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Biomedical subjects

H He

Publications and source records attributed to H He.

At least 181 records · Page 10Linked to original sources
PubMed1997

Adenovirus-mediated gene transfer reconstitutes depressed sarcoplasmic reticulum Ca2+-ATPase levels and shortens prolonged cardiac myocyte Ca2+ transients.

BACKGROUND: Decreased expression of the sarcoplasmic reticulum (SR) Ca2+-ATPase of the cardiac myocyte (SERCA2) and abnormal Ca2+ regulation have been independently linked to human heart failure. This study was designed to determine whether expression of a SERCA2 transgene could reconstitute depressed cardiac myocyte SERCA2 levels, augment SR Ca2+ uptake, and shorten prolonged excitation-contraction (EC)-associated Ca2+ transients in neonatal rat cardiac myocytes (NM). METHODS AND RESULTS: Cultured NM were treated with phorbol-12-myristate-13-acetate (PMA), a compound that decreases endogenous SERCA2 expression and results in prolongation of EC-associated Ca2+ transients. PMA-treated NM had a 75% reduction in SERCA2 mRNA and a 40% reduction in SERCA2 protein levels. SERCA2 adenovirus infection increased SERCA2 mRNA expression to 2.5 times control and reconstituted SERCA2 protein levels in PMA-treated cells. This reconstitution was associated with a 32.4% reduction in the time for decline of the Indo-1 Ca2+ transient to half-maximum levels (t(1/2) [Ca2+]i) (P<.05). A 34.5% augmentation of oxalate-facilitated SR Ca2+ uptake was also documented in SERCA2 adenovirus-infected cells (P<.05). CONCLUSIONS: Adenovirus-mediated expression of a SERCA2 transgene can reconstitute depressed endogenous SERCA2 levels, shorten prolonged Ca2+ transients, and augment SR Ca2+ uptake. It is conceivable that such an approach might be used in vivo to normalize altered Ca2+ regulation in human heart failure.

Adenoviridae↗
PubMed1997

Overexpression of the rat sarcoplasmic reticulum Ca2+ ATPase gene in the heart of transgenic mice accelerates calcium transients and cardiac relaxation.

The Ca2+ ATPase of the sarcoplasmic reticulum (SERCA2) plays a dominant role in lowering cytoplasmic calcium levels during cardiac relaxation and reduction of its activity has been linked to delayed diastolic relaxation in hypothyroid and failing hearts. To determine the contractile alterations resulting from increased SERCA2 expression, we generated transgenic mice overexpressing a rat SERCA2 transgene. Characterization of a heterozygous transgenic mouse line (CJ5) showed that the amount of SERCA2 mRNA and protein increased 2. 6-fold and 1.2-fold, respectively, relative to control mice. Determination of the relative synthesis rate of SERCA2 protein showed an 82% increase. The mRNA levels of some of the other genes involved in calcium handling, such as the ryanodine receptor and calsequestrin, remained unchanged, but the mRNA levels of phospholamban and Na+/Ca2+ exchanger increased 1.4-fold and 1.8-fold, respectively. The increase in phospholamban or Na+/Ca2+ exchanger mRNAs did not, however, result in changes in protein levels. Functional analysis of calcium handling and contractile parameters in isolated cardiac myocytes indicated that the intracellular calcium decline (t1/2) and myocyte relengthening (t1/2) were accelerated by 23 and 22%, respectively. In addition, the rate of myocyte shortening was also significantly faster. In isolated papillary muscle from SERCA2 transgenic mice, the time to half maximum postrest potentiation was significantly shorter than in negative littermates. Furthermore, cardiac function measured in vivo, demonstrated significantly accelerated contraction and relaxation in SERCA2 transgenic mice that were further augmented in both groups with isoproterenol administration. Similar results were obtained for the contractile performance of myocytes isolated from a separate line (CJ2) of homozygous SERCA2 transgenic mice. Our findings suggest, for the first time, that increased SERCA2 expression is feasible in vivo and results in enhanced calcium transients, myocardial contractility, and relaxation that may have further therapeutic implications.

Animals↗
PubMed1997

Characterization of visible dyes for four-decay fluorescence detection in DNA sequencing.

Dyes of several classes were investigated as candidates for use in a multiplex, four-decay fluorescence detection scheme for DNA sequencing. The dyes include nitrobenzofuran dyes, rhodamine dyes, fluorescein dyes, cyanine dyes, Nile Red, and BODIPY dyes. Based on the results of fluorescence spectral and lifetime studies, an initial set of four dyes was selected for further study: NBD-aminohexanoic acid (NBD-HA, r = 1.1 ns), tetramethyl-rhodamine, methyl ester (r = 2.2 ns), rhodamine green (r = 4.3 ns), and BODIPY 505/515 (r = 5.9 ns). Limits of lifetime detection of the four dyes were investigated, and lifetime resolution was demonstrated for mixtures of the free dyes in batch solution. Lifetime of dye-labeled DNA primers also were determined in batch solution and detected on-the-fly in capillary electrophoresis (CE). Conjugation of the dyes to DNA improved the resolution of their individual lifetimes in mixtures in batch measurements. When attached to the primer, tetramethyl-rhodamine exhibited biexponential decay with a dominant lifetime of 3.8 ns, making it unsuitable for four-decay sequencing. Contact with the CE gel lengthened the lifetime of NBD-HA-labeled primer from 1.3 to 2.1 ns but did not affect the lifetimes of the other dyes. Lifetime detectability of labeled primers at individual points along an electrophoretic peak in the attomole range.

Evaluation Studies as Topic↗
PubMed1997

Synergistic induction of CTLA-4 expression by costimulation with TCR plus CD28 signals mediated by increased transcription and messenger ribonucleic acid stability.

T cell activation requires at least two signals transduced by the Ag-specific TCR plus a costimulatory receptor. The CD28 costimulatory molecule has been shown to promote T cell proliferation and cytokine production. CTLA-4, a cell surface molecule homologous to CD28, can function as a repressor of T cell activation. Thus, CTLA-4 and CD28 may have opposing functions during T cell activation. CTLA-4 is expressed at low levels on resting T cells and up-regulated after T cell activation. Regulation of CTLA-4 expression is critical to the normal regulation of immunity. For example, CTLA-4-deficient mice develop early onset lethal autoimmunity. We previously showed that CTLA-4 transcription is increased after T cell activation and that induction was controlled by 335 bp of CTLA-4 upstream sequence. In this work, we show that cell surface CTLA-4 expression is increased synergistically by TCR plus CD28 signals. Synergistic induction is mediated by two mechanisms: an enhanced rate of transcription and increased mRNA stability. In contrast to the regulation of IL-2 and IL-2R expression, which is inhibited by cyclosporin A-, but not rapamycin-dependent signal transduction pathways, CTLA-4 expression is inhibited by either cyclosporin A or rapamycin. Thus, synergistic induction of CTLA-4 expression requires both cyclosporin A- and rapamycin-dependent signals.

Abatacept↗
PubMed1997

The small subunit is required for functional interaction of DNA polymerase delta with the proliferating cell nuclear antigen.

DNA polymerase delta is usually isolated as a heterodimer composed of a 125 kDa catalytic subunit and a 50 kDa small subunit of unknown function. The enzyme is distributive by itself and requires an accessory protein, the proliferating cell nuclear antigen (PCNA), for highly processive DNA synthesis. We have recently demonstrated that the catalytic subunit of human DNA polymerase delta (p125) expressed in baculovirus-infected insect cells, in contrast to the native heterodimeric calf thymus DNA polymerase delta, is not responsive to stimulation by PCNA. To determine whether the lack of response to PCNA of the recombinant catalytic subunit is due to the absence of the small subunit or to differences in post-translational modification in insect cells versus mammalian cells, we have co-expressed the two subunits of human DNA polymerase delta in insect cells. We have demonstrated that co-expression of the catalytic and small subunits of human DNA polymerase delta results in formation of a stable, fully functional heterodimer, that the recombinant heterodimer, similar to native heterodimer, is markedly stimulated (40- to 50-fold) by PCNA and that the increase in activity seen in the presence of PCNA is the result of an increase in processivity. These data establish that the 50 kDa subunit is essential for functional interaction of DNA polymerase delta with PCNA and for highly processive DNA synthesis.

Animals↗
PubMed1997

Expression of Cdk5, p35, and Cdk5-associated kinase activity in the developing rat lens.

We have investigated the expression of Cdk5 and its regulatory subunit, p35, in the developing rat lens from embryonic day 16 (E16) to postnatal day 8 (P8). Reverse transcription and polymerase chain reaction (RT/PCR) detected Cdk5 and p35 mRNA expression in lens epithelial cells and in differentiating lens fibers throughout this developmental period. Subsequent sequencing of the RT/PCR products confirmed their identifies. In sity hybridization with Cdk5 and p35 riboprobes showed especially high expression of both mRNAs in the newly formed lens fiber cells in the bow region of the lens. Immunocytochemistry at E18 showed that Cdk5 was present in the cytoplasm of lens epithelial cells and fiber cells, with especially strong immunostaining at the anterior ends of the fibers. Fiber cells in the final stages of maturation, immediately prior to nuclear degeneration, showed positive staining for Cdk5 in the nucleus. Immunoprecipitation of proteins with Cdk5 antibody followed by immunoblotting with either N-terminal specific or C-terminal specific p35 antibodies demonstrated that p35 is complexed with Cdk5 in lens epithelial cells and lens fibers. Immunoprecipitates of Cdk5 from epithelia and fibers showed kinase activity in vitro using histone H1 as a substrate. These findings demonstrate that p35/Cdk5 activity is not restricted to neurons and raise the possibility that this kinase may play a role in lens fiber cell differentiation.

Amino Acid Sequence↗
PubMed1997

Myocardial alpha1-adrenoceptor: inotropic effect and physiologic and pathologic implications.

Alpha1-adrenergic receptors have been found in myocardium of all mammalian species. Although the exact underlying mechanisms have not been conclusively determined, it would appear that the myocardial effects of alpha1-adrenoceptors may vary in importance according to the pathophysiologic process involved. In physiological conditions, this receptor system plays a role in cardiac growth, cardiac contraction, and has both an antiarrhythmic function as well as a role in cardiac adaptation to various situations. This system is also involved in some pathological processes such as ischemia/reperfusion, ischemic preconditioning, and cardiac hypertrophy. The role of alpha1-adrenoceptors in heart failure is somewhat controversial. Experimental evidence suggests that myocardial alpha1-adrenoceptors can have either beneficial or deleterious effects on the heart. It thus seems possible that the development of agents specific to certain subtypes of alpha1-adrenoceptor and a better understanding of their role in pathophysiologic states could be clinically relevant.

Animals↗
PubMed1997

Ochre suppressor transfer RNA restored dystrophin expression in mdx mice.

The mdx mouse is an animal model for human Duchenne muscular dystrophy. The lack of dystrophin in mdx mice is caused by an ochre mutation in exon 23 of the dystrophin gene. This study tested the feasibility of inhibiting translational termination as an approach for genetic therapy for diseases caused by nonsense mutations. We evaluated both the in vitro and in vivo efficiencies of readthrough of ochre codons in 2 genes with the tRNA suppressor gene. The first target was a CAT reporter gene bearing an ochre mutation at the 5' end (CATochre). The second target was the dystrophin gene in mdx mice. The readthrough efficiencies were about 20% in COS cells and 5.5% in rat hearts. At four weeks after a direct injection of plasmid DNA encoding the tRNA suppressor into mdx mice, dystrophin positive fibers were detected by sarcolemmal immunostaining. This is the first convincing data that a tRNA suppressor gene might be a useful in vivo treatment for the genetic disorders caused by nonsense mutations.

Animals↗
PubMed1997

Comparison of the Ca2+ movement by activation of alpha1-adrenoceptor subtypes in HEK-293 cells.

We studied the Ca2+ movement induced by activation of alpha1A-, alpha1B- and alpha1D-adrenoceptor subtypes in transfected HEK-293 cells with the fura-2 probe. All these alpha1-AR subtypes induced both Ca2+ release and Ca2+ entry. The effect on Ca2+ release in alpha1b transfected HEK-293 cells was bigger than that in alpha1a and alpha1d transfected HEK-293 cells, and the effects on Ca2+ entry were the same in alpha1a, alpha1b and alpha1d transfected HEK-293 cells. The Ca2+ entry was inhibited by 1 mM NiSO4, but not by nifedipine. Cyclopiazonic acid (CPA) produced a biphasic Ca2+ signal response in Ca2+ medium, and only induced a transient response in Ca2+-free medium. After depletion of CPA-sensitive Ca2+ pool by 10 microM CPA in Ca2+-free medium, 10 microM adrenaline (Adr) still transiently increased [Ca2+]i in three different alpha1-adrenoceptor subtype transfected HEK-293 cells. However, after depletion of adrenaline-sensitive Ca2+ pool by 10 microM Adr, CPA transiently elevated [Ca2+]i only in alpha1a and alpha1d transfected HEK-293 cells, not in alpha1b transfected HEK-293 cells. U73122, a phospholipase C (PLC) inhibitor, inhibited both Ca2+ release and Ca2+ entry induced by activation of alpha1A alpha1B and alpha1D subtypes in transfected HEK-293 cells. These results suggest that HEK-293 cell line contains two functionally separate intracellular Ca2+ pools, CPA-sensitive and Adr-sensitive pools. Activation of alpha1B-AR stimulates Ca2+ release from both CPA-sensitive and Adr-sensitive Ca2+ pools. Alpha1A and alpha1D subtypes induce Ca2+ release only from Adr-sensitive Ca2+ pool.

Animals↗
PubMed1997

Assessment of sex chromosome ratio and aneuploidy rate in motile spermatozoa selected by three different methods.

Using three-colour fluorescence in-situ hybridization, sex chromosome ratios and frequencies of diploidy and disomy for chromosomes X, Y and 18 were compared in spermatozoa of good and poor motility after separation by swim-up, glass-wool and two-layer discontinuous Percoll methods. Semen samples were collected from seven normal males aged 26-31 years. A minimum of 6000 sperm nuclei per sample were evaluated for each chromosome for a total of 308,432 sperm nuclei. Hybridization efficiency was 99.8%. A slight change in the ratio of X- to Y-bearing spermatozoa was noted after Percoll separation (from 49.3:49.5 to 50.0:48.9; P = 0.036 and P = 0.046), but not after separation by the other two methods. We did not observe significant differences in the disomy rates for sex chromosomes or chromosome 18 or in the diploidy rate between spermatozoa with good and poor motility after separation by any of the three methods. Our data indicate that separation of motile spermatozoa does not alter the ratio of X- to Y-bearing spermatozoa to a degree that represents sex chromosome selection.

Adult↗
PubMed1997

A phenotypically normal liveborn male after prenatal diagnosis of trisomy 20 mosaicism.

We report on a case of prenatally diagnosed trisomy 20 mosaicism. Conventional cytogenetic analysis and fluorescence in situ hybridization (FISH) using a chromosome 20 specific probe were performed on the lymphocytes and extra-embryonic tissues after birth. All of them revealed normal karyotypes. The baby is developing normally at the age of 2 years.

Adult↗
PubMed1997

Regional blood flow in Dahl-Iwai salt-sensitive rats and the effects of dietary L-arginine supplementation.

The purpose of the present study was to determine 1) whether different organs undergo similar increase in vascular resistance in Dahl-Iwai salt-sensitive (S) rats, and 2) the effects of chronic oral L-arginine supplementation on the regional hemodynamics in S rats. Male 6-wk-old S rats and salt-resistant (R) rats were maintained on an 8% NaCl chow for 4 wk. One group (S or R rats) was maintained on tap water and the other group (S/Arg or R/Arg rats) received tap water containing L-arginine at a concentration of 1.5%. Organ blood flow and cardiac output were measured with microspheres in the conscious condition. Mean blood pressure in S, S/Arg, R, and R/Arg rats was 159 +/- 5, 138 +/- 3, 111 +/- 4, and 112 +/- 4 mmHg, respectively. Urinary excretion of protein and albumin in S/Arg rats was significantly suppressed compared with S rats. Concerning regional hemodynamics, the flow rate of the kidney was lower in S rats than in R rats, but there were no differences between S and R rats in the flow rates of the brain, heart, lung, liver, spleen, intestine, skeletal muscle, and skin. Thus the renal blood flow was solely reduced in S rats on a high-salt diet. The flow rate of the kidney in S/Arg rats was maintained at a higher level compared with that of S rats. L-Arginine treatment tended to produce a recovery in the urinary excretion of guanosine 3',5'-cyclic monophosphate in S rats, but had no effect in R rats. Thus the supplementation of L-arginine prevented the increase in blood pressure in S rats on a high-salt diet and normalized the abnormality of renal hemodynamics accompanying salt-induced hypertension.

Animals↗
PubMed1997

Alpha 1-adrenergic stimulation inhibits 3,5,3'-triiodothyronine-induced expression of the rat heart sarcoplasmic reticulum Ca2+ adenosine triphosphatase gene.

The interactions between the beta-adrenergic system and thyroid hormone (T3) on cardiac function have been investigated in detail. In addition to beta-adrenoceptors, alpha 1-adrenergic receptors are present in the mammalian heart. The interactions between T3 and the alpha 1-adrenergic system remain, however, poorly understood. T3 stimulates the expression and transcription of the sarcoplasmic reticulum Ca2+ adenosine triphosphatase (SERCA2) gene, a protein vital in the control of cardiac calcium transients and contractility. We show that in rat cardiac myocytes, the stimulatory effect of T3 on SERCA2 messenger RNA expression and gene transcription is inhibited by an alpha 1-adrenergic agonist. We demonstrate that direct activation of the alpha 1-adrenergic signaling pathway, using a mutant constitutively active G protein (Gq) similarly down-regulated the T3 effect on SERCA2 transcription. The combined effect of thyroid hormone receptor and retinoid X receptors on T3-stimulated SERCA2 gene transcription was also markedly attenuated by alpha 1-adrenergic stimulation. These results suggested that activation of the alpha 1-adrenergic signaling pathway has an inhibitor effect on T3-dependent SERCA2 gene transcription. As this inhibitory effect of alpha 1-adrenergic stimulation occurs when only one thyroid hormone response element (TRE) drives reporter expression, it is most likely mediated by an alteration of the nuclear factors binding to the TRE or by influencing the interaction of the TRE complex with the basal transcriptional machinery.

Animals↗
PubMed1997

[The effect of chlorhexidine varnish system on Streptococcus mutans in fissure plaques].

A chlorhexidine varnish system and a controlled releasing varnish were prepared by the authors. The release rate of chlorhexidine in vitro and the effects on S. mutans and S. sanguis in vivo were determined. The results showed that the system could selectively inhibit the growth of S. mutans in fissure plaque and had no effect on S. sanguis. The period of interaction between the layer of varnish and plaque was prolonged by the control-releasing reagent. The study suggests that the chlorhexidine varnish system is effective of anticaries and can maintain microflora balance in oral bacteria.

Adolescent↗
PubMed1997

[The establishment of antigen sandwiched ELISA for detection of total antibodies to hepatitis C virus].

An antigen sandwiched ELISA for detection of the total antibodies to HCV was established using genetically engineered proteins and synthesized peptides located in the structural and nonstructural regions of HCV genomes as coated antigens and were labeled by horseradish peroxidase, respectively. A comparison of antigen sandwiched ELISA with indirect ELISA showed that the sensitivity of sandwiched system (1:128) was little higher than that of indirect ELISA (1:64), the specificities of both systems were all 100%. This reagent may detect all classes of antibody to HCV in serum, mainly IgG and IgM and is a new diagnostic reagent for detection and early diagnosis of HCV infection.

Enzyme-Linked Immunosorbent Assay↗
PubMed1997

[Primary analysis of the situation of hepatitis G virus infection in different groups of people].

In order to investigate the situation of hepatitis G virus (HGV) infection among different groups of people, two self-selected peptides from HGV NS5 were used to develop an indirect ELISA for anti-HGV IgG detection. 1209 sera from different peoples were detected and the positive rate of anti-HGV IgG was (3.8%). Among these groups the group of non A-E hepatitis had the highest anti-HGV positive rate (20.5%), higher than that in the groups of natural people (0.8%) and other hepatitis patients (3.3%), the difference was significant. The anti-HGV IgG positive rate in the group of HC patients 8.0% was higher than that in other hepatitis A-E patients, the positive rate (8.0%) in paid donors was higher than that in voluntary donors (0%), and that in groups with venereal diseases and HIV infection were 3.6% and 8.0%, respectively. The results suggested that there has higher rate of HGV infection in our country; HGV is the important factor causing non A-E, the paid donors is the dangerous group, they should be strictly screened, and the people with transfusion have higher rate of HGV infection. Since there is different rate of HGV infection in different human population so it is important to study further the epidemiological characteristics of HGV for its prevention.

China↗
PubMed1997

[The detection of human immunodeficiency virus type 1/2 antibody using synthetic peptides].

Four peptides of human immunodeficiency virus (HIV) located in the regions of gp41, gp120, p24 of HIV-1 and gp36 of HIV-2 have been synthesized according to the published amino acid sequences and the positions of antigenetic determinants of viruses by a solid-phase method. An indirect ELISA for detecting antibody against HIV 1/2 was established using these synthetic peptides as the coating antigens. We detected 41 control sera provided by National Institute for the Control of Pharmaceutical and Biological Products. The results indicated that the general coincident rate were 100%, the variant coefficient was less than 10%. A comparison of our reagent with Huayi, Pasteur and Jinhao reagents in detection of 90 positive sera and 140 normal sera for HIV-1/2 showed that the coincident rate was 100% with Pasteur and Jinhao, 99.57% with Huayi. All of 186 sera from patients with other diseases were negative. The reagent are stable at 37 degrees C for 4 days. The results indicated that our reagent is highly specific, sensitive and stable, it can be used for detection of HIV-1/2 infections.

Antibodies, Viral↗