PubMed Health⌕ Search

Biomedical subjects

M Patel

Publications and source records attributed to M Patel.

At least 19 recordsLinked to original sources
PubMed1992

Transfection of mouse cytotoxic T lymphocyte with an antisense granzyme A vector reduces lytic activity.

Murine CTL have seven serine proteases, known as granzymes, in their lytic granules. Despite considerable effort, convincing evidence that these enzymes play an obligatory role in the lytic process has not been presented. To investigate the function of one of these proteases, granzyme A (GA), we utilized an antisense expression vector to lower the level of the enzyme in the cells. An expression vector containing antisense cDNA for GA and the gene for hygromycin B resistance was constructed and electroporated into the murine CTL line, AR1. Transfectants were selected based on resistance to hygromycin B, and a number of stable lines were developed. One of the antisense lines had greatly reduced levels of GA mRNA, when compared to the parental cells or to control lines transfected with the vector lacking the antisense DNA. The message levels for two other CTL granule proteins, granzyme B and perforin, were unaffected by the antisense vector. The amount of GA, as measured by enzymatic activity, was 3- to 10-fold lower in the transfectant. Most significantly, this line also consistently showed 50 to 70% lower ability to lyse nucleated target cells and to degrade their DNA. Furthermore, it exhibited 90 to 95% lower lytic activity to anti-CD3-coated SRBC. Conjugate formation with target cells, however, was normal. These data provide strong evidence that GA plays an important role in the cytolytic cycle, and that the quantity of enzyme is a limiting factor in these cytolytic cells.

Animals↗
PubMed1992

Isolation, characterization and biological activity of a CRF-related diuretic peptide from Periplaneta americana L.

A diuretic peptide (Periplaneta-DP) has been isolated from extracts of whole heads of the cockroach, Periplaneta americana. The purified peptide increases cyclic AMP production and the rate of fluid secretion by isolated Malpighian tubules in vitro. In the fluid secretion assay, the response to native Periplaneta-DP is comparable to that obtained with crude extracts of cockroach corpora cardiaca, and the EC50 lies between 10(-8) and 10(-9) M. The primary structure of Periplaneta-DP was established as a 46-residue amidated peptide: T G S G P S L S I V N P L D V L R Q R L L L E I A R R R M R Q S Q D Q I Q A N R E I L Q T I-NH2. Periplaneta-DP is a further member of the recently established family of CRF-related insect diuretic peptides.

Amino Acid Sequence↗
PubMed1992

Evaluation of hydrogen-bonding and enantiomeric P2-S2 hydrophobic contacts in dynamic aspects of molecular recognition by papain.

1. 2-(N'-Acetyl-D-phenylalanyl)hydroxyethyl 2'-pyridyl disulphide (compound IV) (m.p. 59 degrees C; [alpha]D20 -6.6 degrees (c 1.2 in methanol)) was synthesized. 2. The results of a study of the pH-dependence of the second-order rate constant (k) for its reaction with the catalytic-site thiol group (Cys-25) of papain (EC 3.4.22.2) together with analogous kinetic data for the reactions of related time-dependent inhibitors, notably the L-enantiomer of compound (IV) (compound III) and the L- and D-enantiomers of 2-(N'-acetylphenylalanylamino)ethyl 2'-pyridyl disulphide (compounds I and II respectively), were used to assess the contributions of the (P1)-NH ... O = C < (Asp-158) and (P2) > C = O ... H-N-(Gly-66) hydrogen bonds and enantiomeric P2-S2 hydrophobic contacts in two manifestations of dynamic molecular recognition in papain-ligand association: (a) signalling to the catalytic-site region to provide for a (His-159)-IM(+)-H-assisted transition state and (b) the dependence of P2-S2 stereoselectivity on hydrogen-bonding interactions outside the S2 subsite. The analysis involved determination of the reactivities of individual ionization states of the reactions (pH-independent rate constants, k) and associated macroscopic pKa values and difference kinetic specificity energies (delta delta GKS = -RT1n(k1/k2), where k1 is the pH-independent second-order rate constant for reaction with one inhibitor and k2 is the analogous rate constant in the same ionization state for reaction with another inhibitor so that, when the structural change provides that k2 > k1, delta delta GKS is positive. 3. The kinetic data further illuminate the nature of the interdependence of binding interactions in papain first noted by Kowlessur, Topham, Thomas, O'Driscoll, Templeton & Brocklehurst [(1989) Biochem. J. 258, 755-764] in the S2 subsite, S1-S2 intersubsite and catalytic-site regions. Of particular note is the apparent dependence of the binding of the N-Ac-D-Phe moiety on the binding of the leaving group to (His-159)-Im+H and the fact that the resulting rate enhancement is more effective when (P1)-N-H is absent than when it is present. This result revealed by kinetic analysis goes beyond the conclusion suggested by model building that it is possible to make all of the binding contacts in complexes involving the D-enantiomers [(II) and (IV)] as in those involving the L-enantiomers [(I) and (III)].(ABSTRACT TRUNCATED AT 400 WORDS)

Catalysis↗
PubMed1992

Septin: a factor in plasma that opsonizes lipopolysaccharide-bearing particles for recognition by CD14 on phagocytes.

We have previously reported that lipopolysaccharide (LPS) binding protein (LBP) opsonizes endotoxin (LPS) for recognition by CD14 on phagocytes. Here we show that normal human plasma contains high titers of an activity that also binds LPS (Re, 595) and mediates recognition by CD14. Opsonization of LPS-coated particles with plasma enables the particles to be bound by phagocytes. Further, opsonization with plasma also enables subnanogram-per-milliliter concentrations of LPS to induce dramatic alterations in the function of leukocyte integrins on polymorphonuclear leukocytes and to induce secretion of tumor necrosis factor by monocytes, suggesting that opsonization by factors in plasma may be important in responses of cells to endotoxin. The opsonic activity in plasma appears distinct from LBP since it is not blocked by neutralizing antibodies against LBP. Surprisingly, the opsonic activity of plasma is not present in a single protein species, but at least two species must be combined to observe activity. Further, the opsonic activity of plasma for LPS is blocked by addition of protease inhibitors, suggesting that proteolytic activity or activities are required for opsonization. These properties are suggestive of the action of a protease cascade, but opsonic activity of plasma is not affected by blockade or depletion of either the complement or clotting cascades. We propose the name "septin" to describe this novel LPS-opsonizing activity in plasma.

Acute-Phase Proteins↗
PubMed1992

Mediated Na(+)-independent transport of L-glutamate and L-cystine in 1- and 2-cell mouse conceptuses.

L-Glutamate and L-cystine appeared to compete for transport via a mediated Na(+)-independent transport process in 1- and 2-cell conceptuses. Not only did these substances competitively inhibit each others' uptake by conceptuses, but their Ki values for inhibition were about equal to their Km values for transport in 1-cell conceptuses. Moreover, the transport process interacted strongly with L-amino acids that had 3-6 atoms in a chain between their negatively charged groups, whereas it interacted weakly or not at all with amino acids that did not have these characteristics or that were N-methylated. Transport of anionic amino acids was not altered greatly by pH in the range 4.5-8.0, but transport of L-cystine was much faster at higher pH values. The slower cystine transport at lower pH values was due primarily to protonation of its second amino group. A small increase in the degree of deprotonation of cystine's carboxyl groups also probably contributed slightly to its faster transport at higher pH values. By all of these criteria, the transport process in conceptuses appears to be a form of amino acid transport system xc-. System xc- activity was several-fold higher in 1- than in 2-cell conceptuses. Similarly, L-glutamate uptake by unfertilized eggs was relatively rapid, whereas it was much slower in immature, fully-grown oocytes. System xc- activity in 1-cell conceptuses also appeared to increase in response to the oxidative stress of culture, whereas no such increase was observed for 2-cell conceptuses. We suggest that this transient increase in the activity of system xc- activity during development of 2-cell conceptuses from immature, fully-grown oocytes could help protect unfertilized and fertilized eggs from oxidative stresses in situ.

Amino Acids↗
PubMed1992

Variation in the P2-S2 stereochemical selectivity towards the enantiomeric N-acetylphenylalanylglycine 4-nitroanilides among the cysteine proteinases papain, ficin and actinidin.

1. Values of the kinetic specificity constant, kcat./Km, for the hydrolysis of N-acetyl-L-phenylalanylglycine 4-nitroanilide (I) and of its D-enantiomer (II) catalysed by ficin (EC 3.4.22.3) and by actinidin (EC 3.4.22.14) at pH 6.0, I 0.1 mol/l, 8.3% (v/v) NN-dimethylformamide and 25 degrees C were determined by using initial-rate data with [S] much less than Km and weighted nonlinear regression analysis as: for ficin, (kcat./Km)L = 271 +/- 6 M-1.s-1, (kcat./Km)D = 2.9 +/- 0.1 M-1.s-1, and for actinidin (kcat./Km)L = 13.3 +/- 0.7 M-1.s-1, (kcat/Km)D = 0.34 +/- 0.01 M-1.s-1.2. These data and analogous values for the corresponding reactions catalysed by papain (EC 3.4.22.2), (kcat./Km)L = 2064 +/- 31 M-1.s-1, (kcat./Km)D = 5.5 +/- 0.1 M-1.s-1, demonstrate marked variation in stereochemical selectivity for substrates (I) and (II) among the three cysteine proteinases with the following values for the index of stereochemical selectivity Iss = (kcat./Km)L/(kcat./Km)D: for papain, 375; for ficin 93; for actinidin 39. 3. Model building suggests ways in which, for the papain-catalysed reactions, binding interactions involving the extended acyl groups of the substrates may need to change as the reaction proceeds from adsorptive complex (ES) to tetrahedral intermediate (THI) before its rate-determining, general acid-catalysed collapse to acylenzyme intermediate. In particular, satisfactory alignment in the catalytic site at the THI stage of the acylation process appears to demand rotation of the substrate moiety about its long axis. 4. The different consequences of this rotation for the L- and D-enantiomers suggest that for closely related systems the greater the extent of this rotational adjustment the greater would be the value of Iss.5. For the actinidin-substrate combinations, model building suggests that even at the ES complex stage of catalysis it is not possible to approach optimized P2-S2 contacts and the three hydrogen-bonding interactions deduced for papain-ligand complexes in the absence of significant movement of protein conformation. Possible binding modes in which some of the interactions deduced for papain are relaxed are discussed. Consideration of postulated binding modes in the various transition states is shown to account for the order of reactivity reflected in values kcat./Km for the four reactions involving papain (Pap) and actinidin (Act) with the L- and D-enantiomeric substrates: Pap-L much greater than Act-L greater than Pap-D much greater than Act-D.(ABSTRACT TRUNCATED AT 400 WORDS)

Anilides↗
PubMed1992

Experimental evaluation of ten clinically used arterial prostheses.

The arterial grafts currently in use are classified into five basic categories; 10 different commercially available prostheses were chosen to represent these categories. The Vascutek and Bionit are made from knitted Dacron and have medium porosity, requiring preclotting by the surgeon. The DeBakey Soft Woven and Plasma TFE grafts are made from woven Dacron and have low porosity, not requiring preclotting under most circumstances. Also studied were woven and knitted grafts with leakage resistance referred to as "impervious" Dacron grafts: the Vascutek Gelseal, the Bard Albumin Coated DeBakey Vasculour II, the Microvel with Hemashield, and the albumin saturated, autoclaved DeBakey Soft Woven graft. Gore-Tex and Impra are expanded polytetrafluoroethylene grafts which do not require preclotting. For each type, five grafts 6 cm long and 8 mm in diameter were implanted in the descending thoracic aorta of healthy adult dogs for 16 weeks. The physical characteristics, biocompatibility, and healing patterns varied according to the structure and treatment of the grafts. Pretreatment with biomaterials during manufacture is quite effective in preventing transinterstices blood loss during implantation, but results in altered physical qualities, increased thrombogenicity and delayed healing in comparison to the effects of preclotting with autogenous blood at the time of implantation.

Anastomosis, Surgical↗
PubMed1992

Comparative evaluation of the elasticity and flexibility of bioimpregnated knitted grafts.

Longitudinal elasticities of whole human blood clot, whole canine blood clot, Factor XIII cross-linked fibrin, glutaraldehyde-fixed human albumin, and formaldehyde-fixed collagen, gelatin and collagen/gelatin were determined and normalized to human whole blood clot. Matrices of a knitted Dacron graft were then impregnated with albumin, collagen, and collagen/gelatin and their longitudinal elasticities were determined and normalized to a preclotted graft. Comparisons were also made for the longitudinal elasticities of a virgin graft, a manipulated control for a preclotted graft, and a manipulated control for the matrix-impregnated grafts. Flexibilities were then calculated based on the weights and elasticities of these grafts and normalized to the flexibility of the preclotted graft. Fibrin had twice and 39 times more longitudinal elasticity than human blood clot and collagen, respectively. The preclotted graft has longitudinal elastic properties similar to a virgin graft, and is 2.8, 2, and 1.4 times more elastic than the albumin, collagen and collagen/gelatin grafts, respectively. The preclotted graft was 2.5, 2, and 1.4 times more flexible than the albumin, collagen and collagen/gelatin grafts, respectively.

Albumins↗
PubMed1992

Inhibitory action of orotate, 2-thioorotate and isoorotate on nucleotide metabolism and nucleic acid synthesis in hepatoma cells.

1. The specificity of the action of orotate on hepatoma cells was investigated. 2. Orotic acid and its methyl ester had similar inhibitory effects on the incorporation of [3H]thymidine into DNA of hepatoma cells. 3. In contrast to previous studies in vivo, incubation of rat kidney cells with orotate caused an increase in the ratio of UTP/ATP concentrations that was similar to effects on hepatic cells. 4. Inhibitory effects of 2-thioorotate and isoorotate on metabolism were found to be less selective and required higher concentrations than with orotate.

Animals↗
PubMed1992

The impact of automated blood pressure devices on the efficiency of clinical trials.

By reducing measurement error, automated blood pressure (BP) devices should enhance the precision of BP estimation and thereby decrease sample size requirements in clinical trials of BP-lowering therapy. Enhanced precision would be particularly relevant to clinical trials assessing the efficacy of nonpharmacological therapies. In the present investigation, resting (clinic) BPs by the Dinamap 8100 (a stationary device) and the Accutracker II (an ambulatory device) were as precise as manual BPs given an equal number of observations by each method. However, both the Dinamap and Accutracker devices underestimated resting diastolic BP in comparison to the manual observers. Estimates of average daytime and 24-hour ambulatory BP, based on large numbers of observations over an extended period of time, were extremely precise. These findings suggest that the use of automated devices to measure resting BP may not reduce samples sizes, whereas use of ambulatory BP devices should reduce samples sizes considerably.

Bias↗
PubMed1992

A simplified technique to resect abnormal bony radiolocalizations using a gamma counter.

A simplified technique for localizing and verifying the correct biopsy site of lesions identified on a bone scan has been utilized. A hand-held gamma counter was used for localization of incision placement, determination of extent of bone to be resected, and verification that appropriate tissue was resected. This technique was used to guide biopsy of bony lesions in five patients and to guide resection of a pubic ramus chondrosarcoma. We conclude that intraoperative use of a gamma counter to guide biopsy of bony lesions minimizes surgery time, increases the confidence of obtaining correct tissue, and makes a frequently frustrating procedure very simple. In addition, the probe may assist with determining adequate margins at definitive resection of tumours which accumulate technetium-99m MDP.

Adult↗
PubMed1992

Variability of acetaminophen metabolism in Caucasians and Orientals.

Acetaminophen (paracetamol) is extensively conjugated with glucuronic acid and sulfate prior to renal excretion. A minor metabolic route involves microsomal oxidation of acetaminophen to a hepatotoxic reactive intermediate, which subsequently undergoes glutathione (GSH) conjugation, yielding cysteine and mercapturate conjugates, both of which are excreted in the urine (Slattery et al., 1987). Data collected by de Morais et al. (1989) indicated that in comparison with normal subjects, glucuronidation of acetaminophen was impaired in subjects with Gilbert's syndrome, a genetically-based impairment of bilirubin glucuronidation. Thus, inter-subject and ethnic differences in acetaminophen disposition have pharmacogenetic and toxicological implications. This study was conceived to explore these differences. Urinary excretion of acetaminophen and its metabolites was observed in 125 Caucasian and 33 Oriental subjects. No appreciable difference was noted in the mean fraction of drug excreted as glucuronide between the two groups (51.5% in Caucasians vs 51.8% in Orientals). However, the data strongly indicated that the excretion of acetaminophen glucuronide was not normally distributed. Bimodality was apparent in both groups, with 20% of Caucasian and 33% of Oriental subjects displaying relatively extensive glucuronidation. In addition, glucuronidation displayed a strong negative correlation with sulfation (r = -0.97), suggesting the existence of a compensatory mechanism between the two metabolic pathways. The mean fractional excretions of cysteine and mercapturate conjugates did show significant differences between Caucasians and Orientals (p < 0.005). In addition, the ratio of mercapturate to total GSH-derived conjugates recovered appeared to be bimodal, indicating possible heterogeneity in the conversion of the cysteine conjugate to mercapturate via N-acetylation.

Acetaminophen↗
PubMed1992

Functional comparisons of gastrin/cholecystokinin receptors in isolated preparations of gastric mucosa and ileum.

1. The gastrin cholecystokinin (CCK) receptors mediating stimulation of acid secretion in rat isolated gastric mucosa (RGM) and contraction in guinea-pig isolated ileum longitudinal muscle-myenteric plexus (GPI) have been characterized by use of peptide agonists and the non-peptide antagonists, lorglumide, devazepide and L-365,260. 2. In RGM, gastrin peptides (sulphated gastrin heptadecapeptide (G-17), non-sulphated (ns) G-17 and pentagastrin) were potent agonists of acid secretion (EC50 values of 4.3, 16 and 27 nM respectively). Sulphated CCK octapeptide (CCK-8) was also a potent agonist, (EC50 = 0.9 nM), but was less efficacious, producing a lower maximal response. In contrast, in GPI, CCK-8 was a potent full agonist (EC50 = 1.4 nM) and was more than 1000 times more potent than the gastrin peptides in producing a sustained contractile response. 3. In GPI, CCK-8 (0.1 to 100 nM) produced sustained contractile responses, whilst CCK-4 (3 to 1000 nM) produced transient responses. These responses had different sensitivities to atropine (1 microM), suggesting that more than one receptor may mediate contraction in this tissue. 4. In RGM, L-365,260 was the most potent antagonist of pentagastrin-stimulated acid secretion (pA2 = 7.6). This functional affinity estimate was similar to that for L-365,260 as an antagonist of excitatory responses in rat ventromedial hypothalamic slices (Kemp et al., 1989) but differed from binding affinity estimates in guinea-pig cortex and gastric glands (Freidinger, 1989). 5. In GPI, devazepide, L-365,260 and lorglumide yielded different affinity estimates when compared against CCK-8 and CCK-4 or pentagastrin respectively.These studies were consistent with the view that the sustained response produced by CCK-8 was mediated by CCKA receptors and the transient response produced by CCK-4 and pentagastrin was mediated by CCKB receptors.6. Affinity estimates for L-365,260 and lorglumide against CCK-4 or pentagastrin in GPI were significantly different from corresponding estimates against pentagastrin in RGM. These studies are consistent with the view that gastrin/CCKB receptors in GPI may differ from those in RGM.

Animals↗
PubMed1992

Clinical-radiographic correlations within the first five hours of cerebral infarction.

Fifty patients, ages 54-79, with ischemic hemispheric strokes productive of hemiparesis, at a minimum, underwent standardized neurological evaluations, computed tomographic scanning and cerebral angiography (N = 38) or carotid ultrasound (N = 12) within 5 h of onset. A second scan was performed at 5-7 days. Clinical scores were not associated with a history of, or the presence of: hypertension, smoking or cardiac disease, including atrial fibrillation, nor with severe internal carotid artery stenosis or occlusion. Clinical scores were adversely affected by early scan abnormalities (especially mass effect), lesion size, intracranial arterial occlusions, elevated serum glucose levels and the subsequent development of hemorrhagic infarction. Glucose levels correlated with infarct size and the development of hemorrhagic infarction. Delayed intracranial arterial filling and collateral flow were associated with reduced infarct size but did not confer clinical protection. We believe that combining the initial glucose level and scan results has prognostic significance, and early angiography is valuable in characterizing infarct etiology and assessing clinical severity.

Aged↗