Goals:

• To describe the functions of saliva and relate these functions to its specific constituents.
• To describe the mechanisms underlying the formation of saliva and the factors that intluence
saliva tlow rate and composition under normal and pathophysiological conditions.
• To describe the role of saliva for oral clearance and pH regulation.
• To describe the role of saliva in pellicle formation and tooth solubility.
• To discuss the identification of patients with salivary dysfunction, the measurement of saliva
praduction and oral dryness, and the management of dry mouth.

Key words:
Salivary glands; salivary reflexes; saliva formation; saliva electrolytes and proteins; buffer capacity; oral
clearance; hyposalivation and oral sequelae; xerostornia; management of dry mouth.

its composition as well as dry mouth symp-

• Introduction
toms combined with a thorough medical
Impaired saliva secretion and changes in the and oral history taking provide valuable clu-
saliva composition whether temporary or es for the dentist in the process of differen-
permanent are common conditions. On a tial diagnosis. Nevertheless, understanding
population basis it is assumed that at least the pathology behind salivary gland dys-
10% of the adults suffer from dryness of the functions and the abnormal saliva compo-
mouth. However, the prevalence of oral dry- sition requires detailed insight into both the
ness increases with age due to a higher fre- normal physiology and the pathophysiology
quency of systemic diseases and intake of of the salivary glands.
medicines amongst the elderly. Thus, about
30% of the population aged 65 years and
above suffers from oral dryness. Impaired • Functions of saliva
saliva secretion (hyposalivation) increases
the risk of oral diseases like dental caries and Saliva protects the teeth and the oro-oeso-
oral candidal infection. In addition, the sen- phageal mucosa through a number of mech-
sation of oral dryness (xerostomia) may anisms. Besides maintenance of the integrity
compromise the patient's health-related of these tissues, saliva also has multiple func-
quality of life. Assessment of saliva flow and tions in relation to digestion in the upper

17
 Bardow, Lynge Pedersen and Nauntofte

gastrointestinal tract. It facilitates food in-
take by dissolving food taste-substances, it
clears and dilutes food detritus and bacterial
matter, rinses the mouth, lubricates oral soft
tissues, and facilitates mastication, swal-
lowing and speech. As shown in Table 1 and
Figure 10, some of these important func-
tions relates to the fluid characteristics and
others to specific components of the saliva.
Clearly, therefore, changes that compromise
salivary gland function are likely to affect
oral function and health.
• The salivary glands
The mixed fluid in the mouth is called whole
saliva. It is derived predominantly from three
pairs of major salivary glands (Figure 1) that
account for about 90% of the total fluid secre-
tion, and from the minor salivary glands in
the oral mucosa that contribute somewhat
less than 10% of the total volume. In addition,
whole saliva also contains gingival crevicular
fluid (depending upon the periodontal status
of the patient), microorganisms frorn dental
plaque, and food debris. Saliva is a hypotonic
fluid relative to plasma and it is composed of
more than 99% water and less than 1% of dry
matter such as proteins and salts. The normal
daily production of saliva ranges between
0.5-1.5 litre.
In the resting state, about two-thirds of the
volume of whole saliva is produced by the
submandibular glands. However, when sali-
vary glands are stimulated, the parotids can
account for at least half of the whole saliva
volume in the mouth. Onlya small percen-
tage of unstimulated or stimulated whole sal-
iva comes from the sublingual glands. AI-
though the minor glands, which are distrib-
uted in groups throughout the oral mucosa,

Table 1. Principal functions of saliva related to fluid characteristics and specific components.

Functions Salivary fluid characteristics and specific components

Protection
Mechanical cleansing of the mouth Water
Clearance of food/microorganisms Water
Lubrication of oral surfaces Water, mucins, proline-rich glycoproteins
Mucosal intergrity and coating Water, mucins, electrolytes, epiderrnal growth factor, nerve growth
factor
Tooth mineralisation Cystatins, histatins, proline-rich proteins, statherins Ca2+, and
phosphate
Buffering Bicarbonate, phosphate and protein
Antimicrobial activity Anti-bacterial: amylases, cystatins, histatins, mucins, lactoperoxidase,
Iysozyme, lactoferrin, calprotectin, irnrnunoglobulins, chromogranin A,
von Ebner glands protein, secretory leukocyte proteinase inhibitor
Anti-fungal: histatins, irnrnunoglobulins, chromogranin A
Anti-viral: cystatins, mucins, irnmunoglobulins, secretory leukocyte
proteinase inhibitor

Digestion and speech

Formation of bolus Water, mucins
Mastication and swallowing Water, mucins
Initial digestion Water, mucins, amylases, lipases, ribonucleases, proteases
Taste Water, gustin, Zn2+
Speech Water, mucins

18
 Saliva

Stensen's duct

Parotid gland

SublinguaJ
caruncula

Sublingual Submandibular gland

gland

Wharton's duct

• Figure 1, The salivary glands.

Human major salivary glands: the parotid, the submandibular, and the sublingual gland. The parotid duct (Stensen's
duct) is about 5 cm long and opens into the mouth in the buccal mucosa opposite the second maxillary molar. The
saliva from the submandibular and sublingual glands enters the mouth mostly through the around 5 cm long
submandibular duct (Wharton's duct), which ends on the sublingual caruncula behind the mandibulary incisors.
But the sublingual gland also secretes through several smal! ducts along the sublingual fold in the f100r of the mouth
lateral to the side of the tongue.

make only a small contribution to the total Some of the salivary glands are pureJy se-
saliva voJume, they play an important roJe in rous (Iike the parotids), others are mucous
the lubrication of the mucosa since they se- (Iike the minor palatine glands), and others
crete a Jarge fraction of the salivary proteins. are mixed gland types (submandibular, sub-

Table 2. Morphological and biochemical characteristics of the salivary glands.

Salivary gland Acinar cel! type Fluid characteristics Innervation"

The majar salivary glands

Parotid gland Serous watery, amylase-rich IX
Submandibular gland Mixed, mainly IllUCOUS viscous, mucin-rich VII
Sublingual gland Mixed, mainly IllUCOUS viscous, mucin-rich VII
The minar salivary glands
Palatinal Mucous mucin rich VII
Buccal Seromucous mucin rich VII
Labial Seromucous mucin rich VII
Lingual (von Ebner's glands) Serous watery, lipase-rich f1uid IX
Retromolar Mainly mucous mucin rich VII/IX
"Parasympathetic nerve supply. The sympathetic nerve supply is derived from the superior cervical ganglion.

19
Bardow, Lynge Pedersen and Nauntofte

lingual and minor buccal glands). Table 2 In general, the acinar cells ("secretory end-
summarizes the acinar characteristics of the pieces") comprise about 80% of the gland
salivary gland types, their innervation and mass. Primary saliva is formed in this region
their fluid characteristics. of the gland. The saliva then moves into the

Blood

Acinar cell

ACh
cc-adr

--++-- Intercalated
duct

In tersti ti um Lumen

ACh
o.-adr
K+ .••
~I--;;=--

CI- "'~I--;=-- Excretory

duct

Arterial

Striated duct cell

20
 Saliva

duct system where it continuously undergoes These ducts are quite long and have rela-
modification until it is secreted into the tively large diameters. Conversely, the sub-
mouth. lingual glands as well as a number of minor
The length and the diameter of the ducts glands have only sparsely distributed interca-
vary depending on the gland type. The duct lated and striated duct segments, which may
system of the human parotid and submandi- even be absent in short, small diameter
bular glands is branched and contains in- ducts. Myoepithelial cells envelop the se-
tercalated, striated and excretory duct seg- cretory end -pieces and intercalated ducts.
ments with the striated ducts making up the When they contract, these cells are believed
bulk of the duct system (Figure 2). In the to promote the flow of primary saliva into
parotid and submandibular glands, each seg- the ducts (Figure 2).
ment of the duct system is well developed. The secretory end-piece consists of polar-
ized acinar cells, which surround a central
lumen that is connected to the intercalated
• Figure 2. lntracellular ion and water transport in differen t ducts ("intercalated" means "placed be-
glal1d segments.
tween", and refers to the location of the
Salivary glands consist of many thousands of secretory
units comprising acini (arrangement of acinar cells) and
ducts, which lie between the acinus and the
an intercalated and a striated duct segment. A large nurn- striated duct). The nucleus and a major part
ber of such secretory units converge into a main excretory of the endoplasmic reticulum (ER) are
duct that drains the saliva into the mouth. To the right: located in the basal part of each acinar cell,
Although the duct system is branched thereby serving
whereas the protein-containing granules are
several secretory units, this illustration is a simplified
schematic drawing of a major gland and its blood supply.
placed at the luminal poleo Gap junctions
The blood flows in opposite direction to the saliva flow. connect the cytoplasm of the acinar cells and
To in crease salivary secretion upon autonomic stimula- allow for electrical coupling within the acini.
tion, vasodilatation and increased blood flow takes place. However, there are also tight junction struc-
To the left: Main membrane transport mechanisms of
tures that connect the acinar cells with each
the acinar and striated duct cells. In both cell types, the
specific neurotransmitters bind to their specific receptor
other. These are selectively leaky to cations
complexes and, via a number of intracellular biochemical and water and serve to separate the luminal
events, the outcome is activation of plasma membrane (primary saliva) and interstitial fluids. Fur-
ion transporters leading to the formation of saliva. For thermore, the acinar cell membranes are
simplistic reasons two acinar cells illustrate in cornbi-
highly permeable to water.
nation the transporters that are activated in a single acin-
ar cells. The upper acinar cell illustrates the Ca2+ -acti-
The intercalated duct cells are believed to
vated loss of K" and cr that occurs initially on stimula- be the stem cells for the acinar and ductal
tion/receptor activation whereas the lower acinar cell cell types, and they also are believed to aid in
illustrates the transporters involved in the re-establish- primary saliva secretion. The striated ducts
ment of the prestirnulatory ion gradients across the
consist of columnar cell types with deep
plasma membrane. Note that the cells have two transport
mechanisms, the parallel operating exchange systems for
membrane infolding and many mitochon-
Na+/H+ and Cl~/HC03~ as well as the Na+/K+/2Cl~ dria in the basal surface facing the interstiti-
cotransporter, that both result in cellular uptake of Na+ um. These duct cells modify the primary sal-
and cr . It should also be noted that the important iva formed in the acinus. In contrast to the
mechanism involved the intracellular pH regulation is
acinar cells, the membranes of duct cells
the conversion of CO2 (arising from the metabolic turn
over) and H20 into equal amount of HC03 ~ and H+
have a low permeability to water, and the
catalyzed by the presence of cytosolic carbonic-anhyd- tight junctions connecting the duct cells
rase. have a very low permeability to both ions

21
Bardow, Lynge Pedersen and Nauntofte
and water. Nevertheless, some water reab-
sorption may occur in the duct cells at very
low flow rates, especially in the intralobular
ducts due to the osmotic gradient or as a
result of the influence of circulating antidi-
uretic hormone (ADH).
Finally, the intralobular ducts drain into
an extralobular excretory duct system that
carries the saliva to the main excretory duct
which is lined with stratified columnar epi-
thelium.
Blood supply to the salivary glands
The rich blood supply of the major salivary
glands is important for saliva production
since the fluid in saliva originates from the
capillaries and the interstitial fluido This
blood supply is organised as a portal system
with two capillary networks in series, a
dense one around the duct system and an-
other around the secretory end-piece (Figure
2). The secretory activity of the salivary
glands is entirely under autonomic control.
The blood vessels of the salivary glands are
controlled by the sympathetic nervous sys-
tem, which makes them constrict. However,
parasympathetic stimulation of the glands
can overcome this sympathetic vasoconstric-
tor tone and lead to vasodilatation and in-
creased blood flow to the secreting gland.
Parasympathetic stimulation induces the
production of a number of substances which
participate in the regulation of local blood
flow and hence the secretory activity of the
gland. These substances include mediators
such as vasoactive intestinal polypeptide
(VIP) and nitric oxide (NO) that are syn-
thesised and released by the salivary glands
and their surrounding tissues comprising
nerves, endothelium, and blood vessels. In
addition, the striated ducts release a serine
pro tease, kallikrein, which acts on circu-
lating plasma proteins to induce the forma-
tion of the vasodilator bradykinin: this in
22
turn participates in the regulation of local
blood flow to the gland.
• Nervous control of salivary
secretion
The secretion of saliva is regulated by re-
flexes involving the autonomic nervous sys-
temo The reflex pathways are unilateral, since
stimulation of one side of the mouth induces
only ipsilateral salivation. The act of chewing
and the sensation of taste initiate action po-
tentials in various sensory receptors (Figure
3). The masticatory-salivary ref/ex involves
sensory inputs mainly from mechanical re-
ceptors in the mouth. These arise predomi-
nantly from mechanoreceptors in the peri-
odontal ligament, but other proprioceptors
in the trigeminal innervation including
muscle spindles in the masticatory muscles
and oral nociceptive stimuli may contribute.
The gustatory-salivary ref/ex, however, util-
ises sensory signals from taste-activated
chemoreceptors in the taste buds within the
lingual papillae and in the tonsillar region,
the epiglottis, the pharyngeal wall and
oesophagus: these are conducted along the
facial (VII), glossopharyngeal (IX), and
vagus (X) nerves to the salivatory nuclei in
the medulla oblongata of the brain. Here,
the signals are integrated and activa te the se-
cretomotor pathways of the reflex that con-
sist of parasympathetic and sympathetic
autonomic nerve bundles that travel along
separate pathways to the salivary glands. The
submandibular and sublingual glands are
controlled by the superior salivatory nu-
cleus, whereas the parotid is controlled by
the inferior salivatory nucleus.
Selective parasympathetic or sympathetic
stimulation of the salivary glands elicits se-
cretion. But the parasympathetic fibres car-
ried in the VII and IX nerves and the sym-
 Saliva

• Figure 3. Regulation of secretion.

cerebral cortex
The reflexes involved in salivary secre-
tion. Afferents nerves carry impulses to
salivary nuclei, the center of salivary
thalamus
seeretion placed in the medulla. From
here signals are directed to the efferent -.-?- -
\II,•......•.•.•.. hypothalamus
part of the reflex, which stands under
autonomic regulation by parasympath- -- Iimbic system
etie and sympathetic nerves. Besides
the afferents shown in this figure, other
afferents arising from olfaction and
stretch of the stomach can initiate sali-
vation. Concerning the efferents, the
sympathetic nerves run from the sym-
pathetic trunk, follow the blood ves seis
supplying the glands, and then separ-
ately innervate the glands. The parotid
gland receives parasympathetic signals
from the glossopharyngeal nerve that
synapses in the otic ganglion placed
relatively close to the gland. The sub-

a
mandibular and sublingual glands re-
eeive parasympathetic signals from the submandibular ganglion
facial nerve that synapse in the sub-
mandibular ganglion. Release of neu-
rotransmitters from the postganglionic
neurons of both branches of the ner-
vous system elicits secretion of saliva. sublingual submandibular
gland gland

superior
cervical ganglion

sympathetic trunk

pathetic fibres following the blood vessels
supplying the glands normally act in concert
to produce saliva. The release of neurotrans-
mitters from autonomic nerve endings acti-
yates specific cell surface membrane recep-
tors on the salivary gland tissue thereby de-
termining the flow rate and composition of
saliva.
The classical neurotransmitters released
from the peripheral postganglionic parasym-
pathetic and sympathetic nerve endings are
acetylcholine (ACh) and noradrenaline (NA)
respectively. However, other substances that
are co-released such as adenosine triphos-
phate (ATP), substance P, VIP, neuropeptide
y also have important modulatory effects on
the formation of saliva.
The saliva flow rate depends not only on
the nature of the stimulus but also on its
duration and intensity. Thus stimuli like

23
 Bardow, Lynge Pedersen and Nauntofte
strongly acidic taste, high chewing frequency
and/or high bite force result in elevated sal-
iva flow rates. Generally, the parasympath-
etic branch provides the main stimulus for
salivation giving rise to a high flow rate of
watery saliva, compared with sympathetic
stimulation which leads to a lower flow rate
of saliva that is much more viscous beca use
of its high content of mucins.
The reflexes mentioned above are known
as "unconditioned" reflexes. However, sali-
vary secretion can also be initiated by "con-
ditioned" reflexes that are programmed in
higher centres in the brain (Figure 3). Thus,
positive experiences with foods in the past
may lead to the secretion of saliva induced
by the sight or thought of food (Pavlov's
classical dog experiments). Emotional state
also influences the saliva flow rateo Dental
students will be familiar with the experience
that fear or anxiety can lead to a dry mouth
due to central inhibition of the reflex path-
way. Salivation may also be diminished in
untreated depression. Furthermore, during
sleep, saliva secretion from the major glands
is normally very low.
In sumrnary, therefore, many signals from
a variety of peripheral receptors and from
higher centres are being constantly inte-
grated in the salivatory nuclei, the result of
which may be either facilitation or inhi-
bition of salivation.
• Formation of saliva
According to the secretion model for forma-
tion of saliva proposed by Thaysen and col-
leagues more than 50 years ago, saliva is
formed basically in two steps. The secretory
end-piece produces primary saliva which is
isotonic, having an ionic composition simi-
lar to that of plasma. This fluid is then
modified in the duct system by selective re-
24
absorption of Na + and Cl- (but not water)
and by a certain secretion of K+ and HC03 -
to yield the final saliva which is secreted into
the mouth. Thus the secretion rate and
thereby the volume of final saliva is deter-
mined directly by the formation rate of pri-
mary saliva by the acinar cells.
Figure 4 shows that the ionic composition
of primary saliva resembles that of plasma
or interstial fluido However, the formation of
primary saliva is not the result of pressure
filtration of blood. Rather, it is the result of
active transport of solutes by the gland tissue
that arises from a dramatic, stimulation-in-
duced increase in metabolic activity.
Stimulus-secretion coupling
The secretion of electrolytes, water and the
exocytotic release of proteins from the acinar
cells upon stimulation involve a multitude
of biochemical signalling processes: a few
important ones are illustrated in Figure 5.
The key event is the rise in the free Ca2+
concentration ([ Ca2+]) in the acinus that is
initiated by specific activation of receptors in
the plasma membrane by neurotransmitters.
However, the mechanism by which the vari-
ous activated receptor systems induce a rise
in Ca2+ involves different signalling routes.
ACh binds to muscarinic cholinergic re-
ceptors and the NA to u¡-adrenergic recep-
tors in the salivary gland cell membranes:
both are G protein-coupled receptors of the
so-called Gq/ll type. Binding induces phos-
pholipase C-mediated hydrolysis of the
plasma membrane component phosphati-
dylinositol 4,5 bisphosphate (PIP2) that
forms the second messengers inositol 1,4,5
trisphosphate (IP3) and diacylglycerol
(DAG).
The water-soluble IP3 binds to specific
IPr receptors on the endoplasmic reticulum
(ER) that induces Ca2+ release from this
store within the cell, giving rise to an in-
 Saliva

Oral cavity Duct Acinus Interstitium

r-----~~~~~~----'l r¡-------------------------, ¡ 1

/
Final saliva (mM)
unstimulated stimulated Primary saliva (mM) Interstitium (mM)
Na+ 3 45 Na+ 146 Na+ 143
K+ 25 21 K+ 4 K+ 4
CI- 24 40 cr 102 cr 109
RCO] 3 26 RCO) 28 RCO) 28
PC02 4.5 4.5 (kPa) PC02 6 (kPa) PC02 6 (kPa)
pH 6.5 7.5
protein 2.0 2.5 (mglml)

• Figure 4. Formation and composition of saliva.

The formation of saliva occurs in two steps. Step 1: The secretory end pieces produce a primary saliva resembling
plasma in ionic cornposition. Step 2: This fluid undergoes major changes and become hypotonie as it passes down
the duct system, primarily by reaborption of Na+ and CI- without water. However, the duetal modification strongly
depends on the secretion rateo The figure also shows typieal eleetrolyte concentrations for unstimulated as well as
stimulated whole saliva.

crease in the free intracellular [Ca2+ 1 in the
range from 10-7 M to 10-6 M. Ihis pathway
is particularly important for initiating elec-
trolyte transport.
Another receptor-coupled signalling path-
way that leads to an increase in the intra-
cellular free [Ca2+ 1 is the ~-adrenergic acti-
vation (by NA) of a Gs-protein and adenyl-
ate cyclase, which elicits the synthesis of
cyclic adenosine monophosphate (cAMP).
This cAMP then activates protein kinase A,
which in turn causes intracellular [Ca2+ 1 to
increase. The cAMP pathway has been impli-
cated in protein synthesis in the rough ER,
and in exocytosis of protein-containing se-
cretory granules across the cell membranes
involving a number of sma11 GIP binding
proteins. In addition, protein synthesis also
occurs as a conseguence of DAG formation:
this activates protein kinase C, another me-
diator of cellular protein synthesis and secre-
tion. Protein secretion from the salivary
gland tissues comprises a continuous so-
called "constitutive exocytosis" of protein-
containing vesicles that contribute to the
protein secretion and is ongoing at a11times.

25
 Bardow, Lynge Pedersen and Nauntofte

activation protein
plasma DAG - of protein kinase C - phosphorylatíon
membrane
_ release of Ca2+ _ electrolyte transport

from ER

muscarinic, secretion of electrolytes,

o-adrenergic, water, and proteins
or substance P ~
stimulation ~

activation protein
cAMP - of protein kinase A - phosphorylation

ATP

f3-adrenergic
stimulation \ ~ ....t....,

• Figure 5. Membrane receptors and cell signaling pathways.

Binding of a variety of neurotransmitters to specific ceLlsurface receptors in the acinar plasma membrane initiates
a number of biochemical cascade reactions within the cell membranes and cytoplasm that leads to secretion of
primary saliva from the secretory end piece. This schematic drawing illustrates some receptor-coupled processes
generating increased cytosolic concentration of the second messengers: calcium and cyclic adenosine monophosphate
(cAMP). Generally, activation of muscarinic cholinergic, u,-adrenergic or substance P receptors leads to an increased
intracellular calcium concentration resulting in electrolyte and water transport and protein synthesis and secretion
whereas activation of the ~-adrenergic receptor results in an increased cAMP concentration, which triggers protein
synthesis and secretion. The basic signaling pathways shown i11this figure are also involved in the ductal cell types
where activation of transport mechanisms occur leading to modification of the primary saliva by the reaborption of
some electrolytes and secretion of others as well as by protein synthesis and release.

The constitutive exocytosis can be acceler-
ated to the regulatory exocytosis that occurs
as a result of appropriate firing impulse fre-
quency and specific receptor activation of
the salivary glands. This regulated exocytotic
protein secretion from the major salivary
glands is controlled by the dual parasym-
pathetic and sympathetic secretomotor in-
nervation. However, the minor glands se-
crete a protein rich (mucin) secretion con-
tinuously.
Electrolyte transport by the acinar cells
The plasma membranes of the acinar cells
are freely permeable to water and to lipid-
soluble substances, but not to small, charged
molecules such as ions. Thus electrolyte
transport across the plasma membrane must
occur through specific transport mechan-
isms such as ion channels, pumps, cotrans-
porters and exchange systems (Figure 2).
The general principle behind the formation
of primary saliva is that the acinar cell re-

26
 Saliva

sponds to a receptor-activated increase in causes water to follow the inward move-

the intracellular free [Ca2+] by losing K+ to ments of ions and the cell swells back to its
the interstitium and cr (and some HC03-) prestimulatory volume. Accordingly, when
to the lumen via activated Ca2+ -regulated the stimulus is removed, the free intracellu-
K+ and cr channels in the basolateral and lar [Ca2+], the cell volumes, the cytoplasmic
luminal surfaces of the cell membrane, re- pH and the activity of the transporters in-
spectively. Because of the simultaneous acti- cluding the ion channels return to their orig-
vation of these ion channels, the membrane inal prestimulatory levels, and the acinus is
potential of about - 60 m V remains virtually again ready to produce substantial amounts
unchanged upon secretion. The accumu- of primary saliva in response to a new
lation of cr ions in the lumen creates a stimulatory challenge.
negative intracellular potential that drives
interstitial Na + into the lumen via a paracel- Duetal modifieation of electrolyte
lular transport route through cation-selec- composition of primary saliva
tive tight junctions to preserve electroneu- As for the secretory end-pieces the parasym-
trality. A transepithelial water flux, probably pathetic and sympathetic nerve fibres con-
occurring by trans- and paracellular path- trol the activity of the saliva ducts. Further-
ways, follows the net movement of salt into more, the membrane transporters and the
the lumen osmotically, resulting in acinar cell-signalling mechanisms of the duct cells
cell shrinkage (presumably by water loss are similar to those of the acinar cells. How-
via water channels "aquaporins") and for- ever, the ductal epitheliums possess both ab-
mation of isotonic, plasma-like primary sorptive and secretory functions. Most of the
saliva. reabsorption of Na + and Cl- occurs in the
As a result of this initial receptor-activated striated duct and, beca use of the low water
acinar cellloss of K+ and Cl- and water (cell permeability of the duct, the final saliva se-
shrinkage), the intracellular Na+ concen- creted into the mouth becomes hypotonic to
tration in the acinar cell increases mainly by plasma, i.e., with much lower concentrations
downhill Na+ influx via aetivation of the of Na+ and cr than primary saliva.
Na +/H+ exchanger and/or Na + /K+ I2Cl- The driving force for the Na + reabsorp-
cotransporter. This elevated acinar Na+ con- tion from the primary saliva across the lu-
centration in turn activates the central mem- minal membrane is generated from the ac-
brane element, the Na+ /K+ -pump (ATPase). tivity of basolateral ATP-consuming Na +/
This active pumping mechanism, utilizing K+ -pumps in the infoldings of the mem-
energy in the form of ATP, then re-establish- brane in the striated duct cells (Figure 2).
es the original, prestimulatory (unstimu- This pump mechanism maintains extrusion
lated) ion gradients across the acinar plasma of Na + from the duct cell to the interstitium
membrane by active uphill extrusion of Na + (and ductal uptake of K+). This creates an
and influx of K+. The prestimulatory acinar inwardly directed Na + gradient, allowing
cr concentration is re-established by uphill Na + to pass into the duct cell from the pri-
influx of the Cl- ion against an electro- mary saliva. The ductal uptake of Na + ,
chemical gradient via the cr /HC03 - which is activated by circulating adrenal
exchangers (that operate in parallel with mineralocorticoids (e.g. aldosterone), occurs
the Na + /H+ exchangers) and/or via the via Na + channels and presumably also by
Na +/K+ 12 cr cotransporter. Osmosis Na + /H+ exchange mechanisms. The uptake

27
 Bardow, Lynge Pedersen and Nauntofte
of Na + into the duct is to a large extent bal-
anced by parallel uptake of CI- vía CI-
channels and Cl-/HC03 - exchange mech-
anisms. Besides the uptake of CI-, some se-
cretion of K+ into the saliva occurs to pre-
serve electraneutrality. These transporters
are the most important mechanisms for the
modification of primary saliva by the ducts.
Stimulation of receptors in the duct cells by
neurotransmitters and peptides induces a
rise in intracellular free [Ca2+] and cAMP
in a manner similar to that in acinar cells,
but the regulatory rale of these messengers
for the modification of saliva in the duct is
still not clear.
Fram a clinical point of view, it is import-
ant to stress that medicines that inhibit the
muscarinergic or adrenergic receptor systems
(like some antidepressants, neuroleptics, an-
tihistamines, and hypertensives) or that di-
rectly affect the membrane transporters in the
salivary gland tissues (like some diuretics),
have the potential to induce changes in the
flow rate and composition of saliva.
• Saliva and its inorganic
composition
The final composition of the saliva ansmg
from the major salivary glands depends
• Figure 6. Inorganic saliva composition. 50
The saliva inorganic and organic com-
position as a function of the saliva flow 40
rateo As shown the salivar y concen-
trations ofNa+, cr , and HC03- in- 30
creases with increasing flow rates,
whereas the saliva concentrations of 20
..e:::.::::::::::.""""':::;,.,.c...------:::::;:::oo-=---
total phosphate and to some extend
K+ decreases with increasing flow 10
rates. The concentrations of total cal-
cium and total protein also increase
upon stimulation although not sub-
O
l
O
i~::;:~~;;:;;;;;;;~;;;;;;~~;;;:~T~o~ta~1
stantially.
28
strongly on the flow rateo However, the final
product secreted to the mouth is always hy-
potonic. Depending on the flow rate, whole
saliva contains 3-6 times less electrolytes
than plasma. As the flow rate increases, the
most dramatic increases occur in the Na +,
cr , and HC03 - concentration (see Figures
4 and 6).
Because the ductal transport mechanisms
involved in Na + (and CI-) reabsorption frorn
the primary saliva have a maximal transport
capacity, and beca use there is limited time
within the duct system to modify the electra-
lyte composition at high stimulated flow
rates, stimulated saliva is less hypotonic than
unstimulated saliva (i.e., it has higher concen-
trations of Na + and cr ).The concentration
of HC03 -, which is the principal buffer in
saliva, also varies with the flow rate: unstimu-
lated saliva contains very little HC03-
whereas stimulated saliva contains much
higher concentrations. With very power-
ful stimulation (including pharmacological
stimulation), the salivary HC03 - concen-
tration certainly exceeds the plasma level and
can even become the predominant anion in
the stimulated saliva. The secreted HC03 - is
derived from CO2 fram salivary gland met-
abolism. It is, however, not clear how much
originates fram the acinar and ductal gland
segments. It is possible that the striated ducts
Total protein
(0.1 . mglml)
HCOj(mM)
K+ (mM)
2
phosphate
Total calcium (mM)
Whole saliva flow rate (mllmin)
 Saliva

• Figure 7. Oral sugar clearance. 100 -,-------;-------------,

'2
Oral sugar c1earance
leve! of sugar) as a function
(to a very low
of the un-
·s'-'
Q)
stimulated saliva flow rate calculated
from a mathematical model (Dawes,
.5
.•... Normal unsumulated
saliva Ilow rate
Q)
1983). In this model the volume of sal- u
50
iva just before swallowing as well as the
volume of saliva swallowed are set to a ~
u
Q)

constant leve!. As shown the oral sugar

clearance is highly dependent on the
unstimulated saliva flow rate increas-
o +-------~--~---.--------.-------~
ing substantially when the
lated saliva flow rate is below 0.2 mI!
unstimu-
o 0.2 0.4 0.6 0.8
rmn. Saliva flow rate (ml/min)

can both absorb HC03 - (at low flow rates) stimulated saliva collected during the first
and secrete it (at high flow rates). few minutes has a composition that is differ-
An essential aspect of the HC03 - buffer ent from that of saliva collected following
system is that it can diffuse freely across the several minutes of sustained stimulation.
epithelial boundaries in form of gaseous CO2 Furthermore, many solutes show circadian
driven by the CO2 gradient within the differ- variations that are independent of the vari-
ent gland compartments. However, because ation in salivary flow rateo
the partial pressure ofC02 (PC02) in saliva re-
mains almost constant at a11flow rates and at
various metabolic rates, its HC03 - concen- • Oral clearance
tration (and thereby pH) increases with flow
rate (see Figure 4 and later for the equilibrium The time taken to clear the food that en-
of the HC03 - buffer system). ters the mouth varies from one subject to
The membrane transporters believed to another (Figure 7). This clearance is pri-
govern the HC03 - concentration in saliva marily due to swallowing and the flushing
may comprise ductal cr /HC03 - ex- effect of saliva. The dilution of substances
changers. In parotid saliva, for example, in the mouth and their removal is crucial
HC03 - concentration correlates positively to for the protection of the oral tissues. lf
that of Cl". Accordingly, at low flow rates the substances that are harmful to teeth such
CI- and HC03 - concentrations and pH are as sugars or acids accumulate in the
low, and vice versa at higher secretion rates. mouth, destructive processes such as dental
Figure 6 shows that as flow rate increases, caries or erosion are accelerated. The nor-
the whole saliva concentrations of Na +, cr , mal procedure for measuring oral clearance
HC03 -, total calcium and total protein in- is to introduce a certain substance (e.g.,
crease to varying extents, while the concen- sugar) into the mouth in high concen-
tration of K+ and total phosphate decreases. tration after an initial swallow. The sub-
However, the saliva flow rate and compo- stance is then diluted by the 0.8 mi or so
sition also depend on the type of gland from of saliva that remains in the mouth after
which it is secreted, as we11as on the nature swallowing (the so-called residual volume).
and the duration of stimulation. Thus, The oral clearance rafe is the time taken

29
 Bardow, Lynge Pedersen and Nauntofte
either to clear the substance from the
mouth or to reduce it to a very low con-
centration. This is strongly dependent on
the saliva flow rate as well as on the vol-
ume of saliva in the mouth before and
after swallowing.
When a substance is introduced to the
mouth in a high concentration it will, de-
pending on its taste, initially stimulate saliva
production. The resulting increase in saliva
flow rate will then in crease the volume of sal-
iva in the mouth. When this volume reaches
the threshold for swallowing (about 1.1 ml),
a swallow will occur, removing part of the in-
gested substance. The remainder will then be
diluted with new saliva coming from the
glands until the threshold is again reached
and the next swallow occurs. Each time the
subject swallows, some of the substance will
be removed which will in turn reduce the
stimulus to secretion. Accordingly, oral clear-
ance occurs non-linearly with time especially
during the first minutes after introduction of
the substance to the mouth. When the sub-
stance in the mouth has reached a certain low
level, it will no longer stimulate saliva flow
and the production of saliva will return to the
unstimulated level. Hence the unstimulated
flow of saliva has the greatest overall impact
on the oral clearance time (Figure 7) beca use
it is present throughout the longest period of
the clearance time. When this is less than 0.2
ml/min, the time taken to clear sugar from the
oral cavity increases dramatically. Therefore
individuals with impaired unstimulated sal-
iva flow rates have a slow oral clearance time
and are at higher risk of developing dental
caries and erosion.
• Saliva buffer capacity
One of the important roles of saliva is to
maintain a non-harmful pH in the mouth
30
during the time taken to clear acids that
are ingested orally, such as carbonated
drinks, fruit juices, and wines. The ability
of the saliva to maintain the pH when
exposed to acids is termed buffer capacity.
This is determined in the laboratory by
titration of the saliva within the pH range
of interest. The saliva pH, which in healthy
individuals varies between 6.0-7.5, depends
strongly on the secretion rate with the
most alkaline fluid being secreted during
stimulated flow. A drop in saliva pH below
5.5-5.0 is potentially harmful to the oral
tissues, particularly enamel/dentine. Thus
from a dental point of view the most inter-
esting pH range for determination of the
buffer capacity is from the saliva's original
pH in the mouth down to pH 5. Typical
titration curves illustrating the saliva pH
as a function of added acid (HCI) are
shown in Figure 8. From such titration
curves the saliva buffer capacity W) can be
determined in mmol H+ /(litre saliva· pH
unit) in a certain pH interval:
Where ~CA is the increase in saliva acid con-
centration and ~pH is the change in saliva
pH caused by the addition of acid.
If addition of large amounts of acid re-
sults in only a minor pH change, the buf-
fer capacity of the saliva is high and vice
versa. Note that, regardless of the concen-
tration of buffers, the buffer capacity will
gradually increase when the pH decreases
beca use of the logarithmic nature of the
pH scale (-log [H+]). Buffer capacity is
not to be confused with the term "buffer
effect" of saliva often used in dental litera-
ture. In the dental office, the buffer effect
is often determined by adding a fixed
amount of acid to a fixed amount of saliva
and then reading off the final pH. Al-
though this gives a handy clue about the
 Saliva

buffering capacity of the saliva, it gives no stimulated saliva at pH 6 is considerable

information about the buffers present in higher than that of tap water. The buffer ea-
the mouth or about variations in the saliva pacity of highly stimulated parotid saliva is
buffer capacity at different pH values. even higher than that of a relatively viscous
The buffer capacity of human saliva in- fluid, like milk. However, the buffer capacity
creases with increasing flow rates and is of human saliva is never as high as that of
maximal around pH 6. Figure 8 shows that blood, which is partIy due to the high pro-
the buffer capacity ofboth unstimulated and tein concentration of blood. The three im-

Titration of whole saliva with strong acid

o
8

----------------------
®
• Figure 8. Titration
oi saliva.
and buffer capacity uws sws ©
4
Titration curves of unstimulated and
stimulated human whole saliva. As O 30
shown stimulated whole saliva (SWS)
is better in maintaining its pH upon mmol H+ / litre saliva
addition of acid compared to unstimu-
lated saliva (UWS) indicating a higher
buffer capacity. The buffer capacity in
(A) originates mainly from the salivary
contents of phosphate, in (B) from the
salivary contents of bicarbonate, and in
(e) from the salivary proteins. The
area where both curves have the f1attest
slope, and therefore the highest buffer
capacity, is around pH 6 indicated at Buffer capacity at pH 6
the figure. Below is shown the buffer
capacity at pH 6 of unstimulated and
70
stimulated
stimulated
whole
parotid
saliva as
saliva (SPS) in corn-
well as e 60
·CIi0 50
parison to the buffer capacity of other
f1uids. It can be seen that the buffer ~
u
40
capacity of saliva is higher than the
~ 30
buffer capacity of water at a11 times,
that the buffer capacity of stimulated
;:l 20
~
parotid saliva is higher than that of 10
milk, and that the buffer capacity of
saliva never becomes as high as it is in O
blood. uws sws sps water milk blood

31
Bardow, Lynge Pedersen and Nauntofte
portant buffer systems in human saliva are
the bicarbonate, the phosphate, and the pro-
tein buffer systems.
The bicarbonate buffer system and saliva
pH regulation
The concentration of bicarbonate, the main
buffer in human saliva, varies under physio-
logical conditions from about 2-5 mM in
unstimulated saliva up to plasma-like levels
of about 28 mM in stimulated saliva. The
equilibrium for the bicarbonate buffer sys-
tem in a partly open compartment like the
mouth is:
CO2 + H20 ~ H2C03 H HC03 - + H+
Where CO2 is the CO2 in saliva and in the
air surrounding the saliva in the mouth and
CA is carbonic anhydrase that catalyses the
hydration of CO2 to carbonic acid.
The bicarbonate buffer system makes .its
highest contribution to the overall buffer ca-
pacity of saliva at the pK value for carbonic
acid that is close to pH 6 in saliva (Figure 8).
At pH 6, the contribution of the bicarbonate
buffer system to the overall buffer capacity
varies from 50% in unstimulated/resting sal-
iva up to more than 90% in stimulated saliva
produced at high flow rates. This difference
is due to the flow-dependent variations in
the saliva bicarbonate concentration.
Figure 4 shows that in the normal pH range
of the parotid saliva (6.0-7.5) the partial
pressure of CO2 (P e02) is 6.0 kPa, which is
equal to that of blood. However, when saliva
enters the mouth, some CO2 is lost which re-
sults in a drop in its Pe02 to around 4.5 kPa.
Nevertheless, the Peo2 of the whole saliva is
still more than 1000 times higher than the
Peo2 in the atmosphere (4 Pa). Accordingly,
any exposure of saliva to the atmosphere
(during saliva collection or during sialochem-
ical measurements) will result in major losses
of saliva CO2 that decrease the concentration
32
of the whole bicarbonate buffer system and
increase the salivary pH shifting the equilib-
rium to the left. CO2 is also lost from the sal-
iva if it is mixed with acid because the equilib-
rium of the bicarbonate buffer system shifts
to the left, which results in an increased sali-
vary Peo2. Thus acidification to pH 4.0 of
whole saliva containing 25 mM of bicarbon-
ate results in a 25-fold increase in its Peo2
level. Such acidification will give rise to a P e02
in saliva that is 25,000 times higher than the
Pe02 in the atmosphere. Accordingly, the
driving force for CO2 loss to the atmosphere
from such acidified saliva is as high as for car-
bonated drink exposed to the atmosphere.
The ability of the bicarbonate buffer system to
go from one form to another during buffering
is called phase buffering and this ability
further increases the buffering of the system.
The saliva bicarbonate concentration is
most commonly determined from the saliva
Peo2 and pH by the Henderson-Hasselbalch
equation:
HC03 - = (0.225 Peo2) (lOPH-pK)
Where P e02 is the saliva P e02 in kPa, and
pK the pK value of carbonic acid in saliva.
To summarise: in order to determine the
bicarbonate concentration and pH of saliva as
well its buffer capacity accurately, it is essen-
tial to avoid the loss of CO2• To achieve this,
the saliva has to be collected and analysed in
closed systems using techniques similar to
those used for arterial blood samples.
The phosphate buffer system
Figure 6 shows that the total phosphate con-
centration in saliva also depends on the flow
rateo However, while the salivary bicarbonate
concentration increases with increasing flow
rates, the saliva total phosphate concen-
tration decreases with increasing flow rates.
Thus resting/unstimulated saliva may con-
tain up to 10 mM of total phosphate,

whereas stimulated saliva secreted at high
flow rates may contain well under 3 mM.
The different ionic forms of phosphate are
determined by their pK values and thus by
the pH value of the saliva (Figure 9).
Within the normal pH range of the saliva
(6.0-7.5), most phosphate will be present as
dihydrogen phosphate (pK around 6.8 in sal-
iva), and hydrogen phosphate. However, the
two other forms of phosphate (i.e. H3P04
and POl-) will also be present in the pH
range from 6.0-7.0, although in very low
concentrations. Accordingly, in saliva phos-
phate is most effective as a buffer at pH 6.8
(Figure 8).
The phosphate buffer system normally
contributes about 50% of the buffer capacity
in unstimulated/resting saliva. However, due
to the flow-dependent decrease in saliva
total phosphate concentration upon stimula-
tion together with the huge increase in the
bicarbonate concentration, phosphate nor-
mally makes only a minor contribution to
the stimulated saliva buffer capacity.
The protein buffer system
Saliva contains a variety of different proteins
with specific biological functions (Figure 10).
Most of these can act as buffers when the pH
is above or below their isoelectric point (pl).
Below their pI they can accept protons and
thereby act as buffers, and above they can re-
lease protons. Because the pI of most salivary
proteins is around pH 7.0, their buffering ef-
fect becomes pronounced mostly at acidic
and alkaline pH values. Thus, they contribute
substantially to the saliva buffer capacity at
pH values ofless than pH 5 (Figure 8). In ad-
dition to their chemical buffering, some of
the saliva proteins also increase the viscosity
of the saliva when the pH becomes acidic and
thereby cover and physically protect the teeth
against the acid loado
Saliva
• Saliva and its saturation with
respect to hydroxyapatite
Under physiological conditions, teeth do not
dissolve in saliva beca use it is supersaturated
with respect to hydroxyapatite CalO(P04)6
(OHh, the main inorganic component of
teeth. The solubility product for hydroxyapa-
tite (KSPHA) is 10- 117 MI8 (where 18 refers to
the 18 ions present in the hydroxyapatite unit
cell). This value corresponds to the ion activ-
ity product for hydroxyapatite (IAPHA) in a
solution saturated with respect to hydroxy-
apatite.
For saliva the IAPHA can be calculated
from its actual salivary ion activities (the free
ionic concentration corrected for electro-
static effects) of Ca2+, PO/-, and OH- as
shown below:
From this expression it can be seen that
IAPHA increases with increasing activities
of these ion s in saliva and vice versa. The
greatest impact on IAPHA, however, comes
from the saliva pH. Thus when pH de-
creases, the activities of both PO/- (see
Figure 9 for the equilibrium of the phos-
phate buffer system) and OH- will de-
crease dramatically.
When KSPHA and IAPHA are known the
degree of saturation of saliva with respect
to hydroxyapatite (DSHA) can be calculated
as:
DSHA = (IAPHA/KSPHA) (1/18)
where (l/L8) al so refers to the 18 ions present
in the hydroxyapatite (i.e. CalO(P04)6(OH)z)
unit cell.
If IAPHA in saliva is larger than KSPHA,
(i.e. DSHA> 1), the saliva is supersaturated
with respect to hydroxyapatite. However, if
IAPHA is smaller than KSPHA (DSHA< 1), the
saliva is undersaturated and the enamel/den-
33
 Bardow, Lynge Pedersen and Nauntofte
• Figure 9. The phosphate buffer system. 100% ~;o:------~o;;;;::-----;---~-..;:------..,
The equilibrium of the phosphate buf-
fer system as a function of pH. In the
normal pH range for saliva most phos-
phate will be present as H2P04 - and
HPO/-. However, the two other
forms of phosphate (i.e. H3P04 and
PO/-) will also be present although
in very low concentrations.
O%L------~---~--_.~~~---._--~
tine will dissolve, leading either to dental
erosion or dental caries, depending on the
origin of the acidic challenge.
Calculation "case" of DSHA in saliva
A patient has an unstimulated saliva
flow rate of 0.3 ml/min and a saliva
pH of 6.0 (OH- activity of lO-s M).
The saliva total calcium concentration
is 1 mM corresponding to a Ca2+ ac-
tivity of OAX 10-3 M and the total
phosphate concentration is S mM cor-
responding to a PO/- activity of
1.7X 10-10 M
IAPHA of saliva=(OAX 10-3)10
(1.7X 10-10)6 (1 X 1O-S)2=2.5X 10-109
MIS
Thus in this case IAPHA is larger than
the corresponding KSPHA of 10-117
MI8
DSHA = (2.5X 10-109/10-117) (1/1S)=2.9
Thus in this case DSHA becomes > 1
and therefore the saliva is supersatu-
rated with respect to hydraxyapatite
34
4 12
~8
pH
Normal saliva pH range
The value of pH in saliva at which IAPHA
equals KSPHA (i.e. the condition where
DSHA = 1) is often denoted the "critica]" pH
in the dental literature. The critical pH in
human saliva is on average 5.5 under normal
physiological conditions, but this is certainly
not a fixed value since it depends on a num-
ber of ion activities that change dynamically
as the saliva flow rates varies. Unstimulated
saliva containing a high phosphate concen-
tration, and therefore having a high phos-
phate activity, has generally a more acidic
critical pH than stimulated saliva containing
a low phosphate concentration. The critical
pH in saliva normally varies from 5.3 to 5.5
depending on the flow rate: however, it may
vary frorn 5.2 to 5.8 in extreme cases where
the phosphate concentration is very high or
very low, respectively.
Nucleation followed by precipitation of
calcium-phosphate salts is likely to occur in
a fluid that is supersaturated with these ions.
Three-dimensional nucleation is the nu-
cleation of the calcium-phosphate salt
within a fluid followed by crystal growth of
the salt (homogeneous and heterageneous
nucleation). Two-dimensional nucleation is
surface- or seed-induced nucleation of the
 Saliva

calcium-phosphate salt followed by crystal rates result in high total protein concen-
growth of the salto Three-dimensional nu- trations). However, the total protein concen-
cleation is not likely to occur in saliva be- tration and composition is much less de-
cause of the low calcium and phosphate con- pendent on the flow rate than are the inor-
centrations as well as the many proteins that ganic components. Instead, it is mainly
act as nucleation inhibitors. Two-dimen- influenced by individual genetic differences.
sional nucleation, however, occurs quite Hence, the profile of saliva proteins in
often in the form of dental calculus where monozygous twins is similar.
substances in the dental plaque and on the Our understanding of the functions of the
tooth surfaces serve as seeds for the process. saliva proteins is based on in vitro experi-
Calculus formation normally occurs on ments rather than clinical trials. So far there-
teeth in the regions of the mouth where the fore little is known about the impact of spe-
salivary supersaturation of calcium-phos- cific saliva proteins on oral health in vivo.
phate salts is highest, i.e. near the orifices of Some studies have shown that differences in
the submandibular, sublingual and parotid protein composition playa role in individ-
ducts. Here the pH is most alkaline due to uals' susceptibility to develop caries while
the high bicarbonate concentration in the other studies have not been able to demon-
secreted saliva, as well as the CO2 evapor- strate this.
ation due to mouth breathing. Accordingly, Mixing dried saliva proteins with water
calculus formation occurs most frequently gives a solution with a texture and viscosity
around the mandibulary incisors and maxil- that is similar to saliva. Not surprisingly,
lary molars. therefore, the distinct texture and viscosity
of saliva is the result of its protein compo-
sition. Some saliva proteins protect the oral
tissues against infections, others coat and lu-
• Organic saliva composition and bricate the oral tissues, some playa role in
its functions maintaining high saliva concentrations of
calcium, other are digestive enzymes, and
Both the acinar and ductal cells secrete pro- some catalyse the hydration of CO2 to car-
teins into saliva (exocrine function) and the bonic acid. Although most have one major
blood circulation (endocrine function); see function, many have multifunctional roles.
also paragraph on stimulus-secretion coup- A few examples are discussed below in the
ling. These salivary proteins are the major context of their function and possible clin-
organic components of saliva. There are ical impacto
more than 40 different proteins in human
saliva whose molecular weights vary from a Digestive enzymes
few kDa to more than one thousand kDa. Human saliva contains u-amylase, an en-
The total protein concentration is on average zyme that hydrolyses the a-l,4 glycosidic
2.0 mg/ml saliva, which is nearly 40 times linkages of starch molecules. Hence the
lower than in plasma. The protein concen- breakdown of ingested starch to simple hex-
tration depends on both the duration of oses occurs in two phases starting in the oral
stimulation (long periods of stimulation re- cavity with salivary n-amylase and continu-
sults in high saliva total protein concen- ing in the intestine with pancreatic o-amy-
trations) and on the flow rate (high flow lase. As shown in Figure l O, the salivary

35
 Bardow, Lynge Pedersen and Nauntofte

Salivary proteins (size and function)

A) Anti-bacterial
1000 -
B) Anti-viral
C) Anti-fungal
D) Tissue coating
E) Lubrication
F) Mineralization
G) Digestion
,-.. 100
H) Buffering
ro
9
"--"
A A
B
A A A A
B B D
a)
N D D G
E E
l
(/)
G G 11 D
10
F

A A D A
13 E e
D F F
I 1- ,~ -, 11

-<en N V> c: V>

'"
V>
c: '"
c:
.:
V>

0'::'"
c:

"6
Q) V>
C) -;;;
V>
E '"
V>
ee -O'" 00; ~
°C
-¡¡;
6 "'"
o¡;;
<2:! :;;.,
>. 2a. N
o
S '"
.J::
-¡¡; Ul
2 ..c
.3
'" '"
>.
N
e .s
u
~ c: u c/í i
t:
0<3
t:
0<3 e,
'" '"
u
<: ~
2'" ~ o
..o '"
e

u
¡;¡
e
e,

• Figure 10. The majar salivary proteins.

Some of the major salivary proteins ranked aeeording to their molecular weight (please note that the seale is logarith-
miel. For eaeh protein its possible funetion in saliva is stated. Thus the high molecular weight muein (MG 1) IS
shown to have (A) anti-baeterial, (B) anti-viral, (O) tissue coating, (E) lubrieating, and (G) digestive effeets.

u-amylase family has molecular weights of can result in elevated plasma o-amylase
around 55-60 kDa and they are therefore concentrations beca use it leaks from the
among the larger proteins found in saliva. salivary glands into the blood circulation.
The a-amylases are secreted particularly Salivary o-amylases are active above pH 6
from the serous cells in the parotid and and become inactivated by the low pH in
submandibular glands in response to para- the stomach. Since this limits the time
sympathetic stimulation and the rx-amy- available for salivary o-arnylases to work,
lases are secreted both to blood circulation it is considered of minor importance in
and the saliva. The concentration of a- starch digestion. It may however be im-
amylases in saliva is high and may consti- portant for starch digestion in neonates
tute as much as 30-40% of the total pro- with an insufficiently developed pancreas.
tein in whole saliva. This is more than Human saliva also contains lipase, an en-
1000 times higher than the total o-arnylase zyme able to break down dietary triglycer-
concentration in human blood. Therefore ides. Salivary lipase is secreted from the se-
injuries to the salivary glands due to ir- rous cells in the parotid gland and from the
radiation, glandular obstruction, or surgery von Ebner's glands on the tongue. Like sali-

36

vary n-amylase, it is considered of minor
importance in healthy individuals, but may
play a role in neonates and patients with
pancreatic dysfunction.
Proteins with lubricating functions
Human saliva contains mucins that are large
glycoproteins, constituting more than 40%
carbohydrate, which have lubricating func-
tions on the oral tissues. Two specific types
of mucins have been characterized in human
saliva. MG1 is produced in the mucous acini
in the submandibular and sublingual glands
as well as in the palatal and labial glands.
MG2 is produced in the serous ce11sin most
glands except for the parotid. The MG1 mu-
cin weighs more than 1000 kDa and is the
largest protein present in saliva, whereas
MG2 weighs 130 kDa (Figure 10). Due to
the differences in weight, the two mucins are
often referred to as high- and low- molecu-
lar weight mucins.
Mucins are hydrophilic and contain water:
they work effectively to moisten and lubri-
cate oral surfaces. They also form disulfide
bridges with other mucin molecules and
thereby create a network that covers and
protects the oral tissues. The hydrophilic and
network-forming abilities of salivary mucins
give saliva its distinct texture and viscosity.
Calcium binding proteins
Human saliva under normal physiological
conditions is always supersaturated with hy-
droxyapatite as well as with other calcium
phosphate salts. However, these salts do not
norma11y precipitate in saliva due to the
presence of calcium-binding proteins. Sta-
therin is an asymmetrical protein and one of
the sma11est proteins in human saliva (4-5
kDa), which is secreted from acinar ce11s.
Statherin inhibits three-dimensional as we11
as two-dimensional nucleation and prevents
the formation of calcified masses (sialoliths)
Saliva
in the salivary ducts and glands, and reduces
the formation of calculus. The acidic proline-
rich proteins (PRPs) are also sma11molecules
(10-30 kDa) that constitute around 30% of
the proteins parotid and submandibular sal-
iva. Like statherin, the PRPs are secreted from
acinar ce11s.They inhibit two-dimensional
nucleation by binding calcium and also bind
tannins that are harmful to the oral tissues.
Interestingly, there appears to be differences
in the amount and quality of acidic proline-
rich proteins in caries-free and caries-active
individuals. Although statherin and the PRPs
are the proteins mostly associated with inhi-
bition of calcium-phosphate salt nucleation,
other proteins like the histatins and cystatins
also inhibit nucleation.
Carbon dioxide hydration
Carbonic anhydrase (CA) is an essential en-
zyme present in a11ce11s.Carbonic anhydrase
catalyse the reversible hydration of CO2 to
carbonic acid (see equilibrium for the bicar-
bonate buffer system). Several isoforms of
CA have been characterized of which one
(CA VI) that has a weight of around 42-45
kDa is present in saliva. In the parotid and
submandibular glands the acinar ce11sse-
crete CA VI. In contrast to the we11-known
cytoplasmic forms of CA, CA VI is the only
isoform of the enzyme that is known to oc-
cur in a secretory formo Some studies have
suggested that CA VI isoforms playa protec-
tive role against dental caries.
Saliva proteins with antirnicrobial
functions
Most immunoglobulins in human saliva are
in the form of secretory IgA (sIgA) that is a
large hydrophilic protein (380 kDa). Salivary
IgA is a product of plasma ce11sthat is modi-
fied and secreted by acinar and duct cells in
the salivary glands. This secretory imrnuno-
globulin is a specific defence factor that is
37
Bardow, Lynge Pedersen and Nauntofte
stimulated by the presence of bacteria. Be-
cause of its hydrophilic abilities, salivary IgA
can mix and function in saliva where it ag-
gregates bacteria. Salivary IgA also has affin-
ity for other antimicrobial components in
saliva like the mucins. This affinity may syn-
ergisticaUy increase the antimicrobial aggre-
gating effects of both proteins. Some studies
have shown a protective effect of salivary IgA
against dental caries. However, other studies
have not been able to show such relation-
ships.
In addition to specific antimicrobial de-
fence mechanisms, human saliva contains a
variety of unspecific or innate antimicrobial
defence mechanisms that have attracted in-
creasing interest in recent years. Mucins, in
addition to their lubricating effect on the
oral tissues, also give protection against mi-
crobes. Thus the coating and network-form-
ing effects of the mucins function as a bar-
rier protecting the underlying tissue. The
mucins also agglutinate large numbers of
bacteria in saliva and thereby increase their
removal by swaUowing.
Lysozyme and lactoferrin are examples of
proteins secreted by duct ceUs. In contrast to
acinar proteins, ductal proteins decrease in
concentration upon increased saliva flow.
Both these proteins belong to the innate anti-
microbial defence mechanisms. Lysozyme
(around 14 kDa) kiUs gram-positive bacteria
by breaking down their ceUwalls. Because of
its strong positive charge, lysozyme may also
be able to activate bacterial autolysin. Lacto-
ferrin is a relatively big protein (around 75-
78 kDa) whose best-known function is to
bind iron and thereby inhibit bacterial
growth by reducing the concentration of this
important co-factor for bacterial enzymes.
Besides having an iron-binding effect, lacto-
ferrin has hidden antimicrobial domains that
are liberated from the protein after proteo-
lytic activity. Thus lactoferrin is believed to
38
have bacteriostatic, bacteriocidal, fungicidal,
and antiviral properties. It has been reported
that low levels of lactoferrin are related to
high levels of potentiaUy pathogenic oral bac-
teria. Human saliva also contains the enzyme
peroxidase (75-78 kDa) another innate anti-
microbial defence mechanisms that is se-
creted by the acini. This enzyme catalyses the
oxygenation of thiocyanate (SCN-) to hypo-
thiocyanate (OSCN-) and thereby reduces
bacterial growth by blocking essential bac-
terial metabolic processes.
Histatins are smaU antimicrobial peptides
of around 3-4 kDa that are known mainly for
their potent lethal effect in vitro on oral fungi
like Candida albicans. Histatins are present in
high concentrations in parotid saliva. The
kiUing effect ofhistatin 5 on Candida albicans
depends on the ionic strength of saliva, and
becomes higher with lower salt concen-
trations such as those in unstimulated saliva
in which histatins bind to Candida albicans.
Histatins have also been shown to kiUbacteria
such as Streptococcus mutans.
Growth factors in saliva
Since ancient times, saliva from various ani-
mals has been applied to wounds to acceler-
ate the healing process. It is now known that
the wound-healing effect of saliva arises
from its intrinsic growth factors. These
growth factors are secreted from ducts cells
in the salivary glands. Some, like epidermal
growth factor (EGF) have a local effect. EGF
is a smaU protein found in the submandibular
and parotid glands whose output is increased
by mastication. When secreted in saliva, EGF
enhances healing of ulcers and plays a protec-
tive role in oesophageal mucosal protection.
Salivary EGF interacts with the innate salivary
protein defence mechanisms to form a mu-
cosal defence barrier. NGF that has stimulat-
ing effect on ganglionic function is also re-
leased from salivary glands.

Transforming growth factors (TGF-o: and
TGF-~) are other small proteins that have
been found in saliva, and which can cause
cell differentiation and growth. Finally,
fibroblast growth factor (FGF), a potent
regulator of wound healing, has also been
found in saliva.
• Formation and function of the
pellicle
Under normal conditions in the mouth, i.e.
with plenty of saliva, an organic film called
the acquired pellicle covers the teeth. The
term "acquired" refers to the fact that this
film develops on the teeth after they have
erupted and not, as assumed earlier, before
eruption. While the exact composition of
this film is not known, it is derived mainly
frorn salivary proteins. Its protective effect is, the purely chemical protection against acids,
however, well known from laboratory
studies in which teeth with and without a
pellicle have been exposed to acid. These
studies have shown that teeth covered by the
pellicle develop less demineralization than
teeth without the pellicle. Studies in which
pure hydroxyapatite crystals have been ex-
posed to acid suggest that formation of a • Other organic components in
salivary protein pellicle on the crystal sur-
faces may decrease the rate of dissolution by
more than 80%. The effect of differences in
saliva protein composition on the protective
effect of this pellicle is not yet fully under-
stood. Nevertheless, the ability of the sali-
vary proteins to form the acquired pellicle is
certainly one of the major mechanisms by
which saliva protects the teeth. Moreover, a
recent study has shown an inverse relation-
ship between the salivary pellicle thickness
and the incidence of cervical erosive lesions,
which suggests that dental erosion may also
be depend on the pellicle-forming ability of
saliva in a site-specific pattern.
Saliva
When completely clean teeth are exposed
to saliva, the pellicle begins to form in less
than a minute and reaches equilibrium
within 2 hours. However, a so-called matu-
ration time of several days is necessary for
it to reach its maximum pratective capacity.
Because the surface of enamel is negatively
charged, the first layer that is formed (the
hydration layer) has a positive charge arising
mainly from calcium. This positively-
charged hydration layer then attracts nega-
tively-charged macromolecuJes. The prateins
that are best known to be attracted to tooth
surfaces and which are present in the pellicle
are the acidic proline-rich proteins, sIgA, cy-
statin, MG 1, lactoferrin, lysozyme, and amy-
lase. The reason why some of the other sali-
vary proteins have not been shown in the
pellicle can be explained by their weaker
ability to bind to hydroxyapatite. Apart from
the acquired pellicle also seems to determine
the initial attachment of bacteria onto the
tooth surface. The acquired pellicle may
thereby favour a non-harmful colonization
pattern and hence further protect the teeth.
saliva
In addition to the proteins, there are numer-
ous other organic components in saliva, in-
cluding the circulating adrenal glucocortico-
id cortisol. The concentration of cortisol in
saliva reflects its concentration in plasma, al-
though its concentration is 10-30 times
lower in saliva (typically 10-20 nM). Under
conditions of high stress, the salivary cortisol
can in crease above 30 nM. Since a collection
of saliva is non-stressful compared with
blood sampling, saliva has become widely
used in measurements of cortisol for moni-
toring stress. Moreover, the cortisol found in
39
 Bardow, Lynge Pedersen and Nauntofte
saliva is unbound; that is, it is not combined
with its carrier protein, so that its concen-
tration is a good indicator of its bioavail-
ability.
Saliva also contains many of the sex hor-
mones present in blood. For example, the
salivary concentration of estriol in pregnant
women also correlates well with its plasma
concentration and can therefore be used to
monitor fetal well-being.
Under normal conditions, the glucose
concentration in saliva follows that of
plasma, although at much lower levels. Thus
glucose is normally present in saliva only in
very low concentrations less than 0.1 mM.
However, patients with high plasma glucose
frorn untreated diabetes have elevated sali-
vary glucose concentrations that can reach
nearly 1.0 mM. Since glucose serve as a bac-
terial substrate such patients are likely to de-
velop dental caries at high rates.
Human saliva normally contains between
2 and 4 mM urea, which is a praduct of pra-
tein metabolismo Urea is not a buffer, but in
the presence of bacterial urease it can alka-
lise pH by being broken down into CO2 and
ammonia (which picks up a proton). Pa-
tients who suffer frorn untreated renal fail-
ure have salivary urea concentrations as high
as 85 mM. The resulting high pH of the sal-
iva and plaque in these patients reduce the
incidence of dental caries.
Finally saliva contains blood group sub-
stances fram the ABO blood graups. Before
genotyping by DNA testing, the presence of
these substances in saliva was an important
tool in forensic and palaeoserological science.
• Saliva as a screening tool
Saliva is a convenient fluid to use for diag-
nostic purposes and has obvious benefits
over plasma beca use it is easy to collect in a
40
non-invasive and non-stressful manner in
the field. The ability of some circulating
drugs to enter the saliva by diffusion is of
special interest in forensic and pharrnaceut-
ical science and in some legal situations.
Only the free and biologically active fraction
of a drug in plasma gets into saliva where
the concentration of prateins is much lower
compared to plasma. When a water-soluble
drug enters saliva that has a lower pH than
plasma, it ionises. Its concentration in saliva,
and thus the ratio of the free drug concen-
tration between plasma and saliva, is then
determined by its pKa, the pH of the saliva
and the pH of plasma. The wide variation
in the rate of salivary secretion and the re-
sulting variation in pH limit its use as a di-
agnostic medium for some drugs (depend-
ing on their pK). Hence, some drugs are
concentrated in saliva while others are pres-
ent in much lower concentrations than in
plasma.
However, for some illegal drugs, the issue
is detection rather than quantification, and
saliva pravides a simple screening assay for
this purpose. The salivary levels of "rec-
reational" drugs like cocaine and marijuana
as well as alcohol give a good reflection of
their plasma levels. Tobacco praducts like
nicotine and cotinine can also be measured
in saliva, and correlate well with their
plasma concentrations. This has made it
possible for the life insurance industry to
verify the smoking status of individuals by
means of salivary tests. Pollutants like lead,
cadmium and copper can also be detected
in saliva and their concentrations show the
extent of the individual's exposure to them.
In another domain, saliva can be used to
determine the profile of infection of the oral
cavity with pathogens such as Candida as well
as giving LactobaciLLus and Streptococcus mut-
ans scores. Finally, saliva is also valuable
for qualitative screening (+ / -) for much

more serious systemic infections diseases like
HIY.
• Salivary gland dysfunction
Salivary gland dysfunction is defined as any
quantitative and/or qualitative change in the
output of saliva. Thus, salivary gland dys-
function includes either an increase in sali-
vary output (hyperfunction) or a decrease
(hypofunction). Genuine salivary hyper-
function, sialorrhoea, is relatively uncom-
mon in adults. It may be caused by mucosal
irritation or be idiopathic. Drooling often
occurs as a result of an underlying neuro-
logical disorder, for example cerebral palsy,
amyotrophic lateral sclerosis, and Par-
kinson's disease. Drooling may also be seen
in mentally-handicapped individuals and as
a side-effect of neuroleptic medication in
adults. The problem is generally the result of
decreased swallowing efficiency and fre-
quency and is rarely related to genuine sali-
vary hyperfunction. Drooling may, however,
be asevere problem to the patient, since it
can cause maceration of the skin at the
angles of the mouth and on the chin fol-
lowed by colonization of opportunistic
microorganisms. Furthermore, constant
drooling may have a negative impact on the
patient's psychological well-being and social
behaviour. Some patients complain of
having too much saliva and develop an ob-
sessive swallowing pattern, i.e. a need to
swallow compulsively at frequent intervals,
but objectively are found to have a normal
production of saliva.
Salivary gland hypofunction, in contrast,
is a common condition especially in people
with other underlying diseases and in those
who take certain types of medication. Sali-
vary hypofunction may develop into hypos-
alivation, a term which is based on objective
Saliva
measures of the saliva production (see sec-
tion on collection of saliva). Hyposalivation
usually results in altered salivary compo-
sition and is often associated with xerostomia
which is defined as the subjective feeling of
dry mouth. Generally, this symptom occurs
when the unstimulated salivary flow is re-
duced to approximately 50% of its normal
value in any given individual, which means
that more than one major salivary gland
must be affected. Unstimulated whole sali-
vary flow rates are more closely associated
with the symptoms of xerostomia than
stimulated rates. However, complaints of
xerostomia can occur in the absence of hy-
posalivation and vice versa. In mouth-
breathing, for example, xerostomia is related
to mucosal dryness arising from increased
evaporation of saliva. It should be empha-
sized that xerostomia is not solely related to
decreased salivary flow rate, but also to
changes in its composition.
• Principal causes of salivary gland
hypofunction and xerostomia
Several medical conditions and medications
can affect the salivary glands and their secre-
tions in a number of ways. The salivary re-
flex can be affected vía effects on the central
or peripheral neural regulation, or through
the receptor systems in the salivary gland
tissues, stimulus-secretion coupling, mem-
brane transporters, protein synthesis and/or
protein release mechanisms. The most com-
mon reasons for salivary gland hypofunction
are listed in Table 3.
Age and hormonal changes
Salivary dysfunctions in children are rare
and mostly temporary conditions. The adult
levels of salivary flow rate are usually
41
Bardow, Lynge Pedersen and Nauntofte

Table 3. Common causes of hyposalivation and/or changes in saliva composition.

Iatrogenic
Autoimmune diseases
Neurological disorders
Hormonal disorders
Infections
Hereditary disorders
Metabolic disturbances
Local salivary diseases
Other conditions
medications, e.g. antidepressants, diuretics, antihistamines,
antihypertensives, antipsychotics and opiates, polypharmacy, chemotherapy;
radiotherapy to the head and neck region, surgical trauma
rheumatoid arthritis, Sjogren's syndrome, sarcoidosis
mental depression, cerebral palsy, Bell's palsy, Holmes-Adie syndrome
diabetes mellitus (Iabile), hyper- and hypothyroidism
HIV infection/AIDS, epidemic parotitis
cystic fibrosis, ectodermal dysplasia
malnutrition, eating disturbances, bulimia, anorexia nervosa, dehydration,
vitamin deficiency
sialolithiasis, sialadenitis, carcinoma
menopause, impaired masticatory performance

reached by the age of 15 years. In healthy
individuals, the stimulated whole saliva flow
rate does not decline with age. However, an
age-related decrease may occur in secretions
from minor and submandibular glands, but
not from the parotids. This functional de-
crease is consistent with age-related changes
of the morphology of these glands in which
the acinar volume may shrink by 40-50%.
On the other hand, diverse morphological
changes can occur in the various salivary
glands with aging including increased adi-
posity, ductal proliferation and fibrosis.
Complaints of oral dryness are common in
older adults and may be explained by the
age-related decrease in minor and submand-
ibular gland secretions, even when chewing-
stimulated whole saliva secretions are nor-
mal. It should be emphasised that, even
though changes in the structure of the sali-
vary gland might suggest hypofunction,
there is no clinically significant reduction in
the overall gland output with aging in
healthy, non-medicated adults. Accordingly,
the increased incidence of subjective symp-
toms and objective signs of salivary gland
hypofunction in the elderly is more likely to
be due to the increased incidence of systemic
diseases and the increased likelihood of
medication in this age group, than to age-
related salivary gland hypofunction.
Menopause is often associated with sali-
vary dysfunction, since the incidence of dry
mouth and burning mouth symptoms is high
among menopausal women. However, obser-
vations on the changes in flow rates and com-
position of saliva have given conflicting re-
sults. Generally, the salivary flow rates are not
significantly decreased in healthy post-
menopausal women compared with those of
premenopausal women. Concentrations of
phosphate in whole saliva are higher in post-
menopausal women than in controls. Hor-
mone replacement therapy increases salivary
flow rates, as well as its buffer capacity and pH
in postmenopausal women.
Iatrogenic-induced salivary hypofunction
Salivary gland hypofunction and xerostomia
are side effects of many prescription drugs,
including antidepressants, antihypertensives,
diuretics, antihistamines, antipsychotics and
opiates. Antidepressants and antihistamines
act on muscarinic cholinergic receptors in
salivary glands resulting in reduced saliva
volume, whereas other drugs such as di-

42

uretics may induce changes in the compo-
sition of saliva through their action on
mechanisms that control fluid and salt bal-
ance, and inhibitory effects on electrolyte
transporters in the salivary gland cells.
There is also a relationship between the
number of medications taken on a regular
daily basis, with xerostomia and hyposaliv-
ation, irrespective of whether the medi-
cations specifically cause hyposalivation.
Thus, patients taking more than four drugs
(polypharmacy) have lower unstimulated
and stimulated whole saliva flow rates than
non-medicated patients.
Radiotherapy of the head and neck region
results in a markedly reduced salivary flow
rate, acidic saliva, poor buffering capacity and
abnormal electrolyte and protein contento
The abnormal electrolyte and protein content
can be due to contribution from their leakage
from the plasma into saliva. The extent of sali-
vary hypofunction depends on the volume of
glandular tissue included in the field of radi-
ation as well as to the total dose of radiation
delivered. Hypofunction induced by doses of
30-50 Gray (Gy) may be reversible, whereas
doses higher than 60 Gy usually induce irre-
versible salivary gland hypofunction and per-
sisting xerostomia. Damage to the glandular
tissue is caused by direct radiation effects on
the acinar and duct cells, but also indirectly
by injury of vascular structures resulting in
increased capillary permeability, interstitial
oedema and inflammatory reactions. Serous
acinar cells are initially more affected by ir-
radiation than mucous acinar cells and duct
cells. Although the salivary glands are con-
sidered to be relatively resistant to irradiation
because of their relatively slow cell division
cycle, radiation induces acute changes in sal-
iva flow rate and composition. However, re-
sults vary, and the long-term effects of the
treatment on oral health are especially uncer-
tain. Finally, patients undergoing chemo-
Saliva
therapy may also experience similar changes
in salivary flow rate and composition.
Systemic diseases and medical conditions
Numerous systemic diseases, and/or the
medication used to treat them, are associated
with impaired secretion of saliva and changes
in its composition. Prominent examples are
autoimmune diseases such as Sjogren's syn-
drome (see textbox) and inherited diseases
like cystic fibrosis where changes in secretion
are in part related to structural changes to the
salivary glands. In neurological disorders like
Bell's palsy (idiopathic paralysis of the facial
nerve) and Holmes-Adie syndrome (auto-
nomic dysfunction), compromised inner-
vation of the glands results in salivary hypo-
function. In inadequately-controlled insulin-
dependent (type 1) diabetes or at the early on-
set of diabetes mellitus type 1 and 2, reduced
salivary flow rate may occur as a result of de-
hydration due to polyuria. The salivary dys-
function may, however, also be related to dia-
betic autonomic neuropathy, and the associ-
ated sialosis (non-inflammatory salivary
gland hypertrophy). Malnutrition, as well as
eating disorders (bulimia and anorexia nerv-
osa) oflong duration reduce salivary flow, in-
duce compositional changes of saliva and
cause enlargement of the major salivary
glands.
• Symptoms and objective findings
in salivary gland hypofunction
Reduced salivary secretion is usually associ-
ated with a sensation of a dry mouth that
persists throughout the day. The dryness
continues at night leading to disturbed sleep
because of the need for regular sips of liquid.
Furthermore, patients with salivary hypo-
function, regardless of the cause, often have
symptoms of burning mouth, difficulties in
43
Bardow, Lynge Pedersen and Nauntofte
Characteristics of Sjogren's syndrome
Definition:
Chronic inflammatory systemic auto-
immune disease that principally causes
dry eyes (keratoconjunctivitis sicca)
and dry mouth (hyposalivation) due
to by lymphocyte-mediated destruc-
tion of the glandular tissue. The syn-
drome has a primary and secondary
formo The latter is usually associated
with rheumatoid arthritis.
Demography:
All ethnic and racial groups, preva-
lence: 1-3%, 90% of the patients are
women in the age of 40-60 years
Aetiology:
Unknown. Probably multifactorial. In-
cluding interactions between genetic,
immunological,
neuroendocrinological, and environ-
mental factors
Symptoms:
Xerostomia, ocular dryness, arthralgia,
myalgia, fatigue.
Clinical findings:
Hyposalivation, keratoconjunctivitis
sicca, parotid enlargement, vasculitis,
dry skin, Raynaud's disease, purpura,
anemia and pharyngitis.
Diagnosis of the oral component:
Symptoms of oral dryness, sialometry .
(whole saliva), salivary scintigraphy,
sialography, labial salivary gland bi-
opsy (focal periductallymphocytic in-
filtration of the salivary gland tissue).
Supplement: sialochemical analysis of
glandular saliva.
44
chewing and swallowing dry foods, difficulty
in speaking, impaired sense of taste, acid re-
flux (heartburn) and halitosis. Removable
dentures may cause discomfort due to the
lack of mucosal lubrication. These distress-
ing sequelae can significantly diminish the
health-related quality oflife. Permanently re-
duced salivary secretion commonly results in
clinically evident changes in the oral soft and
hard tissues. The oral mucosa in patients
with hyposalivation is generally thin and
pale and without the usual glistening ap-
pearance. Saliva frequently appears thick and
foamy and can form whitish threads on mu-
cosal surfaces. The dorsal part of the tongue
looks dry and becomes lobulated with atro-
phy of the filiform papillae. In some cases,
the tongue surface appears fissured. The
most common symptoms and clinical signs
related to permanently reduced salivary flow
rate are shown in Table 4.
Salivary gland infections
Bacterial sialadenitis is a relatively rare con-
dition, which occurs especially in elderly pa-
tients who have reduced salivary flow due to
systemic diseases, medications or dehy-
dration. The condition is usually character-
ized by acute tender swelling of the salivary
gland, particularly the parotid gland, and in
some cases fever and malaise. In medically
or immunologically compromised patients,
the infection may even become life-threaten-
ing due to sepsis. It is assumed that the re-
duced salivary flow allows ascending mi-
crobial colonization of the duct, which pre-
disposes to the development of acute or
chronic suppurative infection. Only limited
information is available about the microflora
of the salivary duct during salivary gland hy-
pofunction, but the microflora in the excret-
ory duct system appear to be more extensive
and complex in patients with Sjogren's syn-
drome than in healthy individuals.
 Saliva

Table 4. Oral symptoms and signs often related to hy- and mostly in the submandibular duct. Most
posalivation and xerostomia.
are asymptomatic and unilateral. The predis-
Oral mucosal dryness and soreness posing factors are changes in saliva compo-
Burning oral sensation sition, infection or inflammation in the
Difficulties in speech (dysphonia)
ducts, or damage to the duct tissue.
Difficulty in chewing dry food
Difficulty in swallowing dry food (dysphagia)
Difficulty in wearing removable dentures Salivary gland tumours
Taste impairment (dysgeusia or hypogeusia) Ranked according to occurrence salivary
Acid reflux, nausea, heartburn gland neoplasms or tumours may occur in
Bad breath (halitosis)
1) the parotid gland, 2) the minor glands, 3)
Sensation of thirst
Dry, glazed and red oral mucosa the submandibular gland, and 4) very sel-
Lobulation or fissuring of the dorsal part of the tongue domly in the sublingual gland. The neo-
Atrophy of filiforrn papillae plasms can arise from both acinar and
Dry, cracked lips ductal cells as well as myoepithelial cells. The
Increased activity of caries with lesions on cervical, inci-
incidence of malignancy is low for the pa-
sal and cuspal tooth surfaces
Increased frequency of oral infections (e.g. recurrent
rotid gland, relatively high for the minor
oral candidiasis, angular cheilitis) glands, and very high for neoplasms in the
Additional non-oral symptoms may occur including
sublingual gland. Symptoms may include a
dryness of the skin, throat, nose and eyes as well as lump in the mouth, swelling in the face, pain
constipation, weight loss and depression. in the jaw or the side of the face, and weak-
ness of the face muscles.

Mumps is an acute systemic viral infec- Saliva and oral microflora

tion caused by paramy:x:ovirus. The parotid
glands, which are primarily affected, become
increasingly swollen and painful over a few
days. As the glands swell, the patients often
develop fever, headache and malaise. Mumps
in adult males may result in the develop-
ment of orchitis, an inflammation of the tes-
ticles, which can result in sterility. The dis-
ease can be prevented by the use of a vaccine
giving long-life immunity. Mumps is the
most common form of viral sialoadenitis,
but parotitis may also be caused other vi-
ruses such as cytomegalovirus and Coxsackie
A virus.
Sialoliths
Calcified masses within the salivary duct
called saliva stones or sialoliths are a rela-
tively common condition that affects as
many as 1% of the population. Sialoliths oc-
cur more frequently in women than men
Salivary gland hypofunction is associated
with impaired antimicrobial capacity of sal-
iva. Changes such as reduced oral tissue pro-
tein protection and lower saliva pH may fa-
vour an aciduric and acidophilic oral micro-
flora, which increase the risk of dental caries
and oral candidiasis. Thus, reduced salivary
flow and/or altered salivary composition are
associated with significant changes in the
oral microflora including increases in acido-
philic microorganisms such as Streptococcus
mutans, Lactobacillus, and yeasts. High sali-
vary Lactobacillus counts are strongly associ-
ated with low salivary secretion and the
number of lactobacilli in saliva is positively
correlated with caries activity.
Saliva and dental caries
Dental caries is the result of an interaction
between many different variables such as
oral hygiene, diet, oral microflora, and tooth

45
Bardow, Lynge Pedersen and Nauntofte
morphology. Nevertheless, several clinical
studies have shown that dental caries is a
common consequence of salivary hypofunc-
tion. In patients with low saliva flow rates,
carious lesions usually develop rapidly, espe-
cially in retention sites for dental plaque
along the gingival margin and adjacent to
dental restorations. Chronic hyposalivation
may even result in early loss of natural teeth
due to progressive caries. In patients with
hyposalivation, the increased caries activity
is mainly attributed to a reduction in oral
clearance. Thus, reduction in salivary flow
rates leads to impairment of the mechanical
flushing and cleansing action of saliva,
which results in slow elimination of oral
bacteria, dietary sugars and dietary acids, as
well as plaque accumulation on the tooth
surfaces (Figure 6).
Saliva and tooth wear
Tooth wear occurs as a result of abrasion
(pathologic wearing of teeth resulting from
an abnormal habit or abnormal use of abras-
ive substances orally), attrition (physiologic
wearing of teeth resulting from mastication)
and erosion (irreversible loss of dental hard
tissue due to a non-bacterial chemical pro-
cess). The aetiology of each of these con-
ditions is like dental caries, i.e., the result of
interactions between a number of different
variables, which make it difficult to make a
clinical differential diagnosis between
chemical and mechanical aetiology of tooth
wear. The relationship between dental ero-
sion and various salivary factors such as pH,
buffer capacity, concentrations of calcium
and phosphate, and mucins as well as un-
stimulated and stimulated salivary flow
rates, have been investigated. However, only
unstimulated salivary flow rate and saliva
buffering capacity have been shown to be
associated with the presence of dental ero-
sion. Thus, patients with unstimulated
46
whole salivary flow rate of 0.10 ml!min or
less have a higher risk of dental erosion
than patients with higher unstimulated sal-
iva flow rates.
Saliva and periodontal disease
A few studies have demonstrated a relation-
ship between salivary flow and periodontal
parameters such as pocket depth, gingival
bleeding, dental plaque, and the extent of
attachment loss. However, most studies do
not support a causal relationship between
periodontal diseases and reduced salivary
flow rates even in patients with Sjogren's
syndrome who have long-term reductions in
saliva flow rates. However, impaired mech-
anical flushing and cleansing due to reduced
salivary flow may accelerate accumulation of
dental plaque that in turn may lead to gingi-
vitis.
Saliva and gastrointestinal functions
Reduced salivary flow results in decreased
bicarbonate concentration in the swallowed
saliva and therefore decreased neutralization
and clearance of gastric acid that enters the
lower oesophagus as the result of normal re-
flux activity: this may increase the risk of
developing gastro-oesophageal reflux and
gastric ulceration. Furthermore, mucosal
protection may be impaired beca use of
diminished lubrication of the oral, oro-
pharyngeal and oesophageal mucosa by sali-
vary mucins. Salivary epidermal growth fac-
tor also plays an important role in the main-
tenance of oro-oesophageal and gastric
tissue integrity. The biological effects of sal-
iva influence the biology of the oesophagus
through their effects on epidermal growth
(including the healing of ulcers), inhibition
of gastric acid secretion (through a feed-
back mechanism), and mucosal protection
against intra-luminal factors such as gastric
acid, bile acids, pepsin and trypsin, as well

as physical, chemical and bacterial factors.
Dysphagia is a common complaint in pa-
tients with hyposalivation and is assumed
to be a consequence of oesophageal dysmo-
tility and altered swallowing pattern due to
reduced salivary flow as well as prolonged
bolus formation. Moreover, salivary gland
hypofunction may indirectly influence gas-
trointestinal functions by causing loss of
appetite, fear of eating, special food prefer-
ences, weight loss, malabsorption and mal-
nutrition.
Saliva and taste
Taste is a major stimulus for the secretion
of saliva. At the same time, tastes are per-
ceived only when saliva is present in the
mouth because taste receptor cells respond
only to dissolved substances (chapter 2).
Furthermore, taste sensitivity is related to
saliva composition as this fluid baths each
receptor cell's oral surface. Complaints of
taste disturbances including abnormal and
impaired sense of taste are common among
patients with Sjogren's syndrome and pa-
tients who have received radiotherapy and
chemotherapy. In these patients, impair-
ment of the threshold for perceiving taste
as well as perception of the intensity of
the four basic taste modalities has been
reported.
• Diagnosis of salivary gland
hypofunction
History-taking and subjective measures of
xerostomia
The procedure for identifying an individual
with potential salivary gland dysfunction re-
quires a careful and systematic evaluation.
The first step is to take a detailed history of
present symptoms, compromised oro-pha-
ryngeal functions related to salivary hypo-
Saliva
function, past and current medication, gen-
eral and oral diseases, trauma to the head,
previous therapies including surgery, radio-
therapy in the head and neck region and/or
chemotherapy (Tables 3 and 4). A ques-
tionnaire including four questions may be
useful in identifying patients with xero-
stomia and salivary gland hypofunction
(Table 5). Dryness of the lips and buccal mu-
cosa, inability to provoke salivary secretion
by palpating the gland as well as total score
of decayed, missing and filled teeth are ad-
ditional predictive measures of salivary
gland hypofunction. The level of oral dry-
ness experienced by the patients may also be
measured by different methods (Table 6).
Extra- and intra-oral examination
The second step in the evaluation of poten-
tial salivary gland dysfunction is a thorough
facial and intraoral examination including
inspection and palpation of the major and
minor salivary glands, the major salivary
duct orifices, the oral mucosa, the dentition,
and the gingivae (see Table 4 for c1inical
signs).
Collection of saliva
The c1inical examination should be followed
by measurement of salivary flow rates.
Stimulated and unstimulated flow rates can
Table S. Selected xerostomia related questions.
1. Does your mouth feel dry when eating a meal?
2. Do you have difficulties swallowing any foods?
3. Do you sip liquids to aid in swallowing dry foods?
4. Does the amount of saliva in your mouth seem toa
little, toa much, or have you not noticed it?
Scientifically validated questions, which may be helpful
in identifying patients with xerostomia and salivary
gland hypofunction. The questions are mainly related
to dry mouth experienced during mealtime and positive
responds are highly predictive of salivary gland hypo-
function.
47
Bardow, Lynge Pedersen and Nauntofte
Table 6. Methods for assessment of subjective
A.
Self-rating categorised questionnaire with
4 degrees of severity*:
0=1 have no feeling of dry mouth
1= 1 have a slight feeling dry mouth
2=1 have asevere feeling of dry mouth
3 =1 have an annoying feeling of dry mouth
Modified according to Beck's inventory scale, item 9.
An ordinal scale based on rank-ordered
estimation. Regarding the latter, the patients
dry mouth on a 100 mm straight line. Verbal labels are assigned
O mrn represents no feeling of oral dryness
methods may also be helpful in assessing
stimulatory treatment.
be measured for whole saliva or individually
for the parotid, submandibular/sublingual
glands using various techniques. This chap-
ter, however, deals with only one method for
measurement of whole saliva secretion (i.e.
the "draining method") for the following
reasons: the method is relatively simple; it
can easily be conducted in the dental office;
it requires only a few tools (Figure 11); it
is internationally accepted, it is the standard
method for measuring unstimulated whole
saliva flow rate (1993 European dassification
criteria for the diagnosis of Sjogren's syn-
drome), and is highly reproducible and re-
liable. Finally, the measurement of whole sal-
iva provides a general assessment of salivary
gland capabilities under basal and stimu-
lated conditions.
Prior to the collection of saliva, the pa-
tient sits relaxed in a chair for a few minutes.
The collection of unstimulated saliva is done
without any masticatory or gustatory stimu-
lus. Movements of the tongue, cheeks, jaws
or lips should be avoided during the collec-
tion periodo For comparisons of salivary
gland function over time, collections in each
patient should be carried out at the same
time of day. The patient must be instructed
48
symptoms of dry mouth.
B.
Visual analogue scale (VAS)
1
100 mm; Worst imaginable
feeling of dry mouth
that makes speech difficult.
O mrn; o feeling of dry mouth
categories (A) and a visual analogue scale (B) representing magnitude
are asked to mark the point that best corresponds to their feeling of
to both ends of the scale to indicate the extremes.
and 100 mm the worst imaginable feeling of oral dryness intensity. The
the intensity of oral dryness over a time span or the effects of saliva
to refrain from eating, drinking, smoking,
and oral hygiene manoeuvres for at least 90
minutes prior to the collection.
Salivary flow rates exhibit large variations
between healthy individuals. Despite the
wide variation in normal whole saliva flow
rates, it is generally accepted that unstimu-
lated rates of 0.3-0.5 ml!min and stimulated
rates of 1.0-1.5 ml!min are within the nor-
mal range. The diagnosis of hyposalivation
is given when the unstimulated, whole sali-
vary flow rate is 0.1 ml/min or less and when
whole salivary flow rates stimulated by
chewing paraffin are 0.5 ml/minute or less
for women, and 0.7 ml!minute or less for
men. Thus, the flow rates in hyposalivation
are much lower than the "normal" values. If
the flow rate measurements suggest salivary
gland dysfunction, a more extensive and spe-
cialized investigation of the patient is indi-
cated. In cases of medication-induced hy-
posalivation and/or xerostomia and polyp-
harrnacy, the patient's physician should be
consulted to discuss the possibility of chang-
ing the medication to another with less ad-
verse effects or, subject to the need to treat
the underlying systemic disease adequately,
changing the dose.
 Saliva

A B e
• Figure 11. Measurement o/ unstimulated and stimulated whole saliva flow rateo
The tools required are a watch, a weight with two digits, a plastic cup, and for saliva stimulation 1 g of paraffin (inert
chewing material) and a metronome. The patient is seated in a relaxed position with his/her head slightly tilted forward.
After an initial swallowing action, the patient is instructed to allow saliva to passively drain from the lower lip into a pre-
weighed plastic cupo The collection starts at time zero. At the end of the collection period, residual saliva is expectorated
from the mouth into the cupo The saliva-containing plastic cup is reweighed, and the flow rate is calculated in g/min,
which is almost equivalent to rnl/rnin. Collection of stimulated whole saliva is performed after collection of unstimulated
saliva following the same the procedure as described above with the exception of shorter collection time and application
of a chewing stimulus. Every 30 seconds the patient should let the saliva drip into the pre-weighed plastic cup for a few
seconds and then saliva collection continues. Measurement of the unstimulated whole saliva flow rate is usually per-
formed for 15 minutes and chewing stimulated saliva for 5 minutes.

Sialochemistry stimulation intensity at which it is collected.

Saliva may also be collected for analysis of It is therefore a prerequisite for inorganic si-
its composition (sialochemistry). The easy alochemistry that the methods of collection
accessibility of salivary glands and their se- are in accordance with the methods de-
cretion has increased the interest in sialoch- scribed earlier in this chapter. Furthermore,
emical analysis. Thus saliva can serve as an determination of saliva pH, bicarbonate
excellent tool for determination of various concentration and buffer capacity, requires
substances like hormones and drugs as well closed systems to avoid loss of CO2 and thus
as for determination of infections of the oral bicarbonate.
cavity. For these purposes a number of chair
side and handy test kits are available on the
market. Nevertheless, when it comes to de-
termination of the inorganic saliva compo- • Management of oral sequelae of
sition it is important to keep the physiology salivary hypofunction
of the salivary glands in mind. Thus, in con-
trast to many other bodily fluids, human sal- First, the patient must be informed about
iva does not have a constant inorganic com- the relationship between salivary hypofunc-
position, because its inorganic composition tion and the increased predisposition for
is strongly influenced by the flow rate and oral disorders. The key concepts of preven-

49
Bardow, Lynge Pedersen and Nauntofte

tive dental management include careful in- ergic agonists that stimulate muscarinic re-
struction of patients in oral hygiene, and ceptors. They should be prescribed only in
regular (at least every 3-month) follow-up at collaboration with the patient's physician in
a dental clinic including dental plaque con- order to avoid unexpected side effects such
trol, dietary instruction and application of as the aggravation of heart disease or inter-
topical fluoride in order to reduce the caries actions with other drugs. Symptoms of oral
activity. Sugar-free chewing gum containing dryness may be alleviated by the use of
fluoride may be useful. In these patients, the mouth gels, oral sprays or artificial saliva.
beneficial effect of fluoride on tooth sub- Some of the latter products contain car-
stance is prolonged due to the low saliva bomethylcellulose, mucins or electrolytes.
flow rate and subsequently, reduced oral However, many patients prefer to take small
clearance. During meals, these patients sips of water rather than use these prepara-
should be advised to sip water when eating tions.
and swallowing. After the meal they should
rinse their mouths thoroughly with water.
At present, due to the lack of controlled • Future perspectives
clinical trials, there is no consensus on which
dental materials are the most appropriate for The most common use of saliva as a diag-
restorative treatment in patients with hypos- nostic tool is to predict a patient's future
alivation. The mouths of denture-wearers caries risk. For this purpose, a simple meas-
should be examined frequently to detect and urement of the unstimulated saliva flow rate
treat possible mucosal ulcerations and den- is still the most reproducible and informa-
ture stomatitis. If present, oral candidiasis tive measure to perform (Figure 6). Al-
should be treated with topical application of though measurements of saliva's buffer ea-
miconazole (2%) ointment or gel, nystatin pacity and saturation with hydroxyapatite
ointment or oral suspension. Systemic treat- were among the first caries-related measure-
ment with fluconazole, ketaconazole or itra- ments performed with saliva, there are prob-
conazole should be reserved for refractory lems with performing both these measure-
cases and immuno-compromised patients. ment for clinical purposes beca use of the
The stimulation of salivary secretion is de- diffusion of CO2 out of the saliva, as dis-
pendent on the presence of residual func- cussed earlier. The possible role of the sali-
tioning salivary gland tissue, and this is de- vary proteins as indicators of caries risk and
termined by measurement of whole saliva progression still needs to be determined. If
flow rate stimulated by chewing. Gustatory it turns out that specific salivary proteins
and/or masticatory stimulation with regular have an impact on caries risk and pro-
use of sugar-free sweets or sugarless chewing gression, modern molecular techniques for
gum may increase secretion of saliva. Acu- protein characterisation could be used as
puncture therapy has been shown to in- tools for individual caries treatment plan-
crease salivary flow rates significantly in ning. Thus, the future challenge for salivary
Sjogren's syndrome patients with remaining researchers is to understand the functions of
functional salivary gland tissue. Pharmaco- the genes and their expression of proteins in
logical sialogogues (e.g., pilocarpine and ce- the salivary glands and saliva.
vimeline) may also stimulate an increase in Several inexpensive and user-friendly di-
salivary secretion. These drugs are cholin- agnostic analysis systems are now appearing

50

for salivary-based diagnosis. There is strong
interest in the development of such systems
for routine monitoring of biomarkers of sys-
temic diseases, since saliva collection is a
non-invasive and painless procedure.
Whole saliva samples contain desqua-
mated epithelial cells from the oral mucosa
from which DNA can be extracted and used
for gene characterization. This may be util-
• Selected references
ised, for example, for gene mapping for
identification of systemic disease poly-
morphisms that will give further insight into
the pathophysiology of the disease. Microar-
ray chip technology now allows for the sim-
ultaneous evaluation of gene expression pat-
terns for thousands of genes, which enables
comparisons to be made of gene expression
profiles between different patient groups.
The combination of the identification of dis-
ease gene polymorphisms and the under-
standing of the expression of these genes will
be important tools for new therapies not
only for salivary gland diseases but also for
systemic diseases.
As the glands serve both exocrine and
some endocrine functions, one strategy be-
hind the use of gene therapy in salivary
glands is to utilise the fact that, in addition
to secreting proteins into saliva, the salivary
gland tissues secrete proteins into the blood
circulation. Although this technology is at a
relative primitive stage today, the future for
using human parotid or submandibular
glands as target organs for gene therapies
using recombinant DNA technology in
which the gene tic material of the host's cells
can be manipulated is promising. Access to
Saliva
the gland via cannulation and retrograde in-
fusion (by viral or non-viral methods) is a
relatively simple procedure due to the easy
accessibility of the main excretory duct of
these glands.
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capacity and buffer systems of human whole saliva
measured without loss of CO2• Arch Oral Biol
2000;45: 1-12.
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Garrett JR, Proctor GB. Control of salivation. In: Lin-
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Lenander-Lumikari M, Loimaranta V. Saliva and dental
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Levine Mi. Development of artificial salivas. Crit Rev
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Nauntofte B. Regulation of electrolyte and fluid secre-
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51