Akitaka Hisatomi

Visceral fat accumulation and insulin resistance are important factors in nonalcoholic fatty liver disease

BackgroundNonalcoholic fatty liver diseases are often associated with obesity, insulin resistance, and excessive visceral fat accumulation. The aims of this study were (1) to evaluate the relationship between the severity of fatty liver and visceral fat accumulation in nonalcoholic fatty liver diseases, and (2) to investigate the relationships of fatty liver with biochemical data and insulin resistance.MethodsOne hundred twenty-nine subjects (63 women) with fatty liver diagnosed by ultrasonography were enrolled. Subjects positive for hepatitis B virus, hepatitis C virus, or autoimmune antibodies and those whose alcohol intake was over 20 g/day were excluded. The visceral fat area at the umbilical level and the liver–spleen ratio were evaluated by computed tomography.ResultsThe severity of fatty liver evaluated by ultrasonography showed a significant positive relationship with the visceral fat area and waist circumstance (fatty liver severity: mild, 92.0 ± 30.9 cm2; moderate, 122.1 ± 32.6 cm2; severe, 161.0 ± 48.4 cm2; P < 0.0001). The visceral fat area and liver–spleen ratio were negatively correlated (r = −0.605, P < 0.0001). The severity of fatty liver showed strong positive relationships with serum aspartate aminotransferase, alanine aminotransferase, fasting plasma glucose, fasting plasma insulin, and insulin resistance. The severity of fatty liver was positively related to the visceral fat area in 49 nonobese subjects (body mass index <25).ConclusionsThe severity of fatty liver was positively correlated with visceral fat accumulation and insulin resistance in both obese and nonobese subjects, suggesting that hepatic fat infiltration in nonalcoholic fatty liver disease may be influenced by visceral fat accumulation regardless of body mass index.

Hepatocyte growth factor induces collagenase (matrix metalloproteinase-1) via the transcription factor Ets-1 in human hepatic stellate cell line

BACKGROUND/AIMS Although hepatocyte growth factor recently has been shown to decrease hepatic fibrosis in animal models, the molecular mechanisms of this effects remain to be elucidated. We investigated regulation of collagenase expression by hepatocyte growth factor in hepatic stellate cells. METHODS A human hepatic stellate cell line, LI90, was treated with hepatocyte growth factor. Expression of collagenase, 72 kDa gelatinase, procollagen alpha 1(I), tissue inhibitor of matrix metalloproteinase-1, transforming growth factor-beta 1, or Ets-1, and carboxyterminal telopeptide of type I collagen was examined. Ets-1 binding activity was determined by gel mobility shift assay, collagenase promoter activity was evaluated by reporter gene assay. LI90 cells were also transfected with Ets-1 antisense oligonucleotides with or without hepatocyte growth factor. RESULTS Hepatocyte growth factor increased expression of collagenase mRNA and protein, and an increase in Ets-1 mRNA preceded the increase in collagenase mRNA. Collagenase activity and protein, and a degradation product of type I collagen were increased in the medium. Nuclear extracts from treated LI90 cells also showed increased Ets-1 binding activity. Hepatocyte growth factor and cotransfection of Ets-1 enhanced promoter activity of collagenase gene. Furthermore, treatment of LI90 cells with Ets-1 antisense oligonucleotides downregulated basal and hepatocyte growth factor-induced Ets-1 and collagenase mRNA expression. CONCLUSIONS Collectively, the results suggest that hepatocyte growth factor increases collagenase expression in hepatic stellate cells via the Ets-1 transcription factor-dependent manner.

Gln27Glu β2-adrenergic receptor variant is associated with hypertriglyceridemia and the development of fatty liver

BACKGROUND Nonalcoholic steatohepatitis (NASH) is associated with the metabolism of lipid, glucose and energy. Beta-adrenergic receptors play an important role in the regulation of energy expenditure, in part, by stimulating lipid mobilization through lipolysis. METHODS To assess whether it is common for the beta2-adrenergic receptor (B2AR) gene polymorphisms in codons 16 and 27 to play a role in the development of fatty liver, we investigated 251 unrelated healthy Japanese males who were drug-free and showed no signs of heavy drinking. RESULTS The allelic frequency of B2AR gene mutation in codons 16 and 27 did not differ between obese subjects (BMI>25.0 kg/m(2), n=151) and non-obese subjects (BMI</=25.0 kg/m(2), n=100). The Gly16 homozygotes had a lower high-density lipoprotein cholesterol (HDL-C) level than the Arg16 homozygotes (1.50+/-0.4 vs. 1.32+/-0.3 mmol/l, p=0.014). However, no significant association with fatty liver was observed in the Gly16 allele frequency. The Gln27Glu27 heterozygotes showed higher concentrations of serum triglycerides (TG) than the Gln27Gln27 homozygotes (1.62+/-0.93 vs. 2.21+/-1.67 mmol/l, p=0.013). This correlation was also observed in all subjects regardless of weight classification. Univariate analysis indicated that subjects with the heterozygous Gln27Glu mutant alleles had a significantly higher prevalence of fatty liver vs. those without the mutation (Glu27 allele frequency, 0.07 vs. 0.12, p=0.047; odds ratio, 1.92; 95% confidence interval, 1.01-3.68). However, multivariate logistic regression models showed the prevalence of fatty liver to be significantly related to the homeostasis model assessment (HOMA) index, BMI, triglyceride and HDL-cholesterol. CONCLUSIONS These results suggest that the amino-terminal polymorphisms of the beta2-adrenergic receptor gene in codon 27 were associated with hypertryglyceridemia and independent of obesity, and thereby could be involved in the molecular pathogenesis of fatty liver.

Insulin resistance by unprocessed insulin proreceptors point mutation at the cleavage site

Failure to cleave the interconnecting site between alpha- and beta-subunit produced insulin proreceptors in the plasma membranes which had markedly low affinity to insulin, leading to extreme insulin resistance in a patient. We performed cDNA sequence analysis of the cleavage site of the insulin proreceptor from the patient. Polymerase chain reaction was used to obtain large amount of cDNA coding for the region including the interconnecting site. A thermostable DNA polymerase, Taq polymerase, successfully produced enough amount of cDNA of the region to be sequenced. The results showed AGG (Arg) to AGT (Ser) point mutation, resulting in the change of interconnecting sequence of the two subunits from -Arg-Lys-Arg-Arg- to -Arg-Lys-Arg-Ser-. These results suggest that the tertial structure change of the cleavage site leads to production of unprocessed insulin proreceptors.

Eradication of hepatitis C virus by interferon improves whole-body insulin resistance and hyperinsulinaemia in patients with chronic hepatitis C.

Background/Aims: To investigate whether eradication of hepatitis C virus (HCV) by interferon (IFN) therapy influences systemic glucose metabolism.

Transforming growth factor‐β1‐induced apoptosis is blocked by β1‐integrin‐mediated mitogen‐activated protein kinase activation in human hepatoma cells

Growth factors and extracellular matrices cooperatively regulate cellular behavior. However, the interactions between transforming growth factor‐β1 (TGF‐β1) and integrins in hepatic cells are not fully understood. We investigated the effects of β1‐integrin on TGF‐β1‐regulated growth of hepatoma cells. Human hepatoma cell lines HepG2, Huh7, and Hep3B were stably transfected with β1‐integrin, and the parental, and mock‐ and β1‐integrin‐transfected hepatoma cells were treated with TGF‐β1. Modulation of apoptosis and pathways involved in the process were investigated. TGF‐β1 suppressed the growth of hepatoma cells, and apoptosis was observed in Hep3B and Huh7. Hepatoma cells transfected with β1‐integrin were protected from TGF‐β1‐induced apoptosis. Mitogen‐activated protein (MAP) kinase inhibitors, PD98059, SB203580, and SP600125, abolished this protective effect of β1‐integrin, but herbimycin A and wortmannin were ineffective. Hepatoma cells overexpressing β1‐integrin showed increased activities of MAP kinases, and TGF‐β1 induced sustained activation of MAP kinases in these cells, but only transient activation in mock‐transfected cells. These data suggest that MAP kinases activated by β1‐integrin provide a strong anti‐apoptotic signal during TGF‐β1‐induced apoptosis in human hepatoma cells. Therefore β1‐integrin‐mediated signals may contribute to the development and progression of hepatocellular carcinoma.

Hepatitis C virus infection enhances insulin resistance induced by visceral fat accumulation

Background/Aims: To clarify the impact of visceral obesity on hepatitis C virus (HCV)‐infected patients, we examined the relationship between insulin resistance development and visceral fat accumulation.

Induction of multiple matrix metalloproteinase genes in human hepatocellular carcinoma by hepatocyte growth factor via a transcription factor Ets-1

Matrix metalloproteinases (MMPs) have been implicated in progression of hepatocellular carcinoma (HCC), as have hepatocyte growth factor (HGF) and its c-Met receptor. We investigated regulation of MMP gene expression by HGF in human HCC. Expression of mRNAs encoding MMPs, HGF and c-Met receptor was examined by quantitative reverse transcription-polymerase chain reaction (RT-PCR) in human HCC and in five human HCC cell lines. HCC cells were treated with HGF, and mRNA expression for MMPs and Ets-1 which activates transcription of MMPs was investigated. Ets binding activity was determined by gel mobility shift assay. MMP promoter activities were evaluated by reporter gene assay. Effects of Ets-1 antisense oligonucleotides were also examined. At the mRNA level, MMP-1, -3, -7 as well as c-Met were overexpressed in HCC compared with corresponding nonneoplastic liver tissues, although MMP-2, -9 or HGF were not. HGF dose-dependently induced Ets-1 together with an increased Ets binding activity, followed by transcription of MMP-1, -3, and -7. HGF increased MMP promoter activity, as did cotransfection with Ets-1. Ets-1 antisense oligonucleotide transfection down-regulated the MMP expression, and abolished induction by HGF. In conclusion, certain MMPs and c-Met, overexpressed in HCC cells, are induced by HGF via Ets-1. This pathway may contribute to tumor progression.

Association of the Pro90Ser CD36 mutation with elevated free fatty acid concentrations but not with insulin resistance syndrome in Japanese

BACKGROUND CD36 deficiency is reportedly an underlying factor about insulin resistance, defective fatty acid metabolism and hypertriglyceridemia in spontaneously hypertensive rat (SHR), and may be involved in the pathogenesis of insulin resistance and hyperlipidemia in humans. METHODS We examined 831 adults undergoing health screening. The majority (780) was Pro90 homozygous for the CD36 gene product, but 51 displayed a CD36 mutation (2 homozygous and 49 heterozygous for Ser90). This is the major mutation site involved in CD36 deficiency in Japanese. RESULTS Among parameters related to insulin resistance, there were no differences in body mass index (BMI), HDL cholesterol, total cholesterol, triglycerides, insulin and insulin resistance index (HOMA IR), or blood pressure between 91 normal subjects (45 male and 46 female) randomly selected from the 780 Pro90 homozygotes and the 51 (29 male and 22 females) CD36-deficient subjects (Ser90 homozygote and Pro90Ser heterozygote). Free fatty acid concentrations, however, were higher in Ser90 CD36 subjects than in Pro90 control subjects. CONCLUSIONS The CD36Pro90Ser mutation is not necessarily related to the insulin resistance syndrome, but is associated with high free fatty acid concentrations in Japanese.

Higher arterial stiffness, greater peripheral vascular resistance and lower blood flow in lower-leg arteries are associated with long-term hyperglycaemia in type 2 diabetic patients with normal ankle-brachial index.

Higher arterial stiffness and greater peripheral vascular resistance reduce blood flow in lower‐leg arteries and contribute to the development of ischaemic limb in diabetic patients even without peripheral artery occlusive disease. The aim of this study was to clarify whether these vascular parameters are associated with long‐term hyperglycaemia in diabetic patients.