Masahide Hamaguchi

The Metabolic Syndrome as a Predictor of Nonalcoholic Fatty Liver Disease

Context The metabolic syndrome is often present in patients with nonalcoholic fatty liver disease (NAFLD), but no one knows whether it precedes NAFLD. Content At baseline, 812 members of a cohort of 4401 apparently healthy Japanese adults had NAFLD on abdominal ultrasonography. In 1 year, the authors identified 308 new cases, and NAFLD had resolved in 113 participants. Participants with the metabolic syndrome were much more likely to develop NAFLD and were less likely to experience disease resolution. Limitations Abdominal ultrasonography is not a perfect gold standard test for NAFLD. Implication The metabolic syndrome appears to predispose people to develop NAFLD. The Editors Nonalcoholic fatty liver disease is increasingly recognized as a major cause of liver-related morbidity and mortality (1-3). Because of its potential to progress to cirrhosis and liver failure (4), interest in this disease is increasing among researchers and clinicians in the relevant basic and clinical science fields. The pathologic picture of nonalcoholic fatty liver disease, ranging from simple steatosis to steatohepatitis, advanced fibrosis, and cirrhosis, resembles that of alcohol-induced liver disease, but it occurs in patients who do not abuse alcohol (3). Nonalcoholic steatohepatitis that is characterized by hepatic steatosis and liver cell injury, hepatic inflammation, and fibrosis and necrosis is believed to be an intermediate stage of nonalcoholic fatty liver disease. (1) This disease is often associated with obesity (5), type 2 diabetes mellitus (6, 7), dyslipidemia (8), and hypertension (9). Each of these abnormalities carries a cardiovascular disease risk, and together they are often categorized as the insulin resistance syndrome or the metabolic syndrome (10). The third report of the National Cholesterol Education Program Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults (Adult Treatment Panel III [ATP III]) (11) recommended the use of 5 variables for diagnosing the metabolic syndrome, namely waist circumference, serum triglyceride level, serum high-density lipoprotein (HDL) cholesterol level, blood pressure, and fasting plasma glucose level. As stated above, the frequent association of nonalcoholic fatty liver disease with individual components of the metabolic syndrome is now well known. However, it is unknown whether the risk for this disease is increased in patients with the metabolic syndrome. This is important because the metabolic syndrome is an emerging problem worldwide and its prevalence is likely increasing (12). The current study was designed first to evaluate the cross-sectional relationship between the metabolic syndrome, defined by the modified ATP III criteria, and the prevalence of nonalcoholic fatty liver disease in Japanese persons. Second, and more important, we addressed longitudinal aspects of the disease and its development and regression and tried to clarify the role of the metabolic syndrome in its pathogenesis. Despite serious concern about a possible future epidemic of this disease in the Asia-Pacific region (13), information regarding a possible association with the metabolic syndrome in Asia is very limited; our study may have special clinical relevance for people who live in this part of the world. Methods Study Participants We designed a prospective cohort study to investigate the role of the metabolic syndrome in the pathogenesis of nonalcoholic fatty liver disease in participants in a medical health checkup program at Murakami Memorial Hospital, Gifu, Japan. Each participant had abdominal ultrasonography. The purpose of the medical health checkup program is to promote public health through early detection of chronic diseases and their risk factors. Medical service of this kind, known as a human dock, is very popular in Japan. The center at which the checkups were performed was founded in 1994 and currently evaluates more than 8000 examinees annually. Of these examinees, 60% repeatedly have annual or biannual examinations and 40% are newly registered examinees. Most of the participants were employees of various companies and local governmental organizations in Gifu, Japan, and their spouses. These companies and organizations recruit employees each year according to a contract with our center. The cost of the medical examination was largely paid for by the employers. Fewer than 10% of the participants individually registered for the program and paid for it themselves, and they are citizens of local communities. Because many participants were expected to have repeated examinations, we took advantage of this opportunity to conduct a follow-up study on nonalcoholic fatty liver disease by using abdominal ultrasonography. The ethics committee of Murakami Memorial Hospital approved the study. All participants who were examined in the health checkup programs between January and December 2001 were invited to enroll in the study. Data Collection and Measurements The health checkup programs included the following: urinalysis, blood cell counts, blood chemistry, measurements of hepatitis B antigen and hepatitis C antibody, electrocardiography, chest radiography, barium examination of the upper gastrointestinal tract, and abdominal ultrasonography. Medical history and lifestyle factors, including physical activity, habits regarding smoking, and habits regarding alcohol consumption, were surveyed by using a self-administered questionnaire. When the participants had difficulty completing the questionnaire, trained nurses provided assistance. Smoking status was expressed by using the Brinkman index, which is calculated as the number of cigarettes smoked per day multiplied by the number of years that the participant smoked. Habits regarding alcohol consumption were evaluated by asking the participants about the amount and type of alcoholic beverages consumed per week, then estimating the mean ethanol intake per day. The diagnosis of fatty liver was based on the results of abdominal ultrasonography, which was done by trained technicians. All ultrasonographic images were stored in the image server. One gastroenterologist reviewed the images and made the diagnosis of fatty liver without reference to any of the participants other individual data. Of 4 known criteria (hepatorenal echo contrast, liver brightness, deep attenuation, and vascular blurring) (14), the participants were required to have hepatorenal contrast and liver brightness to be given a diagnosis of nonalcoholic fatty liver disease. Body mass index (BMI) was calculated as body weight in kilograms divided by the square of the participants height in meters. The ATP III proposed the following 5 abnormalities to define the metabolic syndrome: 1) abdominal obesity (abdominal circumference> 102 cm for men and> 88 cm for women); 2) elevated serum triglyceride level (1.70 mmol/L [150 mg/dL]); 3) decreased HDL cholesterol level (<1.04 mmol/L [<40 mg/dL] for men and <1.30 mmol/L [<50 mg/dL] for women); 4) elevated blood pressure (systolic and diastolic blood pressure 130/85 mm Hg); and 5) an elevated fasting glucose level (6.11 mmol/L (110 mg/dL]). Because waist measurements were not available for the entire study sample, we substituted a BMI of 25 kg/m2 or greater for all participants as an index of obesity. A BMI of 25 kg/m2 or greater has been proposed as a cutoff for the diagnosis of obesity in Asian people (15). Individuals with 3 or more of the 5 abnormalities were considered to have the metabolic syndrome. Exclusion criteria were an alcohol intake of more than 20 g/d, known liver disease, or current use of medication. Regarding liver disease, participants who tested positive for hepatitis B antigen or hepatitis C antibody and those who reported a history of known liver disease, including viral, genetic, autoimmune, and drug-induced liver disease, were also excluded (16). Statistical Analysis The SPSS statistical package, version 11.0.1 J (SPSS, Inc., Chicago, Illinois) was used for all statistical analyses, and a P value less than 0.05 was considered statistically significant. Because the incidence rate of nonalcoholic fatty liver disease was unknown, a formal sample size estimate was not made a priori. Participants with and without follow-up visits were compared to determine the appropriateness of an analysis based on participants with complete data only. Two groups of participants were compared by using the unpaired t-test and the chi-square test. Logistic regression was used to analyze associations between the development and regression of nonalcoholic fatty liver disease and the metabolic syndrome while controlling for potential confounders. The potential confounders were selected from clinical variables, which were different between participants with and without the disease at baseline. As will be described later, weight change was also selected as a confounder because the development and regression of nonalcoholic fatty liver disease generally occurred with weight changes. Unadjusted and adjusted odds ratios and 95% CIs were calculated. Data are expressed as means and SDs for continuous variables. Role of the Funding Source No funding was received for this study. Results Between January and December 2001, we invited 8056 participants in the health checkup program to enroll in the study. A total of 6654 Japanese participants (4601 men and 2053 women) gave informed consent to be included in the study. We excluded 290 participants (216 men and 74 women) who had known liver disease. In addition, 1657 participants (1577 men and 80 women) who consumed more than 20 g of ethanol per day and 306 participants (236 men and 70 women) who were currently receiving medication were excluded. As a result, there were 4401 participants (2572 men and 1829 women). Mean age and BMI were 47.6 years (SD, 8.8) (range, 21 to 80 years) and 22.6 kg/m2 (SD, 3.0) (range, 14.2 to 38.1 kg/m2), respectively. By the end of June 2003, 387

The Severity of Ultrasonographic Findings in Nonalcoholic Fatty Liver Disease Reflects the Metabolic Syndrome and Visceral Fat Accumulation

BACKGROUND:Nonalcoholic fatty liver disease (NAFLD) is closely associated with the metabolic syndrome.AIM:We evaluated the association among the metabolic syndrome, visceral fat accumulation, and the severity of fatty liver with a new scoring system of ultrasonographic findings in apparently healthy Japanese adults.METHODS:Subjects consisted of 94 patients who received liver biopsy and 4,826 participants who were selected from the general population. Two hepatologists scored the ultrasonographic findings from 0 to 6 points. We calculated Cohens kappa of within-observer reliability and between-observer reliability. We evaluated the predictive value of the score by the area under a conventional receiver operating characteristic curve (AUC).RESULTS:Within-observer reliability was 0.95 (95% CI 0.93–0.97, P < 0.001) and between-observer reliability was 0.95 (95% CI 0.93–0.97, P < 0.001). The AUC to diagnose NAFLD was 0.980. The sensitivity was 91.7% (95% CI 87.0–95.1, P < 0.001) and the specificity was 100% (95% CI 95.4–100.0, P < 0.001). The AUC to diagnose visceral obesity was 0.821. The sensitivity was 68.3% (95% CI 51.9–81.9, P = 0.028) and the specificity was 95.1% (95% CI 86.3–99.0, P < 0.001). Adjusted odds ratio of the score for the metabolic syndrome was 1.37 (95% CI 1.26–1.49, P < 0.001).CONCLUSIONS:The scoring system with abdominal ultrasonography could provide accurate information about hepatic steatosis, visceral obesity, and the metabolic syndrome in apparently healthy people who do not consume alcohol.

T Cell Receptor Stimulation-Induced Epigenetic Changes and Foxp3 Expression Are Independent and Complementary Events Required for Treg Cell Development

The transcription factor Foxp3 is essential for the development of regulatory T (Treg) cells, yet its expression is insufficient for establishing the Treg cell lineage. Here we showed that Treg cell development was achieved by the combination of two independent processes, i.e., the expression of Foxp3 and the establishment of Treg cell-specific CpG hypomethylation pattern. Both events were induced by T cell receptor stimulation. The Treg cell-type CpG hypomethylation began in the thymus and continued to proceed in the periphery and could be fully established without Foxp3. The hypomethylation was required for Foxp3(+) T cells to acquire Treg cell-type gene expression, lineage stability, and full suppressive activity. Thus, those T cells in which the two events have concurrently occurred are developmentally set into the Treg cell lineage. This model explains how Treg cell fate and plasticity is controlled and can be exploited to generate functionally stable Treg cells.

Two FOXP3+CD4+ T cell subpopulations distinctly control the prognosis of colorectal cancers

CD4+ T cells that express the forkhead box P3 (FOXP3) transcription factor function as regulatory T (Treg) cells and hinder effective immune responses against cancer cells. Abundant Treg cell infiltration into tumors is associated with poor clinical outcomes in various types of cancers. However, the role of Treg cells is controversial in colorectal cancers (CRCs), in which FOXP3+ T cell infiltration indicated better prognosis in some studies. Here we show that CRCs, which are commonly infiltrated by suppression-competent FOXP3hi Treg cells, can be classified into two types by the degree of additional infiltration of FOXP3lo nonsuppressive T cells. The latter, which are distinguished from FOXP3+ Treg cells by non-expression of the naive T cell marker CD45RA and instability of FOXP3, secreted inflammatory cytokines. Indeed, CRCs with abundant infiltration of FOXP3lo T cells showed significantly better prognosis than those with predominantly FOXP3hi Treg cell infiltration. Development of such inflammatory FOXP3lo non-Treg cells may depend on secretion of interleukin (IL)-12 and transforming growth factor (TGF)-β by tissues and their presence was correlated with tumor invasion by intestinal bacteria, especially Fusobacterium nucleatum. Thus, functionally distinct subpopulations of tumor-infiltrating FOXP3+ T cells contribute in opposing ways to determining CRC prognosis. Depletion of FOXP3hi Treg cells from tumor tissues, which would augment antitumor immunity, could thus be used as an effective treatment strategy for CRCs and other cancers, whereas strategies that locally increase the population of FOXP3lo non-Treg cells could be used to suppress or prevent tumor formation.

Protective effect of alcohol consumption for fatty liver but not metabolic syndrome.

AIM To investigate the effect of alcohol on the metabolic syndrome (MS) and fatty liver in Japanese men and women. METHODS A cross-sectional study was conducted in a medical health checkup program at a general hospital. This study involved 18 571 Japanese men and women, 18-88 years of age, with a mean body mass index of 22.6 kg/m(2). A standardized questionnaire was administered. The total amount of alcohol consumed per week was calculated, and categorized into four grades. Fatty liver was examined by ultrasound modified criteria of the revised National Cholesterol Education Program Adult Treatment Panel III and the new International Diabetes Federation. RESULTS The prevalence of fatty liver decreased in men and women with light to moderate alcohol consumption, whereas the prevalence of MS was not so changed. The prevalence of fatty liver of any grade in men was lower than that in those with no or minimal alcohol consumption. In women with light to moderate alcohol consumption, prevalence of fatty liver was lower than that in women with no or minimal alcohol consumption. By logistic regression analysis, the odds ratio (OR) for MS in women with light alcohol consumption was decreased to < 1.0, but this change was not clear in men. The OR for fatty liver was clearly < 1.0 in men with any level of alcohol consumption and in women with light to moderate consumption. CONCLUSION Light to moderate alcohol consumption has a favorable effect for fatty liver, but not for MS in Japanese men and women.

Detection of self-reactive CD8+ T cells with an anergic phenotype in healthy individuals

Immunological tolerance to self requires naturally occurring regulatory T (Treg) cells. Yet how they stably control autoimmune T cells remains obscure. Here, we show that Treg cells can render self-reactive human CD8+ T cells anergic (i.e., hypoproliferative and cytokine hypoproducing upon antigen restimulation) in vitro, likely by controlling the costimulatory function of antigen-presenting cells. Anergic T cells were naïve in phenotype, lower than activated T cells in T cell receptor affinity for cognate antigen, and expressed several coinhibitory molecules, including cytotoxic T lymphocyte–associated antigen-4 (CTLA-4). Using these criteria, we detected in healthy individuals anergic T cells reactive with a skin antigen targeted in the autoimmune disease vitiligo. Collectively, our results suggest that Treg cell–mediated induction of anergy in autoimmune T cells is important for maintaining self-tolerance. Healthy individuals harbor “silenced” self-reactive T cells. For the immune system, silence is golden For the immune system, balance is key. Immune cells must learn to eliminate invading pathogens but tolerate self. A cell type called regulatory T cells (Tregs) help to maintain this balance, but how they do so, particularly in humans, is unclear. Maeda et al. now report that Tregs “silence” T cells with modest reactivity to self. After culture with Tregs, the silenced T cells proliferated very little and produced almost no cytokines in response to antigen. The authors then examined T cells from healthy donors and from people with an autoimmune disease. Only healthy donors harbored silenced T cells, suggesting that if silencing goes awry, autoimmunity may result. Science, this issue p. 1536

A Critical Role for Allograft Inflammatory Factor-1 in the Pathogenesis of Rheumatoid Arthritis

Rheumatoid arthritis (RA) is characterized by massive synovial proliferation, angiogenesis, subintimal infiltration of inflammatory cells and the production of cytokines such as TNF-α and IL-6. Allograft inflammatory factor-1 (AIF-1) has been identified in chronic rejection of rat cardiac allografts as well as tissue inflammation in various autoimmune diseases. AIF-1 is thought to play an important role in chronic immune inflammatory processes, especially those involving macrophages. In the current work, we examined the expression of AIF-1 in synovial tissues and measured AIF-1 in synovial fluid (SF) derived from patients with either RA or osteoarthritis (OA). We also examined the proliferation of synovial cells and induction of IL-6 following AIF-1 stimulation. Immunohistochemical staining showed that AIF-1 was strongly expressed in infiltrating mononuclear cells and synovial fibroblasts in RA compared with OA. Western blot analysis and semiquantitative RT-PCR analysis demonstrated that synovial expression of AIF-1 in RA was significantly greater than the expression in OA. AIF-1 induced the proliferation of cultured synovial cells in a dose-dependent manner and increased the IL-6 production of synovial fibroblasts and PBMC. The levels of AIF-1 protein were higher in synovial fluid from patients with RA compared with patients with OA (p < 0.05). Furthermore, the concentration of AIF-1 significantly correlated with the IL-6 concentration (r = 0.618, p < 0.01). These findings suggest that AIF-1 is closely associated with the pathogenesis of RA and is a novel member of the cytokine network involved in the immunological processes underlying RA.

Metabolically Healthy Obesity and Risk of Incident CKD

BACKGROUND AND OBJECTIVES Metabolically healthy obesity (MHO) is a unique obesity phenotype that apparently protects people from the metabolic complications of obesity. The association between MHO phenotype and incident CKD is unclear. Thus, this study investigated the association between MHO phenotype and incident CKD. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS A total of 3136 Japanese participants were enrolled in an 8-year follow-up cohort study in 2001. Metabolically healthy status was assessed by common clinical markers: BP, triglycerides, HDL cholesterol, and fasting plasma glucose concentrations. Body mass index ≥25.0 kg/m(2) was defined as obesity. CKD was defined by proteinuria or eGFR of <60 ml/min per 1.73 m(2). To calculate the odds ratio for incident CKD, logistic regression analyses were performed. RESULTS The crude incidence proportions of CKD were 2.6% (56 of 2122 participants) in participants with the metabolically healthy nonobesity phenotype, 2.6% (8 of 302) in those with the MHO phenotype, 6.7% (30 of 445) in those with the metabolically abnormal nonobesity phenotype, and 10.9% (29 of 267) in those with the metabolically abnormal obesity phenotype. Compared with metabolically healthy nonobesity phenotype, the odds ratios for incident CKD were 0.83 (95% confidence interval [95% CI], 0.36 to 1.72; P=0.64) for MHO, 1.44 (95% CI, 0.80 to 2.57; P=0.22) for metabolically abnormal nonobesity, and 2.80 (95% CI, 1.45 to 5.35; P=0.02) for metabolically abnormal obesity phenotype after adjustment for confounders, including age, sex, smoking statues, alcohol use, creatinine, uric acid, systolic BP, HDL cholesterol, and impaired fasting glucose or diabetes. CONCLUSION MHO phenotype was not associated with higher risk of incident CKD.

Aging is a risk factor of nonalcoholic fatty liver disease in premenopausal women

AIM To clarify the relationship between age, menopause, and nonalcoholic fatty liver disease (NAFLD) in women. METHODS We conducted a follow-up study on nonalcoholic fatty liver disease by using abdominal ultrasonography, and investigated the relationship of age and menopause with the development of NAFLD in women. We followed 1829 women and 2572 men (response rate, 86%) selected in 2001 to represent the non-institutionalized adult population of Gifu, Japan. Data collected included self-reported medical history, lifestyle factors, and menopausal status. The postmenopausal state was defined as beginning 1 year after the cessation of menses. We diagnosed NAFLD with the aid of abdominal ultrasonography by using diagnostic criteria described previously. RESULTS The prevalence of NAFLD in women increases with age, but does not alter with age in men. Furthermore, the prevalence of NAFLD in premenopausal women (6%) was lower than that in men (24%) and in postmenopausal women (15%). The associations of the postmenopausal state and hormone replacement therapy with NAFLD were statistically significant in a univariate logistic regression model. At the follow-up examination, 67 women (5%) were newly diagnosed with NAFLD. The incidence of NAFLD was 3.5% (28/802) in premenopausal women, 7.5% (4/53) in menopausal women, 6.1% (24/392) in postmenopausal women, and 5.3% (11/206) in women receiving hormone replacement therapy. The weight gain in premenopausal women was equal to that in postmenopausal women. Metabolic syndrome and weight gain were independent risk factors for NAFLD in pre- and postmenopausal women, but age was an independent risk factor in premenopausal women only. CONCLUSION Aging is a risk factor for NAFLD in premenopausal women, independent of weight gain or influence of metabolic syndrome.

FOXP3+ regulatory T cells: control of FOXP3 expression by pharmacological agents

Naturally arising CD4(+)CD25(+) regulatory T cells (Tregs), which specifically express the forkhead family transcription factor FOXP3, are essential for the maintenance of immunological self-tolerance and immune homeostasis. Tregs can suppress the activation, proliferation and effector function of other lymphocytes in physiological and pathological immune responses. Therefore, control of the development, survival, and function of Tregs is instrumental for effective control of immune responses. For example, cytokines such as interleukin-2 and transforming growth factor-β, monoclonal antibodies to the Treg-associated molecules such as interleukin-2 receptor α chain and cytotoxic T lymphocyte-associated 4, and pharmacological agents that alter signaling pathways for Treg function, can augment or dampen the suppressive activity of Tregs. How these agents control the function of Tregs at the molecular level remains to be elucidated. However, it is envisaged that pharmacological control of the function and development of Tregs by targeting FOXP3 or Treg-associated molecules will enable better control of immune responses in various clinical settings.