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Biomedical subjects

D L Yang

Publications and source records attributed to D L Yang.

At least 19 recordsLinked to original sources
PubMed2003

Molecular genetic and biochemical analysis of woodchuck (Marmota monax) MHC class I polymorphism.

The woodchuck (Marmota monax) is an animal model that is used in the study of human hepatitis B virus ( HBV ) infection. A knowledge of woodchuck MHC class I (Mamo-I) genes and gene products is therefore essential for understanding the antigen-specific T-cell responses in this animal model. A number of Mamo-I genes have been identified by molecular cloning and sequencing. However, the allelic nature of these genes has not been proven by classical genetics like the segregation analysis in families. In this study, we analyzed the allelic diversity of Mamo-I in two three-generation woodchuck families including 15 members by sequencing of Mamo-I genes and immunoblotting of Mamo-I proteins after one-dimensional isoelectric focusing (1D-IEF). In addition to four published Mamo-I alleles, six new alleles that belonged to the same locus as the known Mamo-I alleles (Mamo-A) were found within the two woodchuck families. A typical Mendelian segregation of Mamo-I gene and antigens was observed in the families studied. For simple and rapid detection of allelic variability of Mamo-I gene, a typing method based on the detection of PCR products amplified by sequence specific primers (SSP) has been developed and tested in 41 unrelated animals. The most prevalent allele was Mamo-A*01 with a frequency of 21.9% followed by Mamo-A*07 (12.2%). Our study established Mamo-A as a classical MHC class I locus by the polymorphic and allelic nature of Mamo-I gene in the woodchuck.

Amino Acid Sequence↗
PubMed2001

Evidence for inhibition of low density lipoprotein oxidation and cholesterol accumulation by apolipoprotein H (beta2-glycoprotein I).

Low density lipoprotein (LDL) oxidation and lipid accumulation are thought to enhance the progression of atherosclerosis. Apolipoprotein H (apoH) has been implicated in the development of human atherosclerosis. However, the roles of apoH in the oxidative modification of LDL and cellular accumulation of lipid constituents remained uncharacterized. In this study, the level of plasma apoH was found to be significantly associated with the oxidative susceptibility of LDL in human subjects. Plasma levels of apoH were positively correlated with the lag time but negatively correlated with LDL oxidation rate in conjugated diene formation. By using a J774 A.1 macrophage culture system, we found that apoH could not only inhibit the formation of conjugated diene and thiobarbituric acid-reactive substances, but also reduce the electrophoretic mobility of oxidized LDL. Furthermore, apoH decreased cellular accumulation of cholesterol via a reduction in cholesterol influx and an increase in cholesterol efflux. This is the first demonstration that apoH appears to have "antioxidant"-like effects on LDL oxidation. The results also suggest that apoH can inhibit the translocation of cholesterol from extracellular pools to macrophages, suggesting that apoH may play an important role in the prevention of atherosclerosis.

Arteriosclerosis↗
PubMed2001

[A three-domain antibody fragment VH/L specific to tumor blood vessels].

AA98 is a specificaally anti-angiogenic antibody generated in our lab. The heavy chain variable region (VH) attached with mutagenized 36 nucleotides sequence derived from the heavy chain constant region1 (CH1) was amplified VH and light chain (L) were inserted into high-level expression vector pET21a successively, thus pET21a-VH/L was constructed. VH/L was expressed in E. coli BL21 (DE3) after induction with IPTG. The expression of VH/L was 20% of the total bacterial proteins. The refolding of VH/L was conducted by dilution and gel filtration chromatography. The refolded VH/L could bind to HUVEC specifically. Its affinity to the antigen is similar to that of recombinant AA98Fab, but lower than that of the parent antibody AA98.

Angiogenesis Inhibitors↗
PubMed2001

[RP-HPLC method for determination of protopine in plasma and pharmacokinetics in rats].

AIM: To develop a reversed phase high performance liquid chromatographic method (RP-HPLC) for determination of protopine (Pro) in rat plasma and to investigate the pharmacokinetics of Pro in rats. METHODS: The column was packed with 5 microns C18. The mobile phase (pH 5.6) was a mixture of methanol-water-10% acetic acid (80:20:2). After twice extracted with ether under basic condition, and reextracted with 0.02 mol.L-1 sulfuric acid, protopine in the plasma samples was isolated well. The content of protopine in the plasma sample was measured by UV detector at 285 nm. RESULTS: The lowest limit of detection was 50 ng.mL-1. The intraday and interday precisions were 1.5%-3.0% and 2.1%-6.2%, respectively. The mean recovery was 80.6%-97.6%. A good linear relationship between the peak height and the concentration of protopine in rat plasma was observed. The pharmacokinetics of protopine had been investigated in rats after intravenous administration 10 mg.kg-1. The concentration-time curve of protopine in rat was confirmed to two-compartment open model. The T1/2 alpha, T1/2 beta, Ke, CL, Vd were 0.05 h, 1.85 h, 1.52 h, 6.41 L.h-1 and 17.27 L, respectively. CONCLUSION: This method is suitable for studies on pharmacokinetics of protopine.

Analgesics, Opioid↗
PubMed2000

Molecular cloning and characterization of major histocompatibility complex class I cDNAs from woodchuck (Marmota monax).

The full-length cDNAs of woodchuck major histocompatibility complex (MHC) class I (MhcMamo-I or Mamo-I) genes were cloned by using cellular mRNA isolated from the peripheral blood mononuclear cells and liver tissues of woodchucks. DNA sequence analysis of Mamo-I cDNAs revealed that the coding regions of Mamo-I genes were about 1,080 bp long, encoding 359 amino acid residues. The deduced amino acid sequences of Mamo-I showed structural features like leader, alpha1, alpha2, alpha3, transmembrane and cytoplasmic domains, similar to their homologues in human and other mammals. Analysis of five full-length clones from unrelated woodchucks indicated a polymorphism within the alpha1 and alpha2 domains of Mamo-I heavy chain and a high conservation within the alpha3 and the transmembrane/cytoplasmic domains. Amino acid residues of the alpha2 and alpha3 domains that are supposed to be involved in the binding of MHC class I to CD8 molecule, were largely conserved in Mamo-I genes. Phylogenetic comparison of MHC class I genes of woodchuck and other mammals indicated a close evolutionary relationship between woodchuck and squirrel MHC class I. We tentatively named this region the locus A of Mamo-I genes (Mamo-A). Sequence analysis of 101 clones of alpha1 and alpha2 regions derived from 14 woodchucks revealed that at least 14 different alleles within Mamo-A exist. Among these 14 alleles identified so far, Mamo-A*01 and Mamo-A*09 were of the highest frequency of about 21.5% and 14.5%, respectively. Our results indicate that Mamo-I genes are of a similar molecular structure to those of human and other mammals.

Alleles↗
PubMed1999

Rapid detection of herpes simplex virus by polymerase chain reaction.

This study reports the successful use of three sets of primers, each from different genes on the herpes simplex virus (HSV) genome, 1) the DNA polymerase gene, 2) the glycoprotein B gene, and 3) the glycoprotein D gene, for detection of HSV DNA by polymerase chain reaction (PCR). All three sets of primers detected the same HSV DNA in the throat and genital specimens. Using the conventional viral culture as a standard, PCR provided a sensitivity of 100% and a specificity of 100% in this study. In addition, a nested-PCR protocol using two sets of primers in the glycoprotein D gene, one set internal to the other, was evaluated for the amplification of HSV DNA in cerebrospinal fluid (CSF) from patients suspected of having herpes simplex encephalitis (HSE). Five of the 10 CSF specimens tested were found positive. In conclusion, PCR detection is a valuable tool for rapid diagnosis of HSV infection, especially for CSF specimens.

Cells, Cultured↗
PubMed1997

Meta-analysis of phenotype and genotype of NAT2 deficiency in Chinese populations.

Data on both the incidence of slow acetylator phenotype of probe drugs isoniazid, sulfadimidine or sulfamethazine, caffeine and dapsone in mainland or overseas Chinese, and the distribution of NAT2 genotypes and the frequency of NAT2 alleles in the Chinese populations were summarized and reanalysed using a meta-analysis method. Frequency of the slow acetylator phenotype in 3516 healthy Han Chinese gave an overall mean of approximately 19.9 +/- 4.0%, with the range of the combined data being between 15.8% and 25.5%. In addition, frequencies of the slow acetylator phenotype differ between the different minorities in Chinese populations and the range was between 3.2% and 50.6%, with a mean value of 20.6 +/- 12.9% in a total of 1842 individuals from 17 Chinese minorities. In addition, there was no significant heterogeneity in overseas Chinese between the probe drugs isoniazid and sulfadimidine or sulfamethazine (chi 2 = 5.97, df = 4; p > 0.05), and the mean value of slow acetylator phenotype incidence was 24.5% (119/485; 95% CI: 20.7-28.3%), consistent with that of the native Chinese. As expected, frequency of the slow acetylator genotypes in Chinese populations was 25.4% (112/441; 95% CI: 21.3-29.5%), which was in accordance with that of the slow acetylator phenotype in native or overseas Chinese. For all genotypes, *4/*4 (29.9%, 132/441), *4/*6A (27.4%, 121/441), *4/*7A (12%, 53/441) and *6A/*6A (11.3%, 50/441) occupied 80.6%, but *5A/*7A (0.2%, 1/441), *5A/*5A (1.1%, 5/441) and *7A/*7A (1.8%, 8/441) were not frequently found. From this report, the genotype frequencies of homozygous rapid acetylator, heterozygous rapid acetylator, and homozygous slow acetylator were found to be 0.299 (132/441), 0.447 (197/441) and 0.254 (112/441), respectively. Furthermore, both *4 (52.3%; 95% CI: 49-56%) and *6A (30.5%; 95% CI: 28-34%) were major NAT2 alleles, while *7A (11.2%; 95% CI: 9-13%) and *5A (6%; 95% CI: 4-8%) were uncommonly present. Frequency of the mutant alleles was observed at 0.477 (421/882 alleles). The *7A constituted 23.5% t(99/421) of slow acetylator alleles in Chinese populations, showing that this point mutation exists not only in Oriental or Asiatic, but also in Chinese populations. According to the Hardy-Weinberg equilibrium, in the phenotyped Chinese populations, the mean estimate of predicted allelic frequencies of the genotypes RR, Rr, and rr was 0.294, 0.496, and 0.210 for the Chinese, and the expected frequency of the deficient gene r was 0.458. By comparison, the predicted values are in complete agreement with the observed ones. In conclusion, this meta-analysis determined the accurate population frequencies of phenotype and genotype of the NAT2 genetic deficiency in healthy Chinese subjects.

Arylamine N-Acetyltransferase↗
PubMed1996

Local injection of kainic acid causes widespread degeneration of NADPH-d neurons and induction of NADPH-d in neurons, endothelial cells and reactive astrocytes.

Nitric oxide (NO), a diffusible gas, is a messenger molecule that mediates vascular dilatation and neural transmission. The enzyme nitric oxide synthase (NOS) present in neurons is activated by Ca2+ influx associated with activation of glutamate receptors. Cultured cortical neurons containing NOS are selectively vulnerable to injury by kainic acid (KA). However, the relationship between NOS neurons and excitotoxicity under in vivo conditions is not entirely clear. In the present study, we examined the time course and spatial distribution of changes in NOS neurons caused by an intracortical microinjection of KA in adult rats. NADPH-diaphorase (NADPH-d) histochemistry was used as a marker for NOS and the neuronal changes were correlated with changes in glial cells and endothelial cells. We demonstrated a rapid loss of NADPH-d neurons in the lesion center and degeneration of NADPH-d neurons and nerve terminals throughout ipsilateral cortex and hippocampus; the striatal neurons appeared to be unaffected. Subsequent to cortical neuronal degeneration, new NADPH-d activity appeared in proliferative reactive astrocytes and in endothelial cells at lesion periphery, and in neuronal groups at lesion periphery, in ipsilateral entorhinal cortex and bilateral hippocampus. These findings indicate that neurons expressing NADPH-d in cerebral cortex and hippocampus are selectively vulnerable to KA toxicity in vivo. The subsequent induction of NOS in neural and non-neural cells may be regarded as an adaptive response to the kainate-induced brain lesion.

Animals↗
PubMed1996

[A prevalence study on HBV and HCV infection status in patients with hemopathy].

A prevalence study on Hepatitis B virus (HBV) and Hepatitis C virus (HCV) infection in 144 patients with hemopathy was carried out, using an enzyme linked immunosorbent assay (ELISA). The HBsAg, HBcAb and anti-HCV positive rates in sera of these patients were 9.72%, 36.81% and 24.31% respectively, comparing with data from healthy control subjects, 12.03%, 46.30% and 0.93% respectively. No correlation between HCV infection and patient's age or sex was noticed. Nevertheless the incidence of HCV infection was closely related to the history and the frequency of blood transfusion. The ALT level in sera of patients with HCV infection was higher than that of patients without HCV infection. The ALT level in sera was the highest in patients superinfected with HBV and HCV. Our results suggested that blood transfusion and receiving blood products were important routes of HCV transmission in patients with hemopathy.

Adolescent↗
PubMed1996

Kainate-induced brain lesion: similar local and remote histopathological and molecular changes as in ischemic brain infarct.

Cerebral ischemia/hypoxia induces histopathological changes characterized by nuclear and cytoplasmic condensation and sustained c-fos expression. The ischemic changes are thought to be initiated by excessive glutamate released by the ischemic neurons. However, no comparative study has been made between the pathological and molecular changes caused by local injection of excitotoxin and by ischemia. In the present study, we investigated the histopathological changes in rat brains induced by an intracerebral microinjection of kainic acid, a potent analogue of glutamate using two newly available markers for ischemic neurons: Fos immunohistochemistry and EA 50 stain. The rats were sacrificed at intervals from 1 hour (h) to 28 days. We demonstrated that the neurons at the site of injection developed changes typical of ischemia 1 h post-lesion: nuclear and cytoplasmic condensation, strong Fos immunoreactivity and positive EA 50 stain. By 1 day, the neurons underwent necrosis and an infarct-like picture was produced. The neuronal degeneration rapidly spread to the bilateral neocortex, CA3 and CA4 regions of hippocampus, piriform gyrus, amygdala and cerebellar Purkinje cells. After 3 days, there was neovascularization and macrophage production in the lesion center and astrocytic proliferation at the lesion periphery. The CA1 of hippocampus showed delayed neuronal necrosis typical of ischemia. Thus, intracerebral microinjection of KA induces similar histopathological and molecular changes as those occurring in brain infarct and is a simple and reliable model for studying changes related to focal brain infarct.

Animals↗
PubMed1995

[Study of HCV infection and viremia in patients with posttransfusion hepatitis in China].

One hundred and sixty-nine patients with posttransfusion hepatitis (PTH) were investigated for etiology with serological and PCR methods. Our result showed that HCV was the major causative agent of PTH in this study. A part of patients (24/169) were also HBV markers positive. Three patients were negative for all detected viral markers, looking forwards to progressive study. Anti-HCV was detectable from 7 days to one year after onsetting of clinical symptoms, average time 54.62 days. HCV RNA could be detected in early time of HCV infection ( 6 to 20 days later), average time 8.72 days. A follow-up study of 84 patients showed that all patients remained anti-HCV positive during the observation for 1.5 -- 3 years, but their HCV viremia had three forms: transient, persistent or intermittent. The interaction between HCV and HBV by their superinfection was unknown, awaiting further study.

Adolescent↗
PubMed1995

Infectivity and risk factors of hepatitis C virus transmission through sexual contact.

HCV infection among heterosexuals with multiple partners and family members of patients with hepatitis C, and HCV RNA in the body fluid of these patients were investigated. The results showed that the HCV infection in heterosexuals with multiple partners, which was related to sexual activity, was much higher than that of healthy pregnant women. The HCV RNA in the saliva, semen or vaginal discharge of patients with hepatitis C was at detectable level. Among the patient's relatives, none of the children but 2 spouses were found to be infected with HCV. We concluded that sexual contact might play some role in HCV transmission.

Adolescent↗
PubMed1992

Analysis of antigenic polypeptides of Dane particles and antibody response ability of HBV infected subjects to PreS1 polypeptides.

We demonstrated the constitutive polypeptides (PP) of Dane particles employing Western Blot and investigated the antibody response ability of HBV infected subjects to PreS1 PP in comparison with other serum markers from HBV infected individuals. The results indicated that 1) the major reason for discrepant results may be related to the detergents used in the sample solutions and the degree of denaturation the samples had undergone; 2) there are 12 bands in the PAGE-graph of Dane particles. By Western Blot it was confirmed that 5 PP (P24, P27, P36, P39, P42) are derived from S-open reading frame (S-ORF), P21 is associated with C-ORF, P24-25 possesses some epitopes of Pol protein, and P45 and P76 express similar epitopes to human IgG and IgM; and 3) the prevalence of anti-PreS1 PP was 17.24% in the group of healthy persons following latent HBV infection, much higher than that of HBV infected patients (1.21%). The above findings imply that antibody response ability of the host to PreS1 PP is attributing to the outcome of HBV infection. It may play an important role in the elimination of the virus.

Blotting, Western↗
PubMed1992

[Detection of hepatitis B virus DNA in leucocytes of primary hepatocarcinoma using polymerase chain reaction].

The detection of HBV DNA in sera and leucocytes of 30 primary hepatocarcinoma patients whom were negative for HBsAg by reverse passive hemagglutination using polymerase chain reaction (PCR) were reported. The detectable rates of HBV DNA in leucocytes and sera were 86.7% and 70% respectively and the serological indexes, 43.4%. These indicated that HBV DNA may existent in blood of the patients with negative serological indexes. It was common that HBV DNA presented in peripheral leucocytes of primary hepatocarcinoma, indicated that extrahepatic sites of the HBV genome, and perhaps replication of HBV virus, existed. It was of great importance for the role of HBV DNA in lemology and epidemiology, etiology of hepatocarcinoma, diagnosis and treatment of infection as well as standard of cure.

Adult↗