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Biomedical subjects

A K Chakraborty

Publications and source records attributed to A K Chakraborty.

At least 19 recordsLinked to original sources
PubMed1999

Biochemical effects of some pesticides on lipid peroxidation and free-radical scavengers.

Oxidative stress was studied in blood samples obtained from lindane, malathion and propoxur poisoning cases admitted to the Guru Teg Bahadur Hospital, Delhi and evaluated for lipid peroxidation, oxygen free radical (OFR) scavenging enzymes, and glutathione (GSH) and related enzymes. Acetylcholine esterase (AChE), gamma glutamyl transpeptidase (GGT) and GSH level were also assayed in lymphocytes. The level of thiobarbituric acid reacting substances and activities of superoxide dismutase, catalase, glutathione peroxidase, glutathione-S-transferase and GGT were increased and GSH level was decreased in pesticide poisoning. Apparently lindane (at the concentration examined) was more potent than malathion and propoxur in producing alteration in lipid peroxidation, GSH related parameters and OFR scavenging enzymes. However, AChE activity and GSH level in lymphocytes of malathion poisoning cases were reduced and GGT activity was enhanced in comparison to control subjects. The present results suggest that OFR scavenging enzymes were induced while combating oxidative stress in a differential manner in organochlorine, organophosphate and carbamate poisoning. Increased lipid peroxidation, coupled with altered levels of GSH and OFR scavenging enzymes in the blood are discussed in the light of oxidative stress.

Catalase↗
PubMed1999

The depigmenting effect of alpha-tocopheryl ferulate on human melanoma cells.

Oral vitamin E (alpha-tocopherol, alpha-T) supplementation has been reported to improve facial hyperpigmentation. alpha-Tocopheryl ferulate (alpha-TF) is a compound of alpha-T and ferulic acid connected by an ester bond; ferulic acid is also an antioxidant, and could scavenge free radicals induced by ultraviolet (UV) radiation, and thus maintain the long-lasting antioxidative effect of alpha-T. Our aim was to see whether alpha-TF might be useful as a whitening agent and an antioxidant to improve and prevent facial hyperpigmentation following UV exposure. In this study, the inhibitory effect of alpha-TF on melanogenesis was examined biochemically using human melanoma cells in culture. The results show that alpha-TF, solubilized in ethanol or in 0.5% lecithin, inhibited melanization significantly, as did alpha-T at a concentration of 100 microg/mL, without inhibiting cell growth. This phenotypic change was associated with inhibition of tyrosinase and 5, 6-dihydroxyindole-2-carboxylic acid polymerase activities, and the degree of inhibition was dose dependent. No significant effect on DOPAchrome tautomerase activity was observed. alpha-TF did not directly inhibit tyrosinase activity of the large granule fraction extracted from human melanoma cells, and Western blotting revealed that there were no changes in protein content or in molecular size of tyrosinase, tyrosinase-related protein (TRP)-1 or TRP-2. Therefore, the inhibition of tyrosinase activity by alpha-TF might be due to effects at the post-translational level, and possibly by a secondary molecule activated by alpha-TF. These results suggest that alpha-TF is a candidate for an efficient whitening agent which suppresses melanogenesis and inhibits biological reactions induced by reactive oxygen species.

Antioxidants↗
PubMed1999

Enhanced expression of melanocortin-1 receptor (MC1-R) in normal human keratinocytes during differentiation: evidence for increased expression of POMC peptides near suprabasal layer of epidermis.

Immunohistochemical staining of human skin specimen showed the stronger localization of proopiomelanocortin peptides near the suprabasal layer of the epidermis, where keratinocytes are mostly differentiated. To test the possibilities of whether the production of proopiomelanocortin peptides or their receptor-binding activity or both is increased during differentiation of keratinocytes, we treated the cells in culture with Ca2+ to induce their differentiation. The production of proopiomelanocortin peptides and its gene expression were not induced significantly, but the binding ability of melanocortin receptor, as well as its gene expression were stimulated by Ca2+. Ultraviolet B irradiation, an inducer of differentiation, stimulated both proopiomelanocortin production and melanocortin receptor expression. These data show that normal human keratinocytes express melanocortin receptor similar to melanocytes, and that it is induced during differentiation.

Adrenocorticotropic Hormone↗
PubMed1998

Role of far upstream repressor elements controlling proto-Ha-ras gene transcription.

The far upstream region of the rat Ha-ras gene has been characterized to determine whether possible repressor sequences may control the low level of Ha-ras gene transcription from its TATA-less, GC-rich strong promoter. The chloramphenicol acetyl transferase (CAT) gene under the control of the 3.8-kb Ha-ras upstream promoter was minimally expressed in HeLa cells. Surprisingly, CAT gene expression was increased by the deletion of a 0.7-kb BglII fragment containing non-coding exon minus 2 and TATA box promoter elements located 1.7 kb upstream of the GC-rich strong promoter. Far upstream (CA)25 repeats also appeared to repress Ha-ras gene activity. Sequences within the 0.7-kb BglII fragment suppressed CAT gene expression when placed upstream of a heterologous thymidine kinase (tk) gene promoter. Repressor activity was further localized to a 160-bp AvrII-BglII sub-fragment. Gel shift assays identified two sequence-specific DNA binding proteins. The results demonstrated for the first time that far upstream repressor sequences control normal transcription of the Ha-ras proto-oncogene.

Animals↗
PubMed1998

Immunoreactivity of alpha-melanocyte-stimulating hormone, adrenocorticotrophic hormone and beta-endorphin in cutaneous malignant melanoma and benign melanocytic naevi.

Melanocyte-stimulating hormone (MSH) has been reported to enhance the experimental metastatic behaviour of melanoma cells in the mouse model. alpha-MSH production and MSH receptor (melanocortin 1 receptor gene) expression have been detected in cultured normal human melanocytes and metastasized melanomas. The exact role of MSH in the metastatic behaviour of human melanoma cells is, however, not yet known. To clarify a possible role of proopiomelanocortin (POMC)-derived peptides, including alpha-MSH, in melanoma development and progression, we analysed immunohistochemically the localization of alpha-MSH adrenocorticotrophic hormone (ACTH) and beta-endorphin in various kinds of benign pigmented naevocytic lesions and malignant melanomas. Three of 21 samples of common and dysplastic naevi showed detectable alpha-MSH staining in naevus cells, and five and six of 15 samples were weakly positive for ACTH and beta-endorphin staining, respectively. In melanoma samples, 24 of 45, 23 of 39 and 30 of 42 samples showed positive staining with alpha-MSH, ACTH and beta-endorphin antibodies, respectively. Furthermore, staining for all three antibodies was noted to be more intense and diffuse in samples of nodular melanoma, vertically growing acral lentiginous melanoma and superficial spreading melanoma as well as metastatic lesions compared with those of naevi. Although it is yet to be determined whether or not this strong staining for POMC-derived peptides in advanced melanoma cells indicates a role of autocrine or paracrine regulation, our results suggest a possible involvement of POMC gene products in melanoma progression.

Adrenocorticotropic Hormone↗
PubMed1998

Modulation of melanocyte-stimulating hormone receptor expression on normal human melanocytes: evidence for a regulatory role of ultraviolet B, interleukin-1alpha, interleukin-1beta, endothelin-1 and tumour necrosis factor-alpha.

Melanocyte-stimulating hormone (MSH) receptor binding activity and melanocortin-1 receptor (MC1-R) gene expression on normal human melanocytes have been studied as responses to the effects of ultraviolet B (UVB), interleukin-1 (IL-1), endothelin-1 (ET-1) and tumour necrosis factor-alpha (TNF-alpha), which are known as UV sensitive regulators of melanocytic function. MSH receptor (MSH-R) binding activity was upregulated by UVB, IL-1alpha, -1beta and ET-1, but was downregulated by TNF-alpha. Northern blot analysis showed that MC1-R mRNA expression was induced 24 h after UVB irradiation in a dose-dependent manner, and that 24-h treatment with ET-1 also induced an expression of MC1-R mRNA, whereas TNF-alpha downregulated the expression. In addition, IL-1alpha and -1beta have a small but real inductive effect on MC1-R mRNA expression. Taken together, our results suggest a model in which higher MC1-R mRNA expression is accompanied by upregulation of MSH-R binding activity, and enhanced by UVB or cytokines sensitive to UVB. Such a regulatory system would enable normal human melanocytes to respond to MSH more efficiently and induce an increase of melanization of the skin through the MSH/MSH-R system after UVB radiation.

Adolescent↗
PubMed1998

Effect of arbutin on melanogenic proteins in human melanocytes.

The inhibitory effect of arbutin, a naturally occurring beta-D-glucopyranoside derivative of hydroquinone, on melanogenesis was studied biochemically by using human melanocytes in culture. Cells were cultured in the presence of different concentrations of arbutin. The maximum concentration of arbutin that was not inhibitory to growth of the cells was 100 micrograms/ml. At that concentration, melanin synthesis was inhibited significantly by approximately 20% after 5 days, compared with untreated cells. This phenotypic change was associated with the inhibition of tyrosinase and DHICA polymerase activities, and the degree of inhibition was dose dependent. No significant difference in DOPAchrome tautomerase (DT) activity was observed before or after arbutin treatment. Western blotting experiments revealed there were no changes in protein content or in molecular size of tyrosinase, TRP-1 or TRP-2, indicating that inhibition of tyrosinase activity by arbutin might be due to effects at the post-translational level.

Adolescent↗
PubMed1998

Melanoma x macrophage fusion hybrids acquire increased melanogenesis and metastatic potential: altered N-glycosylation as an underlying mechanism.

We recently reported that a majority of hybrids generated in vitro between weakly metastatic mouse Cloudman S91 melanoma cells and human or mouse macrophages showed enhanced metastatic potential. With few exceptions, hybrids with enhanced metastatic potential also had elevated basal melanin content and increased responsiveness to MSH compared to parental cells. Here we investigated the hybrid melanotic phenotype in more detail, comparing the pigmentary systems of hybrids and parental Cloudman S91 cells by several techniques. Cells were studied by electron microscopy, cell lysates were analyzed for tyrosinase (E.C.1.14.18.1) activity, and melanosomal proteins were analyzed by gel electrophoresis and immunoblotting. Melanosomes in parental Cloudman melanoma cells were few in number and relatively amorphous, whereas those in the hybrids were numerous and heavily pigmented, containing highly organized lattice structures. Both basal and MSH-inducible tyrosinase activities were elevated several fold in hybrids compared to parental cells. Tyrosinase, TRP-2, and LAMP-1 from hybrids migrated more slowly on gels compared to the same proteins from parental melanoma cells, consistent with increased glycosylation. Migration of LAMP-1 from hybrids was similar to that from peritoneal macrophages, which also appeared to be more heavily glycosylated than LAMP-1 from Cloudman cells. By using 3H-glucosamine as a marker of N-glycosylation, its incorporation into tyrosinase and LAMP-1 was found to be elevated in hybrids, suppressed by N-glycosylation inhibitors, and stimulated by MSH to a greater degree in hybrids compared to parental cells. These results indicate N-glycosylation as an important regulatory pathway for MSH-induced melanogenesis and further suggest that altered N-linked glycosylation may be an underlying mechanism for regulation of both melanogenesis and metastasis in macrophage x melanoma hybrids.

Animals↗
PubMed1998

Spectral studies on the interaction of iodine and anthracene sulfonate with bacterial phospholipids from different mutants of Salmonella minnesota.

Using spectrophotometric and spectrofluorometric techniques, the interaction of iodine and 2-anthracene sulfonate (ANS) with the phospholipids (PL) isolated from four genetically correlated Salmonella minnesota isolates viz., a smooth form (S), a deeply rough mutant (Rc) and two intermediate forms (Ra and Rb) were studied. Appearance of an isosbestic point and a new band in absorption spectra indicated charge-transfer (C-T) interaction of iodine with the PL through the formation of 1:1 complex. Stern-Volmer type fluorescence quenching of PL was observed with the addition of iodine to PL, while PL enhanced the fluorescence of anionic dye ANS. The values of the binding constants between iodine/PL and ANS/PL, measured by using suitable equations, showed a systematic gradation in the molecular properties of the PL in the membrane structure in smooth (S) and rough (Ra, Rb and Rc) mutants of Salmonella minnesota.

Anthracenes↗
PubMed1998

Spectrophotometric and spectrofluorometric studies on interaction of cationic dyes with bacterial capsular polysaccharide.

Interaction of Klebsiella K14 capsular polysaccharide with cationic dyes pinacyanol chloride, acridine orange and phenosafranin has been studied by spectrophotometric and spectrofluorometric techniques. The polymer containing both glucuronic acid and pyruvic acid in its repeating unit behaved as a unique polyelectrolyte. It induced blue shift of the absorption band of pinacyanol chloride indicating strong metachromasy. Stoichiometry of the polyanion and the dye cations in the polymer-dye compound (1:2) indicated that both glucuronic acid and pyruvic acid acted as potential anionic sites for interaction with the cationic dye molecules. The stoichiometry of anionic site (of polyanion): cationic site (of dye) in the polymer dye compound was calculated as 1:1. Interaction of the polymer with acridine orange and phenosafranin dyes studied by fluorescence measurements demonstrated Stern-Volmer type of quenching. Equivalent weight of the polymer was determined by spectrophotometric and spectrofluorometric titrations. From the present studies chromotropic property of the polymer was established.

Acridine Orange↗
PubMed1997

Chromatographic separation of DNA dependent RNA polymerases and molecular properties of RNA polymerase II from a Leishmania Spp.

Multiple forms of DNA-dependent RNA polymerases have been isolated and characterized from Leishmania strain UR6 promastigotes. RNA polymerases from this organism fail to resolve into multiple forms by conventional chromatography on DEAE-Sephadex A25, but could be separated by a modification of the method using CM-Sephadex C25. The CM-Sephadex bound enzyme is resistant to alpha-amanitin even up to a concentration of 250 micrograms/ml. The activity which flows through CM-Sephadex further resolves into two forms upon chromatography on DEAE-Sephadex A25. These forms are sensitive to alpha-amanitin to different extent. Enzyme activity in peak I is 50% inhibited by 3 micrograms/ml and in peak II by 50 micrograms/ml of the drug respectively. The enzyme in peak I has been further purified by heparin agarose and fast performance liquid chromatography (FPLC) on MonoQ. The enzyme has Stoke's radius of 70 A, a sedimentation coefficient of 17.6S and an f/fo of 1.35. Analysis of ammonium sulfate and metal ion optima of the enzyme in peak I, relative activities with Mn+2 versus Mg+2 and template specificities gave results similar to those reported for other type II RNA polymerases in eukaryotes. The MonoQ purified enzyme resolves into 16 polypeptides on denaturing polyacrylamide gel and densitometric analysis suggests that 9 major bands are present in the stoichiometry expected of RNA polymerase subunits having molecular weights: 154000; 104000; 77000; 64000; 52000; 48000; 46000; 45000 and 39000 respectively.

Animals↗
PubMed1997

Effect of short-term dermal toxicity of fenvalerate on residue, cell architecture and biochemical profiles in broiler chicks.

Effect of fenvalerate on cell architecture, tissue biochemical parameters and its residual concentration was studied in broiler chicks following dermal application at 0.1 and 1% in ethanol once daily for 31 days. It did neither produce loss of body weight nor clinical signs of toxicity. Kidney contained maximal residue followed by heart, fat, liver and brain after 0.1%; and fat contained maximal residue followed by kidney, heart, liver and brain after 1% application. Fenvalerate (0.1%) increased the aspartate aminotransferase (AST) (except brain), alanine aminotransferase (ALT), alkaline phosphatase (AP), acid phosphatase (AcP) (only brain) activities, glycogen level (only liver) in liver, kidney, heart and brain tissues; and 1% increased the AST (except brain), ALT, AcP (except liver and kidney), AP (only heart), glycogen (only liver) and decreased AP (except heart), AcP (only kidney), cholesterol (except liver and heart), and acetylcholinesterase (AChE) (liver and brain) of liver, kidney, heart and brain tissue homogenates respectively. Histopathological examination in general showed aggregation of mononuclear cells in liver, around the kidney tubules and cardiac muscle fibre. In addition, fibrosis in the periportal area of liver, proliferation of ureter and tubular degeneration, and congestion of endocardial vessels were also observed. The intensity of cellular changes was more marked after 1% dermal application.

Animals↗
PubMed1996

Production and release of proopiomelanocortin (POMC) derived peptides by human melanocytes and keratinocytes in culture: regulation by ultraviolet B.

It is demonstrated that ultraviolet B (UVB) radiation stimulates increased expression of the proopiomelanocortin (POMC) gene which is accompanied by production and release of alpha-melanocyte stimulating hormone (alpha-MSH) and adrenocorticotropin (ACTH) by both normal and malignant human melanocytes and keratinocytes. The production and release of both peptides are also stimulated by dibutyryl cyclic adenosine monophosphate (dbcAMP) and interleukin 1 alpha (IL-1 alpha) but not by endothelin-1 (ET-1) or tumor necrosis factor-alpha (TNF-alpha). N-acetyl-cysteine (NAC), a precursor of glutathione (GSH), an intracellular free radical scavenger, abolishes the UVB-stimulated POMC peptide production and secretion. Conclusions are as follows: (1) Cultured human cells of cutaneous origin, namely keratinocytes and melanocytes, can produce and express POMC; (2) POMC expression is enhanced by exposure to UVB, possibly through a cyclic AMP-dependent pathway; and (3) The action of UVB on POMC production may involve a cellular response to oxidative stress.

Acetylcysteine↗
PubMed1996

Polymerization of 5,6-dihydroxyindole-2-carboxylic acid to melanin by the pmel 17/silver locus protein.

Recent advances in melanogenesis have focused on the role of dihydroxyindole-2-carboxylic acid[(HO)2IndCOOH]. For example, it has been shown that formation of (HO)2IndCOOH from dopachrome is catalyzed by dopachrome tautomerase, that the melanogenic protein tyrosinase-related protein (TRP)-1 can oxidize (HO)2IndCOOH to its indole quinone, that (HO)2IndCOOH-melanins can be synthesized chemically, that mammalian melanins are naturally rich in (HO)2IndCOOH subunits, and that (HO)2IndCOOH is incorporated into melanins of melanomas in mice. The question thus emerges as to the mechanism(s) by which (HO)2IndCOOH and other precursors become incorporated into melanins in vivo. Accordingly, an activity was partially purified that catalyzed melanin formation with (HO)2IndCOOH as a substrate. Analyses of the (HO)2IndCOOH polymerization factor from Cloudman melanoma cells revealed the following: it was proteinaceous in that it was heat labile and destroyed by proteinase K; it was a glycoprotein in that it adhered to wheat germ agglutinin and was eluted with N-acetyl glucosamine; it was located predominantly in the melanosomal fraction of cell homogenates; the activity was reduced by exposure to the metal chelators EDTA and EGTA, but not by phenylthiourea, a tyrosinase inhibitor; the (HO)2IndCOOH polymerization reaction was inhibited by superoxide dismutase. In addition, the activity was found with the mouse pmel 17/silver locus protein immunopurified from human melanoma cells, and was significantly reduced in extracts of mouse melanocytes cultured from silver (si/si) mice compared to extracts from Si/Si melanocytes. In summary, an activity has been identified in human and mouse melanoma cells that catalyzes the superoxide-dependent polymerization of (HO)2IndCOOH to melanin in vitro, and appears to be a function of the pmel 17/silver protein of the human pmel 17 gene and the mouse silver locus.

Animals↗
PubMed1996

The disposition kinetics and residues of fenvalerate in tissues following a single dermal application to black Bengal goats.

The disposition kinetics of fenvalerate were studied in goats after dermal application of 100 ml of 0.25% (w/v) solution. The insecticide persisted in the blood for 72 h. The mean (+/- SEM) Vd(area) and apparent t 1/2 (beta) were 9.92 +/- 1.44 L/kg and 17.51 +/- 2.65 h, while the AUC and ClB values were respectively 82.15 +/- 7.40 micrograms h/ml and 0.56 +/- 0.05 L/(kg h). Four days after the dermal application, the highest concentration of fenvalerate residues was found in the adrenal gland, followed by the biceps muscle, omental fat, liver, kidney, lung and cerebrum in that order. Fenvalerate caused hyperglycaemia but had no effect on serum protein and cholesterol levels. Serum acetylcholinesterase activities were increased after 24 h but were below the initial values from 48 to 120 h.

Acetylcholinesterase↗
PubMed1996

Vitiligo, psoralen, and melanogenesis: some observations and understanding.

Since the etiology of vitiligo is still unknown, we searched for some abnormal biochemical parameters, if any, in subjects with vitiligo. Higher urinary excretion of indole metabolites in vitiliginous patients have been noted, in association with higher dioxygenase, superoxide dismutase, and tyrosine aminotransferase activity in their serum. Similar results have also been found in an animal model, Bufo melanostictus, during induced tyrosinase inhibition. Treatment with psoralen can reverse the parameters, except tyrosine aminotransferase, to a normal level. Although psoralens are not the magic bullet for the therapy of vitiligo, they are still being used as a chemotherapeutic agent against vitiligo on a major scale to date. Tryptophan was found to participate in the pathway of melanogenesis, as a precursor as well as a positive regulator of tyrosinase. Its behavior in this regard is much more similar to the conventional substrates tyrosine and dopa (dihydroxyphenylalanine). In consideration of combined participation of tyrosine and tryptophan in the synthesis of melanin and its breakdown, the possible influence of different enzymatic reactions, like mono-oxygenase, dioxygenase, and deamination, has been suggested.

Animals↗