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Biomedical subjects

G Kulkarni

Publications and source records attributed to G Kulkarni.

17 recordsLinked to original sources
PubMed2006

eEF1A2 activates Akt and stimulates Akt-dependent actin remodeling, invasion and migration.

eEF1A2 (eukaryotic protein elongation factor 1 alpha 2) is a protein translation factor that is likely a human oncogene by virtue of its capacity to transform mammalian cells and its high expression in tumors of the ovary, breast and lung. Here, we show that expression of eEF1A2 is sufficient to stimulate the formation of filopodia in BT549 human breast cancer cells and non-transformed Rat2 cells. Filopodia formation in eEF1A2-expressing cells is dependent on the activity of phosphatidylinositol-3 kinase (PI3K), and the ROCK and Akt kinases. Furthermore, eEF1A2 expression is sufficient to activate Akt in a PI3K-dependent fashion and inactivation of eEF1A2 by short interfering RNA reduces Akt activity. Using breast cancer cell line BT 549, we show that eEF1A2 expression stimulates cell migration and invasion in a largely PI3K- and Akt-dependent manner. These results suggest that eEF1A2 regulates oncogenesis through Akt and PI3K-dependent cytoskeletal remodeling.

Actins↗
PubMed2006

An in vitro comparison of short and long term bond strengths of polyacid modified composite resins to primary human and bovine enamel and dentine.

AIM: This in vitro study compared the shear bond strengths and fracture patterns over 7 and 180 day periods of two PMCRs bonded to both human and bovine primary tooth enamel and dentine to determine if the bovine tooth model is a suitable substitute for the human tooth model. METHODS: Flattened enamel and dentine surfaces were produced using water irrigated #600 grit SiC paper and the teeth randomly placed in groups of N=10. Cylinders of Dyrat AP (Dentsply/DeTrey) or F2000 (3M/ESPE) were bonded to the surfaces that were either etched with 37% H(3)PO(4) (etch, E groups) or treated with their respective conditioners, primers and adhesives (no etch, NE groups). The teeth were stored in distilled water at 37(o)C and shear bond strength (SBS) tested to failure at either 7 (human and bovine) or 180 days (bovine). The modes of failure were assessed under a dissecting microscope at x30. RESULTS: In general the results showed comparable mean SBS for both human and bovine enamel with Dyract AP, E and NE and also for F2000, E. Lower enamel SBS were recorded for F2000 NE but there was no significant differences in mean SBS for bovine enamel after long term storage. For dentine, significantly larger SBS were recorded for human versus bovine teeth for all of the four bonding protocols. There were also significant species differences, the mean SBS for Dyract AP, E and F2000, E for human dentine were higher than bovine but the mean SBS for the respective NE groups showed no significant differences between species. It was concluded that 37% H(3)PO(4) has a detrimental effect on SBS for bovine dentine. Over long term storage SBS of bovine dentine bonds decreased. The modes of failure were related to SBS with varying degrees of significance. CONCLUSION: Bovine primary incisor enamel and dentine, provided the latter is not conditioned with 37% H(3)PO(4), are suitable alternative test SBS substrates for human enamel and dentine.

Analysis of Variance↗
PubMed2005

Cervical intra-/extramedullary solitary fibrous tumour.

A 53-year-old man presented with a 9-month history of symptoms of right-sided weakness, tingling and hypersentivity to clothes on both sides of the body. MRI revealed a large intraspinal intradural tumour at the level of C3-C4 in the cervical cord. The final histology was a solitary fibrous tumour (SFT) of the cervical spinal cord. The radiological diagnosis, surgical management and histology are reviewed.

Cervical Vertebrae↗
PubMed2004

An effective method for improving the fertility of glycerol-exposed poultry semen.

Semen cryopreservation is necessary for banking germplasm from critical poultry stocks. To date, glycerol is the most effective cryoprotectant for poultry sperm; however, the contraceptive effects of glycerol require a significant reduction of the cryoprotectant from thawed semen before artificial insemination (AI). The effectiveness of glycerol reduction by dialysis, Percoll density gradient centrifugation, or washing through 12% (wt/vol) Accudenz was evaluated by fertility trials with highly inbred chicken research lines and commercial turkey lines. Semen was extended 1:1 and then diluted with glycerolized extender to yield a final 11% (vol/vol) glycerol concentration. Glycerolized rooster semen was aliquoted for control, Accudenz centrifugation, and dialysis treatments. A total of 90 pure line and 85 F1 hybrid chicken hens were each inseminated with 100 x 10(6) sperm at 7-d intervals for 4 to 6 wk. All eggs from the glycerolized control semen treatments were infertile, and fertility rates from dialyzed semen decreased steadily from 26.4 to 0% within the first 4 wk for the pure lines. In contrast, fertility rates for Accudenz-processed semen increased from 17.9 to 37.17% during the first 4 wk. Similar fertility rates occurred with the F1 hybrid cross lines. For turkey fertility trials, the dialysis treatment was not used; glycerolized turkey semen was processed by Accudenz or Percoll centrifugation to reduce glycerol. A total of 36 hens were inseminated with 150 x 10(6) sperm at 7-d intervals for 6 wk. Similar to the chicken trials, fertility rates of Accudenz-processed semen steadily increased to 49.4% by the sixth week of insemination. The average fertility of Percoll-processed semen was only 19.1%. These data demonstrate that Accudenz centrifugation is an acceptable glycerol reduction method for nonfrozen poultry semen.

Animals↗
PubMed2001

Does single versus stepped curing of composite resins affect their shear bond strength?

BACKGROUND: Shrinkage occurs during polymerization of all resin composites, affecting not only the physical properties of the composites but also the marginal integrity of the restoration. Stepped photo-polymerization, in contrast to single-cure photo-polymerization, is said to minimize this shrinkage. The thickness of the increments of resin composite being cured may also be a factor in final shear bond strength. This study compared the shear bond strength resulting from these two curing methods with resin composites of various thicknesses. METHODS: Resin composite in increments of 1.5, 3 and 4.5 mm thickness, cured by stepped or single-cure photo-polymerization, was bonded to human third molar dentin with either the Scotchbond Multipurpose adhesive system or the Singlebond adhesive system. Each experimental group consisted of 12 specimens. After 7 days of storage in water, shear bond strength was tested to failure, and the mode of failure was recorded. RESULTS: The thickness of the resin composite and the method of curing had no significant effect on shear bond strength for bonds mediated by Scotchbond Multipurpose adhesive, but significantly lower shear bond strength was recorded for bonds mediated by Singlebond adhesive for resin composite 4.5 mm in thickness. With thicker resin composite, there was a tendency toward a greater proportion of adhesive cohesive bond failures. CLINICAL SIGNIFICANCE: The stepped photo-polymerization system of curing appears to offer no advantages over single-cure photo-polymerization, except that the former reportedly improves marginal adaptation and reduces marginal leakage. These results suggest that increments of resin composite to be cured by either method should be no thicker than 2 mm, particularly when the bond is mediated by a single-bottle adhesive.

Bisphenol A-Glycidyl Methacrylate↗
PubMed1999

Characterization of the human endothelial nitric-oxide synthase promoter.

Understanding transcription initiation of the endothelial nitric-oxide synthase (eNOS) gene appears pivotal to gaining a comprehensive view of NO biology in the blood vessel wall. The present study therefore focused upon a detailed dissection of the functionally important cis-DNA elements and the multiprotein complexes implicated in the cooperative control of constitutive expression of the human eNOS gene in vascular endothelium. Two tightly clustered cis-regulatory regions were identified in the proximal enhancer of the TATA-less eNOS promoter using deletion analysis and linker-scanning mutagenesis: positive regulatory domains I (-104/-95 relative to transcription initiation) and II (-144/-115). Analysis of trans-factor binding and functional expression studies revealed a surprising degree of cooperativity and complexity. The nucleoprotein complexes that form upon these regions in endothelial cells contained Ets family members, Sp1, variants of Sp3, MAZ, and YY1. Functional domain studies in Drosophila Schneider cells and endothelial cells revealed examples of positive and negative protein-protein cooperativity involving Sp1, variants of Sp3, Ets-1, Elf-1, and MAZ. Therefore, multiprotein complexes are formed on the activator recognition sites within this 50-base pair region of the human eNOS promoter in vascular endothelium.

Animals↗
PubMed1999

Decreased cerebral emboli during distal aortic arch cannulation: a randomized clinical trial.

BACKGROUND: Cerebral emboli occur during cardiopulmonary bypass and are a principal cause of postoperative neurologic dysfunction. We hypothesized that arterial cannulation of the distal aortic arch, with placement of the cannula tip beyond the left subclavian artery, will result in fewer cerebral microemboli than conventional cannulation of the ascending aorta. METHODS: Patients undergoing coronary bypass surgery with a single crossclamp technique were randomized to receive cannulation of the distal aortic arch (n = 17) or standard cannulation of the ascending aorta (control group, n = 17). Trendelenburg positioning was used whenever possible. Cerebral emboli were quantified by continuous transcranial Doppler monitoring of the middle cerebral artery. RESULTS: Baseline demographics were similar for the 2 groups of patients, including cardiopulmonary bypass and crossclamp times. Cerebral microemboli were detected during cardiopulmonary bypass in all patients, with a range of 17 to 627 emboli. The total number of detected emboli was lower in the arch cannulation group (152 +/- 33, mean +/- standard error of the mean) than in the conventional cannulation group (249 +/- 35, P =.04). Embolization rates were lower in distal arch patients than in control patients during cardiopulmonary bypass (2.0 +/- 0.3 vs 4.2 +/- 0.9 per minute, respectively, P =.03). Reduction in cerebral emboli by distal arch cannulation was most pronounced during perfusionist interventions. CONCLUSIONS: Cannulation of the distal aortic arch results in less cerebral microembolism than conventional cannulation of the ascending aorta. Provided it is performed safely, distal arch cannulation may be an important surgical option for patients with severe atherosclerosis of the ascending aorta.

Analysis of Variance↗
PubMed1997

Expression, purification, and characterization of the recombinant NAD-malic enzyme from Ascaris suum.

The cDNA encoding the 65-kDa subunit of malic enzyme from Ascaris suum was cloned into the bacterial expression vector pKK223-3 and overproduced in Escherichia coli. A protein with a subunit molecular mass of 65,000 was expressed at a level of up to 3% of the total soluble protein in JM109, as judged by SDS-PAGE. The enzyme was purified using column chromatography on phenyl-Sepharose followed by orange-A agarose. The purification procedure resulted in a 32-fold purification with an overall yield of 51%. The bacterially expressed enzyme exhibits kinetic constants identical to those measured for native A. suum NAD-malic enzyme.

Animals↗
PubMed1997

Expression of Ascaris suum malic enzyme in a mutant Escherichia coli allows production of succinic acid from glucose.

The malic enzyme gene of Ascaris suum, was cloned into the vector pTRC99a in two forms encoding alternative amino-termini. The resulting plasmids, pMEA1 and pMEA2, were introduced into Escherichia coli NZN111, a strain that is unable to grow fermentatively because of inactivation of the genes encoding pyruvate dissimilation. Induction of pMEA1, which encodes the native animoterminus, gave better overexpression of malic enzyme, approx 12-fold compared to uninduced cells. Under the appropriate culture conditions, expression of malic enzyme allowed the fermentative dissimilation of glucose by NZN111. The major fermentation product formed in induced cultures was succinic acid.

Animals↗
PubMed1993

Cloning and nucleotide sequence of a full-length cDNA encoding Ascaris suum malic enzyme.

The nucleotide sequence of a full-length cDNA encoding NAD(+)-malic enzyme from the parasitic nematode Ascaris suum was determined. The entire sequence of 2269 bases comprises a 5'-leader, a single open reading frame of 1851 bases, and the complete 3'-noncoding region of 340 bases. The first 12 amino acids of the translated sequence are hydrophobic, typical of mitochondrial translocation signals, and do not appear in the purified mature protein. The mature protein contains 605 amino acids and has a molecular mass of 68,478 Da. The amino acid sequences of tryptic peptides from the purified protein and also the N-terminal sequence show excellent correspondence with the translated nucleotide sequence. Comparison of the amino acid sequence of the ascarid protein with the human and rat liver NAD(+)-malic enzymes reveals highly conserved regions interrupted with long stretches of lesser homologous sequences. Structural motifs such as the putative nucleotide binding domains and also the malate binding site are clearly identified by alignment of the three protein sequences.

Amino Acid Sequence↗
PubMed1991

Collagenase activity in recurrent periodontitis: relationship to disease progression and doxycycline therapy.

Previous reports have suggested that active progression of periodontitis may be correlated with increased collagenolytic activity, and that improved clinical conditions after tetracycline treatment may be explained by inhibition of host collagenase. Eighty-two patients with a recent history of periodontal abscesses and/or loss of gingival attachment level (GAL) despite active periodontal therapy were enrolled in a double-blind, randomized, placebo-controlled trial. Clinical measurements, sampling of gingival crevicular fluid (GCF) and subgingival scaling were performed every 2 months. If any site exhibited greater than 2 mm loss of GAL or a periodontal abscess, patients were administered either 100 mg doxycycline per day for 3 weeks or placebo. During 12 months of monitoring, 55 patients exhibited recurrent active disease and were then randomly assigned to either the doxycycline (n = 30) or placebo (n = 25) groups. Analysis of active collagenase and latent collagenase in GCF samples were determined by functional assays and quantitated after SDS-PAGE and fluorography. Collagenase activities were assayed at sites exhibiting active destruction (study site), at sites with pocket depth comparable to the study site but without active destruction, and at healthy sites. Clinical measurements of GAL and collagenase activity were made at intervals between 1 wk and 7 months after completion of the drug regime. Within 7 months, 15 out of 19 patients on placebo exhibited recurrent disease compared to 13 out of 29 patients on doxycycline. Collagenase activity exhibited large variations among patients and was analyzed as presence or absence of active collagenase with a logistic model.(ABSTRACT TRUNCATED AT 250 WORDS)

Analysis of Variance↗
PubMed1990

Randomized controlled trial of doxycycline in prevention of recurrent periodontitis in high-risk patients: antimicrobial activity and collagenase inhibition.

82 patients with a recent history of periodontal abscesses and/or loss of gingival attachment (GAL) despite active periodontal therapy were enrolled in a double-blind, randomized, placebo-controlled trial. Clinical measurements and subgingival scaling were performed every 2 months. If any site exhibited greater than or equal to 2 mm loss of GAL or a periodontal abscess, patients were administered either 100 mg Doxycycline per day for 3 weeks or placebo. During 12 months of monitoring, 55 patients exhibited recurrent active disease and were then randomly assigned to either the Doxycycline or placebo groups. Clinical measurements of GAL and microbiological culture of subgingival bacteria were made at intervals between 1 week and 7 months after completion of the drug regime. Within 7 months, 15 out of 19 patients on placebo exhibited recurrent disease compared to 13 out of 29 patients on Doxycycline, a relative risk reduction of 43% (p less than 0.05) for Doxycycline compared to placebo. Minimal inhibitory concentrations of Doxycycline for subgingival plaque samples from active sites ranged between 25-100 micrograms/ml, which are several fold higher than reported crevicular fluid concentrations for this drug. However gingival crevicular fluid collagenase was inhibited in vitro at concentrations of 5-10 micrograms/ml Doxycycline. These data indicate that Doxycycline provides significant risk reduction of recurrent periodontitis in patients with active disease.

Adult↗
PubMed1988

Phosphofructokinase from Dirofilaria immitis. Stimulation of activity by phosphorylation with cyclic AMP-dependent protein kinase.

Phosphofructokinase has been partially purified from the filariid helminth, Dirofilaria immitis, using ion exchange and affinity chromatography. The D. immitis phosphofructokinase cross-reacted with antibodies prepared against the phosphofructokinase from Ascaris suum. These antibodies had been bound to agarose beads. The enzyme was eluted from the immobilized antigen-antibody complex by denaturing agents, and the subunit molecular weight determined by sodium dodecyl sulfate gel electrophoresis was identical to that of the ascarid enzyme, 90,000. At pH 6.8, substrate saturation curves of the filarial phosphofructokinase with ATP revealed that the enzyme was inhibited by ATP. The fructose-6-P saturation curve was sigmoid at all ATP levels tested. Phosphorylation of the D. immitis phosphofructokinase by the catalytic subunit of beef heart cyclic AMP-dependent protein kinase resulted in incorporation of 0.8 mol of phosphate/mol of subunit and in a 3-4-fold increase in catalytic activity when measured at pH 6.8 at inhibitory levels of ATP. Additional kinetic studies revealed that the phosphorylated enzyme was less susceptible to ATP inhibition than was the nonphosphorylated form. It is proposed that phosphorylation of phosphofructokinase plays an important role in the regulation of carbohydrate metabolism in the filarial as well as the intestinal-dwelling nematodes.

Adenosine Triphosphate↗
PubMed1987

Ascaris suum phosphofructokinase. Phosphorylation by protein kinase and sequence of the phosphopeptide.

Phosphorylation of the ascarid phosphofructokinase with the catalytic subunit of beef heart cyclic AMP-dependent protein kinase results in the incorporation of 1 mol of P/mol of subunit. Accompanying the phosphorylation there is a 3-4-fold increase in catalytic activity when measured at pH 6.8 with inhibitory levels of ATP. Studies on the effect of phosphorylation on the ATP saturation curve demonstrated that phosphorylation decreased the inhibitory action of ATP. The apparent Km of the catalytic subunit for the phosphofructokinase was 11.2 microM. Chymotryptic or subtilisin digestion of the labeled enzyme released distinct but overlapping phosphopeptides that were purified by high pressure liquid chromatography and sequenced by gas phase peptide sequencing. The sequence of the chymotryptic peptide was Ala-Lys-Gly-Arg-Ser-Asp-Ser(P)-Ile-Val-Pro-Thr. Based on these results and earlier observations, it is proposed that phosphorylation of phosphofructokinase plays an important role in the regulation of energy metabolism in the parasitic helminth.

Adenosine Triphosphate↗
PubMed1984

Antibodies to ATLV/HTLV-1 in Africa.

Almost 4000 sera from seven African states were examined for antibodies to ATLV/HTLV-1. Between 1% and 8% of healthy people from sub-Saharan Africa have such antibodies. The highest frequency was observed in Gabon. There were considerable variations between villages. The percentage of seropositives and the mean titre increased with age. Our findings suggest that the African continent is the largest endemic area for ATLV.

Adolescent↗