sTNFR concentrations are associated with TNF levels
when the TNF level is relatively low, but not when it is
high. The imbalance between TNF and sTNFRs is
probably pathophysiologically important, as reflected
by the observation that their ratio on admission may
be of predictive value for clinical outcome (Girardin
et al., 1992).
Cancer
TNF was originally characterized as an antitumor pro-
tein inducing necrosis of Meth A sarcomas in vivo
(Carswell et al., 1975). TNF has been widely studied
and explored as an antitumor drug; at present it is
used clinically as a locally delivered intratumoral
agent and has some efficacy (Lienard et al., 1992). The
therapeutic application of TNF as a single and sys-
temic antitumor agent is limited by toxic side-effects
at tumoricidal doses (Blick et al., 1987). Several strate-
gies have been proposed to avoid the limitations
imposed by toxicity, including: (1) high-dose intra-
arterial administration of TNF directly into the tumor
compartment (Ohkawa et al., 1989); (2) administra-
tion of non-toxic levels of TNF to tumors sensitive to
its therapeutic effects; (3) administration of non-toxic
doses of TNF in combination with other antitumor

THE CYTOKINES AND CHEMOKINES

850 TUMOUR NECROSIS FACTOR
agents - synergistic effects have been identified
(Zimmerman et al., 1989), but enhanced side-effects
have also been reported (Schiller etal., 1992); (4) iden-
tification and development of TNF-like molecules
with maximum efficacy as an antitumor agent, but
minimal side-effects. In addition, chemical modifica-
tion of TNF with compounds such as polyethylene
glycol (PEG) has been found selectively to increase the
antitumor potency of TNF while effectively reducing
its systemic toxic side-effects (Tsutsumi et al., 1995).
A few mechanisms have been suggested for the
tumoricidal effect of TNE including direct cytotoxicity
against tumor cells, activation of immune antitumor
response and selective damage of tumor blood vessels
(North and Havell, 1988). Direct cytotoxicity to tumor
cells has been supported by most in vitro studies
(Duerksen-Hughes et al., 1992). TNF also activates
other cytokines (Tracey and Cerami, 1994) and cyto-
toxic factors such as NO (Estrada et al., 1992), which
in turn participate in immune antitumor responses
and direct tumor cell killing. A study by Sato et al.
(1996) reported that it might be feasible to use TNF
gene-transduced tumor cells as a vaccine, since
TNF is a powerful activator of the immune response.
TNF selectively eradicates vascularized tumors but is
much less effective in killing avascular implants
(Yoshimura and Sone, 1987; Thomson, 1997). In con-
trast, TNF is produced by some tumors or tumor cells,
and can act as an autocrine growth factor (Sugarman
et al., 1985; Buck et al., 1990).
ASSOCIATED DISEASE
SUSCEPTIBILITY
The TNF locus with two tandemly arranged and
closely linked genes, TNF and lymphotoxin-tz, lies in
the class III region of the MHC on the short arm of
human chromosome 6. A striking feature of the MHC
is the high degree of polymorphism of genes in this
region. At least five polymorphisms (Partanen and
Koskimies, 1988; D'Alfonso and Richiardi, 1994) and
five microsatellites (Jongeneel et al., 1991; Udalova
et al., 1993) have been described at the TNF locus.
These polymorphisms have been linked to the vari-
ability of TNF production in different individuals
(Molvig et al., 1988). Because TNF production has
been implicated as an important factor in immune
THE CYTOKINES AND CHEMOKINES
regulation and the inflammatory response, and
affects the outcome of human diseases, polymor-
phisms within the TNF locus are considered possible
genetic bases for these diseases. Some studies have
indicated that genetic variation of TNF and closely
linked loci are important in determining suscepti-
bility to a significant number of human diseases,
including autoimmune diseases such as diabetes,
rheumatoid arthritis and multiple sclerosis, parasitic,
bacterial and viral infections and cancer.
TNF responsiveness is controlled by variable
genetic elements within the MHC region. A polymor-
phism located at -308 bp upstream of the TNF tran-
scriptional start site, with two allelic forms referred to
as TNF1 and TNF2, has been demonstrated directly
to affect TNF production (Wilson et al., 1993, 1997);
TNF2 is associated with higher constitutive and
inducible transcriptional levels than the TNF1 allele
(Wilson et al., 1994, 1997). There is also a correlation
of human leukocyte antigen (HLA)-DR2 with low TNF
production (Bendtzen et al., 1988; Molvig et al., 1988),
and of HLA-DR3 and HLA-DR4 with high TNF pro-
duction (Jacob et al., 1990; Abraham et al., 1993).
These polymorphisms are associated with the suscep-
tibility or severity of certain diseases. Thus, the TNF
locus may contribute to the pathogenesis of diseases
by influencing the TNF levels produced in those
diseases.
Susceptibility to infectious diseases
MHC alleles affect outcome in several infectious
diseases. TNF2 homozygotes with associated higher
TNF levels are at increased risk of cerebral malaria
(McGuire et al., 1994) or post-traumatic sepsis
(Majetschak et al., 1999). This is consistent with the
phenotypic observation that Gambian children with
cerebral malaria have higher TNF levels than children
with mild malaria (Kwiatkowski et al., 1990), and that
TNF levels are highest in children who die or develop
neurologic sequelae due to cerebral malaria. In
contrast, the HLA-B53 locus, which correlates with
depressed levels of circulating TNF, appears to be pro-
tective against several forms of malaria (Hill, 1992).
The TNFB2 allele, also associated with higher levels of
TNF production, is linked to lower survival in patients
with multiple organ failure secondary to severe sepsis
(Stuber et al., 1996). Susceptibility to mucotaneous
 ASSOCIATED DISEASE SUSCEPTIBILITY
leishmaniasis, which is accompanied by high circulat-
ing TNF levels, is linked directly with the regulatory
polymorphisms affecting TNF production (Cabrera
et al., 1995). In addition, HLA-DQ and HLA-DP alleles
are associated with generalized and localized forms of
onchocerciasis (Meyer et al., 1994). As the under-
standing of allelic regulatory differences improves, it
should be possible to more effectively translate our
basic research advances in TNF mechanisms into
clinically useful methods to prevent the pathologic
sequelae of excessive TNF, and/or to improve the
potential benefits of its controlled local production
(e.g. cancer).
Susceptibility to cancer
Alleles of HLA-A, -B and -DR loci are associated with
resistance to lung cancer. Investigators have located
one such element to the TNF locus and found that
TNFB2 homozygosity is associated with disease
resistance and a better prognosis in patients with lung
cancer (Shimura et al., 1994), in keeping with a previ-
ous study that demonstrated a strong correlation
between HLA-A1 and a lower i-year survival rate
(Markman et al., 1984). As TNFB2 is associated with
higher levels of TNF production, these results are con-
sistent with the original characterization of TNF as an
antitumor factor. It has been reported that germline
allelic frequencies for the TNF locus are significantly
different in patients with colorectal cancer compared
with those in normal controls. Most strikingly, the R3
allele in patients with colorectal cancer has an allelic
frequency of nearly 30% but is not detected in the
normal control group (Campbell et al., 1994). Other
studies have shown that class II haplotypes such as
DRB 1"1501-DQB 1"0602 are positively associated with
the development of cervical cancer, but that DR13-
positive haplotypes are negatively associated (Apple
et al., 1994).
Susceptibility to a u t o i m m u n e diseases
The outcome of a large number of studies suggests
that TNF genetics is an important contributing factor
to autoimmune disease susceptibility. Celiac disease
(CD) is an immune disease triggered by the cereal
antigen gliadin, resulting in villus atrophy in the small
intestine. Susceptibility to the development of celiac
THE CYTOKINES AND CHEMOKINES
85 1
disease is strongly influenced by the genes in MHC,
and is associated with the HLA-A1-B8-DR3-DQ2
haplotype (Ahmed et al., 1993) and more strongly
associated with the DQA*0501/DQB*0201 het-
erodimer (Sollid et al., 1989). Wilson and colleagues
(1993) found that the TNF2 allele in the TNF locus is
closely associated with HLA-A1-B8-DR3-DQ2, sug-
gesting that the association of this haplotype with
celiac disease and high TNF production may be
related to polymorphisms within the TNF locus. At the
same time, susceptibility to celiac disease is strongly
related to TNF2 (Manus et al., 1996), supporting this
speculation. The TNF2 allele appears to be involved
in a variety of MHC-linked diseases including sys-
temic lupus erythematosus, dermatitis herpetiformis
and insulin-dependent diabetes mellitus (IDDM).
Microsatellite polymorphisms (TNFa2 and TNFb3)
close to the TNF genes are associated with celiac dis-
ease, an observation that could have functional signif-
icance because the TNFa2 allele correlates with high
TNF production (McManus et al., 1996).
Rheumatoid arthritis (RA) is a debilitating disease
characterized by progressive joint dysfunction and
chronic systemic inflammation (Pincus et al., 1994).
TNF has been implicated in the pathogenesis of RA,
although its precise mechanisms of action in the dis-
ease remain poorly understood. Genetic analyses
have implicated the TNFcl allele as well as the -238
GG genotype, but not the TNFa allele, in rheumatoid
arthritis (RA) susceptibility (Fabris et al., 2002). Ele-
vated concentrations of TNF have been identified in
the serum and synovial fluid of RA patients (Chu et al.,
1991), and plasma TNF concentrations correlate with
disease severity in terms of joint function and pain
(Beckham et al., 1992). Because TNF induces the
expression of endothelial cell adhesion molecules
(Moser et al., 1989), TNF may recruit inflammatory
cells into affected joints. TNF also induces the expres-
sion of matrix metalloproteinases from fibroblasts
and chondrocytes, and may therefore accelerate joint
degradation and dysfunction in RA (Shingu et al.,
1993). Therapeutic interventions that inhibit the
activity of TNF have proven efficacious for the treat-
ment of both adult and juvenile RA (Maini et al., 1999;
Weinblatt et al., 1999; Lovell et al., 2000).
The role of various TNF alleles as susceptibility fac-
tors for systemic lupus erythematosus (SLE) is cur-
rently under intense investigation. The alleles TNFB 1
852 T U M O O R NECROSIS FACTOR
(Bettinotti et al., 1993) and TNF2 (Wilson et al., 1993)
are associated with systemic lupus erythematosus
(SLE), but the association is not independent of
HLA-DR3 (Welch et al., 1988). The HLA-DR3 and
TNF-308A loci appear to be independent susceptibil-
ity loci for SLE (Rood et al., 2000; van der Linden et al.,
2001), but others challenge this view (Tsuchiya et al.,
2001). In another study (D'Alfonso et al., 1996), 123
patients with SLE and 199 matched controls were
analyzed. Three TNF-238/A homozygous patients
but no homozygous control were detected, suggesting
that the -238/AA genotype is a marker of a particular
clinical subtype; this has recently been confirmed in
a cohort of 51 Mexican SLE patients (Zuniga et al.,
2001). Because TNF has been genetically implicated
as a mediator of SLE, several studies have attempted
to neutralize TNF as a therapeutic approach to SLE.
Although immunologic or pharmacologic inhibition
of TNF production has reduced the clinical manifesta-
tions of experimental SLE in mice (Segal et al., 2001),
aberrant TNF activity may promote humoral auto-
immunity and allow the development of autoreactive
B cells (Via et al., 2001). These findings suggest that,
while abrogation of TNF activity may be therapeuti-
cally beneficial for the treatment of established SLE,
depressed TNF levels may play a causative role in the
development of this autoimmune disease.
TNF has also been implicated in the pathogenesis
of IDDM as a mediator in the immune-mediated
destruction of pancreatic [3 cells (Mandrup-Poulsen
et al., 1987). Studies of TNF genetics in patients with
IDDM show a weak association between TNF2 and
IDDM (Pociot et al., 1993b; Cox et al., 1994). However,
another study demonstrated a higher frequency of the
TNFa2 allele in DR3-DR4 heterozygotic patients
with IDDM than in controls. As this marker is associ-
ated with higher TNF production independently of
class II alleles, the TNF locus may play a direct role in
susceptibility to IDDM (Pociot et al., 1993a).
Anti-TNF therapy
Several strategies have been proposed to prevent TNF
toxicity in sepsis (Tracey and Cerami, 1994). One
approach is direct inhibition of TNF activity with anti-
bodies or native and chimeric sTNFRs (Tracey et al.,
THE CYTOKINES A N D C H E M O K I N E S
1987b; Ashkenazi et al., 1991; Peppel et al., 1991).
Another is to suppress TNF synthesis using glucocor-
ticoids, pentoxifylline, CNI-1493, spermine, thalido-
mide and cytokines such as IL-10 (Beutler et al., 1986;
Lilly et al., 1989; Sampaio et al., 1991; Cohen et al.,
1996; Zhang et al., 1997). The third approach is to
induce TNF tolerance with repetitive administration
of low-dose TNE Inhibitors of receptor-mediated
endocytosis of TNF decrease TNF-induced gene
expression, thus providing yet another potential point
of intervention (Bradley et al., 1993). In addition,
pharmacologic inhibitors of metalloproteinases have
been developed to inhibit TNF processing, although
clinical efficacy has not been addressed (McGeehan
et al., 1994). Finally, TNF cytotoxicity may be inhibited
by targeting its secondary mediators such as IL-1
(McNamara et al., 1993). Some of these therapies have
been demonstrated to have salutory effects in animal
models of sepsis, yet preliminary clinical trials have
not been as encouraging (van der Poll and Lowry,
1995).
Strategies to directly inhibit TNF activity with anti-
bodies or sTNFRs are effective in animal models
(Mohler et al., 1993) (Table 35.2), and have been
tested in clinical trials of Crohn's diseases (Sandborn
and Hanauer, 1999; Shanahan, 2000; Kam and Targan,
2000), rheumatoid arthritis (Taylor, 2001; Schwarz
et al., 2000; Feldmann and Maini, 2001; Alldred, 2001),
and sepsis (Dhainaut et al., 1995; Cronin et al., 1995;
Abraham et al., 1997, 1998). For instance, a chimeric
monoclonal anti-TNF antibody, infliximab, has
shown efficacy in moderately to severely active
Crohn's disease, and has recently been approved
by the U.S. Food and Drug Administration (FDA)
(Sandborn and Hanauer, 1999). A recombinant
human soluble TNF receptor, Etanercept (Enbrel), has
recently been approved by the U.S. FDA for the
treatment of rheumatoid arthritis, which has shown
a statistically significant reduction in swollen and
inflamed joint counts, and a significant improve-
ments in quality of life measures (Schwarz et al., 2000;
Taylor, 2001; Feldmann and Maini, 2001; Alldred,
2001). Although anti-TNF therapy has been success-
fully employed in the treatment of rheumatoid arthri-
tis, a significant survival advantage has not been
observed in the treatment of sepsis.
 REFERENCES
Regulation of TNF by the cholinergic
anti-inflammatory pathway
The cholinergic a n t i - i n f l a m m a t o r y p a t h w a y is a neu-
ral p a t h w a y t h a t rapidly a n d directly a t t e n u a t e s the
p r o d u c t i o n of TNE In r e s p o n s e to e n d o t o x i n or
i s c h e m i a / reperfusion injury, efferent p a r a s y m p a -
thetic signals t h r o u g h the vagus nerve increase h e a r t
rate, stabilize b l o o d pressure, a n d a t t e n u a t e s e r u m
a n d tissue TNF levels (Borovikova et al., 2000a, 2000b;
Bernik e t al., 2001, 2002). Acetylcholine, the p r i m a r y
n e u r o t r a n s m i t t e r of the vagus nerve, b i n d s alpha-
b u n g a r o t o x i n - s e n s i t i v e nicotinic cholinergic recep-
tors e x p r e s s e d on m a c r o p h a g e s to inhibit the
p r o d u c t i o n of p r o i n f l a m m a t o r y mediators, i n c l u d i n g
TNF a n d IL-1 (Borovikova et al., 2000a, 2000b). Elec-
trical s t i m u l a t i o n of the vagus nerve can r e c a p i t u l a t e
or a u g m e n t the i n n a t e central r e s p o n s e to p e r i p h e r a l
i n f l a m m a t i o n (Borovikova et al., 2000a, 2000b; Bernik
et al., 2001, 2002); b e c a u s e vagus nerve s t i m u l a t o r s
are u s e d clinically for the t r e a t m e n t of d e p r e s s i o n a n d
epilepsy (Goodnick et al., 2001; B e n b a d i s a n d Tatum,
2001), vagus nerve s t i m u l a t i o n m a y provide a n e w
a p p r o a c h for the t r e a t m e n t of i n f l a m m a t o r y diseases,
b e c a u s e it inhibits TNF a n d o t h e r p r o i n f l a m m a t o r y
mediators.
REFERENCES
Abraham, E., Glauser, M.P., Butler, T. et al. (1997). p55 Tumor
necrosis factor receptor fusion protein in the treatment
of patients with severe sepsis and septic shock. A ran-
domized controlled multicenter trial. Ro 45-2081 Study
Group. ]AMA 277, 1531-1538.
Abraham, E., Anzueto, A., Gutierrez, G. et al. (1998). Double-
blind randomised controlled trial of monoclonal anti-
body to human tumour necrosis factor in treatment of
septic shock. NORASEPT II Study Group. Lancet 351,
929-933.
Abraham, L.J., French, M.A. and Dawkins, R.L (1993).
Polymorphic MHC ancestral haplotypes affect the activ-
ity of tumour necrosis factor-a. Clin. Exp. I m m u n o l . 92,
14-18.
Adam-Klages, S., Adam, D., Wiegmann, K. et al. (1996). FAN,
a novel WD-repeat protein, couples the p55 TNF-receptor
to neutral sphingomyelinase. Cell 86, 937-947.
Aderka, D. (1996). The potential biological and clinical sig-
nificance of the soluble tumor necrosis factor receptors.
Cytokine Growth Factor Rev. 7, 231-240.
Ahmed, A.R., Yunis, J.J., Marcus-Bagley, D. et al. (1993). Major
histocompatibility complex susceptibility genes for der-
matitis herpetiformis compared with those for gluten-
sensitive enteropathy. ]. Exp. Med. 178, 2067-2075.
THE CYTOKINES AND CHEMOKINES
853
Alessi, D.R., Cuenda, A., Cohen, P. et al. (1995). PD 098059 is
a specific inhibitor of the activation of mitogen-activated
protein kinase kinase in vitro and in vivo. ]. Biol. Chem.
270, 27489-27494.
Alldred, A. (2001). Etanercept in rheumatoid arthritis. Exp.
Opin. Pharmacother. 2, 1137-1148.
Alloatti, G., Penna, C., De Martino, A. et al. (1999). Role of
nitric oxide and platelet-activating factor in cardiac alter-
ations induced by tumor necrosis factor-alpha in the
guinea-pig papillary muscle. Cardiovasc. Res. 41, 611-619.
An, J., Ribeiro, R.C., Webb, P. et al. (1999). Estradiol repression
of tumor necrosis factor-alpha transcription requires
estrogen receptor activation function-2 and is enhanced
by coactivators. Proc. N a t l Acad. Sci. USA 96,
15161-15166.
Apple, R.J., Erlich, H.A., Klitz, W. et al. (1994). HLA DR-DQ
associations with cervical carcinoma show papillo-
mavirus-type specificity. Nat. Genet. 6, 157-162.
Ashkenazi, A., Marsters, S.A., Capon, D.J. et al. (1991).
Protection against endotoxic shock by a tumor necrosis
factor receptor immunoadhesin. Proc. Natl Acad. Sci. USA
88, 10535-10539.
Auphan, N., Di Donato, J.A., Rosette, C. et al. (1995). Immuno-
suppression by glucocorticoids: inhibition of NFKB activ-
ity through induction of I~:-B synthesis. Science 270,
286-290.
Balboa, M.A., Balsinde, J., Aramburu, J. et al. (1992).
Phospholipase D activation in human natural killer cells
through the KP43 and CD16 surface antigens takes place
by different mechanisms. Involvement of the phospho-
lipase D pathway in tumor necrosis factor-~ synthesis.
J. Exp. Med. 176, 9-17.
Baldwin, R.L, Stolowitz, M.L, Hood, L. et al. (1996). Structural
changes of tumor necrosis factor ~ associated with mem-
brane insertion and channel formation. Proc. Natl Acad.
Sci. USA 93, 1021-1026.
Barbara, J.A., Smith, W.B., Gamble, J.R. et al. (1994). Dissocia-
tion of TNF-R cytotoxic and proinflammatory activities
by p55 receptor- and p75 receptor-selective TNF-~
mutants. EMBO ]. 13, 843-650.
Beckham, J.C., Caldwell, D.S., Peterson, B.L. et al. (1992).
Disease severity in rheumatoid arthritis: relationships of
plasma tumor necrosis factor-alpha, soluble interleukin
2-receptor, soluble CD4/CD8 ratio, neopterin, and fibrin
D-dimer to traditional severity and functional measures.
]. Clin. I m m u n o l . 12, 353-361.
Benbadis, S.R. and Tatum, W.O. IV. (2001). Advances in the
treatment of epilepsy. Am. Faro. Physician 64, 91-98.
Bendtzen, K., Morling, N., Fomsgaard, A. et al. (1988).
Association between HLA-DR2 and production of
tumour necrosis factor (~ and interleukin 1 by mono-
nuclear cells activated by lipopolysaccharide. Scand. J.
I m m u n o l . 28, 599-606.
Bernik, T.R., Ivanova, S.M., Ochani M. et al. (2001). Vagus
nerve stimulation attenuates cardiac TNF production in
endotoxic shock. Shock 15 (Supp.), 27.
Bernik, T.R., DiRaimo, R., Friedman, S.G. et al. (2002).
Systemic anti-inflammatory action of CNI-1493 is
mediated by the cholinergic anti-inflammatory pathway.
]. Exp. Med., 18, 781-788.
Bettinotti, M.P., Hartung, K., Deicher. H. et al. (1993).
Polymorphism of the tumor necrosis factor [3gene in sys-
temic lupus erythematosus: TNFB-MHC haplotypes.
I m m u n o g e n e t i c s 37, 449-454.
854 TUMOUR NECROSIS FACTOR
Beutler, B., Greenwald, D., Hulmes, J.D. et al. (1985). Identity
of tumour necrosis factor and the macrophage-secreted
factor cachectin. Nature 316, 552-554.
Beutler, B., Krochin, N., Milsark, I.W. et al. (1986). Control of
cachectin (tumor necrosis factor) synthesis: mechanisms
of endotoxin resistance. Science 232, 977-980.
Bevilacqua, M.P., Pober, J.S., Majeau, G.R. et al. (1986).
Recombinant tumor necrosis factor induces procoagu-
lant activity in cultured human vascular endothelium:
characterization and comparison with the actions of
interleukin 1. Proc. Natl Acad. Sci. USA 83, 4533-4537.
Bianchi, M., Bloom. 0., Raabe, T. et al. (1996). Suppression of
proinflammatory cytokines in monocytes by a tetravalent
guanylhydrazone. L Exp. Med. 183, 927-936.
Biswas, D.K., Ahlers, C.M., Dezube, B.J. et al. (1994).
Pentoxifylline and other protein kinase C inhibitors
down-regulate HIV-LTR NF-•B-induced gene expression.
Mol. Med. 1, 31-43.
Black, R.A., Rauch, C.T., Kozlosky, C.J. et al. (1997). A metal-
loproteinase disintegrin that releases tumour-necrosis
factor-a from cells. Nature 385, 729-733.
Blick, M., Sherwin, S.A., Rosenblum, M. et al. (1987). Phase I
study of recombinant tumor necrosis factor in cancer
patients. Cancer Res. 47, 2986-2989.
Bogdan, C., Paik, J., Vodovotz, Y. et al. (1992). Contrasting
mechanisms for suppression of macrophage cytokine
release by transforming growth factor-~. ]. Biol. Chem.
267, 23301-23308.
Boise, LH. and Thompson, C.B. (1997). Bcl-x(L) can inhibit
apoptosis in cells that have undergone Fas-induced pro-
tease activation. Proc. Natl Acad. Sci. USA 94, 3759-3764.
Boldin, M.E, Goncharov, T.M., Goltsev, Y.V. et al. (1996).
Involvement of mach, a novel MORT1/FADD-interacting
protease, in Fas/Apo-1- and TNF receptor-induced cell
death. Cell 85, 803-815.
Borovikova, L.V., Ivanova, S., Nardi, D. et al. (2000a). Role of
vagus nerve signaling in CNI- 1493-mediated suppression
of acute inflammation. Auton. Neurosci. 85, 141-147.
Borovikova, L.V., Ivanova, S., Zhang, M. et al. (2000b). Vagus
nerve stimulation attenuates the systemic inflammatory
response to endotoxin. Nature 405, 458-462.
Bradley, J.R., Johnson, D.R. and Pober, J.S. (1993). Four differ-
ent classes ofinhibitors of receptor-mediated endocytosis
decrease tumor necrosis factor-induced gene expression
in human endothelial cells. ]. I m m u n o l . 150, 5544-5555.
Brett, J., Gerlach, H., Nawroth, P. et al. (1989). Tumor necro-
sis factor/cachectin increases permeability of endothelial
cell monolayers by a mechanism involving regulatory
G proteins. ]. Exp. Med. 169, 1977-1991.
Brockhaus, M., Schoenfeld. H.J., Schlaeger, E.J. et al. (1990).
Identification of two types of tumor necrosis factor
receptors on human cell lines by monoclonal antibodies.
Proc. Natl Acad. Sci. USA 87.3127-3131.
Bruce, A.]., Boling, W., Kindy, M.S. et al. (1996). Altered neu-
ronal and microglial responses to excitotoxic and
ischemic brain injury in mice lacking TNF receptors. Nat.
Med. 2, 788-794.
Buck, C., Digel, W., Schoniger, W. et al. (1990). Tumor necro-
sis factor-a, but not lymphotoxin, stimulates growth of
tumor cells in hairy cell leukemia. L e u k e m i a 4, 431-434.
Cabrera, M., Shaw, M.A., Sharpies, C. et al. (1995). Poly-
morphism in tumor necrosis factor genes associated
with mucocutaneous leishmaniasis. 1. Exp. Med. 182,
1259-1264.
THE CYTOKINES AND CHEMOKINES
Calandra, T., Baumgartner, J.D., Grau, G.E. et al. (1990).
Prognostic values of tumor necrosis factor/cachectin,
interleukin-1, interferon-a and interferon-7 in the serum
of patients with septic shock. Swiss-Dutch J5 immuno-
globulin study group. J. Infect. Dis. 161,982-987.
Campbell, D.A., Field, M., McArdle, C.S. et al. (1994).
Polymorphism at the tumour necrosis factor locus: a
marker of genetic predisposition to colorectal cancer?
Lancet 343, 293-294 (letter).
Caput, D., Beufler, B., Hartog, K. et al. (1986). Identification
of a common nucleotide sequence in the 3'-untranslated
region of mRNA molecules specifying inflammatory
mediators. Proc. Natl Acad. Sci. USA. 83, 1670-1674.
Carswell, E.A., Old, L.J., Kassel, R.L. et al. (1975). An endo-
toxin-induced serum factor that causes necrosis of
tumors. Proc. Natl Acad. Sci. USA 72, 3666-3670.
Cha, S.S., Kim, J.S., Cho, H.S. et al. (1998). High resolution crys-
tal structure of a human tumor necrosis factor-a mutant
with low systemic toxicity. ]. Biol. Chem. 273, 2153-2160.
Chainy, G.B., Singh, S., Raju, U. et al. (1996). Differential acti-
vation of the nuclear factor-~:B by TNF muteins specific
for the p60 and p80 TNF receptors. ]. I m m u n o l . 157,
2410-2417.
Cheng, B., Christakos. S. and Mattson, M.P. (1994). Tumor
necrosis factors protect neurons against metabolic-
excitotoxic insults and promote maintenance of calcium
homeostasis. N e u r o n 12, 139-153.
Cheng, G. and Baltimore, D. (1996). Tank, a co-inducer with
TRAF2 of TNF- and CD 401-mediated NF~:B activation.
Genes Dev. 10, 963-973.
Chikanza, I.C., Roux-Lombard, P., Dayer, J.M. et al. (1993).
Tumour necrosis factor soluble receptors behave as acute
phase reactants following surgery in patients with
rheumatoid arthritis, chronic osteomyelitis and
osteoarthritis. Clin. Exp. I m m u n o l . 92, 19-22.
Chinnaiyan, A.M., O'Rourke, K., Tewari, M. et al. (1995).
FADD, a novel death domain-containing protein, inter-
acts with the death domain of Fas and initiates apoptosis.
Cell 81,505-512.
Chu, C.Q., Field, M., Feldman, M. et al. (1991). Localization
of tumor necrosis factor-a in synovial tissues and at the
cartilage-pannus junction in patients with rheumatoid
arthritis. Arthritis R h e u m . 34, 1125-1132.
Coeshott, C., Ohnemus, C., Pilyavskaya, A. et al. (1999).
Converting enzyme-independent release of tumor necro-
sis factor alpha and IL-lbeta from a stimulated human
monocytic cell line in the presence of activated neu-
trophils or purified proteinase 3. Proc. NatlAcad. Sci. USA
96, 6261-6266.
Cohen, O., Inbal, B., Kissil, J.L. et al. (1999). DAP-kinase par-
ticipates in TNF-alpha- and Fas-induced apoptosis and
its function requires the death domain. ]. Cell Biol. 146,
141-148.
Cohen, P.S., Nakshatri. H., Dennis, J. et al. (1996). CNI-1493
inhibits monocyte/macrophage tumor necrosis factor by
suppression of translation efficiency. Proc. Natl Acad. Sci.
USA. 93, 3967-3971.
Condorelli, G., Vigliotta, G., Cafieri, A. et al. (1999). PED/
PEA-15: an anti-apoptotic molecule that regulates
FAS/TNFRl-induced apoptosis. Oncogene 18, 4409-4415.
Cox, A., Gonzalez, A.M., Wilson, A.G. et al. (1994). Compara-
tive analysis of the genetic associations of HLA-DR3
and tumour necrosis factor a with h u m a n IDDM.
Diabetologia 37, 500-503.
 REFERENCES
Crawford, E.K., Ensor, J.E., Kalvakolanu, I. et al. (1997). The
role of 3' poly(A) tail metabolism in tumor necrosis
factor-alpha regulation. 1. Biol. Chem. 272, 21120-21127.
Creasey, AA., Doyle, L.V., Reynolds, M.T. et al. (1987).
Biological effects of recombinant human tumor necrosis
factor and its novel muteins on tumor and normal cell
lines. Cancer Res. 47, 145-149.
Cronin, L., Cook, D.J., Carlet, J. et al. (1995). Corticosteroid
treatment for sepsis: a critical appraisal and meta-
analysis of the literature. Crit. Care Med. 23, 1430-1439.
Crowe, P.D., Walter, B.X., Mohler, K.M. et al. (1995). A metal-
loprotease inhibitor blocks shedding of the 80-kD TNF
receptor and TNF processing in T lymphocytes. I. Exp.
Med. 181, 1205-1210.
Cuenda, A., Rouse, J., Doza, Y.N. et al. (1995). SB 203580 is a
specific inhibitor of a MAP kinase homologue which is
stimulated by cellular stresses and interleukin-1. FEBS
Lett. 364, 229-233.
D'Alfonso, S. and Richiardi, RM. (1994). A polymorphic vari-
ation in a putative regulation box of the TNFA promoter
region. Immunogenetics 39, 150-154.
D'Alfonso, S., Colombo, G., Della Bella, S. et al. (1996).
Association between polymorphisms in the TNF region
and systemic lupus erythematosus in the Italian popula-
tion. Tissue Antigens 47, 551-555.
Dean, J.L., Wait, R., Mahtani, K.R. et al. (2001). The 3' untrans-
lated region of tumor necrosis factor alpha mRNA is a tar-
get of the mRNA-stabilizing factor HuR. Mol. Cell Biol. 21,
721-730.
Dekkers, RE., Lauw, F.N., ten Hove, T. et al. (1999). The effect
of a metalloproteinase inhibitor (GI5402) on tumor
necrosis factor-alpha (TNF-alpha) and TNF-alpha recep-
tors during human endotoxemia. Blood 94, 2252-2258.
Denham, W., Yang, J., Wang, H. et al. (2000). Inhibition of p38
mitogen activated kinase attenuates the severity of pan-
creatitis-induced adult respiratory distress syndrome.
Crit. Care Med. 28, 2567-2572.
Dhainaut, J.E, Vincent, J.L., Richard, C. et al. (1995). CDP571,
a humanized antibody to human tumor necrosis factor-
alpha: safety, pharmacokinetics, immune response, and
influence of the antibody on cytoldne concentrations in
patients with septic shock. CPD571 Sepsis Study Group.
Crit. Care Med. 23, 1461-1469.
Di Marco, S., Hel, Z., Lachance, C. et al. (2001). Polymorphism
in the 3'-untranslated region ofTNFalpha mRNA impairs
binding of the post-transcriptional regulatory protein
HuR to TNFalpha mRNA. Nucleic Acids Res. 29, 863-871.
Duan, H. and Dixit, V.M. (1997). RAIDD is a new 'death' adap-
tor molecule. Nature 385, 86-89.
Duerksen-Hughes, RJ., Day, D.B., Laster, S.M. et al. (1992).
Both tumor necrosis factor and nitric oxide participate in
lysis of simian virus 40-transformed cells by activated
macrophages. ]. I m m u n o l . 149, 2114-2122.
Estrada, C., Gomez, C., Martin, C. et al. (1992). Nitric oxide
mediates tumor necrosis factor-a cytotoxicity in endo-
thelial cells. Biochem.Biophys.Res. C o m m u n . 186,475-482.
Fabris, M., Di, RE., D'Elia, A. et al. (2002). Tumor necrosis
factor-alpha gene polymorphism in severe and mild-
moderate rheumatoid arthritis. ]. R h e u m a t o l . 29,
29-33.
Feldmann, M. and Maini, R.N. (2001). Anti-TNF alpha ther-
apy of rheumatoid arthritis: what have we learned? Annu.
Rev. I m m u n o l . 19, 163-196.
Feng, G.J., Goodridge, H.S., Harnett, M.M. et al. (1999).
THE CYTOKINES AND CHEMOKINES
855
Extracellular signal-related kinase (ERK) and p38
mitogen-activated protein (MAP) kinases differentially
regulate the lipopolysaccharide-mediated induction of
inducible nitric oxide synthase and IL- 12 in macrophages:
Leishmania phosphoglycans subvert macrophage IL-12
production by targeting ERK MAP kinase. ]. I m m u n o l . 163,
6403-6412.
Fillit, H., Ding, W.H., Buee, L. et al. (1991). Elevated circulat-
ing tumor necrosis factor levels in Alzheimer's disease.
Neurosci. Lett. 129, 318-320.
Finkel, M.S., Oddis, C.V., Jacob, T.D. et al. (1992). Negative
inotropic effects of cytokines on the heart mediated by
nitric oxide. Science 257, 387-389.
Fong, C.W., Siddiqui, A.H. and Mark, D.E (1995). Characteri-
zation of protein complexes formed on the repressor
elements of the human tumor necrosis factor alpha gene.
]. Interferon Cytokine Res. 15, 887-895.
Fong, Y., Tracey. K.J., Moldawer, L.L. et al. (1989). Antibodies
to cachectin/tumor necrosis factor reduce interleukin 113
and interleukin 6 appearance during lethal bacteremia.
]. Exp. Med. 170, 1627-1633.
Gadient, R.A., Cron, K.C. and Otten, U. (1990). Interleukin- 113
and tumor necrosis factor-a synergistically stimulate
nerve growth factor (NGF) release from cultured rat
astrocytes. Neurosci. Lett. 117, 335-340.
Gatanaga, T., Hwang, C.D., Gatanaga, M. et al. (1991). The
regulation of TNF receptor mRNA synthesis, membrane
expression, and release by PMA- and LPS-stimulated
human monocytic THP-1 cells in vitro. Cell I m m u n o l .
138, 1-10.
Gearing, A.J., Beckett, P., Christodoulou, M. et al. (1994).
Processing of tumour necrosis factor-a precursor by
metalloproteinases. Nature 370, 555-557.
Gearing. A.J., Beckett, R, Christodoulou, M. et al. (1995).
Matrix metalloproteinases and processing of pro-TNF-a.
]. Leukoc. Biol. 57, 774-777.
Girardin, E., Grau, G.E., Dayer, J.M. et al. (1988). Tumor
necrosis factor and interleuldn-1 in the serum of children
with severe infectious purpura. N. Engl. ]. Med. 319,
397-400.
Girardin, E., Roux-Lombard, R, Grau, G.E. et al. (1992).
Imbalance between tumour necrosis factor-a and soluble
TNF receptor concentrations in severe meningococ-
caemia. The J5 Study Group. I m m u n o l o g y 76, 20-23.
Giroir, B.R, Horton, J.W.,White, D.J. et al. (1994). Inhibition of
tumor necrosis factor prevents myocardial dysfunction
during burn shock. Am. ]. Physiol. 267, H118-H124.
Goldfeld, A.E., McCaffrey, RG., Strominger, J.L. et al. (1993).
Identification of a novel cyclosporin-sensitive element in
the human tumor necrosis factor ~ gene promoter. ]. Exp.
Med. 178, 1365-1379.
Goodnick, RJ., Rush, A.J., George, M.S. et al. (2001). Vagus
nerve stimulation in depression. Expert Opin. Pharmaco-
ther. 2, 1061-1063.
Grell, M., Douni, E., Wajant, H. et al. (1995). The transmem-
brane form of tumor necrosis factor is the prime activat-
ing ligand of the 80 kDa tumor necrosis factor receptor.
Cell 83, 793-802.
Grimaldi, L.M., Martino, G.V., Franciotta, D.M. et al. (1991).
Elevated a-tumor necrosis factor levels in spinal fluid
from HW-l-infected patients with central nervous sys-
tem involvement. Ann. Neurol. 29, 21-25.
Grimm, S., Stanger, B.Z. and Leder, P. (1996). RIP and FADD:
two 'death domain'-containing proteins can induce
856 TUMOUR NECROSIS FACTOR
apoptosis by convergent, but dissociable, pathways. Proc.
Natl Acad. Sci. USA 93, 10923-10927.
Gueydan, C., Droogmans, L., Chalon, P. et al. (1999).
Identification of TIAR as a protein binding to the transla-
tional regulatory AU-rich element of tumor necrosis fac-
tor alpha mRNA. J. Biol. Chem. 274, 2322-2326.
Guha, M., O'Connell, M.A., Pawlinski, R. et al. (2001).
Lipopolysaccharide activation of the MEK-ERK1/2 path-
way in human monocytic cells mediates tissue factor
and tumor necrosis factor alpha expression by inducing
Elk-1 phosphorylation and Egr-1 expression. Blood 98,
1429-1439.
Habib, EM., Springall, D.R., Davies, G.J. et al. (1996). Tumour
necrosis factor and inducible nitric oxide synthase in
dilated cardiomyopathy. Lancet. 347, 1151-1155.
Han, J., Thompson, P. and Beutler, B. (1990). Dexamethasone
and pentoxifylline inhibit endotoxin-induced cachectin!
tumor necrosis factor synthesis at separate points in the
signaling pathway. ]. Exp. Med. 172, 391-394.
Hegewisch, S., Weh, H.J. and Hossfeld, D.K. (1990). TNF-
induced cardiomyopathy. Lancet 335, 294-295 (letter).
Hill, A.V. (1992). Malaria resistance genes: a natural selection.
Trans. R. Soc. Trop. Med. Hyg. 86, 225-226, 232.
Hofman, P.M., Hinton, D.R., Johnson, K. et al. (1989). Tumor
necrosis factor identified in multiple sclerosis brain.
]. Exp.,Med. 170, 607-612.
Hohmann, H.P., Remy. R., Brockhaus, M. et al. (1989). Two
different cell types have different major receptors for
human tumor necrosis factor (TNF-a). J. Biol. Chem. 264,
14927-14934.
Hohmann, H.P., Brockhaus, M., Baeuerle, P.A. et al. (1990).
Expression of the types A and B tumor necrosis factor
(TNF) receptors is independently regulated, and both
receptors mediate activation of the transcription factor
NF-~:B. TNF-a is not needed for induction of a biological
effect via TNF receptors. J. Biol. Chem. 265, 22409-22417.
Hsu, H., Xiong, J. and Goeddel, D.V. (1995). The TNF receptor
1-associated protein TRADD signals cell death and NF-
~:B activation. Cell 81,495-504.
Hsu. H., Shu, H.B., Pan, M.G. et al. (1996). TRADD-TRAF2
and TRADD-FADD interactions define two distinct TNF
receptor 1 signal transduction pathways. Cell 84,
299-308.
Jacob, C.O., Fronek, Z., Lewis, G.D. et al. (1990). Heritable
major histocompatibility complex class II-associated
differences in production of tumor necrosis factor ~:
relevance to genetic predisposition to systemic lupus
erythematosus. Proc. Natl Acad. Sci. USA 87, 1233-1237.
Jones, E.Y., Stuart, D.I. and Walker, N.P. (1989). Structure of
tumour necrosis factor. Nature 338, 225-228.
Jongeneel, C.V., Briant, L., Udalova, I.A. etal. (1991). Extensive
genetic polymorphism in the human tumor necrosis fac-
tor region and relation to extended HLA haplotypes. Proc.
Natl Acad. Sci. USA 88, 9717-9721.
Kam, L.Y. and Targan, S.R. (2000). TNF-alpha antagonists
for the treatment of Crohn's disease. Expert Opin.
Pharmacother. 1, 615-622.
Kapadia, S., Torre-Amione, G., Yokoyama, T. et al. (1995).
Soluble TNF binding proteins modulate the negative
inotropic properties of TNF-~ in vitro. Am. ]. Physiol. 268,
H517-H525.
Katz, S.D., Rao, R., Berman, J.W. "et al. (1994). Patho-
physiological correlates of increased serum tumor necro-
sis factor in patients with congestive heart failure.
THE CYTOKINES AND CHEMOKINES
Relation to nitric oxide-dependent vasodilation in the
forearm circulation. Circulation 90, 12-16.
Kohno, T., Brewer, M.T., Baker, S.L. et al. (1990). A second
tumor necrosis factor receptor gene product can shed a
naturally occurring tumor necrosis factor inhibitor. Proc.
Natl Acad. Sci. USA 87, 8331-8335.
Kouba, D.J., Nakano, H., Nishiyama, T. et al. (2001). Tumor
necrosis factor-a induces distinctive NF-KB signaling
within human dermal fibroblasts. ]. Biol. Chem. 276,
6214-6224.
Kramer, B., Meichle, A., Hensel, G. et al. (1994). Characteri-
zation of a KROX-24/EGR-l-responsive element in the
h u m a n tumor necrosis factor promoter. Biochim.
Biophys.Acta 1219, 413-421.
Kramer. B., Wiegmann, K. and Kronke, M. (1995). Regulation
of the human TNF promoter by the transcription factor
Ets. ]. Biol. Chem. 270, 6577-6583.
Kriegler, M., Perez, C., De Fay, K. et al. (1988). A novel form of
TNF/cachectin is a cell surface cytotoxic transmembrane
protein: ramifications for the complex physiology of TNE
Cell 53, 45-53.
Kruys, V., Marinx, O., Shaw, G. et al. (1989). Trans-
lational blockade imposed by cytokine-derived UA-rich
sequences. Science 245, 852-855.
Kwiatkowski, D., Hill, A.V., Sambou, I. et al. (1990). TNF
concentration in fatal cerebral, non-fatal cerebral, and
uncomplicated P l a s m o d i u m f a l c i p a r u m malaria. Lancet
336, 1201-1204.
Lacraz, S., Nicod, L., Galve-de Rocnemonteix, B. et al. (1992).
Suppression of metalloproteinase biosynthesis in human
alveolar macrophages by intefleukin-4. L Clin. Invest. 90,
382-388.
Lacraz, S., Nicod, L.P., Chicheportiche, R. et al. (1995). IL-10
inhibits metalloproteinase and stimulates TIMP-1 pro-
duction in human mononuclear phagocytes. I. Clin.
Invest. 96, 2304-2310.
Lai, W.S., Carballo, E., Strum, J.R. et al. (1999). Evidence that
tristetraprolin binds to AU-rich elements and promotes
the deadenylation and destabilization of tumor necrosis
factor alpha mRNA. Mol. Cell, Biol. 19, 4311-4323.
Lee, E., Vaughan, D.E., Parikh, S.H. et al. (1996). Regulation of
matrix metalloproteinases and plasminogen activator
inhibitor-1 synthesis by plasminogen in cultured human
vascular smooth muscle cells. Circ. Res. 78, 44-49.
Lee. ES., Hagler. J., Chen, Z.J. et al. (1997). Activation of the
bcBa kinase complex by MEKK1, a kinase of the JNK path-
way. Cell 88, 213-222.
Lee, J.C., Laydon, J.T., McDonnell, P.C. et al. (1994). A protein
kinase involved in the regulation of inflammatory
cytokine biosynthesis. Nature 372, 739-746.
Leeuwenberg, J.E, Jeunhomme, T.M. and Buurman, W.A.
(1994). Slow release of soluble TNF receptors by mono-
cytes in vitro. ]. I m m u n o l . 152, 4036-4043.
Leist, T.P., Frei, I.C., Kam-Hansen, S. et al. (1988). Tumor
necrosis factor-a in cerebrospinal fluid during bacterial,
but not viral, meningitis. Evaluation in murine model
infections and in patients. ]. Exp. Med. 167, 1743-1748.
Leitman, D.C., Mackow, E.R., Williams, T. et al. (1992). The
core promoter region of the tumor necrosis factor a gene
confers phorbol ester responsiveness to upstream tran-
scriptional activators. Mol. Cell Biol. 12, 1352-1356.
Levine, B., Kalman, J., Mayer, L. et al. (1990). Elevated circu-
lating levels of tumor necrosis factor in severe chronic
heart failure. N. Engl]. Med. 323, 236-241.
 REFERENCES
Lienard, D., Ewalenko. P., Delmotte. J.J. et al. (1992).
High-dose recombinant tumor necrosis factor r in com-
bination with interferon 7 and melphalan in isolation
perfusion of the limbs for melanoma and sarcoma.
]. Clin. Oncol. 10, 52-60.
Lilly, C.M., Sandhu, J.S., Ishizaka, A. et al. (1989). Pento-
xifylline prevents tumor necrosis factor-induced lung
injury. Am. Rev. Respir. Dis. 139, 1361-1368.
Lin, L.L., Wartmann, M., Lin, A.Y. etal. (1993). CPLA2 is phos-
phorylated and activated by MAP kinase. Cell 72, 269-278.
Lin, L.S. (1992). In: Beutler, B (ed.) Tumor Necrosis Factor:
The Molecules a n d Their Emerging Role in Medicine. New
York: Raven Press, pp. 33-48.
Lin, R.H., Hwang, Y.W.,Yang, B.C. et al. (1997). TNF receptor-
2-triggered apoptosis is associated with the down-
regulation of Bcl-xL on activated T cells and can be
prevented by CD28 costimulation. ]. I m m u n o l . 158,
598-603.
Liu, T., Clark, R.K., McDonnell, PC. et al. (1994). Tumor
necrosis factor-alpha expression in ischemic neurons.
Stroke 25, 1481-1488.
Liu, Z.G., Hsu, H., Goeddel, D.V. et al. (1996). Dissection of
TNF receptor 1 effector functions: JNK activation is not
linked to apoptosis while NF-~:B activation prevents cell
death. Cell 87, 565-576.
Loetscher, H., Stueber, O., Banner, D. et al. (1993). Human
tumor necrosis factor ~ (TNF ~) mutants with exclusive
specificity for the 55-kDa or 75-kDa TNF receptors. ]. Biol.
Chem. 268, 26350-26357.
Lovell, D.J., Giannini, E.H., Reiff, A. et al. (2000). Etanercept
in children with plyarticular juvenile rheumatoid arthri-
tis. N. Engl ]. Med. 342, 763-769.
Machleidt, T., Kramer, B., Adam, D. et al. (1996). Function of
the p55 tumor necrosis factor receptor 'death domain'
mediated by phosphatidylcholine-specific phospholi-
pase C. ]. Exp. Med. 184, 725-733.
Mackay, E, Rothe, J., Bluethmann, H. et al. (1994).
Differential responses of fibroblasts from wild-type and
TNF-r55-deficient mice to mouse and h u m a n TNF-~
activation. ]. I m m u n o l . 153, 5274-5284.
Maini, R., St. Clair, E.W., Breeveld, F. et al. (1999). Infliximab
(chimeric anti-tumour necrosis factor-~ monoclonal
antibody) versus placebo in rheumatoid arthritis patients
receiving concomitant methotrexate: a randomised
phase III trial. Lancet 354, 1932-1939.
Majetschak, M., Flohe, S., Obertacke, U. et al. (1999).
Relation of a TNF gene polymorphism to severe sepsis
in trauma patients. Ann. Surg. 230, 207-214.
Malinin, N.L, Boldin, M.P., Kovalenko, A.V. et al. (1997).
MAP3K-related kinase involved in NF-KB induction by
TNF, CD95 and IL-1. Nature 385, 540-544.
Mallett, S. and Barclay, A.N. (1991). A new superfamily of cell
surface proteins related to the nerve growth factor recep-
tor. I m m u n o l . Today 12, 220-223.
Mandrup-Poulsen, T., Bendtzen, K., Dinarello, C.A. et al.
(1987). Human tumor necrosis factor potentiates h u m a n
interleukin 1-mediated rat pancreatic [3-cell cytotoxicity.
]. I m m u n o l . 139, 4077-4082.
Manus, R.M., Wilson, A.G., Mansfield, J. et al. (1996). TNF2, a
polymorphism of the tumour necrosis-a gene promoter,
is a c o m p o n e n t of the celiac disease major histo-
compatibility complex haplotype. Eur. ]. I m m u n o l . 26,
2113-2118.
Markman, M., Braine, H.G. and Abeloff, M.D. (1984).
THE CYTOKINES AND CHEMOKINES
857
Histocompatibility antigens in small cell carcinoma of
the lung. Cancer 54, 2943-2945.
Marx, J. (1995). How the glucocorticoids suppress immunity.
Science 270, 232-233.
Mattson, M.P., Zhang, Y. and Bose, S. (1993). Growth factors
prevent mitochondrial dysfunction, loss of calcium
homeostasis, and cell injury, but not ATP depletion in
hippocampal neurons deprived of glucose. Exp. Neurol.
121, 1-13.
McGeehan, G.M., Becherer, J.D., Bast, R.C., Jr. et al.
(1994). Regulation of tumour necrosis factor-R process-
ing by a metalloproteinase inhibitor. N a t u r e 370,
558-561.
McGuire, W., Hill, A.V., Allsopp, C.E. et al. (1994). Variation in
the TNF-a promoter region associated with susceptibility
to cerebral malaria. Nature 371,508-510.
McManus, R., Moloney, M., Borton, M. et al. (1996). Associa-
tion of celiac disease with microsatellite polymorphisms
close to the tumor necrosis factor genes. H u m . I m m u n o l .
45, 24-31.
McNamara, M.J., Norton, J.A., Nauta, R.J. et al. (1993).
Interleukin- 1 receptor antibody (IL-1RAB) protection
and treatment against lethal endotoxemia in mice. ]. Surg.
Res. 54, 316-321.
Mehler, M.F., Rozental., R., Dougherty, M. et al. (1993).
Cytokine regulation of neuronal differentiation of hip-
pocampal progenitor cells. Nature 362, 62-65.
Meistrell, M., III, Botchkina. G.I., Wang, H. et al. (1997). TNF
is a brain-damaging cytokine in cerebral ischemia. Shock
8, 341-348.
Meyer, C.G., Gallin, M., Erttmann, K.D. et al. (1994). HLA-D
alleles associated with generalized disease, localized dis-
ease, and putative immunity in Onchocerca volvulus
infection. Proc. Natl Acad. Sci. USA 91, 7515-7519.
Millar, A.B., Foley, N.M., Singer, M. et al. (1989). Tumour
necrosis factor in b r o n c h o p u l m o n a r y secretions of
patients with adult respiratory distress syndrome. Lancet
ii, 712-714.
Mohler, K.M., Torrance, D.S., Smith, C.A. et al. (1993). Soluble
tumor necrosis factor (TNF) receptors are effective thera-
peutic agents in lethal endotoxemia and function simul-
taneously as both TNF carriers and TNF antagonists.
]. I m m u n o l . 151, 1548-1561.
MoMg, J., Baek, L., Christensen, P. et al. (1988). Endotoxin-
stimulated h u m a n monocyte secretion of interleukin 1,
tumour necrosis factor ~, and prostaglandin E2 shows
stable interindividual differences. Scand. ]. I m m u n o l . 27,
705-716.
Moriarty, T.M., Padrell. E., Carty, D.J. et al. (1990). GO protein
as signal transducer in the pertussis toxin-sensitive phos-
phatidylinositol pathway. Nature 343, 79-82.
Moser, R.B., Schleiffenbaum, B., Groscurth, P. et al. (1989).
Interleukin 1 and tumor necrosis factor stimulate human
vascular endothelial cells to promote transendothelial
neutrophil passage. ]. Clin. Invest. 83, 444-455.
Moss, M.L., Jin, S.L., Milla, M.E. et al. (1997). Cloning of a
disintegrin metalloproteinase that processes precursor
tumour-necrosis factor-~. Nature 385, 733-736.
Mullberg, J., Durie, F.H., Otten-Evans, C. et al. (1995). A
metalloprotease inhibitor blocks shedding of the IL-6
receptor and the p60 TNF receptor. ]. I m m u n o l . 155,
5198-5205.
Mtiller, G., Ayoub, M., Storz, P. et al. (1995). PKC zeta is a
molecular switch in signal transduction of TNF-alpha,
858 TUMOUR NECROSIS FACTOR
bifunctionally regulated by ceramide and arachidonic
acid. EMBO]. 14, 1961-1969.
Muzio, M., Chinnaiyan, A.M., Kischkel, F.C. et al. (1996).
FLICE, a novel FADD-homologous ICE/CED-3-1ike pro-
tease, is recruited to the CD95 (Fas/APO-1) death-
inducing signaling complex. Cell 85, 817-827.
Narhi, L.O., Philo, J.S., Li, T. et al. (1996). Induction of
a-helix in the ~-sheet protein tumor necrosis factor-a:
acid-induced denaturation. Biochemistry 35,
11454-11460.
Natanson, C., Eichenholz, P.W., Danner, R.L. et al. (1989).
Endotoxin and tumor necrosis factor challenges in dogs
simulate the cardiovascular profile of human septic
shock. ]. Exp. Med. 169, 823-832.
Natoli, G., Costanzo, A., Ianni, A. et al. (1997). Activation of
SAPK/JNK by TNF receptor 1 through a noncytotoxic
TRAF2-dependent pathway. Science 275, 200-203.
Nawroth, PP., Bank, I., Handley, D. et al. (1986). Tumor necro-
sis factor/cachectin interacts with endothelial cell recep-
tors to induce release of interleukin 1. ]. Exp. Med. 163,
1363-1375.
North, R.J. and Havell, E.A. (1988). The antitumor function of
tumor necrosis factor (TNF). II. Analysis of the role of
endogenous TNF in endotoxin-induced hemorrhagic
necrosis and regression of an established sarcoma. 1. Exp.
Med. 167, 1086-1099.
Ohkawa, S., Wright, K.C., Mahajan, H. et al. (1989). Hepatic
arterial infusion of human recombinant tumor necrosis
factor-a. An experimental study in dogs. Cancer 63,
2096-2102.
Panagakos, ES., O'Boskey, J.E, Jr. and Rodriguez, E. (1996).
Regulation of pulp cell matrix metalloproteinase produc-
tion by cytokines and lipopolysaccharides. 1. Endod. 22,
358-361.
Partanen. J. and Koskimies. S. (1988). Low degree of DNA
polymorphism in the HIA-linked lymphotoxin (tumour
necrosis factor 13) gene. Scand. 1. I m m u n o l . 28,
313-316.
Pena, L.A., Fuks, Z. and Kolesnick, R. (1997). Stress-induced
apoptosis and the sphingomyelin pathway. Biochem.
Pharmacol. 53, 615-621.
Peppel, K., Crawford, D. and Beutler, B. (1991). A tumor
necrosis factor (TNF) receptor-IGG heavy chain chimeric
protein as a bivalent antagonist of TNF activity. 1. Exp.
Med. 174, 1483-1489.
Pfeffer, K., Matsuyama, T., Kundig, T.M. et al. (1993). Mice
deficient for the 55 kD tumor necrosis factor receptor are
resistant to endotoxic shock, yet succumb to L. monocy-
togenes infection. Cell 73, 457-467.
Piecyk, M., Wax, S., Beck, A.R. et al. (2000). TIA-1 is a transla-
tional silencer that selectively regulates the expression of
TNF-alpha. EMBO ]. 19, 4154-4163.
Pincus, T., Brooks, R.H. and Callahan, L.E (1994). Prediction
of long-term mortality inpatients with rheumatoid
arthritis according to simple questionnaire and joint
count measures. Ann. Intern. Med. 120, 26-34.
Plata-Salaman, C.R. (1991). Immunoregulators in the nerv-
ous system. Neurosci. Biobehav. Rev. 15, 185-215.
Pociot, E, Briant, L, Jongeneel, C.V. et al. (1993a). Association
of tumor necrosis factor (TNF) and class II major histo-
compatibility complex alleles with the secretion of TNF-a
and TNF-[3 by human mononuclear cells: a possible link
to insulin-dependent diabetes mellitus. Eur. 1. I m m u n o l .
23, 224-231.
THE CYTOKINES AND CHEMOKINES
Pociot, E, Wilson, A.G., Nerup, J. et al. (1993b). No independ-
ent association between a tumor necrosis factor-a pro-
moter region polymorphism and insulin-dependent
diabetes mellitus. Eur. ]. I m m u n o l . 23, 3050-3053.
Rood, M.J., van Krugten, M.V., Zanelli, E. et al. (2000). TNF-
308A and HLA-DR3 alleles contribute independently to
susceptibility to systemic lupus erythematosus. Arthritis
Rheum. 43, 129-134.
Rothe, M., Sarma, V., Dixit, V.M. et al. (1995). TRAF2-
mediated activation of NF-KB by TNF receptor 2 and
CD40. Science 269, 1424-1427.
Sampaio, E.E, Sarno, E.N., Galilly, R. et al. (1991). Thalido-
mide selectively inhibits tumor necrosis factor a produc-
tion by stimulated human monocytes. ]. Exp. Med. 173,
699-703.
Sandborn, W.J. and Hanauer, S.B. (1999). Antitumor necrosis
factor therapy for inflammatory bowel disease: a review
of agents, pharmacology, clinical results, and safety.
Inflamm. Bowel Dis. 5, 119-133.
Sato, N., Goto, T., Haranaka, K. et al. (1986). Actions of tumor
necrosis factor on cultured vascular endothelial cells:
morphologic modulation, growth inhibition, and cyto-
toxicity. ]. Natl Cancer Inst. 76, 1113-1121.
Sato, Y., Koshita, Y., Hirayama, M. et al. (1996). Augmented
antitumor effects of killer cells induced by tumor necro-
sis factor gene-transduced autologous tumor cells from
gastrointestinal cancer patients. Hum. Gene Ther. 7,
1895-1905.
Saukkonen, K., Sande, S., Cioffe, C. et al. (1990). The role of
cytokines in the generation of inflammation and tis-
sue damage in experimental Gram-positive meningitis.
]. Exp. Med. 171,439-448.
Scheinman, R.I., Cogswell, EC., Lofquist, A.K. et al. (1995).
Role of transcriptional activation of IKB a in mediation
of immunosuppression by glucocorticoids. Science 270,
283-286.
Schiller, J.H., Win, EL. Storer, B. et al. (1992). Clinical and bio-
logic effects of combination therapy with 7-interferon
and tumor necrosis factor. Cancer69, 562-571.
Schutze, S., Potthoff, K., Machleidt, T. et al. (1992). TNF acti-
vates NF-KB by phosphatidylcholine-specific phospholi-
pase C-induced 'acidic' sphingomyelin breakdown. Cell
71,765-776.
Schwarz, E.M., Looney, R.J. and O'Keefe, R.J. (2000). Anti-
TNF-alpha therapy as a clinical intervention for peripros-
thetic osteolysis. Arthritis Res. 2, 165-168.
Seckinger, P., Isaaz, S. and Dayer, J.M. (1989). Purification
and biologic characterization of a specific tumor necrosis
factor a inhibitor. ]. Biol. Chem. 264, 11966-11973.
Segal, R., Dayan, M., Zinger, H. et al. (2001). Suppres-
sion of experimental systemic lupus erythematosus
(SLE) in mice via TNF inhibition by an anti-TNFa
monoclonal antibody and by pentoxiphylline. Lupus 10,
23-31.
S6gui, B., Bezombes, C., Uro-Coste, E. et al. (2000). Stress-
induced apoptosis is not mediated by endolysosomal
ceramide. FASEB ]. 14, 36-47.
Selmaj, K.W., Farooq, M., Norton, W.T. et al. (1990). Prolifera-
tion of astrocytes in vitro in response to cytokines. A pri-
mary role for tumor necrosis factor. ]. I m m u n o l . 144,
129-135.
Shanahan, E (2000). Anti-TNF therapy for Crohn's disease: a
perspective (infliximab is not the drug we have been
waiting for). Inflamm. Bowel Dis. 6, 137-139.
 REFERENCES
Shaw, G. and Kamen, R. (1986). A conserved AU sequence
from the 3' untranslated region of GM-CSF mRNA medi-
ates selective mRNA degradation. Cell 46, 659-667.
Shimura, T., Hagihara, M., Takebe, K. et al. (1994). The study
of tumor necrosis factor [3 gene polyrnorphism in lung
cancer patients. Cancer 73, 1184-1188.
Shingu, M., Nagai, Y., Isayama, T. et al. (1993). The effects of
cytokines on metalloproteinase inhibitors (TIMP) and
collagenase production by h u m a n chondrocytes and
TIMP production by synovial cells and endothelial cells.
Clin. Exp. I m m u n o l . 94, 145-149.
Singh, I.S., He, J.R., Calderwood, S. et al. (2001). A high affin-
ity HSF- 1 binding site in the 5'-untranslated region of the
murine tumor necrosis factor- (alpha) gene is a transcrip-
tional repressor. 1. Biol. Chem. 15, 4981-4988.
Silver, I.A., Murrills, R.J. and Etherington, D.J. (1988). Micro-
electrode studies on the acid microenvironment beneath
adherent macrophages and osteoclasts. Exp. Cell Res.
175, 266-276.
Smith, R.A. and Baglioni, C. (1987). The active form of tumor
necrosis factor is a trimer. 1. Biol. Chem. 262, 6951-6954.
Smith, R.A. and Baglioni, C. (1992). Characterization of TNF
receptors. I m m u n o l . Ser. 56, 131-147.
Sollid, L.M., Markussen, G., Ek, J. et al. (1989). Evidence for
a primary association of celiac disease to a particular
HLA-DQ~/[3 heterodimer. 1. Exp. Med. 169, 345-350.
Spence, M.W. (1993). Sphingomyelinases. Adv. Lipid Res. 26,
3-23.
Stanger, B.Z., Leder, P., Lee, T.H. etal. (1995). RIP: a novel pro-
tein containing a death domain that interacts with
FAS/APO-1 (CD95) in yeast and causes cell death. Cell81,
513-523.
Stephens, K.E., Ishizaka, A., Wu, Z.H. et al. (1988).
Granulocyte depletion prevents tumor necrosis factor-
mediated acute lung injury in guinea pigs. Am. Rev.
Respir. Dis. 138, 1300-1307.
Stuber, E, Petersen, M., Bokelmann, E et al. (1996). A
genomic polymorphism within the tumor necrosis factor
locus influences plasma tumor necrosis factor-a concen-
trations and outcome of patients with severe sepsis. Crit.
Care Med. 24, 381-384.
Suffredini, A.E, Fromm, R.E., Parker, M.M. et al. (1989). The
cardiovascular response of normal h u m a n s to the
administration of endotoxin. N. Engl 1. Med. 321,
280-287.
Sugarman, B.J., Aggarwal., B.B., Hass, P.E. et al. (1985).
Recombinant human tumor necrosis factor-a: effects on
proliferation of normal and transformed cells in vitro.
Science 230, 943-945.
Tartaglia, L.A., Weber, R.F., Figari, I.S. et al. (1991). The two
different receptors for tumor necrosis factor mediate
distinct cellular responses. Proc. Natl Acad. Sci. USA 88,
9292-9296.
Tartaglia, L.A., Ayres, T.M., Wong, G.H. et al. (1993a). A novel
domain within the 55 kD TNF receptor signals cell death.
Cell 74, 845-853.
Tartaglia, L.A., Pennica, D. and Goeddel, D.V. (1993b). Ligand
passing: the 75-kDA tumor necrosis factor (TNF) receptor
recruits TNF for signaling by the 55-kDa TNF receptor.
]. Biol. Chem. 268, 18542-18548.
Taylor, PC. (2001). Anti-tumor necrosis factor therapies.
Curr. Opin. Rheumatol. 13, 164-169.
Thomson, A.M. (1997). Neuroscience. More than just fre-
quency detectors? Science275, 179-180.
THE CYTOKINES AND CHEMOKINES
859
Tracey, K.J. (1998). Suppression of TNF and other proinflam-
matory cytokines by the tetravalent guanylhydrazone
CNI-1493. Prog. Clin. Biol. Res. 397, 335-343.
Tracey, K.J. and Cerami, A. (1990). Metabolic responses to
cachectin/TNE A brief review. Ann. N Y Acad. Sci. 587,
325-331.
Tracey, K.J. and Cerami, A. (1994). Tumor necrosis factor: a
pleiotropic cytokine and therapeutic target. Annu. Rev.
Med. 45, 491-503.
Tracey, K.J., Beutler, B., Lowry, S.E et al. (1986a). Shock and
tissue injury induced by recombinant h u m a n cachectin.
Science 234, 470-474.
Tracey K.J., Lowry S.E, Beutler B. et al. (1986b). Cachectin/
tumor necrosis factor mediates changes of skeletal
muscle plasma membrane potential. 1. Exp. Med. 164,
1368-1373.
Tracey, K.J., Fong, Y., Hesse, D.G. et al. (1987a). Anti-
cachectin/TNF monoclonal antibodies prevent septic
shock during lethal bacteraemia. Nature 330, 662-664.
Tracey, K.J., Lowry. S.E, Fahey, T.J., III. et al. (1987b).
Cachectin/tumor necrosis factor induces lethal shock
and stress hormone respenses in the dog. Surg. Gynecol.
Obstet. 164, 415-422.
Tracey, K.J., Wei, H., Manogue, K.R. et al. (1988). Cachectin/
tumor necrosis factor induces cachexia, anemia, and
infammation. ]. Fxp. Med. 167, 1211-1227.
Tracey, K.J., Vlassara, H. and Cerami, A. (1989). Cachectin/
tumour necrosis factor. Lancet i, 1122-1126.
Tsai, E.Y., Yie, J., Thanos, D. et al. (1996). Cell-type-specific
regulation of the h u m a n tumor necrosis factor-a gene in
B cells and T cells by NFATP and ATF-2/Jun. Mol. Cell BioL
16, 5232-5244.
Tsuchiya, N., Kawasaki, A., Tsao, B.P. et al. (2001). Analysis of
tbe association of HLA-DRB1, TNFalpha promoter and
TNFR2 (TNFRSF1B) polymorphisms with SLE using
transmission disequilibrium test. Genes I m m u n . 2,
317-322.
Tsutsumi, Y., Kihira, T., Tsunoda, S. et al. (1995). Molecular
design of hybrid tumour necrosis factor-a with polyethyl-
ene glycol increases its anti-tumour potency. Br. I. Cancer
71,963-968.
Udalova, I.A., Nedospasov, S.A., Webb, G.C. et al. (1993).
Highly informative typing of the h u m a n TNF locus
using six adjacent polymorphic markers. Genomics 16,
180-186.
Van Antwerp, D.J., Martin, S.J., Kafri, T. et al. (1996).
Suppression of TNF-a-induced apoptosis by NF-KB.
Science 274, 787-789.
van der Goot, EG., Pugin, J., Hribar, M. et al. (1999). Mem-
brane interaction of TNF is not sufficient to trigger
increase in membrane conductance in mammalian cells.
FEBS Lett. 460, 107-111.
van der Linden, M.W., van der Slik, A.R., Zanelli, E. et al.
(2001). Six microsatellite markers on the short arm of
chromosome 6 in relation to HLA-DR3 and TNF-308A in
systemic lupus erythematosus. Genes I m m u n . 2, 373-380.
van der Poll, T. and Lowry, S.E (1995). Tumor necrosis factor
in sepsis: mediator of multiple organ failure or essential
part of host defense? Shock 3, 1-12 (editorial).
Van Ostade, X., Vandenabeele, P., Everaerdt, B. et al. (1993).
Human TNF mutants with selective activity on the p55
receptor. Nature 361,266-269.
Van Zee, K.J., Stackpole, S.A., Montegut, W.J. et al. (1994). A
human tumor necrosis factor (TNF) a mutant that binds
860 TUMOUR NECROSIS FACTOR
exclusively to the p55 TNF receptor produces toxicity in
the baboon. ]. Exp. Med. 179, 1185-1191.
Via, C.S., Shustov, A., Rus, V. et al. (2001). In vivo neutraliza-
tion of TNF-alpha promotes humoral autoimmunity by
preventing the induction of CTL. 1. I m m u n o l . 167,
6821-6826.
Waage, A., Brandtzaeg, P., Halstensen, A. et al. (1989). The
complex pattern of cytokines in serum from patients with
meningococcal septic shock. Association between inter-
leukin 6, interleukin 1. and fatal outcome. ]. Exp. Med.
169, 333-338.
Wang, C.Y., Mayo, M.W. and Baldwin, A.S., Jr. (1996). TNF-
and cancer therapy-induced apoptosis: potentiation by
inhibition of NF-KB. Science 274, 784-787.
Weinblatt, M.E., Kremer, J.M., Bankhurst, A.D. et al. (1999). A
trial of etanercept, a recombinant tumor necrosis fac-
tor:Fc fusion protein, in patients with rheumatoid arthri-
tis receiving methotrexate. N. Engl]. Med. 340, 253-259.
Welch, T.R., Beischel, L.S., Balakrishnan, K. et al. (1988).
Major histocompatibility complex extended haplotypes
in systemic lupus erythematosus. Dis. Markers 6,
247-255.
Wiegmann, K., Schutze, S., Machleidt, T. et al. (1994).
Functional dichotomy of neutral and acidic sphin-
gomyelinases in tumor necrosis factor signaling. Cell 78,
1005-1015.
Wilson, A.G., de Vries, N., Pociot, E et al. (1993). An allelic
polymorphism within the human tumor necrosis factor-
promoter region is strongly associated with HLA A1, B8,
and DR3 alleles. ]. Exp. Med. 177, 557-560.
Wilson, A.G., Gordon. C., Di Giovine, ES. etal. (1994). A genetic
association between systemic lupus erythematosus and
tumor necrosis factor-a. Eur. ]. I m m u n o l . 24, 191-195.
Wilson, A.G., Symons, J.A., McDowell, T.L. et al. (1997).
Effects of a polymorphism in the human tumor necrosis
factor-a promoter on transcriptional activation. Proc.
Natl Acad. Sci. USA 94, 3195-3199.
Winston, B.W. and Riches, D.X. (1995). Activation of
p42MAPK/ERK2 following engagement of tumor necrosis
factor receptor CD120A (p55) in mouse macrophages.
]. I m m u n o l . 155, 1525-1533.
Wong, G.H. and Goeddel, D.V. (1988). Induction of
THE CYTOKINES AND CHEMOKINES
manganous superoxide dismutase by tumor necrosis
factor: possible protective mechanism. Science 242,
941-944.
Xia, Z., Dickens, M., Raingeaud, J. et al. (1995). Opposing
effects of ERK and JNK-p38 MAP kinases on apoptosis.
Science 270, 1326-1331.
Yokoyama, T., Vaca, L., Rossen, R.D. et al. (1993). Cellular
basis for the negative inotropic effects of tumor necrosis
factor-a in the adult mammalian heart. ]. Clin. Invest. 92,
2303-2312.
Yoshimura, T and Sone, S. (1987). Different and synergistic
actions of human tumor necrosis factor and interferon-7
in damage of liposome membranes. J. Biol. Chem. 262,
4597-4601.
Young, P., McDonnell, P., Laydon, J. et al. (1994). A novel MAP
kinase regulates the production of IL-1 and TNF in LPS
activated human monocytes. Cytokine 6, 564.
Zhang, M., Caragine, T., Wang, K. et al. (1997). Spermine
inhibits proinflammatory cytokine synthesis in human
mononuclear cells. J. Exp. Med. 185, 1759-1768.
Zhang, X.M., Weber, I. and Chen, M.J. (1992). Site-directed
mutational analysis of human tumor necrosis factor-a
receptor binding site and structure-functional relation-
ship. ]. Biol. Chem. 267, 24069-24075.
Zheng, L., Fisher, G., Miller, R.E. et al. (1995). Induction of
apoptosis in mature T cells by tumour necrosis factor.
Nature 377, 348-351.
Zhu, W., Downey, J.S., Gu, J. et al. (2000). Regulation of TNF
expression by multiple mitogen-activated protein kinase
pathways. J. I m m u n o l . 164, 6349-6358.
Zimmerman, R.J., Chan, A. and Leadon, S.A. (1989).
Oxidative damage in murine tumor cells treated in vitro
by recombinant human tumor necrosis factor. Cancer
Res. 49, 1644-1648.
Zubiaga, A.M., Belasco, J.G. and Greenberg, M.E. (1995). The
nonamer UUAUUUAUU is the key AU-rich sequence
motif that mediates mRNA degradation. Mol. Cell Biol.
15, 2219-2230.
Zuniga, J., Vargas-Alarcon, G., Hernandez-Pacheco, G. et al.
(2001). Tumor necrosis factor-alpha promoter polymor-
phisms in Mexican patients with systemic lupus erythe-
matosus (SLE). Genes I m m u n . 2, 363-366.