Salivary exoglycosidases in the detection of early onset of salivary gland involvement in rheumatoid arthritis.

INTRODUCTION: The aim of the present study was to evaluate the possibility of making use of the specific activity of N-acetyl-ß-hexosaminidase, its isoenzymes and ß-glucuronidase--potential indicators of salivary gland damage--in the detection of early onset of salivary gland impairment in RA, which is also demonstrated by xerostomia. MATERIAL/METHODS: For this purpose RA xerostomic salivary patients (unstimulated salivary flow >0.1 mL/min) were compared with RA xerostomic hyposalivary patients (unstimulated salivary flow ≤0.1 mL/min), RA patients without xerostomia (unstimulated salivary flow >0.1 mL/min) and generally healthy controls (unstimulated salivary flow >0.1 mL/min, without xerostomia). Salivary N-acetyl-ß-hexosaminidase, its isoenzymes A and B, and ß-glucuronidase specific activity were determined according to the Marciniak et al. method. The protein content in the unstimulated saliva was determined by the bicinchoninic acid method. RESULTS: In xerostomic rheumatoid arthritis patients, the specific activity of salivary ß-glucuronidase and isoenzyme A was significantly higher than in the healthy controls but the specific activity of salivary N-acetyl-ß-hexosaminidase, its isoenzyme B and ß-glucuronidase was significantly lower than in xerostomic hyposalivary rheumatoid arthritis patients. CONCLUSIONS: We suggest a simple, safe and cheap method for the determination of exoglycosidases as a useful tool for the diagnosis of early stages of salivary gland involvement in rheumatoid arthritis.

Chitinase expression in parotid glands of non-obese diabetic mice.

OBJECTIVE: This investigation was a basal study that used a mouse model of xerostomia to identify protein biomarkers of xerostomia in saliva. We identified genes expressed differently in parotid glands from non-obese diabetic mice with diabetes and those from control mice; subsequently, we investigated expression of the proteins encoded by these genes in parotid glands and saliva. MATERIALS AND METHODS: DNA microarray and real-time PCR analyses were performed to detect differences between NOD/ShiJcl and C57BL/6JJcl (control) female mice in gene expression from parotid glands or parotid acinar cells. Subsequently, protein expression was assessed using immunoblotting and immunohistochemistry. Similarly, enzyme activity in saliva was assessed using zymography. RESULTS: Based on gene expression analyses, Chia expression was higher in diabetic mice than non-diabetic mice and control mice; similarly, expression of chitinase, the protein encoded by Chia, was higher in diabetic mice. Saliva from NOD/ShiJcl mice had more chitinase than saliva from control mice. CONCLUSIONS: Chitinase was highly expressed in parotid acinar cells from diabetic mice compared with non-diabetic and control mice. Increased chitinase expression and enzyme activity may characterize the autoimmune diabetes in mice; however, further investigation is required to assess its use as a biomarker of xerostomia in humans.

Effect of diode laser on enzymatic activity of parotid glands of diabetic rats.

The aim of this study was to evaluate the effect of laser irradiation (LI) on enzymatic activities of amylase, catalase and peroxidase in the parotid glands (PG) of diabetic and non-diabetic rats. Ninety-six female rats were divided into eight groups: D0; D5; D10; D20 and C0; C5; C10; C20, respectively. Diabetes was induced by administration of streptozotocin and confirmed later by the glycemia results. Twenty-nine (29) days after the induction, the PGs of groups D5 and C5; D10 and C10; D20 and C20, were irradiated with 5 J/cm(2), 10 J/cm(2) and 20 J/cm(2) of laser diode (660 nm/100 mW) respectively. On the following day, the rats were euthanized and the enzymatic activity in the PGs was measured. Diabetic rats that had not been irradiated (group D0) showed higher catalase activity (P < 0.05) than those in group C0 (0.14 +/- 0.02 U/mg protein and 0.10 +/- 0.03 U/mg protein, respectively). However, laser irradiation of 5 J/cm(2) and 20 J/cm(2) decreased the catalase activity of the diabetic groups (D5 and D20) to non-diabetic values (P > 0.05). Based on the results of this study, LI decreased catalase activity in the PGs of diabetic rats.

In vitro secreted aspartyl proteinase activity of Candida albicans isolated from oral diseases and healthy oral cavities.

The in vitro secreted aspartyl proteinase (SAP) activity of Candida albicans isolated from a variety of oral conditions, including healthy oral cavities, was determined. SAP activity (units/10(6) cells/ml, +/-SD) was 0.28 +/- 0.33 for pseudomembranous candidosis isolates (n = 18), 0.35 +/- 0.46 for chronic erythematous candidosis isolates (n = 21) and 0.30 +/- 0.32 for chronic hyperplastic candidosis isolates (n = 50). SAP activity of 0.19 +/- 0.22 was recorded for isolates from squamous cell carcinoma (n = 18), 0.26 +/- 0.37 for burning mouth syndrome isolates (n = 29), 0.25 +/- 0.38 for isolates from xerostomia (n = 15) and 0.39 +/- 0.50 for isolates from lichen planus (n = 13). The SAP activity of isolates from oral disease states was significantly (P < 0.05) higher than that recorded for 28 isolates from healthy mouths (activity of 0.04 +/- 0.03). However, there was no significant difference in the SAP activity between the three forms of clinical oral candidosis (P > 0.05). SAP activity was inhibited in control samples containing the SAP inhibitor, pepstatin A. These results indicate that C. albicans strains associated with oral disease have inherently higher SAP activity.

Serum pilocarpine esterase activity and response to oral pilocarpine.

Pilocarpine is used orally to treat xerostomia but patients vary widely in their response and ability to tolerate this drug. To elucidate the potential pharmacokinetic contribution of serum to this variability, the enzymatic hydrolysis of pilocarpine in human serum in vitro was investigated using a stability indicating HPLC assay. The reaction at 37 degrees C follows Michaelis-Menten kinetics (K(m) = 2.78 +/- 0.48 mmol/liter, Vmax = 79 +/- 13 nmol min-1 ml 1; n = 5) and produces pilocarpic acid as the only detectable product. The distribution of pilocarpine esterase activity in a group of healthy young adults at age 21 (n = 163; 87 males, 76 females) was examined by incubating serum samples with pilocarpine (10 mmol/ liter) at 37 degrees C for 60 min. The distribution was positively skewed and ranged from 4 to 132 nmol min-1 ml-1 with a mean value of 55 +/- 23 nmol min-1 ml 1. The means for males and females were not significantly different. Similar measurements in xerostomia patients undergoing treatment with oral pilocarpine showed that those with higher serum esterase activity tolerated pilocarpine well and tended to require higher doses for relief of xerostomia, whereas those with low activity were sensitive to the adverse effects of the drug and were adequately treated with a lower dose. The results suggest that at least some of the variability in response to oral pilocarpine is due to differences in serum pharmacokinetics.

[Analysis of the effectiveness of pilocarpine in the management of xerostomia]. / A pilocarpin hatékonyságágak elemzése a xerostomia kezelésében.

The authors have studied the "sympathetic like" side effect of pilocarpine after single injection (1 mg/100 g b.w., i.p.) of the drug. They developed a system for the continuous automatic recording the amylase activity of the saliva secreted by the parotid glands of rats, "in situ". Pilocarpine stimulus was characterized by a peak in amylase activity--regularly observed in the first 40-60 min--which was additive to the cholinergic amylase secretory response. After this the amylase secretion was continued with lower activity. The role of a beta-adrenergic component in the pilocarpine stimulus appears to be supported by the finding that propranolol (2.5 mg/100 g b.w., i.p.) pretreatment applied 30 min prior to the pilocarpine stimulus prevented the appearance of the characteristic amylase peak. These data support that the beta-adrenergic side effect triggering the periodical synthesis of export proteins during the course of pilocarpine treatment accounts for the selative efficiency of pilocarpine in the therapy of xerostomia.

Biochemical diagnosis of reduced salivary gland function.

The volume of the secretion of saliva is decreased in elderly people. The volume of salivary secretion varies directly with acid DNase activity. Neutral DNase activity, however, shows no significant variation.

Study of the ductal epithelial cell component of human labial salivary glands in vitro.

Salivary gland cultures were propagated from primary explant cultures of 55 human labial gland biopsies. Cultures were maintained for at least 7 days in 199 medium plus 20% newborn calf serum and growth measured at this time both by cell counting and planimetry of the area. Ductal cell population identification was undertaken on the basis of an intra-cellular function, the latter being the histochemical detection of the enzyme 11 beta-hydroxysteroid dehydrogenase. Epithelioid cell lines were derived by enzymatic and mechanical means. Non-invasive quantitation of the proportion of ductal epithelial cells present in primary explant cultures, and derived cell lines, was attempted using a radioimmunoassay to detect conversion in the media of cortisol to cortisone. The cell lines were terminated after undergoing 18 passages over 20 weeks. By this time the epithelioid morphology of cells could no longer be equated to a differentiated epithelial cell origin since conversion of cortisol to cortisone in the growth media no longer occurred.

The effect of radiation-induced xerostomia on saliva and serum lysozyme and immunoglobulin levels.

Saliva and serum lysozyme, immunoglobulin, albumin, and total protein levels were monitored in thirty patients with cancer of the head or neck before, during, and after radiotherapy and compared with those of a group of non-irradiated noncancer control subjects. The mean volume-based saliva lysozyme and total protein concentrations were significantly higher in the cancer patients before radiotherapy than in the control group. During radiotherapy, the mean volume-based concentrations of all protein components assayed increased as the saliva flow rate decreased. Protein-based ratios of saliva albumin, IgG, and lysozyme and the ratio of IgG/IgA increased as the xerostomia intensified. Ratios of saliva total protein and IgA to flow rate paralleled the flow rate decrease. Such increased concentrations in saliva immunoproteins were offset, however, by a greater than 93 per cent reduction in total saliva output. This reduced saliva output, therefore, resulted in an immunoprotein deficit. There were no significant differences between the mean serum lysozyme levels of the cancer and control groups at any point of comparison. The mean serum immunoglobulin concentrations in the cancer patients before radiotherapy were significantly higher than those in the control group. During radiotherapy, there was a decrease in the mean serum total protein, albumin, and immunoglobulin levels which reverted toward the pretreatment values during the postirradiation period.