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Biomedical subjects

A Banerjee

Publications and source records attributed to A Banerjee.

At least 307 records · Page 17Linked to original sources
PubMed1994

Microtubule dynamics in vitro are regulated by the tubulin isotype composition.

The growing and shortening dynamics of individual bovine brain microtubules at their plus ends at steady state in vitro, assembled from isotypically pure alpha beta II, alpha beta III, or alpha beta IV tubulin dimers, were determined by differential interference contrast video microscopy. Microtubules assembled from the purified alpha beta III isotype were considerably more dynamic than microtubules made from the alpha beta II or alpha beta IV isotypes or from unfractionated phosphocellulose-purified tubulin. Furthermore, increasing the proportion of the alpha beta II isotype in a mixture of the alpha beta II and alpha beta III isotypes suppressed microtubule dynamics, demonstrating that microtubule dynamics can be influenced by the tubulin isotype composition. The data support the hypothesis that cells might determine the dynamic properties and functions of its microtubules in part by altering the relative amounts of the different tubulin isotypes.

Animals↗
PubMed1994

Implications of multidrug resistance for the future of short-course chemotherapy of tuberculosis: a molecular study.

Tuberculosis-control programmes are compromised by the increased frequency of multidrug-resistant strains of Mycobacterium tuberculosis. We used the polymerase chain reaction (PCR) and single-strand conformation polymorphism (SSCP) analysis techniques to establish the molecular basis of resistance in 37 drug-resistant isolates of M tuberculosis, and correlated these findings with clinical and antibiotic-sensitivity data. Resistance to isoniazid was found in 36 strains, 16 of which were also resistant to ethionamide. Of the 36 isoniazid-resistant strains, 23 had mutations in the katG gene, and 5 of these also had mutations in the inhA gene. A further 5 strains had alterations in the inhA locus without the katG gene being mutated. Rifampicin resistance was less frequent (13 strains) and usually associated with isoniazid resistance (11 of 13 strains). Mutations in the rpoB gene were detected for all these rifampicin-resistant isolates. Mutations in the rpsL and rrs genes, associated with streptomycin resistance, were found in 13 of 25 and 2 of 25 streptomycin-resistant strains, respectively. The same chromosomal mutations, or combinations of mutations, were found in strains displaying single or multidrug resistance, from cases of both primary and secondary resistance, and from patients infected with human immunodeficiency virus. Thus, multidrug resistance is not due to a novel mechanism and tuberculosis chemotherapy is not subject to a new threat.

AIDS-Related Opportunistic Infections↗
PubMed1994

Cloning, sequencing, and expression of Rhodococcus L-phenylalanine dehydrogenase. Sequence comparisons to amino-acid dehydrogenases.

L-Phenylalanine dehydrogenase catalyzes the NAD(+)-dependent, reversible, oxidative deamination of L-phenylalanine to form ammonia, phenyl pyruvate, and NADH. The enzyme has been purified to homogeneity from Rhodococcus sp. M4, and a partial amino acid sequence was obtained. A cosmid library of Rhodococcus sp. M4 genomic DNA was prepared and used to isolate a 2.5-kilobase PstI fragment that contained the pdh gene. The open reading frame of 1068 nucleotides encodes a polypeptide of 356 amino acids, portions of which match the amino acid sequence determined for the purified enzyme. Expression of the Rhodococcus pdh gene in Escherichia coli, which does not contain a phenylalanine dehydrogenase activity, yields a soluble enzyme exhibiting phenylalanine dehydrogenase activity. Both the enzyme purified from Rhodococcus and the enzyme expressed in E. coli are post-translationally modified by removal of the amino-terminal methionine. The overall amino acid sequence is homologous to previously reported sequences of leucine and phenylalanine dehydrogenases as well as several glutamate dehydrogenases. The amino-terminal portion of the enzyme contains residues involved in L-amino acid binding and catalysis, while the carboxyl-terminal portion contains the presumptive dinucleotide-binding domain. A detailed sequence comparison of Rhodococcus phenylalanine dehydrogenase with leucine, phenylalanine, and glutamate dehydrogenases suggests residues involved in general amino acid binding and others that provide for amino acid discrimination.

Amino Acid Oxidoreductases↗
PubMed1994

Interaction of desacetamidocolchicine, a fast binding analogue of colchicine with isotypically pure tubulin dimers alpha beta II, alpha beta III, and alpha beta IV.

Desacetamidocolchicine (DAAC) is a colchicine analogue that lacks the acetamido side chain in the ring-B of colchicine. Unlike colchicine, it binds to tubulin very quickly, and yet it has powerful antimitotic properties. It has been demonstrated that the beta-tubulin isoforms differ significantly in their interactions with colchicine. In an effort to understand the role of the ring-B of colchicine, we have studied the interaction of DAAC with purified beta-tubulin isoforms. The association was studied fluorometrically using a stopped-flow instrument under pseudo-first-order conditions in the presence of a large excess of drug. The observed pseudo-first-order rate constants increased in a nonlinear way with the drug concentration, indicating that the binding of DAAC to tubulin isoforms occur in two steps as is true for the binding of colchicine to tubulin (Garland, D.L. (1978) Biochemistry 17, 4266-4272), [formula: see text] where the first step is a fast reversible binding and the second step is a slow conformational change leading to the formation of the stable complex (T.DAAC)*. Kinetic analysis shows that the tubulin isoforms exhibit very little differences in their K1 values, which are 5794 +/- 670, 7109 +/- 1800, and 8993 +/- 1780 M-1 for alpha beta II, alpha beta III, and alpha beta IV, respectively. The k2 values for alpha beta II, alpha beta III, and alpha beta IV are 0.67 +/- 0.05, 0.05 +/- 0.006, and 0.59 +/- 0.07 s-1, respectively. The apparent on-rate constants (k(on,app) = K1k2) for alpha beta II, alpha beta III, and alpha beta IV are 3907 +/- 530, 376 +/- 80, and 5305 +/- 1200 M-1 s-1, respectively. The off-rate constants as measured by the loss of fluorescence of drug-tubulin complexes in the presence of a large excess of podophyllotoxin are 6.3 x 10(-4), 5.2 x 10(-4), and 5.7 x 10(-4) s-1, respectively, for alpha beta II, alpha beta III, and alpha beta IV. The affinity constants as determined by Scatchard analyses are 2.5 x 10(6), 1.5 x 10(6), and 4 x 10(6) M-1 for alpha beta II, alpha beta III, and alpha beta IV, respectively.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1994

inhA, a gene encoding a target for isoniazid and ethionamide in Mycobacterium tuberculosis.

Isoniazid (isonicotinic acid hydrazide, INH) is one of the most widely used antituberculosis drugs, yet its precise target of action on Mycobacterium tuberculosis is unknown. A missense mutation within the mycobacterial inhA gene was shown to confer resistance to both INH and ethionamide (ETH) in M. smegmatis and in M. bovis. The wild-type inhA gene also conferred INH and ETH resistance when transferred on a multicopy plasmid vector to M. smegmatis and M. bovis BCG. The InhA protein shows significant sequence conservation with the Escherichia coli enzyme EnvM, and cell-free assays indicate that it may be involved in mycolic acid biosynthesis. These results suggest that InhA is likely a primary target of action for INH and ETH.

Amino Acid Sequence↗
PubMed1994

Endotoxin disrupts beta-adrenergic signal transduction in the heart.

BACKGROUND: Nonsurvivors of septic shock demonstrate impaired myocardial function refractory to the administration of beta-agonists. METHODS: Using the isolated rat heart preparation, the integrity of the beta-adrenergic transduction pathway was tested (rate pressure product, rate of contraction, rate of relaxation, and cyclic adenosine monophosphate content) using isoproterenol hydrochloride (beta-receptor agonist) or colforsin (forskolin) (adenylyl cyclase activator) stimulation following intracoronary endotoxin infusion. RESULTS: Basal rate pressure product, rate of contraction, rate of relaxation, and cyclic adenosine monophosphate concentrations were unaffected by endotoxin infusion. Endotoxin impaired, increases in rate pressure product, rates of contraction and relaxation, and cyclic adenosine monophosphate to isoproterenol (P < .05), but the response to colforsin was unaffected by endotoxin. CONCLUSIONS: Endotoxin disrupts the myocardial response to direct beta-receptor stimulation but not to adenylyl cyclase stimulation in the isolated rat heart. CLINICAL RELEVANCE: Alteration of the proximal beta-adrenoreceptor complex by endotoxin suggests that therapy of the failing heart during refractory septic shock may be directed to intact sites distal in the beta-adrenergic pathway.

Adenylyl Cyclases↗
PubMed1994

Phospholipase A2 regulates critical inflammatory mediators of multiple organ failure.

PLA2 is a family of regulatory enzymes that control eicosanoid synthesis and PAF production. PLA2 must be tightly regulated within the cell or cell destruction results. Circulatory release of PLA2 occurs in states of profound illness including sepsis, shock, severe injury, and pancreatitis, all of which are linked to the development of ARDS and MOF. Experimental and clinical evidence suggests that PLA2 may serve a primary regulatory role in the development of these inflammatory disorders. This evidence suggests that inhibitors of PLA2 activation could play an important role in future intensive care management.

Animals↗
PubMed1994

Endotoxin pretreatment inhibits neutrophil proliferation and function.

Gut ischemia/reperfusion (I/R) induces lung injury by a mechanism that involves neutrophils (PMNs). We have previously shown that endotoxin (LPS), when administered after gut I/R, amplifies this lung injury, while treatment with LPS prior to gut I/R prevents lung injury. The purpose of this study was to determine whether LPS pretreatment (Pre Rx) alters the PMN inflammatory component of the gut I/R injury. Specifically, we focused on whether LPS Pre Rx effected (i) PMN stem cell proliferation, (ii) gut I/R-induced PMN priming, and (iii) gut I/R-induced PMN lung sequestration. Bone marrow was harvested from normal and LPS-pretreated (0.5 mg/kg, ip, 3 days prior) rats, and colony forming units--granulocyte/macrophage (CFU-GM) were quantitated using a soft agar culture technique. In another experiment, normal and LPS-pretreated rats were subjected to gut I/R (45 min superior mesenteric artery occlusion/6 hr reperfusion), and blood and lungs were then harvested. The in vivo priming of PMN was assessed by measuring the difference in superoxide production (O2-) with and without the activating stimulus, N-formylmethionyl-leveyl-phenylalanine (fMLP). The quantity of myeloperoxidase (MPO) was used as an index of the number of PMN sequestered in lung tissue.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Pulmonary vascular smooth muscle relaxation by cGMP- versus cAMP-mediated mechanisms.

The purpose of this study was to examine and compare pulmonary vascular smooth muscle relaxation by clinically used agents as related to cGMP mediation (sodium nitroprusside, nitroglycerin) and cAMP mediation (isoproterenol, prostaglandin E1 (PGE1), and amrinone) in isolated rat pulmonary arterial rings. Isolated rat pulmonary arterial rings were suspended on a fine wire tensiometer in individual organ chambers. After confirming the endothelial integrity of the rings (response to acetylcholine), the rings were preconstricted with phenylephrine 10(-6) M. Dose-response curves for sodium nitroprusside, nitroglycerin, isoproterenol, amrinone, and PGE1 were then generated. Each of these agents was tested on six rings. Each ring was tested with each agent in a random order. The doses of sodium nitroprusside, isoproterenol, and nitroglycerin required to produce relaxation of isolated pulmonary arterial rings were not statistically different, but were significantly less than those required by amrinone and PGE1 (P < 0.05). These data suggest that relaxation of pulmonary vascular smooth muscle is more readily achieved via cGMP by guanylate cyclase activation (sodium nitroprusside, nitroglycerin) and via beta-adrenergic cAMP mediation (isoproterenol) than via cAMP-mediated pathways requiring either prostaglandin receptor activation (PGE1) or phosphodiesterase inhibition (amrinone).

Alprostadil↗
PubMed1994

Postoperative evaluation of pulmonary arteries in congenital heart surgery by magnetic resonance imaging: comparison with echocardiography.

Palliative and corrective operations for the treatment of cyanotic congenital heart disease frequently involve or potentially influence the size of the pulmonary arteries. Echocardiography and magnetic resonance imaging (MRI) are two noninvasive imaging techniques currently used to assess morphologic abnormalities of the pulmonary arteries. The purpose of this study was to evaluate the role of MRI in comparison with echocardiography for defining morphologic changes of the pulmonary arteries after congenital heart surgery. The MRI scans and echocardiograms of 33 patients with surgery involving or affecting the pulmonary arteries were compared. The pulmonary outflow tract, pulmonary confluence, right and left pulmonary arteries, and surgical shunts were separately evaluated. Cineangiography and surgical reports were used to confirm findings. MRI and echocardiography were equivalent for demonstrating abnormalities of the right ventricular outflow tract, main pulmonary artery, and a variety of pulmonary shunts. MRI was superior to echocardiography in demonstrating abnormalities of the right and left pulmonary arterial branches (p < 0.001). MRI is effective for monitoring pulmonary arterial status after surgery and is superior to echocardiography for the evaluation of the right and left pulmonary arteries.

Adolescent↗
PubMed1994

Nonlinearity of the left ventricular end-systolic wall stress-velocity of fiber shortening relation in young pigs: a potential pitfall in its use as a single-beat index of contractility.

OBJECTIVES: We sought to evaluate in the young heart the primary assumptions on which the current use of the mean "velocity of fiber shortening corrected for heart rate" as a noninvasive index of contractility are based. BACKGROUND: End-systolic wall stress-velocity of fiber shortening relation has been applied as a single-beat, load-independent index of contractility in children. This use is based on poorly validated assumptions of linearity, parallel shifts with changing contractile state and inotropic sensitivity of the end-systolic wall stress-velocity of fiber shortening relation. METHODS: In eight anesthetized young piglets, 5F mciromanometric catheters were placed in the ascending aorta and balloon occlusion catheters in the descending aorta. End-systolic wall stress and velocity of fiber shortening were calculated from aortic pressure and M-mode echocardiography under six conditions: in three contractile states 1) baseline, 2) increased contractility during dobutamine infusion (10 micrograms/kg per min), and 3) decreased contractility after propranolol injection (1 mg/kg), each at two afterload states (normal and increased load by partial aortic occlusion). RESULTS: Dobutamine increased and propranolol decreased afterload-matched velocity of fiber shortening corrected for heart rate significantly to 140% and 77% of baseline, respectively. However, the slope of end-systolic wall stress-velocity of fiber shortening relation was much greater (251% of baseline) during dobutamine infusion, which also significantly decreased wall stress, and was much less (27% of baseline) after propranolol injection, which increased wall stress. CONCLUSIONS: The velocity of fiber shortening corrected for heart rate did change predictably with changes in contractility and as such can be used noninvasively in the temporal evaluation of individual patients undergoing therapeutic interventions or to define the natural history of a disease process. However, the relation on which it is based is not defined by parallel straight lines across contractile states, so that abnormal single point measurements may reflect only the nonlinearity of the relation rather than abnormalities in contractility. Thus, we recommend that the end-systolic wall stress-velocity of fiber shortening relation should not be used as a single-beat index of contractility.

Aging↗
PubMed1994

Stereoselective epoxidation of 2,2-dimethyl-2H-1-benzopyran-6-carbonitrile.

The chiral intermediate (3S,4R)-trans-3,4-dihydro-3,4-dihydroxy-2,2-dimethyl- 2H-1-benzopyran-6-carbonitrile [(+)-trans diol 3] was made by the stereoselective microbial epoxidation of 2,2-dimethyl-2H-1-benzopyran-6-carbonitrile 1. This compound is a potential intermediate for the total synthesis of potassium-channel openers. Several microbial cultures were found which catalyzed the transformation of 1 to the corresponding (3S,4S)-epoxide 2 and (+)-trans diol 3. The two best cultures, Corynebacterium sp. SC 13876 and Mortierella ramanniana SC 13840 gave reaction yields of 32 M% and 67.5 M% and optical purities of 88 and 96%, respectively, for (+)-trans diol 3. A single-stage process (fermentation-epoxidation) for the biotransformation of 1 was developed using Corynebacterium sp. SC 13876 and M. ramanniana SC 13840. In a 25-L fermentor, the (+)-trans diol 3 was obtained in 38.6 M% yield with an optical purity of 90% using Corynebacterium SC 13876. The reaction yield of 60.7 M% and optical purity of 92.5% were obtained for (+)-trans diol 3 using M. ramanniana SC 13840. A two-stage process for the preparation of (+)-trans diol 3 was also developed using a 3 L cell-suspension (10% w/v, wet cells) of M. ramanniana SC 13840. The reaction was carried out in a 5-L Bioflo fermentor. The concentration of substrate 1 was 2 g L-1 with glucose present at 10 g L-1. After 48 h, (+)-trans diol 3 was obtained in 76 M% yield with an optical purity of 96%.(ABSTRACT TRUNCATED AT 250 WORDS)

Bacteria↗
PubMed1994

Gut ischemia/reperfusion produces lung injury independent of endotoxin.

OBJECTIVE: Bacterial translocation from the gut has been invoked as a common inciting event for postinjury multiple organ failure. We previously showed that gut ischemia/reperfusion induces remote organ injury. The purpose of this study was to ascertain if endotoxin has a pivotal mechanistic role in this process. DESIGN: Prospective, randomized study. SETTING: Animal laboratory. SUBJECTS: Sprague-Dawley rats weighing 300 to 350 g. INTERVENTIONS: Anesthetized animals underwent 45 mins of superior mesenteric artery occlusion and 2 hrs of reperfusion; sham laparotomy served as controls. Endotoxin was eliminated with the murine immunoglobulin (Ig) M antibody E5, 3 mg/kg i.v. before the study. MEASUREMENTS AND MAIN RESULTS: Plasma endotoxin was measured by the limulus amebocyte lysate assay. At 2 hrs of reperfusion, circulating neutrophil priming was determined by the difference in superoxide generation with and without the activating stimulus, N-formyl-Met-Leu-Phe. Neutrophil sequestration in the lung was quantitated by myeloperoxidase activity, and by lung endothelial permeability by 125I albumin lung/blood ratio. Endotoxin concentrations were not significantly (significance determined as p < .05) different between the gut ischemia/reperfusion and laparotomy groups (n = > or = 5) during ischemia or reperfusion. Circulating neutrophil priming, neutrophil accumulation in the lung, and lung injury were provoked by gut ischemia/reperfusion, but not altered by endotoxin elimination. CONCLUSION: Gut ischemia/reperfusion primes circulating neutrophils and produces lung injury by a mechanism independent of endotoxin.

Animals↗
PubMed1994

The postischemic gut serves as a priming bed for circulating neutrophils that provoke multiple organ failure.

Our trauma research center program entitled, "Trauma Primes Cells" is based on the fundamental hypothesis that prior exposure to multiple, sequential, sublethal cellular insults primes constructive or destructive pathways of cellular responses of subsequent injury. A major objective is to design therapy that will reduce the incidence of multiple organ failure. Although a number of inflammatory cascades have been incriminated in the pathogenesis of multiple organ failure (MOF), diffuse PMN-mediated tissue injury remains an attractive unifying concept. We have developed a sequential insult rodent model in which the priming event consisted of superior mesenteric arterial (SMA) clamping for 45 minutes followed by 6 hours of reperfusion. Following this priming event, activation was induced with a low dose of endotoxin (2.5 mg/kg). We believe that these studies support our hypothesis: mesenteric ischemic/reperfusion primes circulating PMNs. When these have been activated they can then be provoked by endotoxin to provoke distant organ injury. Primed PMNs are released from the postischemic mesenteric bed and enter the systemic circulation. They subsequently sequester in the pulmonary vascular bed where they are relatively harmless until they are activated by low dose endotoxin. These activated PMNs then migrate across the endothelium cell and release reactive oxygen metabolites.

Animals↗
PubMed1994

Trauma primes cells.

Trauma induces many dramatic and complex changes in host cellular response. This complexity arises from the constellation of signals induced by stress, infection, and injury. Cellular priming, defined as altered response to an agonist induced by an antecedent stimulus, appears to be operative after trauma. If the interaction between two signalling pathways is such that one augments (or depresses) the other, subsequent stimulation of the second "primed" pathway can result in an exaggerated (or attenuated) response. Thus, trauma can prime cells in either a constructive or destructive fashion. Receptor stimulation by priming agents results in the activation of receptor-specific cell signalling pathways. These intracellular signalling pathways can "crosstalk" modifying each other in either a positive of negative fashion. The positive or negative character of interpathway crosstalk eventually manifests itself physiologically as constructive or destructive priming. Could the characteristics of the initial priming predict the sense of the final cellular message? Although the magnitude of both the priming stimulus and subsequent stimuli are important, the temporal relationship between the two stimuli as well as the positive or negative character of their interacting signal pathways need to be considered. It is unclear whether any one characteristic alone determines the sense of the priming effect. In general, it is the interaction of two stimuli with a cell at all three levels (magnitude, character, and temporal relationship) that dictates the final cellular response.

Animals↗