Biomedical subjects

J Debnath

Publications and source records attributed to J Debnath.

14 recordsLinked to original sources

PubMed2002

Testicular gametogenic and steroidogenic activities in cyclophosphamide treated rat: a correlative study with testicular oxidative stress.

The present work examined the changes in testicular activities in relation to testicular oxidative stress in cyclophosphamide as well as human chorionic gonadotrophin (hCG) co-treated cyclophosphamide treated Wistar strain rats. Testicular activities were evaluated by the quantification of spermatogenesis and by the measurement of steroidogenic key enzyme activities along with plasma levels of testosterone. Testicular oxidative stress in relation to cyclophosphamide treatment was monitored by the study of products of free radicals like conjugated dienes and malondialdehyde (MDA) as well as the activity of testicular antioxidant enzymes like peroxidase and catalase. Cyclophosphamide treatment at the dose of 5 mg/kg body weight/day for 28 days resulted a significant diminution in the activities of testicular delta 5, 3 beta-hydroxysteroid dehydrogenase (delta 5, 3 beta-HSD), 17 beta-hydroxysteroid dehydrogenase (17 beta-HSD) activities, plasma level of testosterone along with significant reduction in the number of germ cells at stage-VII of spermatogenesis. Levels of testicular MDA and conjugated dienes both were elevated whereas testicular peroxidase and catalase activities both were inhibited significantly in cyclophosphamide treated rats in comparison to control. After hCG co-administration at the dose of 5 I.U./kg body weight/day for 28 days in cyclophosphamide treated rats resulted a significant protection in the activities of testicular peroxidase and catalase along with significant decrease in the levels of MDA and conjugated dienes to the control level. Moreover, the testicular steroidogenic key enzyme activities and spermatogenesis along with plasma levels of testosterone were restored to the control level. Therefore, it may be concluded that there is a correlation between testicular steroidogenic activities as well as spermatogenesis and testicular oxidative stress in cyclophosphamide treated rats. Moreover, as restoration of plasma testosterone to the control level is noted in hCG co-treated cyclophosphamide treated rat, therefore, the results suggest that testosterone may be the key regulator for this correlation.

Animals↗

PubMed2001

Requirements for activation and RAFT localization of the T-lymphocyte kinase Rlk/Txk.

BACKGROUND: The Tec family kinases are implicated in signaling from lymphocyte antigen receptors and are activated following phosphorylation by Src kinases. For most Tec kinases, this activation requires an interaction between their pleckstrin homology (PH) domains and the products of phosphoinositide 3-Kinase, which localizes Tec kinases to membrane RAFTs. Rlk/Txk is a Tec related kinase expressed in T cells that lacks a pleckstrin homology domain, having instead a palmitoylated cysteine-string motif. To evaluate Rlk's function in T cell receptor signaling cascades, we examined the requirements for Rlk localization and activation by Src family kinases. RESULTS: We demonstrate that Rlk is also associated with RAFTs, despite its lack of a pleckstrin homology domain. Rlk RAFT association requires the cysteine-string motif and is independent of PI3 Kinase activity. We further demonstrate that Rlk can be phosphorylated and activated by Src kinases, leading to a decrease in its half-life. A specific tyrosine in the activation loop of Rlk, Y420, is required for phosphorylation and activation, as well as for decreased stability, but is not required for lipid RAFT association. Mutation of this tyrosine also prevents increased tyrosine phosphorylation of Rlk after stimulation of the T cell receptor, suggesting that Rlk is phosphorylated by Src family kinases in response to T cell receptor engagement. CONCLUSIONS: Like the other related Tec kinases, Rlk is associated with lipid RAFTs and can be phosphorylated and activated by Src family kinases, supporting a role for Rlk in signaling downstream of Src kinases in T cell activation.

Cell Line↗

PubMed2001

Protection of sodium arsenite-induced ovarian toxicity by coadministration of L-ascorbate (vitamin C) in mature wistar strain rat.

Arsenic, a major water pollutant in India, produces toxic effects on female reproductive system in rodent models at the dose available in drinking water in arsenic-intoxicated zones. This study examines the coadministration of L-ascorbate (vitamin C) on ovarian steroidogenesis, plasma levels of gonadotrophins, brain monoamines, and ovarian as well as uterine peroxidase activities in sodium arsenite-treated rats. After sodium arsenite treatment, relative ovarian and uterine weights, ovarian Delta5-3beta-HSD and 17beta-HSD activities, plasma levels of gonadotrophins, norepinephrine levels in midbrain and diencephalon, and the activities of peroxidase in ovary and uterus were decreased significantly. On the other hand, serotonin levels in midbrain and diencephalon were increased significantly 28 days after sodium arsenite treatment at the dose of 0.4 ppm/100 g body weight/rat/day. All these parameters were protected significantly and in most cases were unchanged from control level when L-ascorbate at 25 mg/100 g body weight/rat/day was coadministered orally with sodium arsenite. This cotreatment of L-ascorbate with sodium arsenite also restored the estrous cycle in a regular manner. We concluded that L-ascorbate plays a pivotal role in maintaining normal ovarian activities and brain monoamines in arsenic-treated rats.

Animals↗

PubMed2001

Biochemical and genetic analyses of the Tec kinases Itk and Rlk/Txk.

The Tec kinases have been implicated as important components of signalling pathways downstream of lymphocyte antigen receptors. Activation of these kinases requires two steps: (i) phosphorylation by Src family kinases and (ii) plasma membrane localization, which is mediated by interaction between the pleckstrin homology (PH) domains of Tec kinases and the products of phosphoinositide-3 kinase (PI-3K). Itk and Rlk/Txk are Tec kinases expressed in T-lymphocytes. Despite similarity to other Tec kinases, Rlk/Txk lacks a PH domain and instead possesses a palmitoylated cysteine-string motif. We have found that both Rlk/Txk and Itk are phosphorylated in response to T-cell receptor stimulation and can be activated by phosphorylation by Src family kinases. However, consistent with its lack of PH domain, Rlk/Txk is phosphorylated independent of PI-3K activity. Furthermore, we demonstrated that like Itk, Rlk/Txk is associated with lipid RAFTs (detergent-insoluble, cholesterol-rich regions of the membrane), but unlike Itk, Rlk/Txk's RAFT association is independent of PI-3K activity. Despite these differences, Rlk/Txk partially compensates for loss of Itk in gene-targeted animals, suggesting overlapping functions for these kinases.

Animals↗

PubMed2000

Tec family kinases modulate thresholds for thymocyte development and selection.

Tec family kinases are implicated in T cell receptor (TCR) signaling, and combined mutation of inducible T cell kinase (Itk) and resting lymphocyte kinase (Rlk)/Txk in mice dramatically impairs mature T cell function. Nonetheless, mutation of these kinases still permits T cell development. While itk(-)(/)- mice exhibit mild reductions in T cells with decreased CD4/CD8 cell ratios, rlk(-)(/)-itk(-)(/)- mice have improved total T cell numbers yet maintain decreased CD4/CD8 ratios. Using TCR transgenics and an in vitro thymocyte deletion model, we demonstrate that mutation of Tec kinases causes graded defects in thymocyte selection, leading to a switch from negative to positive selection in rlk(-)(/)-itk(-)(/)- animals. The reduction in both positive and negative selection and decreased CD4/CD8 ratios correlates with decreased biochemical parameters of TCR signaling, specifically defects in capacitive Ca(2+) influx and activation of the mitogen-activated kinases extracellular signal-regulated kinase 1 and 2. Thus, Tec kinases influence cell fate determination by modulating TCR signaling, leading to altered thresholds for thymocyte selection. These results provide support for a quantitative model for thymic development and provide evidence that defects in negative selection can substantially alter thymic cellularity.

Animals↗

PubMed1999

Requirement for Tec kinases Rlk and Itk in T cell receptor signaling and immunity.

T cell receptor (TCR) signaling requires activation of Zap-70 and Src family tyrosine kinases, but requirements for other tyrosine kinases are less clear. Combined deletion in mice of two Tec kinases, Rlk and Itk, caused marked defects in TCR responses including proliferation, cytokine production, and apoptosis in vitro and adaptive immune responses to Toxoplasma gondii in vivo. Molecular events immediately downstream from the TCR were intact in rlk-/-itk-/- cells, but intermediate events including inositol trisphosphate production, calcium mobilization, and mitogen-activated protein kinase activation were impaired, establishing Tec kinases as critical regulators of TCR signaling required for phospholipase C-gamma activation.

Animals↗

PubMed1999

rlk/TXK encodes two forms of a novel cysteine string tyrosine kinase activated by Src family kinases.

Rlk/Txk is a member of the BTK/Tec family of tyrosine kinases and is primarily expressed in T lymphocytes. Unlike other members of this kinase family, Rlk lacks a pleckstrin homology (PH) domain near the amino terminus and instead contains a distinctive cysteine string motif. We demonstrate here that Rlk protein consists of two isoforms that arise by alternative initiation of translation from the same cDNA. The shorter, internally initiated protein species lacks the cysteine string motif and is located in the nucleus when expressed in the absence of the larger form. In contrast, the larger form is cytoplasmic. We show that the larger form is palmitoylated and that mutation of its cysteine string motif both abolishes palmitoylation and allows the protein to migrate to the nucleus. The cysteine string, therefore, is a critical determinant of both fatty acid modification and protein localization for the larger isoform of Rlk, suggesting that Rlk regulation is distinct from the other Btk family kinases. We further show that Rlk is phosphorylated and changes localization in response to T-cell-receptor (TCR) activation and, like the other Btk family kinases, can be phosphorylated and activated by Src family kinases. However, unlike the other Btk family members, Rlk is activated independently of the activity of phosphatidylinositol 3-kinase, consistent with its lack of a PH domain. Thus, Rlk has two distinct isoforms, each of which may have unique properties in signaling downstream from the TCR.

Animals↗

PubMed1999

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.

Acid Phosphatase↗

PubMed1999

Effect of sodium arsenite on plasma levels of gonadotrophins and ovarian steroidogenesis in mature albino rats: duration-dependent response.

Effect of arsenic on ovarian steroidogenesis at the dose available in drinking water at wide areas of West Bengal is reported here. Weights of ovary, uterus and vagina along with biochemical activities of ovarian delta 5-3 beta-hydroxysteroid dehydrogenase (delta 5-3 beta-HSD) and 17 beta-hydroxysteroid dehydrogenase (17 beta-HSD) and plasma levels of LH, FSH and estrogen were measured in mature rats of the Wistar strain at diestrous phase following subchronic treatment with sodium arsenite at a dose of 0.4 ppm/rat/day for 16 days (4 estrous cycles) and 28 days (7 estrous cycles). A significant reduction in plasma levels of LH, FSH and estrogen along with significant diminution in the activities of ovarian delta 5-3 beta-HSD and 17 beta-HSD were observed following sodium arsenite treatment for 28 days. This duration of treatment also resulted in a marked degree in diminution in the weights of ovary, uterus and vagina, but 16 days of treatment did not exhibit any significant effect on these above parameters. Arsenic-treated rats exhibited a prolonged diestrous phase in the estrous cycle in contrast to control rats having 4 days of a regular estrous cycle. Deposition of arsenic in ovary, uterus, vagina and plasma was also monitored in arsenic-treated rats. The results of our experiment suggest that duration of arsenic treatment is the critical factor for its adverse effect on ovarian activities at the dose within the range noted in drinking water at several areas of West Bengal in India.

Animals↗

PubMed1995

Introduction of a loss-of-function point mutation from the SH3 region of the Caenorhabditis elegans sem-5 gene activates the transforming ability of c-abl in vivo and abolishes binding of proline-rich ligands in vitro.

We have introduced two loss-of-function point mutations from highly conserved regions of the src homology 3 (SH3) domains of the Caenorhabditis elegans sem-5 gene into the SH3 domain of the murine type IV c-abl tyrosine kinase proto-oncogene. One of the mutations, P131L, activated abl to transform fibroblasts while the other, G128R, did not. When combined with independent activating mutations in the c-abl kinase domain or NH2-terminus, the G128R mutation blocked transformation by the double mutant, suggesting that the G128R mutant was unable to transform cells for trivial reasons. The c-Abl G128R mutant, like wild type c-Abl protein, was localized to the nucleus and actin cytoskeleton and had normal tyrosine kinase activity in vitro, while the transforming c-Abl P131L protein was localized exclusively to the cytoplasm and exhibited decreased in vitro kinase activity. By real-time biospecific interaction analysis, the wild type Abl SH3 domain bound to two proteins containing proline-rich motifs with dissociation constants of 0.2 and 17 microM; the G128R mutant bound with 50-fold lower affinity, and no binding was detected by the P131L mutant. Both mutations completely abolished binding of the Abl SH3 domain to proline-rich target proteins in a filter-binding assay. These results suggest that the transforming activity of Abl is regulated in vivo by an inhibitor protein which associates with the SH3 domain via a proline-rich sequence.

3T3 Cells↗

PubMed1994

Effect of lithium chloride on testicular delta 5-3 beta, 17 beta-hydroxy steroid dehydrogenase, acid phosphatase and gametogenesis in Bufo melanostictus.

Injection of lithium chloride at the dose of 200 micrograms/toad/alternate day for 7, 14 and 21 days caused a significant reduction in the activities of testicular delta 5-3 beta, 17 beta-hydroxysteroid dehydrogenase and acid phosphatase enzymes. There was a marked inhibition in spermatogenesis in lithium chloride treated toad for 14 and 21 days of treatment, but 7 days of treatment has no effect.

17-Hydroxysteroid Dehydrogenases↗

PubMed1993

HPLC-based method for determination of absolute configuration of alpha-chiral amines.

We introduce a novel, HPLC-based method for facile determination of the absolute configuration of alpha-chiral amines. Our method is easily applied to a variety of compounds, including amino acid derivatives. The method involves initial derivatization of the chiral amine analyte with the chiral derivatizing reagent, N-succinimidyl alpha-methoxyphenylacetate (SMPA), to produce the corresponding diastereomeric adducts. Inspection of a particular rotomer of the SMPA adduct and application of simple rules correlates absolute configuration and HPLC elution order. A key aspect of our method is that it can be used to determine absolute configuration without using enantiomeric standards of the amine analytes. Furthermore, it is of utmost significance that our method can also be used to determine absolute configuration even when only one analyte enantiomer of unknown absolute configuration is present, as is often the case for enzymatic products, naturally derived compounds, or enantiomerically enriched compounds prepared via chiral syntheses. We have observed strict adherence between predicted and observed absolute configuration for a wide variety of alpha-chiral amines. The chromatographic method we present in this paper is very practical and has several important advantages over NMR-based approaches which have been previously developed. For example, microgram quantities of an analyte in a complex enzymatic mixture can be directly analyzed by our HPLC-based method while the impurities often preclude definitive proton assignments in the NMR approach.

Amines↗

PubMed1992

Activation of an adrenergic pro-drug through sequential stereoselective action of tandem target enzymes.

The synthetic amino acid, 3,4-dihydroxyphenylserine (DOPS) has been of great interest for many years as an adrenergic pro-drug, since the L-threo diastereomer of DOPS can be a precursor of R-(-)-norepinephrine, the natural form of this neurotransmitter. We now report bioactivation of DOPS to the potent pharmacological agent, noradrenalone (arterenone), via sequential stereoselective action by two target enzymes--dopamine beta-monooxygenase (DBM) and L-aromatic amino acid decarboxylase (AADC)--acting in tandem. Enzymatic activation is stereospecific, with only the L-erythro DOPS diastereomer producing noradrenalone; this is consistent with the known stereospecificities of AADC and DBM. These results provide a heretofore unrecognized rationale for the bioactivity of L-erythro DOPS and provide a basis for the design of new adrenergic pro-drugs.

Adrenal Glands↗

PubMedDate not supplied

Effect of cold exposure on testicular delta 5-3 beta and 17 beta hydroxysteroid dehydrogenase activities and plasma levels of testosterone in toad (Bufo melanostictus) in breeding and hibernating season: duration-dependent response.

The purpose of this study was to show the effect of cold exposure on testicular activities in breeding and hibernating seasons in the toad. Adult male toads were placed in a cold chamber in both breeding and hibernating seasons for periods of 7, 14 and 21 days. At the time of sacrifice on the 15th and 22nd days, cold-exposed animals showed a decrease in testicular weight, testicular delta 5-3 beta and 17 beta-hydroxysteroid dehydrogenase activities and low levels of plasma testosterone both in breeding and hibernating seasons. There was no significant alteration in the above mentioned steroidogenic enzymes and plasma levels of testosterone after 7 days of cold exposure, both in breeding and hibernating seasons in respect to the control animals. The results of our present experiment suggest that environmental cold is an important modulator of breeding activities in the male toad. It also indicates that the breeding and hibernating cycle in the toad (seasonal breeders) may be asynchronous to each other.

17-Hydroxysteroid Dehydrogenases↗