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Biomedical subjects

J Luo

Publications and source records attributed to J Luo.

At least 325 records · Page 18Linked to original sources
PubMed1995

Identification and analysis of domains in proteins.

An automatic algorithm based on inter-residue contacts is presented to identify domains in proteins. The results of the algorithm are compared to an assignment performed by inspection that was guided by the authors' description in the literature. The authors' and the algorithm's assignments for a chain were considered to agree if the same number of domains were identified and if the assignments were the same for at least 95% of the residues. With this criterion, the algorithm agreed with the authors' assignment for 78% of the 284 non-redundant chains considered. When some of the authors' assignments were re-evaluated based on the results of the algorithm, an agreement of 84% was obtained. The algorithm is therefore a useful tool for data validation in domain assignment. The authors assignments of domains were analysed for structural principles of domains. The number of chains forming one, two, three, four and five domains are 197, 67, 13, 6 and 1 respectively. Most domains in multidomain proteins are formed from continuous segments and adopt the same structural class. Distributions of the number of residues and the ellipticity of domains and chains are presented. The relationship between accessible surface area and molecular weight for domains and chains is examined.

Algorithms↗
PubMed1995

NMDA prevents alcohol-induced neuronal cell death of cerebellar granule cells in culture.

Neuronal cell loss is one of the most debilitating effects of alcohol exposure during development of the nervous system. In this study, primary cultures of neuronal cells (cerebellar granule cells) were used to examine mechanisms of alcohol-induced neuronal cell death. Previously, we established that (Pantazis et al., Alcohol Clin Exp Res 17:1014-1021, 1993): (1) alcohol exposure caused neuronal cell death in cultures of cerebellar granule cells and this cell loss was both time-dependent and dose-dependent; and (2) the vulnerability of cerebellar granule cells to alcohol-induced loss changed with the length of time the cells were in culture before initiating alcohol exposure-that is, younger cultures (1 day in vitro) were much more susceptible to alcohol-induced neuronal cell death than older cultures (4 or 7 days in vitro). The primary goal of the present study was to examine the potential role of the NMDA receptor in alcohol-induced death of cerebellar granule cells in culture. Experiments were performed to test the hypothesis that the alcohol-induced death of cerebellar granule cells can be prevented or reduced by NMDA treatment. Our results indicate that stimulation of the NMDA receptor has a neuroprotective effect and can significantly reduce the alcohol-induced neuronal cell death of newly established cerebellar granule cell cultures. This neuroprotective effect of NMDA is blocked by 2-amino-5-phosphonovalerate, a competitive inhibitor of the NMDA receptor, confirming that this neuroprotective effect is mediated via the NMDA receptor. This is the first report that alcohol's neurotoxic effect can be ameliorated by activation of the NMDA receptor.

Animals↗
PubMed1995

Polypyrimidine tract-binding protein and heterogeneous nuclear ribonucleoprotein A1 bind to human T-cell leukemia virus type 2 RNA regulatory elements.

Efficient expression of human T-cell leukemia virus (HTLV) and human immunodeficiency virus structural proteins requires Rx and Rev proteins, respectively. Decreased expression of Gag and Env appears to be due, in part, to intragenic RNA sequences, termed cis-acting repressive sequences (CRS), and may be mediated by binding of specific cellular factors. We demonstrated previously that two cellular proteins, p60CRS and p40CRS, interact with HTLV type 2.5' long terminal repeat CRS RNA and that the interaction of both proteins with CRS RNA correlates with function (A. C. Black, C. T. Ruland, J. Luo, A. Bakker, J. K. Fraser, and J. D. Rosenblatt, Virology 200:29-41, 1994). By radioimmunoprecipitation of HeLa nuclear proteins UV cross-linked to CRS RNAs with murine monoclonal antibodies, we now show that p40CRS is heterogeneous nuclear ribonucleoprotein (hnRNP) A1 and p60CRS is polypyrimidine tract-binding protein or hnRNP I. These immunoprecipitation results were confirmed by an immunobinding assay with hnRNP I and hnRNP AI antibodies and by cross-competition electrophoretic mobility shift experiments. In addition, we mapped a putative hnRNP A1 binding site in U5 RNA and demonstrated that p40CRS (hnRNP A1) binding to that site correlates with CRS function. Since both hnRNP I and hnRNP A1 have been shown to influence splicing and potentially other steps in RNA processing, the binding of both hnRNP I and hnRNP A1 to HTLV RNA regulatory elements may alter retrovirus RNA processing and may be involved in regulation by Rex.

Base Sequence↗
PubMed1995

Towards an intelligent system for the automatic assignment of domains in globular proteins.

The automatic identification of protein domains from coordinates is the first step in the classification of protein folds and hence is required for databases to guide structure prediction. Most algorithms encode a single concept based and sometimes do not yield assignments that are consistent with the generally accepted perception. Our development of an automatic approach to identify reliably domains from protein coordinates is described. The algorithm is benchmarked against a manual identification of the domains in 284 representative protein chains. The first step is the domain assignment by distance (DAD) algorithm that considers the density of inter-residue contacts represented in a contact matrix. The algorithm yields 85% agreement with the manual assignment. The paper then considers how the reliability of these assignments could be evaluated. Finally the use of structural comparisons using the STAMP algorithm to validate domain assignment is reported on a test case.

Algorithms↗
PubMed1995

[The regulation of deep peroneal nerve inputs on depressor response evoked by caudal ventrolateral medulla excitation in rabbits].

Experiments were performed on 33 white male rabbits anaesthetized with chloralose and urethane under artificial ventilation. The results were as follows: (1) depressor response could be evoked by electric stimulation of caudal ventrolateral medulla (cVLM). The inputs of deep peroneal nerve (DPN) could inhibit partly the depressor response evoked by electric stimulation of cVLM (P < 0.001). The regulation effects of DPN disappeared in 25-30 min. (2) after iv naloxone (1.5 mg/kg) or microinjection of naloxone into rostral ventrolateral medulla (rVLM) (2 micrograms/1 microliter NS), the inhibitory effect of DPN inputs was abolished partly (P < 0.01, P < 0.001). These results suggest that: (1) the depressor response elicited by cVLM excitation in rabbits can be regulated by inputs of DPN; (2) opiate peptide in rVLM may take part in the inhibitory effect of DPN inputs on depressor response excited by cVLM.

Animals↗
PubMed1995

[Experience in management of capsular contraction through areolar incision after implantation of prosthesis for augmentation mammaplasty].

Since 1990, 19 cases of capsular contraction of Baker's III, IV have been treated with partial excision of the capsule and replacement of implants through areolar incision. The result was satisfactory. After six months to eighteen months follow-up observation, no capsular contraction was again found. All patients were graded Baker's I. Prevention of capsular contraction was also discussed.

Adult↗
PubMed1994

Effect of E-type prostaglandins on bradykinin-induced plasma extravasation in the knee joint of the rat.

We studied the effect of different E-type prostaglandins on an experimental model of inflammation in the rat. Plasma extravasation was induced in the knee joint of the rat by continuous perfusion of two potent inflammatory mediators, bradykinin (160 nM) or platelet activating factor. Both prostaglandin E1 and prostaglandin E2 (0.5-500 ng ml-1), when perfused with bradykinin, produced a similar dose-dependent enhancement of plasma extravasation. Prostaglandin E2 (0.5-500 ng ml-1) also dose dependently enhanced plasma extravasation induced by platelet activating factor, while prostaglandin E1 significantly enhanced platelet activating factor-induced plasma extravasation only at concentrations above 5 ng ml-1. In contrast, co-perfusion of bradykinin or platelet activating factor with the prostaglandin E1 analogues, enisoprost and misoprostol (0.5-500 ng ml-1) did not enhance plasma extravasation. In fact, misoprostol attenuated plasma extravasation induced by bradykinin. These results demonstrate that in the rat knee joint, misoprostol and enisoprost have different pharmacological actions compared to their parent compound, prostaglandin E1 and to prostaglandin E2.

Animals↗
PubMed1994

Binding of nuclear proteins to HTLV-II cis-acting repressive sequence (CRS) RNA correlates with CRS function.

The shift from viral regulatory to structural gene expression in human T-cell leukemia virus types I (HTLV-I) and II (HTLV-II) is mediated by Rex. We have previously shown that HTLV-II Rex acts through an element in R/U5 of the 5' long terminal repeat (LTR), the Rex-responsive element (RxRE), and that Rex protein binds to specific RNA sequences, the Rex binding element (RBE), contained within the RxRE (Black et al., J. Virol. 65, 6645-6653, 1991b). Rex action through the RBE (nt 405-520) overcomes the inhibition of expression conferred by a contiguous LTR RNA regulatory element, which contains cis-acting repressive sequences (CRS; nt 520-630) that are not bound by Rex protein (Black et al., Virology, 181, 433-444, 1991a). We now show by electrophoretic mobility shift assay (EMSA) that cellular proteins in a HeLa nuclear extract bind specifically to RNA transcripts containing the HTLV-II CRS. Using ultraviolet (uv) crosslinking of gel-retarded bands, we identified a major protein species of approximately 60 kDa, p60CRS, that binds to CRS RNA and, with weaker affinity, to RBE RNA. In addition, a distinct 40-kDa protein, p40CRS, binds to U5 RNA (nt 645-750) downstream from the CRS. Specific deletions within CRS RNA can reduce or abrogate binding to this 60-kDa protein. EMSA and uv crosslinking assays also suggest that both p60CRS and p40CRS interact with CRS RNA. CRS function in a 5' LTR-linked gene expression assay correlates with the ability of both p60CRS and p40CRS to interact with 5' LTR RNA in vitro.

Base Sequence↗
PubMed1994

Inhibition of the regulation of intracellular pH: potential of 5-(N,N-hexamethylene) amiloride in tumour-selective therapy.

The viability of cells within the acidic microenvironment found in solid tumours is expected to depend on the regulation of intracellular pH (pHi). 5-(N,N-hexamethylene) amiloride (HMA) is a potent inhibitor of the Na+/H+ antiport, a major mechanism for the regulation of pHi. We have therefore studied the cytotoxicity of HMA in combination with nigericin, a cell-acidifying agent, for EMT-6 cells in monolayer cell culture, in spheroids and in a murine tumour model. The combination of nigericin and HMA was toxic to cells in tissue culture at extracellular pH (pHe) < or = 6.8 (as may be found in tumours) but not at pH 7.0 or above (as in most normal tissues). Compared with amiloride, the relative potency of HMA in causing in vitro cytotoxicity (approximately 100-fold) was similar to that for inhibition of the Na+/H+ antiport. The fluorescent probe Hoechst 33342 was used with flow cytometry to study the cytotoxicity of HMA and nigericin at different depths in multicellular tumour spheroids. Only small differences in the level of cell survival were observed, but higher concentrations of HMA were required as compared with those giving equal levels of survival in monolayer culture. The pharmacokinetics of HMA in mice was studied by using high-performance liquid chromatography: after intraperitoneal injection of 20 micrograms g-1, the plasma level of HMA peaked at 8 microM after about 15 min and decreased to 1 microM at 120 min; the half-life was 35 min. Nigericin and HMA, at doses of 1.25 micrograms g-1 and 10 micrograms g-1 respectively, failed to cause significant cell killing in the EMT-6 murine tumour, but the surviving fraction was reduced to approximately 0.004 when hydralazine was administered with nigericin and HMA. Local tumour irradiation (15 Gy), followed by treatment with these drugs, led to cell killing that was additive to the effects of drugs and radiation alone, so that hypoxic cells which survived radiation did not appear more sensitive to pH-dependent drug treatment. Acid-mediated therapy can lead to cell death in murine solid tumours, but further measures will be required before the strategy can be exploited clinically.

Amiloride↗
PubMed1994

[Cloning and sequencing of promoter and signal sequence coding regions from Bacillus subtilis].

Promoter and signal sequence coding regions from B. subtilis were cloned in E. coli using a bifunctional and signal sequence selection plasmid pGPB14 as a vector. The Sau3A digested chromosome DNA was ligated with BamHI digested pGPB14. The ligated mixture was used to transform E. coli C600. Ampicillin and erythromycin resistant clones were selected. Recombinant plasmids were isolated from double resistant transformants. Restriction analysis showed that inserts of various length had been cloned and these inserts rendered the E. coli cells resistant to different concentrations of ampicillin. The recombinant plasmids were transformed and showed the same secretion function in B. subtilis. The amount and localization of beta-lactamase were determined in transformed E. coli and B. subtilis. The results indicate that the beta-lactamase activities of E. coli mainly in periplasm while the enzyme produced by B. subtilis secreted extracellularly. 10 of the cloned fragments were sequenced by Sanger's dideoxy chain termination method. The sequencing data show that all fragments contain promoter, ribosome binding site and signal sequence coding region.

Amino Acid Sequence↗
PubMed1994

Endocrine and carbohydrate responses to interleukin-6 in vivo.

The role of interleukin-6 (IL-6) in carbohydrate metabolism beyond its inhibition of hepatic phosphoenolpyruvate carboxykinase has not been widely pursued. To describe such IL-6 effects, we examined in the rat the responses of plasma corticosterone, glucagon, insulin, and glucose levels and the hepatic glycogen content 30, 60, 90, 120, and 180 min after intravenous injection of recombinant human IL-6. The effect to increase plasma corticosterone was consonant with the well-known action of IL-6 on the hypothalamus-pituitary-adrenal cortex. IL-6 produced a transient increase in plasma glucagon that was mirrored by elevated plasma glucose and a depletion of hepatic glycogen. Plasma insulin levels were not elevated within the first hour after IL-6 injection but were significantly elevated 90 min and beyond. We suggest that the stimulus for increased circulating insulin was elevated plasma glucose, rather than a direct effect of IL-6. The results demonstrate that IL-6, acting directly on peripheral organs and/or through the central nervous system (CNS) can alter the hormonal and carbohydrate milieu. We propose that these actions of IL-6 are one aspect of its role in the acute phase response.

Animals↗
PubMed1994

[Anatomy of the eyebrow and upper eyelid region and its significance in ptosis surgery].

The article reports the results of the anatomic observations from dissecting 30 upper eyelids and 8 eyebrows of adult cadavers. In the eyebrow region, the anterior and posterior sheaths of the galea enclose the muscle plane where the frontalis, orbicularis and corrugator muscles are interdigitated. The posterior sheath attaches to supraorbital ridge through the brow fat pad. It was found that a few fibers of the frontalis muscle insert to the skin of the brow. In the upper eyelid, the orbital septum fuses with the aponeurosis at or just below the upper margin of the tarsus. The levator aponeurosis, Muller's muscle, particularly their attachments were carefully observed and measured. Finally, the application of the anatomy in the ptosis surgery is discussed.

Blepharoptosis↗
PubMed1994

Expression and secretion of alpha-amylase and glucoamylase in Saccharomyces cerevisiae.

alpha-Amylase genes of Bacillus licheniformis and glucoamylase cDNA of Aspergillus niger were ligated to a E. coli-yeast shuttle vector. The resultant plasmid was used to transform Saccharomyces cerevisiae to construct starch-degrading yeast strain. The results of enzyme activity assay and enzyme property analysis show that alpha-amylase and glucoamylase genes have been expressed simultaneously in yeast under the control of promoters and terminators of yeast MF-alpha 1 factor and PGK genes and over 99% of enzyme activities were secreted to the medium. The engineered yeast strain hydrolyses 97% of the starch (10%) in the medium after 6 days. The recombinant plasmid exists stably in yeast.

Aspergillus niger↗
PubMed1994

[ELISA for measurement of human serum apolipoprotein A II].

A specific, sensitive and simple ELISA for the measurement of human serum apo A II has been developed. The monospecific antibody was raised in goats. The polytyrene plates coated with purified anti-apo A II goat gamma-globulin together with enzyme labelled goat antibodies against human apo A II conjugate were used in this assay. The conjugate was obtained by binding horseradish perioidate by a simplified periodase method. No cross-reactivity with human apo A I, B100, C I, C II, C III and albumin was observed. The minimum measurable concentration of apo A II was 500ng in each assay. A standard curve with a working range of 0.25-8.0 mg/dl was plotted. The coefficients of variation of the reproducibility of intra- and interassays of apo A II in samples were 5.0-8.6% and 6.8-9.9% respectively. The recovery were 106.0 +/- 2.1% (n = 4). The mean concentrations of apo A II in 41 healthy subjects were 24.4 +/- 5.9mg/dl by our method and 26.7 +/- 4.6 mg/dl by RID method, respectively (r = 0.8000, P < 0.001).

Apolipoprotein A-II↗