👤 Kentaro Honda

Visceral fat (VF), particularly epicardial adipose tissue (EAT), plays a crucial role in the development of coronary artery disease (CAD). Cathelicidin (LL37) is an antimicrobial peptide involved in innate immunity and has been implicated in inflammatory processes. However, the relationship between VF accumulation, cathelicidin, and atherosclerosis remains unclear. Seventy-eight subjects without CAD were enrolled and classified by obesity type: normal-weight (normal; n=20), subcutaneous fat (SF; n=19), and VF (n=39). Plasma LL37 concentrations were compared across groups. LL37 expression in EAT was assessed in 9 patients undergoing open-heart surgery, stratified by CAD status. In animal experiments, angiotensin II-infused wild-type and Apoe LL37 is associated with VF accumulation and CAD. However, these findings are exploratory and warrant prospective validation to determine its potential utility as a biomarker. Show less

Because the prognostic value of lipoprotein(a) [Lp(a)] levels in Japanese patients remains unclear, we assessed their distribution and association with long-term outcomes in ST-segment elevation myocardial infarction (STEMI). In our retrospective analysis of 868 consecutive patients with STEMI, the median serum Lp(a) level was 15.75 mg/dL at admission, and the median follow-up was 736.5 days. Using restricted cubic spline analysis, we stratified patients into high (≥47.26 mg/dL) and low (<47.26 mg/dL) Lp(a) groups. The high Lp(a) group had a higher proportion of older and female patients, with lower body weight, estimated glomerular filtration rate, and stent use, and higher dyslipidemia prevalence than those in the low Lp(a) group. The 5-year cumulative incidence of the composite primary endpoint (cardiovascular death, non-fatal myocardial infarction, non-fatal stroke, or any revascularization) was significantly higher in the high Lp(a) group, primarily because of a high rate of any revascularization. Patients with elevated Lp(a) levels demonstrated higher rates of any revascularization for both de novo and restenotic lesions than those with lower levels. After adjusting for confounders, a high Lp(a) level was identified as an independent predictor of the primary endpoint (hazard ratio:1.932; 95% confidence interval:1.255-2.974). In Japanese patients with STEMI, elevated Lp(a) levels were independently associated with worse long-term outcomes. Show less

Estrogen is synthesized throughout various tissues in the body, and its production is regulated by the rate-limiting enzyme aromatase (encoded by the Cyp19a1 gene). Notably, aromatase is also expressed in central nervous system cells, allowing for localized estrogen synthesis in regions such as the hypothalamus. Estrogens produced within these neurons are referred to as neuroestrogens. In this study, we investigated the role of neuroestrogens in the regulation of appetite through modulation of hypothalamic pathways in OVX, ArKO, and aromatase-restored mice. Estrogen suppresses appetite by influencing the expression of appetite-regulating peptides, including POMC and NPY, via MC4R. We explored the direct effects of neuroestrogens, independent from ovarian estrogen, on appetite suppression and the underlying molecular mechanisms. We monitored body weight and food intake and evaluated the expression of Cyp19a1, Mc4r, and other appetite-related genes. Our findings indicate that OVX and ArKO mice exhibited increased body weight and food consumption, which correlated with altered expression of Mc4r and Cyp19a1. Conversely, restoration of Cyp19a1 expression in a neuron specific manner significantly decreased food intake and increased Mc4r expression in the hypothalamus. Furthermore, neuroestrogens enhanced leptin responsiveness. Our results imply that neuroestrogens likely contribute to appetite regulation and may be relevant for body weight reduction. Show less

Type 2 diabetic patients exhibited an increased secretion of triglyceride-rich lipoproteins and low high-density lipoprotein cholesterol levels with a greater amount of small dense low-density lipoprotein (LDL). Given that apolipoprotein B (apoB), a proatherogenic lipoprotein, exists at both triglyceride-rich lipoproteins and LDL particles, circulating apoB may associate with diabetic coronary atherosclerosis. The OPTIMAL study was a prospective randomized-controlled study which employed serial near-infrared spectroscopy (NIRS)/intravascular ultrasound (IVUS) imaging to evaluate the efficacy of glycemic control on coronary atherosclerosis in 94 statin-treated type 2 diabetic patients with coronary artery disease (CAD) (UMIN000036721). Of these, 78 patients with both serial apoB levels and NIRS/IVUS images at baseline and week 48 were analyzed. NIRS/IVUS-derived plaque measures were compared in those with and without any reduction of apoB levels. All of the study subjects received a statin, and 60.6% of the study subjects exhibited any reduction of apoB levels. There was no significant difference in the atheroma progression rate between the 2 groups (-0.27 ± 0.15% vs -0.33 ± 0.51%, P = .44). However, patients with any reduction of apoB levels exhibited a greater frequency of change in maximal lipid-core burden index at 4-mm segment (maxLCBI In statin-treated type 2 diabetic patients with CAD, a greater delipidation of coronary atherosclerosis was observed in association with a reduction of apoB levels. The current findings indicate a potential anti-atherosclerotic effect of lowering apoB levels, which may ultimately mitigate future coronary events risk in statin-treated type 2 diabetic patients with CAD. Show less

Apolipoprotein B (ApoB) is essential for the assembly and secretion of triglyceride (TG)-rich VLDL particles, and its dysfunction is linked to metabolic disorders, including dyslipidemia and liver steatosis. However, less attention has been paid to whether and how other apolipoproteins play redundant or compensatory roles when the ApoB function is compromised. Here, we investigated the effects of microsomal triglyceride transfer protein (MTP), which mediates lipidation of nascent ApoB, on ApoE function. We observed a paradoxical increase in ApoE secretion resulting from increased expression in MTP inhibitor (MTPi)-treated human hepatoma cells. This phenotype was recapitulated in APOB-knockout cells and was associated with impaired ApoB secretion. While MTP-dependent transfer of neutral lipids is dispensable for ApoE secretion, TG biosynthesis, redundantly catalyzed by DGAT1 and DGAT2, is required for efficient ApoE secretion in hepatoma cells. ApoE colocalizes with lipid droplets near the Golgi apparatus and mediates TG export in an ApoB-independent fashion. We found that simultaneous inhibition of both ApoE and ApoB, but not inhibition of either alone, led to TG accumulation in hepatoma cells, indicating that both proteins function redundantly to control TG content. Validation studies in primary human hepatocytes (PHHs) demonstrated DGAT2-dependent secretion of ApoE. While MTPi treatment did not elevate ApoE secretion, it induced increased sialylation of ApoE in the supernatants of PHHs. These results show that enhanced ApoE secretion compensates for the impaired ApoB function to maintain the lipid homeostasis, providing an alternative route to modulate lipid turnover in hepatoma cells. Show less

Aberrant fibroblast growth factor receptor (FGFR)-driven signaling, predominantly arising from FGFR2 amplification, plays a key role in gastric cancer pathogenesis. This open-label, phase 2 study evaluated the efficacy and safety of futibatinib, an irreversible FGFR1-4 inhibitor, in patients with gastric or gastroesophageal junction (GEJ) cancer harboring FGFR2 amplifications. Patients were treated with futibatinib 20 mg orally once daily in a 28-day cycle. The primary endpoint was objective response rate (ORR) per independent central review. Secondary endpoints included progression-free survival (PFS), overall survival (OS), and safety. Among 28 treated patients, the ORR per independent central review was 17.9 %, comprising five patients with a partial response (median duration of response, 3.9 months), and an additional nine patients with stable disease for a disease control rate of 50.0 %. Median PFS per independent central review and median OS were 2.9 and 5.9 months, respectively. The most common treatment-related adverse events (any grade) were hyperphosphatemia (89.3 %), decreased appetite (32.1 %), and increased aspartate aminotransferase (21.4 %). Only one (3.6 %) patient discontinued study treatment due to an adverse event. Futibatinib demonstrated modest antitumor activity with a safety profile consistent with previous reports in patients with gastric or GEJ cancer harboring FGFR2 amplifications, potentially warranting further investigation. Show less

We tested whether elevated 1 h post-load glucose (1hPG) was associated with decreased leg fat (relative to body fat) and serum concentrations of lipoprotein lipase (LPL), a rate-limiting enzyme for lipid storage in subcutaneous leg fat. Body fat mass and distribution, as measured by DXA, surrogate measures of insulin sensitivity, and insulin secretion inferred from serum insulin kinetics during a 75-g oral glucose tolerance test, as well as serum adipokines and LPL, were assessed in 164 Japanese female university students and 94 middle-aged parents. They all had normal glucose tolerance. Students provided their birth weight. Elevated 1hPG was found in 6% of daughters and 22% of parents. Multivariate logistic regression analyses revealed that log insulinogenic index (IGI) (OR: 0.031, 95% CI 0.003-0.30, p = 0.003) and serum LPL (OR: 0.90, 95% CI 0.83-0.98, p = 0.01) were associated with elevated 1hPG independently of serum adiponectin and birth weight in young Japanese. In middle-aged Japanese, the ratio of leg fat to body fat (OR: 0.66, 95% CI 0.44-0.97, p = 0.03), log IGI (OR: 0.002, 95% CI 0.00003-0.07, p = 0.001), and Matsuda index (OR: 0.67, 95% CI 0.47-0.96, p = 0.03) were related to elevated 1hPG independently of fat mass index, the ratio of trunk fat to body fat, LPL, and homeostasis model assessment insulin resistance. A decreased amount of leg fat in middle-aged Japanese with elevated 1hPG may be explained hypothetically by a prolonged deficiency of LPL. Show less

Generation and accumulation of amyloid-β (Aβ) protein in the brain are the primary causes of Alzheimer's disease (AD). Alcadeins (Alcs composed of Alcα, Alcβ and Alcγ family) are a neuronal membrane protein that is subject to proteolytic processing, as is Aβ protein precursor (APP), by APP secretases. Previous observations suggest that Alcs are involved in the pathophysiology of Alzheimer's disease (AD). Here, we generated new mouse App Show less

As many as 700,000 unique sequences in the human genome are predicted to fold into G-quadruplexes (G4s), non-canonical structures formed by Hoogsteen guanine-guanine pairing within G-rich nucleic acids. G4s play both physiological and pathological roles in many vital cellular processes including DNA replication, DNA repair and RNA transcription. Several reagents have been developed to visualize G4s in vitro and in cells. Recently, Zhen et al. synthesized a small protein G4P based on the G4 recognition motif from RHAU (DHX36) helicase (RHAU specific motif, RSM). G4P was reported to bind the G4 structures in cells and in vitro, and to display better selectivity toward G4s than the previously published BG4 antibody. To get insight into G4P- G4 interaction kinetics and selectivity, we purified G4P and its expanded variants, and analyzed their G4 binding using single-molecule total internal reflection fluorescence microscopy and mass photometry. We found that G4P binds to various G4s with affinities defined mostly by the association rate. Doubling the number of the RSM units in the G4P increases the protein's affinity for telomeric G4s and its ability to interact with sequences folding into multiple G4s. Show less

As many as 700,000 unique sequences in the human genome are predicted to fold into G-quadruplexes (G4s), non-canonical structures formed by Hoogsteen guanine-guanine pairing within G-rich nucleic acids. G4s play both physiological and pathological roles in many vital cellular processes including DNA replication, DNA repair and RNA transcription. Several reagents have been developed to visualize G4s Show less

Lung adenocarcinoma is the most common type of lung cancer. Known risk variants explain only a small fraction of lung adenocarcinoma heritability. Here, we conducted a two-stage genome-wide association study of lung adenocarcinoma of East Asian ancestry (21,658 cases and 150,676 controls; 54.5% never-smokers) and identified 12 novel susceptibility variants, bringing the total number to 28 at 25 independent loci. Transcriptome-wide association analyses together with colocalization studies using a Taiwanese lung expression quantitative trait loci dataset (n = 115) identified novel candidate genes, including FADS1 at 11q12 and ELF5 at 11p13. In a multi-ancestry meta-analysis of East Asian and European studies, four loci were identified at 2p11, 4q32, 16q23, and 18q12. At the same time, most of our findings in East Asian populations showed no evidence of association in European populations. In our studies drawn from East Asian populations, a polygenic risk score based on the 25 loci had a stronger association in never-smokers vs. individuals with a history of smoking (P Show less

A 70-year-old woman with complaints of edema, general malaise, and hypotension was diagnosed with renal amyloidosis, and laser microdissection mass spectrometry revealed her amyloidosis to predominantly comprise the apolipoprotein A-IV type. The M-protein turned from negative to positive during the course, and a bone marrow biopsy showed smoldering myeloma. Treatment with bortezomib and dexamethasone failed to save her from heart failure six months after the onset. Western blotting of urine samples at the time of the renal biopsy showed that amyloid light-chain κ amyloidosis had been present since the onset. Unlike the myeloma, Congo red staining was positive in the plasma cells of the bone marrow. Show less

Exostosin 1 and exostosin 2 (EXT1/EXT2) on glomerular basement membrane (GBM) were recently reported as novel putative antigens in secondary membranous nephropathy with autoimmune disease. However, the clinical significance of glomerular EXT1/EXT2 remains elusive in patients with lupus nephritis (LN). The immunofluorescence staining pattern of glomerular EXT1/EXT2 is also undetermined in membranous LN (MLN) or proliferative LN (PLN). We cross-sectionally analyzed patients with MLN (pure class V, n = 11) and PLN (class III, IV, and mixed class III/IV + V, n = 22) who underwent renal biopsies between 2010 and 2020 at Showa University Hospital. Glomerular EXT1/EXT2 expressions were evaluated by immunofluorescence. T-helper (Th) cell-related serum inflammatory cytokines were measured using enzyme-linked immunosorbent assay. The positivity for both EXT1/EXT2 was higher in patients with MLN than PLN (90.9% vs 63.6%, P = 0.212). MLN showed global and bright granular EXT1/EXT2 expressions along GBM, while PLN showed segmental and moderate expressions on GBM. Additionally, glomerular EXT1/EXT2 positivity was not associated with the degree of proteinuria or renal function in MLN and PLN patients, but the levels of serum anti-dsDNA antibody and circulating immune complexes were lower in patients with EXT1/EXT2-positive MLN than EXT1/EXT2-negative PLN. Moreover, serum complement levels and IL-4/IFN-γ ratios were elevated in EXT1/EXT2-positive MLN than EXT1/EXT2-negative PLN. Collectively, immunofluorescence staining for glomerular EXT1/EXT2 had characteristic patterns between MLN and PLN. Glomerular EXT1/EXT2 expressions tended to be high in Th2-dominant MLN patients without severe hypocomplementemia and elevated autoantibodies. Thus, EXT1/EXT2 might be involved in the unique developmental mechanism of MLN. Show less

Monogenic diabetes is clinically heterogeneous and differs from common forms of diabetes (type 1 and 2). We aimed to investigate the clinical usefulness of a comprehensive genetic testing system, comprised of targeted next-generation sequencing (NGS) with phenotype-driven bioinformatics analysis in patients with monogenic diabetes, which uses patient genotypic and phenotypic data to prioritize potentially causal variants. We performed targeted NGS of 383 genes associated with monogenic diabetes or common forms of diabetes in 13 Japanese patients with suspected (n = 10) or previously diagnosed (n = 3) monogenic diabetes or severe insulin resistance. We performed in silico structural analysis and phenotype-driven bioinformatics analysis of candidate variants from NGS data. Among the patients suspected having monogenic diabetes or insulin resistance, we diagnosed 3 patients as subtypes of monogenic diabetes due to disease-associated variants of INSR, LMNA, and HNF1B. Additionally, in 3 other patients, we detected rare variants with potential phenotypic effects. Notably, we identified a novel missense variant in TBC1D4 and an MC4R variant, which together may cause a mixed phenotype of severe insulin resistance. This comprehensive approach could assist in the early diagnosis of patients with monogenic diabetes and facilitate the provision of tailored therapy. Show less

The Dedicator Of CytoKinesis (DOCK) family of atypical guanine nucleotide exchange factors activates the Rho family GTPases Rac and/or Cdc42 through DOCK homology region 2 (DHR-2). Previous structural analyses of the DHR-2 domains of DOCK2 and DOCK9 have shown that they preferentially bind Rac1 and Cdc42, respectively; however, the molecular mechanism by which DHR-2 distinguishes between these GTPases is unclear. Here we report the crystal structure of the Cdc42-bound form of the DOCK7 DHR-2 domain showing dual specificity for Rac1 and Cdc42. The structure revealed increased substrate tolerance of DOCK7 at the interfaces with switch 1 and residue 56 of Cdc42. Furthermore, molecular dynamics simulations showed a closed-to-open conformational change in the DOCK7 DHR-2 domain between the Cdc42- and Rac1-bound states by lobe B displacement. Our results suggest that lobe B acts as a sensor for identifying different switch 1 conformations and explain how DOCK7 recognizes both Rac1 and Cdc42. Show less

Obesity, a risk factor for all-cause and cardiovascular mortality, is a major health concerns among middle-aged men. The aim of this study was to investigate a possible association of dietary habits and obesity related single nucleotide polymorphisms (SNPs) with obesity and metabolic abnormalities. We conducted a retrospective cohort study using annual health examination data of 5112 male workers, obtained between 2007 and 2011. Average dietary energy was estimated using electronically collected meal purchase data from cafeteria. We examined 8 SNPs related to obesity: GHRL rs696217, PPARG rs1175544, ADIPOQ rs2241766, ADIPOQ rs1501299, PPARD rs2016520, APOA5 rs662799, BRAP rs3782886, and ITGB2 rs235326. We also examined whether SNPs that were shown to associate with obesity affect other metabolic abnormalities such as blood pressure (BP), glucose, and lipid profile. Average dietary energy significantly associated with increased abdominal circumference (AC) and body mass index (BMI). The odds ratios (ORs) of overweight and obesity also increased. The major allele of rs696217 significantly increased BMI and an increased OR with obesity, while the minor allele of rs3782886 was associated with significantly decreased AC and the decreased ORs with overweight and obesity. The minor allele of rs3782886 was also associated with significantly decreased systolic BP (SBP), triglyceride (TG), high-density lipoprotein (HDL), and fasting blood sugar (FBS), while rs696217 was not associated with other metabolic abnormalities. Average dietary energy in lunch, rs3782886, and rs696217 were associated with obesity, and rs3782886 was associated with other metabolic abnormalities. Show less

Several single nucleotide polymorphisms (SNPs) have been implicated in the predisposition to chronic kidney disease (CKD). Atherosclerotic disease is deeply involved in the incidence of CKD; however, whether SNPs related to arteriosclerosis are involved in CKD remains unclear. This study aimed to identify SNPs associated with CKD and to examine whether risk allele accumulation is associated with CKD. We conducted a cross-sectional study using data of 4814 male workers to examine the association between estimated glomerular filtration rate (eGFR) and 59 candidate polymorphisms (17 CKD, 42 atherosclerotic diseases). We defined the genetic risk score (GRS) as the total number of risk alleles that showed a significant association in this analysis and examined the relationship with CKD (eGFR < 60 ml/min/1.73m2). Multivariate logistic regression, discrimination by area under the receiver operating characteristic curve, integrated discrimination improvement (IDI), and category-free net reclassification improvement (cNRI) were evaluated. In total, 432 participants were categorized as having CKD. We found eight candidate SNPs with P value < 0.05 (CX3CR1 rs3732379, SHROOM3 rs17319721, MTP rs1800591, PIP5K1B rs4744712, APOA5 rs662799, BRAP rs3782886, SPATA5L1 rs2467853, and MCP1 rs1024611) in the multivariate linear regression adjusted for age, body mass index, systolic blood pressure, and fasting blood glucose. Among these eight SNPs, BRAP rs3782886 and SPATA5L1 rs2467853 were significantly associated with eGFR (false discovery rate < 0.05). GRS was significantly associated with CKD (odds ratio, 1.17; 95% confidence interval, 1.09-1.26). C-statisics improved from 0.775 to 0.780 but showed no statistical significance. However, adding GRS significantly improved IDI and cNRI (0.0057, P = 0.0028, and 0.212, P < 0.001, respectively). After adjustment for clinical factors, kidney function was associated with BRAP rs3782886 and SPATA5L1 rs2467853 and the GRS for CKD that we developed was associated CKD. Show less

Age-related macular degeneration (AMD) is a major cause of blindness in the elderly. Previous sequencing studies of AMD susceptibility genes have revealed the association of rare coding variants in CFH, CFI, C3 and C9 in European population; however, the impact of rare or low-frequency coding variants on AMD susceptibility in other populations is largely unknown. To identify the role of low-frequency coding variants on exudative AMD susceptibility in a Japanese population, we analysed the association of coding variants of 34 AMD candidate genes in the two-stage design by a multiplex PCR-based target sequencing method. We used a total of 2,886 (1st: 827, 2nd: 2,059) exudative AMD cases including typical AMD, polypoidal choroidal vasculopathy, and retinal angiomatous proliferation and 9,337 (1st: 3,247 2nd: 6,090) controls. Gene-based analysis found a significant association of low-frequency variants (minor allele frequency (MAF) < 0.05) in CETP, C2 and CFB. The association of CETP remained after conditioned with all known genome-wide association study (GWAS) associated variants. In addition, when we included only disruptive variants, enrichment of rare variants (MAF < 0.01) was also observed after conditioned with all GWAS associated variants (P = 1.03 × 10−6, odds ratio (OR) = 2.48). Haplotype and conditional analysis of the C2-CFB-SKIV2L locus showed a low-frequency variant (R74H) in CFB would be individually associated with AMD susceptibility independent of the GWAS associated SNP. These findings highlight the importance of target sequencing to reveal the impact of rare or low-frequency coding variants on disease susceptibility in different ethnic populations. Show less

Although many single nucleotide polymorphisms (SNPs) have been identified to be associated with metabolic syndrome (MetS), there was only a slight improvement in the ability to predict future MetS by the simply addition of SNPs to clinical risk markers. To improve the ability to predict future MetS, combinational effects, such as SNP-SNP interaction, SNP-environment interaction, and SNP-clinical parameter (SNP × CP) interaction should be also considered. We performed a case-control study to explore novel SNP × CP interactions as risk markers for MetS based on health check-up data of Japanese male employees. We selected 99 SNPs that were previously reported to be associated with MetS and components of MetS; subsequently, we genotyped these SNPs from 360 cases and 1983 control subjects. First, we performed logistic regression analyses to assess the association of each SNP with MetS. Of these SNPs, five SNPs were significantly associated with MetS (P < 0.05): LRP2 rs2544390, rs1800592 between UCP1 and TBC1D9, APOA5 rs662799, VWF rs7965413, and rs1411766 between MYO16 and IRS2. Furthermore, we performed multiple logistic regression analyses, including an SNP term, a CP term, and an SNP × CP interaction term for each CP and SNP that was significantly associated with MetS. We identified a novel SNP × CP interaction between rs7965413 and platelet count that was significantly associated with MetS [SNP term: odds ratio (OR) = 0.78, P = 0.004; SNP × CP interaction term: OR = 1.33, P = 0.001]. This association of the SNP × CP interaction with MetS remained nominally significant in multiple logistic regression analysis after adjustment for either the number of MetS components or MetS components excluding obesity. Our results reveal new insight into platelet count as a risk marker for MetS. Show less

7-Dehydrocholesterol (7-DHC) is a common precursor of vitamin D3 and cholesterol. Although various oxysterols, oxygenated cholesterol derivatives, have been implicated in cellular signaling pathways, 7-DHC metabolism and potential functions of its metabolites remain poorly understood. We examined 7-DHC metabolism by various P450 enzymes and detected three metabolites produced by sterol 27-hydroxylase (CYP27A1) using high-performance liquid chromatography. Two were further identified as 25-hydroxy-7-DHC and 26/27-hydroxy-7-DHC. These 7-DHC metabolites were detected in serum of a patient with Smith-Lemli-Opitz syndrome. Luciferase reporter assays showed that 25-hydroxy-7-DHC activates liver X receptor (LXR) α, LXRβ and vitamin D receptor and that 26/27-hydroxy-7-DHC induces activation of LXRα and LXRβ, although the activities of both compounds on LXRs were weak. In a mammalian two-hybrid assay, 25-hydroxy-7-DHC and 26/27-hydroxy-7-DHC induced interaction between LXRα and a coactivator fragment less efficiently than a natural LXR agonist, 22(R)-hydroxycholesterol. These 7-DHC metabolites did not oppose agonist-induced LXR activation and interacted directly to LXRα in a manner distinct from a potent agonist. These findings indicate that the 7-DHC metabolites are partial LXR activators. Interestingly, 25-hydroxy-7-DHC and 26/27-hydroxy-7-DHC suppressed mRNA expression of sterol regulatory element-binding protein 1c, an LXR target gene, in HepG2 cells and HaCaT cells, while they weakly increased mRNA levels of ATP-binding cassette transporter A1, another LXR target, in HaCaT cells. Thus, 7-DHC is catabolized by CYP27A1 to metabolites that act as selective LXR modulators. Show less

This study is a post-hoc analysis of a subset of patients who participated in our multi-institutional case-control study that evaluated the effects of pitavastatin in patients with non-alcoholic fatty liver disease (NAFLD) with hypercholesterolemia. Serum samples of fifteen patients with biopsy-proven NAFLD with dyslipidemia were investigated. Serum markers of lipid metabolism were quantified by liquid chromatography-mass spectrometry (LC-MS)/MS. These data were then compared with those of 36 sex- and age-matched healthy controls. In addition, changes in these markers produced by treatment with pitavastatin were evaluated. Serum non-cholesterol sterols, reflecting intestinal cholesterol absorption, were significantly lower in the NAFLD patients compared to the controls, and the cholesterol synthesis marker, the ratio of lathosterol to cholesterol, was not significantly different between the two groups. Serum proportions of liver X receptor α (LXRα) ligand oxysterols (ratios to cholesterol) were significantly elevated in the NAFLD patients compared to the controls. The sum of oxysterols relative to cholesterol and the homeostasis model assessment as an index of insulin resistance (HOMA-IR) were significantly correlated. The marker representing cholesterol synthesis was significantly suppressed by pitavastatin treatment, from 3 months after initiation of the treatment, and the suppression remained significant during the observation period. The markers representing cholesterol absorption were unchanged at 3 months, but had significantly increased at 12 months. Serum oxysterol levels relative to cholesterol maintained high values and did not change significantly during the 12-month period of treatment. We speculate that serum LXRα ligand oxysterol levels (relative to cholesterol) could be surrogate markers of insulin resistance, and that high oxysterol levels in the circulation may play an important role in the development of hepatic and peripheral insulin resistance followed by NAFLD. Show less

The aim of this study was to investigate the relationship between serum apolipoprotein (apo) A-IV levels and markers for atherosclerosis, including carotid intima-media thickness (CIMT) and the ankle-brachial index (ABI), in hemodialysis patients. We performed a cross-sectional study involving 116 maintenance hemodialysis patients (70 males; median age, 64 years), measuring CIMT, ABI, the usual laboratory examinations, and serum apo A-IV before the dialysis session. The apo A-IV concentration was measured by a noncompetitive ELISA. Serum apo A-IV concentrations were significantly lower in hemodialysis patients with cardiovascular disease and plaque in the carotid artery. The apo A-IV level was positively associated with urea nitrogen and creatinine, and negatively associated with age, interleukin-6, the neutrophil/lymphocyte ratio, and maximum CIMT. Moreover, serum apo A-IV concentrations were significantly lower in the low ABI group. On logistic analysis, patients with high apo A-IV levels had a lower odds ratio for atherosclerosis (maximum CIMT > 1.0) and cardiovascular disease compared to patients with low apo A-IV levels. On stepwise multivariate regression analysis, the serum apo A-IV level was independently associated with creatinine, the neutrophil/lymphocyte ratio, and the maximum CIMT. Serum apo A-IV is associated with atherosclerotic lesions in hemodialysis patients. Apo A-IV levels may be useful for estimating the risk of cardiovascular disease in dialysis patients. Show less

DEC1 (BHLHB2/Stra13/Sharp2)-a basic helix-loop-helix transcription factor-is known to be involved in various biological phenomena including clock systems and metabolism. In the clock systems, Dec1 expression is dominantly up-regulated by CLOCK : BMAL1 heterodimer, and it exhibits circadian rhythm in the suprachiasmatic nucleus (SCN)-the central circadian pacemaker-and other peripheral tissues. Recent studies have shown that the strong circadian rhythmicity of Dec1 in the SCN was abolished by Clock mutation, whereas that in the liver was affected, but not abolished, by Clock mutation. Moreover, feeding conditions affected hepatic Dec1 expression, which indicates that Dec1 expression is closely linked with the metabolic functions of the liver. Among ligand-activated nuclear receptors examined, LXRalpha and LXRbeta with T0901317-agonist for LXR-were found to be potent enhancers for Dec1 promoter activity, and a higher expression level of LXRalpha protein was detected in the liver than in the kidney and heart. T0901317 increased the levels of endogenous Dec1 transcript in hepatoma cells. Chromatin immunoprecipitation assay indicated that LXRalpha bound to the Dec1 promoter, and an LXRalpha-binding site was identified. These observations indicate that hepatic DEC1 mediates the ligand-dependent LXR signal to regulate the expression of genes involved in the hepatic clock system and metabolism. Show less

17beta-Hydroxysteroid dehydrogenase (17beta-HSD) Type3 is an NADPH-dependent membrane-bound enzyme that is specifically expressed in testis and catalyzes the conversion of androstenedione to testosterone. To date, the sequence of Type3 enzymes has been clarified in humans, mice and rats; however, the sequence of the pig enzyme remains unknown. In this study, we determined the cDNA sequence of pig testicular 17beta-HSD Type3. PCR primers for partial pig testicular 17beta-HSD Type3 were designed from rat and human enzyme consensus sequences. Full-length cDNA was obtained by 3'- and 5'-RACE based on partial PCR products. The cDNA coding region was 933 bp in length, which is the same as the human enzyme, and shared 84.7% sequence identity with the human cDNA coding region. The monomer was estimated to have a molecular weight of 34,855 and to contain 310 amino acid residues. The predicted pig amino acid sequence showed 81.9, 75.5 and 72.9% sequence identity with the human, rat and mouse sequences, respectively. To elucidate 17beta-HSD Type3 activity, the expression vector pCMV/pig17beta-HSD3 was established and transfected into human embryo kidney 293 cells. Subsequently, 17beta-HSD activity (androstenedione conversion to testosterone) was strongly detected in cell lysates. Show less

EXT gene family members including EXT1, EXT2, and EXTL2 are glycosyltransferases required for heparan sulfate biosynthesis. To examine the biological functions of rib-2, a member of the Caenorhabditis elegans EXT gene family, we generated a mutant worm lacking the rib-2 gene using the UV-TMP method followed by sib-selection. Inactivation of rib-2 alleles induced developmental abnormalities in F2 and F3 homozygous worms, while F1 heterozygotes showed a normal morphology. The F2 homozygous progeny generated from the F1 heterozygous hermaphrodites somehow developed to adult stage but exhibited abnormal characteristics such as developmental delay and egg-laying defects. The F3 homozygous progeny from the F2 homozygous hermaphrodites showed early developmental defects and most of the F3 worms stopped developing during the gastrulation stage. Whole-mount staining analysis for heparan sulfate using Toluidine blue (pH 2.5) revealed a defect of heparan sulfate biosynthesis in the F2 homozygotes. The analysis using fluorometric post-column high-performance liquid chromatography also uncovered reduced production of heparan sulfate in the rib-2 mutant. These results indicate that rib-2 is essential for embryonic development and heparan sulfate biosynthesis in C. elegans. Show less