Evaluation of metabolic dysfunction-associated fatty liver disease using FibroScan, diet, and microbiota: A large cross-sectional study.

OBJECTIVE: We evaluated the clinical characteristics of metabolic dysfunction-associated fatty liver disease (MAFLD) to evaluate the usefulness of the MAFLD diagnostic criteria in a resident health survey. METHODS: In 1056 participants of a health survey, we compared obesity, diabetes, metabolic dysregulation, FibroScan-aspartate aminotransferase (FAST) score, dietary habits, and gut microbiota between healthy individuals and participants with MAFLD and Nonalcoholic fatty liver disease (NAFLD). RESULTS: The proportion of participants with MAFLD in the fatty liver was higher than that with NAFLD (88.1% vs. 75.5%, respectively). Of 36 participants with a FAST score > 0.35, 29 (80.6%) participants had MAFLD and 23 (63.9%) participants had NAFLD. Of 29 patients with liver fibrosis, 26 (89.7%) participants had obesity and metabolic dysregulation. In the evaluation of diet, the total energy, protein, dietary fiber, and salt intake were significantly higher in participants with MAFLD than those in participants without fatty liver. In the microbiota analysis, the results of the linear discriminant analysis effect size analysis revealed nine bacterial genera that were significantly different in participants with MAFLD in comparison with participants without fatty liver. Of these genera, the relative abundance of Blautia was especially low in participants with MAFLD. CONCLUSION: In a resident health survey, participants with MAFLD had a higher proportion of fatty liver than those with NAFLD. MAFLD criteria could help in improved screening of participants with liver fibrosis. Therefore, the MAFLD criteria could be a useful diagnostic tool for aggressively identifying participants with a high risk of fatty liver. Additionally, Blautia might be involved in the development of MAFLD.

Reduced Equol Production and Gut Microbiota Features in Men With Lean Nonalcoholic Fatty Liver Disease.

Patients with lean nonalcoholic fatty liver disease (NAFLD) may have different metabolic profiles than those with NAFLD. Estrogenic activity is associated with NAFLD pathogenesis. We evaluated the production ability of equol, which has estrogenic activity, in lean NAFLD and assessed their gut microbiota in relation to their equol-producing ability. Among 684 adult participants, 276 (40%) had NAFLD and 293 (43%) were equol producers. The rates of equol producers in the normal and NAFLD groups were 43% and 42%, respectively. Among the patients with NAFLD, 55 (20%) had lean NAFLD of which 18 (33%) were equol producers. The rate of equol production in men with lean NAFLD was 8%, which was the lowest, while the corresponding rate in the other participants was approximately 40%. The gut microbiota composition of equol producers and nonproducers showed many significant differences. The gut microbiota of men with lean NAFLD showed increased abundance of Caulobacter and decreased abundances of Slackia and Terrisporobacter. Thus, almost all men with lean NAFLD lacked equol-producing ability, and their gut microbiota showed a reduced abundance of Slackia, which is related to equol production. The pathology of lean NAFLD in men may be strongly associated with equol-producing ability and gut microbiota.

Association of serum creatinine-to-cystatin C ratio with skeletal muscle mass and strength in nonalcoholic fatty liver disease in the Iwaki Health Promotion Project.

We evaluated the feasibility of using serum creatinine-to-cystatin C ratio in the assessments of muscle mass and strength in nonalcoholic fatty liver disease. In a community-based cross-sectional study, skeletal muscle mass and handgrip strength were assessed in 641 Japanese adults. Low skeletal muscle mass index and low handgrip strength were defined as indicated in the sarcopenia diagnostic criteria of the Japan Society of Hepatology. Nonalcoholic fatty liver disease was defined as fatty liver on ultrasonography in the absence of other causes of steatosis. The creatinine-to-cystatin C ratio was useful for identifying the participants with low skeletal muscle mass index, with an area under the receiver-operating characteristic curve of 0.84 [95% confidence interval (CI), 0.77-0.91] in men and 0.72 in women (95% CI, 0.65-0.78), and those with low handgrip strength, with an area under the receiver-operating characteristic curve of 0.96 (95% CI, 0.93-0.99) in men and 0.79 (95% CI, 0.66-0.92) in women. Moreover, the creatinine-to-cystatin C ratio correlated with skeletal muscle mass index (r = 0.511, p<0.001) and handgrip strength (r = 0.657, p<0.001), whereas it did not correlate with exacerbation of hepatic steatosis. In this study, creatinine-to-cystatin C ratio correlated with muscle mass and strength in nonalcoholic fatty liver disease regardless of hepatic steatosis.

Association between serum ß-carotene-to-retinol ratio and severity of hepatic steatosis in non-alcoholic fatty liver disease in Japan: A cross-sectional study.

OBJECTIVES: Retinol and ß-carotene have been reported to be involved in the pathogenesis of non-alcoholic fatty liver disease (NAFLD). However, clinical studies are limited. The aim of this study was to investigate the relationship between serum the ratio of ß-carotene to retinol (SC/SR) and hepatic steatosis in NAFLD diagnosed by ultrasonography. METHODS: The participants were 606 Japanese adults who were enrolled in a health survey. Clinical profile, dietary nutrition intake, blood biochemistry, serum retinol, and carotenoids were analyzed. NAFLD was defined as fatty liver on ultrasonography in the absence of other causes of steatosis. RESULTS: Women had higher daily intake of α- and ß-carotene, although there were no differences in daily retinol and carotenoid intake between participants with or without NAFLD in both men and women. Women had a higher SC/SR ratio than men regardless of the presence or absence of NAFLD, and the SC/SR ratio in women decreased with exacerbation of hepatic steatosis, whereas the SC/SR ratio in men did not change despite exacerbation of hepatic steatosis. After adjusting for confounding factors, the likelihood of NAFLD among participants in the highest quartile of SC/SR ratio decreased by two-thirds compared with participants in the lowest quartile (adjusted odds ratio, 0.64; 95% confidence interval, 0.21-1.92; P = 0.041). The SC/SR ratio was positively correlated with serum high-density lipoprotein cholesterol level, and negatively correlated with serum triacylglycerol level. CONCLUSIONS: The SC/SR ratio was lower in NAFLD with sex differences, and was associated with the severity of hepatic steatosis and lipid profile. Future studies are needed to expand on these findings.

Changed Amino Acids in NAFLD and Liver Fibrosis: A Large Cross-Sectional Study without Influence of Insulin Resistance.

Altered amino acid levels have been found in nonalcoholic fatty liver disease (NAFLD) and nonalcoholic steatohepatitis (NASH). However, it is not clear whether this alteration is due to altered hepatic metabolism or insulin resistance. The aim of this study was to clarify the association among amino acid levels, fatty liver, and liver fibrosis while eliminating the influence of insulin resistance. NAFLD and liver fibrosis were diagnosed using transient elastography and subjects were divided into three groups: normal, NAFLD, and liver fibrosis. To exclude the influence of insulin resistance, the subjects were matched using the homeostasis model assessment of insulin resistance (HOMA-IR). The amino acid serum levels were compared among the groups. Of 731 enrolled subjects, 251 and 33 were diagnosed with NAFLD and liver fibrosis. Although significant differences were observed among the groups in the serum levels of most amino acids, all but those of glutamate and glycine disappeared after matching for HOMA-IR. The multivariate logistic regression revealed that glutamate, glycine, and HOMA-IR were independent risk factors for liver fibrosis. The altered serum levels of most amino acids were associated with insulin resistance, while the increase in glutamate and the decrease in glycine levels were strongly associated not only with insulin resistance, but also with altered liver metabolism in patients with liver fibrosis.

Association of Bone Metabolism with Fatty Liver Disease in the Elderly in Japan: A Community-based Study.

Objective With the aging of society, both osteoporosis and fatty liver disease (FLD) are becoming important issues. However, the relationship between osteoporosis and FLD remains controversial. We investigated the association between bone metabolism and FLD in a Japanese community in a cross-sectional study. Methods A total of 1,020 participants were enrolled in a health survey. FLD was diagnosed by ultrasonography. Bone metabolism was evaluated based on bone mineral density (BMD), which was assessed using dual-energy X-ray absorptiometry, and with the bone formation index (total type I procollagen N-terminal propeptide/bone-alkaline phosphatase ratio; P1NP/BAP ratio) and the bone resorption index (crosslinked N-telopeptide of type I collagen/tartrate-resistant acid phosphatase-5b ratio; NTx/TRACP-5b ratio) calculated from serum bone turnover markers. Results The BMD (percentage of the young adult mean) was the same level in both male and female participants with and without FLD. Both men and women showed an age-dependent decrease in their bone formation index and bone resorption index values. Men of ≥70 years of age and women of 60-69 years of age with FLD had significantly lower bone formation index values and higher bone resorption index values. However, similar findings were not seen in women of ≥70 years of age. Conclusion Although the BMD levels were the same, regardless of the presence or absence of FLD, elderly participants with FLD showed decreased bone formation and increased bone resorption, with sex differences. Because our results suggest that FLD in elderly individuals is detrimental for bone metabolism, and that it leads to bone loss and osteoporosis, further studies using a cohort population are warranted.

Leptin/adiponectin ratio correlates with hepatic steatosis but not arterial stiffness in nonalcoholic fatty liver disease in Japanese population.

BACKGROUND AND AIMS: Cardiovascular disease (CVD) is a leading cause of mortality in nonalcoholic fatty liver disease (NAFLD). The aim of this study was to investigate the relationship of leptin-to-adiponectin (L/A) ratio with hepatic steatosis and arterial stiffness in NAFLD. METHODS: The subjects were 871 Japanese adults who participated in a health survey. Dietary intake, body composition, lipid profile, serum interleukin-6 (IL-6), leptin, and adiponectin were analyzed. NAFLD was defined as fatty liver on ultrasonography in the absence of other causes of steatosis. Arterial stiffness was evaluated by the brachial-ankle pulse wave velocity (baPWV). RESULTS: The subjects with NAFLD had a greater body mass index (BMI) and body fat percentage (BFP); a higher intake of daily energy (kcal) and carbohydrates; and a higher prevalence of hypertension, diabetes, and hyperlipidemia. The subjects with NAFLD had higher serum leptin and lower serum adiponectin concentrations and a higher L/A ratio than subjects without NAFLD. The L/A ratio increased with increasing severity of steatosis. The L/A ratio showed positive correlations with BMI and BFP, and a negative correlation with age. Women had higher L/A ratio and BFP levels than men regardless of the presence or absence of NAFLD. There was a weak positive correlation between baPWV and severity of steatosis. BaPWV was strongly correlated with age, while no relation was found between baPWV and L/A ratio. IL-6 level was correlated with baPVW and age, while the correlation between Il and 6 level and L/A ratio was very weak. The L/A ratio was correlated with triglycerides and the ratio of total cholesterol to high-density lipoprotein-cholesterol. CONCLUSION: L/A ratio and arterial stiffness were associated with the severity of steatosis, whereas there was no correlation between L/A ratio and arterial stiffness in NAFLD. These findings suggest that not only leptin and adiponectin but also other factors might be involved in the pathogenesis for atherosclerosis in NAFLD.

Significant decrease in Faecalibacterium among gut microbiota in nonalcoholic fatty liver disease: a large BMI- and sex-matched population study.

BACKGROUND: Compositional changes of the gut microbiota are known to occur in patients with nonalcoholic fatty liver disease (NAFLD); however, the changes did not corroborate between the studies. We evaluated the gut microbiota between NAFLD and non-NAFLD participants, excluding the influence of obesity and sex in this study involving a large number of participants. METHODS: In total, 1148 adults participated in the health survey. NAFLD was defined as fatty liver by ultrasonography in the absence of other causes of steatosis. To exclude the influence of obesity and sex, NAFLD participants were matched to non-NAFLD participants based on BMI and sex. The relative abundance of each bacterial taxa in fecal samples was calculated using 16S ribosomal RNA amplification and was compared between NAFLD and non-NAFLD participants. RESULTS: There were 205 (23.5%) participants defined as having NAFLD. Before matching, there were significant differences in the relative abundance of more than 1% in two classes, two orders, three families, and three genera including Faecalibacterium between NAFLD and non-NAFLD participants. After matching, 153 matched pairs were obtained. In terms of the relative abundance of more than 1%, the relative abundance of two taxa, including the family Ruminococcaceae and the genus Faecalibacterium, was significantly lower in NAFLD participants than in non-NAFLD participants (p = 0.016 and p = 0.018). CONCLUSIONS: The significant decrease in Faecalibacterium is a remarkable characteristic on BMI- and sex-matched analysis in NAFLD participants in a large study population. The decrease in Faecalibacterium is related to the pathogenesis of NAFLD.

Interventional Treatment for Giant Hepatic Hemangioma Accompanied by Arterio-portal Shunt with Ascites.

A 73-year-old woman with massive ascites associated with a giant hepatic mass accompanied by arterio-portal (AP) shunt was admitted to our hospital. Based on contrast-enhanced computed tomography (CT) and angiography findings, hepatic hemangioma with AP shunt and ascites due to portal hypertension was diagnosed. Transcatheter arterial embolization (TAE) by N-butyl-2-cyanoacrylate (NBCA) was performed without complications. The patient's ascites disappeared, and her liver function test results improved after the treatment. The patient has maintained a steady state for two years. This case indicates that TAE with NBCA is a safe and effective treatment for hepatic hemangioma accompanied by AP shunt.

Inhibition of Systemic Hyaluronan Synthesis Exacerbates Murine Hepatic Carcinogenesis.

BACKGROUND/AIM: Hyaluronan (HA) is used as a biomarker of liver fibrosis, which is a key risk factor for the development of hepatocellular carcinoma (HCC). We examined the effects of prolonged pharmacological inhibition of HA synthesis on liver carcinogenesis. MATERIALS AND METHODS: Liver tumors were induced in mice by administering 0.03% thioacetamide (TAA) in drinking water over a 12-month period. Animals simultaneously received either a diet containing of an inhibitor of HA synthesis [4-methylumbelliferone (4-MU)], or a control diet. RESULTS: Addition of 4-MU resulted in a significantly higher number of tumors compared to TAA treatment alone. Moreover, addition of 4-MU resulted in a dose-dependent increase in maximum tumor size. CONCLUSION: While local HA suppression has been shown to have an inhibitory effect on HCC in vitro and in tumor cell implantation experiments, the present results indicate that systemic inhibition of HA synthesis by 4-MU supplementation facilitates hepatic carcinogenesis in vivo.

Kidney Injury due to Ureteral Obstruction Caused by Compression from Infected Simple Hepatic Cyst.

Simple hepatic cysts are common and most often asymptomatic. In symptomatic cases, hemorrhage, rupture, and infection are major complications. However, urinary tract obstruction caused by a simple hepatic cyst is rare. We treated an 82-year-old Japanese man with an infected giant hepatic cyst causing right hydronephrosis who had a past history of left nephrectomy for renal cell carcinoma. The patient underwent ultrasound-guided percutaneous drainage and sclerotherapy with minocycline hydrochloride for the infected hepatic cyst. Right hydronephrosis was relieved, and renal dysfunction improved with regression of the hepatic cyst after treatment. This is the first report of hydronephrosis due to ureteral obstruction caused by compression from a hepatic cyst.

Two Cases of Primary Sclerosing Cholangitis Overlapping with Autoimmune Hepatitis in Adults.

Overlap syndrome between primary sclerosing cholangitis (PSC) and autoimmune hepatitis (AIH) is extremely rare in Japan. We herein report two adult patients with PSC-AIH overlap syndrome. They were diagnosed with PSC-AIH overlap syndrome based on the findings of endoscopic retrograde cholangiography and liver biopsy, and using the International Autoimmune Hepatitis Group scoring system. In both cases, PSC preceded AIH, and combination therapy with steroid and ursodeoxycholic acid was effective. Because there are few reported cases in Japan, it is important to study more cases to shed light on the clinical and pathological features of PSC-AIH overlap syndrome.

Identification of a novel alpha-fetoprotein-expressing cell population induced by the Jagged1/Notch2 signal in murine fibrotic liver.

The liver is well known to possess high regenerative capacity in response to partial resection or tissue injury. However, liver regeneration is often impaired in the case of advanced liver fibrosis/cirrhosis when mature hepatocytes can hardly self-proliferate. Hepatic progenitor cells have been implicated as a source of hepatocytes in regeneration of the fibrotic liver. Although alpha-fetoprotein (AFP) is known as a clinical marker of progenitor cell induction in injured/fibrotic adult liver, the origin and features of such AFP-producing cells are not fully understood. Here, we demonstrate a unique and distinct AFP-expressing cell population that is induced by the Jagged1/Notch2 signal in murine fibrotic liver. Following repeated carbon tetrachloride injections, a significant number of AFP-positive cells with high proliferative ability were observed along the fibrous septa depending on the extent of liver fibrosis. These AFP-positive cells exhibited features of immature hepatocytes that were stained positively for hepatocyte-lineage markers, such as albumin and hepatocyte nuclear factor 4 alpha, and a stem/progenitor cell marker Sox9. A combination of immunohistological examination of fibrotic liver tissues and coculture experiments with primary hepatocytes and hepatic stellate cells indicated that increased Jagged1 expression in activated hepatic stellate cells stimulated Notch2 signaling and up-regulated AFP expression in adjacent hepatocytes. The mobilization and proliferation of AFP-positive cells in fibrotic liver were further enhanced after partial hepatectomy, which was significantly suppressed in Jagged1-conditional knockout mice. Finally, forced expression of the intracellular domain of Notch2 in normal liver induced a small number of AFP-expressing hepatocytes in vivo. Conclusion: Insight is provided into a novel pathophysiological role of Jagged1/Notch2 signaling in the induction of AFP-positive cells in fibrotic liver through the interaction between hepatocytes and activated hepatic stellate cells. (Hepatology Communications 2017;1:215-229).

A successful case of transcatheter arterial embolization with n-butyl-2-cyanoacrylate for pancreatic arteriovenous malformation.

A 57-year-old man was referred and admitted to our hospital for treatment of a symptomatic pancreatic mass. Pancreatic arteriovenous malformation (AVM) was diagnosed based on the findings of contrast-enhanced computed tomography (CT) and angiography, and transcatheter arterial embolization (TAE) with N-butyl-2-cyanoacrylate (NBCA) was performed without complications. The patient's symptoms subsequently improved after TAE, and resolution of the pancreatic AVM was detected on contrast-enhanced CT performed six months after the embolization procedure. This case indicates that TAE with NBCA is a safe and effective treatment for pancreatic AVM.

Differential contribution of dermal resident and bone marrow-derived cells to collagen production during wound healing and fibrogenesis in mice.

Recent studies show that bone marrow (BM)-derived cells migrating into a dermal wound promote healing by producing collagen type I. However, their contribution to the repair process has not been fully verified yet. It is also unclear whether BM-derived cells participate in dermal fibrogenesis. We have addressed these issues using transgenic mice that harbor tissue-specific enhancer/promoter sequences of α2(I) collagen gene linked to either enhanced green fluorescent protein (COL/EGFP) or the luciferase (COL/LUC) reporter gene. Following dermal excision or subcutaneous bleomycin administration, a large number of EGFP-positive collagen-producing cells appeared in the dermis of COL/EGFP reporter mice. When wild-type mice were transplanted with BM cells from transgenic COL/EGFP animals and subjected to dermal excision, no EGFP-positive BM-derived collagen-producing cells were detected throughout the repair process. Luciferase assays of dermal tissues from COL/LUC recipient mice also excluded collagen production by BM-derived cells during dermal excision healing. In contrast, a limited but significant number of CD45-positive collagen-producing cells migrated from BM following bleomycin injection. These results indicate that resident cells in the skin are the major source of de novo collagen deposition in both physiological and pathological conditions, whereas BM-derived cells participate, in part, in collagen production during dermal fibrogenesis.

Negligible contribution of bone marrow-derived cells to collagen production during hepatic fibrogenesis in mice.

BACKGROUND & AIMS: Recent studies have reported that bone marrow (BM)-derived cells migrating into fibrotic liver tissue exhibit a myofibroblast-like phenotype and may participate in the progression of liver fibrosis. However, their contribution to collagen production has not been fully verified yet. We revisited this issue by using 2 mechanistically distinct liver fibrosis models introduced into transgenic collagen reporter mice and their BM recipients. METHODS: BM of wild-type mice was replaced by cells obtained from transgenic animals harboring tissue-specific enhancer/promoter sequences of alpha2(I) collagen gene (COL1A2) linked to enhanced green fluorescent protein (EGFP) or firefly luciferase (LUC) gene. Liver fibrosis was introduced into those mice by repeated carbon tetrachloride injections or ligation of the common bile duct. Activation of COL1A2 promoter was assessed by confocal microscopic examination detecting EGFP signals and luciferase assays of liver homogenates. RESULTS: The tissue-specific COL1A2 enhancer/promoter was activated in hepatic stellate cells following a single carbon tetrachloride injection or during primary culture on plastic. A large number of EGFP-positive collagen-expressing cells were observed in liver tissue of transgenic COL1A2/EGFP mice in both liver fibrosis models. In contrast, there were few EGFP-positive BM-derived collagen-producing cells detected in fibrotic liver tissue of COL1A2/EGFP recipients. Luciferase assays of liver tissues from COL1A2/LUC-recipient mice further indicated that BM-derived cells produced little collagen in response to fibrogenic stimuli. CONCLUSIONS: By using a specific and sensitive experimental system, which detects exclusively BM-derived collagen-producing cells, we conclude an unexpectedly limited role of BM-derived cells in collagen production during hepatic fibrogenesis.

Apoptotic body engulfment by hepatic stellate cells promotes their survival by the JAK/STAT and Akt/NF-kappaB-dependent pathways.

BACKGROUND/AIMS: We have previously shown that phagocytosis of apoptotic bodies (AB) by hepatic stellate cells (HSC) is profibrogenic. As HSC survival is central to the progression of liver fibrosis, our goal was to investigate if phagocytosis induces HSC survival. METHODS: Apoptosis of phagocytosing HSC was studied in the presence of known apoptotic agents. The JAK/STAT- and PI3K/Akt-dependent pathways, NF-kappaB activation and expression of the anti-apoptotic proteins Mcl-1 and A1 were evaluated. Apoptosis was assessed after blocking A1 by an siRNA approach. RESULTS: Phagocytosing HSC were resistant to FasL/cycloheximide or TRAIL-induced apoptosis. Inhibition of the JAK/STAT or PI3K-mediated pathways induced apoptosis of HSC. Phagocytosis induced JAK1/STAT3 phosphorylation, and this was prevented by inhibiting JAK. Translocation of STAT3 to the nucleus was also blocked by JAK inhibition. Mcl-1 expression was upregulated in a JAK-dependent manner. PI3K-dependent phosphorylation of Akt depended on NADPH oxidase activity and superoxide production. NF-kappaB activation and subsequent upregulation of A1 was observed, and A1 inhibition induced apoptosis of HSC. CONCLUSION: Phagocytosis of AB promotes HSC survival by two pathways, of which the A1 dependent is more significant. This represents a new mechanism by which engulfment of AB contributes to the propagation of liver fibrosis.

Leptin induces phagocytosis of apoptotic bodies by hepatic stellate cells via a Rho guanosine triphosphatase-dependent mechanism.

UNLABELLED: Leptin, a profibrogenic cytokine, plays an important role in the development of non-alcoholic steatohepatitis. Leptin also regulates immune responses, including macrophage phagocytic activity. Stellate cells are key elements in liver fibrogenesis, and previously we have demonstrated that phagocytosis of apoptotic bodies by stellate cells is profibrogenic. To study the effects of leptin on the phagocytic activity of hepatic stellate cells, we exposed both LX-2 cells and primary stellate cells to leptin, and we have observed increased phagocytic activity. In stellate cells isolated from Zucker (fa/fa) rats, the rate of phagocytosis was significantly decreased. To investigate the mechanism by which leptin induces phagocytosis, we focused on the role of Rho-guanosine triphosphate (GTP)-ases. We found that leptin induced the PI3K-dependent activation of Rac1, and that nicotinamide adenine dinucleotide phosphate, reduced form (NADPH) oxidase activation was also implicated in this process. Leptin also induced RhoA activation and translocation to the phagosomes. Expression of the constitutive active Rac1 and RhoA both increased the phagocytic rate, whereas inhibition of the Rho-dependent kinase decreased the phagocytic activity. CONCLUSION: We describe a novel role of leptin in the fibrogenic process, the induction of phagocytosis of apoptotic bodies by hepatic stellate cells. The data provide strong evidence of a Rho-GTPase-mediated regulation of the cytoskeleton during stellate cell phagocytosis. Leptin-mediated phagocytic activity of stellate cells therefore could be an important mechanism responsible for progression of fibrosis in non-alcoholic steatohepatitis.

Epimorphin, a morphogenic protein, induces proteases in rodent hepatocytes through NF-kappaB.

BACKGROUND/AIMS: Epimorphin, expressed by hepatic stellate cells in the liver, directs normal morphogenesis in various organs. The aim of this study was to clarify the mechanism by which epimorphin functions as a morphogen in vitro. METHODS: Male Balb/c mice and Sprague-Dawley rats were used. First, we explored the relationship between epimorphin expression and distribution of protease-positive cells in carbon tetrachloride-induced acute liver injury. We then examined protease levels in cultured hepatocytes and signal transduction of epimorphin. Finally, we determined the requirement for proteases and NF-kappaB in spheroid formation induced by epimorphin. RESULTS: Epimorphin expression was enhanced in injured areas during late recovery phase, in which protease-positive hepatocytes were localized adjacent to epimorphin-expressing cells. In vitro, epimorphin induced matrix metalloproteinase (MMP) 9, MMP 3 and urokinase type plasminogen activator (uPA) in hepatocytes. NF-kappaB mediated these protease expressions in hepatocytes. These proteases were required for epimorphin-induced and Matrigel induced spheroid. An epimorphin-neutralizing antibody also blocked spheroid formation on Matrigel, which contained epimorphin. In addition, NF-kappaB activation was also required for spheroid formation. CONCLUSION: Epimorphin elicits hepatocyte spheroids by inducing proteases in rodent hepatocytes through NF-kappaB.

Zinc L-carnosine protects against mucosal injury in portal hypertensive gastropathy through induction of heat shock protein 72.

BACKGROUND AND AIMS: Increased susceptibility to gastric mucosal injury is observed in portal hypertensive gastropathy (PHG). In this study, the effects of zinc L-carnosine, an anti-ulcer drug, were evaluated on expression of heat shock protein (hsp) 72 and cytoprotection in gastric mucosa in a rat model of PHG. METHODS: Portal hypertensive gastropathy with liver cirrhosis was induced by bile duct ligation for 4 weeks in male Sprague-Dawley rats. Expression of gastric mucosal hsp72 was evaluated by Western blotting at 6 h after intragastric administration of L-carnosine, zinc sulfate, or zinc L-carnosine. Blood was also collected for determination of serum zinc level. Mucosal protective abilities against hydrochloric acid (HCl) (0.6N) followed by pretreatment with L-carnosine, zinc sulfate or zinc L-carnosine were also studied. RESULTS: L-carnosine, zinc sulfate, and zinc L-carnosine induced hsp72 in gastric mucosa of rats with bile duct ligation. Zinc sulfate and zinc L-carnosine suppressed HCl-induced mucosal injury. However, L-carnosine could not suppress HCl-induced mucosal injury. Serum zinc levels were significantly elevated after zinc L-carnosine administration. Furthermore, pretreatment with zinc L-carnosine (30-300 mg/kg) increased the expression of hsp72 in gastric mucosa and prevented HCl-induced mucosal injury in rats with bile duct ligation in a dose-dependent manner. CONCLUSIONS: Zinc derivatives, especially zinc L-carnosine, protected portal hypertensive gastric mucosa with increased hsp72 expression in cirrhotic rats. It is postulated that zinc L-carnosine may be beneficial to the mucosal protection in PHG as a 'chaperone inducer'.