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Biomedical subjects

K Mukhopadhyay

Publications and source records attributed to K Mukhopadhyay.

At least 19 recordsLinked to original sources
PubMed2001

Effect of cholesterol on interaction of dibucaine with phospholipid vesicles: a fluorescence study.

Interaction of the local anesthetic dibucaine with small unilamellar vesicles of dimyristoylphosphatidylcholine (DMPC) and dioleoyl phosphatidylcholine (DOPC) containing different mol percents of cholesterol has been studied by fluorescence spectroscopy. Fluorescence measurements on dibucaine in presence of phospholipid vesicles containing various amounts of cholesterol yielded a pattern of variation of wavelength at emission maximum and steady-state anisotropy which indicated that the microenvironment of dibucaine is more polar and flexible in membranes that contain cholesterol than in membranes without cholesterol. Experiments on quenching of fluorescence from membrane-associated dibucaine by potassium iodide showed a marked increase in quenching efficiency as the cholesterol content of the vesicles was increased, demonstrating increased accessibility of the iodide quenchers to dibucaine in the presence of cholesterol, when compared to that in its absence. Total emission intensity decay profiles of dibucaine yielded two lifetime components of approximately 1 ns and approximately 2.8--3.1 ns with mean relative contributions of approximately 25 and approximately 75%, respectively. The mean lifetime in vesicles was 20--30% smaller than in the aqueous medium and showed a moderate variation with cholesterol content. Fluorescence measurements at two different temperatures in DMPC SUVs, one at 33 degrees C, above the phase transition temperature and another at 25 degrees C, around the main phase transition, indicated two different mode of dibucaine localization. At 25 degrees C dibucaine partitioned differentially in presence and absence of cholesterol. However, at 33 degrees C the apparent partition coefficients remained unaltered indicating differences in the microenvironment of dibucaine in presence and absence of cholesterol in the phospholipid membranes.

Anesthetics, Local↗
PubMed2001

Separation of carrier-free 181Re produced in 16O-irradiated thulium target.

Heavy ion activation of natural Tm2O3 with 90 MeV 16O beam results in the formation of carrier-free short-lived 181Ir and 181Os which ultimately decay out to 181Re in the matrix. The liquid cation exchanger, HDEHP, has effectively been utilized as an extractant for quantitative separation of bulk thulium target matrix from carrier-free rhenium radionuclide.

Journal Article↗
PubMed2000

Granulocyte macrophage-colony stimulating factor (GM-CSF) in neonatal neutropenia.

Neutropenia in neonates is often associated with sepsis, prematurity and maternal hypertension with increased risk of mortality. We describe two neonates with neutropenia treated with granulocyte macrophage colony stimulating factor. The total and absolute neutrophil counts showed a marked response and led to a favourable outcome. Human granulocyte macrophage colony stimulating factor may be used as an adjuvant therapy for neonatal neutropenia of different aetiologies.

Granulocyte-Macrophage Colony-Stimulating Factor↗
PubMed2000

Overexpression of DeltaFosB transcription factor(s) increases bone formation and inhibits adipogenesis.

Members of the AP-1 family of transcription factors participate in the regulation of bone cell proliferation and differentiation. We report here a potent AP-1-related regulator of osteoblast function: DeltaFosB, a naturally occurring truncated form of FosB that arises from alternative splicing of the fosB transcript and is expressed in osteoblasts. Overexpression of DeltaFosB in transgenic mice leads to increased bone formation throughout the skeleton and a continuous post-developmental increase in bone mass, leading to osteosclerosis. In contrast, DeltaFosB inhibits adipogenesis both in vivo and in vitro, and downregulates the expression of early markers of adipocyte differentiation. Because osteoblasts and adipocytes are thought to share a common precursor, it is concluded that DeltaFosB transcriptionally regulates osteoblastogenesis, possibly at the expense of adipogenesis.

Adipocytes↗
PubMed2000

Use of RNA arbitrarily primed-PCR fingerprinting to identify Vibrio cholerae genes differentially expressed in the host following infection.

Evidence suggests that a repertoire of Vibrio cholerae genes are differentially expressed in vivo, and regulation of virulence factors in vivo may follow a different pathway. Our work was aimed at characterization of in vivo-grown bacteria and identification of genes that are differentially expressed following infection by RNA arbitrarily primed (RAP)-PCR fingerprinting. The ligated rabbit ileal loop model was used. The motility of in vivo-grown bacteria increased by 350% over that of in vitro-grown bacteria. Also, the in vivo-grown cells were more resistant to killing by human serum. By using the RAP-PCR strategy, five differentially expressed transcripts were identified. Two in vitro-induced transcripts encoded polypeptides for the leucine tRNA synthatase and the 50S ribosomal protein, and the three in vivo-induced transcripts encoded the SucA and MurE proteins and a polypeptide of unknown function. MurE is a protein involved in the peptidoglycan biosynthetic pathway. The lytic profiles of in vivo- and in vitro-grown cells suspended in distilled water were compared; the former was found to be slightly less sensitive to lysis. Ultrathin sections of both cells observed under the transmission electron microscope revealed that in contrast to the usual wavy discontinuous membrane structure of the in vitro-grown cells, in vivo-grown cells had a more rigid, clearly visible double-layered structure. The V. cholerae murE gene was cloned and sequenced. The sequence contained an open reading frame of 1,488 nucleotides with its own ribosome-binding site. A plasmid containing the murE gene of V. cholerae was transformed into V. cholerae 569B, and a transformed strain, 569BME, containing the plasmid was obtained. Ultrathin sections of 569BME viewed under a transmission electron microscope revealed a slightly more rigid cell wall than that of wild-type 569B. When V. cholerae 569B and 569BME cells were injected separately into ligated rabbit ileal loops, the transformed cells had a preference for growth in the ileal loops versus laboratory conditions.

Amino Acid Sequence↗
PubMed1999

Sequential search and the influence of male quality on female mating decisions.

The patterns of phenotypic association between mated males and females depend on the decision rules that individuals employ during search for a mate. We generalize the sequential search rule and examine how the shape of the function that relates a male character to the benefit of a mating decision influences the threshold value of the male trait that induces females to terminate search. If the fitness function is linear the optimal threshold value of a male character increases with the slope of the function. The phenotypic threshold criterion declines, all else being equal, if the fitness function is made more concave (or less convex) by an increase of the risk of the function. The expression of the trait in females has no effect on the optimal threshold value of a male character if the fitness function is linear and phenotypic values combine additively to influence the benefit of a mating decision; the phenotypic threshold criterion is ubiquitous among females. A convex fitness function induces females with high trait values to adopt a relatively high phenotypic threshold criterion, whereas a concave fitness function induces such females to adopt a low threshold value for the male trait. Thus, linear, convex and concave fitness functions effect random, assortative and disassortative combinations of phenotypes among mated individuals, respectively. Changes of female search behavior induced by changes of the distribution of a male character similarly depend on the shape of the fitness function. A variance-preserving increase of male trait values produces a relatively small increase of the threshold criterion for the male character if the fitness function is concave, relative to conditions in which the fitness function is either linear or convex. Our results suggest that a sequential search rule can in principle induce the kinds of mating patterns observed in nature and that the phenotypic association between mated individuals is likely to depend on how a male character translates into fitness, the distribution of the trait among males and attributes of searching females.

Animals↗
PubMed1999

The tyrosine phosphatase SHP-1 is a negative regulator of osteoclastogenesis and osteoclast resorbing activity: increased resorption and osteopenia in me(v)/me(v) mutant mice.

Naturally occuring inactivating mutations of the Src homology 2 (SH2) domain-containing tyrosine phosphatase 1 (SHP-1) in mice give rise to the motheaten (me) phenotype. me/me mice have multiple hematopoietic abnormalities, suggesting that this phosphatase plays an important role in hematopoiesis. SHP-1 binds to and is activated by several hematopoietic surface receptors, including the colony-stimulating factor type 1 receptor. We have examined the role of SHP-1 in osteoclastogenesis and osteoclast function using mice with the viable motheaten (me(v)/me(v)) mutation, which has markedly decreased SHP-1 activity. Histomorphometric analysis of 6-week-old me(v)/me(v) mice and control littermates showed a marked osteopenia with an increase in bone resorption indices. The number of formed osteoclast-like cells (OCLs) in cocultures of me(v)/me(v) hematopoietic cells with normal osteoblasts was significantly increased. In contrast, the number of OCLs formed in the coculture of normal bone marrow cells with the me(v)/me(v) osteoblasts was not significantly different from controls. The bone-resorbing activity of me(v)me(v) OCLs and authentic osteoclasts was also found to be increased. Finally, Western blotting of proteins from me(v)/me(v) and control OCLs revealed an overall increase in tyrosine phosphorylation in the me(v)/me(v) lysates. These in vivo and in vitro results suggest that SHP-1 is a negative regulator of bone resorption, affecting both the formation and the function of osteoclasts.

Animals↗
PubMed1999

Tracheal agenesis.

Explore the source record for details and available documents.

Abnormalities, Multiple↗
PubMed1998

Conformation induction in melanotropic peptides by trifluoroethanol: fluorescence and circular dichroism study.

Conformation induction in the two related peptides, alpha-melanocyte stimulating hormone (alpha-MSH) and delta-melanocyte stimulating hormone (delta-MSH), have been studied in solvent media containing varying percentages of the membrane-mimetic solvent 2,2,2-trifluoroethanol (TFE) using fluorescence and circular dichroism (CD) spectroscopy. Singular value decomposition (SVD) analysis of the CD spectra at different TFE concentrations showed that these spectra can be described as linear combinations of only two distinct basis spectra, corresponding to the peptides in the random-coil and 'folded' conformations. For alpha-MSH the spectrum of the folded state is very similar to the standard spectrum of the alpha-helix, while that for delta-MSH has partial resemblance to the helical spectrum. Fitting the data on ellipticity (at 222 nm) as a function of TFE volume fraction to an equation based on a two-state model describing TFE-induced conformation induction in the peptides gave values of (1.1 +/- 0.4) and (4.2 +/- 0.5) kcal mol-1 for alpha-MSH and delta-MSH, respectively, for the free energy of equilibrium between the helix and coil forms in water. Measurement of fluorescence emission parameters (emission maximum, quantum yield, steady-state anisotropy and mean excited-state lifetime) indicated that the microenvironment around the single tryptophan residues of both peptides changes in like manner with increasing concentration of TFE in the solvent. The similarity of fluorescence behaviour of the peptides suggests that their Trp fluorophores do not participate in secondary structure formation in TFE.

Circular Dichroism↗
PubMed1998

Role of early postnatal dexamethasone in respiratory distress syndrome.

OBJECTIVE: To study the effect of early postnatal dexamethasone therapy on severity of hyaline membrane disease. DESIGN: Prospective, randomized, controlled, unblinded study. SETTING: Neonatal Intensive Care Unit. METHODS: 19 babies who had hyaline membrane disease were included in this study. The inclusion criteria were clinical and radiographic diagnosis of RDS, requiring mechanical ventilation and FiO2 > 0.3. Ten babies received injection dexamethasone 0.5 mg/kg/dose 12 hourly for 3 days starting within 6 hours of birth. The control group did not receive any drug. Babies with active infection, bleeding tendency and congenital malformation were excluded. None of the babies received surfactant. The duration of ventilation and AaDO2 and FiO2 requirements from day one to five were calculated. RESULTS: The initial AaDO2 were similar in both the groups but on day 3, 4, 5 AaDO2 were low in study group (201, 85, 70) compared to control group (236, 209, 162). The initial FiO2 were 0.66 and 0.63 in dexamethasone and control groups, respectively and remained high till day 2 and came down in study group on days 3, 4 and 5 (0.41, 0.27, 0.27) compared to control group (0.53, 0.34, 0.42). The mean duration of ventilation was shorter in dexamethasone group (87 hours) vs control group (120 hours). CONCLUSION: Early use of postnatal dexamethasone reduces the disease severity and oxygen requirement in RDS and hence would be useful in the Indian context.

Anti-Inflammatory Agents↗
PubMed1997

Endogenous protein phosphorylation and protein kinase activity in winged bean.

In winged bean (Psophocarpus tetragonolobus) protein kinases (E.C. 2.7.1.37) were found in all tissues studied. There was a significant increase in kinase activity during seed development, with a concomitant enhancement in the phosphorylation of a number of polypeptides; this was reversed in germinating seed cotyledons. Protein phosphorylation was apparently correlated with the increase in the protein content of the developing seed and the growing axis. At least three distinct autophosphorylating proteins could be distinguished in the developing seeds after SDS-PAGE, indicating the presence of different types of protein kinases in winged bean.

Fabaceae↗
PubMed1997

Purification and characterisation of a protein kinase from winged bean.

A soluble, cytoplasmic protein kinase was purified from the developing seeds of winged bean (Psophocarpus tetragonolobus) following conventional methods of protein purification including anion-exchange chromatography, gel-filtration and Blue Sepharose chromatography. The purified enzyme consists of a single polypeptide of M(r) 45,000 as determined by SDS-PAGE and gel-filtration chromatography on Sephacryl S-200. The pH optimum of the protein kinase activity was 7.0, while the optimum concentration of Mg2+ was 5 mM. The enzyme utilised casein as an exogenous phosphate acceptor. The conventional modulators of protein kinases, including the cyclic nucleotides, Ca2+ and calmodulin, did not stimulate the purified enzyme. Heparin and spermine, too, had no effect on its activity. Phosphoamino acid analysis revealed that the enzyme transferred the gamma-phosphate of ATP only to serine residues of casein. All these characteristics, taken together, classifies the purified protein kinase as a member of the casein kinase I group of enzymes.

Amino Acids↗
PubMed1996

An 18-base-pair sequence in the mouse proalpha1(II) collagen gene is sufficient for expression in cartilage and binds nuclear proteins that are selectively expressed in chondrocytes.

The molecular mechanisms by which mesenchymal cells differentiate into chondrocytes are still poorly understood. We have used the gene for a chondrocyte marker, the proalpha1(II) collagen gene (Col2a1), as a model to delineate a minimal sequence needed for chondrocyte expression and identify chondrocyte-specific proteins binding to this sequence. We previously localized a cartilage-specific enhancer to 156 bp of the mouse Col2a1 intron 1. We show here that four copies of a 48-bp subsegment strongly increased promoter activity in transiently transfected rat chondrosarcoma (RCS) cells and mouse primary chondrocytes but not in 10T1/2 fibroblasts. They also directed cartilage specificity in transgenic mouse embryos. These 48 bp include two 11-bp inverted repeats with only one mismatch. Tandem copies of an 18-bp element containing the 3' repeat strongly enhanced promoter activity in RCS cells and chondrocytes but not in fibroblasts. Transgenic mice harboring 12 copies of this 18-mer expressed luciferase in ribs and vertebrae and in isolated chondrocytes but not in noncartilaginous tissues except skin and brain. In gel retardation assays, an RCS cell-specific protein and another closely related protein expressed only in RCS cells and primary chondrocytes bound to a 10-bp sequence within the 18-mer. Mutations in these 10 bp abolished activity of the multimerized 18-bp enhancer, and deletion of these 10 bp abolished enhancer activity of 465- and 231-bp intron 1 segments. This sequence contains a low-affinity binding site for POU domain proteins, and competition experiments with a high-affinity POU domain binding site strongly suggested that the chondrocyte proteins belong to this family. Together, our results indicate that an 18-bp sequence in Col2a1 intron 1 controls chondrocyte expression and suggest that RCS cells and chondrocytes contain specific POU domain proteins involved in enhancer activity.

Animals↗
PubMed1995

Use of a new rat chondrosarcoma cell line to delineate a 119-base pair chondrocyte-specific enhancer element and to define active promoter segments in the mouse pro-alpha 1(II) collagen gene.

We show that a new rat chondrosarcoma (RCS) cell line established in long-term culture from the Swarm tumor displayed a stable differentiated chondrocyte-like phenotype. Indeed, these cells produced the collagen types II, IX, and XI and alcian blue-stainable cartilage-specific proteoglycans, but no type I or type III collagen. To functionally characterize their chondrocytic nature, the cells were stably transfected with a type II collagen/beta geo chimeric gene which confers essentially perfect chondrocyte-specific expression in transgenic mice. RCS cells expressed both beta-galactosidase and G418 resistance, in comparison with similarly transfected 10T1/2 and NIH/3T3 fibroblasts which did not. These cells were then used to perform a systematic deletion analysis of the first intron of the mouse type II collagen gene (Col2a1) using transient expression experiments to determine which segments stimulated expression of a luciferase reporter gene in RCS cells but not in 10T1/2 fibroblasts. Cloning of two tandem copies of a 156-base pair (bp) intron 1 fragment (+2188 to +2343) in a construction containing a 314-bp Col2a1 promoter caused an almost 200-fold increase in promoter activity in RCS cells but no increase in 10T1/2 cells. DNase I footprint analysis over this 156-bp fragment revealed two adjacent protected regions, FP1 and FP2, located in the 3'-half of this segment, but no differences were seen with nuclear extracts of RCS cells and 10T1/2 fibroblasts. Deletion of FP2 to leave a 119-bp segment decreased enhancer activity by severalfold, but RCS cell specificity was maintained. Further deletions indicated that sequences both in the 5' part of the 119-bp fragment and in FP1 were needed simultaneously for RCS cell-specific enhancer activity. A series of deletions in the promoter region of the mouse Col2a1 gene progressively reduced activity when these promoters were tested by themselves in transient expression experiments. However, these promoter deletions were all activated to a similar level in RCS cells by a 231-bp intron 1 fragment that included the 156-bp enhancer. The RCS cell-specific activity persisted even if the Col2a1 promoter was replaced by a minimal adenovirus major late promoter. This 231-bp intron 1 fragment also had strong enhancing activity in transiently transfected mouse primary chondrocytes. Our experiments establish the usefulness of RCS cells as an experimental system for studies of the control of chondrocyte-specific genes, provide an extensive delineation of segments in the Col2a1 first intron involved in chondrocyte-specific activity, and show that promoter sequences are dispensable for chondrocyte specificity.

3T3 Cells↗
PubMed1995

Aging of tubulin monomers using 5,5'-bis(8-anilino-1-naphthalenesulfonate) as a probe.

The fluorescent probe bis(8-anilino-1-naphthalenesulfonate) (bis-ANS) has been used to monitor the time- and temperature-dependent aging of tubulin, whereby new hydrophobic binding sites of lower affinity are generated on the protein [Prasad, A. R. S., et al. (1986a) Biochemistry 25, 739-742]. We carried out a detailed analysis of this phenomenon and found that, in addition to antimitotic drugs like colchicine or vinblastine, other parameters, viz., low temperature and protein stabilizers (e.g., glycerol and sucrose), inhibit the extent of enhanced binding of bis-ANS. Moreover, the generation of additional bis-ANS binding sites are also suppressed at high concentrations of tubulin. Cleavage of the carboxy-termini of tubulin (bound to bis-ANS) by subtilisin causes a significant reduction in the enhanced fluorescence, but has no effect on the high-affinity binding site of bis-ANS. All of these observations can be explained by the correlation of the presence of additional binding with the dissociation of heterodimeric tubulin into monomers. Enhanced binding of bis-ANS is due to tubulin dimers that have undergone dissociation, resulting in a loosening of its tertiary structure with the generation of a plethora of hydrophobic sites.

Anilino Naphthalenesulfonates↗
PubMed1995

A 182 bp fragment of the mouse pro alpha 1(II) collagen gene is sufficient to direct chondrocyte expression in transgenic mice.

Type II collagen is a major chondrocyte-specific component of the cartilage extracellular matrix and it represents a typical differentiation marker of mature chondrocytes. In order to delineate cis-acting elements of the mouse pro alpha 1(II) collagen gene that control chondrocyte-specific expression in intact mouse embryos, we generated transgenic mice harboring chimeric constructions in which varying lengths of the promoter and intron 1 sequences were linked to a beta-galactosidase reporter gene. A construction containing a 3,000 bp promoter and a 3,020 bp intron 1 fragment directed high levels of beta-galactosidase expression specifically to chondrocytes. Expression of the transgene coincided with the temporal expression of the endogenous gene at all stages of embryonic development. Successive deletions of intron 1 delineated a 182 bp fragment which targeted beta-galactosidase expression to chondrocytes with the same specificity as the larger intron 1 fragment. Transgenic mice harboring a 309 bp Col2a1 promoter lacking intron 1 tester sequences showed no beta-galactosidase expression in chondrocytes. Reduction of the 182 bp fragment to a 73 bp subfragment surrounding a decamer sequence previously reported to be involved in chondrocyte specificity, resulted in loss of transgene expression in chondrocytes. When the Col2a1 promoter was replaced with a minimal beta-globin promoter, the 182 bp intron 1 sequence was still able to target expression of the transgene to chondrocytes. We conclude that a 182 bp intron 1 DNA segment of the mouse Col2a1 gene contains the necessary information to confer high-level, temporally correct, chondrocyte expression on a reporter gene in intact mouse embryos and that Col2a1 promoter sequences are dispensable for chondrocyte expression.

Animals↗