[Saliva--its biological role, composition and secretory mechanism. I. The function and secretion of saliva].
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BACKGROUND: Saliva collection can provide clinical information about individual patients. However, a correlation between ranking buffering capacity using resting and stimulated saliva is still unknown. The aim of this study was to evaluate the pH change after HCl titration into resting and stimulated saliva for a salivary buffering capacity test. METHODS: Resting and stimulated saliva (by chewing paraffin wax) were collected from 80 patients. After the pH of both saliva samples was measured using a hand-held pH meter, the saliva samples were titrated with 0.1N HCl to evaluate the buffering capacity. Correlations of ranking buffering capacity (high, medium, low) between stimulated saliva and resting saliva with 30 microL HCl titration and between stimulated saliva and resting saliva with 40 microL HCl titration were statistically analysed by Spearman Rank Correlation Test (p < 0.05). RESULTS: At 50 microL HCl titration, stimulated saliva buffering capacities were ranked into high (above pH 5.5), medium (pH from 5.5 to 4.5) and low (below pH 4.5). At 30-40 microL HCl titration, the resting saliva buffering capacities were ranked into the same categories. Spearman Rank Correlation indicated significant positive coefficients for the stimulated saliva and resting saliva buffering capacity at 30 microL titration and the stimulated saliva and resting saliva at 40 microL titration. CONCLUSION: Stimulated saliva is more resistant to variation in pH change during HCI titration than resting saliva. Stimulated saliva sampling is a good method to determine buffering capacity during a comprehensive oral health assessment.
Five neuropeptides: Substance P (SP), Neurokinin A (NKA), Calcitonin Gene-Related Peptide (CGRP), Neuropeptide Y (NPY) and Vasoactive Intestinal Polypeptide (VIP), were measured in the saliva of eight subjects. The saliva was collected using different stimulation techniques: whole resting saliva, whole paraffin stimulated saliva, whole citric acid stimulated saliva and parotid saliva of different secretion rates -0.25 mL/min, 0.50 mL/min and 1.00 mL/min, also stimulated by citric acid. The neuropeptides were analysed by radioimmunoassay. The results showed that the concentration of all neuropeptides decreased significantly, two- to four-fold (CGRP up to 16-fold) in whole saliva, when the salivary secretion rates increased six- to eight-fold due to stimulation. However, the amounts of all neuropeptides released over time into the whole saliva increased two- to five-fold (ten-fold for CGRP) as the volumes of saliva increased due to chewing-stimulation as compared to resting saliva or citric acid stimulated saliva. There was also more CGRP in the resting saliva than in the citric acid stimulated saliva. The concentration of CGRP in the parotid saliva decreased three- to ten-fold when the salivary flow increased, whereas the concentration of NKA increased three- to four-fold and that of NPY almost two-fold under the same conditions. The concentrations of SP and VIP did not change in the different flows of parotid saliva. The release of all neuropeptides in the parotid saliva over time showed significant increases (3-14-fold) when the secretion rates increased except CGRP, which showed no changes at all. We concluded that neuropeptides are continuously released into the saliva. Their amounts increase with stimulation, but they are diluted by the increased volume of saliva, and they are also affected by the mode of stimulation-muscular activity leads to a greater release than citric acid stimulation. As the neuropeptides play an important role in the control of salivary secretory mechanisms, their normal occurrence and release are of fundamental importance for the understanding of the function of the salivary glands.
Glucose uptake was examined by using whole-cell suspensions of Streptococcus mutans (strains BHT, Ingbritt, and GS-5), Streptococcus mitis (strains 9811 and 72x41), and Actinomyces viscosus (strains T6 and WVU626) incubated for up to 90 min in 0 to 82% (vol/vol) human whole salivary supernatant. Glucose uptake by the S. mutans strains was completely inhibited at all saliva concentrations. Dithiothreitol (DTT), present during saliva incubation, prevented saliva inhibition. Glucose uptake was also restored when saliva-inhibited cells were subsequently exposed to DTT. The inclusion of catalase in the saliva incubation mixtures resulted in protection equal to that obtained with DTT. The S. mitis strains were also inhibited by saliva but to a far lesser extent that S. mutans. DTT and catalase also protected S. mitis from saliva inhibition. Both A. viscosus strains were completely refractory to saliva inhibition of glucose uptake. Based on (i) the sensitivity of the catalase-negative streptococci and the resistance of catalase-positive actinomyces to saliva inhibition and (ii) the equal and complete protection to saliva inhibition afforded by DTT and catalase, we conclude that the lactoperoxidase-SCN(-)-H(2)O(2) system in saliva was the only antibacterial system expressed under our experimental conditions. The relative resistance of S. mitis 9811 (compared with S. mutans BHT) to saliva inhibition was shown not to result from poor H(2)O(2) production in either glucose-supplemented buffer or saliva solutions. S. mitis produced inhibitory quantities of H(2)O(2) that equaled or exceeded S. mutans H(2)O(2) accumulation. It is suggested that S. mitis might possess a greater ability to repair lactoperoxidase-mediated damage than does S. mutans. Every organism studied exhibited a saliva concentration-dependent, cell growth-independent stimulation of glucose uptake after 60 to 90 min of incubation. The A. viscosus and S. mitis strains showed saliva stimulation (or stabilization) of glucose uptake with unsupplemented saliva. In the case of S. mutans, saliva stimulation was only observed when DTT was present. The possible role of salivary lactoperoxidase as a modulator of the intraoral site specificities exhibited by S. mutans is discussed.
SALIVA COLLECTION: Whole saliva is a product of secretion of 3 major glands (parotid, submandibular, sublingual) and many minor glands (labial, buccal, palatal). Unstimulated saliva is usually obtained as the patient spits out every 60 sec. or by forward bended head the patient allows saliva to drip off the lower lip into a cylinder. By collection of saliva in the tube the flow rate per unit time can be measured. When volume measurement is not required the saliva can be collected on cotton rolls, gauze or filter paper. For evaluating salivary gland function or when large volumes of saliva are required for analytic purposes, stimulated whole saliva is used. Method of collection is the same as for unstimulated saliva. The usual masticatory stimuli are paraffin wax or a washed rubber band. A standard gustatory stimulus is obtained by 2% citric acid applied directly to the tongue every 15 to 60 sec. Parotid saliva can be collected by aspiration from the duct opening with a micropipette. Parotid saliva is best collected with Lashley's vacuum chamber. Submandibular and sublingual saliva can be collected by cannulation of the duct with micropipette, but in practice this is both uncomfortable for the patients and technically difficult since the duct orifice is mobile and has a strong sphincter. Because of that, alginate and silicone impression material is used for retention of the collecting tube. As alternative and simple technique is to block off secretion from the parotid glands with absorbent swabs and collect mixed submandibular and sublingual saliva by pipette from the floor of the mouth. Saliva from labial and palatal glands can be collected by filter paper disc or disc of other synthetic materials. SALIVARY IMMUNOGLOBULIN A: The most significant characteristics of the salivary immunoglobulin system are quantitative domination of immunoglobulin A, local synthesis and specific structure. Immunofluorescence studies have shown that immunoglobulin A is produced by plasma cells locally in the salivary glands. There is still little convincing evidence for the origin of predominantly immunoglobulin A secreting plasma cells in salivary glands. DETECTION OF IMMUNOGLOBULIN A IN SALIVA: Radial immunodiffusion (RID) was the most applicable method for detecting salivary immunoglobulin A. However, there are more sensitive and automatic methods such as nephelometry and ELISA. A standard level of immunoglobulin in saliva is still in question since the concentration varies in relation to origin of saliva, method of collection and stimulation of secretion (Table 1). PERIODONTAL DISEASE: Studies of the salivary immunoglobulin A in patients with periodontal disease and healthy persons showed that there are differences which can be used in detection of high-risk groups and individuals. If the bacterial adherence to the mucosa is a prerequisite for bacterial evolution in subgingival or any other region of the oral cavity respectively introduction in periodontitis development, than it is to be presumed that the basic function of salivary immunoglobulin A is inhibition of bacterial adherence rather than antigens destruction. Several bacterial species frequently isolated from the oral cavity of patients with periodontitis have been identified as producers of IgA protease. These enzymes cleave serum IgA and secretory IgA equally well. Additionally, most of the IgA proteases studied have cleaved the A1 and A2 subclass. Several studies have demonstrated that cleavage of human IgA occurs in vivo, resulting in generation of intact Fab alpha and (Fc alpha)2 fragment. Moreover, when bacteria are exposed to Fab alpha fragments released from IgA after cleavage by IgA protease, their surface antigens are likely to be occupied by Fab alpha fragments. These Fab alpha fragments left on the bacterial surface may mediate adhesion. Together, these results indicate that IgA proteases, by promoting adherence, contribute the pathogenic potential of bacteria in the oral c
Hyaluronan (hyaluronic acid) is a glycosaminoglycan that functions as a constituent of ground substance, a mediator of cell proliferation and would healing, and that plays a prominent part in tumorigenesis as well as in embryogenesis. Its presence and possible role in saliva has been subjected to little study. Unstimulated and stimulated pure parotid and mixed saliva was obtained from 10 volunteers. The protein content of the samples was assayed and the hyaluronan concentration was evaluated by means of an enzyme immunosorbent-like assay using a hyaluronan-binding peptide. Stimulated whole saliva had the highest protein content (mean 1.26 mg/ml) followed by unstimulated parotid saliva (1.15 mg/ml), stimulated parotid saliva (0.95 mg/ml) and unstimulated whole saliva (0.93 mg/ml). Absolute hyaluronan concentrations were highest in unstimulated whole saliva (mean 459.2 ng (nanograms)/ml), and lowest in stimulated parotid saliva (82.7 ng/ml). When hyaluronan concentrations are expressed as ng/mg of protein, the highest are in the unstimulated whole saliva (mean 477.5 ng/mg protein) followed by stimulated parotid saliva (229.7 ng/mg), unstimulated parotid saliva (179.6 ng/mg) and stimulated whole saliva (159.9 ng/mg). There are wide variations in the levels of hyaluronan in human saliva depending on the type of saliva and the conditions at the time of collection. Regulation of hyaluronan metabolism represents an intricate balance between production and degradation, and it is unclear whether elevated concentrations of hyaluronan in response to tissue proliferation, regeneration or repair. The hyaluronan may contribute to the healing properties of saliva, assisting in protecting the oral mucosa and adding to the lubricating properties of saliva.
The aims of this study were to determine the relationship between saliva and plasma methadone concentrations and the influence of variability in saliva pH. Saliva and plasma samples were taken before the daily dose of methadone in 60 patients undergoing methadone maintenance treatment (MMT). Saliva pH was measured immediately after sampling, and concentrations of (RS)-, (R)-, and (S)-methadone in saliva and plasma were assayed by LC/MS. In addition, unbound (R)- and (S)-methadone concentrations were measured in plasma samples by ultrafiltration. Plasma binding and pH differences between plasma and saliva were then used to estimate methadone saliva/plasma ratios and to compare them with observed values. Saliva pH ranged from 5.1 to 7.6 (mean +/- SD, 6.7 +/- 0.5). Plasma and saliva concentrations correlated weakly [(RS)-, r = 0.14, P = 0.007, n = 44; (R)-, r = 0.10, P = 0.04, n = 43; (S)-, r = 0.22, P = 0.002, n = 43], and the mean saliva-to-plasma methadone concentration ratios were 1.1 (+/-1.3 SD), 1.5 (+/-1.5), and 0.8 (+/-0.8), for (RS)-, (R)-, and (S)-methadone, respectively. Corresponding values based on unbound concentrations of methadone in plasma were 21 (+/-20.6, n = 31), 21 (+/-19, n = 34), and 17 (+/-15, n = 36). The salivary concentration-to-dose ratios showed statistically significant but weak inverse correlations with saliva pH [(RS)-, r = 0.27, P < 0.001; (R)-, r = 0.25, P < 0.001; (S)-, r = 0.29, P < 0.001, respectively]. There were significant correlations between predicted and observed saliva/plasma ratios [(RS)-, r = 0.44, P < 0.001, n = 31; (R)-, r = 0.58, P < 0.001, n = 32; (S)-, r = 0.10, P = 0.04, n = 34], but the mean predicted saliva concentrations were about 5 times lower than the mean observed values. The poor correlations between salivary and plasma methadone concentrations observed in this study are partly related to the effect of variable saliva pH. However, saliva pH explained only 10%-36% of the total variation. As a conclusion, monitoring methadone concentrations in saliva may not be a useful alternative to plasma concentration measurements. Correction for saliva pH measured immediately after collection improves the relationship between saliva and plasma methadone concentration, but most of the variation remains unexplained.
Ionized (saliva-Ca2+) and total calcium (saliva-CaT), inorganic phosphate (saliva-P), magnesium (saliva-Mg), and zinc (saliva-Zn) levels in parotid saliva were studied in eight healthy volunteers during a 3-h calcium infusion, directly regulated to obtain a pre-settled standardized blood calcium level. Frequent determinations of whole blood ionized calcium (B-Ca2+) were used for continuous adjustments of the calcium infusion rate ('calcium clamp'). Basal B-Ca2+ (1.26 +/- 0.04 mmol X l-1, mean +/- SD) was thus elevated to and maintained at 1.51 +/- 0.05 mmol X l-1 from +30 to +180 min. Saliva samples were taken prior to the infusion and at regular intervals during the infusion period. Saliva-Ca2+ decreased from 0.38 +/- 0.06 to 0.33 +/- 0.05 mmol X l-1 (P less than 0.01) and saliva-Ca2+ as a percentage of saliva CaT from 63 +/- 7% to 53 +/- 10% (P less than 0.001) during the infusion. No changes could be observed in saliva-CaT, saliva-P, saliva-Mg, saliva-Zn, saliva-Na and saliva-K. Possible mechanisms behind these findings might be decreased parathyroid hormone and increased calcitonin concentrations in serum.
A study of saliva and its tooth-protective components reveals at least four important functions of saliva: (1) buffering ability, (2) a cleansing effect, (3) antibacterial action, and (4) maintenance of a saliva supersaturated in calcium phosphate. Several salivary constituents subserve one or more of these functions. Research has yielded important information about organic and inorganic secretory products. It is also clear that saliva as a unique biologic fluid has to be considered in its entirety to account fully for its effects on teeth. Saliva is greater then the sum of its parts. One reason for this is that salivary components display redundancy of function, each often having more than one function. This redundancy, however, does not imply that proteins that share functional roles all contribute to the same degree. For instance, when comparing proteins that inhibit calcium phosphate precipitation, statherin and acidic proline-rich proteins are most potent, whereas histatins, cystatins, and mucins appear to play lesser roles. The complex interaction between proteins is another major factor contributing to saliva's function. In this regard, heterotypic complexes of various proteins have been shown to form on hydroxyapatite. Mucin binding to other salivary proteins, including proline-rich proteins, histatins, cystatins, and statherin, is well documented. The complexes, whether adsorbed to the tooth surface or in saliva, have important implications for bacterial clearance, selective bacterial aggregation on the tooth surface, and control of mineralization and demineralization. Finally, proteolytic activity of saliva generates numerous products whose biologic activities are often different from their parent compounds. Fluoride is another important component of saliva that is discussed separately in other articles in this issue. The ability of saliva to deliver fluoride to the tooth surface constantly makes salivary fluoride an important player in caries protection largely by promoting remineralization and reducing demineralization. Some key properties of salivary components discussed in this article are listed in Table 1. Saliva is well adapted to protection against dental caries. Saliva's buffering capability; the ability of the saliva to wash the tooth surface, to clear bacteria, and to control demineralization and mineralization; saliva's antibacterial activities; and perhaps other mechanisms all contribute to its essential role in the health of teeth. The fact that the protective function of saliva can be overwhelmed by bacterial action indicates the importance of prevention and therapy as in other infectious diseases. The knowledge of functional properties of saliva as well as those of its separate components may permit a better assessment of dental caries susceptibility. Future research is essential to characterize more fully salivary components and their interactions and how these affect the caries process. With such knowledge, the use of modified oral molecules as therapeutic agents may become a reality. Equally intriguing is the prospect of influencing the secretion of salivary components by greater knowledge and control over the secretory processes responsible for the delivery of those components.
Saliva is an alternate biological matrix for drug testing that has several advantages over more traditional fluids such as blood and urine. Collection is rapid, noninvasive, and relatively easy to obtain. Several reports have detailed the appearance of drugs of abuse in saliva, but few have compared the excretion profiles of drugs administered by different routes. In this study, subjects were administered three smoked and three intravenous doses of heroin in an ascending dose design. Blood and saliva were collected periodically after drug administration and analyzed by gas chromatography-mass spectrometry (GC-MS) for heroin, 6-acetylmorphine, and morphine. In a second study, subjects were administered a single, smoked dose of 40 mg cocaine base and an intravenous dose of 44.8 mg cocaine HO on separate occasions. Plasma and saliva were collected and analyzed by CC-MS for cocaine, anhydroecgonine methyl ester (AEME), and seven additional metabolites. Heroin and 6-acetylmorphine were detected in the first saliva sample collected (2 min) following drug administration by both routes. Peak heroin concentrations were achieved quickly, between 2 and 5 min after intravenous administration and at 2 min after smoke heroin. Peak heroin concentrations in saliva after smoking heroin base ranged from 3534 (2.6 mg) to 20,580 ng/mL (5.2 mg), and after intravenous administration, concentrations ranged from 6 (10 mg heroin HCl to 30 ng/mL (12 mg heroin HCl. Saliva concentrations of heroin declined rapidly after intravenous administration, reaching the limit of sensitivity of the assay (1 ng/mL) by 60 min. Heroin concentrations in saliva after smoking declined slowly; detection times ranged from 4 to 24 h. Cocaine was the major analyte detected in saliva and plasma after smoked and intravenous administration. Peak saliva cocaine concentrations after intravenous administration ranged from 428 to 1927 ng/mL (N = 7); after smoking, they ranged from 15,852 to 504,880 ng/mL (N = 7). Peak plasma cocaine concentrations after intravenous administration ranged from 122 to 442 ng/mL A = 7), and after smoking, concentrations ranged from 46 to 291 ng/mL A = 7). The thermal degradation product of cocaine, AEME, was detected in saliva but not in plasma after smoking. Peak saliva AEME concentrations were achieved at 2 min and ranged from 558 to 4374 ng/mL (N = 7). These are the first reported observations of heroin and metabolites in saliva following heroin smoking and of AEME in saliva after smoking cocaine base. The presence of AEME in saliva may be useful as a marker of the smoked route following cocaine administration.