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R Sharma

Publications and source records attributed to R Sharma.

At least 289 records · Page 16Linked to original sources
PubMed1996

Conformationally constrained analogues of diacylglycerol. 10. Ultrapotent protein kinase C ligands based on a racemic 5-disubstituted tetrahydro-2-furanone template.

5,5-Bis(hydroxymethyl)tetrahydro-2-furanone and its isomer 4,4-bis(hydroxymethyl)tetrahydro-2-furanone were investigated as possible templates for the construction of conformationally constrained analogues of the biologically important second messenger, diacylglycerol (DAG). The former lactone contains embedded within its structure an exact glycerol moiety, while in the latter the ring oxygen has been transposed to the other side of the carbonyl group. All target compounds were synthesized as racemates from 1,3-dihydroxy-2-propanone. The 5,5-bis(hydroxymethyl)tetrahydro-2-furanone proved to be the better template for the construction of DAG surrogates that were demonstrated to have high binding affinities for the biological target, protein kinase C (PK-C). The simplest target compounds derived from this template (3e and 3f) have one of the hydroxyl moieties functionalized either as a myristate or as an oleate ester. The simplest target compound (9c) derived from the ineffective 4,4-bis-(hydroxymethyl)tetrahydro-2-furanone template was investigated only with a myristoyl acyl chain. Reducing the long acyl chain to an acetyl moiety and attaching a compensating lipophilic chain to the lactone ring as an alpha-alkylidene moiety produced compounds 10e and 10f (Z-isomers) and 11e and 11f (E-isomers), which were constructed on the more effective 5,5-bis(hydroxymethyl)tetrahydro-2-furanone template. Targets 14c (Z-isomer) and 15c (E-isomer) were derived, in turn, from 4,4-bis(hydroxymethyl)tetrahydro-2-furanone. The affinities of these ligands for PK-C were assessed in terms of their ability to displace bound [3H-20]phorbol 12,13-dibutyrate (PDBU) from the single isozyme PK-C alpha. The biological data support the hypothesis that the increase in binding affinity for PK-C shown by some of these constrained DAG mimetics appears to be entropic in nature. Two of the designed ligands (10e and 10f) showed the highest affinities (34 and 24 nM, respectively) reported so far for a DAG analogue. Assuming that the interaction between these racemic compounds and PK-C is stereospecific, the potency of the active enantiomer is anticipated to double.

4-Butyrolactone↗
PubMed1996

Conformationally constrained analogues of diacylglycerol. 11. Ultrapotent protein kinase C ligands based on a chiral 5-disubstituted tetrahydro-2-furanone template.

Conformationally constrained analogues of diacylglycerol (DAG) built on a racemic 5(-)[(acyloxy)-methyl]-5-(hydroxymethyl)tetrahydro-2-furanone template were shown previously to have excellent binding affinities for protein kinase C (PK-C). Since the interaction of PK-C with DAG is stereospecific, it was anticipated that PK-C would bind tightly to only one enantiomeric form of the compounds constructed with this new lactone template. Separation of enantiomers by chiral HPLC was discarded due to the ease with which acyl migration occurs in these class of compounds, and a total chiral synthesis was undertaken. Prior to chemical synthesis, the selection of the "correct" enantiomeric template was predicted by a molecular conformational analysis that compared the two enantiomers of DAG in their presumed "active" conformation with the two enantiomeric lactone templates. This presumed "active" conformation for DAG was derived from a previously developed pharmacophore model that uses the molecule of a potent phorbol diester as the ideal rigid template. The results from this analysis indicated that the "correct" lactone template corresponded to the inactive (R)-isomer of DAG. This analysis also predicted that the lactone template corresponding to the active (S)-DAG enantiomer would not fit adequately into the pharmacophore. The chiral syntheses of target compounds 2, 4, and 6, constructed on the selected, and presumably "correct" lactone template, were achieved from a common bicyclic intermediate (5R,8R,9R)-8,9-O-isopropylidene-2-keto-1,7-dioxaspiro[4.4]nonane (10) that was synthesized from commercially available 1,2:3,5-di-O-isopropylidene-alpha-D-threo-apiofuranose (7) by a very effective spirolactonization approach. On the basis of their ability to inhibit the binding of [3H-20]phorbol 12,13-dibutyrate (PDBU) to PK-C alpha, the enantiomeric ligands 2, 4, and 6 were twice as potent as the corresponding racemates. These results confirm that binding of these lactones is stereospecific and consistent with a binding mechanism similar to that of DAG.

4-Butyrolactone↗
PubMed1996

Conformationally constrained analogues of diacylglycerol. 12. Ultrapotent protein kinase C ligands based on a chiral 4,4-disubstituted heptono-1,4-lactone template.

Conformationally constrained analogues of diacylglycerol (DAG) built on a 5(-)[(acyloxy)methyl]-5-(hydroxymethyl)tetrahydro-2-furanone template (1, Chart 1) were shown previously to bind tightly to protein kinase C alpha (PK-C alpha) in a stereospecific manner. These compounds, however, racemized readily through rapid acyl migration and lost biological potency. In order to circumvent this problem, the "reversed ester" analogues were designed as a new set of PK-C ligands. This reversal of the ester function produced some new DAG mimetics that are embedded in a C-4 doubly-branched heptono-1,4-lactone template. The reversed ester analogues were impervious to racemization, and their chemically distinct branches facilitated the enantiospecific syntheses of all targets. Compound 2, the simplest reversed ester analogue of 1 (Chart 1), exhibited a 3.5-fold reduction in binding affinity toward PK-C alpha which we attributed to the loss of a stabilizing gauche interaction that caused the ester branch in 2 to be more disordered than in the normal ester 1. However, conversion of the propanoyl branch of 2 into a propenoyl branch restored binding affinity (3 versus 5). As expected, the compounds bound to the enzyme with strict enantioselectivity (3 and 5 versus 4 and 6). Functionalization of the propenoyl-branched compounds as alpha-alkylidene lactones, in a manner which proved successful with the 5(-)[(acyloxy)methyl]-5-(hydroxymethyl)tetrahydro-2-furanone template (9 and 10), produced stable compounds with equivalent ultrapotent binding affinities for PK-C alpha (7 and 8). The additional incorporation of the propenoyl-branched carbonyl into a gamma-lactone ring was performed (11-14) not only to derive a possible additional entropic advantage but also to confirm the spatial disposition of this carbonyl function in the ligand-enzyme complex. Although no additional entropic advantage was derived, the high binding affinities displayed by compounds 11 and 12 helped to establish the correct orientation of the equivalent carbonyl group in PK-C-bound DAG. As expected, these DAG analogues activated PK-C alpha. The most potent agonist, compound 8, stimulated phosphorylation of the alpha-pseudosubstrate peptide, and in primary mouse keratinocytes it caused inhibition of binding of epidermal growth factor with an ED50 of approximately 1 microM. In contrast to the phorbol esters, compound 8 did not induce acute edema or hyperplasia in skin of CD-1 mice, and its pattern of downregulation with several PK-C isozymes was different from that of phorbol 12-myristate 13-acetate (PMA).

4-Butyrolactone↗
PubMed1996

Lectin binding reveals divergent carbohydrate expression in human and mouse Peyer's patches.

The nature of cell-associated carbohydrates in the human intestine that may mediate transepithelial transport of bacterial and dietary lectins and their processing by the lymphoid cells of Peyer's patches is not known. Because the cell surface carbohydrate receptors for lectins may vary in different species, the glycoconjugates of human and mouse follicle-associated epithelium and gut-associated lymphoid tissue were compared. A panel of 27, mainly recently isolated, lectins were used to identify glycoconjugate expression in M-cells, enterocytes, goblet cells, lymphocytes and macrophages in mouse and human intestine. Mouse M-cells were exclusively labelled by fucose-specific lectins but in human follicle-associated epithelium no distinct M-cell staining pattern was observed. In the human Peyer's patches, Bryonia dioica lectin bound selectively to paracortical T-lymphocytes and Chelidonium majus lectin to germinal centre B-cells. Certain mannose-specific lectins (Galanthus nivalis, Hippeastrum hybrid) stained the tingible body macrophages in the germinal centre of human Peyer's patches but labelled the macrophages in the paracortical T-cell region of the mouse. The results indicate distinct differences in glycosylation between mouse and human Peyer's patches and their associated lymphoid cells. When considering cell surface glycoconjugates as target molecules for the gut immune system, care has to be taken to choose the appropriate lectin for each species.

Animals↗
PubMed1996

Surgical experience with total correction of tetralogy of Fallot in infancy.

Fifty two patients less than one year old with tetralogy of Fallot underwent primary repair between January 1991 and December 1994. Age range was three to twelve months (mean 10.09 +/- 2.01 months) and body weight ranged from 4.5 to 9 kg (mean 8.38 +/- 2.79 kg). Transatrial-transpulmonary repair was performed in 36 patients and the classical transventricular approach was used in 16 patients. Six patients underwent emergency surgery for severe cyanosis and spells. Five patients had left pulmonary artery plasty for pulmonary artery bifurcation stenosis and two out of the five patients who had anomalous coronary arteries needed a right ventricle to pulmonary artery conduit. Mean post repair peak right ventricular/systemic pressure ratio was 0.74 +/- 0.18 in the transventricular group and 0.71 +/- 0.26 in the transatrial-transpulmonary group. There were three hospital deaths. Follow-up ranged from 3 to 46 months (mean 21.18 months). Forty patients underwent echocardiography and twenty patients underwent cardiac catheterisation six to eighteen months after surgery. Mean right ventricular outflow tract gradient on echocardiography was 20.35 +/- 10.12 and, at cardiac catheterisation, 17.51 +/- 13.49 mmHg with mean post repair peak right ventricle/left ventricle pressure ratio of 0.44 +/- 0.11. These were significantly less than the values obtained in the operating room. Only one patient had residual ventricular septal defect with left to right shunt of 1.6:1 at cardiac recatheterisation. There was one late death after reoperation for residual obstruction. Encouraging results with primary repair of tetralogy of Fallot in infancy prompt us to continue this policy in suitable cases.

Body Weight↗
PubMed1996

Proton MRS in Pott's spine--a case report.

Proton Magnetic Resonance Spectroscopy was carried out at 1.5 T on a patient with histologically proven Pott's spine affecting D11 vertebral body. In addition to the significantly reduced signals from the lipids in the region between 1 and 2 ppm, a prominent resonance at 5.1 ppm is seen. The spectrum is very different from that recorded on the one hand for a normal spine and, on the other, for a tumor arising from the vertebral body of a patient.

Adult↗
PubMed1996

The application of a multirod collimator in fast neutron therapy.

The application of a novel multirod collimator in fast neutron therapy to produce irregularly shaped fields with partial transmission and fully blocked areas is discussed. The transmission through the tungsten rods is measured in free space in broad beam geometry. A model based on scatter analysis is applied in calculating the thickness of the rods required to produce partially blocked areas. The effect of a full beam block is also measured. Measurements have been made in a water phantom to confirm the computational model. The good agreement between the measured and calculated values demonstrates that the scatter model can be safely applied to fast neutron beam dose computations.

Cyclotrons↗
PubMed1996

Matrix metalloproteinase (stromelysin-1) increases the albumin permeability of isolated rat glomeruli.

Matrix metalloproteinases (MMPs) secreted by connective tissue cells are capable of acting on extracellular matrix components of glomerular basement membrane at a slow rate and thus may play a role in the control of protein permeability and in the progression of certain kinds of glomerulonephritis. We have used an in vitro assay to measure the direct effect of three MMPs and human neutrophil elastase on glomerular albumin permeability (Palbumin). Glomeruli were isolated from normal male Sprague-Dawley rats and suspended in isolation medium with or without interstitial collagenase, gelatinase-A, stromelysin-1, or elastase and were incubated at 37 degrees C for up to 4 hours. A tissue-specific inhibitor of matrix metalloproteinases (TIMP-1) and a plasma proteinase inhibitor, alpha2-macroglobulin (alpha2M), were used to block the activity of MMPs. Palbumin was calculated from the change in glomerular volume in response to an applied oncotic gradient. In this study stromelysin-1 (10 microg/ml) and elastase (5 microg/ml) increased Palbumin significantly. Stromelysin-1 increased Palbumin after 4 hours, whereas elastase had an effect after 2 hours. Lower concentrations of stromelysin-1 or shorter incubation time had no effect on Palbumin. Incubation for up to 4 hours with interstitial collagenase (10 microg/ml) or gelatinase-A (10 microg/ml) had no effect on Palbumin. Coincubation with TIMP-1 and alpha2M blocked the stromelysin-1-mediated increase in Palbumin. We conclude that stromelysin-1 is capable of affecting the glomerular filtration barrier directly and that it may play an important role in causing proteinuria in glomerular diseases.

Albumins↗
PubMed1996

Melorheostosis of the hand.

We report a case of melorheostosis of the hand which showed a sclerotomal pattern of involvement, with swelling and pain. Surgical debulking resulted in correction of the deformity and relief of pain.

Adult↗
PubMed1996

Hepatitis B virus infection in pregnant women and its transmission to infants.

HBsAg was screened by Reverse Passive Haemagglutination Test (RPHA) and was confirmed by ELISA test in 157 pregnant females and their newborns. Anti-HBc and IgM anti-HBc was done in these cases by enzyme immuno-assay. The overall prevalence of HBsAg in mothers was 16 out of 157 (10 per cent) and in cord blood of newborns 5 per cent. The transplacental transmission was found in eight of 16 (50 per cent) HBsAg positive mothers. Anti-HBc was present in 12 out of 16 (75 per cent) HBsAg positive mothers and, of these, seven (58 per cent) neonates acquired HBsAg infection. IgM anti-HBc was present in seven out of eight (88 per cent) HBsAg positive neonates, suggesting active in utero infection. Fourteen out of 16 (88 per cent) neonates born to HBsAg positive mothers were alive and healthy, one was stillborn and one had a congenital anomaly.

Adult↗
PubMed1996

In situ stimulation of topoisomerase II-induced cleavage sites in the c-myc protooncogene by antitumor agent pMC540 is associated with gene expression.

The antitumor activity of pMC540 has been shown to be mediated via its interaction with topoisomerase (Topo) II eventually leading cells into apoptosis. This agent was also found to down regulate the expression of the c-myc oncogene in L1210 leukemia cells. To investigate the possibility that damage within select genomic regions may contribute to the antiproliferative activity of pMC540, differential damage in regions surrounding the c-myc locus as well as other select genes was determined. Southern blot hybridization experiments show that pMC540 treatment induces in situ DNA cleavage products in the 5' end of the c-myc oncogene of L1210 leukemia cells. In cells pre-treated with 50 microM ethidium bromide, an inhibitor of the Topo II-dependent DNA cleavage, a subsequent treatment with pMC540 failed to induce DNA cleavage, suggesting that the cleavage activity of pMC540 was Topo II dependent. pMC540-induced cleavage does not appear to correlate with the over-expression of the c-myc oncogene in these cells as another over-expressed gene c-myb was not affected. Thus, it is proposed that the c-myc gene may be a preferred target for pMC540 may mediated antiproliferative activity.

Animals↗
PubMed1996

Radiolabeled denatured RBC scintigraphy in autologous splenic transplantation.

An 18-year-old man underwent surgery for blunt abdominal trauma requiring a total splenectomy. Heterotopic splenic autotransplantation was done in the omentum. Immediate platelets counts, cell morphology, and serum immunoglobulin levels were suggestive of impaired splenic function. Radiolabeled denatured RBC splenic imaging showed evidence of functioning autotransplanted splenic tissue in the omentum. The blood parameters were found to be normal 4 weeks after transplant. Radionuclide denatured RBC imaging has been found to be useful in the localization of functional autotransplanted splenic tissue.

Abdominal Injuries↗
PubMed1996

Variations in the fluorescence intensity of intact DAPI-stained bacteria and their implications for rapid bacterial quantification.

As current techniques for the quantification of bacteria are laborious and often imprecise, instrumental approaches such as sedimentation field-flow fractionation (SdFFF) are attractive. In this technique, fluorogenic dyes specific for nucleic acids are used to identify bacterial cells. Bacterial biomass can be quantified directly with SdFFF if the specific fluorescence of bacterial cells is constant. The effect of different growth conditions on the specific fluorescence of one strain each of Escherichia coli, Pseudomonas aeruginosa, Proteus mirabilis and Staphylococcus epidermidis stained with 4',6-diamidino-2-phenylindole was examined. Specific fluorescence varied over a 500-fold range, from 0.22 to 103 arbitrary fluorescence units per cell. Specific fluorescence was highest when cells were in log phase, and lowest when cells were in stationary phase. Specific fluorescence decreased when cells harvested in log phase were starved for 7 d in a carbon-free minimal medium, and increased rapidly (within 2 h) after cells were relieved from carbon limitation. Such variations in specific fluorescence must be considered when using gross fluorescence as a direct indicator of bacterial numbers in the SdFFF technique for quantifying bacterial biomass. Moreover, they have serious implications for the application of fluorescence techniques in other instrumental approaches for bacterial enumeration in environmental samples.

Bacteria↗