Histological and biochemical characterization of von Ebner's glands in the Syrian hamster; comparison with rat von Ebner's glands.

We report here for the first time a morphological description and observations on some of the secretory proteins of the von Ebner's lingual salivary glands (VEG) of the Syrian hamster. Hamster VEG were macroscopically less distinct, but histologically similar to rat VEG. VEG extracts of hamster and rat were assayed for lipase, alpha-amylase and peroxidase activities. Unlike rat VEG, which is rich in lipase activity, hamster VEG extract had no detectable lipase activity and did not react with antibodies to either rat lingual lipase or human gastric lipase in Western blots. Immunohistochemical reactions with the anti-rat lingual lipase antibody were very weak in hamster VEG and strong in rat VEG. Moderate alpha-amylase enzyme activities and immunohistochemical reactions were demonstrated in both hamster and rat VEG. Peroxidase activity was negligible in the VEG, unlike the high activity in the submandibular glands of both species. An 18 kDa von Ebner's gland protein (VEGP), a member of the lipocalin superfamily of hydrophobic ligandbinding proteins, was abundant in rat VEG, but not detected in hamster VEG. Thus, hamster VEG differs from rat VEG in macroscopic appearance and the absence of lipase and VEGP. It is similar to rat VEG histologically and with regard to the presence of alpha-amylase and absence of peroxidase.

Combined role of Casoni test and indirect haemagglutination test in the diagnosis of hydatid disease.

PURPOSE: To study the combined role of Casoni test and indirect haemagglutination (IHA) test in the diagnosis of hydatid disease. METHODS: Twenty eight suspected cases of hydatid disease were subjected to Casoni intradermal test using commercially available antigen (Span Diagnostics, India) after collecting pre-test serum samples. The serum samples were tested by IHA using an indigenously developed IHA test. RESULTS: Considering the clinical diagnosis of hydatid disease as the gold standard, the specificity of both Casoni test and IHA was 47%, however, the sensitivity of IHA was higher (81.8%) than Casoni test (63.8%). With the two tests considered together the sensitivity was 90.9%. CONCLUSIONS: Combined use of Casoni test and IHA test could establish presumptive and cost effective diagnosis in upto 90.9% of clinically suspected cases of hydatid disease.

Abundant secretory lipocalins displaying male and lactation-specific expression in adult hamster submandibular gland. cDNA cloning and sex hormone-regulated repression.

We have previously identified massively expressed 24- and 20.5-kDa male-specific proteins in submandibular salivary glands (SMG) of adult hamsters. Here we report the cloning of the cDNA encoding the 24-kDa protein which we have now found to be a heterogenously N-glycosylated form of the 20.5-kDa protein. The deduced amino acid sequence indicated that the protein is a member of the lipocalin family, the two most related lipocalins being rat odorant-binding protein of nasal mucosa and aphrodisin, a pheromonal protein present in vaginal discharge and saliva of female hamsters. Northern blot analysis showed that cognate mRNA is expressed in hamster SMG and lacrimal gland (LG) displaying marked sex-hormonal repression. The sex-hormonal repression patterns showed similarities and dissimilarities between SMG and LG but they were, respectively, similar to the sex-hormonal repression pattern noted for the SMG 24/20.5-kDa male-specific proteins and for an abundant female-specific 20-kDa LG secretory protein. These SMG and LG proteins were found to be immunologically similar and secretion of the SMG proteins in saliva and their excretion in urine was detected. The male-specific and abundant expression of the SMG proteins were seen at and after sexual maturity but was not dependent on androgens. Surprisingly, a temporary male-like expression of these SMG proteins was seen in lactating females which was obliterated by oestrogen administration. Our results show that despite differences in their repression by sex hormones, the gene for SMG 24/20.5-kDa proteins is similar or identical to that of LG 20-kDa protein and their marked repression by both androgens and oestrogens might be at the transcriptional level. Moreover, they might be excellent models with which to study sex hormone repression of gene expression at the molecular level. The results of homology search and the male- and lactation-specific pressure of the SMG proteins in adult saliva and urine suggests a possibility of their involvement in olfaction-mediated chemical communication between hamsters.

Hormonal effects on hamster lacrimal gland female-specific major 20 kDa secretory protein and its immunological similarity with submandibular gland major male-specific proteins.

Hormonal regulation of a major 20 kDa protein of hamster exorbital lacrimal gland (LG) was studied by SDS-PAGE profile analysis and the purified protein's antisera was used to screen tissues of hamster and other species for crossreacting proteins. This protein was seen in female LG but not in males and late-pregnant or hCG-treated females. Low estrogen state in females after gonadectomy, prolonged light-deprivation, prolonged starvation or lactation increased its level several folds to approximately 20% of LG soluble proteins and similar levels were induced in males after gonadectomy (low androgen state). However, light-deprivation or melatonin treatment-induced low androgen state in males had no effect. In gonadectomized hamsters, this LG protein was obliterated on treatment with androgens, estrogens or thyroid hormones. Only estrogen inhibition of LG 20 kDa was prevented by simultaneous tamoxifen administration. Simultaneous treatment of gonadectomized hamsters with gonadotrophins and estrogen/androgen did not prevent the LG 20 kDa protein's inhibition. Relative potencies of estrogens (3.6 microg daily dose) were: estradiol-17beta approximately diethylstilbestrol > estrone > estradiol-17alpha, while estriol and chlorotrianisene had no effect. Dexamethasone, progesterone, prolactin, hypothyroid state or adrenalectomy had no effect on LG 20 kDa expression. Western blot studies confirmed the marked repression of LG 20 kDa by estrogen androgen and thyroid hormone and detected the protein in tears of females and gonadectomized hamsters but not in males. Interestingly, among other tissues tested, crossreaction was only seen with the estrogen-repressed 24 and 20.5 kDa major male-specific secretory proteins of hamster submandibular glands (SMG) which were previously reported by us. This strongly indicated that the LG and SMG proteins are products of the same or closely related genes. A possible role for these hamster sex-specific LG and SMG major secretory proteins in olfactory communication is suggested.

Natural history of severe duodenal ulcer disease.

BACKGROUND: A subset of patients with chronic duodenal ulcer has severe ulcer diathesis in the form of frequent relapses and complications like perforation and hemorrhage. We observed the effect of drug treatment on the natural history of this subset. METHODS: Of 526 patients diagnosed to have chronic duodenal ulcer by endoscopy, 23 patients with severe diathesis were available for long follow-up (mean period 36 months). Each patient was assessed clinically and endoscopically every 2 months for at least 12 months and then every 3 months or when symptomatic. Helicobacter pylori status was assessed during endoscopy. The effect of antisecretory drugs and anti-H. pylori therapy on natural history was determined. RESULTS: Thirteen of 23 patients (56%) had refractory ulcers; six responded to double dose of H2-receptor antagonists (H2RA) for 8 weeks and six to omeprazole 40 mg daily for 4-8 weeks. Of 20 patients (87%) who were H. pylori-positive, 15 completed triple-drug therapy; of these, 10 patients eradicated H. pylori. These 10 patients were followed up for 24 months; there were no ulcer relapses within the first 12 months but 8 of them relapsed between 12 and 24 months (total number of relapses 8). Reinfection with H. pylori occurred in 3 patients. In the other 10 patients who remained H. pylori-positive, there were 19 episodes of ulcer relapse in 7 patients over 24 months, in spite of maintenance therapy with H2RA (p < 0.05). CONCLUSIONS: Refractoriness in patients with severe ulcer disease is usually episodic and amenable to larger doses of omeprazole or H2RA. Anti-H. pylori therapy improves the natural history but its effect in preventing ulcer relapse is short lasting (less than 12 months). Recurrence of infection is a problem in our population.

Clinicopathologic study of lupus nephritis.

Needle biopsies of kidney were done in 35 cases of systemic lupus erythematosus (SLE) with renal lesions. The lupus nephritis were classified according to WHO classification and were correlated with response to therapy and prognosis. Detailed clinical features, routine haematological, biochemical tests (e.g., serum urea, creatinine, total protein and albumin, cholesterol, etc), examination of urine (degree of proteinuria and cells) and occurrence of various auto-antibodies e.g., antinuclear antibody (ANA), anti double stranded DNA (anti DsDNA) by enzyme immunoassay (EIA) method, LE cells and rheumatoid factor (RF) were studied in all cases. Clinically hypertension was present in 19 (54.3%) cases and nephrotic range of proteinuria was detected in 20 (57.2%) cases. ANA was found in 31 (88.5%) cases, anti DsDNA 24 (68.5%) and LE cells were detected in 25 (71.5%) cases. RF was detected in 2 (5.7%) cases. Histologically the most frequent lesions were class IV occurring in 15 cases (42.8%) with initial complete remission achieved only 4 cases by immunosuppressive therapy. Active lesions were also most frequent in this class. Class III lesions were found in 8 (22.8%) cases with 6 cases had complete remission. The best prognosis was noted in class II cases with 4 out of 5 (14.3%) cases had complete remission. Class V lesions were found in 6 (17.2%) cases with complete remission achieved in 3 cases. Only one patient presented with class VI lesion. RF positive cases had milder renal lesions.

Sex-hormonal regulation of 20.5 and 24 kDa major male-specific proteins in Syrian hamster submandibular gland.

Examination of SDS-PAGE protein profiles of adult hamster submandibular glands revealed a marked sexual dimorphism in levels of 20.5 and 24 kDa proteins. These proteins, together apparently constituting 40% of soluble proteins, were present in intact and castrated males but not in females. This sexual dimorphism was also confirmed in two-dimensional gels which revealed that the protein at 24 kDa was heterogenous and consisted of at least four species. In females, ovariectomy induced the 20.5 and 24 kDa proteins to male levels. Estrogen administration to gonadectomized hamsters of either sex or to intact males obliterated these proteins. Whereas androgens had no effect in males, they markedly inhibited these proteins in ovariectomized females. Androgens were less potent than estrogens in this regard. Progesterone or dexamethasone had no effect on these proteins. Only male saliva contained 20.5 and 24 kDa major proteins. These submandibular male-specific proteins are useful markers to study the hormonal regulation of this gland, and they provide a model system to study how both androgens and estrogens mediate inhibition of protein synthesis. The possible functions of these submandibular male-specific proteins are discussed.

Pulmonary infection caused by Mycobacteria other than M. tuberculosis in and around Calcutta.

A study was undertaken to determine the prevalence rate of atypical mycobacteria in indolent fibrocavitary pulmonary diseases. 450 sputum specimens were examined and cultured, of which 15 were identified as atypical mycobacteria on repeated culture. No previous study was recorded from Calcutta. Compared to the results of previous workers from different regions of India, the prevalence rate seems high in this part of the country.

PIP: The prevalence of atypical mycobacteria-induced indolent fibrocavity pulmonary disease was investigated in 450 patients attending the outpatient chest clinic at S.S.K.M. Hospital in Calcutta, India, with clinical symptoms suggestive of pulmonary tuberculosis. Early morning sputum specimens were collected and cultured. 86 (19.1%) of the 450 specimens were positive for AFB. Of these, 15 (17.4%) were confirmed to be atypical mycobacteria on repeated testing (3 cultures) and the remaining 71 were Mycobacterium tuberculosis. The 17.4% prevalence rate of atypical mycobacteria identified in this study is substantially higher than that reported from other parts of India (0-8.4%).

Comparative in vitro activity of fluoroquinolones against Mycobacterium tuberculosis.

Sixty clinical isolates of M. tuberculosis, 24 susceptible and 36 resistant to conventional primary antituberculous drugs, were tested against four fluoroquinolones (ciprofloxacin, ofloxacin, pefloxacin and norfloxacin). Ofloxacin and ciprofloxacin were found to be the most active, with minimum inhibitory concentration (MIC) of 0.6 mg/l or less to all strains tested. Strains resistant to isoniazid and other antitubercular drugs also showed more or less equal MICs for these two drugs. Mycobacterium tuberculosis H37 Rv showed MIC < 0.6 mg/l on each occasion. Other agents viz., norfloxacin and pefloxacin showed lesser activity against all these strains tested in comparison to ciprofloxacin and ofloxacin.

Androgens and estrogens markedly inhibit expression of a 20-kDa major protein in hamster exorbital lacrimal gland.

We report here for the first time a 20-kDa hamster lacrimal gland major protein whose expression is markedly inhibited by physiological levels of both androgens and estrogens. This novel protein was present in adult females but not in males. In females, its level was several fold elevated on ovariectomy to apparently 20% of total soluble proteins. Castration in males induced this 20-kDa protein from undetectable to ovariectomized female levels. Administration of only androgens or estrogens to gonadectomized hamsters of both sexes obliterated this major lacrimal protein and estrogens were more potent than androgens. The 20-kDa lacrimal protein was secreted only in female tears. This lacrimal 20-kDa protein of yet unknown function is a useful marker to study the hormonal regulation of the lacrimal gland and it provides a model system to study how both androgens and estrogens mediate inhibition of a specific protein's expression.

Thiocyanate, a plausible physiological electron donor of gastric peroxidase.

Gastric peroxidase (GPO) was purified to apparent homogeneity to characterize its major physiological electron donor. The enzyme (RZ = 0.7), with a subunit molecular mass of 50 kDa, is a glycoprotein, with a relative abundance of aspartic and glutamic acid over arginine and lysine. It has a Soret maximum at 412 nm, which is shifted to 426 nm by H2O2 due to formation of compound II. Although the physiological electron donors I-, Br- and SCN-, but not Cl-, are oxidized by GPO optimally at acid pH, only I- and SCN- are oxidized appreciably at physiological pH. Considering that the I- concentration in stomach is less than 1 microM, whereas the SCN- concentration is about 250 microM, SCN- may act as a major electron donor for GPO. Moreover, SCN- oxidation remains unaltered in the presence of physiological concentrations of other halides. The second-order rate constant for the reaction of GPO with H2O2 (k1) and compound I with SCN- (k2) at pH 7 was found to be 8 x 10(7) M-1.s-1 and 2 x 10(5) M-1.s-1 respectively. GPO has significant pseudocatalase activity also in the presence of I- or Br-, but it is blocked by SCN-. The SCN- oxidation product OSCN- may be reduced back to SCN- by cellular GSH, and GSSG may be reduced back to GSH by glutathione reductase and NADPH. In a system reconstituted with pure glutathione reductase, NADPH, GSH, SCN- and H2O2. GPO-catalysed SCN- oxidation could be coupled to NADPH oxidation. This system where GPO utilizes SCN- as the major physiological electron donor may operate efficiently to scavenge intracellular H2O2.

Comparative efficacy of fluoroquinolones, aminoglycosides, ureidopenicillins & newer cephalosporins against Pseudomonas species.

A total of 74 strains of Pseudomonas aeruginosa and 18 strains of Ps. putrefaciens were tested for sensitivity to 14 different antimicrobial agents. Ps. aeruginosa were mostly sensitive to netilmicin (81%), piperacillin (78%), amikacin (73%), azlocillin (70%), ceftazidime (69%) and pefloxacin (65%). Only 66 per cent strains of Ps. putrefaciens were sensitive to netilmicin, ceftazidime and ceftriaxone. The MIC values of the different drugs for the sensitive strains were comparable with the results of susceptibility testing. The Ps. putrefaciens strains showed greater resistance than Ps. aeruginosa.

Tissue distribution of constitutive and induced soluble peroxidase in rat. Purification and characterization from lacrimal gland.

A thorough search for a soluble peroxidase in 31 different tissues of rat indicated the presence of a constitutive activity only in lacrimal, preputial and submaxillary gland. An induced soluble peroxidase activity was also detected in the lactating mammary gland and in the estrogen-induced uterine secretory fluid. The lacrimal gland was the richest source of the enzyme. No peroxidase activity was detected in the lactating mammary gland of mouse and hamster nor in the preputial gland of mouse and uterine fluid of hamster. The three constitutive and two induced soluble peroxidases of rat had a native molecular mass of 73 kDa by gel filtration and they showed a similar mobility in native PAGE. Lactoperoxidase of cow's milk and solubilized rat membrane-bound peroxidases of uterus, intestine and bone marrow showed in native PAGE a mobility which was distinctly different from that of rat soluble peroxidases. As the lacrimal gland of rat was the richest source of soluble peroxidase, the enzyme was purified from this gland to apparent homogeneity; SDS/PAGE then showed a single band of molecular mass 75 kDa which was similar to that obtained by gel filtration. Peroxidase also purified from preputial and submaxillary gland, as well as commercial lactoperoxidase, had a similar molecular mass on SDS/PAGE to purified lacrimal peroxidase. The visible spectrum of lacrimal peroxidase was similar to that of lactoperoxidase but different from membrane-bound peroxidase of rat neutrophils. On isoelectric focussing, purified lacrimal peroxidase resolved into about 14 multiple forms spanning a pI range of 6.5-3.5 while lactoperoxidase focussed at the cathode. Evidence presented suggests that the multiple forms are possibly due to differences in glycosylation. Immunodiffusion, immunoprecipitation and Western blot using antilacrimal peroxidase serum showed a similar interacting species for all five soluble peroxidases of rat while membrane-bound peroxidases showed no interaction. Although in immunodiffusion, the antiserum failed to cross-react with lactoperoxidase it did interact with lactoperoxidase on Western blot. The results indicate that the various constitutive and induced soluble peroxidases of rat tissues are similar to lacrimal peroxidase but are distinctly different from the known membrane-bound peroxidases of rat. However the lacrimal peroxidase shows both similarities as well as dissimilarities with bovine lactoperoxidase. This soluble peroxidase system of rat could be useful to study tissue-specific regulation of gene expression at the molecular level.

Purification and characterization of a soluble peroxidase of rat preputial gland: comparison with lactoperoxidase.

A highly active soluble peroxidase (donor: H2O2 oxidoreductase EC 1.11.1.7) has been purified from the preputial gland of the rat by hydroxylapatite chromatography, ammonium sulfate fractionation, Sephadex gel filtration and affinity chromatography on con A-Sepharose. The enzyme shows apparent homogeneity when analysed by acid and alkaline-PAGE. Its molecular, spectral, kinetic and catalytic properties were compared with those of bovine lactoperoxidase (LPO). Preputial gland peroxidase (PPO) is a glycoprotein of molecular weight of 70-73 kDa slightly lower (78 kDa) than that of LPO. Using isoelectric focussing, PPO was resolved into eight different closely spaced protein species spanning a pI range of 5.4 to 6.4, while LPO focuses into several closely spaced protein bands in the pI range 8.5-9.3. PPO is similar to LPO in its spectral (Soret) and some kinetic properties, but it differs significantly from LPO in substrate (H2O2) tolerance and substrate inactivation. PPO also differs from LPO in showing differential inactivation by SDS. Both enzymes are glycoproteins and although concanavalin A (con A) showed a variable interaction with both enzymes, wheat germ agglutinin interacted specifically with LPO only. We suggest that PPO, the secretory peroxidase of the preputial gland, differs significantly from LPO in its molecular and catalytic properties.

Incidence of neonatal bacterial infection in cases of prolonged rupture of amniotic membrane.

Thirty cases of premature rupture of amniotic membrane were studied bacteriologically. Twenty cases showed growth of organisms from one or more sites at birth. From 20 control cases no pathogenic organism could be isolated. Out of these 20 cases showing growth of organisms 8 neonates developed clinical infection (deep and superficial) subsequently. The organisms grown from vaginal swab and umbilical swab culture showed maximum correlation to the organism from the infected neonates. Esch coli was found to be most common organism causing infections, next to it was Staph aureus. Growth of anaerobic organisms was found only in cases of prolonged rupture of membrane in mother, but not isolated from infected babies. So bacteriological studies of vaginal, placental, umbilical and nasal swabs and cord blood just after the birth of baby may be of some help in predicting the onset of neonatal infection.

Immunological characterization of soluble peroxidases from rat tissues including preputial gland.

A highly active soluble peroxidase has been identified in the preputial gland of rats and characterized immunologically along with other soluble peroxidases of a number of rat tissues such as submaxillary gland, exorbital lacrimal gland and also of the uterine fluid of the estrogen treated rats. All these peroxidases have the native molecular weight around 73K as determined by gel filtration on Sephadex G-150. An antiserum raised against the pure bovine lactoperoxidase interacts with all these soluble peroxidases and immunoprecipitates the enzyme activity in a similar fashion when titrated against varied concentration of the antiserum. Following electrophoretic transfer to nitrocellulose by Western blotting, the antiserum crossreacts with the preputial, submaxillary and lacrimal gland protein of molecular weight around 73K and with the uterine fluid protein of molecular weight of 80K. An additional crossreacting protein of molecular weight of 80K is also evident in the lacrimal gland. All these enzyme preparations, however, contain another immunoreactive protein of molecular weight of about 64K. While 73-80K molecular weight interacting proteins may represent different forms of peroxidase, presumably with varied carbohydrate moieties, 64K molecular weight protein may be a precursor of the peroxidase which after posttranslational modification such as heme conjugation and glycosylation leads to formation of native enzyme. Rat harderian gland, unlike bovine origin, does not contain any detectable peroxidase activity.(ABSTRACT TRUNCATED AT 250 WORDS)