Kung Chia Young

Andrographolide exerts anti-hepatitis C virus activity by up-regulating haeme oxygenase-1 via the p38 MAPK/Nrf2 pathway in human hepatoma cells.

This study aimed to evaluate the anti‐hepatitis C virus (HCV) activity of andrographolide, a diterpenoid lactone extracted from Andrographis paniculata, and to identify the signalling pathway involved in its antiviral action.

Incisional Hernia after Laparotomy: Prospective Randomized Comparison between Early-absorbable and Late-absorbable Suture Materials

Abstract. Incisional hernia is a serious postoperative complication of laparotomy. Selecting an appropriate suture material may lessen such morbidity. This study undertook a prospective, randomized comparison of early-absorbable polyglactin 910 suture versus late-absorbable polydioxanone loop suture for fascial closure after abdominal surgery. A series of 340 consecutive patients undergoing elective laparotomy were randomized to have fascial closure with either polyglactin 910 suture or polydioxanone loop suture between October 1993 and August 1996. A 2-year follow-up revealed that 23 patients had died, and the overall mortality rate was 6.8% (23/340). Ten (10/340, 2.9%) patients, including seven with polyglactin 910 suture and three with polydioxanone loop suture, developed incisional hernias. The early postoperative evaluation revealed an incidence of wound infection of 4.1% (14/340). The development of incisional hernia was not secondary to postoperative wound infection in this study. Among these 340 patients, 192 had malignant diseases and 148 had nonmalignant ones. Fascial closure with polyglactin 910 suture was associated with more incisional hernias than that with polydioxanone loop suture, with marginal significance for patients in the malignant group (4.7% versus 0%, p= 0.07) but not in the nonmalignant group (2.6% versus 4.2%, p= 0.67). In conclusion, abdominal closure with a late-absorbable polydioxanone loop suture may be beneficial to patients with a malignant disease for preventing incisional hernia.

Estrogen receptor-α polymorphism in a Taiwanese clinical breast cancer population: a case–control study

IntroductionReceptor-mediated estrogen activation participates in the development and progression of breast cancer. Estrogen receptor (ER)-α polymorphism has been found to be associated with breast cancer and clinical features of the disease in Caucasians. Epidemiologic studies have revealed that age–incidence patterns of breast cancer in Asians differ from those in Caucasians. Genomic data for ER-α in either population is therefore of value in the clinical setting for that ethnic group.MethodsA case–control study was conducted to establish a database of ER-α polymorphisms in a Taiwanese population in order to compare Western and Taiwanese (Asian) distributions and to evaluate ER-α polymorphism as an indicator of clinical outcome. The ER-α gene was scanned in a Taiwanese clinical breast cancer group (189 patients) and in healthy individuals (177 healthy control individuals). PCR single-strand conformation polymorphism technology was employed and real-time PCR melting curve analysis was performed.ResultsThree sites of silent single nucleotide polymorphism (SNPs) were found, as reported previously in Western studies, but at significantly different frequencies. Among the three SNPs, the frequency of allele 1 (TCT → TCC) in codon 10 was significantly lower in breast cancer patients (32.0%) than in control individuals (40.4%; P = 0.018). We found that allele 1 (ACG → ACA) in codon 594 was less common in breast cancer patients with a family history of breast cancer (5.9%) than in those without such a history (19.6%; P = 0.049). Individually, both allele 1 in codon 325 (CCC → CCG) and allele 1 in codon 594 exhibited a reverse association with the occurrence of lymph node metastasis. Furthermore, incorporation of both SNP markers further increased predictive accuracy.ConclusionsOur data suggest that ER-α polymorphisms are correlated with various aspects of breast cancer in Taiwan. ER-α genotype, as determined during presurgical evaluation, might represent a surrogate marker for predicting breast cancer lymph node metastasis.

Microarray profiling of gene expression patterns in bladder tumor cells treated with genistein.

Microarray technology was used to gain an insight into the molecular events of tumor cell growth inhibition mediated by the soy isoflavone genistein. For this, a susceptible bladder tumor line TCCSUP was treated with the inhibitory dose (50 microM) of genistein for various periods of time, followed by mRNA isolations, cDNA probe preparations, and hybridization individually to cDNA chips containing 884 sequence-verified known human genes. After analyzing the hybridization signals with a simple quantitative method developed by this study, we detected that egr-1, whose expression has been associated with proliferation and differentiation, was transiently induced and this expression pattern was later confirmed by RT-PCR. Thus, microarray technology is a reliable and powerful tool for profiling gene expression patterns in many biological systems related to cancer. We further detected many groups of genes with distinct expression profiles and most of them encode for proteins that regulate the signal transduction or the cell cycle pathways. These genes warrant further investigation as regards their roles in the susceptibility of the tumor cell line to the antitumor drug.

Comparative proteomic profiling of plasma very-low-density and low-density lipoproteins

BACKGROUND Low-density lipoprotein (LDL) is a natural metabolite of very-low-density lipoprotein (VLDL) in the circulation. Systematic investigation of total protein components and dynamics might provide insights into this normal metabolic process. METHODS VLDL and LDL were purified from normolipidemia pooled plasma by gradient ultracentrifugation with either ionic or non-ionic media. The protein contents were compared by liquid chromatography tandem mass analyses based on isobaric tag for relative and absolute quantitation and two-dimensional gel electrophoresis. RESULTS Our comparative lipoproteomes revealed 21 associated proteins. Combined with Western blot analysis, and on the basis of the differential expression levels we classified them into 3 groups: (i) VLDL>LDL [apolipoprotein (apo) A-IV, apo(a), apoCs, apoE, apoJ and serum amyloid A-4]; (ii) VLDL<LDL [albumin, alpha-1-antitrypsin, apoD, apoF, apoM, and paraoxonase-1]; and (iii) VLDL=LDL [apoA-I, apoA-II, apoB-100, apoL-I and prenylcysteine oxidase-1]. The apoA-I level positively correlated with PCYOX1 but negatively with apoM in VLDL and LDL. Furthermore, the two-dimensional maps displayed 5 apoA-I isoforms in which phosphorylation at Ser55, Ser166, Thr185, Thr221 and Ser252 residues were identified. CONCLUSIONS This study revealed the VLDL- and LDL lipoproteomes and the full-spectrum protein changes during physiological VLDL-to-LDL transition. It provides a valuable dataset VLDL and LDL proteomes potentially applied to the development of diagnostics.

Simultaneous Quantification and Genotyping of Hepatitis B Virus for Genotypes A to G by Real-Time PCR and Two-Step Melting Curve Analysis

ABSTRACT Both the viral titer and the genotype significantly determine clinical outcomes and responses to antiviral treatment in chronic hepatitis B virus (HBV) infection. A method was developed for large-scale A-to-G genotyping with simultaneous viral quantification. The assay was run on a LightCycler instrument using hybridization probes. The genotype was determined from the melting points of the probes in a two-step manner. Set 1 amplicons differentiated genotypes B, E, and F from A, C, D, and G and simultaneously quantified viremia by real-time PCR. Melting curve analysis using the set 2-1 amplicon or the set 2-2 amplicon reaction mixture was then used to differentiate these genotype groups into single genotypes. HBV DNA quantification was consistent with that of the Amplicor assay and linear in a range from 102 to 1013 copies/ml. By comparison with the restriction fragment length polymorphism method, 92.3% of 441 samples were accurately genotyped by the current assay. The method should be useful for genotyping and quantification of HBV DNA in areas where all genotypes exist.

Very low-density lipoprotein/lipo-viro particles reverse lipoprotein lipase-mediated inhibition of hepatitis C virus infection via apolipoprotein C-III

Objective Circulating hepatitis C virus (HCV) virions are associated with triglyceride-rich lipoproteins, including very low-density lipoprotein (VLDL) and low-density lipoprotein (LDL), designated as lipo-viro-particles (LVPs). Previous studies showed that lipoprotein lipase (LPL), a key enzyme for hydrolysing the triglyceride in VLDL to finally become LDL, may suppress HCV infection. This investigation considers the regulation of LPL by lipoproteins and LVPs, and their roles in the LPL-mediated anti-HCV function. Design The lipoproteins were fractionated from normolipidemic blood samples using iodixanol gradients. Subsequent immunoglobulin-affinity purification from the canonical VLDL and LDL yielded the corresponding VLDL-LVP and LDL-LVP. Apolipoprotein (apo) Cs, LPL activity and HCV infection were quantified. Results A higher triglyceride/cholesterol ratio of LDL was found more in HCV-infected donors than in healthy volunteers, and the triglyceride/cholesterol ratio of LDL-LVP was much increased, suggesting that the LPL hydrolysis of triglyceride may be impaired. VLDL, VLDL-LVP, LDL-LVP, but not LDL, suppressed LPL lipolytic activity, which was restored by antibodies that recognised apoC-III/-IV and correlated with the steadily abundant apoC-III/-IV quantities in those particles. In a cell-based system, treatment with VLDL and LVPs reversed the LPL-mediated inhibition of HCV infection in apoC-III/-IV-dependent manners. A multivariate logistic regression revealed that plasma HCV viral loads correlated negatively with LPL lipolytic activity, but positively with the apoC-III content of VLDL. Additionally, apoC-III in VLDL was associated with a higher proportion of HCV-RNA than was IgG. Conclusion This study reveals that LPL is an anti-HCV factor, and that apoC-III in VLDL and LVPs reduces the LPL-mediated inhibition of HCV infection.

Identification of hnRNPH1, NF45, and C14orf166 as novel host interacting partners of the mature hepatitis C virus core protein.

The hepatitis C virus core protein (HCVc) forms the viral nucleocapsid and is involved in viral persistence and pathogenesis, possibly by interacting with host factors to modulate viral replication and cellular functions. Here, we identified 36 cellular protein candidates by one-dimensional SDS-PAGE and LC-MS/MS-based proteomics after affinity purification with HCVc174, a matured form of HCVc from HCV-1b genotype, tagged with biotin and calmodulin-binding peptide/protein A at N- and C-termini, respectively. By pull-down and confocal imaging techniques, we confirmed that heterogeneous nuclear ribonucleoprotein H1 (hnRNPH1), nuclear factor 45 (NF45), and C14orf166 are novel HCVc174-interacting host proteins, known to participate in mRNA metabolism, gene regulation, and microtubule organization, respectively. Unlike the other 2 proteins, NF45 interacted with HCVc174 in an RNA-dependent manner. These 3 proteins colocalized with ectopic HCVc-1b in both the cytoplasm and nucleus, which demonstrated their spatial interaction with naturally translocated HCVc174 after HCVc biogenesis. Such colocalization, however, shifted to the cytoplasm in cells with replicating virus of 1b or 2a genotype, indicating that active viral replication confined these interacting proteins in the cytoplasm. Collectively, our findings suggest that spatial interactions of hnRNPH1, NF45, and C14orf166 with HCVc174 likely modulate HCV or cellular functions during acute and chronic HCV infection.

Association of intrahepatic cccDNA reduction with the improvement of liver histology in chronic hepatitis B patients receiving oral antiviral agents.

Covalently closed circular DNA (cccDNA) of hepatitis B virus (HBV) is difficult to eradicate using current antiviral therapy. This study compares cccDNA reduction with relation to liver histology in nucleoside/nucleotide‐naïve chronic hepatitis B patients receiving oral antiviral monotherapy (n = 35), including entecavir (ETV, n = 13), adefovir dipivoxil (ADV, n = 22) or placebo (n = 14). Serum HBV DNA, intrahepatic total HBV DNA and cccDNA are quantified. Histological hepatic examination is performed at baseline and at 48 weeks of treatment. Treatment with ETV or ADV shows significant median reduction in serum HBV DNA (−6.21 and −4.27 log10 copies/mL) and intrahepatic total HBV DNA (−1.69 and −1.23 log10 copies/cell). Intrahepatic cccDNA levels are reduced slightly in the ETV and the ADV groups, but do not differ statistically from the placebo group (−0.17 vs. −0.01 vs. 0.02 copies/cell). Only the level of intrahepatic cccDNA correlates with Knodell necroinflammation activity (r = 0.527, P < 0.001) and Ishak fibrosis severity (r = 0.348, P = 0.015) before treatment. Multivariate logistic regression analysis indicates that treatment‐induced cccDNA reduction is associated with improved necroinflammation (P = 0.041) and fibrosis (P = 0.026). In conclusion, baseline intrahepatic cccDNA loads correlate with histologic activity. Although one‐year ETV or ADV treatment is insufficient for cccDNA eradication, oral antiviral therapies may improve liver histology, probably by suppressing intrahepatic cccDNA. J. Med. Virol. 83:602–607, 2011.

Apolipoprotein J, a glucose-upregulated molecular chaperone, stabilizes core and NS5A to promote infectious hepatitis C virus virion production

BACKGROUND & AIMS Hepatitis C virus (HCV) infection leads to glucose abnormality. HCV depends on lipid droplets (LDs) and very-low density lipoproteins for assembly/releasing; however, the components and locations for this process remain unidentified. Apolipoprotein J (ApoJ), upregulated by glucose, functions as Golgi chaperone of secreted proteins and resides abundantly in very-low density lipoproteins. This study investigates the interplay between glucose, ApoJ and HCV virion production. METHODS The effects of high glucose on ApoJ expression and HCV production were evaluated with cultivated HuH7.5, primary human hepatocytes, and in treatment naive chronic hepatitis C patients. How ApoJ affects HCV lifecycle was assessed using siRNA knockdown strategy in JFH1 infected and subgenomic replicon cells. The interactions and locations of ApoJ with viral and host components were examined by immunoprecipitation, immunofluorescence and subcellular fractionation experiments. RESULTS HCV infection increased ApoJ expression, which in parallel with HCV infectivity was additionally elevated with high glucose treatment. Serum ApoJ correlated positively with fasting blood glucose concentration and HCV-RNA titre in patients. ApoJ silencing reduced intracellular and extracellular HCV infectivity and extracellular HCV-RNA, but accumulated intracellular HCV-RNA in HCV-infected cells. ApoJ interacted with HCV core and NS5A and stabilized the dual protein complex. HCV infection dispersed cytoplasmic ApoJ from the compact zones of the Golgi to encircle LDs, where co-localization of the core, NS5A, HCV-RNA, subcellular markers for LDs, endoplasmic reticulum (ER), Golgi, and membrane contact sites occurred. CONCLUSIONS ApoJ facilitates infectious HCV particle production via stabilization of core/NS5A, which might surround LDs at the ER-Golgi membrane contact site.