Rechargeable anticandidal denture material with sustained release in saliva.

OBJECTIVE: Candida-induced denture stomatitis is a common debilitating problem among denture wearers. Previously, we described the fabrication of a new denture material that released antifungal drugs when immersed in phosphate buffered saline. Here, we use more clinically relevant immersion conditions (human saliva; 37°C) and measure miconazole release and bioactivity. MATERIALS AND METHODS: Disks were prepared by grafting PNVP [poly(N-vinyl-2-pyrrolidinone)] onto PMMA [poly(methylmethacrylate)] using plasma initiation (PMMA-g-PNVP) and then loaded with miconazole. Drug-loaded disks were immersed in 10-100% human saliva (1-30 days). Miconazole release was measured and then tested for bioactivity vs miconazole-sensitive and miconazole-resistant Candida isolates. RESULTS: HPLC was used to quantify miconazole levels in saliva. Miconazole-loaded disks released antifungal drug for up to 30 days. Higher drug release was found with higher concentrations of saliva, and, interestingly, miconazole solubility was increased with higher saliva concentrations. The released miconazole retained its anticandidal activity. After immersion, the residual miconazole could be quenched and the disks recharged. Freshly recharged disks displayed the same release kinetics and bioactivity as the original disks. Quenched disks could also be charged with chlorhexidine that displayed anticandidal activity. CONCLUSIONS: These results suggest that PMMA-g-PNVP is a promising new denture material for long-term management of denture stomatitis.

Anticandidal activity and biocompatibility of a rechargeable antifungal denture material.

OBJECTIVES: Candida-associated denture stomatitis is a recurrent and debilitating oral mucosal disease. Development of anticandidal denture materials represents a promising strategy to manage this condition. We have previously shown that miconazole incorporated in methacrylic acid (MAA) copolymerized diurethane dimethacrylate (UDMA) denture materials has long-term anticandidal activity. In this study, we examined the ability of culture medium conditioned with drug-free- or miconazole-MAA-UDMA discs to prevent Candida infection in an in vitro oral epithelial cell/Candida albicans coculture system. MATERIALS AND METHODS: Candida albicans (C. albicans)-induced OKF6/TERT-2 cell damage was quantified by the release of lactate dehydrogenase from epithelial cells, cytokine production was quantified using protein cytokine arrays, and the expression of C. albicans genes was measured by RT-qPCR. RESULTS: Candida albicans had limited growth with altered expression levels of secreted aspartyl proteinase-2 and -5 in culture medium conditioned by miconazole-MAA-UDMA discs. Significantly, the ability of C. albicans to induce oral epithelial cell damage and trigger epithelial proinflammatory cytokine production was also inhibited by miconazole disc conditioned media. CONCLUSION: Miconazole released from MAA-UDMA denture materials effectively prevents the development of candidal infection in an in vitro oral epithelial system. Further characterization of this drug-rechargeable denture material is warranted.

Cellular signals underlying ß-adrenergic receptor mediated salivary gland enlargement.

We examined the cellular signaling pathways involved in parotid gland enlargement induced by repeated isoproterenol administration in rats. Immunoblot analysis revealed early (1h) activation of the mitogen activated protein kinase (MAPK) ERK1/2, and progressive activation of epidermal growth factor receptor (EGFR), p38MAPK and p70S6 kinase (p70S6K) during 72h of isoproterenol treatment. Expression of ß-adrenergic receptors (ARs) of the ß2, but not ß1, subtype increased over time in parallel with increases in the proliferation marker PCNA and parotid gland weight. Levels of ß2-AR mRNA, assessed by quantitative RT-PCR and Northern blot analysis, were upregulated in parotid glands of isoproterenol treated rats. cAMP response element binding protein (CREB), a positive regulator of ß2-AR transcription, was activated at 1h after isoproterenol administration, as evidenced by increased nuclear translocation and DNA binding using immunohistochemical staining and electrophoretic mobility shift assay. ELISA of NF-κB, also a ß2-AR transcriptional regulator, revealed an increase in p65 and p50 subunits in nuclear protein extracts from parotid glands of isoproterenol treated rats. Together, these results demonstrate that ß-adrenergic stimulation activates diverse cell survival and progrowth signaling pathways, including cAMP and EGFR linked activation of ERK1/2, p38MAPK, and p70S6K, and also induction of ß2-ARs, possibly mediated by CREB and NF-κB, resulting in salivary gland enlargement. We propose that during isoproterenol treatment activation of the ß1-AR, the predominant ß-AR subtype in unstimulated salivary glands, initiates proliferative signaling cascades, and that upregulation of the ß2-AR plays an essential role in later stages of salivary gland growth.

Rechargeable infection-responsive antifungal denture materials.

Candida-associated denture stomatitis (CADS) is a significant clinical concern. We developed rechargeable infection-responsive antifungal denture materials for potentially managing the disease. Polymethacrylic acid (PMAA) was covalently bound onto diurethane dimethacrylate denture resins in the curing step. The PMAA resins bound cationic antifungal drugs such as miconazole and chlorhexidine digluconate (CG) through ionic interactions. The anticandidal activities of the drug-containing PMAA-resin discs were sustained for a prolonged period of time (weeks and months). Drug release was much faster at acidic conditions (pH 5) than at pH 7. Drugs bound to the denture materials could be "washed out" by treatment with EDTA, and the drug-depleted resins could be recharged with the same or a different class of anticandidal drugs. These results suggest clinical potential of the newly developed antifungal denture materials in the management of CADS and other infectious conditions.

Salivary secretion, mucin concentrations and Candida carriage in HIV-infected patients.

OBJECTIVES: To test whether the submandibular/sublingual (SMSL) salivary secretion, mucin concentration and candida carriage status were altered in human immunodeficiency virus-positive (HIV+) patients. SUBJECTS AND METHODS: SMSL saliva collected from 48 HIV-infected and 31 HIV-negative men were analyzed for flow rates, total protein and mucin concentrations. Salivary cultures were performed for Candida assessment. RESULTS: The salivary flow rate and protein secretion of the HIV+ patients was 37% and 32% less than that of the controls (P < 0.0001, P = 0.0087). The mucin concentrations (MG1 and MG2) were higher in the HIV+ subjects compared with controls (P = 0.0186, P = 0.0014); however, the mucin secretions were not different. The frequency of Candida-positive cultures was higher in the HIV+ subjects than in the controls (61.4%vs 24.1%, P = 0.0018). In the HIV-infected group, the unstimulated SMSL flow rates were lower in Candida-positive than in Candida-negative patients (P = 0.0158). CONCLUSION: The salivary secretion of the SMSL glands was reduced in HIV infection. Although the mucin concentration increased in HIV+ subjects, mucin secretion was not altered. Highly active antiviral therapy had no effect on salivary function. We found an association between the level of candida carriage and salivary flow rate in HIV-infected patients.

Alteration in salivary function in early HIV infection.

The etiology of salivary gland hypofunction in HIV(+) patients is unclear. This study was designed to determine the effect of early-stage HIV(+) infection (CD4(+) > 200 cells/ micro L; n = 139) on salivary gland function and the relationship of this dysfunction to the taking of xerostomic medications. Salivary flow rates and the content of electrolytes and antimicrobial proteins in stimulated parotid and submandibular/sublingual saliva were determined. Compared with healthy controls (n = 50), the HIV(+) group showed significant reductions in flow rates of unstimulated whole (35%), stimulated parotid (47%), unstimulated submandibular/sublingual (23%), and stimulated submandibular/sublingual (39%) saliva. The flow rates for the HIV(+) patients taking xerostomic medications did not differ from those of patients who did not. Concentrations of some salivary gland components were altered in the HIV(+) group. Analysis of these data suggests that salivary gland function is adversely affected early in HIV infection and that these changes do not appear to be compounded by the taking of xerostomic medications.

Activated caspase 3 and cleaved poly(ADP-ribose)polymerase in salivary epithelium suggest a pathogenetic mechanism for Sjögren's syndrome.

OBJECTIVE: Apoptosis is an organized energy-dependent process of cellular self-destruction carried out by proteolytic enzymes such as the caspases. These enzymes may play a role in epithelial cell apoptosis in Sjögren's syndrome (SS). A classical caspase substrate is poly(ADP-ribose)polymerase (PARP), a DNA repair enzyme. To elucidate the molecular mechanisms responsible for salivary gland dysfunction in SS, we studied the expression of caspase and PARP in SS salivary gland biopsies. METHODS: The presence of activated caspases (caspases 3 and 9) and cleaved PARP (85 kDa) in SS biopsies was demonstrated by immunohistochemistry using specific polyclonal antibodies. RESULTS: Initial studies performed with an antibody reagent that recognizes both active and inactive forms of caspase 3 identified this enzyme in SS salivary ductal and acinar cells. Activated caspase 3 and cleaved PARP were strongly expressed in ductal and acinar cells in SS salivary glands (13/15). Ductal and acinar cells from normal salivary glands (n=5) stained with less intensity compared with SS tissue. Staining for activated caspase 9 was negative in all samples. Likewise, infiltrating lymphocytes were negative for caspase 3, caspase 9 and cleaved PARP. CONCLUSION: This study shows that caspase 3 is important in the salivary dysfunction of SS, while caspase 9 appears not to be involved.

Salivary anticandidal activity and saliva composition in an HIV-infected cohort.

This study investigated salivary anticandidal activity and salivary composition in stimulated whole saliva of 18 advanced HIV-infected patients and compared these values to healthy controls. Stimulated whole saliva from HIV-infected patients showed decreased anticandidal activity. The flow rate was reduced by 40% as compared with controls. The saliva flow rate for HIV-infected patients who had recoverable yeast in their saliva was reduced as compared to HIV-infected patients without recoverable yeast. For HIV-infected patients, the saliva concentrations of lactoferrin, secretory IgA and Cl- were increased while the secretion rate of lysozyme, total protein and K+ were reduced. There was no difference in any parameter as a function of taking the antifungal drug fluconazole. There was no association between salivary anticandidal activity and any salivary component. This study shows reduced anticandidal activity and salivary flow rate in HIV-infected patients. These alterations may contribute to their increased incidence of oral candidal infections.

Measuring short-term gamma-irradiation effects on mouse salivary gland function using a new saliva collection device.

A restraining device was designed specifically for the collection of whole saliva from mice without using anesthesia. As the procedure does not involve surgical cannulation of the salivary glands, saliva can be collected from the same mouse at different times. The time between the injection of a secretory stimulant (pilocarpine) and the appearance of saliva in the mouth (lag time) was 100.5 +/-8.5 s (mean+/-S.E.M., n=10) for control mice. The volume of saliva collected in the first 5 min was three times greater than that collected between 15 and 20 min. The average flow rate for a collection period of 15 min was 16.7 +/-1.8 microl/min (n=10). The flow rate was decreased 50% (P<0.005) whereas the lag time was increased more than 300% (P<0.05) at 24 h after irradiation. The concentrations of a 23.5-kDa protein and a mucin were decreased after irradiation whereas there was no significant effect on the concentration of amylase or peroxidase.

On velocity estimation using speckle decorrelation.

Quantitative estimation of blood velocity using Doppler techniques is fundamentally limited because only the axial component can be detected. Speckle decorrelation resulting from scatterer motion may be used to compute non-axial components and to obtain quantitative flow information. Based on both simulations and experimental results, it is shown that the decorrelation technique is feasible only for constant flows. If flow gradients are present, the correlation between two signals along the same line of observation may be significantly affected by the gradients. Therefore, the decorrelation method cannot be used for quantitative flow estimation if flow gradients are not accurately measured and effects on signal correlation are not fully compensated. Results in this paper show that accurate estimation of flow gradients is practically difficult. It is further shown that effects of signal-to-noise ratio (SNR) on the correlation must also be taken into account for quantitative flow analysis.

Feasibility study of time-intensity-based blood flow measurements using deconvolution.

Ultrasonic contrast agents have been used to enhance the acoustic backscattered intensity of blood and to assist the assessment of blood flow parameters. One example is the time-intensity method based on the indicator-dilution theory. In this case, a mixing chamber model can be employed to describe the concentration of the contrast agent as a function of time. By measuring the time intensities at both the input and output of the blood mixing chamber, blood flow information can be obtained if proper deconvolution techniques are applied. Note that most deconvolution techniques assume a linear and time invariant (LTI) system for the mixing of the contrast agent with blood. In this paper, the hypothesis that a blood mixing chamber is an LTI system was tested. Several aspects were studied. One aspect was the linear relationship between the concentration of the contrast agent and the backscattered intensity. The other aspect was the dependence of the derived time constants on the concentration. The concept of an effective mixing volume was also introduced and evaluated. Finally, the input and the output time constants were measured and compared to theory under the LTI assumption. Extensive experiments were performed. Two in vitro flow models were constructed and two contrast agents were used. Results indicated that the LTI assumption does not hold and quantitative flow estimation is generally not possible. Nonetheless, the indicator-dilution theory can still be applied if only relative measurements of the flow rate are required.

Salivary alterations in type 2 (non-insulin-dependent) diabetes mellitus and hypertension.

OBJECTIVES: The aim of this study was to determine whether saliva output and composition are altered in type 2 diabetes mellitus by comparison with a healthy, non-medicated control group, and also a group of hypertensives. METHODS: From a community-dwelling cohort of Mexican American and European American subjects enrolled in the OH:SALSA oral aging study, we identified 233 subjects with type 2 diabetes mellitus, 227 with hypertension, and 240 healthy control subjects. We collected unstimulated whole (UW) and submandibular/ sublingual (US) saliva, as well as stimulated parotid (SP) and submandibular/ sublingual (SS) saliva. Flow rates were determined, yeast carriage was assayed in UW saliva, and SP and SS saliva samples were analyzed for protein composition. ELISA was used to determine concentrations of an array of specific protein components, with both antimicrobial and other activities. RESULTS: Both diabetic and hypertensive subjects had reduced output of both stimulated and unstimulated submandibular/sublingual saliva. 30% of the diabetic subjects had high oral yeast counts (> or =1000 cfu/mL) compared with 17% of the healthy subjects and 20% of the hypertensives. Significant increases in the concentrations of a number of the protein components were found in the diabetic subjects, specifically, SP lactoferrin, myeloperoxidase (MPO), and salivary peroxidase (SPO), as well as SS total protein, albumin, lactoferrin and secretory IgA. CONCLUSIONS: The pattern of decreased flow rates and increased protein concentrations were similar, but consistently greater in diabetics than hypertensives, suggesting that disease-specific mechanisms may be responsible. Diabetics may be more prone to oral dryness and infections than non-diabetics.

Association of salivary flow rates with maximal bite force.

Mean salivary secretion and bite force decrease with advancing age. Previous studies have shown that salivary flow rates are influenced by mastication. In the present study, we examined the relationship between salivary flow rates and maximal bite force in a community-based sample of men and women 35 years of age or older. Salivary flow rates for unstimulated whole and unstimulated submandibular/sublingual (SMSL) saliva as well as citrate-stimulated parotid and SMSL saliva were measured in 399 subjects. Bite force was assessed with a bilateral force transducer. Pearson correlation analysis yielded significant positive correlations between bite force and flow rates for unstimulated whole saliva (r = 0.24, p < 0.0001), stimulated parotid saliva (r = 0.13, p < 0.03), unstimulated SMSL (r = 0.14, p < 0.0001), and stimulated SMSL (r = 0.16, p < 0.003). When adjusted for age and gender, the partial correlations between bite force and salivary flow rates remained significant for unstimulated whole saliva (r = 0.10, p < 0.05), stimulated parotid saliva (r = 0.13, p < 0.02), and stimulated SMSL saliva (r = 0.14, p < 0.006). Subjects were divided into four groups based on their maximal bite force score (low, medium low, medium high, and high). For each saliva type, the flow rate of the high-bite-force group was significantly greater than that of the low-bite-force group as well as that of the medium-high-bite-force group. These results confirm an age-related decrease in bite force and salivary flow rates and show that, regardless of age or gender, bite force is correlated with salivary flow.

EGF inhibits muscarinic receptor-mediated calcium signaling in a human salivary cell line.

The effects of epidermal growth factor (EGF) on intracellular calcium ([Ca(2+)](i)) responses to the muscarinic agonist carbachol were studied in a human salivary cell line (HSY). Carbachol (10(-4) M)-stimulated [Ca(2+)](i) mobilization was inhibited by 40% after 48-h treatment with 5 x 10(-10) M EGF. EGF also reduced carbachol-induced [Ca(2+)](i) in Ca(2+)-free medium and Ca(2+) influx following repletion of extracellular Ca(2+). Under Ca(2+)-free conditions, thapsigargin, an inhibitor of Ca(2+) uptake to internal stores, induced similar [Ca(2+)](i) signals in control and EGF-treated cells, indicating that internal Ca(2+) stores were unaffected by EGF; however, in cells exposed to thapsigargin, Ca(2+) influx following Ca(2+) repletion was reduced by EGF. Muscarinic receptor density, assessed by binding of the muscarinic receptor antagonist L-[benzilic-4,4'-(3)HCN]quinuclidinyl benzilate ([(3)H]QNB), was decreased by 20% after EGF treatment. Inhibition of the carbachol response by EGF was not altered by phorbol ester-induced downregulation of protein kinase C (PKC) but was enhanced upon PKC activation by a diacylglycerol analog. Phosphorylation of mitogen-activated protein kinase (MAP kinase) and inhibition of the carbachol response by EGF were both blocked by the MAP kinase pathway inhibitor PD-98059. The results suggest that EGF decreases carbachol-induced Ca(2+) release from internal stores and also exerts a direct inhibitory action on Ca(2+) influx. A decline in muscarinic receptor density may contribute to EGF inhibition of carbachol responsiveness. The inhibitory effect of EGF is mediated by the MAP kinase pathway and is potentiated by a distinct modulatory cascade involving activation of PKC. EGF may play a physiological role in regulating muscarinic receptor-stimulated salivary secretion.

Effect of donor age on the concentrations of histatins in human parotid and submandibular/sublingual saliva.

Histatins are small proteins of human glandular saliva that have antifungal properties. Recent studies show that oral candidal infections increase with age, suggesting an age-associated compromise in oral host defence. Here, the effect of age and of physiological gland stimulation on the concentration and secretion of salivary histatins was investigated. Parotid and submandibular/sublingual salivas were collected from six young adults under unstimulated, mechanical (chewing) and gustatory (0.025 M and 0.1 M citric acid) stimulation, and the concentration and secretion of histatins was measured by cationic polyacrylamide gel electrophoresis with subsequent densitometric scanning of the stained gels. With gland stimulation, parotid saliva showed no significant increase in histatin concentration (microg/ml); however, histatin secretion (microg/min) increased up to 26-fold (p<0.005; ANOVA). Stimulation of submandibular/sublingual saliva resulted in significant increases in both histatin concentration (p<0.005) and secretion (p<0.0005). Ageing effects on salivary histatins were determined in citric acid (0.1 M)-stimulated parotid and submandibular/sublingual saliva samples collected from 80 individuals (divided into four age groups having approximately equal numbers of males and females: 35-44 years; 45-54 years; 55-64 years and 65-76 years). None of the patients was taking medications or wore dentures. ANOVA showed no sex differences in histatins. Regression analysis showed significant age-associated decreases for parotid saliva histatin concentration (p<0.002) and secretion (p<0. 002) as well as for submandibular/sublingual saliva histatin concentration (p<0.0001) and secretion (p<0.0001). Both saliva types showed significant (p<0.0001) decreases in the histatin concentration per mg of total protein, suggesting a preferential decrease in salivary histatins compared to total salivary protein. These results suggest that the salivary histatin component of the oral host defence system is compromised with increasing age.

G-protein signaling abnormalities mediated by CD95 in salivary epithelial cells.

Salivary epithelial cells from patients with primary Sjögren's syndrome (SS) undergo Fas-mediated apoptosis. Bcl-2 and Bcl-xL are apoptosis suppressing oncogenes. Very little is known about the role of these oncogene molecules in salivary epithelial cells. To investigate the possible prevention of salivary glandular destruction in SS by Bcl-2 and Bcl-xL, stable transfectants expressing these molecules were made from HSY cells, a human salivary epithelial cell line. HSY cells were transfected with an expression vector for human Bcl-2 or Bcl-xL. Stable transfectants were selected and apoptosis was induced by anti-Fas antibody. Apoptosis was quantified by propidium iodide staining followed by flow cytometry. Caspase activity was detected by immunohistochemical analysis and enzyme cleavage of DEVD-AMC, a fluorescent substrate. Response to carbachol, a muscarinic receptor agonist, and EGF was measured by Ca2+ mobilization and influx. Fas-mediated apoptosis was significantly inhibited in Bcl-2 and Bcl-xL transfectants compared to wild-type and control transfectants (empty vector). Surprisingly, caspase activity was not inhibited in Bcl-2 and Bcl-xL transfectants. Activation of the Fas pathway in the Bcl-2 and Bcl-xL transfectants by antibody also inhibited carbachol and EGF responsiveness (i.e., Ca2+ mobilization and/or influx) by 50-60%. This Fas-mediated inhibition of cell activation was partially or completely restored by specific peptide interference of caspase enzyme activity. The prevention of Fas-mediated apoptosis by the overexpression of Bcl-2 and Bcl-xL in salivary gland epithelial cells results in injured cells expressing caspase activity and unable to respond normally to receptor agonists. Such damaged cells may exist in SS patients and could explain the severe dryness out of proportion to the actual number of apoptotic cells seen on salivary gland biopsy.

Further characterization of human salivary anticandidal activities in a human immunodeficiency virus-positive cohort by use of microassays.

Salivary anticandidal activities play an important role in oral candidal infection. R. P. Santarpia et al. (Oral Microbiol. Immunol. 7:38-43, 1992) developed in vitro anticandidal assays to measure the ability of saliva to inhibit the viability of Candida albicans blastoconidia and the formation of germ tubes by C. albicans. In this report, we describe modifications of these assays for use with small volumes of saliva (50 to 100 microl). For healthy subjects, there is strong inhibition of blastoconidial viability in stimulated parotid (75%), submandibular-sublingual (74%), and whole (97%) saliva, as well as strong inhibition of germ tube formation (>80%) for all three saliva types. The susceptibility of several Candida isolates to inhibition of viability by saliva collected from healthy subjects is independent of body source of Candida isolation (blood, oral cavity, or vagina) or the susceptibility of the isolate to the antifungal drug fluconazole. Salivary anticandidal activities in human immunodeficiency virus (HIV)-infected patients were significantly lower than those in healthy controls for inhibition of blastoconidial viability (P < 0.05) and germ tube formation (P < 0. 001). Stimulated whole-saliva flow rates were also significantly lower (P < 0.05) for HIV-infected patients. These results show that saliva of healthy individuals has anticandidal activity and that this activity is reduced in the saliva of HIV-infected patients. These findings may help explain the greater incidence of oral candidal infections for individuals with AIDS.

Shape-based grey-level image interpolation.

The three-dimensional (3D) object data obtained from a CT scanner usually have unequal sampling frequencies in the x-, y- and z-directions. Generally, the 3D data are first interpolated between slices to obtain isotropic resolution, reconstructed, then operated on using object extraction and display algorithms. The traditional grey-level interpolation introduces a layer of intermediate substance and is not suitable for objects that are very different from the opposite background. The shape-based interpolation method transfers a pixel location to a parameter related to the object shape and the interpolation is performed on that parameter. This process is able to achieve a better interpolation but its application is limited to binary images only. In this paper, we present an improved shape-based interpolation method for grey-level images. The new method uses a polygon to approximate the object shape and performs the interpolation using polygon vertices as references. The binary images representing the shape of the object were first generated via image segmentation on the source images. The target object binary image was then created using regular shape-based interpolation. The polygon enclosing the object for each slice can be generated from the shape of that slice. We determined the relative location in the source slices of each pixel inside the target polygon using the vertices of a polygon as the reference. The target slice grey-level was interpolated from the corresponding source image pixels. The image quality of this interpolation method is better and the mean squared difference is smaller than with traditional grey-level interpolation.

Impact of aging on human salivary gland function: a community-based study.

A comprehensive evaluation of salivary flow rates and composition was undertaken in an age- and community-stratified population. A nonmedicated subpopulation was used to assess the effect of "primary aging" on salivary gland function. Unstimulated whole, parotid and submandibular/sublingual (SMSL) saliva, as well as citrate-stimulated parotid and SMSL saliva were collected from 1006 subjects. Flow rates were determined, and the total protein concentrations measured. Height and caloric intake were documented. Subjects were divided into six age groups from 35 to 75+ years old. Significant age-related decreases in the secretion rates of unstimulated whole (p < 0.001), stimulated parotid (p < 0.01) and unstimulated and stimulated SMSL (both p < 0.0001) saliva were observed in the total population. In the non-medicated subpopulation, age-related decreases in salivary secretions were observed in unstimulated whole (p < 0.01) and unstimulated and stimulated SMSL (p < 0.01 and p < 0.0001, respectively). Multiple regression analysis revealed that, as well as age, caloric intake was related to unstimulated SMSL and stimulated SMSL saliva in the whole population, and height was a contributor to unstimulated whole saliva and unstimulated parotid saliva flow rate variances. In the non-medicated population, caloric intake was the significant independent variable for unstimulated and stimulated parotid secretion, as was height for unstimulated whole and SMSL flow rates. Age-related increases in the total protein concentration of unstimulated parotid (p < 0.001) and unstimulated SMSL (p < 0.05) saliva were evident in the whole population, but not in the non-medicated subgroup. These data suggest that there are significant age-related alterations in salivary function.