[Surgical Left Atrial Ablation Through the Left Atrial Appendage During Aortic Valve Replacement].

This is a 2-case report of concomitant left atrial( LA) surgical ablation through the left atrial appendage( LAA) for atrial fibrillation( AF) during aortic valve replacement( AVR). Case 1, a 74-year-old man, and Case 2, an 85-years-old woman, were presented for AVR and AF. Under the cardio-pulmonary bypass, right and left pulmonary vein isolations( PVIs) were performed with radiofrequency( RF) ablation devices. Through the opened LAA, an RF lesion was placed to connect the bilateral PVIs and the LAA, and another RF line was placed toward the mitral annulus. A right atrial( RA) isthmus ablation was also performed through a RA incision. Postoperatively, sinus rhythm and both RA and LA contraction were obtained, and have been maintained for 7.5 and 5 years without classⅠ/Ⅲ antiarrhythmic drugs in Case 1 and 2, respectively. This procedure is considered to be effective and safe, although further study is needed.

[Tricuspid Valve Ring Annuloplasty with Posterior Annular Plication for Functional Tricuspid Regurgitation].

This report presents a modified procedure of tricuspid valve ring annuloplasty (R-TAP) with posterior annular plication for functional tricuspid regurgitation (TR). Sutures on the native annulus were placed by a standard fashion in R-TAP, and those on the posterior annulus and its bilateral commissures were passed through in a narrow range between the 3 and 4 o'clock positions of the 26-mm ring. The other sutures were done with an usual manner and the ring was fixed to the annulus, resulting in the posterior annular plication( bicuspidization). Follow-up was performed for more than 5 years( mean: 7.9 years, range:5.5~11.5 years) by echocardiography in 13 cases. Postoperative TR reduced significantly to less than moderate, which was maintained during the entire follow-up period, even in the case with atrial fibrillation. There was no sign of tricuspid stenosis. R-TAP with posterior annular plication was feasible, reproducible, and effective, although further investigation is needed.

Characterisation of Ppy-lineage cells clarifies the functional heterogeneity of pancreatic beta cells in mice.

AIMS/HYPOTHESIS: Pancreatic polypeptide (PP) cells, which secrete PP (encoded by the Ppy gene), are a minor population of pancreatic endocrine cells. Although it has been reported that the loss of beta cell identity might be associated with beta-to-PP cell-fate conversion, at present, little is known regarding the characteristics of Ppy-lineage cells. METHODS: We used Ppy-Cre driver mice and a PP-specific monoclonal antibody to investigate the association between Ppy-lineage cells and beta cells. The molecular profiles of endocrine cells were investigated by single-cell transcriptome analysis and the glucose responsiveness of beta cells was assessed by Ca2+ imaging. Diabetic conditions were experimentally induced in mice by either streptozotocin or diphtheria toxin. RESULTS: Ppy-lineage cells were found to contribute to the four major types of endocrine cells, including beta cells. Ppy-lineage beta cells are a minor subpopulation, accounting for 12-15% of total beta cells, and are mostly (81.2%) localised at the islet periphery. Unbiased single-cell analysis with a Ppy-lineage tracer demonstrated that beta cells are composed of seven clusters, which are categorised into two groups (i.e. Ppy-lineage and non-Ppy-lineage beta cells). These subpopulations of beta cells demonstrated distinct characteristics regarding their functionality and gene expression profiles. Ppy-lineage beta cells had a reduced glucose-stimulated Ca2+ signalling response and were increased in number in experimental diabetes models. CONCLUSIONS/INTERPRETATION: Our results indicate that an unexpected degree of beta cell heterogeneity is defined by Ppy gene activation, providing valuable insight into the homeostatic regulation of pancreatic islets and future therapeutic strategies against diabetes. DATA AVAILABILITY: The single-cell RNA sequence (scRNA-seq) analysis datasets generated in this study have been deposited in the Gene Expression Omnibus (GEO) under the accession number GSE166164 ( www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE166164 ).

Development of monoclonal mouse antibodies that specifically recognize pancreatic polypeptide.

Pancreatic polypeptide (PP) is a 36-amino acid peptide encoded by the Ppy gene, which is produced by a small population of cells located in the periphery of the islets of Langerhans. Owing to the high amino acid sequence similarity among neuropeptide Y family members, antibodies against PP that are currently available are not convincingly specific to PP. Here we report the development of mouse monoclonal antibodies that specifically bind to PP. We generated Ppy knockout (Ppy-KO) mice in which the Ppy-coding region was replaced by Cre recombinase. The Ppy-KO mice were immunized with mouse PP peptide, and stable hybridoma cell lines producing anti-PP antibodies were isolated. Firstly, positive clones were selected in an enzyme-linked immunosorbent assay for reactivity with PP coupled to bovine serum albumin. During the screening, hybridoma clones producing antibodies that cross-react to the peptide YY (PYY) were excluded. In the second screening, hybridoma clones in which their culture media produce no signal in Ppy-KO islets but detect specific cells in the peripheral region of wild-type islets, were selected. Further studies demonstrated that the selected monoclonal antibody (23-2D3) specifically recognizes PP-producing cells, not only in mouse, but also in human and rat islets. The monoclonal antibodies with high binding specificity for PP developed in this study will be fundamental for future studies towards elucidating the expression profiles and the physiological roles of PP.

[Mitral Valve Replacement with the On-X Valve].

This is a 3-case report of mitral valve replacement (MVR) with an On-X mechanical valve followed up to 10 years. Case 1(64-year-old man) and case 2 (66-year-old woman) experienced traffic accident and traumatic event, respectively, in their chronic phase after MVR. Case 1 had multiple bone fractures of the bilateral lower limbs, which was followed by systemic infection and pyogenic spondylitis. He needed long-term antibiotics therapy for more than 4 years. Case 2 fell down at home and severely hit her head, which resulted in a traumatic subarachnoid hemorrhage. She was in a deep coma, and needed discontinuation of anticoagulation therapy for 4 weeks. Case1, 2, and 3(54-year-old man) are doing well in New York Heart Association functional class I without any valve-related thromboembolic or hemorrhagic events at 10, 9 and 8 years after MVR, respectively. On-X valve performance has also been found well maintained in all cases by echocardiography, even after traumatic accident or discontinuation of anticoagulation in Case 1 and 2. In this report, the On-X mechanical valve demonstrated good midterm result of its valve performance in the mitral position and its potential advantages in antithrombogenicity.

Normal pancreatic ß-cell function in mice with RIP-Cre-mediated inactivation of p62/SQSTM1.

Recent studies have suggested that decreased pancreatic ß-cell function and mass are common features of patients with type 2 diabetes mellitus. Pancreatic ß-cell homeostasis is regulated by various types of signaling molecules and stress responses. Sequestosome 1/p62 (SQSTM1, hereafter referred to as p62) is a ubiquitin-binding adaptor protein involved in cell signaling, oxidative stress, and autophagy. Because p62 appears to play an important role in maintaining mitochondrial quality control, it is possible that the loss of p62 in pancreatic ß cells contributes to mitochondrial dysfunction, and thus leading to impaired glucose tolerance. In this study we investigated the physiological roles of p62 by inactivating p62 in a ß-cell specific manner. We found that firstly, rat insulin-2 promoter-Cre (RIP-Cre)-mediated p62 inactivation did not cause body weight gain, although ubiquitous inactivation of p62 was previously shown to result in severe obesity. Secondly, we found no gross structural disorganization of the islets of p62-deficient mice. Consistent with normal islet morphology, no impairment in glucose tolerance was observed in mice with RIP-Cre-mediated p62 deletion. These results suggest that p62 is dispensable for normal islet organization and ß-cell function.

[Intraoperative Endoscopic Evaluation of the Aortic Valve in Aortic Valve Repair].

This report discusses intraoperative endoscopic evaluation of the aortic valve performed in 2 cases of aortic valve repair. The "direct" or real image by the endoscopy helped to confirm the preoperatively-known lesion and even to detect a new legion which was not detected preoperatively. The endoscopy also enabled the evaluation of the aortic valve under the pressure-loaded condition without releasing the aortic clamp. Postoperative aortic regurgitation was grade I or less in both cases, although it progressed to grade II at 1 year in case 2. Echocardiographic parameters demonstrated no change in the size of the aortic root configuration for 8 and 5 years in case 1 and case 2, respectively. Intraoperative aortic endoscopy was useful to define the pathogenesis of aortic regurgitation and to evaluate the cusp repair procedures, which may contribute to a good mid-term result of aortic valve repair.

Increased expression of ERp57/GRP58 is protective against pancreatic beta cell death caused by autophagic failure.

Autophagy is a tightly regulated self-digestion system. As in other cell types, autophagy plays an essential role in the homeostasis of pancreatic beta cells. However, the mechanisms involved in the deterioration of beta cell function caused by autophagic failure have not yet been fully elucidated. To gain insight into its mechanisms, we compared the protein expression of islets from beta cell-specific Atg7-deficient mice (Atg7(Δß-cell) mice) and their controls (Atg7(f/f) mice). Liquid chromatography/mass spectrometry after 1-dimensional electrophoresis identified the increased expression of ERp57/GRP58 in islets isolated from Atg7(Δß-cell) mice compared with those from Atg7(f/f) mice. The expression level of ERp57 was also elevated in rat insulinoma INS-1 cells by inducible knock-down of the atg7-gene. In Atg7 knock-down INS-1 cells, the suppression of ERp57 expression by siRNA resulted in an increase in the level of cleaved Caspase-3 protein and a decrease in the number of live cells. Furthermore, cell cycle analyses demonstrated that the suppressed expression of ERp57 increased the sub-G1 population. These data reveal that increased expression of ERp57 may contribute to the protection from beta cell death caused by autophagic failure.

Beneficial effects of vildagliptin combined with miglitol on glucose tolerance and islet morphology in diet-controlled db/db mice.

Dipeptidyl peptidase-4 (DPP-4) inhibitors improve glycemic control in patients with type 2 diabetes primarily by increasing plasma active glucagon-like peptide-1 (GLP-1) levels. While various combination therapies based on DPP-4 inhibitors have been proposed for treatment of type 2 diabetes, the effects of combination therapy of DPP-4 inhibitors and alpha-glucosidase inhibitors on ß-cell function are less characterized. We evaluated the effects of long-term treatment with vildagliptin, a DPP-4 inhibitor, on metabolic parameters and ß-cell function, in combination with miglitol, an alpha-glucosidase inhibitor, in diet-controlled db/db mice. In this study, 6-week-old male db/db mice were provided with standard chow twice a day for 6 weeks. Meal tolerance tests and glucose tolerance tests showed that the combination therapy of vildagliptin with miglitol, but not each alone, suppressed postprandial glycemic excursion, enhanced postprandial active GLP-1 levels and prevented deterioration of glucose tolerance in the db/db mice. The combination treatment did not alter ß-cell mass, but resulted in preserved expression of glucose transporter 2, Zinc transporter 8 and MafA and reduced the number of α cells. These results suggest that the combination of vildagliptin and miglitol prevents the development of overt diabetes in diet-controlled pre-diabetic db/db mice by normalizing postprandial glucose and incretin response, and by preserving ß-cell structure and the expression of factors essential for ß-cell function.

Involvement of the parasympathetic nervous system in the initiation of regeneration of pancreatic ß-cells.

The mechanism that initiates regeneration of pancreatic ß-cells is not clear at present. The vagal nerve is implicated in the regulation of gastrointestinal functions, glucose metabolism and proliferation of pancreatic ß-cells under physiological conditions. To elucidate the triggering mechanism of the regeneration of pancreatic ß-cells, we examined the involvement of the vagal nerve. To this end, we employed a rat pancreatic duct ligation (DL) model, in which profound ß-cell neogenesis and ß-cell proliferation were observed within a week. We administered atropine to block the vagal nerve. Administration of atropine inhibited proliferation of ß-cells in both islets and islet-like cell clusters (ICC), without affecting ductal cell proliferation in the ligated pancreas. The numbers of PDX-1 and MafB-positive cells in or attaching to the ducts were significantly reduced by atropine. MafB/glucagon and MafB/insulin double-positive cells were also decreased by atropine. Finally, atropine reduced the number of MafA-positive ductal cells, all of which were positive for insulin, by 50% on day 5. These results strongly suggest that the vagal nerve is involved in ß-cell proliferation, induction of endocrine progenitors and neogenesis of α- and ß-cells.

Extracellular matrix modulates insulin production during differentiation of AR42J cells: functional role of Pax6 transcription factor.

Extracellular matrix (ECM) modulates differentiation of pancreatic ß-cells during development. However, the mechanism by which ECM proteins modulate differentiation is not totally clear. We investigated the effect of ECM proteins on differentiation ß-cells in vitro. We investigated the effect of basement membrane ECM on differentiation of AR42J cells and rat ductal cells. First, we examined the effect of reconstituted basement membrane, Matrigel on differentiation of AR42J cells induced by activin and betacellulin. Matrigel augmented insulin production and increased the expression of GLUT2, SUR1, and glucokinase. Among various transcription factors investigated, Matrigel markedly upregulated the expression of Pax6. When Pax6 was overexpressed in cells treated with activin and betacellulin, the expression of insulin was upregulated. Conversely, knockdown of Pax6 significantly reduced the insulin expression in cells cultured on Matrigel. The effects of Matrigel on insulin-production and induction of Pax6 were reproduced partially by laminin-1, a major component of Matrigel, and inhibited by anti-integrin-ß1 antibody. Matrigel also enhanced the activation of p38 mitogen-activated kinase induced by activin and betacellulin, which was inhibited by anti-ß1 antibody. Finally, the effect of Matrigel on differentiation was reproduced in rat cultured ductal cells, and Matrigel also increased the expression of Pax6. These results indicate that basement membrane ECM augments differentiation of pancreatic progenitor cells to insulin-secreting cells by upregulating the expression of Pax6. .

Glucokinase and IRS-2 are required for compensatory beta cell hyperplasia in response to high-fat diet-induced insulin resistance.

Glucokinase (Gck) functions as a glucose sensor for insulin secretion, and in mice fed standard chow, haploinsufficiency of beta cell-specific Gck (Gck(+/-)) causes impaired insulin secretion to glucose, although the animals have a normal beta cell mass. When fed a high-fat (HF) diet, wild-type mice showed marked beta cell hyperplasia, whereas Gck(+/-) mice demonstrated decreased beta cell replication and insufficient beta cell hyperplasia despite showing a similar degree of insulin resistance. DNA chip analysis revealed decreased insulin receptor substrate 2 (Irs2) expression in HF diet-fed Gck(+/-) mouse islets compared with wild-type islets. Western blot analyses confirmed upregulated Irs2 expression in the islets of HF diet-fed wild-type mice compared with those fed standard chow and reduced expression in HF diet-fed Gck(+/-) mice compared with those of HF diet-fed wild-type mice. HF diet-fed Irs2(+/-) mice failed to show a sufficient increase in beta cell mass, and overexpression of Irs2 in beta cells of HF diet-fed Gck(+/-) mice partially prevented diabetes by increasing beta cell mass. These results suggest that Gck and Irs2 are critical requirements for beta cell hyperplasia to occur in response to HF diet-induced insulin resistance.

Administration of conophylline and betacellulin-delta4 increases the beta-cell mass in neonatal streptozotocin-treated rats.

The present study was conducted to examine the effect of administration of conophylline (CnP) and betacellulindelta4 (BTCdelta4) on the beta-cell mass in neonatal streptozotocin-treated rats (neonatal STZ rats). STZ (100 microg/g) was injected into neonatal rats, and then CnP (2 microg/g) and/or BTCdelta4 (200 pmol/g) were administered to neonatal STZ rats for 1 week. The plasma glucose concentration was monitored, and an intraperitoneal glucose tolerance test (ipGTT) was performed on day 8 and at 8 weeks after the STZ injection. In neonatal STZ rats treated with control solution (S group), the plasma glucose concentration increased for several days after the STZ injection, returned to nearly normal levels, and then increased gradually after six weeks of age. Eight weeks after the STZ-injection, the plasma glucose concentration was increased significantly compared to that of normal rats. The glucose response to ipGTT was significantly reduced in neonatal STZ rats treated with CnP (CnP group), BTCdelta4 (delta4 group) and CnP+BTCdelta4 (CnP+delta4 group). The beta-cell mass and the insulin content of the pancreas were significantly increased in the CnP group and delta4 group. The effect of CnP+delta4 was greater than that of CnP alone or BTCdelta4 alone. CnP+BTCdelta4 significantly increased the number of PDX-1-positive ductal cells and the number of insulin/BrdU double-positive ductal cells. These results indicate the efficacy of CnP and BTCdelta4 in increasing the beta-cells mass of neonatal STZ-treated rats.

Enhanced Expression of the Key Mitosis Regulator Cyclin B1 Is Mediated by PDZ-Binding Kinase in Islets of Pregnant Mice.

The proliferation of pancreatic ß cells is enhanced to enable an increase in ß-cell mass and to compensate for insulin resistance during pregnancy. To elucidate the mechanisms involved, we previously investigated islets from pregnant and nonpregnant mice by gene expression profiling and found that the expression of postsynaptic density-95/Discs large/zonula occludens-1 (PDZ)-binding kinase (Pbk), a member of the mitogen-activated protein kinase kinase family, is increased in pregnant mouse islets compared with control mouse islets. Among the pregnancy hormones, treatment with estradiol upregulated Pbk expression. Inhibition of Pbk expression using a small interfering RNA for Pbk reduced bromodeoxyuridine incorporation in mouse insulinoma 6 cells, which was accompanied by a decreased expression of Ccnb1, a regulatory gene involved in mitosis. Ccnb1 expression was augmented in mouse islets during pregnancy. The forced expression of Pbk using an adenovirus system in isolated mouse islets increased Ccnb1 expression, and the Pbk inhibitor HI-TOPK-032 suppressed Ccnb1 expression in islets isolated from pregnant mice. Our results suggest that Pbk contributes to the expansion of islets during pregnancy and that Ccnb1 may assist Pbk in its role in ß-cell proliferation.

Emx2 and Pax6 function in cooperation with Otx2 and Otx1 to develop caudal forebrain primordium that includes future archipallium.

One of the central issues in developmental neurobiology is how the forebrain is organized ontogenetically. The traditional view is that the anterior neuroectoderm first develops into mesencephalic and prosencephalic vesicles; the latter vesicle subsequently develops into the diencephalon and secondary prosencephalon, of which dorsal parts protrude to generate the telencephalon. The diencephalon yields the pretectum, thalamus, and prethalamus, and the telencephalon produces the archipallium, neopallium, and ganglionic eminences. By identifying cell descendants that once expressed Emx2 with use of the Cre knock-in mutant into the Emx2 locus and analyzing phenotypes of double mutants between Emx2 and Otx2/Otx1 and between Emx2 and Pax6, we propose that at the 3-6 somite stage, the anterior neuroectoderm develops into three primordia: midbrain, caudal forebrain, and rostral forebrain. The caudal forebrain primordium generates not only the pretectum, thalamus, and prethalamus but also the archipallium, cortical hem, choroid plexus, choroidal roof, and eminentia thalami. The primordium corresponds to the Emx2- or Pax6-positive region at the 3-6 somite stage that most probably does not include the future neopallium or commissural plate. Otx2 and Otx1 that are expressed in the entire future forebrain and midbrain cooperate with this Emx2 and Pax6 expression in the development of the caudal forebrain primordium; Emx2 and Pax6 functions are redundant. In the embryonic day 9.5 Emx2-/-Pax6-/- double mutant, the caudal forebrain remained unspecified and subsequently transformed into tectum in a mirror image of the endogenous one.

Expression mechanism of tryptophan hydroxylase 1 in mouse islets during pregnancy.

Serotonin signaling plays key roles in augmentation of pancreatic ß-cell function during pregnancy. Increased expression of tryptophan hydroxylase 1 (Tph1), a rate-limiting enzyme for serotonin synthesis by lactogenic hormones, is involved in this phenomenon. To investigate its mechanisms, we here performed 5'-RACE and identified ß-cell-specific transcription initiation sites for Tph1. Prolactin enhanced the expression of mRNA containing these exons; however, reporter gene plasmids containing the proximal 5'-flanking region of these exons did not show prolactin responsiveness in MIN6 cells. Prolactin-induced Tph1 expression was inhibited by a Jak2 inhibitor and was partially inhibited by an MEK1/2 or PI3K inhibitor. Therefore, we analyzed interferon γ-activated sequences (GAS) and found GAS-A about 9-kbp upstream of the transcription start site. The reporter gene plasmid containing the GAS-A region linked to a heterologous promoter showed increased promoter activity by prolactin, which was inhibited by the forced expression of a dominant-negative mutant form of Stat5A and a Jak2 inhibitor. Chromatin immunoprecipitation analysis showed that prolactin treatment augmented Stat5 binding to the GAS-A region in MIN6 cells, as well as in isolated mouse islets, and that Stat5 recognized the GAS-A region in pregnant mouse islets. In addition, the transactivation activity of Stat5 was enhanced by prolactin through the Erk and PI3K pathways in MIN6 cells. Finally, serotonin expression was attenuated in islets of ß-cell-specific Stat5-deficient mice compared with that of control littermates during pregnancy. Our findings suggest that prolactin-induced Tph1 expression is mediated by the activation of Jak2/Stat5, Erk, and PI3K pathways in ß cells.

Human IAPP-induced pancreatic ß cell toxicity and its regulation by autophagy.

Pancreatic islets in patients with type 2 diabetes mellitus (T2DM) are characterized by loss of ß cells and formation of amyloid deposits derived from islet amyloid polypeptide (IAPP). Here we demonstrated that treatment of INS-1 cells with human IAPP (hIAPP) enhances cell death, inhibits cytoproliferation, and increases autophagosome formation. Furthermore, inhibition of autophagy increased the vulnerability of ß cells to the cytotoxic effects of hIAPP. Based on these in vitro findings, we examined the pathogenic role of hIAPP and its relation to autophagy in hIAPP-knockin mice. In animals fed a standard diet, hIAPP had no toxic effects on ß cell function; however, hIAPP-knockin mice did not exhibit a high-fat-diet-induced compensatory increase in ß cell mass, which was due to limited ß cell proliferation and enhanced ß cell apoptosis. Importantly, expression of hIAPP in mice with a ß cell-specific autophagy defect resulted in substantial deterioration of glucose tolerance and dispersed cytoplasmic expression of p62-associated toxic oligomers, which were otherwise sequestrated within p62-positive inclusions. Together, our results indicate that increased insulin resistance in combination with reduced autophagy may enhance the toxic potential of hIAPP and enhance ß cell dysfunction and progression of T2DM.

Exendin-4 improves ß-cell function in autophagy-deficient ß-cells.

Autophagy is cellular machinery for maintenance of ß-cell function and mass. The implication of autophagy failure in ß-cells on the pathophysiology of type 2 diabetes and its relation to the effect of treatment of diabetes remains elusive. Here, we found increased expression of p62 in islets of db/db mice and patients with type 2 diabetes mellitus. Treatment with exendin-4, a glucagon like peptide-1 receptor agonist, improved glucose tolerance in db/db mice without significant changes in p62 expression in ß-cells. Also in ß-cell-specific Atg7-deficient mice, exendin-4 efficiently improved blood glucose level and glucose tolerance mainly by enhanced insulin secretion. In addition, we found that exendin-4 reduced apoptotic cell death and increased proliferating cells in the Atg7-deficient islets, and that exendin-4 counteracted thapsigargin-induced cell death of isolated islets augmented by autophagy deficiency. Our results suggest the potential involvement of reduced autophagy in ß-cell dysfunction in type 2 diabetes. Without altering the autophagic state in ß-cells, exendin-4 improves glucose tolerance associated with autophagy deficiency in ß-cells. This is mainly achieved through augmentation of insulin secretion. In addition, exendin-4 prevents apoptosis and increases the proliferation of ß-cells associated with autophagy deficiency, also without altering the autophagic machinery in ß-cells.

The diabetes-susceptible gene SLC30A8/ZnT8 regulates hepatic insulin clearance.

Recent genome-wide association studies demonstrated that common variants of solute carrier family 30 member 8 gene (SLC30A8) increase susceptibility to type 2 diabetes. SLC30A8 encodes zinc transporter-8 (ZnT8), which delivers zinc ion from the cytoplasm into insulin granules. Although it is well known that insulin granules contain high amounts of zinc, the physiological role of secreted zinc remains elusive. In this study, we generated mice with ß cell-specific Slc30a8 deficiency (ZnT8KO mice) and demonstrated an unexpected functional linkage between Slc30a8 deletion and hepatic insulin clearance. The ZnT8KO mice had low peripheral blood insulin levels, despite insulin hypersecretion from pancreatic ß cells. We also demonstrated that a substantial amount of the hypersecreted insulin was degraded during its first passage through the liver. Consistent with these findings, ZnT8KO mice and human individuals carrying rs13266634, a major risk allele of SLC30A8, exhibited increased insulin clearance, as assessed by c-peptide/insulin ratio. Furthermore, we demonstrated that zinc secreted in concert with insulin suppressed hepatic insulin clearance by inhibiting clathrin-dependent insulin endocytosis. Our results indicate that SLC30A8 regulates hepatic insulin clearance and that genetic dysregulation of this system may play a role in the pathogenesis of type 2 diabetes.

Conophylline suppresses pancreatic stellate cells and improves islet fibrosis in Goto-Kakizaki rats.

Activin A is a differentiation factor for ß-cells and is effective to promote ß-cell neogenesis. Activin A is also an autocrine activator of pancreatic stellate cells, which play a critical role in fibrogenesis of the pancreas. Conophylline (CnP) is a natural compound, which reproduces the effect of activin on ß-cell differentiation and promotes ß-cell neogenesis when administered in vivo. However, its effect on stellate cells is not known. We therefore investigated the effect of CnP on stellate cells both in vitro and in vivo. Unlike activin A, CnP inhibited activation of cultured stellate cells and reduced the production of collagen. We then analyzed the involvement of stellate cells in islet fibrosis in Goto-Kakizaki (GK) rats, a model of type 2 diabetes mellitus. In pancreatic sections obtained from 6-wk-old GK rats, CD68-positive macrophages and glial fibrillary acidic protein- and α-smooth muscle actin-positive stellate cells infiltrated into islets. Later, the number of macrophages was increased, and the α-smooth muscle actin staining of stellate cells became stronger, indicating the involvement of stellate cells in islet fibrosis in GK rats. When CnP was administered orally for 4 wk, starting from 6 wk of age, invasion of stellate cells and macrophages was markedly reduced and islet fibrosis was significantly improved. The insulin content was twice as high in CnP-treated rats. These results indicate that CnP exerts antifibrotic actions both in vitro and in vivo and improves islet fibrosis in Goto-Kakizaki rats.