High chronic fructose consumption is linked to a nonalcoholic steatohepatitis (NASH) type of hepatotoxicity. Oxalate is the major endpoint of fructose metabolism, which accumulates in the kidney causing renal stone disease. Both diseases are life-threatening if not treated. Our objective was to study the molecular cytotoxicity mechanisms of fructose and some of its metabolites in the liver. Fructose metabolites were incubated with primary rat hepatocytes, but cytotoxicity only occurred if the hepatocytes were exposed to non-toxic amounts of hydrogen peroxide such as those released by activated immune cells. Glyoxal was most likely the endogenous toxin responsible for fructose induced toxicity formed via autoxidation of the fructose metabolite glycolaldehyde catalyzed by superoxide radicals, or oxidation by Fenton’s hydroxyl radicals. As for hyperoxaluria, glyoxylate was more cytotoxic than oxalate presumably because of the formation of condensation product oxalomalate causing mitochondrial toxicity and oxidative stress. Oxalate toxicity likely involved pro-oxidant iron complex formation.

High chronic fructose consumption is linked to a nonalcoholic steatohepatitis (NASH) type of hepatotoxicity. Oxalate is the major endpoint of fructose metabolism, which accumulates in the kidney causing renal stone disease. Both diseases are life-threatening if not treated. Our objective was to study the molecular cytotoxicity mechanisms of fructose and some of its metabolites in the liver. Fructose metabolites were incubated with primary rat hepatocytes, but cytotoxicity only occurred if the hepatocytes were exposed to non-toxic amounts of hydrogen peroxide such as those released by activated immune cells. Glyoxal was most likely the endogenous toxin responsible for fructose induced toxicity formed via autoxidation of the fructose metabolite glycolaldehyde catalyzed by superoxide radicals, or oxidation by Fenton’s hydroxyl radicals. As for hyperoxaluria, glyoxylate was more cytotoxic than oxalate presumably because of the formation of condensation product oxalomalate causing mitochondrial toxicity and oxidative stress. Oxalate toxicity likely involved pro-oxidant iron complex formation.

Indiana University-Purdue University Indianapolis (IUPUI) / A burgeoning area of focus within liver disease research is centered on the concomitant muscle atrophy present in end stage liver disease patients which shows a correlation to severity of hepatic fibrosis and transplant survival outcomes. Of particular interest, nonalcoholic steatohepatitis (NASH) is a form of liver disease that is characterized as the hepatic manifestation of metabolic syndrome. If left untreated, the disease can progress to the state of cirrhosis and hepatocellular carcinoma requiring transplant. Concordant with increasing global prevalence of obesity, NASH is projected to become the leading cause for liver transplants by 2020. Due to a lack of therapeutic options, these patients represent a large unmet medical need in the western world. A major hurdle to therapeutic research is the lack of a quick, reproducible, and cost effective in vivo model that recapitulates the plethora of pathologies and their molecular underpinnings manifested by this disorder. Our studies attempted to validate and expand upon a two-hit model of NASH, which incorporated both the integral comorbidities associated with metabolic challenges of obesity along with liver injury. The two-hit model manifests not only the hepatic morphohistological characteristics of the disease, but also incorporates the obligatory muscle atrophy. To further elaborate on the potential direct link between liver and skeletal muscle and remove any confounding issues associated with the model, in vitro administration of hepatotoxins representing various pathologies associated with liver disease, were used to recapitulate the liver-muscle endocrine signaling that exists in vivo. Our data shows that a variety of hepatoxins can elicit hepatocellular damage which releases factors that inhibits myotube size in vitro. The two hit model also preserves many of conserved molecular underpinnings observed in clinical hepatic fibrosis. Of particular interest, the TGFβ superfamily has been demonstrated to play an important regulatory role in the progression of fibrosis in NASH patients. TGFβ, Activin A, and Follistatin are members of the highly conserved family that are increased in NASH patients. Furthermore, these proteins have a well-studied role in muscle health, regeneration, and mass that has been hypothesized to be conserved between liver and muscle tissues. Surprisingly, novel expression of the myokine and negative regulator of muscle mass Gdf8 (myostatin) was increased in our in vivo model as well. Our studies focused on the molecular interactions of these TGFβ superfamily members and their role on liver disease progression. Through specific inhibition of these proteins (Activin A and Gdf8), we demonstrated that they appear to play key individual roles in the progression of the concomitant muscle atrophy observed in NASH patients. Interestingly, superior efficacy was gained with the treatment of a pan inhibitor of these proteins (Activin A, B, Gdf8 etc.) via a soluble decoy receptor (ActRIIB-Fc), suggesting an additional unaccounted for ligand. Activin B, was found to be increased in two separate in vivo models of liver fibrosis (two-hit model and BDL), has been implicated in regulating muscle mass. Our data suggest a pivotal role for several members of the TGFβ superfamily in NASH associated muscle atrophy. Therapies designed to treat liver fibrosis and the resultant decrements in muscle mass and force must account for these agents which will require pan inhibition of TGFβ superfamily ligands that signal through the ActRIIB receptor.

Nonalcoholic steatohepatitis (NASH) has been identified as a source of significant inter individual variation in drug metabolism. A previous ex vivo study demonstrated significant changes in hepatic Cytochrome P450 (CYP) activity in human NASH. This study evaluated the in vivo activities of multiple CYP isoforms simultaneously in prominent diabetic NASH mouse models. The pharmacokinetics of CYP selective substrates: caffeine, losartan, and omeprazole changed significantly in a diabetic NASH mouse model, indicating attenuation of the activity of Cyp1a2 and Cyp2c29, respectively. Decreased mRNA expression of Cyp1a2 and Cyp2c29, as well as an overall decrease in CYP protein expression, was found in the diabetic NASH mice. Overall, these data suggest that the diabetic NASH model only partially recapitulates the human ex vivo CYP alteration pattern. Therefore, in vivo determination of the effects of NASH on CYP activity should be conducted in human, and more appropriate models are required for future drug metabolism studies in NASH.

Correct dosing in pharmacotherapeutics is based on the idea that too much of a drug will cause toxicity, while too little will result in failure to elicit the desired response. A major factor in the ability of a patient to handle any dose of a drug is the capacity to metabolize and eliminate that drug from the body. For the vast majority of drugs, the liver plays a key role in determining the rate at which drugs are eliminated. First, drugs must be taken up across the cell membrane into hepatocytes by uptake transporters. Once inside the hepatocyte, biotransformation enzymes metabolize and conjugate the drug to a more water-soluble compound, the distribution of which is more easily controlled. These water-soluble metabolites are then transported out of the hepatocyte by additional drug transporters either into bile for elimination, or back into the blood.More than 2 million severe adverse drug reactions occur in the US each year and often result from interindividual variation in the ability to metabolize and eliminate drugs. This number does not include medical errors, but rather circumstances where an individual is unable to handle the standard dose of the correctly prescribed drug. Although genetics plays an important role, the greatest source of variation comes from other environmental factors such as disease states. Nonalcoholic fatty liver disease (NAFLD) is a chronic condition that comprises a spectrum of histopathologies that range from simple steatosis to the more severe steatohepatitis. Specifically, nonalcoholic steatohepatitis (NASH) has become one of the leading causes for liver transplantation in the United States, and thus clearly become a considerable burden to the U.S. healthcare system.It is not known whether the capacity of the liver to metabolize and excrete drugs is altered in patients with NASH. Because the liver plays such a critical role in drug metabolism and disposition, any disease state that disrupts or modifies these functions will alter the fate of a given drug within the body. It is therefore very likely that the ability of the liver to metabolize and excrete clinically relevant drugs is compromised in NASH patients.

Background/objectives The relationship between nonalcoholic fatty liver disease (NAFLD) and albuminuria has been shown in many epidemiologic studies, although the results were inconsistent. This meta-analysis was conducted to summarize all available data and to estimate the risk of albuminuria among patients with NAFLD. Methods Comprehensive literature review was conducted utilizing Medline and Embase database through January 2018 to identify studies that compared the risk of albuminuria among patients with NAFLD versus those without NAFLD. Effect estimates from each study were extracted and combined using the random-effect, generic inverse variance method of DerSimonian and Laird. Results Nineteen studies (17 cross-sectional studies and two cohort studies) with 24 804 participants fulfilled the eligibility criteria and were included in this meta-analysis. The risk of albuminuria among patients with NAFLD was significantly higher than those without NAFLD with the pooled odds ratio (OR) of 1.67 [95% confidence interval (CI): 1.32-2.11]. Subgroup analysis demonstrated the significantly increased risk of albuminuria among patients with NAFLD without diabetes with pooled OR of 2.25 (95% CI: 1.65-3.06). However, we found no significant association between albuminuria and NAFLD among diabetic patients [pooled OR 1.28 (95% CI: 0.94-1.75)]. Conclusion A significantly increased risk of albuminuria among patients with NAFLD was observed in this meta-analysis. Physicians should pay more attention to the early detection and subsequent treatment of individuals with microalbuminuria especially in patients with NAFLD.

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第18147号 / 医博第3867号 / 新制||医||1002(附属図書館) / 31005 / 京都大学大学院医学研究科医学専攻 / (主査)教授 川口 義弥, 教授 坂井 義治, 教授 千葉 勉 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DGAM

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The body encounters an innumerable amount of foreign substances, termed xenobiotics, which it must remove in order to prevent damage to cells and organs. This system of removal is a collection of processes known as ADME (absorption, distribution, metabolism, and excretion). The dynamics of ADME ultimately determine the fate, or pharmacokinetics, of a xenobiotic in the body whether it be an administered pharmaceutical or a potentially harmful toxicant. The major cellular effectors of ADME are the drug metabolizing enzymes (DMEs) and transporters. DMEs function to transform xenobiotics into a metabolite that is more suitable for excretion, whereas drug transporters serve a two-fold function. They may facilitate the uptake of the xenobiotic into the cell so that it can be acted upon by DMEs, or they may function to actively secrete xenobiotics and metabolites from the cell, encouraging their removal from the body. Any perturbations in the expression or function of these critical cellular effectors can result in the diminished therapeutic effect of a pharmaceutical via accelerated removal from the body, or increased toxicity of a pharmaceutical or toxicant due to retention in the body and increased exposure.Perturbations in the ADME processes may result in adverse drug reactions (ADRs) which are an unintended response to a pharmaceutical when administered at the recommended dose. In the last reporting year, the USFDA documented 471,291 serious ADRs causing hospitalization or permanent disabilities, of which 82,724 resulted in death. ADRs can be categorized as two types: dose-related ADRs, and those that are generally unpredictable and mostly occur in susceptible individuals. The major factors that make a person susceptible to ADRs are genetics and disease; however, genetics account for only a small proportion. This dissertation is focused on the contribution of an environmentally-derived component, particularly liver disease, to the occurrence of ADRs. Nonalcoholic fatty liver disease (NAFLD) is the most common liver disease of industrialized nations. It represents a spectrum of damage progressing to the severe stage of nonalcoholic steatohepatitis (NASH), and is closely related to obesity and type 2 diabetes. The following studies have determined the effect of NAFLD and NASH on DMEs and transporters, and demonstrated the propensity for NASH to result in serious ADRs.