Hepatocyte growth factor promotes retinal pigment epithelium cell activity through MET/AKT signaling pathway

AIM:To explore the effects of hepatocyte growth factor(HGF)on retinal pigment epithelium(RPE)cell behaviors.METHODS:The human adult retinal pigment epithelial cell line-19(ARPE-19)were treated by HGF or mesenchymalepithelial transition factor(MET)inhibitor SU11274 in vitro.Cell viability was detected by a Cell Counting Kit-8 assay.Cell proliferation and motility was detected by a bromodeoxyuridine incorporation assay and a wound healing assay,respectively.The expression levels of MET,phosphorylated MET,protein kinase B(AKT),and phosphorylated AKT proteins were determined by Western blot assay.The MET and phosphorylated MET proteins were also determined by immunofluorescence assay.RESULTS:HGF increased ARPE-19 cells’viability,proliferation and migration,and induced an increase of phosphorylated MET and phosphorylated AKT proteins.SU11274 significantly reduced cell viability,proliferation,and migration and decreased the expression of MET and AKT proteins.SU11274 suppressed HGF-induced increase of viability,proliferation,and migration in ARPE-19 cells.Additionally,SU11274 also blocked HGF-induced phosphorylation of MET and AKT proteins.CONCLUSION:HGF enhances cellular viability,proliferation,and migration in RPE cells through the MET/AKT signaling pathway,whereas this enhancement is suppressed by the MET inhibitor SU11274.HGF-induced MET/AKT signaling might be a vital contributor of RPE cells survival.

Research progress on the development of hepatocyte growth factor/c-Met signaling pathway in gastric cancer:A review

Hepatocyte growth factor(HGF)and its receptor,c-Met,play important roles in the occurrence,development,and treatment of gastric cancer(GC).This review explored the function of the HGF/c-Met signaling pathway in GC and its potential targeted therapeutic mechanisms.As one of the most common malignant tumors worldwide,GC has a complex pathogenesis and limited therapeutic options.Therefore,a thorough understanding of the molecular mechanism of GC is very important for the development of new therapeutic methods.The HGF/c-Met signaling pathway plays an important role in the proliferation,migration,and invasion of GC cells and has become a new therapeutic target.This review summarizes the current research progress on the role of HGF/c-Met in GC and discusses targeted therapeutic strategies targeting this signaling pathway,providing new ideas and directions for the treatment of GC.

Hepatocyte growth factor enhances the ability of dental pulp stem cells to ameliorate atherosclerosis in apolipoprotein E-knockout mice

BACKGROUND Atherosclerosis(AS),a chronic inflammatory disease of blood vessels,is a major contributor to cardiovascular disease.Dental pulp stem cells(DPSCs)are capable of exerting immunomodulatory and anti-inflammatory effects by secreting cytokines and exosomes and are widely used to treat autoimmune and inflam-mation-related diseases.Hepatocyte growth factor(HGF)is a pleiotropic cytokine that plays a key role in many inflammatory and autoimmune diseases.AIM To modify DPSCs with HGF(DPSC-HGF)and evaluate the therapeutic effect of DPSC-HGF on AS using an apolipoprotein E-knockout(ApoE-/-)mouse model and an in vitro cellular model.METHODS ApoE-/-mice were fed with a high-fat diet(HFD)for 12 wk and injected with DPSC-HGF or Ad-Null modified DPSCs(DPSC-Null)through tail vein at weeks 4,7,and 11,respectively,and the therapeutic efficacy and mechanisms were analyzed by histopathology,flow cytometry,lipid and glucose measurements,real-time reverse transcription polymerase chain reaction(RT-PCR),and enzyme-linked immunosorbent assay at the different time points of the experiment.An in vitro inflammatory cell model was established by using RAW264.7 cells and human aortic endothelial cells(HAOECs),and indirect co-cultured with supernatant of DPSC-Null(DPSC-Null-CM)or DPSC-HGF-CM,and the effect and mechanisms were analyzed by flow cytometry,RT-PCR and western blot.Nuclear factor-κB(NF-κB)activators and inhibitors were also used to validate the related signaling pathways.RESULTS DPSC-Null and DPSC-HGF treatments decreased the area of atherosclerotic plaques and reduced the expression of inflammatory factors,and the percentage of macrophages in the aorta,and DPSC-HGF treatment had more pronounced effects.DPSCs treatment had no effect on serum lipoprotein levels.The FACS results showed that DPSCs treatment reduced the percentages of monocytes,neutrophils,and M1 macrophages in the peripheral blood and spleen.DPSC-Null-CM and DPSC-HGF-CM reduced adhesion molecule expression in tumor necrosis factor-αstimulated HAOECs and regulated M1 polarization and inflammatory factor expression in lipopolysaccharide-induced RAW264.7 cells by inhibiting the NF-κB signaling pathway.CONCLUSION This study suggested that DPSC-HGF could more effectively ameliorate AS in ApoE-/-mice on a HFD,and could be of greater value in stem cell-based treatments for AS.

Erythropoietin-induced hepatocyte receptor A2 regulates effect of pyroptosis on gastrointestinal colorectal cancer occurrence and metastasis resistance

Erythropoietin-induced hepatocyte receptor A2(EphA2)is a receptor tyrosine kinase that plays a key role in the development and progression of a variety of tumors.This article reviews the expression of EphA2 in gastrointestinal(GI)colorectal cancer(CRC)and its regulation of pyroptosis.Pyroptosis is a form of programmed cell death that plays an important role in tumor suppression.Studies have shown that EphA2 regulates pyrodeath through various signaling pathways,affecting the occurrence,development and metastasis of GI CRC.The overexpression of EphA2 is closely related to the aggressiveness and metastasis of GI CRC,and the inhibition of EphA2 can induce pyrodeath and improve the sensitivity of cancer cells to treatment.In addition,EphA2 regulates intercellular communication and the microenvironment through interactions with other cytokines and receptors,further influencing cancer progression.The role of EphA2 in GI CRC and its underlying mechanisms provide us with new perspectives and potential therapeutic targets,which have important implications for future cancer treatment.

Microvesicles derived from mesenchymal stem cells inhibit acute respiratory distress syndrome-related pulmonary fibrosis in mouse partly through hepatocyte growth factor

BACKGROUND Pulmonary fibrosis is one of the main reasons for the high mortality rate among acute respiratory distress syndrome(ARDS)patients.Mesenchymal stromal cell-derived microvesicles(MSC-MVs)have been shown to exert antifibrotic effects in lung diseases.AIM To investigate the effects and mechanisms of MSC-MVs on pulmonary fibrosis in ARDS mouse models.METHODS MSC-MVs with low hepatocyte growth factor(HGF)expression(siHGF-MSC-MVs)were obtained via lentivirus transfection and used to establish the ARDS pulmonary fibrosis mouse model.Following intubation,respiratory mechanics-related indicators were measured via an experimental small animal lung function tester.Homing of MSC-MVs in lung tissues was investigated by near-infrared live imaging.Immunohistochemical,western blotting,ELISA and other methods were used to detect expression of pulmonary fibrosis-related proteins and to compare effects on pulmonary fibrosis and fibrosis-related indicators.RESULTS The MSC-MVs gradually migrated and homed to damaged lung tissues in the ARDS model mice.Treatment with MSC-MVs significantly reduced lung injury and pulmonary fibrosis scores.However,low expression of HGF(siHGF-MSC-MVs)significantly inhibited the effects of MSC-MVs(P<0.05).Compared with the ARDS pulmonary fibrosis group,the MSC-MVs group exhibited suppressed expression of type I collagen antigen,type III collagen antigen,and the proteins transforming growth factor-βandα-smooth muscle actin,whereas the siHGF-MVs group exhibited significantly increased expression of these proteins.In addition,pulmonary compliance and the pressure of oxygen/oxygen inhalation ratio were significantly lower in the MSC-MVs group,and the effects of the MSC-MVs were significantly inhibited by low HGF expression(all P<0.05).CONCLUSION MSC-MVs improved lung ventilation functions and inhibited pulmonary fibrosis in ARDS mice partly via HGF mRNA transfer.

Vitamin D 1,25-Dihydroxyvitamin D_(3) reduces lipid accumulation in hepatocytes by inhibiting M1 macrophage polarization

BACKGROUND Non-alcoholic fatty liver disease(NAFLD),which is a significant liver condition associated with metabolic syndrome,is the leading cause of liver diseases globally and its prevalence is on the rise in most nations.The protective impact of vitamin D on NAFLD and its specific mechanism remains unclear.AIM To examine the role of vitamin D in NAFLD and how vitamin D affects the polarization of hepatic macrophages in NAFLD through the vitamin D receptor(VDR)-peroxisome proliferator activated receptor(PPAR)γpathway.METHODS Wild-type C57BL/6 mice were provided with a high-fat diet to trigger NAFLD model and administered 1,25-dihydroxy-vitamin D[1,25(OH)_(2)D_(3)]supplementation.1,25(OH)_(2)D_(3) was given to RAW264.7 macrophages that had been treated with lipid,and a co-culture with AML12 hepatocytes was set up.Lipid accumulation,lipid metabolism enzymes,M1/M2 phenotype markers,proinflammatory cytokines and VDR-PPARγpathway were determined.RESULTS Supplementation with 1,25(OH)_(2)D_(3) relieved hepatic steatosis and decreased the proinflammatory M1 polarization of hepatic macrophages in NAFLD.Administration of 1,25(OH)_(2)D_(3) suppressed the proinflammatory M1 polarization of macrophages induced by fatty acids,thereby directly relieving lipid accumulation and metabolism in hepatocytes.The VDR-PPARγpathway had a notable impact on reversing lipid-induced proinflammatory M1 polarization of macrophages regulated by the administration of 1,25(OH)_(2)D_(3).CONCLUSION Supplementation with 1,25(OH)_(2)D_(3) improved hepatic steatosis and lipid metabolism in NAFLD,linked to its capacity to reverse the proinflammatory M1 polarization of hepatic macrophages,partially by regulating the VDRPPARγpathway.The involvement of 1,25(OH)_(2)D_(3) in inhibiting fatty-acid-induced proinflammatory M1 polarization of macrophages played a direct role in relieving lipid accumulation and metabolism in hepatocytes.

Effect of hepatocyte growth factor on inflammatory factors associated with CCL_(4)-induced hepatocyte injury

Objective:The aim of this study is to investigate the effects of Hepatocyte Growth Factor(HGF)on the expression levels of IL-8,TNF-α,IL-4,and IL-21 in mice with liver injury induced by CCL_(4).Methods:An acute liver injury mouse model was established using CCL_(4),and hepatocytes and white blood cells were separated by gradient density centrifugation.Different concentrations of HGF were added in vitro,and the expression levels of cytokines were detected using ELISA.Results:In the in vivo injury model,the hepatocyte experiment results showed that the expression level of IL-8 was reduced in the 10 ng/mL HGF group compared to the injured hepatocyte group(P<0.05),and increased in the 50 ng/mL HGF group compared to the 10 ng/mL HGF group(P<0.05).For IL-4,the expression levels were reduced in both the 25 ng/mL HGF group(P<0.05)and the 50 ng/mL HGF group(P<0.05)compared to the injured hepatocyte group.The white blood cell experiment results showed that the expression levels of TNF-αwere reduced in both the 10ng/ml HGF group(P<0.05)and the 25 ng/mL HGF group(P<0.05)compared to the injured white blood cell group.In the in vitro injury model,hepatocyte experiment results showed that the expression levels of TNF-αwere reduced in both the 25 ng/mL HGF group(P<0.05)and the 50 ng/mL HGF group(P<0.05)compared to the normal control group.For IL-4,the expression level was reduced in the 25 ng/mL HGF group compared to the normal control group(P<0.05).The white blood cell experiment results showed that the expression level of TNF-αwas increased in the 50 ng/mL HGF group compared to the 10 ng/mL HGF group(P<0.001);for IL-21,the expression levels were reduced in the CCL_(4) model group(P<0.05),10 ng/mL HGF group(P<0.05),25 ng/mL HGF group(P<0.05),and 50 ng/mL HGF group(P<0.05)compared to the normal control group.Conclusion:when the liver of mice is acutely damaged by CCL_(4),HGF can reduce the expression levels of inflammatory cytokines IL-8,TNF-α,IL-4 in hepatocytes,and TNF-αin liver white blood cells.

Dexamethasone potentiates the insulin-induced Srebp-1c expression in primary rat hepatocytes

The impacts of dexamethasone(Dex)and thyroid hormone T3 on the insulin-stimulated Srebp-1c expression were studied in primary rat hepatocytes. Primary hepatocytes from Sprague-Dawley rats were isolated, cultured and treated with insulin in the presence or absence of the indicated reagents over time. The mRNA levels of indicated genes were determined using real-time PCR. Insulin treatment induced the Srebp-1c expression and suppressed the Pck1 expression in a time-dependent manner. Dex treatment alone reduced the Srebp-1c expression, whereas potentiated the insulin-induced its expression, which reached to a level that was higher than the insulin alone group. On the other hand, insulin treatment completely suppressed the Dex-induced Pck1 expression in the same cells. T3 treatment did not affect the expressions of Srebp-1c and Pck1 alone or in the presence of absence of insulin or Dex. Interestingly, insulin treatment induced the Rxrg m RNA expression level in the absence or presence of T0901317, a specific agonist for the liver X receptor. Dex and insulin mutually affect each other's ability to regulate the expression levels of hepatic genes involved in glucose and fatty acid metabolism. Insulin induced Rxrg expression in primary hepatocytes, which may contribute to the induction of Srebp-1c expression in the same cells.

Purification and Characterization of Hepatocyte Regeneration Stimulatory Factor from Shark Liver

Aim To purify hepatocyte regeneration stimulatory factor from shark liver and research its molecular feature and activity. Methods and Results Hepatocyte regeneration stimulatory factor (sHRSF) was isolated from healthy shark livers and separated by homogenization, freezing melting, heat treating, centrifugation, and ultrafiltration. HRSF activity was found mainly in the subfraction of molecular weight less than 30 000 daltons. This crude ultrafiltrate was further purified successively by DEAE Sepharose fast flow chromatography, FPLC Resource 30Q, Resource Q and Mono Q chromatography. A single band was displayed on sodium dodecyl sulfate polyacrylamide gel electrophoresis, which corresponds to molecular weight of 14 600 daltons. The characteristic absorption was obtained at the wavelength 276 nm. The isoelectric point was about 5 1. It contained 18 amino acids and the 15 N terminal amino acid residues were LVGPIGAVGPAGKDG. It had a significant activity in stimulating liver to regenerate. Conclusion We obtained an unknown new active protein, that is hepatocyte regeneration stimulatory factor from shark liver (sHRSF).

Cyclic feeding regime may delay aging in animals by enhancing the hepatocytes nuclei structure

Background:The liver is fundamental for keeping up the entire body’s homeostasis.The liver hepatocytes have been shown to undergo genomic instability with aging.The stability of the hepatocytes depends on its nuclear architecture.Calorie restriction has been shown to extend life-span favorably and this may be through the reorganization of the nuclear structure.Objective:To study the effect of cyclic feeding regime on the chromatin assembly anchored to the nuclear membrane scaffold of rat models hepatocytes nuclei.Method:Rats models underwent cyclic feeding regime,after which nuclei were isolated;then,we investigated the chromatin decondensation and nuclear membrane disintegration of the hepatocytes using fluorescence imaging methods.Results:In 60 seconds,protease decondensed the chromatin and disintegrated the nuclear membrane structure of controls.After the first fasting,the time increased to 145 seconds in 3-month-old rats.The first refeeding increased the time to 156 seconds with a further rise to 340 seconds following the second fasting,then dropped to 116 seconds by the second refeeding.20 months old rats showed 186 seconds increase in the time of chromatin decondensation and nuclear membrane disintegration after the first fasting,with a decrease to 140 seconds observed after first refeeding.The second fasting increased the time to 165 seconds,which then slightly decreased to 163 seconds after the second refeeding.Conclusion:These results show that intermittent fasting may have acted on chromatin histone interactions and the structural lamin networks of the nuclear membranes in bringing about nuclear stability,which is essential for normal cellular function.

Reversible immortalization of human hepatocytes mediated by retroviral transfer and site-specific recombination

AIM: To establish a method for the reversible immortalization of human hepatocytes, which may offer a good and safe source of hepatocytes for practical applications.

Hepatocyte transplantation program:Lessons learned andfuture strategies

This review aims to share the lessons we learned over time during the setting of the hepatocyte transplantation(HT) program at the Hepatic Cell Therapy Unit at Hospital La Fe in Valencia. New sources of liver tissue for hepatocyte isolation have been explored. The hepatocyte isolation and cryopreservation procedures have been optimized and quality criteria for assessment of functionality of hepatocyte preparations and suitability for HT have been established. The results indicate that:(1) Only highly viable and functional hepatocytes allow to recover those functions lacking in the native liver;(2) Organs with steatosis(≥ 40%) and from elderly donors are declined since low hepatocyte yields, viability and cell survival after cryopreservation, are obtained;(3) Neonatal hepatocytes are cryopreserved without significant loss of viability or function representing high-quality cells to improve human HT;(4) Cryopreservation has the advantage of providing hepatocytes constantly available and of allowing the quality evaluation and suitability for transplantation; and(5) Our results from 5 adults with acute liver failure and 4 from children with inborn metabolic diseases, indicate that HT could be a veryuseful and safe cell therapy, as long as viable and metabolically functional human hepatocytes are used.

肿瘤组织中GPC3、Hepatocyte、CD_(34)检测对AFP阴性肝细胞癌的辅助诊断作用

目的评价肿瘤组织中磷脂酰肌醇蛋白聚糖3(GPC3)、Hepatocyte、CD34蛋白检测对甲胎蛋白(AFP)阴性肝细胞癌(HCC)的辅助诊断作用。方法回顾性分析48例癌细胞AFP阴性的HCC组织及其癌旁组织中GPC3、Hepatocyte、CD34蛋白表达特点。结果 48例HCC及其癌旁组织中GPC3蛋白阳性率分别为91.7%(44/48)、2.1%(1/48),两者比较P<0.01;Hepatocyte蛋白阳性率分别为95.8%(46/48)、100.0%(48/48),两者比较P>0.05;CD34蛋白阳性率分别为100.0%(48/48)、4.2%(2/48),两者比较P<0.01。HCC组织中GPC3与Hepatocyte表达呈正相关(r=0.314,P<0.05)。结论检测肿瘤组织中GPC3、Hepatocyte和CD34蛋白有助于AFP阴性原发性HCC的诊断。

Porcine hepatocyte isolation and reversible immortalization mediated by retroviral transfer and site-specific recombination

AIM: To develop a hepatocyte cell line, we immortalized primary porcine hepatocytes with a retroviral vector SSR#69 containing the Simian Virus 40 T antigen (SV40T ag). METHODS: We first established a method of porcine hepatocyte isolation with a modified four-step retrograde perfusion technique. Then the porcine hepatocytes were immortalized with retroviral vector SSR#69 expressing SV40T and hygromycin-resistance genes flanked by paired loxP recombination targets. SV40T cDNA in the expanded cells was subsequently excised by Cre/LoxP site-specific recombination. RESULTS: The resultant hepatocytes with high viability (97%) were successfully immortalized with retroviral vector SSR#69. One of the immortalized clones showed the typical morphological appearance, TJPH-1, and was selected by clone rings and expanded in culture. After excision of the SV40T gene with Cre-recombinase, cells stopped growing. The population of reverted cells exhibited the characteristics of differentiated hepatocytes. CONCLUSION: In conclusion, we herein describe a modified method of hepatocyte isolation and subsequently established a porcine hepatocyte cell line mediated by retroviral transfer and site-specific recombination.

Protective effects of ACLF sera on metabolic functions and proliferation of hepatocytes co-cultured with bone marrow MSCs in vitro

AIM： To investigate whether the function of hepatocytes co-cultured with bone marrow mesenchymal stem cells （MSCs） could be maintained in serum from acute-on- chronic liver failure （ACLF） patients.METHODS： Hepatocyte supportive functions and cy- totoxicity of sera from 18 patients with viral hepatitis B-induced ACLF and 18 healthy volunteers were evalu- ated for porcine hepatocytes co-cultured with MSCs and hepatocyte mono-layered culture, respectively. Chemo- kine profile was also examined for the normal serum and liver failure serum.RESULTS： Hepatocyte growth factor （HGF） and Tumor necrosis factor; tumor necrosis factor （TNF）-a were re- markably elevated in response to ACLF while epidermal growth factor （EGF） and VEGF levels were significantly decreased. Liver failure serum samples induced a higher detachment rate, lower viability and decreased liver sup- port functions in the homo-hepatocyte culture. Hepato-cytes co-cultured with MSCs could tolerate the cytotoxic- ity of the serum from ACLF patients and had similar liver support functions compared with the hepatocytes cul- tured with healthy human serum in vitro. In addition, co- cultured hepatocytes maintained a proliferative capability despite of the insult from liver failure serum.CONCLUSION： ACLF serum does not impair the cell morphology, viability, proliferation and overall metabolic capacities of hepatocyte co-cultured with MSCs in vitro.

Pediatric non-alcoholic steatohepatitis:The first report on the ultrastructure of hepatocyte mitochondria

AIM: To investigate the ultrastructure of abnormal hepatocyte mitochondria, including their cellular and hepatic zonal distribution, in bioptates in pediatric non-alcoholic steatohepatitis (NASH).

Hepatocytes isolated from neoplastic liver-immunomagnetic purging as a new source for transplantation

AIM： To investigate whether hepatocytes isolated from macroscopically normal liver during hepatic resection for neoplasia could provide a novel source of healthy hepatocytes, including the development of reliable protocols for malignant cells removal from the hepatocyte preparation. METHODS： Hepatocytes were procured from resected liver of 18 patients with liver tumors using optimised digestion and cell-enrichment protocols. Suspensions of various known quantities of the HT-29 tumor cell line and patient hepatocytes were treated or not with Ep-CAM-antibody-coated immunomagnetic beads in order to investigate the efficacy of tumor-purging by immunomagnetic depletion, using a semi-quantitative RT-PCR method developed to detect tumor cells. Immunomagnetic bead-treated or bead-untreated tumor cell-hepatocyte suspensions were transplanted intra-peritoneally in Balb/C nude mice to assess the rates of tumor development. RESULTS： Mean viable hepatocyte yield was 9.3×10^6 cells per gram of digested liver with mean viability of 70.5%. Immunomagnetic depletion removed tumor cells to below the RT-PCR detection-threshold of 1 tumor cell in 10^6 hepatocytes, representing a maximum tumor purging efficacy of greater than 400000-fold. Transplanted, immunomagnetic bead-purged tumor cell-hepatocyte suspensions did not form peritoneal tumors in Balb/C nude mice. Co-transplantation of hepatocytes with tumor cells did not increase tumorigenesis of the tumor cells. CONCLUSION： Immunomagnetic depletion appears to be an effective method of purging contaminating tumor cells to below threshold for likely tumorigenesis. Along with improved techniques for isolation of large numbers of viable hepatocytes, normal liver resected for neoplasia has potential as another clinically useful source of hepatocytes for transplantation.

Controlled and reversible induction of differentiation and activation of adult human hepatocytes by a biphasic culture technique

AIM:Clinical application of human hepatocytes (HC) is hampered by the progressive loss of growth and differentiation in vitro. The object of the study was to evaluate the effect of a biphasic culture technique on expression and activation of growth factor receptors and differentiation of human adult HC. METHODS: Isolated HC were sequentially cultured in a hormone enriched differentiation medium (DM) containing nicotinamide, insulin, transferrin, selenium, and dexame-thasone or activation medium (AM) containing hepatocyte growth factor (HGF), epidermal growth factor (EGF), and granulocyte-macrophage colony-stimulating factor (GMCSF). Expression, distribution and activation of the HC receptors (MET and EGFR) and the pattern of characteristic cytokeratin (CK) filaments were measured by fluorometry, confocal microscopy and Western blotting. RESULTS: In the biphasic culture system, HC underwent repeated cycles of activation (characterized by expression and activation of growth factor receptors) and re-differentiation (illustrated by distribution of typical filaments CK-18 but low or absent expression of CK-19). In AM increased expression of MET and EGFR was associated with receptor translocation into the cytoplasm and induction of atypical CK-19. In DM low expression of MET and EGFR was localized on the cell membrane and CK-19 was reduced. Receptor phosphorylation required embedding of HC in collagen type I gel. CONCLUSION: Control and reversible modulation of growth factor receptor activation of mature human HC can be accomplished in vitro, when defined signals from the extracellular matrix and sequential growth stimuli are provided. The biphasic technique helps overcome dedifferentiation, which occurs during continuous stimulation by means of growth factors.

Hepatotropic growth factors protect hepatocytes during inflammation by upregulation of antioxidative systems

AIM:To investigate effects of hepatotropic growth factors on radical production in rat hepatocytes during sepsis.METHODS:Rat hepatocytes,isolated by collagenase perfusion,were incubated with a lipopolysaccharide(LPS)-containing cytokine mixture of interleukin-1β,tumor necrosis factor-α and interferon-γ to simulate sepsis and either co-incubated or pre-incubated with hepatotropic growth factors,e.g.hepatocyte growth factor,epidermal growth factor and/or transforming growth factor-α.Cells were analyzed for glutathione levels.Culture supernatants were assayed for produc-tion of reactive oxygen intermediates(ROIs) as well as NO2-,NO3-and S-nitrosothiols.To determine cellular damage,release of aspartate aminotransferase(AST) into the culture medium was analyzed.Activation of nuclear factor(NF)-κB was measured by electrophoretic mobility shift assay.RESULTS:Rat hepatocytes treated with the LPS-containing cytokine mixture showed a significant increase in ROI and nitrogen oxide intermediate formation.AST leakage was not significantly increased in cells treated with the LPS-containing cytokine mixture,independent of growth-factor co-stimulation.However,pretreatment with growth factors significantly reduced AST leakage and ROI formation while increasing cellular glutathione.Application of growth factors did not result in increased NF-κB activation.Pretreatment with growth factors further increased formation of NO2-,NO3-and S-nitrosothiols in hepatocytes stimulated with LPS-containing cytokine mixture.Thus,we propose that,together with an increase in glutathione increased NO2-,NO3-formation might shift their metabolism towards non-toxic products.CONCLUSION:Our data suggest that hepatotropic growth factors positively influence sepsis-induced hepatocellular injury by reducing cytotoxic ROI formation via induction of the cellular protective antioxidative systems.

Unexpected discovery of 2 cases of hepatocyte nuclear factor 1α-mutated infracentimetic adenomatosis

We present 2 cases of hepatocyte nuclear factor 1α (HNF1α)-mutated adenomatosis, discovered for reasons unrelated to this disease, and identified using immunohistochemical methods. These new tools may further our understanding of the link between adenomas/adenomatosis subtypes and their complications, and their association with other abnormalities.