Long-term exposure to diesel exhaust particles induces concordant changes in DNA methylation and transcriptome in human adenocarcinoma alveolar basal epithelial cells.

BACKGROUND: Diesel exhaust particles (DEP), which contain hazardous compounds, are emitted during the combustion of diesel. As approximately one-third of the vehicles worldwide use diesel, there are growing concerns about the risks posed by DEP to human health. Long-term exposure to DEP is associated with airway hyperresponsiveness, pulmonary fibrosis, and inflammation; however, the molecular mechanisms behind the effects of DEP on the respiratory tract are poorly understood. Such mechanisms can be addressed by examining transcriptional and DNA methylation changes. Although several studies have focused on the effects of short-term DEP exposure on gene expression, research on the transcriptional effects and genome-wide DNA methylation changes caused by long-term DEP exposure is lacking. Hence, in this study, we investigated transcriptional and DNA methylation changes in human adenocarcinoma alveolar basal epithelial A549 cells caused by prolonged exposure to DEP and determined whether these changes are concordant. RESULTS: DNA methylation analysis using the Illumina Infinium MethylationEPIC BeadChips showed that the methylation levels of DEP-affected CpG sites in A549 cells changed in a dose-dependent manner; the extent of change increased with increasing dose reaching the statistical significance only in samples exposed to 30 µg/ml DEP. Four-week exposure to 30 µg/ml of DEP significantly induced DNA hypomethylation at 24,464 CpG sites, which were significantly enriched for DNase hypersensitive sites, genomic regions marked by H3K4me1 and H3K27ac, and several transcription factor binding sites. In contrast, 9,436 CpG sites with increased DNA methylation levels were significantly overrepresented in genomic regions marked by H3K27me3 as well as H3K4me1 and H3K27ac. In parallel, gene expression profiling by RNA sequencing demonstrated that long-term exposure to DEP altered the expression levels of 2,410 genes, enriching 16 gene sets including Xenobiotic metabolism, Inflammatory response, and Senescence. In silico analysis revealed that the expression levels of 854 genes correlated with the methylation levels of the DEP-affected cis-CpG sites. CONCLUSIONS: To our knowledge, this is the first report of genome-wide transcriptional and DNA methylation changes and their associations in A549 cells following long-term exposure to DEP.

RHEB is a potential therapeutic target in T cell acute lymphoblastic leukemia.

T cell acute lymphoblastic leukemia (T-ALL) is an aggressive malignancy of immature T lymphocytes. Although various therapeutic approaches have been developed, refractoriness of chemotherapy and relapse cause a poor prognosis of the disease and further therapeutic strategies are required. Here, we report that Ras homolog enriched in brain (RHEB), a critical regulator of mTOR complex 1 activity, is a potential target for T-ALL therapy. In this study, we established an sgRNA library that comprehensively targeted mTOR upstream and downstream pathways, including autophagy. CRISPR/Cas9 dropout screening revealed critical roles of mTOR-related molecules in T-ALL cell survival. Among the regulators, we focused on RHEB because we previously found that it is dispensable for normal hematopoiesis in mice. Transcriptome and metabolic analyses revealed that RHEB deficiency suppressed de novo nucleotide biosynthesis, leading to human T-ALL cell death. Importantly, RHEB deficiency suppressed tumor growth in both mouse and xenograft models. Our data provide a potential strategy for efficient therapy of T-ALL by RHEB-specific inhibition.

A Pilot Study to Assess Glucose, Insulin, and Incretin Responses Following Novel High Resistant Starch Rice Ingestion in Healthy Men.

INTRODUCTION: A newly developed resistant starch (RS) rice line with double mutation of starch synthase IIIa and branching enzyme IIb (ss3a/be2b) exhibits a tenfold greater percentage RS value than the wild-type rice line. Currently, the effects of cooked rice with such high RS content on secretion and action of glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) are unclear. Therefore, we conducted a pilot study to assess postprandial responses of GLP-1 and GIP along with glucose and insulin and also gastric emptying after ingestion of the high-RS cooked rice with ss3a/be2b in healthy subjects. METHODS: In a non-randomized crossover design, five healthy men ingested two test foods, control (low-RS) and high-RS cooked rice, with at least 1-week washout period between testing days. Plasma glucose, serum insulin, plasma total GLP-1, plasma total GIP, and also gastric emptying rate were measured after ingestion of each test food, and the incremental area under the curves (iAUC) was calculated for each biochemical parameter using the values from 0 to 180 min after ingestion. RESULTS: The high-RS cooked rice ingestion tended to reduce iAUC-glucose (p = 0.06) and significantly reduced iAUC-insulin (p < 0.01) and iAUC-GLP-1 (p < 0.05) but not iAUC-GIP (p = 0.21) relative to control cooked rice ingestion. In addition, the high-RS cooked rice ingestion did not affect gastric emptying. CONCLUSIONS: The present results indicate that the suppressive effects of the high-RS cooked rice ingestion on postprandial responses of glucose and insulin may be provided through attenuation in GLP-1 secretion along with its low digestibility into glucose. We suggest that the high-RS rice with ss3a/be2b may serve as a better carbohydrate source and also as a novel functional food for dietary interventions to improve postprandial hyperglycemia and hyperinsulinemia without both enhancing GLP-1 secretion and affecting gastric emptying in patients with diabetes.

Genome-wide Survival Analysis for Macular Neovascularization Development in Central Serous Chorioretinopathy Revealed Shared Genetic Susceptibility with Polypoidal Choroidal Vasculopathy.

PURPOSE: To identify susceptibility genes for macular neovascularization (MNV) development in central serous chorioretinopathy (CSC). DESIGN: Genome-wide survival analysis using a longitudinal cohort study. PARTICIPANTS: We included 402 and 137 patients with CSC but without MNV at their first visit from the Kyoto CSC Cohort and Kobe CSC dataset, respectively. All patients underwent detailed ophthalmic examinations, including multimodal imaging, such as fundus autofluorescence, spectral-domain OCT, and fluorescein angiography/indocyanine green angiography or OCT angiography. METHODS: We conducted a genome-wide survival analysis using the Kyoto CSC Cohort. We applied the Cox proportional hazard model to adjust for age, sex, and the first principal component. Single nucleotide polymorphisms (SNPs) with P values < 1.0 × 10-5 were carried forward to the replication in the Kobe CSC dataset. Moreover, we evaluated the contribution of previously reported age-related macular degeneration (AMD) susceptibility loci. We used FUMA and ToppFun for the functional enrichment analysis. MAIN OUTCOME MEASURES: The association between SNPs and MNV development in patients with CSC. RESULTS: Rs370974631 near ARMS2 displayed a genome-wide significant association in the meta-analysis of discovery and replication result (hazard ratio [HR]meta, 3.63; Pmeta = 5.76 × 10-9). Among previously reported AMD susceptibility loci, we additionally identified CFH rs800292 (HR, 0.39, P = 2.55 × 10-4), COL4A3 rs4276018 (HR, 0.26, P = 1.56 × 10-3), and B3GALTL rs9564692 (HR, 0.56, P = 8.30 × 10-3) as susceptibility loci for MNV development in CSC. The functional enrichment analysis revealed significant enrichment of 8 pathways (GO:0051561, GO:0036444, GO:0008282, GO:1990246, GO:0015272, GO:0030955, GO:0031420, and GO:0005242) related to ion transport. CONCLUSIONS: ARMS2, CFH, COL4A3, and B3GALTL were identified as susceptibility genes for MNV development in CSC. These 4 genes are known as susceptibility genes for AMD, whereas COL4A3 and B3GALTL were previously reported to be polypoidal choroidal vasculopathy (PCV)-specific susceptibility genes. Our findings revealed the shared genetic susceptibility between PCV and MNV secondary to CSC.

Polygenic risk scores for low-density lipoprotein cholesterol and familial hypercholesterolemia.

Familial hypercholesterolemia (FH) is an autosomal dominant monogenic disorder characterized by elevated levels of low-density lipoprotein cholesterol (LDL-C) and an increased risk of premature coronary artery disease (CAD). Recently, it has been shown that a high polygenic risk score (PRS) could be an independent risk factor for CAD in FH patients of European ancestry. However, it is uncertain whether PRS is also useful for risk stratification of FH patients in East Asia. We recruited and genotyped clinically diagnosed FH (CDFH) patients from the Kanazawa University Mendelian Disease FH registry and controls from the Shikamachi Health Improvement Practice genome cohort in Japan. We calculated PRS from 3.6 million variants of each participant (imputed from the 1000 Genome phase 3 Asian dataset) for LDL-C (PRSLDLC) using a genome-wide association study summary statistic from the BioBank Japan Project. We assessed the association of PRSLDLC with LDL-C and CAD among and within monogenic FH, mutation negative CDFH, and controls. We tested a total of 1223 participants (376 FH patients, including 173 with monogenic FH and 203 with mutation negative CDFH, and 847 controls) for the analyses. PRSLDLC was significantly higher in mutation negative CDFH patients than in controls (p = 3.1 × 10-13). PRSLDLC was also significantly linked to LDL-C in controls (p trend = 3.6 × 10-4) but not in FH patients. Moreover, we could not detect any association between PRSLDLC and CAD in any of the groups. In conclusion, mutation negative CDFH patients demonstrated significantly higher PRSLDLC than controls. However, PRSLDLC may have little additional effect on LDL-C and CAD among FH patients.

Effects of GLP-1 receptor agonist on changes in the gut bacterium and the underlying mechanisms.

There is a close relationship between the gut microbiota and metabolic disorders. In this study, acute administration of the glucagon-like peptide-1 receptor agonist (GLP-1RA) liraglutide to mice increased the cecal levels of caseinolytic protease B, a component of Escherichia coli, and of norepinephrine. Chemical sympathectomy blocked these events. Norepinephrine was found to pass into the intestinal lumen in vitro. c-Fos staining of the intermediolateral nucleus was identified as indirect evidence of sympathetic nervous system activation of the intestinal tract by GLP-1RA. Under normal conditions, the increase in E. coli did not affect the host. However, in mice with colitis, bacterial translocation was observed with attenuation of tight junction gene expression. This is the first study to investigate the unique underlying mechanisms related the effects of GLP-1RA on changes in the gut bacterium.

ASK1 suppresses NK cell-mediated intravascular tumor cell clearance in lung metastasis.

Tumor metastasis is the leading cause of death worldwide and involves an extremely complex process composed of multiple steps. Our previous study demonstrated that apoptosis signal-regulating kinase 1 (ASK1) deficiency in mice attenuates tumor metastasis in an experimental lung metastasis model. However, the steps of tumor metastasis regulated by ASK1 remain unclear. Here, we showed that ASK1 deficiency in mice promotes natural killer (NK) cell-mediated intravascular tumor cell clearance in the initial hours of metastasis. In response to tumor inoculation, ASK1 deficiency upregulated immune response-related genes, including interferon-gamma (IFNγ). We also revealed that NK cells are required for these anti-metastatic phenotypes. ASK1 deficiency augmented cytokine production chemoattractive to NK cells possibly through induction of the ligand for NKG2D, a key activating receptor of NK cells, leading to further recruitment of NK cells into the lung. These results indicate that ASK1 negatively regulates NK cell-dependent anti-tumor immunity and that ASK1-targeted therapy can provide a new tool for cancer immunotherapy to overcome tumor metastasis.

The solubility of N-acetyl amino acid amides in organic acid and alcohol solutions: Mechanistic insight into structural protein solubilization.

Structural proteins such as spider silk and silkworm silk are generally poorly soluble in aqueous and organic solutions, making them difficult to manipulate in manufacturing processes. Although some organic acids and alcohols, such as formic acid and hexafluoroisopropanol (HFIP), effectively solubilize poorly soluble proteins, little is known about their protein solubilization mechanism. In this study, the solubility of N-acetyl amino acid amide compounds in organic solvents-formic acid, acetic acid, HFIP and isopropanol-was measured to clarify the protein solubilization mechanism at the amino acid residue level. On the basis of thermodynamic analyses of the solubility in terms of the transfer free energy (from water to organic solvents), every organic solvent was found to be effective in thermodynamically stabilizing hydrophobic amino acid side chains in the liquid phase. Formic acid and HFIP were comparably effective in the stabilization of the polypeptide backbone, whereas acetic acid and isopropanol were ineffective. Therefore, the significant solubilizing effect of formic acid and HFIP on the structural proteins was attributed to their favorable interactions with hydrophobic amino acid side chains and with the polypeptide backbone of the proteins. The present findings are useful for the optimization of protein manipulation and amino acid sequence design.

Inhibition of GIP signaling extends lifespan without caloric restriction.

AIMS/INTRODUCTION: Caloric restriction (CR) promotes longevity and exerts anti-aging effects by increasing Sirtuin production and activation. Gastric inhibitory polypeptide (GIP), a gastrointestinal peptide hormone, exerts various effects on pancreatic ß-cells and extra-pancreatic tissues. GIP promotes glucose-dependent augmentation of insulin secretion and uptake of nutrients into the adipose tissue. MATERIALS AND METHODS: Gipr-/- and Gipr+/+ mice were used for lifespan analysis, behavior experiments and gene expression of adipose tissue and muscles. 3T3-L1 differentiated adipocytes were used for Sirt1 and Nampt expression followed by treatment with GIP and α-lipoic acid. RESULTS: We observed that GIP receptor-knockout (Gipr-/-) mice fed normal diet showed an extended lifespan, increased exploratory and decreased anxiety-based behaviors, which are characteristic behavioral changes under CR. Moreover, Gipr-/- mice showed increased Sirt1 and Nampt expression in the adipose tissue. GIP suppressed α-lipoic acid-induced Sirt1 expression and activity in differentiated adipocytes. CONCLUSIONS: Although maintenance of CR is difficult, food intake and muscle endurance of Gipr-/- mice were similar to those of wild-type mice. Inhibition of GIP signaling may be a novel strategy to extend the lifespan of diabetic patients.

A new targeted capture method using bacterial artificial chromosome (BAC) libraries as baits for sequencing relatively large genes.

To analyze a specific genome region using next-generation sequencing technologies, the enrichment of DNA libraries with targeted capture methods has been standardized. For enrichment of mitochondrial genome, a previous study developed an original targeted capture method that use baits constructed from long-range polymerase chain reaction (PCR) amplicons, common laboratory reagents, and equipment. In this study, a new targeted capture method is presented, that of bacterial artificial chromosome (BAC) double capture (BDC), modifying the previous method, but using BAC libraries as baits for sequencing a relatively large gene. We applied the BDC approach for the 214 kb autosomal region, ring finger protein 213, which is the susceptibility gene of moyamoya disease (MMD). To evaluate the reliability of BDC, cost and data quality were compared with those of a commercial kit. While the ratio of duplicate reads was higher, the cost was less than that of the commercial kit. The data quality was sufficiently the same as that of the kit. Thus, BDC can be an easy, low-cost, and useful method for analyzing individual genome regions with substantial length.

Genome-wide association study identifies ERBB4 on 2q34 as a novel locus associated with sperm motility in Japanese men.

BACKGROUND: The decrease in sperm motility has a potent influence on fertilisation. Sperm motility, represented as the percentage of motile sperm in ejaculated sperms, is influenced by lifestyle habits or environmental factors and by inherited factors. However, genetic factors contributing to individual differences in sperm motility remain unclear. To identify genetic factors that influence human sperm motility, we performed a genome-wide association study (GWAS) of sperm motility. METHODS: A two-stage GWAS was conducted using 811 Japanese men in a discovery stage, followed by a replication study using an additional 779 Japanese men. RESULTS: In the two-staged GWAS, a single nucleotide polymorphism rs3791686 in the intron of gene for erb-b2 receptor tyrosine kinase 4 (ERBB4) on chromosome 2q34 was identified as a novel locus for sperm motility, as evident from the discovery and replication results using meta-analysis (ß=-4.01, combined P=5.40×10-9). CONCLUSIONS: Together with the previous evidence that Sertoli cell-specific Erbb4-knockout mice display an impaired ability to produce motile sperm, this finding provides the first genetic evidence for further investigation of the genome-wide significant association at the ERBB4 locus in larger studies across diverse human populations.

GLP-1 release and vagal afferent activation mediate the beneficial metabolic and chronotherapeutic effects of D-allulose.

Overeating and arrhythmic feeding promote obesity and diabetes. Glucagon-like peptide-1 receptor (GLP-1R) agonists are effective anti-obesity drugs but their use is limited by side effects. Here we show that oral administration of the non-calorie sweetener, rare sugar D-allulose (D-psicose), induces GLP-1 release, activates vagal afferent signaling, reduces food intake and promotes glucose tolerance in healthy and obese-diabetic animal models. Subchronic D-allulose administered at the light period (LP) onset ameliorates LP-specific hyperphagia, visceral obesity, and glucose intolerance. These effects are blunted by vagotomy or pharmacological GLP-1R blockade, and by genetic inactivation of GLP-1R signaling in whole body or selectively in vagal afferents. Our results identify D-allulose as prominent GLP-1 releaser that acts via vagal afferents to restrict feeding and hyperglycemia. Furthermore, when administered in a time-specific manner, chronic D-allulose corrects arrhythmic overeating, obesity and diabetes, suggesting that chronotherapeutic modulation of vagal afferent GLP-1R signaling may aid in treating metabolic disorders.

ASK1 facilitates tumor metastasis through phosphorylation of an ADP receptor P2Y12 in platelets.

Tumor metastasis is the major cause of deaths in cancer patients and is modulated by intertwined stress-responsive signaling cascades. Here we demonstrate that deletion of stress-responsive apoptosis signal-regulating kinase 1 (Ask1) in platelets results in unstable hemostasis and drastic attenuation of tumor lung metastasis, both of which are attributable to platelet dysfunction. Platelet-specific deletion of Ask1 in mice leads to defects in ADP-dependent platelet aggregation, unstable hemostasis and subsequent attenuation of tumor metastasis. We also revealed that activating phosphorylation of Akt is attenuated in Ask1-deficient platelets, contrary to the previous reports suggesting that Akt is negatively regulated by ASK1. Mechanistically, ASK1-JNK/p38 axis phosphorylates an ADP receptor P2Y12 at Thr345, which is required for the ADP-dependent sustained Akt activity that is vital to normal platelet functions. Our findings offer insight into positive regulation of Akt signaling through P2Y12 phosphorylation as well as MAPK signaling in platelets by ASK1 and suggest that ASK1-JNK/p38 axis provides a new therapeutic opportunity for tumor metastasis.

Food Intake Affects Sperm-Egg Fusion Through the GIP/PSG17 Axis in Mice.

In addition to overeating, starvation also reduces fecundity in mammals. However, little is known about the molecular mechanisms linking food intake to fertility, especially in males. Gastric inhibitory polypeptide (GIP), which is released from intestinal K-cells after meal ingestion, stimulates insulin secretion from pancreatic ß-cells through the action of incretin and has several extrapancreatic effects. Here, we identified GIP receptor (Gipr) expression in mouse spermatids. Microarray analysis revealed that pregnancy-specific glycoprotein 17 (Psg17), a potential CD9-binding partner, was significantly decreased in GIP receptor-knockout (Gipr-/-) testes. Glycosylphosphatidylinositol-anchored PSG17 was expressed on the surface of acrosome-reacted sperm, and Gipr-/- sperm led to a lower fertilization rate in vitro, compared with that of Gipr+/+ sperm, both in the absence and presence of the zona pellucida. Plasma GIP concentrations and Psg17 messenger RNA (mRNA) were immediately increased in the testis after a single meal, whereas ingestion of a chronic high-fat diet markedly decreased Gipr and Psg17 mRNA. These results suggest that reduced GIP signaling, by decreased GIP levels or the downregulation of Gipr, is associated with the reduction of fecundity due to starvation or overeating. Thus, proper regulation of GIP signaling in the testis could be a potential unique therapeutic target for male infertility in obese and diabetic individuals.

Stromal cell-derived factor-1 is upregulated by dipeptidyl peptidase-4 inhibition and has protective roles in progressive diabetic nephropathy.

The role of stromal cell-derived factor-1 (SDF-1) in the pathogenesis of diabetic nephropathy and its modification by dipeptidyl peptidase-4 (DPP-4) inhibition are uncertain. Therefore, we studied this independent of glucagon-like peptide-1 receptor (GLP-1R) signaling using two Akita diabetic mouse models, the diabetic-resistant C57BL/6-Akita and diabetic-prone KK/Ta-Akita. Increased SDF-1 expression was found in glomerular podocytes and distal nephrons in the diabetic-prone mice, but not in kidneys from diabetic-resistant mice. The DPP-4 inhibitor linagliptin, but not the GLP-1R agonist liraglutide, further augmented renal SDF-1 expression in both Glp1r(+/+) and Glp1r(-/-) diabetic-prone mice. Along with upregulation of renal SDF-1 expression, the progression of albuminuria, glomerulosclerosis, periglomerular fibrosis, podocyte loss, and renal oxidative stress was suppressed in linagliptin-treated Glp1r(+/+) diabetic-prone mice. Linagliptin treatment increased urinary sodium excretion and attenuated the increase in glomerular filtration rate which reflects glomerular hypertension and hyperfiltration. In contrast, selective SDF-1 receptor blockade with AMD3100 reduced urinary sodium excretion and aggravated glomerular hypertension in the Glp1r(+/+) diabetic-prone mice. Thus, DPP-4 inhibition, independent of GLP-1R signaling, contributes to protection of the diabetic kidney through SDF-1-dependent antioxidative and antifibrotic effects and amelioration of adverse renal hemodynamics.

Novel extrapancreatic effects of incretin.

The hormonal factors implicated as transmitters of signals from the gut to pancreatic ß-cells are referred to as incretins. Gastric inhibitory polypeptide (GIP) and glucagon-like peptide-1 (GLP-1) are incretins. In addition to the insulinotropic effects, we have shown, using the GIP receptor and GLP-1 receptor-deficient mice, that GIP and GLP-1 have direct actions on adipocytes and the kidney, respectively. Because GIP receptors and GLP-1 receptors are differentially expressed in a tissue-specific manner, GIP and GLP-1 have specific physiological activities, and further comprehensive characterization of the extrapancreatic actions of GIP and GLP-1 is anticipated, as dipeptidyl peptidase IV inhibitors activate both GIP and GLP-1 signaling.

The protective roles of GLP-1R signaling in diabetic nephropathy: possible mechanism and therapeutic potential.

Glucagon-like peptide-1 (GLP-1) is a gut incretin hormone that has an antioxidative protective effect on various tissues. Here, we determined whether GLP-1 has a role in the pathogenesis of diabetic nephropathy using nephropathy-resistant C57BL/6-Akita and nephropathy-prone KK/Ta-Akita mice. By in situ hybridization, we found the GLP-1 receptor (GLP-1R) expressed in glomerular capillary and vascular walls, but not in tubuli, in the mouse kidney. Next, we generated C57BL/6-Akita Glp1r knockout mice. These mice exhibited higher urinary albumin levels and more advanced mesangial expansion than wild-type C57BL/6-Akita mice, despite comparable levels of hyperglycemia. Increased glomerular superoxide, upregulated renal NAD(P)H oxidase, and reduced renal cAMP and protein kinase A (PKA) activity were noted in the Glp1r knockout C57BL/6-Akita mice. Treatment with the GLP-1R agonist liraglutide suppressed the progression of nephropathy in KK/Ta-Akita mice, as demonstrated by reduced albuminuria and mesangial expansion, decreased levels of glomerular superoxide and renal NAD(P)H oxidase, and elevated renal cAMP and PKA activity. These effects were abolished by an adenylate cyclase inhibitor SQ22536 and a selective PKA inhibitor H-89. Thus, GLP-1 has a crucial role in protection against increased renal oxidative stress under chronic hyperglycemia, by inhibition of NAD(P)H oxidase, a major source of superoxide, and by cAMP-PKA pathway activation.

Apoptosis signal-regulating kinase 1 inhibits hepatocarcinogenesis by controlling the tumor-suppressing function of stress-activated mitogen-activated protein kinase.

UNLABELLED: The stress-activated mitogen-activated protein kinases (MAPKs), c-Jun NH2-terminal kinase (JNK), and p38 have been implicated in hepatocarcinogenesis. Although the many interrelated functions of JNK and p38 are precisely regulated by upstream signaling molecules, little is known about upstream regulators. We investigated the role of apoptosis signal-regulating kinase 1 (ASK1), a major player in the regulation of JNK and p38 activities, in hepatocarcinogenesis using a mouse hepatocellular carcinoma (HCC) model. ASK1-deficient (ASK1(-/-) ) and wildtype (WT) mice were treated with diethylnitrosamine on postnatal day 14. Strikingly, after 7 months, approximately three times as many tumors developed in ASK1(-/-) mice as in WT mice. Although JNK and p38 activation were attenuated in ASK1(-/-) HCCs relative to WT HCCs, cell proliferation was comparable in HCCs from both types of mice. On the other hand, both cancer cell apoptosis and hyperphosphorylation of BimEL, a proapoptotic Bcl-2 family member, were suppressed in the ASK1(-/-) HCCs. ASK1(-/-) mice showed remarkable resistance to Fas-induced hepatocyte apoptosis in vivo, probably because of attenuated JNK-mediated BimEL phosphorylation and mitochondrial apoptotic pathway activation. The reintroduction of ASK1 to ASK1(-/-) mouse liver using an adenoviral vector restored Fas-induced hepatocyte death and phosphorylation of JNK and BimEL. Similar findings were obtained in tumor necrosis factor alpha-induced hepatocyte apoptosis. Furthermore, ASK1 was involved in DNA damage-induced p21 up-regulation through a p38 pathway. CONCLUSION: ASK1 is involved in death receptor-mediated apoptosis and DNA-damage response by way of stress-activated MAPK in the liver, and thus acts as a tumor suppressor in hepatocarcinogenesis. This study provides new insight into the regulation of stress- activated MAPK signaling in hepatocarcinogenesis.

A case of insulinoma following total gastrectomy--effects of an alpha-glucosidase inhibitor on suppressing GIP and GLP-1 elevations.

A 61-year-old woman with fasting hypoglycemia following total gastrectomy was diagnosed as insulinoma. GIP and GLP-1 levels after a mixed meal were extremely increased. Administration of miglitol, an alpha-glucosidase inhibitor, suppressed the GIP and GLP-1 elevations.

Allele frequencies of the ABCC11 gene for earwax phenotypes among ancient populations of Hokkaido, Japan.

Human earwax is classified into wet and dry types, which are determined by a single-nucleotide polymorphism in the adenosine triphosphate-binding cassette, sub-family C11 (ABCC11) gene locus. To investigate the allele frequencies of the ABCC11 locus within ancient populations on the Northern Japanese island of Hokkaido, amplified product-length polymorphisms were analyzed for 50 specimens of the Okhotsk people and 35 specimens of the Jomon and Epi-Jomon people excavated from various archaeological sites of Hokkaido. Of these specimens, 31 Okhotsk and 19 Jomon/Epi-Jomon samples were genotyped successfully. Frequencies of the wet-type allele in the Jomon/Epi-Jomon people, considered a major ancestor of the Ainu, were higher than those of other Northeastern Asian populations, including the modern Ainu. By contrast, in the Okhotsk people, believed to originate from East Siberia, frequencies of the dry-type allele were relatively higher than those in the Ainu and Jomon/Epi-Jomon people. These results suggest that gene flow from the Northeastern Asian Continent to descendants of the Jomon/Epi-Jomon people of Hokkaido through the Okhotsk people occurred, resulting in the establishment of the Ainu.