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Biomedical subjects

S K Ghosh

Publications and source records attributed to S K Ghosh.

At least 55 records · Page 3Linked to original sources
PubMed1999

Cytotoxic effector T cells elicited by the killed tumor vaccine differ significantly from the effectors generated during active growth of a murine B-cell lymphoma.

Active specific immunotherapy of neoplastic diseases is an elusive goal. Using a murine B lymphoma 2C3, we showed that vaccination with the killed tumor cells effectively induces protective immunity and a cytotoxic T cell (CTL) response. Similar protection, however, is rarely observed in mice bearing live tumor cells. These animals usually succumb to the progressively growing tumor. In this study, we inquired whether the splenic CTL induced during tumor progression in mice differ from those evoked by the killed tumor cells. Here we demonstrate that the CTL generated following vaccination are significantly different from those induced in the tumor-bearing hosts. Adding to the complexity, the CTL from the early tumor bearers also differ significantly from those induced at the late stages. These differences are based on their cytotoxic activity, MHC allele specificity, mitogen responsiveness, cytokine secretion profile and T cell receptor Vbeta gene expression. The results clearly indicate that passive immunization with killed tumor is most effective, possibly because the CTL induced are not subject to the same regulatory pressure as those induced during active tumor growth. This decreasing effectiveness of CTL could be due to greater variability in antigenic stimulus, less involvement of innate immunity, changes in cytokine milieu and/or costimulatory factors.

Alleles↗
PubMed1999

Chiral recognition in dipeptides containing 1-aminocyclopropane carboxylic acid or alpha-aminoisobutyric acid: NMR studies in solution.

Protected dipeptides containing 1-aminocyclopropane carboxylic acid (Ac3c) or alpha-aminoisobutyric acid (Aib) residues at the C-terminus and Phe, Val or Ala residues at the N-terminus displayed different proton NMR spectra for the pure enantiomers and the racemic mixtures in deuterochloroform (CDCl3) solution. An unequal mixture of enantiomers showed two sets of resonances (NMR nonequivalence), one corresponding to major and the other to minor enantiomer. The NMR nonequivalence was originated by the presence of the C-terminal Ac3c or Aib residues, which have been known for their unique spatial preferences in avoiding an extended (C5) conformation. When a C5 conformation favoring residue such as glycine was incorporated in place of Ac3c or Aib, negligible NMR nonequivalence was observed. The magnitude of the NMR nonequivalence depended on the side chain as well as on the protecting groups at N-terminus alpha-amino acid. For the same peptide, the magnitude of nonequivalence increased with increasing solution concentration and/or with decreasing the solution temperature. The NMR nonequivalence disappeared in polar solvent-like deuterated dimethylsulfoxide (DMSO-d6). A preference for hetero-chiral recognition leading to dimeric association under fast exchange conditions had been invoked to explain the observed phenomenon. The dipeptides thus prepared could well serve as 'model peptides' for the evaluation of any preparative methods.

Amino Acids↗
PubMed1999

Acetylation of alpha-crystallin with N-acetylimidazole and its influence upon the native aggregate and subunit reassembly.

PURPOSE: An attempt has been made to investigate the involvement and importance of some of the hydrogen bond forming amino acid side chains in intra and inter subunit interactions in alpha-crystallin assembly. METHODS: For this, alpha-crystallin has been acetylated, partially or completely, using N-acetylimidazole. The apparent molecular size, electrophoretic mobility, conformational properties, surface hydrophobicity and chaperone activity of the modified proteins have been determined and compared with those of unmodified native protein as well as of the aggregates reassembled from the modified subunits. RESULTS: Acetylation of the surface-exposed tyrosine side chains has been found to destabilize the integrity of the native assembly with the formation of a somewhat smaller aggregate. This acetylated aggregate appears to adopt a molten globule-like conformation as evidenced from its almost unaltered secondary structure with some detectable alterations in its tertiary structure as well as from its enhanced chaper-one activity exhibited by the reduction assay compared to the native alpha-crystallin. Reassociation studies from either partially or completely acetylated subunits indicate that acetylation perturbs the information needed for native refolding of the subunits from their unfolded state as well as that needed for the normal mode of subunit reassembly. Acetylated subunits exhibit abnormal gel electrophoretic band pattern with distinctly retarded migration compared to the unmodified subunits. However, in spite of the partial/complete acetylation of the subunits or their reassociation from the denatured state, the tryptophan fluorescence emission maxima of the modified proteins and also that of the reassociated aggregates appear to remain unaffected. CONCLUSIONS: Results tend to indicate that the unperturbed hydrogen bonding capability of the relevant side chains in alpha-crystallin is needed for the integrity of the native alpha-crystallin assembly, for the normal refolding of its denatured subunits and also for the correct mode of subunit reassembly.

Acetylation↗
PubMed1999

Bayesian analysis and model selection for interval-censored survival data.

Interval-censored data occur in survival analysis when the survival time of each patient is only known to be within an interval and these censoring intervals differ from patient to patient. For such data, we present some Bayesian discretized semiparametric models, incorporating proportional and nonproportional hazards structures, along with associated statistical analyses and tools for model selection using sampling-based methods. The scope of these methodologies is illustrated through a reanalysis of a breast cancer data set (Finkelstein, 1986, Biometrics 42, 845-854) to test whether the effect of covariate on survival changes over time.

Algorithms↗
PubMed1999

Chitinase secretion by encysting Entamoeba invadens and transfected Entamoeba histolytica trophozoites: localization of secretory vesicles, endoplasmic reticulum, and Golgi apparatus.

Entamoeba histolytica, the protozoan parasite that phagocytoses bacteria and host cells, has a vesicle/vacuole-filled cytosol like that of macrophages. In contrast, the infectious cyst form has four nuclei and a chitin wall. Here, anti-chitinase antibodies identified hundreds of small secretory vesicles in encysting E. invadens parasites and in E. histolytica trophozoites overexpressing chitinase under an actin gene promoter. Abundant small secretory vesicles were also identified with antibodies to the surface antigen Ariel and with a fluorescent substrate of cysteine proteinases. Removal of an N-terminal signal sequence directed chitinase to the cytosol. Addition of a C-terminal KDEL peptide, identified on amebic BiP, retained chitinase in a putative endoplasmic reticulum, which was composed of a few vesicles of mixed sizes. A putative Golgi apparatus, which was Brefeldin A sensitive and composed of a few large, perinuclear vesicles, was identified with antibodies to ADP-ribosylating factor and to epsilon-COP. We conclude that the amebic secretory pathway is similar to those of other eukaryotic cells, even if its appearance is somewhat different.

ADP-Ribosylation Factors↗
PubMed1999

Feline leukemia virus long terminal repeat activates collagenase IV gene expression through AP-1.

Leukemia and lymphoma induced by feline leukemia viruses (FeLVs) are the commonest forms of illness in domestic cats. These viruses do not contain oncogenes, and the source of their pathogenic activity is not clearly understood. Mechanisms involving proto-oncogene activation subsequent to proviral integration and/or development of recombinant viruses with enhanced replication properties are thought to play an important role in their disease pathogenesis. In addition, the long terminal repeat (LTR) regions of these viruses have been shown to be important determinants for pathogenicity and tissue specificity, by virtue of their ability to interact with various transcription factors. Previously, we have shown that, in the case of Moloney murine leukemia virus, the U3 region of the LTR independently induces transcriptional activation of specific cellular genes through an LTR-generated RNA transcript (S. Y. Choi and D. V. Faller, J. Biol. Chem. 269:19691-19694, 1994; S.-Y. Choi and D. V. Faller, J. Virol. 69:7054-7060, 1995). In this report, we show that the U3 region of exogenous FeLV LTRs can induce transcription from collagenase IV (matrix metalloproteinase 9) and monocyte chemotactic protein 1 (MCP-1) promoters up to 12-fold. We also show that AP-1 DNA-binding activity and transcriptional activity are strongly induced in cells expressing FeLV LTRs and that LTR-specific RNA transcripts are generated in those cells. Activation of mitogen-activated protein kinase kinases 1 and 2 (MEK1 and -2) by the LTR is an intermediate step in the FeLV LTR-mediated induction of AP-1 activity. These findings thus suggest that the LTRs of FeLVs can independently activate transcription of specific cellular genes. This LTR-mediated cellular gene transactivation may play an important role in tumorigenesis or preleukemic states and may be a generalizable activity of leukemia-inducing retroviruses.

3T3 Cells↗
PubMed1998

Interaction of H(+)-ions with alpha-crystallin: solvent accessibility of ionizable side chains and surface charge.

Interaction of H(+)-ions with alpha-crystallin from goat lens has been studied at three different ionic strengths using the potentiometric titration method. Titrations have also been carried out in the presence of 1.5 M and 6 M GuHCl (guanidine hydrochloride). The isoionic pH of the protein in water and the effect of KCl on it have been determined. Titration curves have been found to be reversible between pH 3 to 9.25 at all ionic strengths. To aid in the data analyses, the reactivities of alpha-crystallin lysine residues to trinitrobenzenesulfonic acid have been determined in this work. For alpha-crystallin aggregate, 130 +/- 2 histidine side chains out of a total of 300 and about 134 +/- 4 lysine side chains out of 310 have been found to be inaccessible to the solvent in the native condition. The remaining titratable side chains determine the surface charge of the native protein. In 1.5 M GuHC1, however, the nontitratable histidine side chains are found to be available for titration as are the nontitratable lysine and tyrosine side chains in 6M GuHC1. The theoretical titration curve computed on the basis of Linderstrøm-Lang model is found to fit quite comfortably with the experimental one between pH 4.6 and 9.25. The pKint value for beta gamma-carboxyl side chains has been found to be 5.18 which is somewhat higher than usual indicating that the carboxyl groups in the protein are probably in some constrained condition which is released in the presence of a denaturant. Below pH 4.6, there begins a conformational change in the alpha-crystallin aggregate as is corroborated from the circular dichroism studies. The value of electrostatic interaction factor w which remains more or less constant between pH 4.6 and 9.25 is also found to gradually fall off below pH 4.6.

Acid-Base Equilibrium↗
PubMed1998

Influence of chemical modification of cysteine and histidine side chains upon subunit reassembly of alpha crystallin.

Alpha crystallin, the important multimeric structural protein of mammalian eye lens, is an assembly composed of 30 alpha-A and 10 alpha-B subunits. The influence of either partial or complete chemical modification of two important amino acid side chains, cysteine and histidine, upon the integrity of native alpha crystallin assembly and also upon the mode of subunit reassembly has been investigated. It has been found that chemical modification of surface-exposed cysteine and histidine side chains does not affect the subunit-subunit interactions stabilizing the native aggregate. Cysteine modifications, either partial or complete, unlike histidine modifications, do not seem to affect the backbone conformation of the subunits refolded after denaturation. Both cysteine and histidine modifications, however, affect the packing of the refolded structural elements forming the tertiary structure of the subunits and also the mode of oligomeric reorganization. The most striking effect of histidine modification is the considerable increase in size of the aggregates upon reassociation of the modified subunits. The chaperone activity, however, has been found to remain almost unaffected in spite of these chemical modifications.

Chromatography, Gel↗
PubMed1998

The MCM16 gene of the yeast Saccharomyces cerevisiae is required for chromosome segregation.

We have cloned and characterized the MCM16 gene required for the maintenance of minichromosomes in the yeast Saccharomyces cerevisiae. This gene corresponds to a 181-amino acid ORF, YPR046W, on chromosome XVI. Mutant cells carrying minichromosomes accumulate them in higher copy numbers than do wild-type cells. Intact dicentric plasmid could be recovered from the mutant, in contrast to the wild-type, in which the plasmid suffered frequent deletions. A wild-type centromere, CEN6, acts as a block to the transcription of a reporter gene, such as beta-galactosidase. This block was less effective in the mutant than in the wild-type strain, suggesting alterations in kinetochore assembly in the former. The mutant also showed increased sensitivity to the antimitotic drugs benomyl and thiabendazole. The mcm16 mutation caused a high rate of loss of chromosome III, without any significant increase in the recombination frequency. A strain carrying a deletion-disruption derivative of the MCM16 gene was viable and, when compared to the wild-type, did not show any significant changes in growth rate or cell morphology at 16, 23 and 37 degrees C. These properties show that MCM16 is required for an important but nonessential role that governs the kinetochore-microtubule mediated process of chromosome segregation.

Chromosome Deletion↗
PubMed1998

Targets of oxidative stress in cardiovascular system.

Although oxidants such as superoxide (O2.) and hydrogen peroxide (H2O2) play a role in host-mediated destruction of foreign pathogens yet excessive generation of oxidants may lead to a variety of pathological complications in the cardiovascular system. An important mechanism by which oxidants cause dysfunction of the cardiovascular system appears to be due to the increase in intracellular free Ca2+ concentration. Oxidants cause cellular Ca2+ mobilization by modulating activities of a variety of regulators such as Na+/H+ and Na+/Ca2+ exchangers, Na+/K+ ATPase and Ca2+ ATPase and Ca2+ channels that are associated with Ca2+ transport in the plasma membrane and the sarco(endo)plasmic reticular membrane of myocardial cells. Recent research have suggested that the increase in Ca2+ level by oxidants plays a pivotal role in inducing several protein kinases such as protein kinase C, tyrosine kinase and mitogen activated protein kinases. Oxidant-mediated alteration of different signal transduction systems and their interations eventually regulate a variety of pathological conditions such as atherosclerosis, apoptosis and necrosis in the myocardium.

Animals↗
PubMed1998

Characterization of MboI satellites in Cirrhina mrigala and Clarias batrachus (Pisces).

We have cloned and characterized two highly reiterated, tandemly repeated, and A+T rich MboI DNA fragments, one in Cirrhina mrigala (Cyprinidae), with a monomer size of 266 bp, and one in Clarias batrachus (Clariidae), with a monomer size of 227 bp. The MboI fragment in C. mrigala is species-specific and absent in other carps, such as Catla catla and Labeo rohita. The MboI fragment in C. batrachus was also present in two other catfishes tested, namely Clarias gariepinus and Heteropneustes fossilis. In C, mrigala x C. catla and C. mrigala x L. rohita hybrids, the C. mrigala specific MboI fragment is inherited uniparentally. In the reciprocal hybrids of C. batrachus x H. fossilis, the satellite ladder contains the bands of both parental species. The MboI satellite of carp may be useful in genetic introgression analysis and that of catfish in distinguishing between gynogenetic progeny and true hybrids.

Animals↗
PubMed1998

The effect of glutathione upon chaperone activity of alpha-crystallin is probably mediated through target modulation.

We studied the effect of the presence of glutathione, both reduced (GSH) and oxidized (GSSG), on the chaperone activity of alpha-crystallin towards thermal aggregation of gamma 2-crystallin. Results showed that while GSH enhanced the chaperone activity of alpha-crystallin, GSSG appeared to diminish it to some extent. When, however, the effect of the presence of glutathione was studied in the absence of alpha-crystallin, the nature of their action was found to be almost similar to that observed in its presence. These results suggest that glutathiones probably modulate the target protein which ultimately influences the chaperone activity of alpha-crystallin.

Animals↗
PubMed1998

Evaluation of lambdacyhalothrin-impregnated bednets in a malaria endemic area of India. Part 3. Effects on malaria incidence and clinical measures.

In Indian villages with high malaria endemicity use of nylon bednets treated at 25 mg/m2 at 6-month intervals for 3 years caused significant reductions in malaria incidence, slide positivity rate, slide falciparum rate, annual parasite index, and parasite rate in the entire population, as well as reductions in rates of splenomegaly and anemia in children. In villages with untreated nets, considerable reduction also occurred in these parameters except for the rate of splenomegaly. In the village without nets, a relatively small drop occurred in the parasite rate and anemia but no change occurred in malaria incidence, and an increase occurred in the rate of splenomegaly. The trial thus showed the efficacy of impregnated bednets against malaria in the forested hills of Orissa State where the existing control strategy based on indoor residual spraying of DDT has remained incapable of interrupting malaria transmission.

Anemia↗
PubMed1997

Selective up-regulation of cytokine-induced RANTES gene expression in lung epithelial cells by overexpression of IkappaBR.

We previously reported the cloning of a cDNA for IkappaBR (for IkappaB-related) from human lung alveolar epithelial cells. IkappaBR is related to the IkappaB proteins that function as regulators of the NF-kappaB family of transcription factors. Here, we investigated the consequence of IkappaBR overexpression on the expression of NF-kappaB-regulated chemokine genes in lung alveolar epithelial cells. Chemokines play an important role in many inflammatory diseases such as asthma and AIDS. Overexpression of IkappaBR in the lung cells resulted in a rapid 50-100-fold greater production of the RANTES (regulated upon activation, normal T expressed and presumably secreted) protein upon cytokine-induction compared with control cells. IkappaBR overexpression, however, did not enhance interleukin-8 or MIP-1alpha gene expression, despite the fact that the expression of all three chemokine genes are regulated by NF-kappaB. The up-regulation of RANTES gene expression resulting from overexpression of IkappaBR correlated with increased amounts of a unique RANTES-kappaB binding activity and decreased binding of p50 homodimers. Previous studies have shown that p50 homodimers function as repressors of certain kappaB sites. Our studies suggest that IkappaBR can aid activation of select NF-kappaB-regulated genes by sequestering p50 homodimers.

Base Sequence↗
PubMed1997

Effects of mutating specific residues present near the amino terminus of 2'-5'-oligoadenylate synthetase.

In this study, we investigated the role of specific amino acid residues present near the amino terminus of the 9-2 isozyme of 2'-5'-oligoadenylate synthetase. In vitro expression of deletion mutants showed that residues 1-9 are required for enzyme activity. Within this region, residues 3, 7, and 8 were found to be conserved among all known isozymes of 2'-5'-oligoadenylate synthetase. Mutation of these residues singly or in combination resulted in partial or total loss of enzyme activity. Substitution of the proline residue at position 7 by different residues caused a partial or complete loss of activity. The properties of the inactive P7Q mutant were further explored by expressing the protein in bacteria. The bacterially expressed protein was also enzymatically inactive. The mutant protein could bind the substrate ATP and the activator double-stranded RNA normally. Oligomerization properties of the protein were examined by an affinity-based interaction assay and by glycerol gradient centrifugation; there was no detectable difference between the wild type and the P7Q mutant. These results demonstrated the importance of the proline residue at position 7 in conferring enzyme activity to the protein without affecting its other properties.

2',5'-Oligoadenylate Synthetase↗
PubMed1997

Repression of interleukin-6 gene expression by 17 beta-estradiol: inhibition of the DNA-binding activity of the transcription factors NF-IL6 and NF-kappa B by the estrogen receptor.

The cytokine interleukin-6 (IL-6), a key mediator of immune and acute phase responses of the liver, has also been implicated in uterine functions. Estrogens are potent repressors of IL-6 production by uterine stromal cells. In the endometrial adenocarcinoma cell line Ishikawa, phorbol ester-induced activation of the IL-6 promoter was inhibited to basal levels by 17 beta-estradiol (E2) in a wild-type receptor-dependent fashion. Although tamoxifen has been shown to have estrogenic effects on the endometrium, it did not inhibit induction of the IL-6 promoter. We previously showed that inhibition of IL-6 gene expression by E2 does not involve high-affinity binding of the estrogen receptor (ER) to IL-6 DNA. We now report that the ER can directly interact with the transcription factors NF-IL6 and NF-kappa B and can inhibit their ability to bind DNA which might be the molecular basis for repression of IL-6 gene expression by estrogens.

Adenocarcinoma↗