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Biomedical subjects

S Kumar

Publications and source records attributed to S Kumar.

At least 757 records · Page 42Linked to original sources
PubMed1999

Sequence context effects on mutational properties of cis-opened benzo[c]phenanthrene diol epoxide-deoxyadenosine adducts in site-specific mutation studies.

Diastereomeric N6-substituted dAdo adducts (cis B[c]PhDE-2/1R and cis B[c]PhDE-2/1S) that correspond to cis-opening at C-1 of the enantiomeric benzo[c]phenanthrene 3,4-diol 1,2-epoxides in which the epoxide oxygen and the benzylic hydroxyl group are trans (DE-2) were synthetically incorporated into oligonucleotide 16-mers. Each adduct was placed at the fourth nucleotide from the 5'-end of each of two different oligonucleotide sequences derived from the E. coli supF gene. Each adduct was also placed in two additional oligonucleotide sequences that were constructed by interchanging the adduct site and the immediately adjacent nucleotides between the two original sequences. These oligonucleotides were designed for use in site-specific mutation studies, with a single-stranded bacteriophage M13mp7L2 vector, to determine if the effects of sequence context on types and frequencies of base substitution mutations are attributable only to nucleotides immediately adjacent to these polycyclic aromatic hydrocarbon diol epoxide-dAdo adducts, or whether more distant nucleotide residues also affect the mutagenic response. In SOS-induced Escherichia coli SMH77, total base substitution mutation frequencies for the cis B[c]PhDE-2/1R-dAdo adduct were relatively low (0.62-5.6%) compared with those for the cis B[c]PhDE-2/1S-dAdo adduct (11.9-56.5%). Depending on sequence context, cis B[c]PhDE-2/1R-dAdo gave predominantly A-->T or a more equal distribution of A-->T and A-->G mutations whereas cis B[c]PhDE-2/1S-dAdo gave either predominantly A-->T or predominantly A-->G base substitutions. Our results clearly indicate that nucleotides that are distal as well as those that are proximal to the adduct site are capable of influencing both the mutation frequency and the distribution of base substitution mutations.

Bacteriophage M13↗
PubMed1999

TL1, a novel tumor necrosis factor-like cytokine, induces apoptosis in endothelial cells. Involvement of activation of stress protein kinases (stress-activated protein kinase and p38 mitogen-activated protein kinase) and caspase-3-like protease.

TL1 is a recently discovered novel member of the tumor necrosis factor (TNF) cytokine family. TL1 is abundantly expressed in endothelial cells, but its function is not known. The present study was undertaken to explore whether TL1 induces apoptosis in endothelial cells and, if so, to explore its mechanism of action. Cultured bovine pulmonary artery endothelial cells (BPAEC) exposed to TL1 showed morphological (including ultrastructural) and biochemical features characteristic of apoptosis. TL1-induced apoptosis in BPAEC was a time- and concentration-dependent process (EC50 = 72 ng/ml). The effect of TL1 was not inhibited by soluble TNF receptors 1 or 2. TL1 up-regulated Fas expression in BPAEC at 8 and 24 h after treatment, and significantly activated stress-activated protein kinase (SAPK) and p38 mitogen-activated protein kinase (p38 MAPK). The peak activities of SAPK and p38 MAPK in TL1-treated BPAEC were increased by 9- and 4-fold, respectively. TL1-induced apoptosis in the BPAEC was reduced by expression of a dominant-interfering mutant of c-Jun (62.8%, p < 0.05) or by a specific p38 inhibitor, SB203580 (1-10 microM) dose-dependently. TL1 also activated caspases in BPAEC, and TL1-induced apoptosis in BPAEC was significantly attenuated by the caspase inhibitor, ZVAD-fluromethyl-ketone. The major component activated by TL1 in BPAEC was caspase-3, which was based on substrate specificity and immunocytochemical analysis. These findings suggest that TL1 may act as an autocrine factor to induce apoptosis in endothelial cells via activation of multiple signaling pathways, including stress protein kinases as well as certain caspases.

Animals↗
PubMed1999

Expectant management of stage A-1 (T1a) prostate cancer utilizing serum PSA levels: a preliminary report.

BACKGROUND AND OBJECTIVES: The natural history of stage A-1 (Tla) prostate cancer remains unpredictable. Although stage A-1 (Tla) was traditionally considered an indolent lesion, recent reports have documented the potential progression of stage A-1 (Tla) cancer. METHODS: Eight men aged 65 to 76 years at the time of diagnosis with stage A-1 (Tla) prostate cancer received no therapy. These men have been followed from 3 to 9.5 years (mean, 6.25 years). During this period, the patients have been followed with periodic rectal examinations and prostate-specific antigen (PSA) levels. RESULTS: The PSA levels of five of the eight patients remain in the normal range and no patient has had a change in his rectal examination. In the three patients who had elevated PSA levels, there was no evidence of metastatic disease. No patient has died from prostate cancer; one patient died from cerebrovascular causes. CONCLUSIONS: Patients with stage A-1 (Tla) prostate cancer have an unpredictable natural history. PSA levels can be used to monitor disease progression and identify those patients where observation is no longer appropriate.

Aged↗
PubMed1999

Regulatory light chain phosphorylation and the assembly of myosin II into the cytoskeleton of microcapillary endothelial cells.

During the crawling movements of non-muscle cells, myosin II-containing structures assemble and disassemble with a high degree of spatial and temporal heterogeneity. In order to understand how this is controlled, we examined factors that influence the association of myosin II with detergent-resistant cytoskeletons of cultured endothelial cells. Treatment of cells with 0.05% Triton X-100 in an actin-stabilizing buffer released approximately 42% of the myosin II from the cytoplasm. Most remaining myosin II was dissociated from the cytoskeleton by treatment with ATP or AMPPNP, but not ADP, suggesting that myosin II is retained as ATP-sensitive filaments or via rigor-like binding to F-actin. Disruption of actin filaments with cytochalasin or latrunculin prior to detergent permeabilization sharply decreased the amount of myosin II retained, suggesting the latter type of association. Because phosphorylation of myosin II affects filament assembly and actin binding in vitro, phosphorylation levels in soluble and cytoskeletal myosin II were measured. Phosphorylation of myosin heavy chains was not significantly different between the two fractions, but regulatory light chains of cytoskeletal myosin II were 5 times more phosphorylated than in soluble myosin II. Tryptic-peptide mapping showed that cytoskeletal light chains were phosphorylated predominantly at serine 19/threonine 18, which regulates myosin II assembly in vitro, whereas soluble light chains were not phosphorylated or were phosphorylated at threonine 9. Treating cells with the kinase inhibitor, staurosporine, prior to permeabilization decreased light-chain phosphorylation with concomitant reduction in myosin retention. These observations suggest that assembly of myosin II into cytoskeletal structures, where it can generate and resist forces, is regulated in vivo by phosphorylation of myosin light chains at serine 19/threonine 18.

Actins↗
PubMed1999

Fine-needle aspiration biopsy of orbital meningioma.

Fine-needle aspiration was performed on an orbital mass in a 53-yr-old woman. The cytologic diagnosis was meningioma, which was later confirmed by histologic study of biopsy material. The cytologic features of the aspirated material (nuclear pseudoinclusions, psammoma bodies, and cells arranged in syncytial sheets and whorls) allowed a rapid, reliable, outpatient diagnosis to be formulated. The cytologic features are discussed and the literature is reviewed. Diagn. Cytopathol. 1999;21:402-404.

Biopsy, Needle↗
PubMed1999

Cervical radiculopathies: relationship between symptom duration and spontaneous EMG activity.

The purpose of this multicenter study was to prospectively examine whether denervation in paraspinal muscles (PSM) and in other major proximal and distal muscles is related to symptom duration in cervical radiculopathies (CRs). Information was collected on 93 electrodiagnostically confirmed CRs using standardized history, physical examination, and electromyographic (EMG) screens. Multivariate, maximum-likelihood estimates showed no evidence of correlation between PSM spontaneous activity and symptom duration. Symptom duration was also nonsignificant in eight of the remaining nine upper limb muscles analyzed. We conclude that the probability of spontaneous activity is not related to symptom duration. Clinicians, therefore, should refrain from interpreting electrodiagnostic findings based upon duration of symptoms.

Adolescent↗
PubMed1999

Relative roles of T-cell receptor ligands and interleukin-2 in driving T-cell proliferation.

Stimulation of T cells by the T-cell receptor (TCR)/CD3 complex results in interleukin-2 (IL-2) synthesis and surface expression of the IL-2 receptor (IL-2R), which in turn drive T-cell proliferation. However, the significance of the requirement of IL-2 in driving T-cell proliferation, when TCR stimulation itself delivers potential mitogenic signals, is unclear. We show that blocking of IL-2 synthesis by Cyclosporin A (CsA) suppressed both the Concanavalin A (Con A)- and phorbol myristate acetate (PMA)/ionomycin-induced proliferation of T cells. The latter is also inhibited by anti-IL-2R. Kinetic studies showed that T-cell proliferation begins to become resistant to CsA inhibition by about 12 h and became largely resistant by 18 h of stimulation. PMA, the protein kinase C activator, enhanced Con A-induced T-cell proliferation if added only within first 12 h of stimulation, and not after that. Given the fact that, in the present study, TCR is downregulated within 2 h of Con A stimulation and T cells entered the S phase of cell cycle by about 18 h of stimulation, the above results suggest that TCR stimulation provides the initial trigger to the resting T cells, which allows the cells to traverse the first two third portions of G1 phase of cell cycle and become proliferation competent. IL-2 action begins afterward, delivering the actual proliferation signal(s), allowing the cells to traverse the rest of G1 phase and enter the S phase of the cell cycle.

Animals↗
PubMed1999

Inhibition of keratinocyte growth by different Nepalese zanthoxylum species.

A total of 11 methanol extracts obtained from four different Nepalese Zanthoxylum species were screened for their antiproliferative activity against the growth of human keratinocytes (HaCaT cells). The extract obtained from Z. armatum barks was highly active with an IC50 value of 11 micrograms/mL. Also, the extracts obtained from Z. oxyphyllum barks and roots with IC50 values of 53 and 57 micrograms/mL, respectively, showed potent activity. Their antiproliferative activity was not due to cytotoxic effects on cell membranes, as documented by the activity of lactate dehydrogenase released from the cytoplasm of keratinocytes, which did not exceed that of the control value. Rather, they also protected against radical-induced damage to model membranes stimulated with 2,2'-azo-bis(2-amidinopropane) dihydrochloride.

Cell Division↗
PubMed1999

beta-Carbolinedione derivatives as topoisomerase I inhibitors.

Pyrrolo[3,4-c]-beta-carbolinedione dimers 5-14 were synthesized from furo[3,4-c]-beta-carbolinediones and diamines by solvent-free TaCl5/silica catalyzed reaction under microwave irradiation. The inhibitory property of these target compounds, the starting materials 2, 31, 32, and the N-alkylated pyrrolo[3,4-c]-beta-carbolinediones 16, 17, 20-30 was tested against the relaxation of supercoiled pRB322 DNA by calf thymus topoisomerases I and II. Some of these compounds, especially 7 and 23 proved to be selective inhibitors of topoisomerase I.

Carbolines↗
PubMed1999

Shear mechanical properties of human lumbar annulus fibrosus.

Function, failure, and remodeling of the intervertebral disc are all related to the stress and strain fields in the tissue and may be calculated by finite element models with accurate material properties, realistic geometry, and appropriate boundary conditions. There is no comprehensive study in the literature investigating the shear material properties of the annulus fibrosus. This study obtained shear material properties of the annulus fibrosus and tested the hypothesis that these properties are affected by the amplitude and frequency of shearing, applied compressive stress, and degenerative state of the tissue. Cylindrical specimens with an axial orientation from seven nondegenerated and six degenerated discs were tested in torsional shear under dynamic and static conditions. Frequency sweep experiments over a physiological range of frequencies (0.1-100 rad/sec) at a shear strain amplitude of 0.05 rad were performed under three different axial compressive stresses (17.5, 25, and 35 kPa). At the largest compressive stress, shear strain sweep experiments (strain amplitude range: 0.005-0.15 rad at a frequency of 5 rad/sec) and transient stress-relaxation tests (shear strain range: 0.02-0.15 rad) were performed. The annulus fibrosus material was less stiff and more dissipative at larger shear strain amplitudes, stiffer at higher frequencies of oscillation, and stiffer and less dissipative at larger axial compressive stresses. The dynamic shear modulus, /G*/, had values ranging from 100 to 400 kPa, depending on the experimental condition and degenerative level. The shear behavior was also predominantly elastic, with values for the tangent of the phase angle (tandelta) ranging from 0.1 to 0.7. The annulus material also became stiffer and more dissipative with degenerative grade; however, this was not statistically significant. The results indicated that nonlinearities, compression/shear coupling, intrinsic viscoelasticity, and, to a lesser degree, degeneration all affect the shear material behavior of the annulus fibrosus, with important implications for load-carriage mechanisms in the intervertebral disc. These material complexities should be considered when choosing material constants for finite element models.

Cadaver↗
PubMed1999

Activated astrocytes display increased 5-HT2a receptor expression in pathological states.

In human brain tissues from patients dying with cerebral infarction, hypertensive encephalopathy, Alzheimer's disease, Huntington's disease, frontotemporal dementia, and Creutzfeldt-Jakob disease there is an activation of astrocytes. Such activated astrocytes display GFAP and strong 5-HT(2A), but not 5-HT(2B) or 5-HT(2C), receptor immunoreactivity; this 5-HT(2A) reaction has not been observed in normal, nonactivated astrocytes. It is suggested that an up-regulation of 5-HT(2A) receptors may be part of an early response reaction in astrocytes, possibly designed to maintain homeostasis or to induce secondary message pathways involving trophic factors or glycogenolysis.

Aged↗
PubMed1999

Production of "authentic" poliovirus RNA-dependent RNA polymerase (3D(pol)) by ubiquitin-protease-mediated cleavage in Escherichia coli.

The first amino acid of "authentic" poliovirus RNA-dependent RNA polymerase, 3D(pol), is a glycine. As a result, production of 3D(pol) in Escherichia coli requires addition of an initiation codon; thus, a formylmethionine is added to the amino terminus. The formylmethionine should be removed by the combined action of a cellular deformylase and methionine aminopeptidase. However, high-level expression of 3D(pol) in E. coli yields enzyme with a heterogeneous amino terminus. To preclude this problem, we developed a new expression system for 3D(pol). This system exploits the observation that proteins fused to the carboxyl terminus of ubiquitin can be processed in E. coli to produce proteins with any amino acid as the first residue when expressed in the presence of a ubiquitin-specific, carboxy-terminal protease. By using this system, authentic 3D(pol) can be obtained in yields of 30-60 mg per liter of culture. While addition of a single glycine, alanine, serine, or valine to the amino terminus of 3D(pol) produced derivatives with a specific activity reduced by at least 25-fold relative to wild-type enzyme, addition of a methionine to the amino terminus resulted in some processing to yield enzyme with a glycine amino terminus. Addition of a hexahistidine tag to the carboxyl terminus of 3D(pol) had no deleterious effect on the activity of the enzyme. The utility of this expression system for production of other viral polymerases and accessory proteins is discussed.

Amino Acid Sequence↗
PubMed1999

Technique of distortion correction in endoscopic images using a polynomial expansion.

A new technique to correct non-linear distortion of endoscopic images based on L2-norm approximation is proposed. A mathematical model is defined that maps the endoscopic images from the distorted image plane onto an undistorted image plane. A set of model parameters is defined, consisting of the image distortion centre, corrected centre and expansion polynomial coefficients representing radial distortion correction. A new method to find the image distortion centre based on a curvature criterion is also developed. The expansion coefficients are estimated on the basis of the degree of straightness of the grid lines in an image of a test grid consisting of dots in a rectangular pattern. The corrected image centre is computed by ensuring that the distances between neighbouring test dots of different grid lines in the corrected image are the same. The quantitative data providing the results and errors of the expansion characteristics are presented. The performance of the proposed distortion correction algorithm is validated with grid patterns of different orientations. The algorithm is also applied to typical endoscopic images, and the results are presented. The high-speed response of the proposed technique is a key step towards on-line camera calibration.

Endoscopy, Gastrointestinal↗
PubMed1999

Real-time automatic extraction of lumen region and boundary from endoscopic images.

A new approach to the automatic extraction of the lumen region and its boundary for gastrointestinal (GI) endoscopic images is presented. First, a quasi region of interest, the darker regions of the image, is segmented using a region splitting scheme termed progressive thresholding. The centre of mass of this segmented region acts as a seed for further processing. Then the lumen region is obtained using a region growing technique called the integrated neighbourhood search (INS). A new quad structure based technique is introduced to enhance the INS speed significantly. A back projection algorithm is suggested to optimise the search for pixels belonging to the lumen region and boundary. A boundary-thinning algorithm is also proposed to remove the redundant pixels from the lumen boundary and to generate a connected single pixel width boundary. The proposed approach does not need a priori knowledge about the image characteristics. The experimental results indicate that the proposed technique enhances the speed of conventional INS by 45.5% to 28.6% based on the lumen size varying from 22,709 pixels to 4947 pixels. The main advantage of the proposed technique is its high-speed response that facilitates real-time analysis of endoscopic images.

Algorithms↗
PubMed1999

Folding funnels and conformational transitions via hinge-bending motions.

In this article we focus on presenting a broad range of examples illustrating low-energy transitions via hinge-bending motions. The examples are divided according to the type of hinge-bending involved; namely, motions involving fragments of the protein chains, hinge-bending motions involving protein domains, and hinge-bending motions between the covalently unconnected subunits. We further make a distinction between allosterically and nonallosterically regulated proteins. These transitions are discussed within the general framework of folding and binding funnels. We propose that the conformers manifesting such swiveling motions are not the outcome of "induced fit" binding mechanism; instead, molecules exist in an ensemble of conformations that are in equilibrium in solution. These ensembles, which populate the bottoms of the funnels, a priori contain both the "open" and the "closed" conformational isomers. Furthermore, we argue that there are no fundamental differences among the physical principles behind the folding and binding funnels. Hence, there is no basic difference between funnels depicting ensembles of conformers of single molecules with fragment, or domain motions, as compared to subunits in multimeric quaternary structures, also showing such conformational transitions. The difference relates only to the size and complexity of the system. The larger the system, the more complex its corresponding fused funnel(s). In particular, funnels associated with allosterically regulated proteins are expected to be more complicated, because allostery is frequently involved with movements between subunits, and consequently is often observed in multichain and multimolecular complexes. This review centers on the critical role played by flexibility and conformational fluctuations in enzyme activity. Internal motions that extend over different time scales and with different amplitudes are known to be essential for the catalytic cycle. The conformational change observed in enzyme-substrate complexes as compared to the unbound enzyme state, and in particular the hinge-bending motions observed in enzymes with two domains, have a substantial effect on the enzymatic catalytic activity. The examples we review span the lipolytic enzymes that are particularly interesting, owing to their activation at the water-oil interface; an allosterically controlled dehydrogenase (lactate dehydrogenase); a DNA methyltransferase, with a covalently-bound intermediate; large-scale flexible loop motions in a glycolytic enzyme (TIM); domain motion in PGK, an enzyme which is essential in most cells, both for ATP generation in aerobes and for fermentation in anaerobes; adenylate kinase, showing large conformational changes, owing to their need to shield their catalytic centers from water; a calcium-binding protein (calmodulin), involved in a wide range of cellular calcium-dependent signaling; diphtheria toxin, whose large domain motion has been shown to yield "domain swapping;" the hexameric glutamate dehydrogenase, which has been studied both in a thermophile and in a mesophile; an allosteric enzyme, showing subunit motion between the R and the T states (aspartate transcarbamoylase), and the historically well-studied lac repressor. Nonallosteric subunit transitions are also addressed, with some examples (aspartate receptor and BamHI endonuclease). Hence, using this enzyme-catalysis-centered discussion, we address energy funnel landscapes of large-scale conformational transitions, rather than the faster, quasi-harmonic, thermal fluctuations.

Adenylate Kinase↗
PubMed1999

Regulation of nitric oxide and prostaglandin E2 production by CSAIDS (SB203580) in murine macrophages and bovine chondrocytes stimulated with LPS.

OBJECTIVE AND DESIGN: To compare two anti-inflammatory drugs: CSAIDS (SB203580) and hydrocortisone on iNOS and COX-2 expression. MATERIAL OR SUBJECTS: Murine macrophages and bovine chondrocytes stimulated with LPS and human OA-affected cartilage were used in this study. TREATMENT: The macrophages and chondrocytes were preincubated (30 min) with 0.1-1.0 microM CSAIDS or 10 microM of hydrocortisone before stimulating them with 1-100 microg/ml LPS. METHODS: The end products of iNOS and COX-2: nitric oxide (NO) and PGE2 were estimated by Greiss method and RIA, respectively. RESULTS: CSAIDS (1 microM) inhibited the production of NO and PGE2 (p< or =0.01) in bovine chondrocytes, but not in murine macrophages (RAW 264.7) (p< or =0.1). In fact, CSAIDS (in murine macrophages) marginally augmented nitrite accumulation (approximately 20%) at 14-24 h of LPS stimulation. Western blot analysis of COX-2 in bovine chondrocytes show decrease in COX-2 expression by hydrocortisone but not CSAIDS, although hydrocortisone and CSAIDS inhibit PGE2 accumulation. Hydrocortisone inhibited both PGE2 and NO production significantly (p< or =0.01) in murine macrophages. Furthermore, hydrocortisone significantly inhibited (p< or =0.01) PGE2 but marginally (p< or =0.05) NO in bovine chondrocytes. CONCLUSION: These experiments demonstrate differential action of CSAIDS and hydrocortisone on NO and PGE2 production in bovine chondrocytes and RAW 264.7 cells.

Animals↗