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Biomedical subjects

S Kumar

Publications and source records attributed to S Kumar.

At least 1,063 records · Page 59Linked to original sources
PubMed1997

Patterns of physician use of angiotensin converting enzyme inhibitors in the inpatient treatment of congestive heart failure.

Randomized clinical trails have demonstrated that angiotensin-converting enzyme (ACE) inhibitors reduce mortality, improve symptoms, and decrease hospitalization rates in congestive heart failure (CHF) patients with impaired left ventricular systolic function. Guidelines from the Agency for Healthcare Policy and Research (AHCPR) endorse the use of ACE inhibitors in eligible CHF patients and note their underutilization in practice. Randomly selected records of 1,212 Medicare CHF patients in Louisiana, discharged between July 1993 and October 1993, were reviewed. Abstracted data were used to characterize practice patterns and pertinent clinical factors influencing current ACE inhibitor utilization by practicing physicians in eligible Medicare CHF patients admitted to acute care hospitals. A total of 1,133 patients admitted were discharged alive; mean age was 77.6 years (64% female; 68% white). One third of the patients (34%) were already receiving ACE inhibitors on admission; of these, 85% were discharged on ACE inhibitors. The remaining 66% of patients were not on an ACE inhibitor on admission; only 35% of these are documented to have been placed on an ACE inhibitor(s) at discharge. Overall, a significantly large number of CHF patient charts (48%) lacked documentation of LV systolic function assessments. On multivariate logistic regression modeling, the following key clinical variables were positively related to the prescription of ACE inhibitors: low ejection fraction, dyspnea and orthopnea, normal creatinine levels, high diastolic blood pressure, cardiomegaly, and increasing age. Among patients with low ejection fraction, factors contributing to not being discharged on ACE inhibitors included: high creatinine levels, history of myocardial infarction or ischemic heart disease, renal failure, and being African American. This study documents the underutilization of ACE inhibitors in patients with impaired left ventricular systolic function. Results suggest the need for increased physician-based educational efforts concerning the use of ACE inhibitors in CHF patients, and also for increasing left ventricular systolic function assessments and documentation of findings in patient charts.

Aged↗
PubMed1997

Quantifying utilization and practice variation of electroconvulsive therapy.

The objective of this study was to quantify inpatient electroconvulsive therapy (ECT) utilization and its practice variation within the State of Louisiana using Medicare data for beneficiaries age 65 years and older. The Louisiana Medicare claims (MedPar) history and the Medicare beneficiary denominator files for fiscal years 1993 and 1994 were used for analysis. Statistical techniques used were: chi 2 to determine significance of the proposed null hypothesis, and the modified systemic component of variance (SCV) to determine the magnitude of variation between the individual parish utilization rates for ECT. The ECT utilization rate for the Louisiana Medicare population was found to be 2.38 per 10,000 person-years, falling well within the range of previous ECT utilization studies in the United States. The chi 2 value was 0.0003 when comparing parishes, indicating the presence of significant nonrandom variation. The SCVs of inpatient treatment for major depression and impatient ECT were 0.47 and 1.34, respectively. Inpatient ECT in this population demonstrates high geographic variability. Further research is required to determine and quantitate the factors responsible for the geographic variation in inpatient ECT utilization within the Louisiana Medicare population.

Aged↗
PubMed1997

Interaction of ethanol and ascorbic acid on lipid metabolism in guinea pigs.

Influence of excessive intake of ascorbic acid (AA) on alcohol induced hyperlipidemia was investigated. In the present study four groups of male guinea pigs were maintained for 30 days as follows: (1) Control group (1 mg AA/100 g body wt). (2) Ethanol group (I mg AA + 9 g ethanol/100 g body wt). (3) AA group (25 mg AA/100 g body wt). (4) AA + Ethanol group (25 mg AA/100 g body wt + 9 g ethanol 100 g body wt). It was shown that tissue ascorbic acid concentration increased with the intake of mega dose of AA. Alcohol administration depleted tissue ascorbic acid content. But coadministration of AA and alcohol enhanced AA levels in comparison with the ethanol group. Alcohol induced hyperlipidemia was reduced in almost all the tissues by the intake of ascorbic acid. This was observed to be due to increased hepatic catabolism of cholesterol to bile acids. However cholesterogenesis was enhanced as evidenced by the increased HMG CoA activity. Thus the results indicate that the mega dose of AA ingestion is beneficial in reducing alcohol induced hyperlipidemia and AA deficiency.

Animals↗
PubMed1996

Activation of the HIV-1 long terminal repeat by cytokines and environmental stress requires an active CSBP/p38 MAP kinase.

The human immunodeficiency virus, type 1 (HIV-1) promoter is known to be activated by proinflammatory cytokines and UV light. These stimuli also activate various members of the mitogen-activated protein kinase family, including JNK/SAPK and CSBP/p38. In HeLa cells containing an integrated HIV-1 long terminal repeat (LTR) -driven reporter, we now show that the specific p38 inhibitor, SB203580, inhibits activation of the HIV-1 LTR by interleukin-1, tumor necrosis factor, UV light, and osmotic stress. Inhibition was 70-90% in all but the case of tumor necrosis factor stimulation, where inhibition was 50%. Each of these stimuli activated p38, which was inhibited by SB203580 in vitro and in vivo with an IC50 (between 0.1 and 1 microM) similar to that required to inhibit transcription. In contrast, SB203580 had no effect on JNK, which was also activated by these stimuli. The NFkappaB sites in the HIV-1 LTR were required for a response to cytokines but not to UV, and SB203580 remained capable of inhibiting UV activation in the absence of the NFkappaB sites. Studies in which SB203580 was added at different times relative to UV stimulation suggested that the critical p38-mediated phosphorylation event occurred between 2 and 4 h after UV treatment. These data indicate that p38 is required for HIV-1 LTR activation but that the action of p38 is delayed, presumably due to substrate unavailability or inaccessibility.

Calcium-Calmodulin-Dependent Protein Kinases↗
PubMed1996

Heteronuclear ribonucleoproteins C1 and C2, components of the spliceosome, are specific targets of interleukin 1beta-converting enzyme-like proteases in apoptosis.

Apoptosis induced by a variety of agents results in the proteolytic cleavage of a number of cellular substrates by enzymes related to interleukin 1beta-converting enzyme (ICE). A small number of substrates for these enzymes have been identified to date, including enzymes involved in DNA repair processes: poly(ADP-ribose) polymerase and DNA-dependent protein kinase. We describe here for the first time the specific cleavage of the heteronuclear ribonucleoproteins (hnRNPs) C1 and C2 in apoptotic cells induced to undergo apoptosis by a variety of stimuli, including ionizing radiation, etoposide, and ceramide. No cleavage was observed in cells that are resistant to apoptosis induced by ionizing radiation. Protease inhibitor data implicate the involvement of an ICE-like protease in the cleavage of hnRNP C. Using recombinant ICE-like proteases and purified hnRNP C proteins in vitro, we show that the C proteins are cleaved by Mch3alpha and CPP32 and, to a lesser extent, by Mch2alpha, but not by ICE, Nedd2, Tx, or the cytotoxic T-cell protease granzyme B. The results described here demonstrate that the hnRNP C proteins, abundant nuclear proteins thought to be involved in RNA splicing, belong to a critical set of protein substrates that are cleaved by ICE-like proteases during apoptosis.

Apoptosis↗
PubMed1996

Differential gene expression of extracellular matrix components in dilated cardiomyopathy.

Extracellular matrix metalloproteinases (MMPs) are activated in dilated cardiomyopathic (DCM) hearts [Tyagi et al. (1996): Mol Cell Biochem 155:13-21]. To examine whether the MMP activation is occurring at the gene expression level, we performed differential display mRNA analysis on tissue from six dilated cardiomyopathy (DCM) explanted and five normal human hearts. Specifically, we identified three genes to be induced and several other genes to be repressed following DCM. Southern blot analysis of isolated cDNA using a collagenase cDNA probe indicated that one of the genes induced during DCM was interstitial collagenase (MMP-1). Northern blot analysis using MMP-1 cDNA probe indicated that MMP-1 was induced three- to fourfold in the DCM heart as compared to normal tissue. To analyze posttranslational expression of MMP and tissue inhibitor of matrix metalloproteinase (TIMP) we performed immunoblot, immunoassay, and substrate zymographic assays. TIMP-1 and MMP-1 levels were 37 +/- 8 ng/mg and 9 +/- 2 ng/mg in normal tissue specimens (P < 0.01) and 2 +/- 1 ng/mg and 45 +/- 11 ng/mg in DCM tissue (P < 0.01), respectively. Zymographic analysis demonstrated lytic bands at 66 kDa and 54 kDa in DCM tissue as compared to one band at 66 kDa in normal tissue. Incubation of zymographic gel with metal chelator (phenanthroline) abolished both bands suggesting activation of neutral MMP in DCM heart tissue. TIMP-1 was repressed approximately twentyfold in DCM hearts when compared with normal heart tissue. In situ immunolabeling of MMP-1 indicated phenotypic differences in the fibroblast cells isolated from the DCM heart as compared to normal heart. These results suggest disruption in the balance of myopathic-fibroblast cell ECM-proteinase and antiproteinase in ECM remodeling which is followed by dilated cardiomyopathy.

Blotting, Northern↗
PubMed1996

Utilization of the same DNA replication origin by human cells of different derivation.

In the past, a highly sensitive and efficient method was developed to map DNA replication origins in human cells, based on quantitative PCR performed on nascent DNA samples. This method allowed the identification of a replication origin in the myeloid HL-60 cell line, located on chromosome 19 within an approximately 500 bp segment near the lamin B2 gene [Giacca et al. (1994) Proc. Natl. Acad. Sci. USA, 91, 7119]. The same procedure has now been further simplified and extended to a variety of other exponentially growing human cells of different histological derivation (three neural, one connectival and one epithelial), with a nearly diploid chromosomal content. In all the six cell lines tested, the origin activity within the lamin B2 gene domain was localized to the same region. Furthermore, the lamin B2 origin was also found to be active in stimulated, but not in quiescent, peripheral blood lymphocytes.

Bone Marrow Cells↗
PubMed1996

Autophosphorylation of type Ibeta cGMP-dependent protein kinase increases basal catalytic activity and enhances allosteric activation by cGMP or cAMP.

Autophosphorylation of purified bovine Ibeta isozyme of cGMP-dependent protein kinase (Ibeta cGK) in the presence of cGMP or cAMP increased basal kinase activity (-cGMP) as much as 4-fold and reduced the Ka for both cGMP and cAMP; maximum catalytic activity (+cGMP) was not altered. Autophosphorylation proceeded with at least two rate components. The faster rate correlated with phosphorylation of Ser-63. The slower rate, as well as the increase in basal kinase activity and decrease in Ka for cyclic nucleotides, correlated with phosphorylation of Ser-79. Autophosphorylation of either residue was an intramolecular reaction. Autophosphorylation of a proteolytically generated Ibeta cGK monomer lacking amino-terminal residues 1-64 increased basal activity (3-fold) and decreased Ka for cAMP (15-fold). This indicated that autophosphorylation of Ser-79 did not require dimeric cGK and that the phosphorylation of Ser-79 in the monomer was sufficient to alter enzymatic characteristics of Ibeta cGK. These studies suggested that increases in intracellular cGMP or cAMP could result in autophosphorylation of Ibeta cGK, which would increase basal kinase activity as well as the sensitivity of cGK to activation by cGMP or to cross-activation by cAMP. Autophosphorylation could also prolong the increased kinase activity after decline of the second messenger.

Allosteric Regulation↗
PubMed1996

Arginine 75 in the pseudosubstrate sequence of type Ibeta cGMP-dependent protein kinase is critical for autoinhibition, although autophosphorylated serine 63 is outside this sequence.

Autoinhibitory domains in many protein kinases include either a phosphorylatable substrate-like sequence or a pseudosubstrate sequence. This study shows that Ibeta cGMP-dependent protein kinase (cGK) autophosphorylates Ser-63, which is in an atypical cGK substrate sequence (-59AQKQAS-) that is amino-terminal to the pseudosubstrate motif (-74KRQAI-). cGMP increases the rate of autophosphorylation (approximately 0.8 phosphate/cGK monomer) approximately 3-fold. Autophosphorylation is an intramolecular process since it is independent of cGK concentration. cGMP activation of cGK enhances proteolysis within and near the pseudosubstrate site; treatment of dimeric cGK with three proteases produces three cGK monomers (approximately 67-70 kDa each). Their amino-terminal sequences are 75RQAISAEPT-, 76QAISAEPTAF-, and 86DIQDLSXV-, respectively. cGMP stimulates these kinases by 10-, 2.5-, and 1.4-fold, respectively, compared with a 10-fold effect on intact cGK. Increased basal activity accounts for the diminished stimulation. Thus, the primary autophosphorylation site of Ibeta cGK is well outside the pseudosubstrate site, but Arg-75 in the pseudosubstrate site is critical for autoinhibition. Autoinhibition also involves elements that are carboxyl-terminal to Arg-75.

Amino Acid Sequence↗
PubMed1996

Interleukin-1 beta-converting enzyme-like protease cleaves DNA-dependent protein kinase in cytotoxic T cell killing.

Cytotoxic T cells (CTL) represent the major defense mechanism against the spread of virus infection. It is believed that the pore-forming protein, perforin, facilitates the entry of a series of serine proteases (particularly granzyme B) into the target cell which ultimately leads to DNA fragmentation and apoptosis. We demonstrate here that during CTL-mediated cytolysis the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs), an enzyme implicated in the repair of double strand breaks in DNA, is specifically cleaved by an interleukin (IL)-1 beta-converting enzyme (ICE)-like protease. A serine protease inhibitor, 3,4-dichloroisocoumarin (DCl), which is known to block granzyme B activity, inhibited CTL-induced apoptosis and prevented the degradation of DNA-PKcs in cells but failed to prevent the degradation of purified DNA-PKcs by CTL extracts. However, Tyr-Val-Ala-Asp-CH2Cl (YVAD-CMK) and other cysteine protease inhibitors prevented the degradation of purified DNA-PKcs by CTL extracts. Furthermore, incubation of DNA-PKcs with granzyme B did not produce the same cleavage pattern observed in cells undergoing apoptosis and when this substrate was incubated with either CTL extracts or the ICE-like protease, CPP32. Sequence analysis revealed that the cleavage site in DNA-PKcs during CTL killing was the same as that when this substrate was exposed to CPP32. This study demonstrates for the first time that the cleavage of DNA-PKcs in this intact cell system is exclusively due to an ICE-like protease.

Amino Acid Sequence↗
PubMed1996

Genetic regulation of protective immune response in congenic strains of mice vaccinated with a subunit malaria vaccine.

The C-terminal 19-kDa, epidermal growth factor-like region of the merozoite surface protein 1 (MSP1) has been used as a vaccine to induce protective immunity to Plasmodium yoelii in mice and to Plasmodium falciparum in monkeys. To analyze the mechanisms and genetic regulation of this MSP1 vaccine-induced protection, we studied the immunologic correlates of protection in H-2 recombinant and congenic mouse strains on the B10 background. Multiple H-2-linked loci were found to contribute, each with a different mechanism. One locus mapped to the I-A region based on the strong protection in C57BL/10 mice compared with intermediate protection in B10.A(4R) mice and the lack of a difference between B10.AKM and B10.MBR mice. Differences in efficacy of passively transferred antisera from vaccinated C57BL/10 vs B10.A(4R) mice indicated that the protection regulated by the I-A locus was at least in part Ab dependent. Two loci mapped to the right of I-A (FE, H-2S, or H-2D) based on a correlation with the number of H-2k loci to the right of I-A in mice that were I-Ak. One effect was Ab independent and may correspond to a possible negative effect of the I-Ek locus. T cells from protected and nonprotected strains differed in their production of IFN-gamma and TNF-alpha following immunization with MSP1(19), but it was unclear how the differential patterns of cytokine expression related to the level of protection. Thus, MSP1(19) vaccine-induced protection is regulated by H-2-linked loci corresponding to two different immune mechanisms. These findings may indicate the need for more than one Ag in a vaccine to protect an HLA-diverse population.

Animals↗
PubMed1996

Using antibodies to perturb the coordination sphere of a transition metal complex.

Metal ions in the active sites of many metalloenzymes exhibit distinctive spectral and chemical features which are different from those of small inorganic complexes. These features are the result of the unusual geometric and electronic constraints that are imposed on the metal ion within the protein environment. Much effort has been invested to try to mimic this feature of metalloenzymes in synthetic systems, but this remains a formidable task. Here we show that one of the key lessons learned from the science of catalytic antibodies--that binding energy can be converted into chemical energy--can be exploited to 'fine-tune' the physicochemical properties of a metal complex. We show that an antibody's binding site can reversibly perturb the coordination geometry of a metal ion, and can stabilize a high-energy coordinated species. Specifically, antibodies designed to bind the organosilicon compound 1 also bind the geometrically similar Cu(I) complex 2. However, the antibody binds a slightly compressed form of 2, which is closer in size to 1. This distortion is manifested by a spectral shift--an 'immunochromic' effect.

Animals↗
PubMed1996

Determination of the number and location of the manganese binding sites of DNA quadruplexes in solution by EPR and NMR in the presence and absence of thrombin.

The interaction of a DNA quadruplex with thrombin has been studied by first determining the sites of manganese binding to the quadruplex in the absence of thrombin. This has been followed by determining if the interactions with thrombin displace the bound manganese. A different DNA quadruplex has also been studied as a control. The refined solution structures of two DNA quadruplexes have been used to predict the electrostatic potentials of these DNAs. The calculated electrostatic potentials have been used to predict the locations of the binding sites of the paramagnetic ion manganese to these DNAs. The enhanced relaxation of DNA protons due to the binding of the paramagnetic metal ion Mn2+ has been used to experimentally determine the locations of the binding sites. The NMR results and the predictions based on the electrostatic potentials both place the binding sites of the manganese in the narrow grooves of these quadruplex DNAs. The predicted locations are spatially close to those experimentally observed, and the predicted and experimental locations also have similar electrostatic potential energy. These results have allowed a validation of the predictions of electrostatic potentials from structure. The 15mer quadruplex has two strong Mn2+ binding sites with one in each narrow groove. Both Mn2+ are released when the 15mer is complexed with thrombin, indicating that both narrow grooves are involved in the 15mer-thrombin interactions. The dimer quadruplex has a different structural motif than the 15mer and the presence of thrombin does not appreciably affect its interactions with Mn2+.

Base Sequence↗