Cytosolic phospholipase A2α (cPLA2α) is a rate-limiting key enzyme controlling the release of arachidonic acid (AA) substrate for the synthesis of prostaglandins and leukotrienes. This study was designed to explore the role of hepatocyte cPLA2α in Fas-mediated liver injury, in vivo.
Transgenic mice with targeted expression of cPLA2α under control of the albumin-promoter enhancer and wild-type mice were injected intraperitoneally with anti-Fas antibody Jo2 or lipopolysaccharide plus D-galactosamine and monitored for liver injury and survival at various time points.
The cPLA2α Tg mice resist Fas-induced liver failure, as reflected by the lower serum transaminase levels, fewer apoptotic hepatocytes, reduced caspase activation, and reduced PARP cleavage when compared to the matched wild type mice. Inhibition of cPLA2α by its pharmacological inhibitor, pyrrolidine, enhanced Jo2-induced liver injury in both cPLA2α Tg and wild type mice. Hepatic overexpression of cPLA2α increases the expression of EGFR in the liver and the EGFR inhibitor, AG1478, exacerbated Jo2-mediated liver injury. The cPLA2α transgenic mice develop more prominent liver tissue damage than wild-type mice after LPS/D-galactosamine injection.
Hepatocyte cPLA2α protects against Fas-induced liver injury and this effect is mediated at least in part through upregulation of EGFR.
Cytosolic phospholipase A2; liver; Fas; apoptosis; epidermal growth factor receptor; LPS
The novel supervised learning method of vertex discriminant analysis (VDA) has been demonstrated for its good performance in multicategory classification. The current paper explores an elaboration of VDA for nonlinear discrimination. By incorporating reproducing kernels, VDA can be generalized from linear discrimination to nonlinear discrimination. Our numerical experiments show that the new reproducing kernel-based method leads to accurate classification for both linear and nonlinear cases.
Gaussian reproducing kernels; nonlinear classifier; regular simplex; reproducing kernel Hilbert space
Docosahexaenoic acid (DHA) has been reported to induce tumor cell death by apoptosis. However, little is known about the effects of DHA on autophagy, another complex well-programmed process characterized by the sequestration of cytoplasmic material within autophagosomes. Here we show that DHA increased both the level of microtubule-associated protein 1 light chain 3 and the number of autophagic vacuoles without impairing autophagic vesicle turnover, indicating that DHA induces not only apoptosis but also autophagy. We also observed that DHA-induced autophagy was accompanied by p53 loss. Inhibition of p53 increased DHA-induced autophagy and prevention of p53 degradation significantly led to the attenuation of DHA-induced autophagy, suggesting that DHA-induced autophagy is mediated by p53. Further experiments showed that the mechanism of DHA-induced autophagy associated with p53 attenuation involved an increase in the active form of AMP-activated protein kinase and a decrease in the activity of mammalian target of rapamycin. In addition, compelling evidence for the interplay between autophagy and apoptosis induced by DHA is supported by the findings that autophagy inhibition suppressed apoptosis and further autophagy induction enhanced apoptosis in response to DHA treatment. Overall, our results demonstrate that autophagy contributes to the cytotoxicity of DHA in cancer cells harboring wild-type p53.
DHA; autophagy; apoptosis; p53; cancer; mTOR; AMPK; p27
Microsomal prostaglandin E synthase-1 (mPGES-1) is a key enzyme that couples with cyclooxygenase-2 (COX-2) for the production of PGE2. Although COX-2 is known to mediate the growth and progression of several human cancers including hepatocellular carcinoma (HCC), the role of mPGES-1 in hepatocarcinogenesis is not well established. This study provides novel evidence for a key role of mPGES-1 in HCC growth and progression. Forced overexpression of mPGES-1 in two HCC cell lines (Hep3B and Huh7) increased tumor cell growth, clonogenic formation, migration and invasion, whereas knockdown of mPGES-1 inhibited these parameters, in vitro. In a SCID mouse tumor xenograft model, mPGES-1 overexpressed cells formed palpable tumors at earlier time points and developed larger tumors when compared to the control (p<0.01); in contrast, mPGES-1 knockdown delayed tumor development and reduced tumor size (p<0.01). Mechanistically, mPGES-1-induced HCC cell proliferation, invasion and migration involve PGE2 production and activation of early growth response 1 (EGR1) and β-catenin. Specifically, mPGES-1-derived PGE2 induces the formation of EGR1-β-catenin complex, which interacts with TCF4/LEF1 transcription factors and activates the expression of β-catenin downstream genes. Our findings depict a novel crosstalk between mPGES-1/PGE2 and EGR1/β-catenin signaling that is critical for hepatocarcinogenesis.
Microsomal prostaglandin E synthase-1 (mPGES-1); β-catenin; early growth response 1 (EGR1); hepatocellular carcinoma (HCC); liver
Haploidentical hematopoietic cell transplantation (HCT) has been used to treat hematologic malignancies but it is unknown whether the procedure is more effective in adults or children. To address this question, we analyzed patients aged 1–65 years old receiving myeloablative conditioning regimens followed by family 2 to 3 antigen HLA-mismatched HCT and reported to the Center for International Blood and Marrow Transplant Research (CIBMTR, n=137) or performed in Dao-Pei Hospital in Beijing, China (n=181). The Dao-Pei cohort had more acute and chronic GVHD, less relapse, lower transplant related mortality (TRM) and better leukemia-free survival (LFS) than the CIBMTR cohort. Overall survival (OS) and outcomes were similar between adults and children. In the CIBMTR cohort receiving ex vivo T cell depletion (TCD), adults had higher TRM (RR 2.71, 95% CI 1.29–5.69, p=0.008) and lower overall survival (RR 1.75, 95%CI 1.08–2.84, p=0.023) than children. In the CIBMTR subset that did not receive ex vivo TCD, relapse was lower in adults compared to children (RR 0.24, 95% CI 0.07–0.80, p=0.020) but TRM, LFS and OS were similar. We conclude that outcomes in adults and children are similar overall, although children have better survival than adults if ex vivo TCD is used.
HLA-mismatched; haploidentical; blood and marrow transplantation; leukemia; antithymocyte globulin
Background & Aims
Microsomal prostaglandin E synthase-1 (mPGES-1) is a rate-limiting enzyme that is coupled with cyclooxygenase-2 (COX-2) in the synthesis of prostaglandin E2 (PGE2). Although COX-2 is involved in development and progression of various human cancers, the role of mPGES-1 in carcinogenesis has not been determined. We investigated the role of mPGES-1 in human cholangiocarcinoma growth.
We used immunohistochemical analyses to examine the expression of mPGES-1 in formalin-fixed, paraffin-embedded human cholangiocarcinoma tissues. The effects of mPGES-1 on human cholangiocarcinoma cells were determined in vitro and in SCID mice. Immunoblotting and immunoprecipitation assays were performed to determine the levels of PTEN and related signaling molecules in human cholangiocarcinoma cells with overexpression or knockdown of mPGES-1.
mPGES-1 is overexpressed in human cholangiocarcinoma tissues. Overexpression of mPGES-1 in human cholangiocarcinoma cells increased tumor cell proliferation, migration, invasion, and colony formation; in contrast, RNAi knockdown of mPGES-1 inhibited tumor growth parameters. In SCID mice with tumor xenografts, mPGES-1 overexpression accelerated tumor formation and increased tumor weight (P<0.01), whereas mPGES-1 knockdown delayed tumor formation and reduced tumor weight (P<0.01). mPGES-1 inhibited the expression of PTEN, leading to activation of the EGFR–PI3K–AKT–mTOR signaling pathways in cholangiocarcinoma cells. mPGES-1–mediated inhibition of PTEN is regulated through blocking of EGR-1 sumoylation and binding to the 5′-UTR of the PTEN gene.
mPGES-1 promotes experimental cholangiocarcinogenesis and tumor progression by inhibiting PTEN.
cancer cell signaling; biliary tract cancer; bile duct; liver
Here, for the first time, we evaluate the hypothesis that the proliferative abilities of satellite cells (SCs) isolated from Lantang (indigenous Chinese pigs) and Landrace pigs, which differ in muscle characteristics, are different. SCs were isolated from the longissimus dorsi muscle of neonatal Lantang and Landrace pigs. Proliferative ability was estimated by the count and proliferative activity of viable cells using a hemocytometer and MTT assay at different time points after seeding, respectively. Cell cycle information was detected by flow cytometry. Results showed that there was a greater (P<0.05) number of SCs in Lantang pigs compared with Landrace pigs after 72 h of culture. The percentage of cell population in S phase and G2/M phases in Lantang pigs were higher (P<0.05), while in G0/G1 phase was lower (P<0.05) in comparison with the Landrace pigs. The mRNA abundances of MyoD, Myf5, myogenin and Pax7 in SCs from Lantang pigs were higher (P<0.05), while those of myostatin, Smad3 and genes in the mammalian target of rapamycin (mTOR) pathway (with the exception of 4EBP1) were lower (P<0.05) than the Landrace pigs. Protein levels of MyoD, myogenin, myostatin, S6K, phosphorylated mTOR and phosphorylated eIF4E were consistent with the corresponding mRNA abundance. Collectively, these findings suggested that SCs in the two breeds present different proliferative abilities, and the proliferative potential of SCs in Lantang pigs is higher than in Landrace pigs.
In this article we study a semiparametric additive risks model (McKeague and Sasieni (1994)) for two-stage design survival data where accurate information is available only on second stage subjects, a subset of the first stage study. We derive two-stage estimators by combining data from both stages. Large sample inferences are developed. As a by-product, we also obtain asymptotic properties of the single stage estimators of McKeague and Sasieni (1994) when the semiparametric additive risks model is misspecified. The proposed two-stage estimators are shown to be asymptotically more efficient than the second stage estimators. They also demonstrate smaller bias and variance for finite samples. The developed methods are illustrated using small intestine cancer data from the SEER (Surveillance, Epidemiology, and End Results) Program.
Censored data; correlation; efficiency; measurement errors; missing covariates
We report here the epitaxial growth of III-nitride material on freestanding HfO2 gratings by molecular beam epitaxy. Freestanding HfO2 gratings are fabricated by combining film evaporation, electron beam lithography, and fast atom beam etching of an HfO2 film by a front-side silicon process. The 60-μm long HfO2 grating beam can sustain the stress change during the epitaxial growth of a III-nitride material. Grating structures locally change the growth condition and vary indium composition in the InGaN/GaN quantum wells and thus, the photoluminescence spectra of epitaxial III-nitride grating are tuned. Guided mode resonances are experimentally demonstrated in fabricated III-nitride gratings, opening the possibility to achieve the interaction between the excited light and the grating structure through guided mode resonance.
PACS: 78.55.Cr; 81.65.Cf; 81.15.Hi.
InGaN/GaN QWs; fast atom beam etching; molecular beam epitaxy
Hepatocellular carcinoma often develops in the setting of abnormal hepatocyte growth associated with chronic hepatitis and liver cirrhosis. Transforming growth factor-βs (TGF-βs) are multifunctional cytokines pivotal in the regulation of hepatic cell growth, differentiation, migration, extracellular matrix production, stem cell homeostasis and hepatocarcinogenesis. However, the mechanisms by which TGF-βs influence hepatic cell functions remain incompletely defined. We report herein that TGF-β regulates the growth of primary and transformed hepatocytes through concurrent activation of Smad and phosphorylation of cPLA2α, a rate-limiting key enzyme that releases arachidonic acid for production of bioactive eicosanoids. The interplays between TGF-β and cPLA2α signaling pathways were examined in rat primary hepatocytes, human hepatocellular carcinoma cells and hepatocytes isolated from the newly developed cPLA2α transgenic mice. Our data show that cPLA2α activates PPAR-γ and thus counteracts Smad2/3-mediated inhibition of cell growth. Therefore, regulation of TGF-β signaling by cPLA2α and PPAR-γ may represent an important mechanism for control of hepatic cell growth and hepatocarcinogenesis.
Transforming growth factor-β; cytosolic phospholipase A2α; peroxisome proliferator activated receptor-γ; hepatocyte; liver
We report here the fabrication of freestanding HfO2 grating by combining fast atom beam etching (FAB) of HfO2 film with dry etching of silicon substrate. HfO2 film is deposited onto silicon substrate by electron beam evaporator. The grating patterns are then defined by electron beam lithography and transferred to HfO2 film by FAB etching. The silicon substrate beneath the HfO2 grating region is removed to make the HfO2 grating suspend in space. Period- and polarization-dependent optical responses of fabricated HfO2 gratings are experimentally characterized in the reflectance measurements. The simple process is feasible for fabricating freestanding HfO2 grating that is a potential candidate for single layer dielectric reflector.
PACS: 73.40.Ty; 42.70.Qs; 81.65.Cf.
HfO2 film; grating; fast atom beam etching
Genetic variants in TRAF1C5 and PTPN22 genes have been shown to be significantly associated with arthritis rheumatoid in Caucasian populations. This study investigated the association between single nucleotide polymorphisms (SNPs) in TRAF1/C5 and PTPN22 genes and rheumatoid arthritis (RA) in a Han Chinese population. We genotyped SNPs rs3761847 and rs7021206 at the TRAF1/C5 locus and rs2476601 SNP in the PTPN22 gene in a Han Chinese cohort composed of 576 patients with RA and 689 controls. The concentrations of anti-cyclic citrullinated peptide antibodies (CCP) and rheumatoid factor (RF) were determined for all affected patients. The difference between the cases and the controls was compared using χ2 analysis.
Significant differences in SNPs rs3761847 and rs7021206 at TRAF1/C5 were observed between the case and control groups in this cohort; the allelic p-value was 0.0018 with an odds ratio of 1.28 for rs3761847 and 0.005 with an odds ratio of 1.27 for rs7021206. This significant association between rs3761847 and RA was independent of the concentrations of anti-CCP and RF. No polymorphism of rs2476601 was observed in this cohort.
We first demonstrated that genetic variants at the TRAF1/C5 locus are significantly associated with RA in Han Chinese, suggesting that TRAF1/C5 may play a role in the development of RA in this population, which expands the pathogenesis role of TRAF1/C5 in a different ethnicity.
rheumatoid arthritis; genetics; TRAF1/C5; association study; Chinese
AIM: To develop a hepatocellular carcinoma (HCC) xenograft model for studying hepatitis C virus (HCV) replication in a mice, and antiviral treatment.
METHODS: We developed a stable S3-green fluorescence protein (GFP) cell line that replicated the GFP-tagged HCV sub-genomic RNA derived from a highly efficient JFH1 virus. S3-GFP replicon cell line was injected subcutaneously into γ-irradiated SCID mice. We showed that the S3-GFP replicon cell line formed human HCC xenografts in SCID mice. Cells were isolated from subcutaneous tumors and then serially passaged multiple times in SCID mice by culturing in growth medium supplemented with G-418. The mouse-adapted S3-GFP replicon cells were implanted subcutaneously and also into the liver of SCID mice via intrasplenic infusion to study the replication of HCV in the HCC xenografts. The tumor model was validated for antiviral testing after intraperitoneal injection of interferon-α (IFN-α).
RESULTS: A highly tumorigenic S3-GFP replicon cell line was developed that formed subcutaneous tumors within 2 wk and diffuse liver metastasis within 4 wk in SCID mice. Replication of HCV in the subcutaneous and liver tumors was confirmed by cell colony assay, detection of the viral RNA by ribonuclease protection assay and real-time quantitative reverse transcription polymerase chain reaction. High-level replication of HCV sub-genomic RNA in the tumor could be visualized by GFP expression using fluorescence microscopy. IFN-α cleared HCV RNA replication in the subcutaneous tumors within 2 wk and 4 wk in the liver tumor model.
CONCLUSION: A non-infectious mouse model allows us to study replication of HCV in subcutaneous and metastatic liver tumors. Clearance of HCV by IFN-α supports use of this model to test other anti-HCV drugs.
Hepatitis C virus; Hepatocellular carcinoma; Tumor xenograft; SCID mouse; Interferon-α; Antiviral agent; Virus replication
Insight into the mechanisms of organ engraftment and acquired tolerance has made it possible to facilitate these mechanisms, by tailoring the timing and dosage of immunosuppression in accordance with two therapeutic principles: recipient pretreatment, and minimum use of post-transplant immunosuppression. We aimed to apply these principles in recipients of renal and extrarenal organ transplants.
82 patients awaiting kidney, liver, pancreas, or intestinal transplantation were pretreated with about 5 mg/kg of a broadly reacting rabbit antithymocyte globulin during several hours. Post-transplant immunosuppression was restricted to tacrolimus unless additional drugs were needed to treat breakthrough rejection. After 4 months, patients on tacrolimus monotherapy were considered for dose-spacing to every other day or longer intervals.
We frequently saw evidence of immune activation in graft biopsy samples, but unless this was associated with graft dysfunction or serious immune destruction, treatment usually was not intensified. Immunosuppression-related morbidity was virtually eliminated. 78 (95%) of 82 patients survived at 1 year and at 13–18 months. Graft survival was 73 (89%) of 82 at 1 year and 72 (88%) of 82 at 13–18 months. Of the 72 recipients with surviving grafts, 43 are on spaced doses of tacrolimus monotherapy: every other day (n=6), three times per week (11), twice per week (15), or once per week (11).
The striking ability to wean immunosuppression in these recipients indicates variable induction of tolerance. The simple therapeutic principles are neither drug-specific nor organ-specific. Systematic application of these principles should allow improvements in quality of life and long-term survival after organ transplantation.
Hepatocellular carcinoma (HCC) is a common human cancer with high mortality and currently there is no effective chemoprevention or systematic treatment. Recent evidence suggests that COX-2-derived PGE2 and Wnt/β-catenin signaling pathways are implicated in hepatocarcinogenesis. Here we report that ω-3 PUFAs, docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), inhibit HCC growth through simultaneously inhibition of COX-2 and β-catenin. DHA and EPA treatment resulted in a dose-dependent reduction of cell viability with cleavage of PARP, caspase-3 and caspase-9 in three human HCC cell lines (Hep3B, Huh-7, HepG2). In contrast, arachidonic acid (AA), a ω-6 PUFA, exhibited no significant effect. DHA and EPA treatment caused dephosphorylation and thus activation of GSK-3β, leading to β-catenin degradation in Hep3B cells. The GSK3-β inhibitor, LiCl, partially prevented DHA-induced β-catenin protein degradation and apoptosis. Additionally, DHA induced the formation of β-catenin/Axin/GSK-3β binding complex, which serves as a parallel mechanism for β-catenin degradation. Furthermore, DHA inhibited PGE2 signaling through downregulation of COX-2 and upregulation of the COX-2 antagonist, 15-hydroxyprostaglandin dehydrogenase (15-PGDH). Finally, the growth of HCC in vivo was significantly reduced when mouse HCCs (Hepa1–6) were inoculated into the Fat-1 transgenic mice which express a Caenorhabditis elegans desaturase converting ω-6 to ω-3 PUFAs endogenously. These findings provide important preclinical evidence and molecular insight for utilization of ω-3 PUFAs for the chemoprevention and treatment of human HCC.
hepatocellular carcinoma; omega-3 polyunsaturated fatty acid; beta-catenin; cyclooxygenase-2; prostaglandin E2; 15-PGDH
COX-2-derived PGs participate in a number of pathophysiological responses such as inflammation, carcinogenesis, and modulation of cell growth and survival. This study utilized complementary approaches of COX-2 transgenic and knockout mice models to evaluate the mechanism of COX-2 in Fas-induced hepatocyte apoptosis and liver failure, in vivo. We generated transgenic mice with targeted expression of COX-2 in the liver by using the albumin promoter-enhancer driven vector. The COX-2 transgenic (Tg), COX-2 knockout (KO), and wild type mice were treated with the anti-Fas antibody Jo2 (0.5 µg/g body weight) for 4–6 hours and the extent of liver injury was assessed by histopathology, serum transaminases, TUNEL staining and caspase activation. The COX-2 Tg mice showed resistance to Fas-induced liver injury when compared to the wild type mice, as reflected by the lower ALT and AST levels, less liver damage and less hepatocyte apoptosis (p<0.01). In contrast, the COX-2 KO mice showed significantly higher serum ALT and AST levels, more prominent hepatocyte apoptosis, and higher levels of caspase-8, 9, 3 activities than the wild type mice (p<0.01). The COX-2 Tg livers express higher levels of epidermal growth factor receptor (EGFR) than the wild type controls; the COX-2 KO livers express lowest levels of EGFR. Pretreatment with the COX-2 inhibitor (NS-398) or the EGFR inhibitor (AG1478) exacerbated Jo2-mediated liver injury and hepatocyte apoptosis. These findings demonstrate that COX-2 prevents Fas-induced hepatocyte apoptosis and liver failure at least in part through upregulation of EGFR.
Cyclooxygenase-2; liver; Fas; apoptosis; epidermal growth factor receptor
Reactive oxygen species (ROS) activate retinoid-containing quiescent hepatic stellate cells (qHSCs) to retinoid-deficient fibrogenic myofibroblast-like cells (aHSCs). However, ROS also cause apoptosis of aHSCs, and apoptotic aHSCs are observed in inflammatory fibrotic liver. Here, we investigated mechanisms of the effects of oxidative stress on the survival of qHSCs and aHSCs. HSCs from normal rat liver were used after overnight culture (qHSCs), or in 3–5 passages (aHSCs). For in vivo induction of oxidative stress, tert-butylhydroperoxide was injected into control and CCl4-induced cirrhotic rats. Spontaneous caspase-3 activation and apoptosis, observed in cultured qHSCs, decreased with time and were unaffected by superoxide. In contrast, superoxide caused caspase-3 and p38-MAPK activation, reduction in Bcl-xL expression, and apoptosis in aHSCs. Inhibition of caspase-3 and p38-MAPK did not affect the viability of qHSCs in the absence or presence of superoxide, but inhibited superoxide-induced death of aHSCs. Glutathione (GSH) level and activities of superoxide dismutase (SOD), catalase and glutathione peroxidase (GPx) were lower in aHSCs than qHSCs. Superoxide increased GSH content, and activities of SOD, catalase and GPx in qHSCs but not in aHSCs. Incubation of 13-cis-retinoic acid (RA)-treated aHSCs with superoxide increased their GSH content significantly, and prevented superoxide-induced p38-MAPK and caspase-3 activation while dramatically reducing the extent of apoptosis. Finally, oxidative stress induced in vivo caused apoptosis of aHSCs in cirrhotic but not of qHSCs in control rats. These results suggest that the absence of retinoids render aHSCs susceptible to superoxide-induced apoptosis via caspase-3 and p38-MAPK activation.
Hepatocytes express adrenergic receptors (ARs) that modulate several functions, including liver regeneration, hepatocyte proliferation, glycogenolysis, gluconeogenesis, synthesis of urea and fatty acid metabolism. Adrenergic hepatic function in adults is mainly under the control of α1-ARs; however, the mechanism through which they influence diverse processes remains incompletely understood. This study describes a novel α1-AR-mediated transactivation of signal transducer and activator of transcription-3 (Stat3) in primary and transformed hepatocytes. Treatment of primary rat hepatocytes with the α1-AR agonist, phenylephrine (PE), induced a rapid phosphorylation of Stat3. PE also increased Stat3 phosphorylation, DNA binding and transcription activity in transformed human hepatocellular carcinoma cells (Hep3B). The PE-induced Stat3 phosphorylation, DNA binding and reporter activity were completely blocked by the selective α1-AR antagonist, prazosin. In addition, transfection of Hep3B cells with human α1B-AR expression vector also enhanced Stat3 phosphorylation and reporter activity. Moreover, overexpression of RGS2, a protein inhibitor of Gq/11 signaling, blocked PE-induced Stat3 phosphorylation and reporter activity. The observations that PE induced the formation of c-Src-Stat3 binding complex and phosphorylation of epidermal growth factor receptor (EGFR) and that inhibiting Src and EGFR prevented PE-induced Stat3 activation indicate the involvement of Src and EGFR. Taken together, these observations demonstrate a novel α1-AR-mediated Stat3 activation that involves Gq/11, Src and EGFR in hepatic cells.
alpha1-adrenergic receptor; signal transducer and activator of transcription-3; hepatocyte; epidermal growth factor receptor; phenylephrine
Motivation: In ordinary regression, imposition of a lasso penalty makes continuous model selection straightforward. Lasso penalized regression is particularly advantageous when the number of predictors far exceeds the number of observations.
Method: The present article evaluates the performance of lasso penalized logistic regression in case–control disease gene mapping with a large number of SNPs (single nucleotide polymorphisms) predictors. The strength of the lasso penalty can be tuned to select a predetermined number of the most relevant SNPs and other predictors. For a given value of the tuning constant, the penalized likelihood is quickly maximized by cyclic coordinate ascent. Once the most potent marginal predictors are identified, their two-way and higher order interactions can also be examined by lasso penalized logistic regression.
Results: This strategy is tested on both simulated and real data. Our findings on coeliac disease replicate the previous SNP results and shed light on possible interactions among the SNPs.
Availability: The software discussed is available in Mendel 9.0 at the UCLA Human Genetics web site.
Supplementary information: Supplementary data are available at Bioinformatics online.
Bacterial lipopolysaccharide (LPS; endotoxin) is implicated in the pathogenesis of acute liver failure and several chronic inflammatory liver diseases. To evaluate the effect of hepatocyte cyclooxygenase-2 (COX-2) in LPS-induced liver injury, we generated transgenic mice with targeted expression of COX-2 in the liver by using the albumin promoter-enhancer driven vector and the produced animals were subjected to a standard experimental protocol of LPS-induced acute fulminant hepatic failure (intraperitoneal injection of low dose of LPS in combination with D-galactosamine (GalN)). The COX-2 transgenic mice exhibited earlier mortality, higher serum ALT and AST levels and more prominent liver tissue damage (parenchymal hemorrhage, neutrophilic inflammation, hepatocyte apoptosis and necrosis) than wild type mice. Western blot analysis of the liver tissues showed that LPS/D-GalN treatment for 4 hours induced much higher cleavage of PARP, caspase-3 and caspase-9 in COX-2 transgenic mice than in wild type mice. Increased hepatic expression of JNK2 in COX-2 transgenic mice suggest that upregulation of JNK2 may represent a potential mechanism for COX-2-mediated exacerbation of liver injury. Blocking the prostaglandin receptor, EP1, prevented LPS/D-GalN-induced liver injury and hepatocyte apoptosis in COX-2 transgenic mice. Accordingly, the mice with genetic ablation of EP1 showed less LPS/D-GalN-induced liver damage and less hepatocyte apoptosis with prolonged survival when compared to the wild type mice. These findings demonstrate that COX-2 and its downstream prostaglandin receptor EP1 signaling pathway accelerates LPS-induced liver injury. Therefore, blocking COX-2/EP1 pathway may represent a potential approach for amelioration of LPS-induced liver injury.
Lipopolysaccharide; endotoxin; cyclooxygenase-2; EP1 receptor; liver; acute hepatic failure
Cytosolic phospholipase A2α (cPLA2α) is a rate-limiting key enzyme that releases arachidonic acid (AA) from membrane phospholipid for the production of biologically active lipid mediators including prostaglandins, leukotrienes and platelet-activating factor. cPLA2α is translocated to nuclear envelope in response to intracellular calcium increase and the enzyme is also present inside the cell nucleus; however, the biological function of cPLA2α in the nucleus remains unknown. Here we show a novel role of cPLA2α for activation of peroxisome proliferator-activated receptor-δ (PPARδ) and β-catenin in the nuclei. Overexpression of cPLA2α in human cholangiocarcinoma cells induced the binding of PPARδ to β-catenin and increased their association with the TCF/LEF response element. These effects are inhibited by the cPLA2α siRNA and inhibitors as well as by siRNA knockdown of PPARδ. Overexpression of PPARδ or treatment with the selective PPARδ ligand, GW501516, also increased β-catenin binding to TCF/LEF response element and increased its reporter activity. Addition of AA and GW501516 to nuclear extracts induced a comparable degree of β-catenin binding to TCF/LEF response element. Furthermore, cPLA2α protein is present in the PPARδ and β-catenin binding complex. Thus the close proximity between cPLA2α and PPARδ provides a unique advantage for their efficient functional coupling in the nucleus, where AA produced by cPLA2α comes immediately available for PPARδ binding and subsequent β-catenin activation. These results depict a novel interaction linking cPLA2α, PPARδ and Wnt/β-catenin signaling pathways and provide insight for further understanding the roles of these key molecules in human cells and diseases.
cPLA2; PPAR-δ; arachidonic acid; β-catenin; cholangiocarcinoma
Hepatitis C virus (HCV) infection is often associated with chronic liver disease, which is a major risk factor for the development of hepatocellular carcinoma (HCC). To study the HCV-host cell relationship on the molecular level, HepG2 and Huh7 cells were stably transfected with an infectious cDNA clone of HCV or with empty vector. Evidence for HCV replication was obtained in both culture systems. HCV also stimulated growth in vitro. To identify genes whose altered expression by HCV are important to the pathogenesis of infection, RNAs were isolated from HepG2-HCV and HepG2-vector cells, and subjected to microarray analysis. The results showed that arginase 1 mRNA and protein were elevated about 3-fold in HCV positive compared to negative cells (P < 0.01). Arginase 1 expression was elevated in more than 75% of HCV infected liver samples compared to paired HCC from the same patients (> 33% positive) and to uninfected liver tissues (0% positive). Arginase 1 specific siRNA inhibited the ability of HCV to stimulate hepatocellular growth in culture by > 70%, suggesting that the metabolism of arginine to ornithine may contribute to HCV mediated stimulation of hepatocellular growth. Introduction of arginase specific siRNA also resulted in increased nitric oxide synthase (iNOS) (>1.2 fold), nitric oxide (NO) production (> 3 fold) and increased cell death (>2.5-fold) in HCV positive compared to negative cells, suggesting that these molecules potentially contribute to hepatocellular damage. Hence, an important part of the mechanism whereby HCV regulates hepatocellular growth and survival may be through altering arginine metabolism.
arginine; arginase 1; hepatocellular carcinoma; hepatitis C virus; nitric oxide
Deficient biliary epithelial cell (BEC) expression of small proline rich protein (SPRR) 2A in IL-6-/- mice is associated with defective biliary barrier function after bile duct ligation. And numerous gene array expression studies show SPRR2A to commonly be among the most highly upregulated genes in many non-squamous, stressed and remodeling barrier epithelia. Since the function of SPRR in these circumstances is unknown, we tested the exploratory hypothesis that BEC SPRR2A expression contributes to BEC barrier function and wound repair.
The effect of SPRR2A expression was studied in primary mouse BEC cultures; in BEC cell line after forced over expression of SPRR2A; and in human livers removed at the time of liver transplantation.
Forced SPRR2A over-expression showed that it functions as a SH3 domain ligand that increases resistance to oxidative injury and promotes wound restitution by enhancing migration and acquisition of mesenchymal characteristics. Low confluency non-neoplastic mouse BEC cultures show a phenotype similar to the stable transfectants, as did spindle-shaped BEC participating in atypical ductular reactions in primary biliary cirrhosis.
These observations suggest that SPRR2A-related BEC barrier modifications represent a novel, but widely utilized and evolutionarily conserved, response to stress that is worthy of further study.
Cholangiocytes; Ductular Reaction; Wound Healing; Oxidative Stress; Repair; SH3 domain; EMT
To investigate the adverse effect of type 2 diabetes on coronary artery anatomy.
PATIENTS AND METHODS:
The coronary angiograms of 1564 consecutive patients were analyzed. The coronary lesions of patients with and without type 2 diabetes were compared.
Seventy-four diabetic and 824 nondiabetic patients were found to have coronary artery disease. Diabetic patients had more multivessel (48.7% versus 34.7%, P<0.01), multilesion (64.9% versus 46.1%, P<0.05), extensive (51.4% versus 7.8%, P<0.01) and small vessel disease (95.2% versus 39.8%, P<0.01) than nondiabetic patients.
Type 2 diabetes is a significant risk factor for coronary artery disease. These findings may provide useful information for the future development of therapeutic strategies.
Coronary angiogram; Coronary artery disease; Diabetes mellitus