Biomedical subjects
Publications and source records attributed to S K Nayak.
Two-particle correlations of direct photons were measured in central 208Pb+208Pb collisions at 158A GeV. The invariant interferometric radii were extracted for 100<K(T)<300 MeV/c and compared to radii extracted from charged pion correlations. The yield of soft direct photons, K(T)<300 MeV/c, was extracted from the correlation strength and compared to theoretical calculations.
Antigenic sharing among the most commonly bacterial pathogens such as Aeromonas hydrophila, Edwardsiella tarda and Pseudomonas fluorescens of Indian major carps has been studied using immunological reactions such as cross-agglutination, disc diffusion and indirect enzyme linked immunosorbent assay (ELISA). The data were analysed using statistical analysis (SAS), version 6.12. The results showed high antigenic similarities among the bacterial whole cells, whole cell lysates, somatic 'O' antigens, lipopolysaccharides (LPS) and extracellular products (ECP). However, few or no similarities were observed in ECP components of <20kD. The present study indicates a need to develop differential diagnostic methods based on serology by choosing the highly specific less cross-reactive ECP antigen.
Different serological tests viz. indirect ELISA, indirect blocking ELISA, competitive ELISA and serum agglutination tests were evaluated to detect antibodies against Edwardsiella tarda in naturally infected fish sera for seromonitoring and epizootiological studies. Approximately 66.6, 62.5, 57.6 and 16.6% of the field sera samples were found to be positive by indirect ELISA, competitive ELISA, indirect blocking ELISA and serum agglutination test, respectively. The percentage of serum samples positive for E. tarda antibodies in serum agglutination, competitive ELISA and indirect blocking ELISA, when compared with indirect ELISA, were 33.3, 83.6 and 66.6%, respectively, but its use was restricted due to the requirement of several conjugates against different fish species and the difficulty in assaying large numbers of serum samples from different fish species in a limited time to enable seromonitoring of the disease prevalence. No significant difference (P<0.05) in the mean optical density value was found in indirect and competitive ELISA. Although the competitive ELISA was slightly less sensitive than the indirect ELISA, it could accommodate a large number of serum samples with one anti-rabbit conjugate, and the need for different fish conjugates as required in indirect ELISA was eliminated. As in medical and veterinary practices, these tests can now be used in aquaculture practices for seromonitoring and study of pre-exposure of Indian major carps to pathogens in enzootic areas.
Density-functional calculations of charge distribution on negatively and positively charged nanotubes result in charge density profiles characterized by a significant increase of charge density at the tube ends. These results are in quantitative agreement with classical electrostatic analysis, which assumes constant electrostatic potential on the conductive tube surface. At high charging levels, the tube ends are observed to be unstable due to Coulomb repulsion. By combining ab initio calculations with classical electrostatics, we determine, as a function of tube length and geometry of the tube end, the critical voltage beyond which nanotubes are unstable.
Larval mortality in Indian major carps is one of the major problems encountered in the pond culture system. The present investigation was carried out to investigate the proper age, duration of exposure, and optimum bacterin concentration for vaccinating rohu (Labeo rohita) and catla (Catla catla) at their early stages with a formalin killed Edwardsiella tarda bacterin suspension. The development of immunological competence was recorded with spawn of rohu and catla of 3 weeks of age exposed to a bacterin at a concentration 10(9) cfu ml(-1) for 15 min, where it persisted up to 4 weeks post vaccination. They showed significant resistance against challenge with virulent E. tarda bacteria. Significant antibody titre could be recorded in advanced fries and fingerlings exposed to 10(9) cfu/ml(-1) bacterin concentration for 45 and 60 min, respectively.
Restrictocin, a member of the fungal ribotoxin family, specifically cleaves a single phosphodiester bond in the 28S rRNA and potently inhibits eukaryotic protein synthesis. Residues Tyr47, His49, Glu95, Phe96, Pro97, Arg120, and His136 have been predicted to form the active site of restrictocin. In this study, we have individually mutated these amino acids to alanine to probe their role in restrictocin structure and function. The role of Tyr47, His49, Arg120, and His136 was further investigated by making additional mutants. Mutating Arg120 or His136 to alanine or the other amino acids rendered the toxin completely inactive, whereas mutating Glu95 to alanine only partially inactivated the toxin. Mutation of Phe96 and Pro97 to Ala had no effect on the activity of restrictocin. The Tyr47 to alanine mutant was inactive in inhibiting protein synthesis, and had a nonspecific ribonuclease activity on 28S rRNA similar to that shown previously for the His49 to Ala mutant. Unlike the His136 to Ala mutant, the double mutants containing Tyr47 or His49 mutated to alanine along with His136 did not compete with restrictocin to cause a significant reduction in the extent of cleavage of 28S rRNA. In a model of restrictocin and a 29-mer RNA substrate complex, residues Tyr47, His49, Glu95, Arg120, and His136 were found to be near the cleavage site on RNA. It is proposed that in restrictocin Glu95 and His136 are directly involved in catalysis, Arg120 is involved in the stabilization of the enzyme-substrate complex, Tyr47 provides structural stability to the active site, and His49 determines the substrate specificity.
[70]Fullerene has been shown to form 1:1 molecular complexes with toluene, p-xylene, m-xylene, 1,2,4,5-tetramethyl benzene (durene) and pentamethyl benzene (PMB) in CCl4 medium by absorption spectroscopic method. Isosbestic points have been detected in case of complexes with PMB and durene. Charge transfer absorption band could not be detected but the intensity of the broad absorption band of C70 in CCl4 decreases systematically with increase in the concentration of the added methylbenzenes. From this trend the formation constants (Kc) of the complexes have been determined at three different wavelengths. The constancy of Kc with respect to change in the wavelength of measurement supports the view that complex of a single stoichiometry (1:1) is formed in each case.
OBJECTIVE: We tried to establish short-term cultures of autologous tumors from patients with breast carcinoma for potential use as active specific immunotherapy (i.e., autologous vaccine) after resection of primary breast cancer, and/or for the treatment of metastases. METHODS: Between 10/90 and 12/99 the cell biology laboratory of the Hoag Cancer Center attempted to establish short-term tumor cell lines from 115 breast cancer specimens from 56 primary breast lesions, 17 axillary nodes, 14 other lymph node/soft tissue sites, 10 chest wall recurrences, and 6 thoracentesis of malignant pleural effusions. Success was defined by growth of 5 x 10(7) viable cells whose malignant nature and breast cancer origin was confirmed by histology of the submitted tissue, cell morphology and antigenic phenotyping. Variables associated with successful growth of short-term cell lines were examined. RESULTS: Expansion to 5 x 10(7) cells was achieved for only 8/115 samples [7%] including two from chest wall recurrences, and one each from a supraclavicular node, an umbilical node, liver, omentum, and pleural fluid. Two of the successful cell lines were established from tissue that originally had been cryopreserved; the others were initiated from fresh tumor. The success rate was better from regional/distant metastases 7/55 (13%) compared to primary tumors 1/56 (1.8%) (p = 0.063). The success rate for tumors harvested at Hoag Hospital was 4/97 (4%) compared to 4/14 from (31%) distant sites, but all but one of the tumors from a distant geographic site was a metastatic lesion. Tumor cell lines were successfully established from metastatic lesions ranging in size from < 1.0 g to 19 g. Four patients were treated with their autologous vaccine in the setting of chemotherapy-refractory metastatic disease without any significant toxicity. CONCLUSIONS: We were unable to establish short-term cell lines for most patients with primary or metastatic breast cancer using this methodology. However, two long-term cell lines have been established and characterized. Treatment with the autologous irradiated cell product was not associated with acute toxicity.
BACKGROUND: We established short-term cultures of autologous tumors from patients with renal carcinoma for use as active specific immunotherapy (i.e., autologous vaccine). METHODS: Between 9/91 and 9/99 the cell biology laboratory of the Hoag Cancer Center received 69 kidney tumor samples that had been surgically excised, including 43 primary tumors and 26 metastatic lesions. Efforts were made to establish short-term tumor cell cultures to use as autologous tumor cell vaccines. Prior to treatment, patients underwent a baseline skin test for delayed tumor hypersensitivity (DTH) and then received s.c. injections of 10 million irradiated tumor cells that were given with various adjuvants weekly x3 and then monthly x5. RESULTS: Cell lines were established for 55/69 patients (80%) including 36/43 (84%) from primary tumors and 19/26 (73%) from distant metastases. Vaccines were prepared for 41 patients; 27 were treated. At the time of this analysis, follow up data was available for 26 patients with a median follow up > 5 years. Treatment was well-tolerated. Of 10 patients who had no evident disease at the time of treatment, nine were alive 1-8 years later; 5/8 had conversion of their DTH test from negative to positive. For 16 patients with measurable metastatic disease at the time of treatment, there were no objective tumor responses; their median survival was 5.0 months. Among these 16 patients, only 1/8 DTH tests converted, but three had a positive baseline DTH test; one was previously treated with interleukin-2 and tumor infiltrating lymphocytes and two others were previously treated with autolymphocyte therapy. CONCLUSIONS: Vaccine therapy with short-term cultures of autologous tumor cells is feasible, well-tolerated and associated with conversion of DTH and long-term survival in patients who are free of disease at the time treatment is initiated. However, significant anti-tumor responses were not seen in patients with measurable disease at the time vaccine treatment was initiated.
Pathological alterations in various organs of rohu (L. rohita) fingerlings following acute (0, 7.50, 11.25 and 13.75 mg/kg body weight) and subchronic (0, 1.25 and 2.50 mg/kg body weight) single i.p. aflatoxin B1 exposure for 10 and 90 days, respectively, were investigated. Mortality (dose-dependent) was marked only during acute toxicosis. The changes observed in various organs were dose and time dependent. The acute dose groups revealed toxic changes viz., necrotic and vascular changes in liver and gill lamellae; meningitis, congestion in brain, degeneration and inflammatory reaction in heart along with degenerative to necrotic changes in kidney tubules and sloughing of the intestinal mucosa. During subchronic exposure to this toxin, preneoplastic lesions in liver along with changes in spleen, intestine, gill and pancreas were recorded. With low doses of aflatoxin, the fish did not reveal any mortality or external signs other than catchexia and increased pigmentation on scales. In composite culture practice of Indian major carps, this could be of economic significance.
Mucinous carcinoma of the male breast is a very rare tumour. So far only about 8 cases have been reported in the literature most of which were diagnosed after surgical resection. Fine-needles aspiration biopsy even though a well established procedure for the evaluation of breast masses in female there are only occasional reports of aspirates form male breast masses. We herein-present one case of mucinous carcinoma of breast in a 65 year old male diagnosed by fine needle and confirmed by histopathology.
Three-particle correlations have been measured for identified pi(-) from central 158A GeV Pb+Pb collisions by the WA98 experiment at CERN. A substantial contribution of the genuine three-body correlation has been found as expected for a mainly chaotic and symmetric source.
Restrictocin, produced by the fungus Aspergillus restrictus, is a highly specific ribonucleolytic toxin which cleaves a single phosphodiester bond between G4325 and A4326 in the 28S rRNA. It is a nonglycosylated, single-chain, basic protein of 149 amino acids. The putative catalytic site of restrictocin includes Tyr47, His49, Glu95, Arg120 and His136. To map the catalytic activity in the restrictocin molecule, and to study the role of N- and C-terminus in its activity, we have systematically deleted amino-acid residues from both the termini. Three N-terminal deletions removing 8, 15 and 30 amino acids, and three C-terminal deletions lacking 4, 6, and 11 amino acids were constructed. The deletion mutants were expressed in Escherichia coli, purified to homogeneity and functionally characterized. Removal of eight N-terminal or four C-terminal amino acids rendered restrictocin partially inactive, whereas any further deletions from either end resulted in the complete inactivation of the toxin. The study demonstrates that intact N- and C-termini are required for the optimum functional activity of restrictocin.
OBJECTIVE: We have tried to establish short-term cultures of autologous tumors from patients with renal cell carcinoma that could be used as active specific immunotherapy (i.e., autologous vaccine) in such patients after resection of primary kidney cancer, and/or for the treatment of metastatic cancer. METHODS: Between 10/90 and 9/99 the cell biology laboratory of the Hoag Cancer Center received 69 kidney tumor samples that had been surgically excised, including 43 primary tumors and 26 metastatic lesions. Efforts were made to establish short-term tumor cell cultures, as defined by the growth of 10(8) cells; malignant nature and renal cell origin were confirmed by morphology and antigenic phenotyping. Variables associated with successful growth of short-term cell lines were examined. RESULTS: Short-term cell lines were successfully established from 55/69 samples [80%] including 36/43 (84%) from primary tumors and 19/26 (73%) from metastatic lesions. The success rate for tumors harvested at Hoag Hospital was 40/50 (80%); the success rate for tumors obtained from other geographic areas was 15/19 (79%). Tumor cell lines were successfully established from metastatic lesions ranging in size from a 0.5 g vertebral lesion to a 22 g rib/lung chest wall metastasis, and from primary renal cell lesions ranging in size from 1.5 g to 39.7 g. CONCLUSIONS: Short-term cell lines can be established for most patients with primary or metastatic renal cell carcinoma making a pure autologous tumor-cell vaccine approach feasible. Vaccines have been prepared for 41 patients and a vaccine therapy trial is in progress.
Cell lines are valuable resources for the study of the malignancy and potential therapy of human breast cancer. A major problem with adapting fresh breast tumor specimens to grow in vitro is contamination by fibroblasts. Previously, we have reported a technique to overcome this problem (Nayak, S. K; Dillman, R. O. Clin. Biotechnol. 3:237-242; 1991). We have recently established two new breast cancer cell lines, HH315 and HH375, that were derived from abdominal and supraclavicular lymph node metastases from two patients. They were characterized by (1) growth kinetics; (2) staining with monoclonal antibodies (MoAbs) to cytokeratin-19, epithelial membrane antigen (EMA), anticarcinoembryonic antigen (CEA), breast cancer antigen 1 (BRST-1), breast cancer antigen 2 (BRST-2), Her2/neu, and p53; (3) expression of domains of urinary plasminogen activator (uPA), neural cell adhesion molecule (NCAM), and haptoglobin (Hp) (Harvey et al., 1997); and (4) karyotypic analysis. Growth kinetic studies showed that doubling times for both lines ranged from 48 to 96 h. These two cell lines were found to have characteristics of the metastatic breast cancer cells. Both lines stained positive with MoAbs to cytokeratin-19 and EMA, thus confirming their epithelial origin. They also strongly reacted with the pan-breast carcinoma MoAbs BRST-1 and BRST-2, and carcinoembryonic CEA MoAb. Both cell lines overexpressed the oncogene proteins Her2/neu and p53. The tumor cells were negative for estrogen and progesterone receptors. HH315 cells were poorly differentiated, whereas the HH375 cells exhibited adenocarcinoma morphology. Both cell lines showed intense cell surface and some cytoplasmic staining for uPA, NCAM, and Hp domains, which is a characteristic of malignant neoplasms (Harvey et al., 1997). The HH375 cell line showed two cell types, of which 60% were hyperdiploids with 60-70 chromosomes and 5-10 marker chromosomes. The remaining cells were polyploid with more than 200 chromosomes. Cell line HH315 consisted of only a polyploid population. These cell lines may be useful in breast cancer research.
Restrictocin, produced by the fungus Aspergillus restrictus, belongs to the group of ribonucleolytic toxins called ribotoxins. It specifically cleaves a single phosphodiester bond in a conserved stem and loop structure in the 28S rRNA of large ribosomal subunit and potently inhibits eukaryotic protein synthesis. Restrictocin contains 149 amino acid residues and includes four cysteines at positions 5, 75, 131, and 147. These cysteine residues are involved in the formation of two disulfide bonds, one between Cys 5 and Cys 147 and another between Cys 75 and Cys 131. In the current study, all four cysteine residues were changed to alanine individually and in different combinations by site-directed mutagenesis so as to remove one or both the disulfides. The mutants were expressed and purified from Escherichia coli. Removal of any cysteine or any one of the disulfide bonds individually did not affect the ability of the toxin to specifically cleave the 28S rRNA or to inhibit protein synthesis in vitro. However, the toxin without both disulfide bonds completely lost both ribonucleolytic and protein synthesis inhibition activities. The active mutants, containing only one disulfide bond, exhibited relatively high susceptibility to trypsin digestion. Thus, none of the four cysteine residues is directly involved in restrictocin catalysis; however, the presence of any one of the two disulfide bonds is absolutely essential and sufficient to maintain the enzymatically active conformation of restrictocin. For maintenance of the unique stability displayed by the native toxin, both disulfide bonds are required.
Transduction of renal cell carcinoma (RCC) cells in vitro with a gene for human interferon-gamma (IFN-gamma) results in enhanced expression of key molecules needed for immune recognition. In this study, we investigated the ability of soluble IFN-gamma protein to enhance expression of these key molecules. RCC cells were incubated with and without IFN-gamma, then analyzed by flow cytometry for the percent positive and mean intensity fluorescence (MIF), a measurement of mean antigen density, of HLA-I, HLA-II, ICAM-1, and tumor antigens (URO-2, URO-3, and URO-4). Results were compared to those seen for RCC cells transduced by IFN-gamma. The ability of cytotoxic T lymphocytes (CTL) to lyse RCC targets was measured using a standard [51Cr]lytic assay. Control cells were 99% (MIF 751) and 98% (MIF 315) positive for HLA-I and ICAM-1, respectively, with 2% (MIF 8) positive for HLA-II. Incubation with IFN-gamma protein resulted in 98% positive HLA-I (MIF 2288), 98% positive ICAM-1 (MIF 1132), and 95% positive HLA-II (MIF 287). The results for the cells incubated with IFN-gamma protein were similar to those for the transduced line. Importantly, the enhanced expression was maintained for several days after irradiation and cryopreservation. Expression of the URO tumor markers was not affected. Protein-treated RCC targets showed superior CTL lysis compared with untreated cells. Our results show that IFN-gamma protein incubation in vitro enhances expression of important immune recognition molecules to levels expressed by transduced cells. Increased expression may enhance tumor recognition by the host's immune system, as in the case of tumor cell vaccines. There may be no advantage to IFN-gamma transduction over in vitro incubation with IFN-gamma protein.
OBJECTIVE: We have tried to establish short-term cultures of autologous tumors from patients with stage III and IV ovarian cancer, which could be used as active specific immunotherapy, (i.e., autologous vaccine) in such patients after debulking surgery & combination chemotherapy. METHODS: Between 5/93 and 11/97 the Hoag cell biology laboratory received 53 ovarian tumor samples that had been surgically excised at the time of laparotomy, and four samples of malignant ascites. Efforts were made to establish short-term tumor cell cultures as confirmed by morphology & phenotype. RESULTS: Short-term proliferating cultures were successfully established from 21/57 samples [37%] which included 8/24 [33%] successes from samples obtained at diagnosis compared to 13/33 [37%] samples obtained at the time of a relapse [p = 0.45]. The probability of successful culture was not related to tumor size for samples with a range of 0.8-34 g (mean 5.8 g). One patient was treated in the setting of metastatic disease and one in the adjuvant setting; both received repeated injections of irradiated autologous tumor cells plus granulocyte macrophage stimulating factor (GM-CSF). In one patient a delayed tumor hypersensitivity skin test converted from negative to positive. CONCLUSIONS: Short-term cultures of autologous tumor cells for use as tumor cell vaccines can be established for about one-third of patients with ovarian cancer using this methodology and the treatment approach is feasible.