Protein phosphatase 2A-B55ß mediated mitochondrial p-GPX4 dephosphorylation promoted sorafenib-induced ferroptosis in hepatocellular carcinoma via regulating p53 retrograde signaling.

Rationale: As a key endogenous negative regulator of ferroptosis, glutathione peroxidase 4 (GPX4) can regulate its antioxidant function through multiple post-translational modification pathways. However, the effects of the phosphorylation/dephosphorylation status of GPX4 on the regulation of inducible ferroptosis in hepatocellular carcinoma (HCC) remain unclear. Methods: To investigate the effects and molecular mechanism of GPX4 phosphorylation/dephosphorylation modification on ferroptosis in HCC cells. Sorafenib (Sora) was used to establish the ferroptosis model in HCC cells in vitro. Using the site-directed mutagenesis method, we generated the mimic GPX4 phosphorylation or dephosphorylation HCC cell lines at specific serine sites of GPX4. The effects of GPX4 phosphorylation/dephosphorylation modification on ferroptosis in HCC cells were examined. The interrelationships among GPX4, p53, and protein phosphatase 2A-B55ß subunit (PP2A-B55ß) were also explored. To explore the synergistic anti-tumor effects of PP2A activation on Sora-administered HCC, we established PP2A-B55ß overexpression xenograft tumors in a nude mice model in vivo. Results: In the Sora-induced ferroptosis model of HCC in vitro, decreased levels of cytoplasmic and mitochondrial GPX4, mitochondrial dysfunction, and enhanced p53 retrograde signaling occurred under Sora treatment. Further, we found that mitochondrial p53 retrograded remarkably into the nucleus and aggravated Sora-induced ferroptosis. The phosphorylation status of GPX4 at the serine 2 site (GPX4Ser2) revealed that mitochondrial p-GPX4Ser2 dephosphorylation was positively associated with ferroptosis, and the mechanism might be related to mitochondrial p53 retrograding into the nucleus. In HCC cells overexpressing PP2A-B55ß, it was found that PP2A-B55ß directly interacted with mitochondrial GPX4 and promoted Sora-induced ferroptosis in HCC. Further, PP2A-B55ß reduced the interaction between mitochondrial GPX4 and p53, leading to mitochondrial p53 retrograding into the nucleus. Moreover, it was confirmed that PP2A-B55ß enhanced the ferroptosis-mediated tumor growth inhibition and mitochondrial p53 retrograde signaling in the Sora-treated HCC xenograft tumors. Conclusion: Our data uncovered that the PP2A-B55ß/p-GPX4Ser2/p53 axis was a novel regulatory pathway of Sora-induced ferroptosis. Mitochondrial p-GPX4Ser2 dephosphorylation triggered ferroptosis via inducing mitochondrial p53 retrograding into the nucleus, and PP2A-B55ß was an upstream signal modulator responsible for mitochondrial p-GPX4Ser2 dephosphorylation. Our findings might serve as a potential theranostic strategy to enhance the efficacy of Sora in HCC treatment through the targeted intervention of p-GPX4 dephosphorylation via PP2A-B55ß activation.

Aflatoxin B1 exposure triggers hepatic lipotoxicity via p53 and perilipin 2 interaction-mediated mitochondria-lipid droplet contacts: An in vitro and in vivo assessment.

Aflatoxin B1 (AFB1) is one of the most toxic mycotoxins widely found in food contaminants, and its target organ is the liver. It poses a major food security and public health threat worldwide. However, the lipotoxicity mechanism of AFB1 exposure-induced liver injury remains unclear and requires further elucidation. Herein, we investigated the potential hepatic lipotoxicity of AFB1 exposure using in vitro and in vivo models to assess the public health hazards of high dietary AFB1 exposure. We demonstrated that low-dose of AFB1 (1.25 µM for 48 h, about one-fifth of the IC50 in HepG2 and HepaRG cells, IC50 are 5.995 µM and 5.266 µM, respectively) exposure significantly induced hepatic lipotoxicity, including abnormal lipid droplets (LDs) growth, mitochondria-LDs contacts increase, lipophagy disruption, and lipid accumulation. Mechanistically, we showed that AFB1 exposure promoted the mitochondrial p53 (mito-p53) and LDs-associated protein perilipin 2 (PLIN2) interaction-mediated mitochondria-LDs contacts, resulting in lipid accumulation in hepatocytes. Mito-p53-targeted inhibition, knockdown of PLIN2, and rapamycin application efficiently promoted the lysosome-dependent lipophagy and alleviated the hepatic lipotoxicity and liver injury induced by AFB1 exposure. Overall, our study found that mito-p53 and PLIN2 interaction mediates three organelles-mitochondria, LDs, and lysosomal networks to regulate lipid homeostasis in AFB1 exposure-induced hepatotoxicity, revealing how this unique trio of organelles works together and provides a novel insight into the targeted intervention in inter-organelle lipid sensing and trafficking for alleviating hazardous materials-induced hepatic lipotoxicity.

Rab7a-mTORC1 signaling-mediated cholesterol trafficking from the lysosome to mitochondria ameliorates hepatic lipotoxicity induced by aflatoxin B1 exposure.

Aflatoxin B1 (AFB1) is a common contaminant in many foodstuffs and is considered a public health concern worldwide due to its hepatotoxicity caused by lipid metabolism disorders. However, the molecular mechanism underlying AFB1-induced lipotoxicity-dependent liver injury via regulating cholesterol metabolism remains unclear. We established a cholesterol trafficking disorder-mediated hepatic lipotoxicity model with AFB1 mixture exposure in vitro (HepaRG and HepG2 cells, 1.6 µM for 36 h) and in vivo (C57BL/6 mice, 3 mg kg-1, i.g., every other day for 6 weeks). In vitro, the interaction between lysosomal Niemann-Pick type C1 (NPC1) protein and mitochondrial translocator protein (TSPO) regulated lipotoxicity induced by AFB1 mixture exposure, including lysosomal membrane permeabilization and mitochondria-dependent necroptosis. Moreover, the downregulation of lysosomal Ras-associated protein 7a (Rab7a) enhanced the mammalian target of rapamycin complex 1 (mTORC1)-mediated disorders of cholesterol trafficking from the lysosome to mitochondria. Furthermore, cholesterol trafficking disorder-mediated hepatic lipotoxicity induced by the low-dose level of AFB1 exposure was relieved by genetic or pharmaceutic activation of Rab7a to inhibit mTORC1 in vitro and ex vivo. In vivo, mTORC1 inhibitor (Torin1, 4 mg kg-1, i.p., every other day for 3 weeks) alleviated the cholesterol trafficking disorder-mediated hepatic lipotoxicity via upregulating the molecular machinery of lysosomes and mitochondria contact mediated by NPC1 and TSPO interaction in the low dose of AFB1 exposure. Altogether, our data suggested a novel mechanism that lysosomal Rab7a-mTORC1 signaling determined the cholesterol trafficking regulated by NPC1-TSPO from the lysosome to mitochondria, which promoted hepatic lipotoxicity via lysosomal quality control and mitochondria-dependent necroptosis signaling pathways in chemical mixture exposure.

Effect of prenatal PFOS exposure on liver cell function in neonatal mice.

Perfluorooctane sulfonate (PFOS), a hepatotoxic pollutant, is detected in the human cord blood, and it may induce health risk to an embryo. In this study, we established intrauterine exposure to PFOS in mice to evaluate potential impacts of PFOS on postnatal day 1 (PND1) offspring through conducting biochemical tests, quantitative PCR, and immunostaining. As results, PFOS-exposed maternal mice showed marked hepatomegaly and induced liver steatosis in a high dose of 5 mg PFOS/kg. In PND1 mice, intrahepatic contents of triglyceride, total cholesterol, and LDL were elevated by high-dose PFOS exposure, while intracellular HDL content was decreased. As shown in quantitative PCR, functional messenger RNAs of cytochrome P4A14 (CYP4A14) for fatty acid oxidation, CD36 for hepatic fatty acid uptake, and apolipoprotein B100 (APOB) and fibroblast growth factor 21 (FGF21) for hepatic export of lipids in PND1 livers were changed when compared to those in PFOS-free controls. In further validations, immunofluorescence stains showed that hepatic CYP4A14 and CD36 immunoreactive cells were increased in PFOS-exposed PND1 mice. In addition, reduced immunofluorescence-positive cells of APOB and FGF21 were observed in PND1 livers. Collectively, these preliminary findings demonstrate that prenatal exposure to PFOS may affect lipid metabolism in liver cells of PND1 mice.

Immunophenotypes of Ductal Epithelial Cells in Advanced Pancreatic Ductal Adenocarcinoma.

BACKGROUND/AIMS: In this report, we aimed to investigate the distribution and characterization of biomarker-immunolabeled ductal epithelium in advanced pancreatic ductal adenocarcinoma (PDAC). DESIGN: Eighteen patients with PDAC were medically diagnosed prior to being treated periodically. All clinical records were collected and assayed. The PDAC samples were subjected to routine biochemical tests and immuno-stains of immunohistochemistry and immunofluorescence. RESULTS: As results, immunohistochemical findings indicated pancreatic ductal cells in PDAC showed plenty of Ki-67, P53, RP, SAM positive cells expressed in nuclei. In addition, ductal epithelial cells exhibited numerous positive cells of CK7, 18, 19, 20 biomarkers, respectively. Interestingly, compared to non-PDAC controls, immunostaining results showed that endocrine hormones were positively expressed in pancreatic ducts of PDAC, characterized with increased insulin-, Ngn3-, PDX1-fluorescence-labeled cells, and reduced F-box and WD-40 domain protein 7 (Fbxw7)-labeled cells in ducts. CONCLUSION: These clinicopathologic findings preliminarily disclose that there may be a potential for insulin-producing cells in PDAC, possibly through negatively inducing Fbxw7 ubiquitination in pancreatic ducts.

Antihepatotoxic benefits of Poria cocos polysaccharides on acetaminophen-lesioned livers in vivo and in vitro.

In our previous study, preliminary data indicates that Poria cocos polysaccharides (PCP) shows beneficial hepatoprotection against acetaminophen (APAP)-induced liver injury in mice. However, biological molecular mechanism warrants to be further discussed. In current study, a number of biochemical tests and immunoassays were subjected to respective PCP-dosed mice in vivo and liver cells in vitro. As a result, PCP-treated mice showed reduced contents of inflammatory cytokines (tumor necrosis factor [TNF]-ß and TNFsR-I), enzymological molecules (alanine aminotransferase, aspartate aminotransferase, and lactate dehydrogenase [LDL]), and heat shock protein 90 (Hsp90) after APAP exposure. Additionally, immunostaining assays exhibited that lowered-positive cells of cleaved-caspase-3, cleaved-poly ADP ribose polymerase, and Hsp90-labeled cells in PCP-treated livers were observed, and increased cluster of differentiation 29 (CD29), CD73-positive cells in the spleen were detected. Further, PCP-treated mouse liver cells resulted in increased cell growth, reduced LDL level. Increased proliferating cell nuclear antigen (PCNA), P38 mitogen-activated protein kinase (MAPK)-labeled cells and decreased Hsp90-positive cells in APAP-exposed liver cells were observed dose-dependently after PCP cotreatments. Collectively, our present experimental findings elucidate that PCP beneficially play hepatoprotective effects against APAP-lesioned liver cells in vivo and in vitro, potentially through the molecular mechanisms of suppressing cell death, reducing hepatocellular inflammatory stress and Hsp90 bioactivity.

Hepatoprotective benefits of vitamin C against perfluorooctane sulfonate-induced liver damage in mice through suppressing inflammatory reaction and ER stress.

Our previous studies show that vitamin C (VC) plays promising hepatoprotection in mice. Intrahepatic exposure of perfluorooctane sulfonate (PFOS) can induce dose-dependent cytotoxicity. However, pharmacology-based assessment of VC on PFOS remains uninvestigated. This study aimed to evaluate the therapeutic benefits of VC on inhibiting PFOS-induced liver steatosis in mice, followed by representative biochemical analysis and immunoassay. As results, VC was beneficial for reduced PFOS-induced liver damages, as showed in reductions of serological levels of transaminases (ALT and AST), lipids (TG and TC), fasting glucose and insulin, inflammatory cytokines (TNF-α and IL6), while content of fibroblast growth factor 21 (FGF21) in serum was increased. In addition, VC reduced histiocytic changes of PFOS-lesioned livers, as revealed in reduced TNF-α-labeled cells and increased FGF21-labeled cells in immunofluorescence assay. Further, intrahepatic expressions of endoplasmic reticulum (ER) stress-based ATF6, eIF2α, GRP78, XBP1 proteins were down-regulated by treatments of VC. Taken together, our preliminary findings set forth that VC exerts pharmacological benefits against PFOS-induced liver steatosis in mice, and the underlying biological mechanism may be linked to suppressing hepatocellular inflammatory reaction and ER stress.

Prenatal exposure to perfluorooctanoic acid induces nerve growth factor expression in cerebral cortex cells of mouse offspring.

Previous studies have showed perfluorooctanoic acid (PFOA) inducing cytotoxicity in an organ. In addition, epidemiological data show that high level of PFOA in cord blood of a pregnant woman is detected. Therefore, we extrapolate that circulating PFOA may affect organogenesis in offspring, such as the brain. In this study, intrauterine exposure to PFOA in mice was used to characterize the potential impacts of prenatal PFOA exposure on cerebral cortex cells of postnatal 21 (PND21) offspring. In an ex vivo cell model, PND21-based cortex cells were exposed to PFOA or/and nerve growth factor (NGF)-specific inhibitor before further biochemical assays. As results, biochemical data showed increased trends of liver metabolic enzymes in sera of PFOA-treated PND21 mice. Interestingly, PFOA-treated PND21 mice resulted in increased levels of NGF in sera and cortex cells. In addition, PFOA-exposed cerebral cortex cells induced NGF and proliferating cell nuclear antigen (PCNA) expressions, while exposure to PFOA/NGF-specific inhibitor downregulated expressions of NGF and PCNA. In addition, Nissl-labeled, NGF-positive cells, and NGF protein expression in cortex cells of PFOA-treated PND21 mice were upregulated, respectively. Further, immunoblotting assays showed that intracephalic poly (ADP-ribose) polymerase (PARP) and p42/44 mitogen-activated protein kinase (MAPK) proliferation-regulated protein levels were elevated in PFOA-treated cortex cells. Taken together, our current findings indicate that the prenatal PFOA exposure may induce proliferation of cerebral cortex cells in PND21 mice through promoting intracephalic NGF expression in the cortex.

Hepatoprotective effects exerted by Poria Cocos polysaccharides against acetaminophen-induced liver injury in mice.

Our study was to investigate the potential pharmacological activity of Poria Cocos polysaccharides (PCP) against acetaminophen (APAP)-induced liver injury in mice. PCP-dosed mice were used to conducting biochemical assays of serological liver enzyme (ALT), lactate dehydrogenase (LD), inflammatory cytokines (TNF-α, IL-6), and immunoassays for functional proteins in the livers. Consequently, APAP-exposed mice resulted in elevated levels of ALT, LD, TNF-α, IL-6 in sera. Interestingly, PCP-dosed mice exhibited reduced ALT, LD and inflammatory cytokines in blood. Inflammatory infiltration and cell death in liver tissue were decreased following by PCP treatments. Furthermore, immunofluorescence staining showed that AKR7A, c-Jun, Bcl-2-positive cells were increased in PCP-dosed livers in mice, while Bax-labeled cells were decreased. In addition, hepatocellular down-regulated NF-κBp65, IkBα expressions were observed dose-dependently in PCP-dosed livers in mice. Taken together, the current findings indicate that Poria Cocos polysaccharides exert pharmacological bioeffects against APAP-induced liver injury in mice, and the underlying molecular mechanism is associated to suppressing inflammatory response and apoptosis in liver cells.

Adverse bioeffect of perfluorooctanoic acid on liver metabolic function in mice.

Perfluorooctanoic acid (PFOA), a kind of manufactured material, is widely accumulated around environmental system and into wildlife, including human beings. Toxicologically, PFOA induces hepatomegaly (liver enlargement) in the dose- and time-dependent manners. However, biological mechanism of hepatotoxicity warrants to be further investigated. In the present study, mature male mice were exposed to dosed PFOA for 21 days before conducting biochemical tests and immunoassays. As results, decreased fast blood glucose and elevated insulin contents were observed in PFOA-dosed mice. In addition, PFOA-dosed mice resulted in increased liver functional enzymes (GPT, GOT) in serum. And lipid contents (TG, lipoproteins) in serum and liver were changed abnormally. As shown in immunohistochemistry, increased insulin- and poly (ADP-ribose) polymerase (PARP)-positive cells were showed in PFOA-exposed pancreatic tissues. Moreover, CD36-positive cells were increased in PFOA-exposed livers, while ApoB-labeled cells were reduced. Further, immunoblot data showed that hepatocellular fibroblast growth factor 21 (FGF21) in PFOA-exposed liver was down-regulated dose-dependently. Taken together, our preliminary findings demonstrated that PFOA-induced hepatocellular lipotoxicity may be linked to impairing lipid-regulated proteins, as well as inducing insulin expression from pancreatic tissue.

Cytoprotective effects of glycyrrhetinic acid liposome against cyclophosphamide-induced cystitis through inhibiting inflammatory stress.

This study was designed to investigate the pharmacological efficacy of glycyrrhetinic acid liposome (GAL) against female mice with nonbacterial cystitis induced by cyclophosphamide (CPS). Mice in different groups were subjected to tests for lactate dehydrogenase (LD), cytokine contents (IL-6, TNF-α) in serum, and histological changes in bladder tissue and to immunoassays. As a result, cyclophosphamide-induced cystitis in mice showed an increased LD level in serum, and the contents of cytokines (IL-6, TNF-α) were elevated. Interestingly, GAL-treated mice showed decreased LD and inflammatory cytokines of IL-6 and TNF-α in blood. Inflammatory infiltration and cell death in bladder tissue were reduced by GAL treatments. In addition, intravesical mRNAs of NF-κB and TNF-α were lowered dose-dependently in GAL-treated mice. As shown in cytohistological staining, the number of intravesical caspase-3, PARP-positive cells decreased in GAL-treated mice. Furthermore, a GAL-treated bladder showed down-regulated NF-κB and TNF-α expressions in a dose-dependent manner. In conclusion, our current findings may be the first to provide scientific evidence demonstrating that glycyrrhetinic acid liposomes provide benefits against cyclophosphamide-induced cystitis, which possibly occurs through underlying mechanisms that inhibit cell death and inflammatory stress.

Biocharacterization of Heat Shock Protein 90 in Acetaminophen-Treated Livers Without Conspicuous Drug Induced Liver Injury.

BACKGROUND/AIMS: Acetaminophen (APAP) refers to a medication used to manage pain and fever symptoms. Heat shock protein 90 (Hsp90) is to be expressed during various stresses, such as wound healing and tissue remodeling. Recently, it is discovered that Hsp90 is a potential modifier of cytogenesis. In comparison to clinical references of liver damage, this study was designed to assess the potential bioeffect of Hsp90 in APAP-treated livers without conspicuous drug induced liver injury (DILI). METHODS: In our current study, human plasma samples of APAP-used patients were collected for biochemical assays in clinical parameters. Adult male mice were used to investigate the biocharacterization of Hsp90 in APAP-treated livers through serological tests and immunoassays. Further, a mouse liver cell strain was employed in assessment of bioeffect of APAP on hepatocellular Hsp90 expression. RESULTS: Correspondingly, the clinical data showed APAP-administered patients resulted in increased Hsp90 levels in serum when compared to other clinical parameters of liver injury. In adult mice study, APAP-treated livers showed unchanged hepatocellular and metabolic functions, as highlighted in biochemical analysis and immunoassay. Notably, Hsp90 expression in APAP-treated mice were elevated in the serum and liver samples. In quantitative western blot assay, the present data suggested that hepatocellular Hsp90 level was up-regulated followed by APAP treatment. In mouse cell strain study, APAP-treated liver cells had increased trend of aminotransferase contents and apoptotic counts. Further, endogenous Hsp90 expression in APAP-exposed cells was increased dose- and time-dependently. CONCLUSIONS: In conclusion, our current findings disclose that Hsp90 biomolecule may be a potential indicator for APAP-induced inconspicuous DILI, in which it seems to be characterized with more sensitive than other diagnosis criteria.

Effect of acute exposure to PFOA on mouse liver cells in vivo and in vitro.

Increasingly, epidemiological evidences indicate chemosynthetic perfluorooctanoic acid (PFOA), an environmental pollutant, induces potential adverse effect on human health after long-term exposure. However, less study has been performed for assessment of acute effect of PFOA exposure on metabolic homeostasis. In experimental designs, PFOA-exposed liver cells in vivo and in vitro were used to discuss underlying mechanism related to PFOA-induced metabolic dysfunction. In serological tests, PFOA-exposed mice showed increased treads of liver functional enzymes in alanine transaminase (ALT), aspartate transaminase (AST), and total bilirubin (T-BIL), trypsinase, low density lipoprotein-cholesterol (LDL-C), and insulin, while blood glucose, high density lipoprotein-cholesterol (HDL-C), and glucagon levels were reduced. In histocytological observations, PFOA-exposed liver showed visible cytoplasmic vesicles, and intact pancreatic islets were observed in PFOA-exposed pancreas. Additionally, increased insulin-positive cells and reduced glucagon-positive cells were detected in PFOA-exposed islets. As shown in immunoassays, PFOA-exposed liver resulted in elevations of cluster of differentiation 36 (CD36)-labeled cells and CD36 protein. In mouse liver cell study, PFOA-exposed cells showed increased cell apoptotic count, and increased phosphorylated levels of Bcl-2 and Bad in the cells. Furthermore, PFOA-exposed liver cells exhibited elevations of CD36-labeled cells and CD36 protein. Taken together, the present data demonstrate that acute exposure to PFOA-impaired liver function is associated with inducting CD36 expression and apoptosis, as well as disrupting key hormones in the pancreas.

FGF21 functions as a sensitive biomarker of APAP-treated patients and mice.

Acetaminophen (APAP) is a common medication that induces hepatocellular damage in a time- or dose-dependent manner. Fibroblast growth factor 21 (FGF21) exerts a series of biological effects, including cellular repair. Compared to clinical diagnosis parameters, we aimed to evaluate whether FGF21 can serve as a sensitive biomarker for APAP-induced liver impairment. In the present study, we discussed comparable data from APAP-treated patients and parallelly established APAP-exposed mice for investigation. The resulting human serological data showed that APAP-treated patients have a visible reduction of FGF21 expression in undetected liver impairment of clinical diagnosis. In the animal study, APAP-exposed livers exhibited normal metabolic functions and liver functions, as revealed by biochemical test and histopathological examination. Endogenous FGF21 concentrations in APAP-treated mice were decreased in sera and liver cells. Moreover, comparable immunoassay data showed that hepatocellular FGF21 expression was reduced in a time-dependent manner. Taken together, these findings elucidate the involvement of abnormal FGF21 expression in early APAP-induced liver impairment. Interestingly, FGF21 may be a promising biomarker of APAP-exposed livers.

Valproate acid (VPA)-induced dysmetabolic function in clinical and animal studies.

BACKGROUND: Valproate acid (VPA), a commonly used antiepileptic medicine, is found to be linked to developing dysmetabolic risk. However, the proposed mechanism remains completely unknown. METHODS: In this study, we collected data from patients with epilepsy and further investigated the preclinical study in mice. RESULTS: As results, the clinical data showed that VPA-used patients resulted in higher levels of blood glucose, urine acid, triglyceride (TG), and immune cells (leucocyte, neutrophil leucocyte) when compared to clinically diagnosed references, while circulating apolipoprotein A1 (Apob A1) and fatty acid binding protein 4 (FABP4) were reduced without visible drug-induced liver injury (DILI). In rodent study, short-term treatment of VPA to mice showed developing trend of metabolic dysfunction, as revealed in increased serum insulin and free fatty acid and TG, and decreased liver TG content. As shown in immunoassay, FABP4 of epididymal white adipose tissue (eWAT) was down-regulated dose-dependently. CONCLUSION: Collectively, our present findings indicate that VPA can induce FABP4-impaired lipid dysmetabolism without DILI. More notably, FABP4 may function as a sensitive indicator for VPA-induced metabolic dysfunction.

Clinical biocharacterization of immunophenotype in hepatocellular carcinoma patients.

In current observations, we investigated the clinical immunophenotypes in liver cells to assess the metastasic predisposition in advance hepatocellular carcinoma patients. In method, we harvested the clinically diagnosed data from 8 liver cancer subjects. Definitely, all patients were received standard chemotherapeutics when being confirmed as advanced liver cancer via clinical diagnosis. In parallel, biopsy liver samples were subjected to histopathological and immunoblotting assays. Representatively, clinical laboratory results showed that blood parameters resulted in notable elevations of aminotransferases (ALT, AST), hepatitis B e antibody (HBeAb), alpha-fetoprotein (APF) and carcinoembryonic antigen (CEA). In addition, immunoassays exhibited significant hepatocellular expressions of Ki-67 (cell proliferation), CD34 (angiogenesis), as well as strong production of CK8, CK10 (metastasis) in liver cells, as revealed in both the immunostaining and Western blotting analyses. Collectively, the present clinical findings elucidate that the metastasis of hepatocellular carcinoma relates to unregulated cell proliferation and angiogenesis. In particular, current representative immunophenotypes may be served as potential biomarkers for screening metastasis of advanced liver cancer.

MicroRNA-421 mediates immunosensitivity in late-stage human liver cancer.

This report showed the novel clinical evidence to evaluate the pathological predisposition of microRNA-421 (miR-421) on immuno-insufficiency to cancer progression of human hepatocellular carcinoma. Some hospitalized patients with liver cancer were recruited when they were diagnosed by clinical parameters. As expected, all patients were subjected to routine biochemical analysis and histocytological inspection prior to receiving medical treatment. Clinical findings suggested that basal parameters of plasmatic aminotransferases and hepatitis B e antibody (HBeAb), alpha-fetoprotein (AFP) were elevated significantly. Further, immune cells and T lymphocytes in peripheral blood were abnormally altered in a time-dependent manner. More specifically, endogenous expressions of miR-421 level in liver cancer, immune-specific glycoprotein CD4 and CD8 were increased when compared to nontumor control. Furthermore, intracellular immunocytochemical staining showed that proliferative marker of Ki-67, metastasized marker of CK19, as well as phenotypic markers of hepatocellular carcinoma for HBeAb and AFP were positively expressions with disseminated distribution. To sum up, these biochemical data and histopathological evidences demonstrate that miR-421-mediated immune hypersensitivity may function as a promising indicator for advanced liver cancer, even in further metastasis. Therefore, inhibition of intracellular miR-421 expression is an effective strategy for management of terminal hepatocellular carcinoma.

Characterization of acetaminophen-induced cytotoxicity in target tissues.

Acetaminophen (APAP), commonly used in clinical prescription, has time- and dose-dependent side effects. Thus, further animal study warrants to be investigated to assess possible adverse effect of APAP application. Here, we conducted pre-clinical research to elucidate the molecular mechanism regarding APAP-mediated toxicological action. Our data showed that serous/urinary and hepatic/renal APAP concentrations were significantly increased when compared with normal control, which the liver tissue showed the highest level. As an acute liver damage model induced by APAP, absolute liver weight, serum enzyme (ALT), urine protein content were notably elevated. Representatively, APAP-damaged liver resulted in increased pro-apoptotic Bax and compensatory Ki-67 positive cells, while the number of anti-apoptotic Bcl2 positive cells was reduced. In addition, the immunoactivity markers for NF-κB, TRL4, TNF-α in the kidney were increased, respectively. Furthermore, intracellular TRL4 and TNF-α mRNAs in the liver and kidney showed significant up-regulation. In summary, our current findings demonstrate that APAP-mediated the specific cytotoxicity is linked to the molecular mechanisms of facilitating apoptosis and inflammatory stress in the liver and kidney.

Puerarin exerts the protective effect against chemical induced dysmetabolism in rats.

In this study, we aim to explore the potential benefits of puerarin on metabolic function of liver fibrosis (LF) rat induced by carbon tetrachloride (CCl4), and to investigate with the underlying molecular mechanism targeted on liver and pancreas tissues. In methodology, The LF rats were prepared through intragastrically giving CCl4 twice each week (2ml/kg, v/w) for 8weeks, and dosed puerarin (20, 40mg/kg) were given three times each week via intraperitoneal injection. After being conducted with oral glucose tolerance test (OGTT), the blood samples of rat were harvested for biochemical tests, as well as the liver and pancreas were isolated for histological examination and biochemical assays. The findings showed that puerarin-administered rats resulted in reduced glucose tolerance, blood insulin level, sero-enzymes of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), and increased plasma level of high-density lipoprotein cholesterol (HDL-C) and reduced low-density lipoprotein cholesterol (LDL-C) content in serum. Further, the intrahepatic collagen deposits were lessened and positive cell of alpha smooth muscle actin (α-SMA) was lessened in puerarin treatment, while the pro-apoptotic cell numbers of Caspase 3, Bax in pancreatic islets were reduced dose-dependently. Moreover, the mRNA expressions of toll-like receptor 4 (TLR4) and tumor necrosis factor (TNF-α) in liver and pancreas were downregulated. In addition, TNF-α protein in the CCl4-lesioned liver and pancreas was reduced. Our findings demonstrate that puerarin contributes to attenuating the metabolic dysfunctions of CCl4-damaged liver and pancreas, in which the possible mechanisms may be linked to inhibition of inflammatory stress and normalization of metabolic homoeostasis in the liver and pancreas.

Impact of perinatal exposure to acetaminophen on hepatocellular metabolic function in offspring.

Acetaminophen (APAP), an over the counter (OTC) medication, is widely used in antipyretic treatment. Although the risk of dose-dependent cytotoxicity has been known, the potential effect of perinatal exposure to acetaminophen on metabolic function in offspring remains uninvestigated. Therefore, we established a prenatally APAP-exposed pregnancy mouse model to assess the possible adverse effect on liver metabolic function in offspring. Biochemical assays were applied in analysis of basic metabolic parameters in postnatal mice. Further, immunoblotting assay was used to assess the expressions of insulin receptor ß (IRß), insulin receptor substrate 1 (IRS1), phospho-Akt and phospho-GSK-3ß proteins in liver cells. In addition, hepatic glucose transporter 2 (GLUT2) immunoactivity was determined by using immunohistochemistry staining. Compared with untreated postnatal mice, APAP-exposed offspring induced impaired glucose metabolism, increased plasma insulin level, and reduced liver glycogen content. In addition, APAP exposure decreased the expressions of IRS1 and phospho-GSK-3ß, phospho-AKT proteins and down-regulated the level of glucose-import regulator GLUT2 in the liver. Taken together, our preliminary findings indicate that perinatal APAP exposure-impaired hepatic glucose metabolism in offspring may be associated with disturbance of insulin-dependent AKT signaling in the liver.