Antibacterial activity of polyphenolic fraction of Kombucha against Vibrio cholerae: targeting cell membrane.

The present study was undertaken to determine the mechanism of antibacterial activity of a polyphenolic fraction, composed of mainly catechin and isorhamnetin, previously isolated from Kombucha, a 14-day fermented beverage of sugared black tea, against the enteropathogen Vibrio cholerae N16961. Bacterial growth was found to be seriously impaired by the polyphenolic fraction in a dose-dependent manner. Scanning Electron Microscopy demonstrated morphological alterations in bacterial cells when exposed to the polyphenolic fraction in a concentration-dependent manner. Permeabilization assays confirmed that the fraction disrupted bacterial membrane integrity in both time- and dose-dependent manners, which were proportional to the production of intracellular reactive oxygen species (ROS). Furthermore, each of the polyphenols catechin and isorhamnetin showed the ability to permeate bacterial cell membranes by generating oxidative stress, thereby suggesting their role in the antibacterial potential of Kombucha. Thus, the basic mechanism of antibacterial activity of the Kombucha polyphenolic fraction against V. cholerae involved bacterial membrane permeabilization and morphological changes, which might be due to the generation of intracellular ROS. To the best of our knowledge, this is the first report on the investigation of antibacterial mechanism of Kombucha, which is mostly attributed to its polyphenolic content. SIGNIFICANCE AND IMPACT OF THE STUDY: The emergence of multidrug-resistant Vibrio cholerae strains has hindered an efficient anti-Vibrio therapy. This study has demonstrated the membrane damage-mediated antibacterial mechanism of Kombucha, a popular fermented beverage of sugared tea, which is mostly attributed to its polyphenolic content. This study also implies the exploitation of Kombucha as a potential new source of bioactive polyphenols against V. cholerae.

Attenuation of the cyproterone acetate-induced testicular hypofunction by a novel nutraceutical lycopene: a genomic approach.

This study was designed to explore the cyproterone acetate (CPA)-induced andrological hypofunction and its correction by oral administration of lycopene. In this concern, spermatogenic, biochemical, histological and genomic profiles were studied. Cyproterone acetate administration for 1 month helped to develop infertile model rats. A significant recovery was noted in sperm motility, sperm count, sperm viability, hypo-osmotic swelling tail-coiled spermatozoa; activities of testicular ∆5 , 3ß-hydroxysteroid dehydrogenase (HSD), 17ß-HSD, catalase (CAT) and superoxide dismutase (SOD); and levels of conjugated diene (CD), malondialdehyde (MDA), testicular cholesterol and serum testosterone after the administration of lycopene at 1.5 mg/0.5 ml Tween-80/100 g body weight/day for last 1 month to infertile model rats. Simultaneously, qRT-PCR study of Bax, Bcl-2, caspase-3, ∆5 , 3ß-HSD and 17ß-HSD genes in testicular tissue showed a significant rectification towards the control in CPA-pre-treated cum CPA-lycopene-cotreated rats. Side-by-side histological and histometric studies showed a significant correction in qualitative analysis of spermatogenesis and seminiferous tubular diameter (STD) in CPA-pre-treated cum CPA-lycopene-cotreated rats. Lycopene showed outstanding efficacy in the management of CPA-induced testicular hypofunction with special reference to correction in oxidative stress-induced testicular apoptosis at genomic level.

Antiapoptotic efficacy of seed of Eugenia jambolana on testicular germ cell in experimental diabetic rat: a genomic study.

This study was designed to focus the genetic regulation of diabetes-induced testicular hypofunction and its amelioration by ethyl acetate fraction of seed of Eugenia jambolana. In this regard, we have assessed relevant biosensors such as biochemical, spermiological, histological and gene expression of antioxidant enzymes, germ cell apoptosis and androgenic key enzymes along with in situ end labelling and DNA fragmentation study. After 60 days administration of said fraction, significant recovery in the glycated haemoglobin, serum testosterone, sperm viability, hypo-osmotic swelling and nuclear chromatin decondensation were noted in fraction-treated diabetic group in comparison with diabetic control. Besides this, a significant recovery in the expression of Bax, Bcl-2, caspase-9, caspase-3, catalase, peroxidase, ∆(5) , 3ß-hydroxy steroid dehydrogenase and 17ß-hydroxy steroid dehydrogenase genes was noted towards the control in ethyl acetate fraction-treated group. Testicular histology focused a significant recovery in the number of different generation of germ cells at stage VII of spermatogenesis in fraction-treated group. In situ end labelling and DNA fragmentation study of testicular tissues also showed a significant recovery in fraction-treated group towards the control. These findings indicate that the ethyl acetate fraction showed outstanding antiapoptotic activity by neutralising oxidative stress as well as by the improvement in glycaemic sensors.

Al2O3 influence on structural, elastic, thermal properties of Yb(3+) doped Ba-La-tellurite glass: evidence of reduction in self-radiation trapping at 1µm emission.

Ba-La-tellurite glasses doped with Yb(3+) ions have been prepared through melt quenching technique by modifying their composition with the inclusion of varied concentration of Al2O3 to elucidate its effects on glass structural, elastic, thermal properties and Yb(3+) ion NIR luminescence performance. The FTIR spectral analysis indicates Al2O3 addition is promoting the conversion of BOs from NBOs which have been generated during the process of depolymerisation of main glass forming TeO4 units. The elastic properties of the glass revealed an improved rigidity of the glass network on addition of Al2O3. In concurrence to this, differential thermal analysis showed an increase in glass transition temperature with improved thermal stability factor. Also, Yb(3+) fluorescence dynamics demonstrated that, Al2O3 inclusion helps in restraining the detrimental radiation trapping of ∼1µm emission.

Corrective role of Eugenia jambolana on testicular impairment in streptozotocin-induced diabetic male albino rat: an approach through genomic and proteomic study.

The present study was conducted to explore the effect of ethyl acetate fraction of hydro-methanolic (40 : 60) extract of seed of Eugenia jambolana on testicular impairment in diabetic rats. In this respect, biomarkers of oxidative stress, genomics and proteomics in testicular tissue were assessed. Side by side, glycated haemoglobin, serum testosterone, activities of glutamate oxaloacetate transaminase and glutamate pyruvate transaminase in serum, epididymal sperm count including reproductive organosomatic indices were evaluated. Results indicate that a significant recovery (P < 0.05) in the levels of these parameters in fraction-treated diabetic group in comparison with diabetic control. A significant recovery was noted (P < 0.05) in the expression of Bax and Bcl-2 gene towards the control after the treatment of said fraction. Histological study also focused a significant recovery (P < 0.05) in the number of different generation of germ cells at stage VII of spermatogenesis in fraction-treated diabetic group. The said fraction treatment to diabetic rat can recover the activities of serum glutamate oxaloacetate transaminase and glutamate pyruvate transaminase significantly towards the control (P < 0.05). Finally, it may be concluded that ethyl acetate fraction of seed of E. jambolana has a promiseable remedial effect on diabetes-induced testicular dysfunctions in male rat without inducing any metabolic toxicity.

Bioefficacy of hydromethanolic extract of tuber of Chlorophytum borivilianum (Safed Musli) for the management of male infertility in cyproterone acetate-treated albino rats.

Increase in male sexual dysfunction, and its treatment with conventional aphrodisiac drugs with side effects lead to investigate the spermatogenesis and androgenesis augmentative efficacy of hydromethanolic (40 : 60) extract of root of Chlorophytum borivilianum (family - Liliaceae) against cyproterone acetate-induced subfertility in Wistar strain male albino rat. For this purpose, experimental rats were divided into three treatment groups: vehicle (received distilled water), cyproterone acetate (gastric intubation at 250 mg kg(-1) twice daily for 35 days) and cyproterone acetate plus root extract of C. borivilianum (gastric intubation at 250 mg kg(-1) plus 400 mg kg(-1) with an interval of 20 min twice daily for 35 days). After 35-day treatment, all rats were euthanised. Reproductive deviations towards negative side were investigated by screening the spermatogenic and steroidogenic biosensors. Oxidative stress profile in reproductive organs and sperm pellet was evaluated by biochemical assessment of antioxidative enzyme activities and level of end products of the lipid peroxidation. Apoptosis profile was evaluated by Western blot study, TUNEL assay and DNA fragmentation study of testicular tissues. Evaluation of toxicity profile was included for experimental investigation. After cyproterone acetate treatment, the pituitary-testicular axis was deviated towards the negative side and its tuning system was affected by oxidative stress and apoptosis-mediated process, which reduced the quality of semen and finally led to subfertility. Co-administration of C. borivilianum root extract enhanced male reproductive potentiality and prevented the negative deviations after the treatment with cyproterone acetate by means of increasing oxidative defence and maintaining homeostasis in testicular apoptosis process.

Associations between pollen counts, pollutants, and asthma-related hospital admissions in a high-density Indian metropolis.

BACKGROUND: The seasonal pattern of asthma-related hospitalization has often been correlated with ambient allergen/pollutant levels. OBJECTIVE: To examine the relationship between asthma-related hospital admissions (ARHA) and outdoor pollen, spore, and pollutant levels for adult patients in a densely populated Indian megacity Kolkata. METHODS: ARHA data were obtained from two major teaching hospitals of the city. Pollen and spores causing allergic sensitization were identified by skin prick tests (SPTs) among respiratory allergic subjects (N = 1353). Outdoor concentrations of aeroallergens were determined using a Burkard sampler for five consecutive years (2004-2009). Levels of NO(2), SO(2), suspended particulate matters (SPMs), and respirable particulate matters (RPMs) were made available by West Bengal Pollution Control Board (WBPCB, Government of West Bengal). Poisson multivariate Poisson regression (with adjustments for overdispersion) was used to model the data. Results. We found that ARHA in Kolkata increased with predictable regularity in March and September, while remaining low in January and July. SPT showed highly positive skin reactions with grass/weed and palm pollens in respiratory allergic patients, while Aspergilli spores also evoked good sensitivity. In our regression model, the airborne pollen types, Cheno-Amaranthaceae and Cyperaceae, and the inorganic pollutant, SO(2) and RPM, were significantly associated with ARHA (p < .05). CONCLUSION: ARHA in the megacity of Kolkata shows two seasonal peaks that can be correlated with outdoor grass/weed pollen and RPM concentrations. In contrast, the city's ambient fungal spore counts were not found to be significantly associated.

Effect of dielectric dispersion on potentised homeopathic medicines.

BACKGROUND: This paper reports dielectric dispersion occurring in potentised homeopathic medicines subjected to variable frequency electric field using an instrumentation method developed by the authors. Oscillations occur in the direction of electric field, and are usually termed longitudinal/acoustic-mode vibrations. METHODS: The test material was lactose soaked with homeopathic medicine. Multiple resonance frequencies, forming a frequency-set, were observed repeatedly for each medicine. RESULTS: We report experimental results for three potencies of Cuprum metallicum (Cuprum met) in the frequency range of 100kHz-1MHz. Each exhibits a set of resonance frequencies, which may be termed as its characteristic set. As the frequency-set of each medicine is different from those of others, each medicine may, therefore, be identified by its characteristic frequency-set. This suggests that potentised homeopathic medicines, which are chemically identical with the vehicle, differ from one another in the arrangement of vehicle molecules.

Therapeutic effect of ferulic acid, an ethereal fraction of ethanolic extract of seed of Syzygium cumini against streptozotocin-induced diabetes in male rat.

Diabetic therapeutic and antioxidative effects of an ethereal fraction of the ethanolic extract of the seed of Syzygium cumini was studied in streptozotocin (STZ)-induced diabetic rats. Diabetes resulted in a significant elevation in the fasting blood glucose level and in the activity of hepatic glucose-6-phosphatase. There was diminution in the levels of glycogen in the liver and skeletal muscle along with diminution in the activities of hepatic glucose-6-phosphate dehydrogenase, catalase and peroxidase in diabetic rats when compared with controls. Hepatic levels of thiobarbituric acid reactive substance (TBARS) and conjugated dienes (CD) were elevated in respect to control. Oral coadministration of the above fraction to diabetic rats resulted in significant protection in all these parameters. Histological studies of the pancreas showed a qualitative diminution in the area and volume of the islet's of Langerhans, but coadministration of the specific fraction resulted in a significant recovery of the islet's of Langerhans. Chromatography study revealed that the used fraction was ferulic acid (FA). Treatment with FA in normoglycemic rats did not show any significant change in the levels of the selected biosensors. The possible hypothesis for the therapeutic effect of FA against diabetes may be due to its pancreatic beta-cell regenerative effect and/or due to its antioxidant properties.

Aerobiological and immunochemical studies on Carica papaya L. pollen: an aeroallergen from India.

BACKGROUND: Carica papaya L. is a fruit yielding tree, wildly grown or cultivated in the tropics and subtropics. Its pollen grain has been reported to be airborne and cause immunoglobulin E (IgE)-mediated hypersensitivity. OBJECTIVE: To conduct long-term aerobiological study on Carica pollen, along with aeroallergenic particles originating from it and to identify vis-a-vis characterize an important IgE-reactive component present in this pollen. METHODS: The seasonal and diurnal periodicities of airborne C. papaya pollen were recorded in a 5-year survey using a Burkard volumetric sampler. The allergenic potential was studied by skin prick tests, IgE-enzyme-linked immunosorbent assay (ELISA) and also by aeroallergen immunoblotting. The total pollen extract was fractionated by Sephacryl S-200 column, and out of the eluted five fractions, the maximum IgE-reactive fraction (as found in ELISA inhibition) was resolved into five major subfractions in reverse-phase high-performance liquid chromatography (RP-HPLC). The subfraction with optimum IgE reactivity was studied by activity gel, native and nonreducing sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). The homogeneity of the isolated protein fraction was checked by crossed immunoelectrophoresis with rabbit antisera and IgE reactivity was confirmed by ELISA inhibition and immunoblotting using individual patient sera. RESULTS: The Carica pollen occurred in the air round the year with peaks during January and September-October. Among a patient population of 1000, skin-test results showed 27.8% +1 level and 5.6% +2/+3 level reactions. In aeroallergen immunoblotting of exposed Burkard tape segments, the detected allergen spots showed a significant correlation with airborne pollen count recorded. The pollen extract elicited loss of IgE reactivity when treated with reducing agent-like beta-mercaptoethanol and heat, but showed six IgE-reactive components in nonreducing IgE-immunoblot. The fraction 1 eluted from Sephacryl S-200 column showed highest IgE reactivity and resolved into five major components in RP-HPLC. Out of these, the fraction showing optimum IgE reactivity in IgE-ELISA inhibition and immunoblotting with patient antisera, elicited esterase activity and found to be a homogenous protein of 100 kDa. CONCLUSION: Carica papaya tree contributes significantly to the aeropollen and aeroallergen load of the suburban outskirts of Calcutta metropolis, India. The pollen extract contains an important IgE-reactive protein component of 100 kDa molecular weight with esterase activity.

An investigation into the acute and long-term effects of selected yogic postures on fasting and postprandial glycemia and insulinemia in healthy young subjects.

The study was conducted to examine the hypothesis that yogasanas help in the treatment of diabetes mellitus by releasing insulin from the pancreas. Twenty healthy young voluntees (17 male, 3 female; age 19-31 years) participated in the study. Each volunteer performed four sets of asanas in random order for 5 consecutive days each with a 2-day gap between consecutive sets of asanas. The four sets of asanas were: (I) dhanurasana + matsyendrasana, (II) halasana + vajrasana, (III) naukasana + bhujangasana, and (IV) setubandhasana + pavanamuktasana. Blood samples were collected on days 4 and 5 of each set of asanas for measurement of glucose and insulin levels before the asanas, within 10 min after performing the asanas, and 30 min after ingestion of 75 g glucose, which in turn was ingested immediately after the second blood sample. A standard 75 g oral glucose tolerance test (OGTT) was also done before and after the study. On the days of the pre-study or post-study OGTT, no asanas were done. The serum insulin levels after the asanas were lower (P<0.05) than those before the asanas. However, the serum insulin level 0.5 h after the post-asana oral 75 g-glucose challenge was higher (P<0.05) in Set IV than the 0.5 h postprandial insulin level in the pre-study OGTT; the same trend was observed in other sets as well although statistically not significant. The observations suggest that the performance of asanas led to increased sensitivity of the B cells of pancreas to the glucose signal. The increased sensitivity seems to be a sustained change resulting from a progressive long-term effect of asanas. The study is significant in that it has for the first time attempted to probe the mechanism by which yogasanas help diabetes mellitus.

Antidiabetic effect of aqueous extract of seed of Tamarindus indica in streptozotocin-induced diabetic rats.

In Indian traditional system of medicine, herbal remedies are prescribed for the treatment of diseases including diabetes mellitus. In recent years, plants are being effectively tried in a variety of pathophysiological states. Tamarindus indica Linn. is one of them. In the present study, aqueous extract of seed of Tamarindus indica Linn. was found to have potent antidiabetogenic activity that reduces blood sugar level in streptozotocin (STZ)-induced diabetic male rat. Supplementation of this aqueous extract by gavage at the dose of 80 mg/0.5 ml distilled water/100 g body weight per day in STZ-induced diabetic rat resulted a significant diminution of fasting blood sugar level after 7 days. Continuous supplementation of this extract for 14 days resulted no significant difference in this parameter from control level. Moreover, this supplementation produced a significant elevation in liver and skeletal muscle glycogen content, activity of liver glucose-6-phosphate dehydrogenase in respect to diabetic group. Activities of liver glucose-6-phosphatase, liver and kidney glutamate oxaloacetate transaminase (GOT) and glutamate pyruvate transaminase (GPT) activities were decreased significantly in the aqueous extract supplemented group in respect to diabetic group. All these parameters were not resettled to the controlled level after 7 days of this extract supplementation but after 14 days of this supplementation, all the above mentioned parameters were restored to the control level.

Effect of Stephania hernandifolia leaf extract on testicular activity in rats.

AIM: The testicular inhibitory effect of the aqueous fraction of methanol extract of Stephania hernandifolia leaf was studied in male Wistar rats. METHODS: The supernatent and the precipitate part of aqueous fractions of the methanol extract of the leaf were gavaged separately to rat at a similar dose of 200 mg/mL per 100 g body weight per day for 28 days. After cessation of treatment, various observations were conducted. RESULTS: In both treated groups, there were significant decreases in the relative weights of the sex organs, the testicular key androgenic enzymes activities, the plasma level of testosterone, the number of different germ cells at stage VII of seminiferous epithelial cell cycle and the seminiferous tubular diameter in comparison to the controls. Neither of the parts had somatic, renal and hepatic toxicity. This study suggested that the active molecules present in the aqueous fraction of methanol extract of Stephania hernandifolia leaves might be steroids as indicated by thin layer chromatography using specific staining substance for steroid molecules. CONCLUSION: In rats, the aqueous fraction of methanol extract of the S. hernandifolia leaves possesses certain testis-inhibitory substances, which may be steroid-like agents.

Effects of leaf extract of Stephania hernandifolia on testicular gametogenesis and androgenesis in albino rats: a dose-dependent response study.

The present study was undertaken to evaluate the dose-dependent effects of a leaf extract of Stephania hernandifolia on testicular activities in albino rats. Whether this leaf extract has any toxic effect on metabolic organs or on the liver or kidney was studied. Adult male Wistar rats, maintained under standard laboratory conditions, were forcefully fed with the aqueous extract of these leaves at the dose of 2 g or 4 g of leaves/mL distilled water/100 g body weight/day for 28 days. All the animals, along with vehicle-treated controls, were killed on the Day 29 of the experiment. Treatment with this leaf extract at both doses resulted in significant reduction in relative weight in the testis, the seminal vesicles, the prostate, and the epididymis without any significant change in the liver and kidney weight in comparison to control. Activities of testicular steroidogenic key enzymes and plasma testosterone level were decreased significantly, along with a significant reduction in the number of germ cells at stage VII of the spermatogenic cycle and in the seminiferous tubular diameter in both treated groups in comparison to control. Activities of glutamate oxaloacetate transaminase, glutamate pyruvate transaminase, acid phosphatase, and alkaline phosphatase were not altered significantly in the liver and kidney in both treated groups compared with controls. We concluded that treatment with an aqueous extract of leaves resulted in diminution in the activities of testicular androgenic key enzymes and plasma level of testosterone along with inhibition of spermatogenesis without any induction of hepatic and renal toxicity.

Host response to infection: the role of CpG DNA in induction of cyclooxygenase 2 and nitric oxide synthase 2 in murine macrophages.

Depending on sequence, bacterial and synthetic DNAs can activate the host immune system and influence the host response to infection. The purpose of this study was to determine the abilities of various phosphorothioate oligonucleotides with cytosine-guanosine-containing motifs (CpG DNA) to activate macrophages to produce nitric oxide (NO) and prostaglandin E(2) (PGE(2)) and to induce expression of NO synthase 2 (NOS2) and cyclooxygenase 2 (COX2). As little as 0.3 microg of CpG DNA/ml increased NO and PGE(2) production in a dose- and time-dependent fashion in cells of the mouse macrophage cell line J774. NO and PGE(2) production was noted by 4 to 8 h after initiation of cultures with the CpG DNA, with the kinetics of NO production induced by CpG DNA being comparable to that induced by a combination of lipopolysaccharide and gamma interferon. CpG DNA-treated J774 cells showed enhanced expression of NOS2 and COX2 proteins as determined by immunoblotting, with the relative potencies of the CpG DNAs generally corresponding to those noted for the induction of NO and PGE(2) production as well as to those noted for the induction of interleukin-6 (IL-6), IL-12, and tumor necrosis factor. Extracts from CpG DNA-treated cells converted L-arginine to L-citrulline, but the NOS inhibitor N(G)-monomethyl-L-arginine (NMMA) inhibited this reaction. The COX2-specific inhibitor NS398 inhibited CpG DNA-induced PGE(2) production and inhibited NO production to various degrees. The NOS inhibitors NMMA, 1400W, and N-iminoethyl-L-lysine effectively blocked NO production and increased the production of PGE(2) in a dose-dependent fashion. Thus, analogues of microbial DNA (i.e., CpG DNA) activate mouse macrophage lineage cells for the expression of NOS2 and COX2, with the production of NO and that of PGE(2) occurring in an interdependent manner.

Familial hypertrophic cardiomyopathy mutations in the regulatory light chains of myosin affect their structure, Ca2+ binding, and phosphorylation.

The effect of the familial hypertrophic cardiomyopathy mutations, A13T, F18L, E22K, R58Q, and P95A, found in the regulatory light chains of human cardiac myosin has been investigated. The results demonstrate that E22K and R58Q, located in the immediate extension of the helices flanking the regulatory light chain Ca(2+) binding site, had dramatically altered Ca(2+) binding properties. The K(Ca) value for E22K was decreased by approximately 17-fold compared with the wild-type light chain, and the R58Q mutant did not bind Ca(2+). Interestingly, Ca(2+) binding to the R58Q mutant was restored upon phosphorylation, whereas the E22K mutant could not be phosphorylated. In addition, the alpha-helical content of phosphorylated R58Q greatly increased with Ca(2+) binding. The A13T mutation, located near the phosphorylation site (Ser-15) of the human cardiac regulatory light chain, had 3-fold lower K(Ca) than wild-type light chain, whereas phosphorylation of this mutant increased the Ca(2+) affinity 6-fold. Whereas phosphorylation of wild-type light chain decreased its Ca(2+) affinity, the opposite was true for A13T. The alpha-helical content of the A13T mutant returned to the level of wild-type light chain upon phosphorylation. The phosphorylation and Ca(2+) binding properties of the regulatory light chain of human cardiac myosin are important for physiological function, and alteration any of these could contribute to the development of hypertrophic cardiomyopathy.

Steroid dehydrogenase structures, mechanism of action, and disease.

Steroid dehydrogenase enzymes influence mammalian reproduction, hypertension, neoplasia, and digestion. The three-dimensional structures of steroid dehydrogenase enzymes reveal the position of the catalytic triad, a possible mechanism of keto-hydroxyl interconversion, a molecular mechanism of inhibition, and the basis for selectivity. Glycyrrhizic acid, the active ingredient in licorice, and its metabolite carbenoxolone are potent inhibitors of human 11 beta-hydroxysteroid dehydrogenase and bacterial 3 alpha, 20 beta-hydroxysteroid dehydrogenase (3 alpha, 20 beta-HSD). The three-dimensional structure of the 3 alpha, 20 beta-HSD carbenoxolone complex unequivocally verifies the postulated active site of the enzyme, shows that inhibition is a result of direct competition with the substrate for binding, and provides a plausible model for the mechanism of inhibition of 11 beta-hydroxysteroid dehydrogenase by carbenoxolone. The structure of the ternary complex of human 17 beta-hydroxysteroid dehydrogenase type 1 (17 beta-HSD) with the cofactor NADP+ and the antiestrogen equilin reveals the details of binding of an inhibitor in the active site of the enzyme and the possible roles of various amino acids in the catalytic cleft. The short-chain dehydrogenase reductase (SDR) family includes these steroid dehydrogenase enzymes and more than 60 other proteins from human, mammalian, insect, and bacterial sources. Most members of the family contain the tyrosine and lysine of the catalytic triad in a YxxxK sequence. X-ray crystal structures of 13 members of the family have been completed. When the alpha-carbon backbone of the cofactor binding domains of the structures are superimposed, the conserved residues are at the core of the structure and in the cofactor binding domain, but not in the substrate binding pocket.

Effect of ascorbic acid supplementation on liver and kidney toxicity in cyclophosphamide-treated female albino rats.

Effects of ascorbic acid supplementation on the activity of acid phosphatase (ACP), alkaline phosphatase (ALP), glutamic oxaloacetic transaminase (GOT) and glutamic pyruvic transaminase (GPT) on liver, kidney and serum in cyclophosphamide-treated female virgin rats were investigated. Oral administration of cyclophosphamide at the dose of 5 mg/kg body weight/day for 12 days resulted in a significant elevation in ACP and ALP activities in liver, kidney and serum. Ascorbic acid supplementation at the dose of 25 mg/kg body weight/day showed a significant protection in the activity of ACP in liver, kidney and serum, but only in ALP activity in kidney. ALP activities in liver and serum were not restored to control level by ascorbic acid supplementation. Activities of GOT and GPT were elevated significantly in liver, kidney and serum after cyclophosphamide treatment, and were protected and restored to control level by ascorbic acid supplementation.

Structure and mechanism of action and inhibition of steroid dehydrogenase enzymes involved in hypertension.

Members of the NADPH-dependent short chain dehydrogenase/reductase (SDR) family control blood pressure, fertility, and natural and neoplastic growth. Despite the fact that only one amino acid residue is strictly conserved in the 100 known members of the family, all appear to have a dinucleotide-binding Rossmann fold and homologous catalytic residues including the conserved tyrosine. Variation in the binding pocket creates specificity for steroids, prostaglandins, sugars and alcohols. The critically important tyrosine appears to maintain a fixed position relative to the scaffolding of the Rossmann fold and the cofactor position, while the substrate-binding pocket alters in such a way that the dehydrogenation/reduction reaction site is brought into bonding distance of the tyrosine hydroxyl group. Licorice induces high blood pressure by inhibiting an SDR in the kidney. The crystal structure of the complex of 3alpha,20beta-hydroxysteroid dehydrogenase and carbenoxolone reveals the mechanism of enzyme inhibition by licorice. The most potent dehydrogenase enzyme inhibitors are those that displace substrate and cofactor and form strong hydrogen bonds to one or more amino acid residues involved in catalysis.

Origin of substrate specificity of human and rat 17beta-hydroxysteroid dehydrogenase type 1, using chimeric enzymes and site-directed substitutions.

Human 17beta-hydroxysteroid dehydrogenase (17-HSD) type 1 predominantly catalyzes the 17beta-reduction of estrone to estradiol. The present results, however, show that rat 17-HSD type 1 equally uses both estrone and androstenedione as substrates. Analyzing the activity of various rat/human chimeric enzymes indicated that the region between amino acids 148 and 268 is responsible for the difference in substrate specificity, which is in line with the structural data showing that the recognition end of the active site is primarily at residues 185-230. The enzymes are highly conserved between amino acids 148-191, and the data indicate that in this region Asn152HisAsp153Glu and Pro187Ala variations are most closely related to the differential steroid specificity. The structural analyses furthermore suggested that the presence of His instead of Asn at position 152 of the human enzyme might result in considerable rearrangement of the loop located close to the beta-face of the A- and B-rings of the bound substrate, and that the Pro187Ala variation could modify the flexible region involved in substrate recognition and access of the substrate to the active site. Altogether, our results indicate that the Asn152His and Pro187Ala variations, together with several amino acid variations at the recognition end of the catalytic cleft built by residues 190-230, alter the structure of the active site of rat 17-HSD type 1 to one more favorable to an androgenic substrate.