14 Semba RD, Graham NMH, Caiaffa WT, et al. Increased mortality 22 Campos FACS, Flores H, Underwood BA.
s H, Underwood BA. Effect of an infection on
associated with vitamin A deficiency during human immunodeficiency
virus type 1 infection. Arch Intern Med 1993; 153: 2149-54.
15 Dushimimana A, Graham NMH, Humphrey JH, et al. Maternal
vitamin A levels and HIV-related birth outcome in Rwanda. VIII
Int Conf on AIDS, Amsterdam, July 1992. Abstr.
16 Tang A, Graham NMH, Kirby J, et al. Dietary micronutrient intake
and risk of progression to AIDS in HIV-1 infected homosexual men.
Am J Epidemiol 1993; 138: 937-51.
17 Bray GA. Definition, measurement, and classification of the
syndromes of obesity. Int J Obesity 1978; 2: 99-112.
18 Miotti PG, Liomba G, Dallabetta G, et al. T lymphocyte subsets
during and after pregnancy: analysis in HIV-1 infected and uninfected
Malawian mothers. J Infect Dis 1992; 165: 1116-19.
19 Lepage P, Van de Perre P, Msellati P, et al. Mother-to-child
transmission of HIV-1 and its determinants: a cohort study in Kigali,
Rwanda. Am J Epidemiol 1993; 137: 589-99.
20 Dabis F, Msellati P, Dunn D, et al. Estimating the rate of mother-to-
child transmission of HIV. Report of a workshop on methodological
issues. Ghent (Belgium), 17-20 Feb 1992. AIDS 1993; 7: 1139-48.
21 Miotti PG, Canner JK, Liomba G, et al. Rate of new HIV infection in
a cohort of women of childbearing age in Malawi. IX Int Conf on
AIDS, Berlin, June 6-11, 1993, Abstr 652.
Genetic linkage of T-cell receptor
IgE responses
Summary
IgE responses to inhaled proteins underlie the clinical
syndrome of allergic (atopic) asthma and rhinitis. We have
investigated genetic linkage between specific IgE reactions to
highly purified major allergens and the T-cell receptor (TCR) &agr;
and &bgr; gene complexes on chromosome 14 and 7, respectively.
Antigens tested included highly purified proteins from the
housedust mite Dermatophagoides pteronyssinus, the
domestic cat and dog, grass pollen, and the mould Alternaria
alternata. Affected sibling-pair methods were used in two
independent sets of families, one in the UK and one in
Australia. No linkage of IgE serotypes to TCR-&bgr; was detected,
but significant linkage to TCR-&agr; was seen in both family
groups. For several of the IgE phenotypes investigated
(positive responses to whole allergen sources or purified
antigens or serum IgE above the 70th percentile in the
population) the affected sibling-pairs showed significant
sharing of TCR-&agr; microsatellite alleles from both parents.
The results show that a gene (or genes) in the TCR-&agr; region
modifies specific IgE responses.
Nuffield Department of Clinical Medicine, John Radcliffe Hospital,
Headington, Oxford OX3 9DU, UK (M F Moffatt DPhil, M R Hill DPhil,
J A Faux PhD, R P Young DPhil, W O C M Cookson MD);
Molecular Immunology Group, Institute for Molecular Medicine,
Oxford (F Cornélis MD); ALK Laboratories, Hørsholm, Denmark
(C Schou PhD); Department of Respiratory Medicine,
Sir Charles Gairdner Hospital, Perth, Western Australia
(A L James MD, G Ryan MD, A W Musk MD);
University Department of Paediatrics, Princess Margaret Hospital
for Children, Perth (P le Souef MD); and Osler Chest Unit,
Churchill Hospital, Oxford (J M Hopkin MD)
Correspondence to: Dr W O C M Cookson
vitamin A statusof children as measured by the relative dose response
(RDR). Am J Clin Nutr 1987; 46: 91-94.
23 Sommer A, Tarwotjo I, Katz J. Increased risk of xerophthalmia
following diarrhea and respiratory disease. Am J Clin Nutr 1987; 45:
977-80.
24 Gal I, Parkinson CE. Effects of nutrient and other factors on
pregnant women’s serum vitamin A levels. Am J Clin Nutr 1974; 27:
688-95.
25 Garbe A, Buck J, Hämmerling U. Retinoids are important cofactors in
T cell activation. J Exp Med 1992; 176: 109-17.
26 Buck J, Ritter G, Dannecker L, et al. Retinol is essential for growth of
activated human B cells. J Exp Med 1990; 171: 1613-24.
27 Graham NMH, Sorenson D, Odaka N, et al. Relationship of serum
copper and zinc levels to HIV-1 seropositivity and progression to
AIDS. J Acquir Immune Defic Syndr 1991; 4: 976-80.
28 Baum MK, Mantero-Atienza E, Shor-Posner G, et al. Association of
vitamin B6 status with parameters of immune function in early HIV-1
infection. J Acquir Immune Defic Syndr 1991; 4: 1122-32.
29 Beach RS, Mantero-Atienza E, Shor-Posner G, et al. Specific
micronutrient abnormalities in asymptomatic HIV-1 infection. AIDS
1992; 6: 701-8.
&agr;/&dgr; complex to specific
Introduction
Atopy, a familial clinical syndrome of asthma, rhinitis, and
eczema, is characterised by IgE-mediated allergy, which
results from genetic and environmental events. Atopic
individuals differ in the allergens to which they react. The
difference is clinically important, since asthma and
bronchial hyper-responsiveness are associated with allergy
to housedust-mite (HDM) antigen but not to grass
pollens.1.2
Genetic regulation of IgE responses to particular
allergens may differ from that of the general atopic
response. Inhaled allergens such as HDM are complex
mixtures of proteins. Several major allergens, to which IgE
responses are consistently found, have been identified from
each allergen source. It is likely that genetic associations
will be more readily detected with reactions to purified
major allergens, rather than with complex allergen sources.
Specific IgE reactions might be constrained by variation
in HLA or T-cell receptor (TCR) proteins, since these
molecules are central to the handling and recognition of
foreign antigens. However, HLA genes alone’ do not
account for the differences in individual IgE reactions to
allergens.3
The role of the TCR in allergic reactions is not yet clear.
Most of these receptors are made up of a and P chains,
although within any individual 5% consists of y and 8
chains. The P chain arises from chromosome 7 and the a
chain from chromosome 14. The õ chain genes are found
within the a chain locus.
An enormous potential for TCR variety arises from the
many variable (V) and junctional (J) segments within the
TCR loci. However, the use of TCR Vex and V(3 segments
by lymphocytes is not random and may be under genetic
control.4-6
To find out whether the TCR genes influence
susceptibility to particular allergens, we tested for genetic
1597
 Table 1: Primer sequences

linkage between IgE responses and the TCR-(X/8 and

TCR-(3 regions. Two independent sets of families, one
British and one Australian, were studied. Because the mode
of inheritance was unknown, and because of interactions
from the environment and other loci, affected sibling-pair
methods were used.

Subjects and methods

410 British individuals from 66 nuclear families and 5 extended
pedigrees were ascertained through family members with asthma
*2 affected siblings in a sibship give 1 sibling-pair,3 give 3 sibling-pairs, 4 give 6, and 5 give 10.
or rhinitis. 413 Australian subjects from 88 nuclear families each
with 2 or more atopic siblings were identified from a random Table 2: Affected siblings and affected sibling-pairs with each
population sample of the country town of Busselton, Western IgE phenotype
Australia. Together the samples provided 823 subjects in 176
sibships. The pedigrees contained 312 sibling pairs, although not
all siblings had positive IgE responses. Phenotyping was carried out by individuals unaware of genotype
For British subjects, serum samples were tested for the presence and vice versa. Linkage was explored with a limited number of

of high IgE titres to the highly purified major allergens Der p I and predetermined phenotype definitions and assessed in affected
Der p II (25’4 kDa and 14.1kDa, from Dermatophagoides sibling-pairs. The number and distribution of affected sibling-
pteronyssinus), Fel d I (18 kDa, from the cat Felis domesticus), pairs in the pedigrees is shown in table 2. Only sibling-pairs in
Phl p V (30 kDa, from timothy grass Phleum pratense), Alt a I which all four parental alleles could be unambiguously identified
(28 kDa, from the mould Alternaria alternata), and Canf (25 kDa, were counted (ie, identity by descent). All sibling-pairs from
from the dog Canis familiaris). IgE titres were measured by multiple sibships were considered as independent. 12 The
chemiluminescence assay with monoclonal anti-IgE (ALK frequency of sharing of two, one, or no alleles was compared with
Laboratories, Horsholm, Denmark): a positive response was taken that expected under a hypothesis of non-linkage (25%, 50%, 25 %)
as equivalent to 1 RAST unit or greater. The total serum IgE was by X2 goodness-of-fit testing.
measured (Phadezym PRIST, Pharmacia, Uppsala, Sweden) and a Lod scores (log of odds in favour of linkage) and recombination
raised level was taken to be greater than the 70th percentile of a fraction (8) between FCA.TAI and D14S50 were assessed by the
general population sample adjusted for age (data available from the LINKAGE computer program.
authors on request). These values approximate to published
normal values for children and adults. 7,8
Australian subjects were tested for Der pI, Der p II, and Fel d I Results
and total serum IgE. Reactivity to whole D pteronyssinus and
No linkage with any phenotype was seen with the TCR-(3
P pratense pollen extract was tested in all subjects by ELISA
microsatellite in either group of families (table 3). There
(Phadezym RAST, Pharmacia).
was no association of any TCR-(3 microsatellite allele with
Genomic DNA was obtained from peripheral blood
lymphocytes of all subjects by phenol-chloroform extraction. reactivity to a particular allergen.
Genotypes were determined by polymerase chain reaction for two Significant excess sharing of TCR-a microsatellite alleles
microsatellite DNA sequences close to the TCR loci-FCA.TA1 was, however, seen in British sibling-pairs with IgE
on chromosome 14 and V06.7CA on chromosome 7.9,10 The responses to HDM (table 4). In Australian subjects there
primers used in a simultaneous reaction are listed in table 1. was excess sharing of alleles in siblings responsive to grass
Each 25 L reaction mixture contained 200 ng human genomic
pollen or with a total serum IgE greater than the 70th
DNA, 2 U Taq polymerase (Boehringer Mannheim) with the percentile (table 4). In the whole study population there was
buffer recommended by the supplier (final concentration 1-5
mmol/L magnesium chloride) and 200 Ilmol/L of each dNTP. Each
primer was at a final concentration of 0-125 fimoI/L. We added
105 cpm of each L primer (FCA.TA1-L and V/36. 7CA-L),
end-labelled withy32PATP.
Amplification conditions were 94°C for 5 min, 65°C for 1 min,
72°C for 1 min, followed by 22 cycles of 94°C for 1 min, 65°C for 1
min, and 72°C for 1 min. Amplified products were resolved on 6°o
polyacrylamide gels and detected by autoradiography. Sizes of the
alleles were determined by comparison with the known genotype of
CEPH individual 1702. Typing for another microsatellite, Data given for Australian and British groups together.
D14S50, which is close to TCR-a,’1 was also carried out (table 1).
Table 3: Sharing of parental TCR-(3 microsatellite alleles by
Genotypes for all markers were independently assigned by two
investigators. sibling-pairs with IgE reponses

1598
Table 4: Sharing of TCR-a microsatellite alleles from both parents
significant excess sharing of TCR-a alleles by affected
sibling-pairs for each phenotype.
Linkage to TCR-a was then tested with IgE responses to
highly purified major allergens. Because only 6 British
sibling-pairs shared positive responses to Phlp V, and 5 and
2 shared responses to Alt a V and Can f I, respectively,
Australian subjects were not tested with these antigens.
Reactions to the purified domestic allergens Der p I,
Der p II, and Fel d I were all associated with excess
sibling-pair sharing of TCR-a microsatellite alleles in the
British subjects (table 4). In the Australian subjects excess
sharing was seen in sibling-pairs who reacted to Fel d I and
Der p I but not in those who reacted to Der p II(table 4).
When the two sets of families were pooled, allele sharing
was highly significant for all phenotypes. The level of
significance was greater than that seen with IgE responses
to whole allergen sources.
The pattern of allele sharing for Der p I and Fel d I was
consistent with a recessive influence on IgE responses in
both sets of families (table 4), since a greater proportion
shared two parental alleles than shared one.
Non-affected sibling-pairs did not show significant allele
sharing, which suggests low penetrances of the traits.
Sibling-pairs discordant for the serotypes did not show
excess allele sharing. There was no association between IgE
serotypes and particular alleles of the TCR-a
microsatellite.
As a control for the correctness of genotyping, the
families were typed for the microsatellite D14S50, for
which a recombination fraction of 0 029 from TCR-a has
been reportedY In our subjects D14S50 was in close
linkage with the TCR-a microsatellite, with a
recombination fraction of 0 019 and a lod score of 61. This
result confirms the accuracy of genotyping.
Discussion
We have found that a locus in the TCR-a/8 region
modulates IgE responses. The close correlation between
total and specific IgE makes it difficult to find out whether
the locus controls specific IgE reactions or confers general
IgE responsiveness. However, linkage was strongest with
highly purified allergens, which suggests that the locus
primarily influences specific responses. The pattern of
allele sharing seen for some phenotypes suggests a recessive
genetic effect; this linkage could correspond to the recessive
atopy locus implied by previous segregation analyses. 13,14
Replication of positive results of linkage in a second set of
subjects is important in interpretation of this study.
Differences between the populations for the serotypes
showing TCR-a allele sharing may be due to different
allergen exposure, since grass pollen responses were much
more common in Australian subjects. In addition, British
subjects were recruited through clinics, whereas those in
Australia were not selected by symptoms. The families did
not general have many affected siblings, and linkage is
in
not an artifact arising from a few large pedigrees: indeed,
there was no linkage between TCR-a and the phenotype
defined by a high IgE among British subjects when several
siblings in a family were affected.
The absence of association between particular IgE
responses and specific TCR-M microsatellite alleles
suggests that the microsatellite is not immediately next to
the IgE-modulating elements. The degree of linkage
disequilibrium across the TCR-a/8 locus seems low,15 and
the microsatellite has been localised only within a 900 kb
yeast artificial chromosome.9 The observed linkage may
therefore be with any elements of TCR-a or TCR-8 or
with other genes in the region.
Several V Cl genes are polymorphic,16 and limitation of
the response to an allergen may correspond to these
polymorphisms. Particular TCR-Va use may induce
interleukin-4 dominant (Th2) helper T-cells, which
enhance IgE production." Reported non-random use of
Val3 in Lol p I specific T-cell clones supports the
possibility that Va genes control IgE responses.18
The TCR-S locus is also a candidate for this linkage. The
function of TCR-y/8 cells is not known, but their location
on mucosal surfaces, where allergens initiate IgE responses,
could suggest a role in IgE regulation.19
This study has therefore identified a further genetic locus
that affects atopy. The genetic restriction of specific IgE
responses may be important clinically and in understanding
of the control of humoral immunity. Further localisation of
this genetic effect requires the identification of TCR-a/8
elements that show population associations with specific
IgE responses. Studies are also needed to investigate the
interactions between this chromosome 14 linkage, the HLA
genes, and the previously identified chromosome llq atopy
locus.2o
We thank all the families who took part in this study; the many individuals
who helped in testing the Busselton population sample;
Ms Gitte N Hansen (ALK Research) for carrying out much of the IgE
serology to purified allergens; and Prof J Bell, Dr P Moss, and DrJ Todd
for discussions of the results. The study was supported by the Wellcome
Trust, and WOCMC is a Welcome senior clinical research fellow.
References
1 Sears MR, Herbison GP, Holdaway MD, Hewitt CJ, Flannery EM,
Silva PA. The relative risks of sensitivity to grass pollen, house dust
mite and cat dander in the development of childhood asthma.
Clin Exp Allergy 1989; 19: 419-24.
2 Cookson WOCM, De Klerk NH, Ryan GR, James AL, Musk AW.
Relative risks of bronchial hyper-responsiveness associated with
skin-prick test responses to common antigens in young adults.
Clin Exp Allergy 1991; 21: 473-79.
3 Young RP, Dekker JW, Wordsworth BP, et al. HLA-DR and
HLA-DP genotypes and immunoglobulin E responses to common
major allergens. Clin Exp Allergy 1994; 24: 431-39.
4 Loveridge JA, Rosenberg WMC, Kirkwood TBL, Bell JI. The genetic
contribution to human T-cell receptor repertoire. Immunology 1991;
74: 246-50.
1599
 5 Moss PAH, Rosenberg WMC, Zintzaras E, Bell JI. Characterization of
the human T cell receptor &agr;-chain repertoire and demonstration of a
genetic influence on V&agr; usage. Eur J Immunol 1993; 23: 1153-59.
6 Gulwani-Akolar B, Posnett DN, Janson CH, et al. T cell
receptor V-segment frequencies in peripheral blood T cells correlate
with human leukocyte antigen type. J Exp Med 1991;
174: 1139-46.
7 Holford-Strevens V, Warren P, Wong C, Manfreda J. Serum total
immunoglobulin E levels in Canadian adults. J Allergy Clin Immunol
1984; 73: 516-22.
8 Kjellman N-IM, Johannson SJO, Roth A. Serum IgE levels in healthy
children quantified by a sandwich technique (PRIST). Clin Allergy
1976; 6: 51-59.
9 Cornélis F, Hashimoto L, Loveridge J, et al. Identification of a CA
repeat at the TCRA locus using yeast artificial chormosomes: a general
method for generating highly polymorphic markers at chosen loci.
Genomics 1992; 13: 820-25.
10 Li Y, Szabo P, Robinson MA, Dong B, Posnett DN. Allelic variations
in the human T cell receptor V&bgr;6.7 gene products. J Exp Med 1990;
171: 221-30.
11 NIH/CEPH Collaborative Mapping Group. A comprehensive genetic
linkage map of the human genome. Science 1992; 258: 67-86.
12 Blackwelder WC, Elston RC. A comparison of sib-pair linkage tests for
disease susceptibility loci. Genet Epidemiol 1985; 2: 85-97.
Randomised trial of preventive nasal ventilation in Duchenne
muscular dystrophy
Jean-Claude Raphael, Sylvie Chevret, Claude Chastang, Françoise Bouvet, for the French Multicentre
on Home Mechanical Ventilation Assistance in Duchenne de Boulogne Muscular Dystrophy*
Summary
Duchenne muscular dystrophy (DMD) is the most common
muscular dystrophy in children. Paralysis of respiratory
muscles causes a decrease in forced vital capacity (FVC) from
age 12 years, and death occurs between 20 and 25 years old
and is usually related to respiratory insufficiency. Uncontrolled
studies suggest that early home use of nasal intermittent
positive-pressure ventilation (NIPPV) in DMD patients free of
respiratory failure could limit progression of the restrictive
syndrome and therefore improve survival Because efficacy of
preventive NIPPV has not been demonstrated in a controlled
trial, we undertook a randomised multicentre study in which 70
patients with DMD were included.
Patients were free of daytime respiratory failure and FVC
was between 20 and 50% of predicted values. At least 6 h of
nocturnal NIPPV (n=35) was compared with conventional
treatment (n = 35). During a mean follow-up of 52 months, 10
patients died,8 in the NIPPV group and 2 in the control group
(p=0·05, log-rank test). No differences were observed
between the two groups for occurrence of hypercapnia,
decrease of FVC below 20% of initial values, or use of
necessary mechanical ventilation.
Preventive NIPPV did not improve respiratory handicap and
reduced survival of DMD patients. Use of NIPPV for preventive
purposes should be avoided in patients with FVC between 20
and 50% of predicted values.
*Collaborators are listed at the end of the article.
Service de Réanimation Médicale, Hôpital Raymond Poincaré,
92380 Garches, France (Prof J-C Raphael MD, F Bouvet MD); and
Département de Biostatistique et Informatique Médicale, Hôpital
Saint-Louis, Paris (S Chevret MD, C Chastang MD)
Correspondence to: Prof J-C Raphael
1600
13 Dizier MH, Hill M, James A, et al. Genetic control of IgE level after
accounting for specific atopy. Genet Epidemiol 1993; 10: 333-34 (abstr).
14 Gerrard JW, Rao DC, Morton NE. A genetic study of
immunoglobulin E. Am J Hum Genet 1978; 30: 46-58.
15 Robinson MA, Kindt TJ. Genetic recombination within the human
T-cell receptor alpha-chain complex. Proc Natl Acad Sci USA 1987;
84: 9089-93.
16 Cornélis F, Pile K, Loveridge J, et al. Systematic study of human &agr;&bgr;
T-cell receptor V segments shows allelic variations resulting in a large
number of distinct TCR haplotypes. Eur J Immunol 1993; 23: 1277-83.
17 Heinzel FP, Sadick MD, Mutha SS, Locksley RM. Production of
interferon gamma, interleukin 2, interleukin 4, and interleukin 10 by
CD4 + lymphocytes in vivo during healing and progressive murine
leishmaniasis. Proc Natl Acad Sci USA 1991; 88: 7011-15.
18 Mohapatra SS, Mohapatra S, Yang M, et al. Molecular basis of
cross-reactivity among allergen-specific human T cells: T-cell receptor
V&agr; gene usage and epitope structure. Immunology 1994; 81: 15-20.
19 Holt PG, McMenamin C. IgE and mucosal immunity: studies on the
role of intraepithelial Ia+ dendritic cells and &dgr;/&g r; T-lymphocytes in
regulation of T-cell activation in the lung. Clin Exp Allergy 1991; 21
(suppl): 148-52.
20 Sandford AJ, Shirakawa T, Moffatt MF, et al. Localisation of atopy
and &bgr;-subunit of high-affinity IgE receptor (Fc&egr;RI) on chromosome
11q. Lancet 1993; 341: 332-34.
Cooperative Group
Introduction
Duchenne muscular dystrophy (DMD), the most common
muscular dystrophy of childhood, is found in 1 in every
5000 boys. Paralysis of limbs is progressive, with loss of
walking ability occurring between 8 and 12 years old.’
Cardiac failure, mostly observed in late stages of the disease,
accounts for 10-20% of deaths.2,3 Paralysis of respiratory
muscles causes a decrease in forced vital capacity (FVC)
from 12 years old, and worsens inexorably. Daytime
hypercapnia appears between 18 and 20 and is associated
with a poor outcome.4 Death occurs between 20 and 25, and
is usually related to respiratory insufficiency. Despite
progress in the pathophysiological understanding of the
disease,s no specific treatment has been found to delay
death. Although a controlled trial of the effect of 6 months’
therapy with corticosteroids on muscular strength showed
short-term6 and long-term7 benefit, there is no proof that
corticosteroids can stop the course of respiratory handicap.
The only available palliative treatment of respiratory
failure is home mechanical ventilation assistance; however,
this treatment remains controversial. 2,4,SEarly intermittent
positive-pressure ventilation by nasal mask (NIPPV) in
patients free of respiratory failure may limit progression of
the restrictive syndrome and therefore prolong survival.9-11
We have called this intervention preventive ventilation as
opposed to necessary ventilation when assistance is
imperative. 12,13 However, the efficacy of preventive NIPPV
has not been tested prospectively, its use is constraining to
the patient and costly for the community, and the lack of
assessment of NIPPV causes patients and their families to
question the treatment. 14,15 Thus, we have done a
multicentre trial that compared at least 6 h of nocturnal
preventive NIPPV with conventional treatment.
Patients and methods
The trial was approved by the ethics committee of the Societe de
Reanimation de Langue Francaise. Patients with DMD admitted