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Biomedical subjects

S Khandekar

Publications and source records attributed to S Khandekar.

At least 19 recordsLinked to original sources
PubMed2000

Temperature-sensitive differential affinity of TRAIL for its receptors. DR5 is the highest affinity receptor.

TRAIL is a member of the tumor necrosis factor (TNF) family of cytokines which induces apoptotic cell death in a variety of tumor cell lines. It mediates its apoptotic effects through one of two receptors, DR4 and DR5, which are members of of the TNF receptor family, and whose cytoplasmic regions contain death domains. In addition, TRAIL also binds to 3 "decoy" receptors, DcR2, a receptor with a truncated death domain, DcR1, a glycosylphosphatidylinositol-anchored receptor, and OPG a secreted protein which is also known to bind to another member of the TNF family, RANKL. However, although apoptosis depends on the expression of one or both of the death domain containing receptors DR4 and/or DR5, resistance to TRAIL-induced apoptosis does not correlate with the expression of the "decoy" receptors. Previously, TRAIL has been described to bind to all its receptors with equivalent high affinities. In the present work, we show, by isothermal titration calorimetry and competitive enzyme-linked immunosorbent assay, that the rank order of affinities of TRAIL for the recombinant soluble forms of its receptors is strongly temperature dependent. Although DR4, DR5, DcR1, and OPG show similar affinities for TRAIL at 4 degrees C, their rank-ordered affinities are substantially different at 37 degrees C, with DR5 having the highest affinity (K(D) </= 2 nm) and OPG having the weakest (K(D) = 400 nm). Preferentially enhanced binding of TRAIL to DR5 was also observed at the cell surface. These results reveal that the rank ordering of affinities for protein-protein interactions in general can be a strong function of temperature, and indicate that sizeable, but hitherto unobserved, TRAIL affinity differences exist at physiological temperature, and should be taken into account in order to understand the complex physiological and/or pathological roles of TRAIL.

Animals↗
PubMed2000

Inhibition of CD23 processing correlates with inhibition of IL-4-stimulated IgE production in human PBL and hu-PBL-reconstituted SCID mice.

BACKGROUND: CD23, the low affinity serum immunoglobulin E (IgE) receptor, is upregulated on B cells following interleukin (IL)-4 stimulation and is concomitantly cleaved to generate soluble CD23 (sCD23) fragments with cytokine-like activity. OBJECTIVE: Compounds that selectively inhibit the proteolytic release of CD23 to generate sCD23 were assessed for their ability to inhibit IgE production in order to evaluate the contribution of sCD23 in the production of human IgE as well as the ability of such compounds to block IgE production. METHODS: IgE production was measured in IL-4-stimulated human peripheral blood lymphocytes (PBL) and PBL-reconstituted SCID mice in the presence of a broad-spectrum matrix metalloprotease (MMP) inhibitor, a compound selective for inhibition of CD23 processing over MMPs and an anti-CD23 mAb, MHM6. RESULTS: The two compounds were equipotent in inhibiting IgE production without inhibition of IgG production by IL-4/anti-CD40-stimulated PBL. Soluble CD23 release was also shown to precede IgE accumulation in the cell-free medium. Addition of compound at later times other than day 0 in the 14 day assay resulted in progressively less inhibition of both IgE and sCD23, and exactly paralleled the effect of an anti-CD23 mAb, MHM6 on IgE levels. Both compounds also inhibited the release of CD23 from human RPMI 8866 cells adoptively transferred i. p. to mice. Doses required for inhibition of CD23 correlated well with the doses required for inhibition of IgE production in IL-4-challenged hu-PBL-SCID mice. IgE was selectively inhibited over total IgG in the SCID mice as well. CONCLUSIONS: Inhibition of CD23 processing alone is sufficient to inhibit IL-4-stimulated IgE production both in vitro and in vivo.

Animals↗
PubMed2000

Endoscopic therapy for stenosis of the biliary and pancreatic duct orifices.

BACKGROUND: Manipulation of the duodenal papilla may lead to symptomatic stenosis of the orifices of bile duct, main pancreatic duct or accessory pancreatic duct. METHODS: Seventeen patients with stenosis of the orifice (bile duct 7, bile duct/main pancreatic duct 7, accessory pancreatic duct 3) underwent sphincterotomy and/or dilation and stent placement for a median of 140 days (range 30 to 1080 days). Patients were interviewed at a median of 720 days (range 120 to 990 days) after removal of the final stent. RESULTS: Median age was 50 years (range 17 to 68 years); 78% were women. The etiology of stenosis of the orifice was sphincterotomy in 8, sphincteroplasty in 7 and papillectomy in 2 patients. Indications for treatment were abdominal pain (100%), dilated bile duct and/or main pancreatic duct (14 patients) and pancreas divisum (3 patients). Sixty procedures (median 4 per patient) were performed with mild morbidity (hospital stay less than 3 days) in 17% of procedures and 35% of patients. Symptoms improved in 100%, 57% and 33% of patients with bile duct, bile duct/main pancreatic duct and accessory pancreatic duct, respectively. Surgery was ultimately needed in 3 (43%) patients with bile duct/main pancreatic duct and 2 (67%) with accessory pancreatic duct stenosis. CONCLUSIONS: Endoscopic therapy successfully relieves pain due to biliary stenosis of the orifice but less frequently relieves pain due to pancreatic stenosis of the orifice.

Adolescent↗
PubMed1999

Crystal structure of beta-ketoacyl-acyl carrier protein synthase III. A key condensing enzyme in bacterial fatty acid biosynthesis.

Beta-ketoacyl-acyl carrier protein synthase III (FabH), the most divergent member of the family of condensing enzymes, is a key catalyst in bacterial fatty acid biosynthesis and a promising target for novel antibiotics. We report here the crystal structures of FabH determined in the presence and absence of acetyl-CoA. These structures display a fold that is common for condensing enzymes. The observed acetylation of Cys(112) proves its catalytic role and clearly defines the primer binding pocket. Modeling based on a bound CoA molecule suggests catalytic roles for His(244) and Asn(274). The structures provide the molecular basis for FabH substrate specificity and reaction mechanism and are important for structure-based design of novel antibiotics.

3-Oxoacyl-(Acyl-Carrier-Protein) Synthase↗
PubMed1999

Scintigraphic detection of post-pneumonectomy bronchopleural fistulae.

A total of 20 ventilation studies [16 with xenon-133 and four with technetium-99m diethylenetriamine pentaacetic acid (DTPA)] were performed in 11 patients with suspected post-pneumonectomy bronchopleural fistulae. The findings on the ventilation scan were correlated with bronchoscopy, taken as the gold standard for purposes of comparison. The sensitivity and specificity for 133Xe scans were 83% and 100% respectively, while the sensitivity for 99mTc-DTPA aerosol studies was poor at 0%. Special techniques for optimal visualization of the fistulae are enumerated.

Aerosols↗
PubMed1998

The immune response to soluble D10 TCR: analysis of antibody and T cell responses.

In order to evaluate the potential of TCR as vaccines for immunomodulation, the immunogenicity of soluble versions of D10 TCR has been investigated in mice. Soluble D10 TCR containing the extracellular domains were produced either as dual chain (dc) TCR lacking transmembrane and cytoplasmic regions or as a TCR-IgG1 chimeric protein. Soluble single chain (sc) D10 TCR contained only the Valpha and Vbeta segments joined by a peptide linker. Syngeneic D10 dcTCR or D10 TCR-IgG1 immunizations of AKR mice induced antibody responses to D10 clonotypic epitopes and to constant region epitopes that are not exposed on D10 cells. Only clonotypic antibodies were produced after D10 scTCR immunizations. Immunization of AKR mice with D10 dcTCR and D10 TCR-IgG1 primed I-Ak- and I-Ek-restricted CD4+ T cells recognizing constant region epitopes, but there was no detectable response to the variable region. Comparison of the in vitro proliferative responses of CD4+ T cells from D10 scTCR-primed H-2 congenic mice revealed that H-2u was a responder haplotype for the variable region. How the immunogenicity of particular regions of the TCR appears to be shaped by tolerance induction in vivo and the implications for immunotherapy with soluble TCR vaccinations are discussed.

Adjuvants, Immunologic↗
PubMed1997

Maternal immunization with a soluble TCR-Ig chimeric protein: long term, V beta-8 family-specific suppression of T cells by maternally transferred antibodies.

Maternal transfer of TCR clonotypic Ab protected young NOD mice against the adoptive transfer of diabetes by the BDC 2.5 T cell clone. The effect of maternal anti-TCR Vbeta-8 Ab on T cell development and function has now been investigated. SJL/J mice, which lack TCR Vbeta-8, were immunized with soluble, chimeric D10 TCR-IgG1 containing Vbeta-8.2. The (SJL/J x AKR/J) F1 offspring of immunized female SJL/J mice were severely depleted of peripheral T cells bearing Vbeta-8 until 11 to 17 wk of age. The loss of Vbeta-8 expression did not appear to be due to modulation of cell surface TCR. Since the Vbeta-8+ T cell population was unperturbed in the (AKR/J x SJL/J) F1 offspring of D10 TCR-IgG1-immunized AKR/J mothers making D10 clonotypic Ab, the effect was immunologically specific. The deletion of Vbeta-8+ T cells had functional consequences. In the in vitro response to the superantigen, staphylococcal enterotoxin B, the usually observed participation of Vbeta-8.2+ T cells was largely suppressed, whereas the recruitment of Vbeta-3+ T cells remained unaltered. In control mice, T cell responses to the 134- to 146-residue peptide of conalbumin (pCA(134-146)) were biased toward use of Valpha-2/Vbeta-8.2 TCR. In D10 TCR-IgG1 maternally immunized (SJL x AKR/J) F1 mice, the T cell responses to pCA(134-146) were suppressed, and T cell lines derived from these in vitro were devoid of Vbeta-8.2 expression. With an increased understanding of TCR V gene usage in autoimmune diseases, similar strategies for the depletion of autoreactive T cells may become feasible in humans.

Animals↗
PubMed1996

Immunization with soluble BDC 2.5 T cell receptor-immunoglobulin chimeric protein:antibody specificity and protection of nonobese diabetic mice against adoptive transfer of diabetes by maternal immunization.

The BDC 2.5 T cell clone is specific for pancreatic beta-cell antigen presented by I-Ag7, and greatly accelerates diabetes when injected into 10-21-d-old nonobese diabetic (NOD) mice. The BDC 2.5 T cell receptor (TCR) has been solubilized as a TCR-IgG1 chimeric protein. All NOD mice immunized against BDC 2.5 TCR-IgG1 produced antibodies recognizing TCR C alpha/C beta epitopes that were inaccessible on the T cell surface. 56% of the mice produced antibodies against the BDC 2.5 clonotype that specifically blocked antigen activation of BDC 2.5 cells. We have used the adoptive transfer model of diabetes to demonstrate that maternal immunization with soluble TCR protects young mice from diabetes induced by the BDC 2.5 T cell clone.

Adoptive Transfer↗
PubMed1994

Human Hsp60: bacterial expression, purification, development of monoclonal antibodies and a sandwich ELISA for quantitation.

Bacterial hsp60 proteins are major targets of immune responses during infection, and the highly conserved nature of bacterial and mammalian hsp60 has led to speculation that immune reactivity to these stress proteins may be a component of certain autoimmune diseases. We have developed recombinant proteins and monoclonal antibodies to facilitate further study of human hsp60 and its association with disease. The human hsp60 gene was expressed in Escherichia coli and a method for purification of the recombinant protein essentially devoid of E. coli GroEL was developed. Using the purified protein we have generated a number of monoclonal antibodies which are specific for human hsp60 and do not cross-react with its counterparts from E. coli and mycobacteria. A highly sensitive sandwich ELISA was developed to quantitate hsp60 levels and was used to study hsp60 accumulation in cells due to vaccinia virus infection and heat shock. This ELISA will be useful for monitoring hsp60 levels in body fluids or tissues during autoimmune reactions and/or inflammatory responses.

Animals↗
PubMed1991

46 XX male with dysgenetic testes.

A 46 XX male with dysgenetic gonad is described. The patient was brought up as a girl and reported with primary amenorrhoea and hirsutism. The pathogenesis and management strategy of this condition are briefly discussed.

Adult↗
PubMed1990

Effect of sulphonylurea therapy on insulin sensitivity in non insulin dependent diabetics.

Insulin tolerance test was done to assess the insulin sensitivity in a group of untreated non insulin dependent diabetic subjects before and after sulfonylurea (tolbutamide, glibenclamide, glipizide) therapy. The parameters studied were KITT, slope of the blood glucose fall, summation values, and area under the curve of glucose response. All the three commonly used sulphonylurea drugs improved insulin sensitivity after 4 weeks of therapy.

Blood Glucose↗
PubMed1990

Screening for late onset congenital adrenal hyperplasia due to 21-hydroxylase deficiency.

Basal and post-ACTH levels of 17 alpha hydroxy-progesterone (17 OHP) were determined in 53 subjects with hirsutism. Late onset congenital adrenal hyperplasia (LOCAH) was detected in five (10.6%) on the basis of elevated basal and/or ACTH stimulated levels of 17 OHP. Of the five patients, two were considered to have a heterogygous state on account of a small rise in stimulated 17 alpha OHP. Screening tests for LOCAH are essential as the clinical diagnosis is not otherwise possible for this treatable and often familial disorder.

17-alpha-Hydroxyprogesterone↗
PubMed1990

Ketoconazole in the treatment of Cushing's disease.

Six patients with Cushing's disease received ketoconazole for 7-10 weeks following ketoconazole challenge test. Clinical and hormonal alterations in circulating cortisol, 17 OH progesterone, androstenedione and testosterone were assessed during the therapy. There was significant clinical improvement which corresponded to a fall in cortisol, androstenedione and testosterone. The rise in 17 OH progesterone and the fall in cortisol suggest a blockade at 17,20 desmolase and 11 B-hydroxylase in the cortisol bio-synthetic pathway.

17-alpha-Hydroxyprogesterone↗
PubMed1990

Persistent Cushing's disease in a patient with invasive pituitary adenoma.

A patient with invasive basophilic pituitary adenoma is described, in whom the hypercortisolaemia and clinical features persisted despite two attempts to reduce the tumour mass by transfrontal pituitary surgery and external beam radiotherapy. The patient responded to oral ketoconazole with clinical and biochemical improvement.

Adenocarcinoma↗
PubMed1990

Circulating cortisol & related steroids following ketoconazole challenge in Cushing's disease.

Circulating levels of cortisol, 17 alpha hydroxy progesterone and delta 4 androstenedione were analysed by specific radioimmunoassays in 11 patients with Cushing's disease before and at 2, 4, 6, 12 and 24# h following an oral dose of 400 mg ketoconazole. A significant fall in cortisol (26 - 79.6%, 59.8 +/- 18.7 SEM), delta 4 androstenedione (21.5 - 63.3%, 47.8 +/- 4.87) with a concomitant rise in 17 alpha hydroxy progesterone (113 - 218%, 116 +/- 11.43) were noted, suggesting inhibition of 17, 20 desmolase and 11 beta-hydroxylase enzymes in cortisol biosynthetic pathway.

17-alpha-Hydroxyprogesterone↗
PubMed1989

Circulating androgens in hirsutism.

The androgen status in fifty consecutive women with hirsutism in the reproductive age group referred to the endocrine clinic was studied. Mean BMI was 24.3 +/- 5.6 kg/M2, 42% had menstrual disturbances and 30%, had severe hirsutism. The hirsute patients had significantly higher estimates of testosterone (48%), dihydrotestosterone (38%) and androstenedione (86%). Based on clinical and hormonal data, polycystic ovarian disease was diagnosed in 20 patients, nonclassical 21 hydroxylase deficiency in one and idiopathic hirsutism in 29.

Adult↗