Search PubMed⌕ Search

Biomedical subjects

K K Sharma

Publications and source records attributed to K K Sharma.

At least 19 recordsLinked to original sources
PubMed1998

Identification of 1,1'-bi(4-anilino)naphthalene-5,5'-disulfonic acid binding sequences in alpha-crystallin.

The hydrophobic binding sites in alpha-crystallin were evaluated using fluorescent probes 1,1'-bi(4-anilino)naphthalenesulfonic acid (bis-ANS), 8-anilino-1-naphthalene sulfonate (ANS), and 1-azidonaphthalene 5-sulfonate (1,5-AZNS). The photolysis of bis-ANS-alpha-crystallin complex resulted in incorporation of the probe to both alphaA- and alphaB-subunits. Prior binding of denatured alcohol dehydrogenase to alpha-crystallin significantly decreased the photoincorporation of bis-ANS to alpha-crystallin. Localization of bis-ANS incorporated into alphaA-crystallin resulted in the identification of residues QSLFR and HFSPEDLTVK as the fluorophore binding regions. In alphaB-crystallin, sequences DRFSVNLNVK and VLGDVIEVHGK were found to be the bis-ANS binding regions. Of the bis-ANS binding sequences, HFSPEDLTVK of alphaA-crystallin and DRFSVNLNVK and VLGDVIEVHGK of alphaB-crystallin were earlier identified as part of the sequences involved in their interaction with target proteins during the molecular chaperone-like action. The hydrophobic probe, 1,5-AZNS, also interacted with both subunits of alpha-crystallin. Localization of 1,5-AZNS binding site in alphaB-crystallin lead to the identification of HFSPEEK sequence as the interacting site in this subunit of alpha-crystallin. Glycated alpha-crystallin displayed decreased ANS fluorescence and loss of chaperone-like function, suggesting the involvement of glycation site as well as ANS binding site in chaperone-like activity display.

Amino Acid Sequence↗
PubMed1998

Characterization of trypsin-modified bovine lens acylpeptide hydrolase.

Acylpeptide hydrolase, which removes the N-acetylated amino acids from peptide substrates was purified from bovine lens, truncated in vitro to a 55 kDa enzyme by trypsin digestion and characterized. The activity of the trypsin-modified enzyme was investigated using alpha A-crystallin and oxidized insulin A chain. The trypsin-modified enzyme was able to unblock alpha A-crystallin and displayed endoprotease activity unlike the native enzyme. SDS-PAGE analysis and amino acid sequencing of (3H)iPr2P-F labeled bovine lens acylpeptide hydrolase showed that the lens has a 55 kDa truncated form of the enzyme. The in vivo truncated form of the enzyme was generated by the cleavage of the Gly203-Asp204 peptide bond in the native enzyme.

Amino Acid Sequence↗
PubMed1998

Interaction of 1,1'-bi(4-anilino)naphthalene-5,5'-disulfonic acid with alpha-crystallin.

The hydrophobic sites in alpha-crystallin were evaluated using a fluorescent probe 1,1'-bi(4-anilino)naphthalenesulfonic acid (bis-ANS). Approximately one binding site/subunit of alpha-crystallin at 25 degrees C was estimated by equilibrium binding and Scatchard analysis (Kd = 1.1 microM). Based on fluorescence titration, the dissociation constant was 0.95 microM. The number of bis-ANS binding sites nearly doubled upon heat treatment of the protein at 60 degrees C. Likewise, the exposure of alpha-crystallin to 2-3 M urea resulted in increased binding of bis-ANS. Above 3 M urea there was a rapid loss in the fluorescence indicating the loss of interaction between bis-ANS and protein. The alpha-crystallin refolded from 6 M urea showed tryptophan fluorescence emission similar to the native alpha-crystallin. However, the refolded alpha-crystallin showed a 60% increase in bis-ANS binding, suggesting distinct changes on the protein surface resulting from exposure to urea similar to the changes occurring due to heat treatment. The fluorescence of tryptophan in native alpha-crystallin was quenched by the addition of bis-ANS. The quenching was inversely related to the amount of bis-ANS bound to alpha-crystallin. Additionally, the binding of bis-ANS reduced the chaperone-like activity of the protein. Photolysis of bis-ANS-alpha-crystallin complex resulted in incorporation of the probe to both A- and B-subunits, indicating that both subunits in native alpha-crystallin contribute to the surface hydrophobicity of the protein.

Anilino Naphthalenesulfonates↗
PubMed1998

Real-time intraoperative transesophageal echocardiography--how useful? Experience of 5,016 cases.

OBJECTIVE: To evaluate transesophageal echocardiography (TEE) as an intraoperative monitoring modality and to assess its safety, reliability, and overall utility in real-time use during cardiac surgery. DESIGN: Prospective, observational cohort study performed from January 1993 to June 1997. SETTING: Operating room of a tertiary care hospital for cardiology and cardiovascular surgery. PARTICIPANTS: Five thousand and sixteen adult patients with acquired heart disease, who underwent 1,356 valve procedures and 3,660 coronary artery bypass graftings (CABGs). INTERVENTIONS: All patients were monitored with radial artery and pulmonary artery catheters, along with continuous TEE monitoring with a multiplane transducer. MEASUREMENTS AND MAIN RESULTS: Prebypass imaging yielded unsuspected findings that either helped or modified the surgical plan in 158 of 1,356 valve procedures (11.65%) and in 993 of 3,660 CABGs (27.13%). There were 3,217 TEE-guided hemodynamic interventions in 944 patients (25.79%) in the CABG group and 629 in 142 patients (10.47%) in the valve group. TEE was the sole guiding factor in initiating therapy in 23.53% of events, whereas it was supportive to other monitoring modalities in 76.46% of events. Postbypass TEE identified the need for graft revision in 29 patients (0.8%), intra-aortic balloon pump (IABP) requirement in 29 patients (0.8%), and inadequate valve repair in 28 patients (2.08%). For the entire series, 38.78% of patients benefited from prebypass and 39.16% from postbypass use of TEE. There were no complications attributable to the use of TEE in the entire series. There was 87% concordance between online interpretation by a trained anesthesiologist and offline analysis by a cardiologist. CONCLUSION: Intraoperative TEE is useful in formulating the surgical plan, guiding various hemodynamic interventions, and assessing the immediate results of surgery. It is safe and the results are reliable in the hands of trained anesthesiologists.

Cardiac Surgical Procedures↗
PubMed1998

Malingering uncommon psychiatric symptoms among defendants charged under California's "three strikes and you're out" law.

This paper describes an epidemic of uncommon psychiatric symptoms among nine criminal defendants charged under California's new "Three Strikes and You're Out" law. The defendants were facing a minimum sentence of 25 years to life in prison. The defendants exhibited the following uncommon psychiatric symptoms: coprophagia (eating feces), eating cockroaches and many reported seeing little green men. The defendants, all of whom we believe were malingering, were evaluated by the authors for competency to stand trial. Thus far, eight of the nine defendants were found competent to stand trial; only one defendant was found incompetent to stand trial. The authors created a database which included information on the defendants from court documents and from our interview with the defendants. We summarized the data and outcomes of the case. Also included is a brief review of the "Three Strikes Law" and a paradigm for how we ruled out relevant psychiatric diagnoses before we arrived at our opinion of malingering.

Adult↗
PubMed1998

Forensic implications and medical-legal dilemmas of maternal versus fetal rights.

The purpose of this paper is to review the issue of fetal rights from primarily a legal perspective, with consideration of morals and professional ethics. The practice of medicine is fraught with numerous bioethical dilemmas. These dilemmas often leave the physician wondering if he has made the correct decision. A physician's morals and professional ethics may influence his or her decision in resolving bioethical dilemmas. The case example is a 34-year-old female with a 41-week intra-uterine pregnancy. The mother was refusing induction of labor. Without the labor induction, the fetus may die. Despite this risk, the mother desired to pursue a vaginal delivery. The AMA's ethics state that a competent, pregnant mother's wishes should prevail and the court should not be involved unless there are unusual circumstances. The mother in the case example was competent and informed consent was provided. Case law does not specifically address the dilemma of the case example. However, there is case law regarding court-ordered cesarean sections which reveals different opinions. The difference in court opinion encompasses the relative degree of weight given to the fetus's right to be born healthy and alive versus the mother's privacy rights. Some courts describe this "balancing test," whereas others state that the mother's privacy rights prevail unless there are exceptional circumstances, which will be extremely rare. The fetus has acquired rights in other areas of the law; for example, abolishment of the intra-family immunity doctrine and the definition of murder in most states. In considering the legal arena of fetal versus maternal rights, a decision tree is presented to assist physicians in assessing cases of a pregnant mother refusing medical treatment. There is no precise demarcation in assessing fetal and maternal rights. The greater the degree of fetal viability, the greater degree of fetal rights. Consideration must also be given to the relative degree of invasiveness to the mother for the proposed procedure; the more invasive, the greater degree of maternal rights. Each case must be evaluated on an individual basis and the decision tree can assist a clinician with this process.

Adult↗
PubMed1998

Tramadol, a centrally acting opioid: anticonvulsant effect against maximal electroshock seizure in mice.

The present study was designed to investigate the pro- or anticonvulsant effect of tramadol using maximal electroshock (MES) test. An attempt was also made to determine the possible opioid receptor mechanism involved. MES seizures were induced through transauricular electrodes (60 mA, 0.2s) and the seizure severity was assessed by the duration of tonic hindlimb extensor phase. Intraperitoneal (i.p.) administration of tramadol resulted in a dose-dependent anticonvulsant action; the ED50 for the effect was 33 mg/kg. The anti-MES effect of tramadol was antagonized by the low doses (0.05 and 0.1 mg/kg, s.c.) of MR 2266, a selective kappa receptor antagonist and also by the high doses (1.0 and 5.0 mg/kg, i.p.) but not the low doses (0.1 and 0.25 mg/kg) of naloxone. The results suggest that the anti-MES effect of tramadol is mediated by kappa receptors, since MR 2266 and naloxone (in high doses) are known to block these receptors.

Animals↗
PubMed1997

Functional elements in molecular chaperone alpha-crystallin: identification of binding sites in alpha B-crystallin.

alpha-Crystallin, the predominant eye lens protein with sequence homology to small heat shock proteins, acts like a molecular chaperone by suppressing the aggregation of damaged crystallins and proteins. To gain an insight into the amino acid sequences in alpha-crystallin involved in chaperone-like function, we used a cleavable, fluorescent, photoactive, crosslinking agent, sulfosuccinimidyl-2 (7-azido-4-methylcoumarin-3-acetamido)-ethyl-1,3' dithiopropionate (SAED), to derivatize yeast alcohol dehydrogenase (ADH) and allowed it to complex with bovine alpha-crystallin at 48 degrees C. The complex was photolyzed and reduced with DTT and the subunits of alpha-crystallin, alpha A- and alpha B-, were separated. Fluorescence analysis showed that both alpha A- and alpha B-crystallins interacted with ADH during chaperone-like function. Tryptic digestion, amino acid sequencing, and mass spectral analysis of alpha B-crystallin revealed that APSWIDTGLSEMR (57-69) and VLGDVIEVHGKHEER (93-107) sequences were involved in binding with ADH.

Amino Acid Sequence↗
PubMed1997

Acquired gastric outlet obstruction during infancy and childhood: a report of five unusual cases.

Infantile hypertrophic pyloric stenosis is a common, specific, and well-understood entity in the pediatric age group. Prepyloric webs and diaphragms are the other rare causes of gastric outlet obstruction followed by still rarer causes, eg, pyloric stenosis secondary to acid peptic disease. The present report describes a newer, nonspecific, clinical condition of pyloric obstruction in five patients of varying age who responded very well to Henieke-Mikulicz' pyloroplasty. The authors have suggested some modifications in the classification of gastric outlet obstruction in children to include these patients.

Child↗
PubMed1997

Forensic analysis and psycholegal implications of parricide and attempted parricide.

In a retrospective archive study, 64 adjudicated adult cases involving the murder or attempted murder of at least one parent, referred for forensic evaluations are described. Biographic, demographic, diagnostic, crime scene, psycholoegal opinion, and disposition data are presented. Results indicated a 40% rate of insanity acquitees. Attempted parricide subjects were more likely to have inpatient psychiatric histories, witnesses present during the criminal act, nonresponsiveness towards their actions, competency raised, and a hospital disposition. Gender and ethnicity were found to have a significant effect on ultimate disposition. Fifty-four percent of the sample opined psychotic were sentenced to prison, suggesting other factors considered by judge and jury. Profile characteristics and typologies are presented. The findings are compared to studies involving parricide and legal strategies involving similar cases.

Adolescent↗
PubMed1996

Identification of new lens protease(s) using peptide substrates having in vivo cleavage sites.

Bovine lens extracts were incubated in pH 7.5 buffer with succinylated VSREEKPSSAPSS, SGVDAGHS, GKPTSAPSS and GKHNERQD, the peptides having age-dependent in vivo cleavage sites in bovine and human alpha A-crystallin. The reaction products were analyzed to identify the initial protease cleavage sites. The results showed that bovine lens extracts contain protease activity that can cleave. Thr-Ser, Ser-Ala, Ser-Ser bonds in test substrates which correspond to peptide bonds Ser168-Ser169, Ser169-Ala170, Ser172-Ser173 in bovine alpha A-crystallin and Thr-Ser, Ser169-Ala170, Ser172-Ser173 in human alpha A-crystallin. In addition, the same extracts were also capable of hydrolyzing Asp-Ala and Asn-Glu bonds corresponding to Asp151-Ala152 and Asn101-Glu102 in alpha A-crystallin from bovine and human lenses. The cleavage specificity of the newly discovered protease(s) suggests that the in vivo truncation of alpha A-crystallin reported earlier may be due to the action of proteases. The newly discovered lens protease(s) were resistant to inactivation by E-64 and DFP. However, prior treatment of the lens extracts with leupeptin and chymostatin resulted in partial loss of Asn-Glu hydrolytic activity. N-ethylmaleimide treatment completely abolished in the Asn-Glu hydrolyzing activity.

Amino Acid Sequence↗
PubMed1996

Peptide hydrolysis in lens: role of leucine aminopeptidase, aminopeptidase III, prolyloligopeptidase and acylpeptidehydrolase.

The distribution of leucine aminopeptidase, aminopeptidase III, prolyloligopeptidase and acylpeptidehydrolase activities in different regions of a bovine lens was determined and correlated with the distribution of crystallin fragments (measured as < 18 kDa protein) and water-insoluble proteins in the same lens. A gradient of activity was observed for all the peptidases tested, with the highest specific activity present in the cortical fibers which decreased to one half or below in the inner cortical fibers and nucleus. An inverse correlation between peptidase activities and the amount of crystallin fragments was observed in different regions of the lens. However, a direct correlation between the water-insoluble protein content and the crystallin fragments was observed in all fibers of the same lens. The amount of crystallin fragments and the amount of water-insoluble proteins increased from 2.7% and 8% in the outer cortical fibers to 13% and 68% in the nucleus of the same lens. The water-insoluble fraction from both cortical and nuclear fibers however displayed 4-5 fold more crystallin fragments compared to that present in the water-soluble fraction of the same preparation. When the bovine lens cortical and nuclear extracts were tested for their ability to hydrolyze the peptide substrate, Ile-Ser-bradykinin, the cortical extract was found to be at least ten times superior to the nuclear extract. Prior inactivation of prolyloligopeptidase and other serine proteases by diisopropylfluorophosphate however diminished the ability of the cortical extract to hydrolyze peptide substrates. Bovine lens cortical extract was able to completely hydrolyze alpha-melanocyte stimulating hormone as well as N-Acetyl-Met-Asp-Arg-Val-Leu-Ser-Arg-Tyr showing the presence of active acylpeptidehydrolase facilitating the complete hydrolysis of N-terminally blocked peptides. The human lens extract was found to contain both diisopropylfluorophosphate sensitive and resistant enzymes capable of hydrolyzing peptide substrates.

Amino Acid Sequence↗
PubMed1996

Comparison of leucine aminopeptidase and aminopeptidase III activities in lens.

PURPOSE: To evaluate the relative contribution of leucine aminopeptidase and aminopeptidase III activities to the total aminopeptidase activity in bovine and human lenses under in vivo pH conditions. METHODS: Bovine and human lens extracts were fractionated on a Sephadex G-200 column at pH 6.9 and 8.5 and all the fractions were assayed with Leu-pNA and Arg-pNA as substrates at in vivo lens pH (6.9) and optimum pH for leucine aminopeptidase, (8. 5). The major peptidases were purified and their activities compared with that of LAP and AP III isolated from bovine lens. The ability of bovine and human lens extracts and purified bovine lens LAP and AP III to hydrolyze various peptide bonds in synthetic peptides, VHLPTVEK, bradykinin and Ile-Ser-bradykinin was determined by amino acid analysis of the reaction products. RESULTS: Sephadex G-200 gel chromatography and assay of all the fractions at pH 6.9 showed that the elution volume for the predominant aminopeptidase present in bovine lens extract is the same as that of purified AP III from the same lenses. However, when the assays were done at pH 8.5, the major activity eluting from the Sephadex G-200 column was found in fractions having LAP. A similar study of human lens extracts at pH 6. 9 and 8.5 showed one major peak with elution volume corresponding to that of purified bovine lens AP III: The human lens extracts displayed a very low level of LAP activity. The hydrolysis pattern of peptide substrates by AP III paralleled that of bovine and human lens extract at pH 6.9. The X-Pro bond resistant to LAP in peptide substrate, VHLTPVEK was hydrolyzed by AP III as well as lens extracts. CONCLUSION: Both bovine and human lenses have very low LAP activity compared to AP III activity at in vivo pH 6.9. AP III, by its higher activity, broad specificity and its ability to cleave peptide bonds that are resistant to LAP, is likely to play a major role in lens during epithelial cell differentiation into fiber cells and complete hydrolysis of peptides generated in vivo.

Amino Acid Sequence↗
PubMed1995

Effect of cross-linking on the chaperone-like function of alpha crystallin.

Alpha crystallin can function as a molecular chaperone in suppressing the heat-induced aggregation of other crystallins and proteins. During cataractogenesis, alpha-crystallin becomes a water-insoluble, high-molecular-weight, cross-linked aggregate. To determine whether the chaperone activity of alpha crystallin is lost during this age-related modification, extracts were prepared by sonication of water-insoluble proteins isolated from aged bovine lenses and human cataract lenses. All the preparations were tested for chaperone-like activity using beta L-crystallin as the target protein and the percentage of alpha-crystallin in water-insoluble sonicated supernatant (WISS) was determined by slot blot immunoassay. The WISS from bovine as well as human lenses were still effective in protecting beta L-crystallin aggregation at 56 degrees C. The bovine cortical WISS with 50% immunoreactive alpha-crystallin showed 62% of the chaperone-like activity displayed by native alpha-crystallin. The WISS from bovine lens nucleus and human lenses with 17% and 5% immunoreactive alpha-crystallin showed 19% and 4% chaperone-like activity compared to native alpha-crystallin. Prior treatment of the WISS of both bovine and human lenses with dithiothreitol resulted in nearly 50% increase in chaperone-like activity suggesting possible loss of chaperone-like activity due to disulfide cross-links. To see if the chaperone-like activity of alpha-crystallin can be altered by non-disulfide cross-linking, native alpha-crystallin isolated from bovine lenses was cross-linked with dimethylsuberimidate (DMS) and dimethyl 3,3'-dithiobispropionimidate (DTBP) and tested for chaperone-like activity. The DMS cross-linked alpha-crystallin was effective in inhibiting the aggregation of beta L-crystallins at 56 degrees C, but required a two- to five-fold higher concentration than the native alpha-crystallin. alpha-Crystallin with higher degree of cross-linking showed lower chaperone-like activity. alpha-Crystallin cross-linked with DTBP, a cleavable cross-linking agent, also showed a 80% loss in chaperone-like activity. However, when the DTBP cross-linked alpha-crystallin was treated with dithiothreitol to cleave the cross-links there was a 50% recovery in the chaperone-like activity. These data suggest that the age-related cross-linking, which restricts the molecular flexibility of alpha-crystallin decreases its chaperone-like function.

Aged↗
PubMed1995

A double-blind randomized placebo cross-over controlled trial using the antioxidant vitamin E to treat reactive oxygen species associated male infertility.

OBJECTIVE: To determine the effectiveness of the in vivo administration of vitamin E as treatment for reactive oxygen species-associated male infertility. SETTING: University-based center for reproductive medicine. DESIGN: Double-blind randomized placebo cross-over controlled trial. PATIENTS, PARTICIPANTS: Thirty healthy men with high levels of reactive oxygen species generation in semen and a normal female partner. INTERVENTIONS: Patients were allocated to two groups according to the blinded randomization. Each patient received either 600 mg/d of vitamin E (Ephynal, 300 mg tablets; F. Hoffman-La Roche Ltd., Basle, Switzerland) (order A) or identical placebo tablets (order B) for 3 months. Then after a 1-month wash-out period the patients were crossed-over to the other treatment. MAIN OUTCOME MEASURES: Improvement in the in vitro function of the spermatozoa measured by conventional semen analysis, computerized motility assessment, determination of reactive oxygen species generation, binding to the zona pellucida of the unfertilized human oocyte in a competitive zona binding assay, development of hyperactivated motility (both spontaneous and in the presence of 20% of the natural agonist, human follicular fluid) and pregnancy. RESULTS: Rise in the blood serum vitamin E levels after treatment accompanied by improvement in one of the sperm function tests: the zona binding assay. The zona binding ratio for order A improved from 0.2 (range 0 to 0.5) before treatment to 0.5 (range 0.1 to 1.0) after treatment, the corresponding values for order B were 0.2 (range 0 to 1.0) before treatment and 0.3 (range 0.1 to 0.7) after treatment. CONCLUSION: Oral administration of vitamin E significantly improves the in vitro function of human spermatozoa as assessed by the zona binding test.

Adult↗
PubMed1995

Oral steroid therapy for subfertile males with antisperm antibodies in the semen: prediction of the responders.

This study was performed to examine the effectiveness of steroid therapy in subfertile men with antisperm antibodies and infertility lasting > 1 year, to predict those who would respond positively, and to evaluate the effect of the therapy on semen parameters and antisperm antibodies. The patients included 48 subfertile couples in whom the male partner had > or = 20% motile spermatozoa with bound antibodies of immunoglobulin (Ig)G, IgA or a combination of both, and were treated with prednisolone, 40 mg a day, for the first 10 days, then 5 mg on days 11 and 12 of the partner's cycle for 9 months. Twelve couples became pregnant; a cumulative conception rate of 30.2% was achieved at 9 months. The pregnant group started with significantly higher concentrations of IgG (tail) and grade I motility (P = 0.03 and P = 0.02 respectively). Multi-covariate discrete logistic regression analysis on the initial screening semen samples predicted a higher chance of conception for those with high levels of IgG (tail) (P = 0.006, sensitivity = 33%, specificity = 93%, correct = 75%, false positive = 33% and false negative = 24%). In the pregnant group, prednisolone caused a significant increase in grade I motility (P = 0.03). In the non-pregnant group, there was a significant increase in grade I motility (P = 0.0002), amplitude of lateral head displacement (P = 0.03), curvilinear velocity (P = 0.02) and decrease in grade IV motility (P = 0.03) following prednisolone treatment. In both groups there was suppression of the total antisperm antibody concentrations.(ABSTRACT TRUNCATED AT 250 WORDS)

Administration, Oral↗