Yoshihiko Sugioka

Glycyrrhizin and its metabolite inhibit Smad3-mediated type I collagen gene transcription and suppress experimental murine liver fibrosis.

AIMS Glycyrrhizin has been widely used for the treatment of chronic hepatitis C. It decreases the serum levels of aminotransferases, and suppresses progression of liver fibrosis as well as subsequent occurrence of hepatocellular carcinoma. Although previous studies have shown that glycyrrhizin and its metabolite inhibit collagen gene expression, its underlying mechanisms are virtually unknown. This study was aimed to explore molecular mechanisms responsible for the inhibitory effect of glycyrrhizin on type I collagen gene transcription. MAIN METHODS Effects of glycyrrhizin and its metabolite, glycyrrhetinic acid, on collagen promoter activity were examined by using transgenic reporter mice harboring alpha2(I) collagen gene (COL1A2) promoter. Their effects on the TGF-beta/Smad signaling pathway were studied by cell transfection assays and immunofluorescence studies using cultured hepatic stellate cells. KEY FINDINGS Administration of glycyrrhizin or its metabolite, glycyrrhetinic acid, significantly suppressed COL1A2 promoter activation and progression of liver fibrosis induced by repeated carbon tetrachloride injections. In cultured hepatic stellate cells, glycyrrhetinic acid, but not glycyrrhizin, inhibited type I collagen synthesis mostly at the level of gene transcription. This inhibitory effect of glycyrrhetinic acid was abolished by a mutation introduced into a Smad3-binding region within the COL1A2 promoter. Glycyrrhetinic acid did not affect gene expression of TGF-beta receptors or Smad proteins, but inhibited nuclear accumulation of Smad3 in activated hepatic stellate cells. In addition to those direct inhibitory effects on COL1A2 transcription, glycyrrhetinic acid also suppressed activation of quiescent hepatic stellate cells in primary culture. SIGNIFICANCE The results provide a molecular basis for the anti-fibrotic effect of glycyrrhizin treatment.

Matrix metalloproteinase-13 promotes recovery from experimental liver cirrhosis in rats.

Objective: To evaluate the role of matrix metalloproteinase (MMP)-13 gene expression in the early phase of recovery from liver fibrosis/cirrhosis. Methods: Liver fibrosis was induced in male Wistar rats by administration of carbon tetrachloride (CCl4) for 10 weeks. Recombinant adenovirus-mediated human MMP-13 gene transfer (RAdMMP-13) was performed via the femoral vein on day 3 after the last CCl4 injection. The role of MMP-13 in stably expressing cell lines was also analyzed. Results: Fibrous deposition in the liver was decreased in RAdMMP-13-injected rats by day 3 after gene transfer compared with empty vector RAd66-injected rats. Furthermore, MMP-2 and MMP-9 enzymatic activity was markedly enhanced in the liver of RAdMMP-13 injected rats. Hepatocyte growth factor (HGF) induction was also increased in RAdMMP-13 injected rats. In established stable HT-1080 cells transfected with MMP-13, HGF-α expression and MMP-2 and MMP-9 enzymatic activity were increased. The conversion of precursor HGF into mature HGF was also increased in the MMP-13 expressing cell lines. Conclusion: Forced MMP-13 expression effectively accelerated recovery from liver cirrhosis via the effects of MMP-13-mediated HGF, MMP-2, and MMP-9 expression, which induced the degradation of collagen fibers and promoted hepatic regeneration.

c-jun NH2-terminal kinase pathway is involved in constitutive matrix metalloproteinase-1 expression in a hepatocellular carcinoma-derived cell line

Transcription factor c‐Jun serves for cellular proliferation, survival, differentiation and transformation and is recognized as an important factor in cancer development, including hepatocellular carcinoma (HCC). The purpose of present study is to determine the involvement of c‐Jun in matrix metalloproteinase‐1 (MMP‐1) expression, which is previously reported by us to be expressed only in the early stage of human HCC showing stromal invasion. Of 5 human HCC cell lines examined, only HLE cells revealed mRNA and protein expression as well as enzymatic activity of MMP‐1. Transient transfection of an MMP‐1 promoter/luciferase construct (including 4.4 kb full promoter region) into HLE and HCC‐T cells (MMP‐1 nonproducer) showed that high promoter activity was observed only in HLE cells without inducers, and that this promoter activity was still observed when a shorter 0.6 kb proximal promoter construct was transfected. The 0.6 kb promoter region contained 3 AP‐1 sites, and c‐jun mRNA was constitutively expressed in HLE cells without inducers. Furthermore, phosphorylated c‐Jun and c‐Jun NH2‐terminal kinase (JNK) were detected in HLE cells. Promoter activity of the 0.6 kb construct was suppressed with SP600125, a potent inhibitor of JNK, but not with PD98059 and SB203580, potent inhibitors of MEK1/2 and p38, respectively. The inhibitory effect of SP600125 was also observed at protein expression level and in enzymatic activity of MMP‐1. Taken together, this study suggests that the JNK pathway is involved in the expression of MMP‐1 in HCC cells and may represent a new functional role of c‐Jun for HCC development.

A Novel Role of the NRF2 Transcription Factor in the Regulation of Arsenite-Mediated Keratin 16 Gene Expression in Human Keratinocytes

Background Inorganic sodium arsenite (iAs) is a ubiquitous environmental contaminant and is associated with an increased risk of skin hyperkeratosis and cancer. Objectives We investigated the molecular mechanisms underlying the regulation of the keratin 16 (K16) gene by iAs in the human keratinocyte cell line HaCaT. Methods We performed reverse transcriptase polymerase chain reaction, luciferase assays, Western blots, and electrophoretic mobility shift assays to determine the transcriptional regulation of the K16 gene by iAs. We used gene overexpression approaches to elucidate the nuclear factor erythroid-derived 2 related factor 2 (NRF2) involved in the K16 induction. Results iAs induced the mRNA and protein expression of K16. We also found that the expression of K16 was transcriptionally induced by iAs through activator protein-1–like sites and an antioxidant response element (ARE) in its gene promoter region. Treatment with iAs also enhanced the production and translocation of the NRF2 transcription factor, an ARE-binding protein, into the nucleus without modification of its mRNA expression. In addition, iAs elongated the half-life of the NRF2 protein. When overexpressed in HaCaT cells, NRF2 was also directly involved in not only the up-regulation of the detoxification gene thioredoxin but also K16 gene expression. Conclusions Our data clearly indicate that the K16 gene is a novel target of NRF2. Furthermore, our findings also suggest that NRF2 has opposing roles in the cell—in the activation of detoxification pathways and in promoting the development of skin disorders.

Aldehyde dehydrogenase 2 and β3-adrenergic receptor gene polymorphisms: their association with elevated liver enzymes and metabolic syndrome

Recent studies indicate that some patients with nonalcoholic fatty liver have ongoing liver injury that may progress from steatosis to steatohepatitis or fibrosis. The present study was designed to clarify the clinical features of liver dysfunction observed in the course of workplace physical check-ups in relation to multiple risk factor syndrome including obesity, hyperlipidemia, hypertension, and impaired glucose tolerance, and to clarify the involvement of aldehyde dehydrogenase 2 (ALDH2) and beta(3)-adrenergic receptor (beta3-AR) gene polymorphisms in elevation of liver enzymes. One hundred forty-eight male workers 35 years of age were enrolled. They were requested to answer questionnaires about drinking and smoking habits, and underwent urinalysis, physical and peripheral blood examinations, blood chemistry, electrocardiogram and chest x-rays. The genotypes of ALDH2 and beta3-AR were analyzed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). The subjects were divided into active ALDH2 or inactive ALDH2 groups. They were also divided into 2 groups according to the beta3-AR genotype. The relationships between ALDH2 and beta3-AR gene polymorphism and the results of the physical examination including liver function tests were analyzed. The subjects were also divided according to the number of components of metabolic syndrome. The prevalence of elevated alanine aminotransferase (ALT) level increased with the accumulation of components of metabolic syndrome. Active ALDH2 was associated with elevated ALT level to a greater degree than beta3-AR polymorphism. Among those with normal body mass index (BMI), the genotypes of ALDH2 and beta3-AR were strongly associated with elevated ALT level. Logistic regression analysis revealed that BMI, triglyceride level, and ALDH2 genotype were associated with ALT elevation. In conclusion, evaluating the genotype of ALDH2 and beta3-AR may assist in predicting and preventing the development of fatty liver which may be related to multiple risk factor syndrome.

Relation of dampness to sick building syndrome in Japanese public apartment houses

ObjectivesThe effect of dampness on sick building syndrome (SBS) symptoms has not been fully investigated in Japan. The purpose of this study is to elucidate the possible effects of dampness on SBS symptoms among residents in Japanese public apartment houses.MethodsA questionnaire was used to investigate the degree of dampness in public apartment houses in Asahikawa, Japan, and its effect on SBS symptoms, involving 480 residents in 64 buildings. Dampness indicators were as follows: condensation on the windowpanes, condensation on the walls and/or closets, visible mold in the bathrooms, visible mold on the walls, window frames, and/or closet, moldy odor, slow drying of the wet towels in bathrooms, water leakage, and bad drainage in bathrooms. ResultsAll dampness indicators except for visible mold in bathrooms had significantly higher odds ratios (ORs) for all or any SBS symptoms after adjustment. The dampness index, the number of positive dampness indicators, was significantly related to all SBS symptoms after adjustment.ConclusionsThere are serious problems relating to dampness in Japanese public housing, which affects the health of residents. There is a need to educate the residents about the relationship between dampness and SBS, and building problems should be rectified.

Dampness, food habits, and sick building syndrome symptoms in elementary school pupils

ObjectivesWe investigated dampness/mold in schools and dwellings, and food habits and subjective symptoms in elementary school pupils, in order to clarify the effect of dampness and food habits on subjective symptoms in elementary school pupils.MethodsQuestionnaires were used to investigate dampness in classrooms and dwellings in Hokkaido, Japan, and its effect on subjective symptoms in 1,077 pupils in 8 elementary schools. We used a dampness index for both the home and classroom; the index was the sum of the presence of four dampness indicators: (1) visible mold, (2) moldy odor, (3) water leakage, and (4) condensation on windowpanes. The questionnaire also contained queries about food habits, as follows: the frequency of eating breakfast, whether the energy provided by the school lunch was sufficient, and whether eating too many snacks and/or sweets were consumed. Adjusted logistic regression was used to determine whether dampness and food habits were related to the subjective symptoms.ResultsIn fully adjusted models, the home dampness index was significantly related to cough, general symptoms, and having at least one symptom; the classroom dampness index was significantly related to nasal symptoms. In addition, usually not eating breakfast was significantly related to eye symptoms, and too many snacks and/or sweets was significantly related to eye, nasal, and general symptoms.ConclusionsBoth home and classroom dampness can affect pupils’ health. Home dampness, in particular, was significantly related to cough and general symptoms, and classroom dampness was significantly related to nasal symptoms. Furthermore, favorable food habits have a positive effect on pupils’ subjective symptoms.

Job stress and burnout among urban and rural hospital physicians in Japan

OBJECTIVE To elucidate the differences in job stress and burnout status of Japanese hospital physicians between large cities, small cities, and towns and villages. DESIGN Cross-sectional study. SETTING Postal self-administered questionnaires were distributed to 2937 alumni of Asahikawa Medical University. PARTICIPANTS Four hundred and twenty-two hospital physicians. MAIN OUTCOME MEASURES The Brief Job Stress Questionnaire was used to evaluate job demand, job control and social support. The Japanese version of the Maslach Burnout Inventory-General Survey (MBI-GS) was used to evaluate burnout. An analysis of covariance was conducted on the mean scores on the Brief Job Stress Questionnaire and the MBI-GS scales after adjusting for sex, age and specialties. RESULTS In adjusted analyses, the job demand score was significantly different among physicians in the three areas. In Bonferroni post-hoc tests, scores in large cities was significantly higher than those in small cities and towns and villages. The job control score showed a significant difference and a marginally significant trend, with large cities associated with lower job control. There were significant differences in support from supervisors and that from family/friends, and scores in large cities was significantly higher than those in small cities in the post-hoc test. There was a significant effect on the exhaustion scale of the MBI-GS, with large cities associated with higher exhaustion, and scores in large cities was significantly higher than those in small cities. CONCLUSIONS Urban hospital physicians had more job demand, less job control and exhaustion caused by burnout, and rural hospital physicians had less social support.

Prevalence of Burnout among Public Health Nurses in Charge of Mental Health Services and Emergency Care Systems in Japan

ObjectivesThe Community Health Act came into effect in 1997 in Japan. This act altered the work system for public health nurses (PHNs) in public health centers (PHCs) nationwide from region-specific to service-specific work. Such major changes to working environment in the new system seem to be exposing PHNs to various types of stress. The present study examined whether prevalence of burnout is higher among PHNs in charge of mental health services (psychiatric PHNs) than among PHNs in charge of other services (non-psychiatric PHNs), and whether attributes of emergency mental health care systems in communities are associated with increased prevalence of burnout.MethodsA questionnaire including the Pines burnout scale for measuring burnout was mailed to 525 psychiatric PHNs and 525 non-psychiatric PHNs. The 785 respondents included in the final analysis comprised 396 psychiatric PHNs and 389 non-psychiatric PHNs.ResultsPrevalence of burnout was significantly higher for psychiatric PHNs (59.2%) than for non-psychiatric PHNs (51.5%). When prevalence of burnout in each group was analyzed in relation to question responses regarding emergency service and patient referral systems, prevalence of burnout for psychiatric PHNs displayed significant correlations to frequency of cases requiring overtime emergency services, difficulties referring patients, and a feeling of “restriction”.ConclusionsPrevalence of burnout is high among psychiatric PHNs, and inadequate emergency mental health service systems contribute to burnout among these nurses. Countermeasures for preventing such burnout should be taken as soon as possible.

Effects of work burden, job strain and support on depressive symptoms and burnout among Japanese physicians

ObjectivesDays off, on call, night duty, working hours and job stress can affect physicians’ mental health, and support from supervisors and co-workers may have a buffering effect. This study elucidates whether job strain and job factors affect physicians’ mental health, and whether support from supervisors and co-workers has a protective effect on their mental health.Material and MethodsThe subjects included 494 physicians. The Brief Job Stress Questionnaire (BJSQ) was used to evaluate job demand, job control and support. High job strain was defined as a combination of high job demand and low job control. Depressive symptoms were assessed using the Patient Health Questionnaire-9. The Maslach Burnout Inventory-General Survey was used to evaluate burnout. Possible confounder adjusted logistic regression analyses were performed to obtain odds ratios for depressive symptoms and burnout.ResultsAs per the analysis, high job strain had significantly higher odds ratios, and support from co-workers had significant protective odds ratios for depressive symptoms. High job strain and having only 2–4 days off per month (compared to > 8 days off per month) had significantly higher odds ratios, and support from co-workers had significant protective odds ratios for burnout.ConclusionsHigh job strain was related to depressive symptoms and burnout, and support from co-workers had a buffering effect on depressive symptoms and burnout. An inadequate number of days off was related to burnout. Assessment of job strain may be a good tool to measure physicians’ mental health, and a sufficient number of days off may be needed to prevent burnout.