Cholangiopathies share common features, including bile duct proliferation, periportal fibrosis, and intrahepatic cholestasis. Damage of biliary epithelium by autoimunne disorder, virus infection, toxic compounds, and developmental abnormalities causes severe progressive hepatic disorders responsible for high mortality. However, the etiologies of these cholestatic diseases remain unclear because useful models to study the pathogenic mechanisms are not available. In the present study, we have found that ezrin knockdown (Vil2kd/kd ) mice develop severe intrahepatic cholestasis characterized by extensive bile duct proliferation, periductular fibrosis, and intrahepatic bile acid accumulation without developmental defects of bile duct morphology and infiltration of inflammatory cells. Ezrin is a membrane cytoskeletal cross‐linker protein, which is known to interact with transporters, scaffold proteins, and actin cytoskeleton at the plasma membrane. We found that the normal apical membrane localizations of several transport proteins including cystic fibrosis transmembrane conductance regulator (CFTR), anion exchanger 2 (AE‐2), aquaporin 1 (AQP1), and Na⁺/H⁺ exchanger regulatory factor were disturbed in bile ducts of Vil2kd/kd mice. Stable expression of a dominant negative form of ezrin in immortalized mouse cholangiocytes also led to the reduction of the surface expression of CFTR, AE‐2, and AQP1. Reduced surface expression of these transport proteins was accompanied by reduced functional expression, as evidenced by the fact these cells exhibited decreased CFTR‐mediated Cl⁻ efflux activity. Furthermore, bile flow and biliary HCO₃ ⁻ concentration were also significantly reduced in Vil2kd/kd mice. Conclusion: Dysfunction of ezrin mimics important aspects of the pathological mechanisms responsible for cholangiopathies. The Vil2kd/kd mouse may be a useful model to exploit in the development and testing of potential therapies for cholangiopathies. (Hepatology 2015;61:1660‐1671)

Ezrin is highly expressed in glomerular podocytes and is reported to form a multi-protein complex with scaffold protein Na⁺/H⁺ exchanger regulatory factor 2 (NHERF2) and podocalyxin, a major sialoprotein. Podocalyxin-knockout mice died within 24 h of birth with anuric renal failure, whereas NHERF2-knockout mice show no apparent changes in the glomerular functions. However, the physiological roles of ezrin in glomerular podocytes remain unclear. Here, we investigated the importance of ezrin in the regulation of glomerular podocyte function using ezrin-knockdown mice (Vil2 ^kd/kd ). The Vil2 ^kd/kd mice did not exhibit apparent glomerular dysfunction, morphological defects or abnormal localisation of podocalyxin and NHERF2 in podocytes. Thus, we investigated the influence of ezrin defects on Rho-GTPase activity, as ezrin interacts with the Rho-GTPase dissociation inhibitor (Rho-GDI), which plays a key role in the regulation of podocyte actin organisation. In Vil2 ^kd/kd glomeruli, Rac1 activity was significantly reduced compared to wildtype (WT) glomeruli at baseline. Furthermore, Vil2 ^kd/kd mice showed reduced susceptibility to glomerular injury. In WT glomeruli, Rac1 activity was enhanced in nephrotic conditions, but remained at baseline levels in Vil2 ^kd/kd glomeruli, suggesting that loss of ezrin protects podocytes from injury-induced morphological changes by suppressing Rac1 activation.

Ursodeoxycholic acid (UDCA) is a hydrophilic bile acid that possesses many pharmacological effects, including increasing bile flow, changing the hydrophobicity of the bile acid pool, and modulation of the immune response. UDCA has been approved for treating cholestatic liver disease, such as primary biliary cholangitis. However, several unanticipated severe side effects of UDCA are observed in cholestatic patients, and its pharmacological benefits remain controversial. We reported that ezrin-knockdown (Vil2kd/kd) mice exhibited severe hepatic injury because of a functional disorder in bile duct fluidity and alkalinity regulation, resembling human intrahepatic cholestatic disease. Here we used Vil2kd/kd mice as a cholestatic model to investigate the pharmacological effects of UDCA. We investigated the effects of oral and parenteral administration of UDCA on Vil2kd/kd mice. In Vil2kd/kd mice, fed a 0.5% (w/w) UDCA diet for 3 weeks, hepatic injury was exacerbated, although oral administration of a lower dose of UDCA slightly improved hepatic function in Vil2kd/kd mice. On the other hand, intraperitoneal administration of UDCA (50 mg/kg/d) ameliorated hepatic function and markedly reduced periductal fibrosis and cholangiocyte proliferation in Vil2kd/kd mice although biliary pH and HCO3- concentration were not improved. The expression levels of inflammatory and profibrotic genes were also significantly decreased in these mice. Furthermore, UDCA prevented cholangiocytes from hydrophobic bile acid-induced cytotoxicity independent of extracellular pH in in vitro experiments. These results suggest that an appropriate dosage of UDCA can ameliorate the intrahepatic cholestasis in Vil2kd/kd mice without changing the biliary bicarbonate secretion.

As glucose-dependent insulinotropic polypeptide (GIP) possesses pro-adipogenic action, the suppression of the GIP hypersecretion seen in obesity might represent a novel therapeutic approach to the treatment of obesity. However, the mechanism of GIP hypersecretion remains largely unknown. In the present study, we investigated GIP secretion in two mouse models of obesity: High-fat diet-induced obese (DIO) mice and leptin-deficient Lep^ob/ob mice. In DIO mice, plasma GIP was increased along with an increase in GIP mRNA expression in the lower small intestine. Despite the robust alteration in the gut microbiome in DIO mice, co-administration of maltose and the α-glucosidase inhibitor (α-GI) miglitol induced the microbiome-mediated suppression of GIP secretion. The plasma GIP levels of Lep^ob/ob mice were also elevated and were suppressed by fat transplantation. The GIP mRNA expression in fat tissue was not increased in Lep^ob/ob mice, while the expression of an interleukin-1 receptor antagonist (IL-1Ra) was increased. Fat transplantation suppressed the expression of IL-1Ra. The plasma IL-1Ra levels were positively correlated with the plasma GIP levels. Accordingly, although circulating GIP levels are increased in both DIO and Lep^ob/ob mice, the underlying mechanisms differ, and the anti-obesity actions of α-GIs and leptin sensitizers may be mediated partly by the suppression of GIP secretion.

Previously we estimated the prevalence of essential hypertension (EH) as around 0.1% and suggested that male gender, obesity, and strong genetic background (hypertension in parents) were risk factors for EH in a young population aged less than 30 based on targeted screening for hypertension at a university health check-up. This study also revealed a high incidence of white coat hypertension (WCH) in university students, and thus, we continued this screening for four consecutive years, and examined the prognosis and clinical characteristics of young-onset WCH. Three occasions of casual blood pressure (BP) measurement and additional home BP measurement revealed 72 WCH and 15 EH students (all males) during the 4-year study period. None of the WCH students had elevated home BP to the level of hypertension during their stay at university, and 26 out of 38 WCH students participating screening in the following years showed normal casual BP. Although WCH students showed a significantly higher pulse rate than controls, WCH could not be fully differentiated from EH either by pulse rate or by correlation between casual BP value and pulse rate. These findings indicate the requirement of longer follow-up after graduation to determine the prognosis of young-onset WCH, though EH and WCH in the young population share the same risk factors and, possibly, autonomic nervous system dysfunction. Since diagnosis of WCH has limited importance for university students, screening of EH following a general health check-up would elevate the clinical validity of casual BP measurement at the university.

Background: α-cyclodextrin (α-CD) is one of the dietary fibers that may have a beneficial effect on cholesterol and/or glucose metabolism, but its efficacy and mode of action remain unclear. Methods: In the present study, we examined the anti-hyperglycemic effect of α-CD after oral loading of glucose and liquid meal in mice. Results: Administration of 2 g/kg α-CD suppressed hyperglycemia after glucose loading, which was associated with increased glucagon-like peptide 1 (GLP-1) secretion and enhanced hepatic glucose sequestration. By contrast, 1 g/kg α-CD similarly suppressed hyperglycemia, but without increasing secretions of GLP-1 and insulin. Furthermore, oral α-CD administration disrupts lipid micelle formation through its inclusion of lecithin in the gut luminal fluid. Importantly, prior inclusion of α-CD with lecithin in vitro nullified the anti-hyperglycemic effect of α-CD in vivo, which was associated with increased intestinal mRNA expressions of SREBP2-target genes (Ldlr, Hmgcr, Pcsk9, and Srebp2). Conclusions: α-CD elicits its anti-hyperglycemic effect after glucose loading by inducing lecithin inclusion in the gut lumen and activating SREBP2, which is known to induce cholecystokinin secretion to suppress hepatic glucose production via a gut/brain/liver axis.