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W Liao

Publications and source records attributed to W Liao.

At least 55 records · Page 3Linked to original sources
PubMed1999

APOBEC-2, a cardiac- and skeletal muscle-specific member of the cytidine deaminase supergene family.

APOBEC-1, which mediates the editing of apolipoprotein (apo) B mRNA, is the only known member of the C (cytidine)-->U (uridine) editing enzyme subfamily of the cytidine deaminase supergene family. Here we report the cloning of APOBEC-2, another member of the subfamily. Human and mouse APOBEC-2 both contain 224 amino acid residues, and their genes are mapped to syntenic regions of human chromosome 6 (6p21) and mouse chromosome 17. By phylogenetic analysis, APOBEC-2 is shown to be evolutionarily related to APOBEC-1, and analysis of substitution rates indicates that APOBEC-2 is a much better conserved gene than APOBEC-1. APOBEC-2 mRNA and protein are expressed exclusively in heart and skeletal muscle. APOBEC-2 does not display detectable apoB mRNA editing activity. Like other editing enzymes of the cytidine deaminase superfamily, APOBEC-2 has low, but definite, intrinsic cytidine deaminase activity. The identification of APOBEC-2 indicates that APOBEC-1 is not the only member of the C-->U editing enzyme subfamily, which, like the A (adenosine)-->I (inosine) subfamily of editing enzymes, must encompass at least two and possibly more different deaminase enzymes. It suggests that the C-->U editing affecting apoB mRNA and other RNAs is not an isolated event mediated by a single enzyme but involves multiple related proteins that have evolved from a primordial gene closely related to the housekeeping enzyme cytidine deaminase.

APOBEC Deaminases↗
PubMed1999

Roles of the three major phosphorylation sites of hepatitis B virus core protein in viral replication.

Hepatitis B virus (HBV) core protein is a phosphoprotein. Its three major phosphorylation sites have been identified at the serine residues located at amino acids 157, 164, and 172. In order to investigate the role of core protein phosphorylation in HBV replication, these three serine residues were mutated to alanine to mimic nonphosphorylated serine or to glutamic acid to mimic phosphoserine. The nonphosphorylated core protein analog did not package the HBV pregenomic RNA, and the phosphorylated analog packaged the pregenomic RNA but failed to support viral DNA replication. These results indicate that the core protein phosphorylation may be important for pregenomic RNA packaging and that its dephosphorylation may be important for viral DNA replication. The individual roles of these three major phosphorylation sites in HBV replication were further investigated by being mutated to alanine in different combinations. The results showed that the serine residue at amino acid 157 was not essential for pregenomic RNA packaging, whereas the serine residues at amino acids 164 and 172 were more important. Furthermore, the serine residue at amino acid 157 was not essential for viral DNA replication or viral maturation.

Carcinoma, Hepatocellular↗
PubMed1999

Genetic deficiency of acylation stimulating protein (ASP(C3ades-Arg)) does not cause hyperapobetalipoproteinemia in mice.

The acylation stimulating protein (ASP) is a 76-amino acid peptide that has been proposed as a potent mediator of triglyceride synthesis and, when functionally impaired, as a major cause of hyperapobetalipoproteinemia (HyperapoB). Purification and sequence analysis of ASP from human sera have revealed that ASP is identical to the complement C3-derived activation peptide C3ades-Arg. Because C3 is the precursor for C3ades-Arg and therefore ASP, a deficiency in C3 would be predicted to result in a phenotype characteristic of HyperapoB. To test this hypothesis in vivo, the current study was undertaken in which ASP(C3ades-Arg)-deficient mice were used as a model system. No significant differences were found in the triglyceride, cholesterol, or free fatty acid concentrations in the plasma of fasted normal and ASP(C3ades-Arg)-deficient animals. In addition, plasma lipoprotein analyses indicated that the very low density lipoprotein, low density lipoprotein, and high density lipoprotein cholesterol and triglyceride concentrations as well as the apolipoprotein B-48 and B-100 levels were not significantly different in the plasma of ASP(C3ades-Arg)-deficient and wild type mice. Furthermore, when challenged with an oral fat load, the ASP(C3ades-Arg)-deficient mice showed no impaired ability to clear triglycerides and free fatty acids from their circulation when compared with their wild-type littermates. Collectively, these results indicate that ASP(C3ades-Arg) deficiency does not cause HyperapoB in mice and that the physiological importance of impaired ASP(C3ades-Arg) function as a cause of hyperapobetalipoproteinemia needs to be reevaluated.

Animals↗
PubMed1999

Endogenous histamine reduces plasma insulin-like growth factor I via H1 receptor-mediated pathway in the rat.

Endotoxin has been recently shown to reduce plasma insulin-like growth factor I. As it was reported that histamine can induce gut-derived endotoxemia, we wanted to determine whether histamine has a similar effect on plasma insulin-like growth factor I. Compound 48/80 (a histamine releaser) was injected subcutaneously into rats, then blood was taken for plasma insulin-like growth factor I assay and the livers were assayed for insulin-like growth factor I mRNA. Like endotoxin, injection of compound 48/80 significantly reduced plasma insulin-like growth factor I. Six hours post-injection, plasma insulin-like growth factor I was reduced by 61% (P < 0.001), and 24 h post-injection, it was still lower (by 35% P < 0.001) than in the control group. Hepatic insulin-like growth factor I mRNA was not reduced by this treatment. The effect of compound 48/80 on plasma insulin-like growth factor I was significantly attenuated by oral administration of the histamine H1 receptor antagonist (chlorpheniramine), but not by the histamine H2 receptor antagonists (cimetidine and ranitidine). Oral administration of polymyxin B (an antiendotoxin antibiotic) did not attenuate the effect of compound 48/80 on plasma insulin-like growth factor I at all. In conclusion, endogenous histamine reduces plasma insulin-like growth factor I via H1 receptor-mediated pathway. Our study suggests a novel role of histamine in the regulation of insulin-like growth factor I metabolism in vivo.

Animals↗
PubMed1999

Adenovirus-mediated overexpression of microsomal triglyceride transfer protein (MTP): mechanistic studies on the role of MTP in apolipoprotein B-100 biogenesis.

The intracellular concentration of the microsomal triglyceride transfer protein large subunit (lMTP), the abetalipoproteinemia gene product, is tightly controlled. To date, attempts at overexpressinglMTP in vivo or in vitro have been unsuccessful. We successfully overexpressed lMTP in HepG2 cells using an adenoviral vector containing an lMTP cDNA, AdMTP. AdMTP-transduced HepG2 cells overexpressed MTP activity. They secreted increased amounts of apoB-100 lipoproteins with LDL and HDL density into the medium. lMTP overexpression alone minimally changed the density profile of apoB-containing lipoproteins, but addition of oleic acid shifted the profile toward lower densities. Oleic acid had a greater stimulatory effect on apoB-100 secretion in control HepG2 cells than in AdMTP-transduced cells, because (i) adenoviral transduction per se suppressed protein synthesis, affecting apoB-100 and albumin equally, and (ii) adenoviral transduction partially attenuated the increase in triglyceride synthesis in response to oleic acid supplementation. AdMTP treatment greatly diminished the intracellular degradation of apoB-100, but in comparison with recombinant virus containing luciferase cDNA (AdLuc), it caused no change in its biosynthetic rate. It greatly reduced, but did not eliminate, its proteasomal degradation. Our study constitutes the initial demonstration that adenovirus-mediated transfer of lMTP markedly stimulates MTP expression which in turn stimulates apoB-100 production. The mechanism involves a downregulation of ubiquitin-proteasome-mediated degradation without any change in synthetic rate.

Adenoviruses, Human↗
PubMed1999

Liver-specific inactivation of the abetalipoproteinemia gene completely abrogates very low density lipoprotein/low density lipoprotein production in a viable conditional knockout mouse.

Conventional knockout of the microsomal triglyceride transfer protein large subunit (lMTP) gene is embryonic lethal in the homozygous state in mice. We have produced a conditional lMTP knockout mouse by inserting loxP sequences flanking exons 5 and 6 by gene targeting. Homozygous floxed mice were born live with normal plasma lipids. Intravenous injection of an adenovirus harboring Cre recombinase (AdCre1) produced deletion of exons 5 and 6 and disappearance of lMTP mRNA and immunoreactive protein in a liver-specific manner. There was also disappearance of plasma apolipoprotein (apo) B-100 and marked reduction in apoB-48 levels. Wild-type mice showed no response, and heterozygous mice, an intermediate response, to AdCre1. Wild-type mice doubled their plasma cholesterol level following a high cholesterol diet. This hypercholesterolemia was abolished in AdCre1-treated lMTP-/- mice, the result of a complete absence of very low/intermediate/low density lipoproteins and a slight reduction in high density lipoprotein. Heterozygous mice showed an intermediate lipoprotein phenotype. The rate of accumulation of plasma triglyceride following Triton WR1339 treatment in lMTP-/- mice was <10% that in wild-type animals, indicating a failure of triglyceride-rich lipoprotein production. Pulse-chase experiments using hepatocytes isolated from wild-type and lMTP-/- mice revealed a failure of apoB secretion in lMTP-/- animals. Therefore, the liver-specific inactivation of the lMTP gene completely abrogates apoB-100 and very low/intermediate/low density lipoprotein production. These conditional knockout mice are a useful in vivo model for studying the role of MTP in apoB biosynthesis and the biogenesis of apoB-containing lipoproteins.

Abetalipoproteinemia↗
PubMed1999

Endotoxin suppresses mouse hepatic low-density lipoprotein-receptor expression via a pathway independent of the toll-like receptor 4.

Endotoxin provokes an inflammatory state in the infected host. C3H/HeJ mice are tolerant to endotoxin because of an Lps gene mutation. Recent studies have identified that this gene encodes the Toll-like receptor 4. Endotoxin also induces hyperlipidemia and suppresses hepatic low-density lipoprotein (LDL)-receptor expression. In the current study, we investigated whether a defective Lps gene would impair the hepatic LDL-receptor response to endotoxin in C3H/HeJ mice. Eighteen hours after an intraperitoneal injection of endotoxin, the hepatic LDL-receptor expression and the plasma lipoprotein pattern were analyzed. Endotoxin increased plasma triglyceride and apoE in very low-density lipoproteins (VLDL) and intermediate-density lipoproteins, and decreased apoAI in high-density lipoproteins (HDL) in the endotoxin-sensitive mice (C3H/HeN), but not in the endotoxin-resistant mice (C3H/HeJ). These data indicate that a defective Lps gene impairs the endotoxin signaling to alter these lipoproteins. However, the hepatic LDL-receptor response to endotoxin in the endotoxin-resistant mice was similar to that in the endotoxin-sensitive mice. Thus, at a dose of 5 microg/mouse, endotoxin reduced hepatic LDL-receptor expression by 35% in C3H/HeN mice and by 52% in C3H/HeJ mice. At a dose of 50 microg/mouse, endotoxin reduced hepatic LDL-receptor expression by 61% in C3H/HeN mice and by 63% in C3H/HeJ mice. It is concluded that endotoxin suppresses hepatic LDL-receptor expression in vivo via a pathway independent of the Toll-like receptor 4.

Animals↗
PubMed1999

A comparison among four regimens of itraconazole treatment in onychomycosis.

The purpose of this study was to compare the efficacy of different dosage regimens in the management of onychomycosis with itraconazole and to determine the results of a further 1-week intermittent pulse treatment in non-cured patients. In this study, 153 patients were randomly allocated to four groups. Patients in group A were treated with daily doses of 100 mg for 3 months in the case of fingernail onychomycosis and for 4 months in the case of toenail onychomycosis. Patients in the other groups received a intermittent pulse therapy, in which the drug was taken for 1 week, then discontinued for 3 weeks, three cycles for fingernail and four cycles for toenail infection. The daily doses were 400 mg (group B), 300 mg (group C) and 200 mg (group D). After therapy, non-cured patients were treated further with one cycle in which the daily dose was 400 mg. Patients were subsequently observed for 9 months and efficacy was assessed by mycological examination and the growth of unaffected nails. At the end of the therapy, the cure rates in the four groups were 19.1% (A), 15.2% (B), 18.9% (C) and 17.9% (D), and no significant differences were found between each of B, C, D and A. At the end of the study, the cure rates were 76.2%, 91.3%, 78.4%, 28.6% respectively. The group that received further treatment had a cure rate of 55.6% at the end of the first month and of 83.3% in the second month. Drug tolerability was equally good in the four groups. Intermittent pulse therapy with a daily dose of 400 mg had the highest cure rate. Treatment of improved but non-cured patients with a dose of 400 mg intermittent pulse therapy markedly increased the cure rate. All treatment regimens were well tolerated.

Adolescent↗
PubMed1999

Association of oestrogen receptor gene polymorphisms with outcome of ovarian stimulation in patients undergoing IVF.

Oestrogen plays an important role in follicular formation and oocyte maturation via its receptor (ER). Many studies have shown association of the ER gene polymorphisms with a variety of pathological conditions. In this study we have examined the relationship of a common PvuII and a rare BstUI polymorphism in the ER gene to the mean numbers of follicles and oocytes, their mean ratios, mean number of embryos, mean oestrogen concentrations, mean size of the follicles and pregnancy rates. Analyses were carried out in 200 local Chinese patients undergoing in-vitro fertilization (IVF) and embryo transfer in three consecutive cycles. The mean follicular number, oocyte number, embryo number, follicular size and pregnancy rate were significantly smaller in patients homozygous for PvuII polymorphism (P < 0.001). These results indicate that PvuII polymorphism may be associated with ovarian follicular development and subsequently with the pregnancy rate. This study supports the view that genetic variability in the ER gene may have a role in the quality of the ovarian follicles in stimulation, which may affect implantation. However BstUI polymorphism was not found in either the IVF or control groups, suggesting that it has no role in the local Chinese population.

Adult↗
PubMed1999

Respiratory chain inhibition induces tolerance to focal cerebral ischemia.

The authors show that the inhibitor of the succinate dehydrogenase, 3-nitroproprionic acid (3-NPA), which in high doses and with chronic administration is a neurotoxin, can induce profound tolerance to focal cerebral ischemia in the rat when administered in a single dose (20 mg/kg) 3 days before ischemia. Infarcts were approximately 70% and 35% smaller in the 3-NPA preconditioned groups of permanent and transient focal cerebral ischemia, respectively. This regimen of 3-NPA preconditioning neither induced necrosis, apoptosis, or any other histologically detectable damage to the brain, nor did it affect behavior of the animals. 3-NPA led to an immediate (1-hour) and long-lasting (3-day) decrease in succinate dehydrogenase activity (30% reduction) throughout the brain, whereas only a short metabolic impairment occurred (ATP decrease of 35% within 30 minutes, recovery within 2 hours). The authors found that 3-NPA induces a burst of reactive oxygen species and the free radical scavenger dimethylthiourea, when administered shortly before the 3-NPA stimulus, completely blocked preconditioning. Inhibition of protein synthesis with cycloheximide given at the time of 3-NPA administration completely inhibited preconditioning. The authors were unsuccessful in showing upregulation of mRNA for the manganese superoxide dismutase, and did not detect increased activities of the copper-zinc and manganese superoxide dismutases, prototypical oxygen free radicals scavenging enzymes, after 3-NPA preconditioning. The authors conclude that it is possible to pharmacologically precondition the brain against focal cerebral ischemia, a strategy that may in principal have clinical relevance. The data show the relevance of protein synthesis for tolerance, and suggests that oxygen free radicals may be critical signals in preconditioning.

Animals↗
PubMed1999

Fosinopril and hydrochlorothiazide combination versus individual components: lack of a pharmacokinetic interaction.

OBJECTIVE: To evaluate the pharmacokinetic interaction and bioequivalence of a combination formulation of the angiotensin-converting enzyme inhibitor fosinopril and the diuretic hydrochlorothiazide (HCTZ). METHODS: In an open-label, balanced, randomized incomplete block, three-way crossover fashion, healthy men received single doses of three of four regimens in one of two independent studies. Regimens for study A (36 subjects): (1) fosinopril 10-mg tablet, (2) HCTZ 12.5-mg tablet, (3) a combination tablet of fosinopril 10 mg plus HCTZ 12.5 mg, or (4) coadministered tablets of fosinopril 10 mg and HCTZ 12.5 mg. Study B (40 subjects) received: (1) fosinopril 20-mg tablet, (2) HCTZ 12.5-mg tablet, (3) a combination tablet of fosinopril 20 mg plus HCTZ 12.5 mg, or (4) coadministered tablets of fosinopril 20 mg and HCTZ 12.5 mg. RESULTS: There was no evidence of any significant effect of HCTZ on the pharmacokinetics of fosinoprilat, based on maximum concentration value, AUC, or cumulative urinary recovery over 24 hours. Fosinoprilat had no clinically important effect on the pharmacokinetics of HCTZ only slightly decreasing its AUC by 14% in study A. Coadministration was well tolerated; no new adverse events were reported with the combination tablet. CONCLUSIONS: Fosinopril and HCTZ in a combination tablet display pharmacokinetic profiles similar to those achieved when either drug is administered alone or when coadministered in separate tablets. When used with HCTZ, the favorable pharmacokinetic feature of fosinopril, dual and compensatory pathways of renal and hepatic elimination, is preserved.

Adolescent↗
PubMed1999

[Detection of stable expression of human interlukin-2 gene in transfected keratinocytes].

OBJECTIVE: To investigate the stable expression and secretion of human interlukin-2 gene in transfected keratinocytes. METHODS: Keratinocytes were transfected with lipofectamine and selected by G418. Then the samples were analyzed with the techniques of DNA dot blot, RNA dot blot, hybridization in situ, immunohistochemistry, Western blot and MTT. RESULTS: The positive signals were observed in transfected keratinocytes by DNA dot blot, RNA dot blot, hybridization in situ and immunohistochemistry. With Western blot analysis, a specific band exhibiting a molecular weight of 15,000 was detected in transfected keratinocytes, which was in acordance with that of IL-2. The expression of IL-2 can maintain for up to 1 month. The amounts of IL-2 in the supernatants of two and four passages transfected keratinocytes were 27.7 U/ml and 15.0 U/ml, respectively. CONCLUSION: Keratinocytes have the potential for stable gene expression and secretion of active transgene products. Thus, it is possible to use keratinocytes as a target cell for gene transfection, gene expression and even gene therapy.

Cells, Cultured↗
PubMed1999

Effect of integrin on procollagen synthesis by fibroblasts from scleroderma.

OBJECTIVE: To investigate the effect of integrin on fibroblasts from scleroderma in the production of procollagen. METHODS: Phosphorothioate modified antisense oligonucleotides were used to interfere with the expression of integrin alpha 5 or beta 1 subunit on fibroblasts from 10 cases of scleroderma, and then the changes of procollagen mRNA due to the decline in level of integrin were observed by using RT-PCR. RESULTS: Expression of integrin alpha 5 or beta 1 subunit was specifically inhibited by their corresponding antisense oligonucleotides. Fibroblasts with decreased expression of alpha 5 or beta 1 subunit produced lower level of procollagen alpha 1 (I), alpha 1 (III) mRNA in comparison to the control. CONCLUSION: Overproduction of procollagen may be inhibited at the level of transcription by lowering the expression of integrin alpha 5 or beta 1 subunit on fibroblasts in scleroderma.

Adult↗
PubMed1999

[Animal experiment of the ketamine effects on traumatic brain injury resulting from impact maxillofacial injury].

OBJECTIVE: To test the effects of ketamine (KET) on traumatic brain injury resulting from impact maxillofacial injury. METHODS: The rat models with middle serious traumatic brain injury were obtained by falling impaction at maxillofacial region of the rats, then the animals were treated with KET in the early stage, and the alteration of the neural function in those rats was observed at the same time. RESULTS: There was obvious difference between the rats in the KET-treating groups and the control group. It took about 2 days fewer than that of the control group for rats in the KET-treating group to recover. CONCLUSION: KET can shorten the recovery time of rats with brain injury resulting from impact maxillofacial injury.

Animals↗
PubMed1999

[Separation and detection of tryptophan metabolites in biological samples].

A reversed phase HPLC method for separation and determination of the major tryptophan (TRP) metabolites in both pyrrolas pathway and TRP hydroxylase pathway, including TRP, kynurenine(KN), 3-hydroxykynurenine(3-HKN), kynurenic acid(KA), xanthurenic acid(XA), 5-hydroxytryptophan(5-HTP), 5-hydroxytryptamin(5-HT) and 5-hydroxyindoleacetic acid(5-HIAA), has been developed by sequential optimization of mobile phase based on acetate buffer and methanol. Trichloroacetic acid(TCA) was used as ion-pairing reagent to increase the retention of 3-HKN. The effects of pH and concentrations of TCA on separation were studied. Good separation can be achieved at pH 4.0-5.0 of mobile phase in less than 25 min. When TCA was not used, 3-HKN was hard to be detected in biological samples. The maximum retention of 3-HKN was obtained at pH 4.0 with mobile phase containing 50 mmol/L TCA. Combination of electrochemical (ED) and ultraviolet (UV) detection was used in which ED was responsible for detection of 3-HKN, 5-HTP, 5-HT and 5-HIAA and UV for the others. The influences of potential of ED and wavelength of UV on detection were studied. The optimization of conditions for separation and detection in different biological samples was also discussed.

Chromatography, High Pressure Liquid↗
PubMed1998

Utilizing the C-terminal cleavage activity of a protein splicing element to purify recombinant proteins in a single chromatographic step.

A conventional affinity protein purification system often requires a separate protease to separate the target protein from the affinity tag. This paper describes a unique protein purification system in which the target protein is fused to the C-terminus of a modified protein splicing element (intein). A small affinity tag is inserted in a loop region of the endonuclease domain of the intein to allow affinity purification. Specific mutations at the C-terminal splice junction of the intein allow controllable C-terminal peptide bond cleavage. The cleavage is triggered by addition of thiols such as dithiothreitol or free cysteine, resulting in elution of the target protein while the affinity-tagged intein remains immobilized on the affinity column. This system eliminates the need for a separate protease and allows purification of a target protein without the N-terminal methionine. We have constructed general cloning vectors and demonstrated single-column purification of several proteins. In addition, we discuss several factors that may affect the C-terminal peptide bond cleavage activity.

Affinity Labels↗
PubMed1998

Proteasome-mediated degradation of apolipoprotein B targets both nascent peptides cotranslationally before translocation and full-length apolipoprotein B after translocation into the endoplasmic reticulum.

A major portion of newly synthesized apolipoprotein B (apoB) is degraded intracellularly. This degradation has been demonstrated to be mediated largely by the ubiquitin-proteasome pathway. We examined whether nascent apoB polypeptides or full-length apoB is selectively retrotranslocated from the endoplasmic reticulum into the cytosol for degradation. Herein, we found that full-length apoB as well as partial-length apoB peptides are ubiquitinated in HepG2 cells, and ubiquitination is an exclusively cytosolic process. Calnexin, which binds specifically to glycoproteins, has been postulated to promote apoB folding and complete translocation; we found that ubiquitinated apoB is bound to calnexin, suggesting that ubiquitinated apoB is glycosylated. In addition to calnexin binding, we have other pieces of evidence that the full-length intracellular ubiquitinated apoB is glycosylated, because (i) it binds to concanavalin A, and (ii) glycan can be demonstrated in the full-length ubiquitinated apoB by a chemical detection method involving oxidation of adjacent hydroxyl groups in the glycan moiety. Because glycosylation occurs inside the endoplasmic reticulum, the full-length glycosylated apoB must have been retrotranslocated into the cytosol for ubiquitination and proteasome-mediated degradation. Next we synchronized translation in HepG2 cells by puromycin treatment. A pulse-chase experiment using [35S]methionine labeling of intracellular apoB in these synchronized cells demonstrated that nascent partial-length apoB peptides are also ubiquitinated cotranslationally. We conclude that the ubiquitin proteasome-mediated degradation of apoB targets both nascent peptides cotranslationally before translocation as well as full-length apoB after its translocation into the endoplasmic reticulum.

Acetylcysteine↗