Calpain-Dependent Degradation of Nucleoporins Contributes to Motor Neuron Death in a Mouse Model of Chronic Excitotoxicity.

Glutamate-mediated excitotoxicity induces neuronal death by altering various intracellular signaling pathways and is implicated as a common pathogenic pathway in many neurodegenerative diseases. In the case of motor neuron disease, there is significant evidence to suggest that the overactivation of AMPA receptors due to deficiencies in the expression and function of glial glutamate transporters GLT1 and GLAST plays an important role in the mechanisms of neuronal death. However, a causal role for glial glutamate transporter dysfunction in motor neuron death remains unknown. Here, we developed a new animal model of excitotoxicity by conditionally deleting astroglial glutamate transporters GLT1 and GLAST in the spinal cords of mice (GLAST+/-/GLT1-cKO). GLAST+/-/GLT1-cKO mice (both sexes) exhibited nuclear irregularity and calpain-mediated degradation of nuclear pore complexes (NPCs), which are responsible for nucleocytoplasmic transport. These abnormalities were associated with progressive motor neuron loss, severe paralysis, and shortened lifespan. The nuclear export inhibitor KPT-350 slowed but did not prevent motor neuron death, whereas long-term treatment of the AMPA receptor antagonist perampanel and the calpain inhibitor SNJ-1945 had more persistent beneficial effects. Thus, NPC degradation contributes to AMPA receptor-mediated excitotoxic motor neuronal death, and preventing NPC degradation has robust protective effects. Normalization of NPC function could be a novel therapeutic strategy for neurodegenerative disorders in which AMPA receptor-mediated excitotoxicity is a contributory factor.SIGNIFICANCE STATEMENT Despite glial glutamate transporter dysfunction leading to excitotoxicity has been documented in many neurological diseases, it remains unclear whether its dysfunction is a primary cause or secondary outcome of neuronal death at disease state. Here we show the combined loss of glial glutamate transporters GLT1 and GLAST in spinal cord caused motor neuronal death and hindlimb paralysis. Further, our novel mutant exhibits the nuclear irregularities and calpain-mediated progressive nuclear pore complex degradation. Our study reveals that glial glutamate transporter dysfunction is sufficient to cause motor neuronal death in vivo.

Regulation of expression and trafficking of perforin-2 by LPS and TNF-α.

Perforin-2 is constitutively expressed in macrophages that are required for bacterial control. In this study, we found that perforin-2 is expressed in human macrophages with two isoforms: full-length perforin-2a and a splice variant, perforin-2b. Two isoforms show different subcellular distributions. Perforin-2a was predominantly localized to the membrane of endosome-like vesicles by a C-terminal transmembrane domain. In contrast, the short isoform perforin-2b lacking the transmembrane domain failed to localize to the membrane of vesicles. Furthermore, we determined that the pro-inflammatory stimuli LPS and TNF-α induced perforin-2a expression via the NF-κB pathway and triggered perforin-2a vesicles fusion with lysosomes. On the other hand, we detected the secretion of perforin-2b in response to LPS stimulation. Taken together, our data provide the evidence that membrane-bound and secretory isoforms of perforin-2 are present in human macrophages and may play important roles in immune defense.

Expression of programmed cell death ligand 1 is associated with poor overall survival in patients with diffuse large B-cell lymphoma.

Programmed cell death ligand 1 (PD-L1) is expressed on both select diffuse large B-cell lymphoma (DLBCL) tumor cells and on tumor-infiltrating nonmalignant cells. The programmed cell death 1 (PD-1)/PD-L1 pathway inhibits host antitumor responses; however, little is known about how this pathway functions in the tumor microenvironment. The aim of this study was to determine the clinicopathological impact of PD-L1(+) DLBCL. We performed PD-L1/PAX5 double immunostaining in 1253 DLBCL biopsy samples and established a new definition of PD-L1(+) DLBCL. We also defined the criteria for microenvironmental PD-L1(+) (mPD-L1(+)) DLBCL (ie, PD-L1(-) DLBCL in which PD-L1(+) nonmalignant cells are abundant in the tumor microenvironment). Of the 273 patients whose clinical information was available, quantitative analysis of PD-1(+) tumor-infiltrating lymphocytes (TILs) was performed. The prevalence rates of PD-L1(+) and mPD-L1(+) DLBCL were 11% and 15.3%, respectively. Both PD-L1(+) and mPD-L1(+) DLBCL were significantly associated with non-germinal center B-cell (GCB) type and Epstein-Barr virus positivity. The number of PD-1(+) TILs was significantly higher in GCB-type tumors and lower in mPD-L1(-) and PD-L1(+) DLBCL. Patients with PD-L1(+) DLBCL had inferior overall survival (OS) compared with that in patients with PD-L1(-) DLBCL (P = .0009). In contrast, there was no significant difference in OS between mPD-L1(+) and mPD-L1(-) DLBCL (P = .31). The expression of PD-L1 maintained prognostic value for OS in multivariate analysis (P = .0323). This is the first report describing the clinicopathological features and outcomes of PD-L1(+) DLBCL. Immunotherapy targeting the PD-1/PD-L1 pathway should be considered in this distinct DLBCL subgroup.

Spalt-like transcription factor 4 immunopositivity is associated with epithelial cell adhesion molecule expression in combined hepatocellular carcinoma and cholangiocarcinoma.

AIM: Combined hepatocellular carcinoma (HCC) and cholangiocarcinoma (CC) (cHCC-CC) is a rare biphasic liver cancer. Recent studies have demonstrated that cHCC-CC originates from hepatic progenitor cells (HPCs). Spalt-like transcription factor 4 (SALL4) is a marker for a progenitor subclass of HCC with an aggressive phenotype. However, little has been revealed about SALL4 expression in cHCC-CC. The aims of this study were to report SALL4 immunopositivity and the results of clinicopathological analysis in cHCC-CC, and to examine the two different nuclear immunostaining patterns for SALL4. METHODS AND RESULTS: We defined the diffuse finely granular nuclear immunostaining pattern as immunopositive for SALL4; this was observed in eight (8.9%) of 90 cHCC-CCs. SALL4 immunopositivity was significantly associated with immunopositivity for α-fetoprotein, glypican 3, and epithelial cell adhesion molecule (EpCAM). There was no relationship between SALL4 immunopositivity and prognosis. We confirmed SALL4 mRNA expression in samples with a punctuate/clumped immunostaining pattern, which showed a significantly lower rate of immunopositivity for EpCAM than those with a diffuse finely granular pattern. CONCLUSIONS: SALL4 immunopositivity is not a prognostic factor in cHCC-CC; however, it is associated with α-fetoprotein, glypican 3 and EpCAM immunopositivity, indicating the mechanism of carcinogenesis. Further study is necessary to interpret the immunostaining pattern for SALL4.

Short- and long-term outcomes of endoscopic resection of rectal neuroendocrine tumours: analyses according to the WHO 2010 classification.

OBJECTIVE: Although the World Health Organisation (WHO) defined a novel classification of gastroenteropancreatic neuroendocrine tumours (NETs) in 2010, indications for endoscopic resection of rectal NETs in the guidelines were based on evidence accumulated for carcinoid tumours defined by a previous classification. This study was designed to clarify indications for endoscopic resection of rectal NETs corresponding to the new WHO classifications. MATERIAL AND METHODS: One hundred-seventy rectal NETs resected endoscopically from April 2001 to March 2012 were histologically re-classified according to the WHO 2010 criteria. The clinicopathological features of these lesions were analysed, and the short- and long-term outcomes of endoscopic resection were evaluated. RESULTS: Of the 170 rectal NETs, 166 were histopathologically diagnosed as NET G1 and four as NET G2. Thirty-eight tumours (22.4%) were positive for lymphovascular invasion, a percentage higher than expected. Although the curative resection rate was low (65.3%), en bloc (98.8%) and complete (85.9%) resection rates were high. Modified endoscopic mucosal resection (88.0%) and endoscopic submucosal dissection (92.2%) resulted in significantly higher complete resection rates than conventional endoscopic mucosal resection (36.4%). No patient experienced tumour recurrence, despite the low curative resection rate. CONCLUSION: Despite the low curative resection rate, prognosis after endoscopic resection of rectal NETs was excellent. Prospective large-scale, long-term studies are required to determine whether NET G2 and tumours >1 cm should be included in the indication for endoscopic resection and whether tumours with lymphovascular invasion can be followed up without additional surgery.

Clinical characteristics associated with esophageal motility function.

BACKGROUND AND AIM: Esophageal motility disorders (EMDs) affect coordinated esophageal contractility. Recent developments in high-resolution manometry have improved diagnosis of EMDs; however, the etiology of EMDs remains to be determined. This study aimed to determine which clinical characteristics are associated with esophageal motility. METHODS: From May 2013 to July 2014, 97 patients (54 women, 43 men; age, 16-89 years) with suspected EMDs were assessed by high-resolution manometry in Kyushu University Hospital. Esophageal motility was evaluated by measuring the distal contractile integral (DCI), basal lower esophageal sphincter pressure, and integrated relaxation pressure (IRP). Data on age, gender, body mass index (BMI), Brinkman Index, and blood tests were retrospectively collected and analyzed. RESULTS: Fifty patients were diagnosed as normal, nine with achalasia, twelve with esophagogastric junction outflow obstruction, four with distal esophageal spasm, one with jackhammer esophagus, six with absent peristalsis, ten with frequent failed peristalsis, and five with weak peristalsis. The median DCI was 1229.0 mmHg-s-cm, the median basal lower esophageal sphincter pressure was 25.3 mmHg, and the median IRP was 9.6 mmHg. Patients with major motility disorders were excluded from analysis. By multivariate regression analysis, BMI (P = 0.029) and total cholesterol (P = 0.023) were negatively associated with DCI, while BMI (P = 0.007) was negatively associated with IRP and glucose (P = 0.044) was positively associated with IRP. CONCLUSIONS: Both BMI and total cholesterol could be highly predictive factors for esophageal body contractility, while BMI and glucose could be predictive factors for lower esophageal sphincter contractile function.

Eplerenone improves carotid intima-media thickness (IMT) in patients with primary aldosteronism.

Primary aldosteronism (PA) is associated with a higher rate of cardiovascular events than essential hypertension. Although adrenalectomy has been reported to reduce carotid intima-media thickness (IMT) in patients with PA, the effects of the selective aldosterone blocker, eplerenone, on vascular damage in these patients remains unclear. To evaluate the effects of eplerenone on vascular status in PA patients, we sequentially measured carotid IMT (using computer software to calculate an average IMT for accurate and reproducible evaluation) in 22 patients including 8 patients treated by unilateral adrenalectomy and 14 patients treated with eplerenone for 12 months. Patients who underwent adrenalectomy showed significant reductions in aldosterone concentration (from 345 ± 176 pg/mL to 67 ± 34 pg/mL; P<0.01) and IMT (from 0.67 ± 0.07 mm to 0.63 ± 0.09 mm; P<0.05) 6 months after surgery. Patients treated with eplerenone showed significant reductions in IMT from baseline (0.75 ± 0.10 mm) to 6 (0.71 ± 0.11 mm; P<0.05) and 12 (0.65 ± 0.09 mm; P<0.01) months, although plasma aldosterone level increased significantly, from 141 ± 105 pg/mL to 207 ± 98 pg/mL (P<0.05). Eplerenone treatment of patients with PA reduces blood pressure, increases serum potassium level, and improves vascular status. Carotid IMT may be a useful marker for evaluating the effectiveness of eplerenone in patients with PA.

Endoscopic Submucosal Dissection is Feasible for Very Elderly Patients with Early Gastric Cancer : Comparison of Short-Term and Long-Term Outcomes in Very Elderly and Non-Elderly Patients.

Background/Aims: Endoscopic submucosal dissection (ESD) has become a standard procedure for the resection of early gastric cancer (EGC). However, the feasibility of ESD for very elderly patients, aged ≥ 80 years, has not been determined. Methodology: The study population included 67 non-elderly (NE) patients aged ≤ 65 years (80 lesions) and 22 very elderly (VE) patients ≥ 80 years (26 lesions) with EGC who underwent ESD and met the criteria for absolute or expanded indications. Eighteen patients (18 lesions) who underwent ESD but did not meet the criteria for absolute and expanded indications were defined as the outside the indications (OI) group. Results: En bloc and complete resection rates were excellent in both the VE and NE groups, without differing significantly. Although the rates of ischemic heart disease and antithrombotic agent use were higher in the VE than in the NE group, procedure-related complication rates did not differ significantly. Of the seven very elderly patients in the OI group, two underwent additional gastrectomy, and the other five were followed-up without surgery. No patient in any group experienced local recurrence, metastasis or disease-specific death. Conclusions: Short- and long-term outcomes of ESD for VE patients with EGC were favorable and did not differ significantly from outcomes in NE patients. ESD may therefore be a good therapeutic option for both VE and NE patients with EGC.

Glial dysfunction in the mouse habenula causes depressive-like behaviors and sleep disturbance.

The lateral habenula (LHb) regulates the activity of monoaminergic neurons in the brainstem. This area has recently attracted a surge of interest in psychiatry because studies have reported the pathological activation of the habenula in patients with major depression and in animal models. The LHb plays a significant role in the pathophysiology of depression; however, how habenular neurons are activated to cause various depression symptoms, such as reduced motivation and sleep disturbance, remain unclear. We hypothesized that dysfunctional astrocytes may cause LHb hyperactivity due to the defective uptake activity of extracellular glutamate, which induces depressive-like behaviors. We examined the activity of neurons in habenular pathways and performed behavioral and sleep analyses in mice with pharmacological and genetic inhibition of the activity of the glial glutamate transporter GLT-1 in the LHb. The habenula-specific inhibition of GLT-1 increased the neuronal firing rate and the level of c-Fos expression in the LHb. Mice with reduced GLT-1 activity in the habenula exhibited a depressive-like phenotype in the tail suspension and novelty-suppressed feeding tests. These animals also displayed increased susceptibility to chronic stress, displaying more frequent avoidant behavior without affecting locomotor activity in the open-field test. Intriguingly, the mice showed disinhibition of rapid eye movement sleep, which is a characteristic sleep pattern in patients with depression. These results provide evidence that disrupting glutamate clearance in habenular astrocytes increases neuronal excitability and depressive-like phenotypes in behaviors and sleep.

Disruption of mitochondrial fission in the liver protects mice from diet-induced obesity and metabolic deterioration.

AIM/HYPOTHESIS: Mitochondria and the endoplasmic reticulum (ER) physically interact by close structural juxtaposition, via the mitochondria-associated ER membrane. Inter-organelle communication between the ER and mitochondria has been shown to regulate energy metabolism and to be central to the modulation of various key processes such as ER stress. We aimed to clarify the role of mitochondrial fission in this communication. METHODS: We generated mice lacking the mitochondrial fission protein dynamin-related protein 1 (DRP1) in the liver (Drp1LiKO mice). RESULTS: Drp1LiKO mice showed decreased fat mass and were protected from high-fat diet (HFD)-induced obesity. Analysis of liver gene expression profiles demonstrated marked elevation of ER stress markers. In addition, we observed increased expression of the fibroblast growth factor 21 (FGF21) gene through induction of activating transcription factor 4, master regulator of the integrated stress response. CONCLUSIONS/INTERPRETATION: Disruption of mitochondrial fission in the liver provoked ER stress, while inducing the expression of FGF21 to increase energy expenditure and protect against HFD-induced obesity.

Fatty acid-binding protein 5 regulates diet-induced obesity via GIP secretion from enteroendocrine K cells in response to fat ingestion.

Gastric inhibitory polypeptide (GIP) is an incretin released from enteroendocrine K cells in response to nutrient intake, especially fat. GIP is one of the contributing factors inducing fat accumulation that results in obesity. A recent study shows that fatty acid-binding protein 5 (FABP5) is expressed in murine K cells and is involved in fat-induced GIP secretion. We investigated the mechanism of fat-induced GIP secretion and the impact of FABP5-related GIP response on diet-induced obesity (DIO). Single oral administration of glucose and fat resulted in a 40% reduction of GIP response to fat but not to glucose in whole body FABP5-knockout (FABP5(-/-)) mice, with no change in K cell count or GIP content in K cells. In an ex vivo experiment using isolated upper small intestine, oleic acid induced only a slight increase in GIP release, which was markedly enhanced by coadministration of bile and oleic acid together with attenuated GIP response in the FABP5(-/-) sample. FABP5(-/-) mice exhibited a 24% reduction in body weight gain and body fat mass under a high-fat diet compared with wild-type (FABP5(+/+)) mice; the difference was not observed between GIP-GFP homozygous knock-in (GIP(gfp/gfp))-FABP5(+/+) mice and GIP(gfp/gfp)-FABP5(-/-) mice, in which GIP is genetically deleted. These results demonstrate that bile efficiently amplifies fat-induced GIP secretion and that FABP5 contributes to the development of DIO in a GIP-dependent manner.

Dehydroepiandrosterone-enhanced dual specificity protein phosphatase (DDSP) prevents diet-induced and genetic obesity.

Dehydroepiandrosterone (DHEA) exerts a wide variety of therapeutic effects against medical disorders, such as diabetes and obesity. However, the molecular basis of DHEA action remains to be clarified. Previously, we reported that DHEA-enhanced dual specificity protein phosphatase, designated DDSP, is one of the target molecules of DHEA. To examine the role of DDSP in DHEA signaling, we generated mice that carry a DDSP transgene in which expression is driven by the CAG promoter (DDSP-Tg). DDSP-Tg mice weighed significantly less than wild-type (WT) control mice when a high fat diet was supplied (p < 0.01). No difference in food-intake or locomotor activity was found between DDSP-Tg and WT mice. Oxygen consumption of DDSP-Tg mice was higher than that of WT mice (p < 0.01), which suggested an increase in basal metabolism in DDSP-Tg mice. To further investigate the role of DDSP in genetic obese mice, DDSP-Tg mice with a db/db background were generated (DDSP-Tg db/db). We observed cancellation of obesity by the db/db mutation and development of a cachexic phenotype in DDSP-Tg db/db mice. In conclusion, our study shows that expression of DDSP leads to prevention of diet-induced and genetic (db/db) obesity. Anti-obese effects of DHEA might be mediated through DDSP, which might be a therapeutic target for intervention of obesity.

Dietary vitamin D3 improves postprandial hyperglycemia in aged mice.

Type 2 Diabetes is closely associated with our daily diets and has become a global health problem with an increasing number of patients. Recent observational and randomized studies on vitamin D3 suggested that higher plasma 25-hydroxyvitamin D3 [25(OH)D3] concentrations and more vitamin D3 intake are associated with lower risk of type 2 diabetes, which is characterized by postprandial hyperglycemia due to inappropriate glucose stimulated insulin secretion (GSIS) and its age-dependent increase of onset. However, rapid action of dietary vitamin D3 on the postprandial glucose profile has not been analyzed. When vitamin D3 is orally ingested in mice aged 12-14 weeks during an oral glucose tolerance test (OGTT), the serum glucose profile was not changed. In contrast, when OGTT was performed with old mice aged 30-34 weeks, the glucose profile was dramatically improved with increased insulin secretion, suggesting that orally ingested vitamin D3 potentiated GSIS in aged mice. Interestingly, there was also a significant increase in plasma GLP-1 in these aged mice. Our results suggest that orally ingested dietary vitamin D3 in aged mice improves glucose metabolism as a GLP-1 enhancer.

Suitability of the expanded indication criteria for the treatment of early gastric cancer by endoscopic submucosal dissection: Japanese multicenter large-scale retrospective analysis of short- and long-term outcomes.

OBJECTIVE: The criteria for endoscopic resection for early gastric cancer include absolute and expanded indications. Consensus already exists for the absolute indications. However, the suitability of the expanded indications must be validated by long-term outcome analyses since such lesions have only recently become resectable with the development of endoscopic submucosal dissection. The aim of this study is to clarify the suitability of the expanded indications for the treatment of early gastric cancer with endoscopic submucosal dissection. MATERIALS AND METHODS: The medical records of 1161 patients with early gastric cancers (1332 lesions) treated by endoscopic submucosal dissection and meeting the criteria for absolute or expanded indications without additional treatment with gastrectomy were divided into absolute indication group or expanded indication group. RESULTS: Complete resection rates were 96.4% and 93.4% in absolute and expanded indication groups, respectively, with no significant differences between the groups. Delayed bleeding rates were significantly higher in the expanded indication group, whereas all cases were successfully managed conservatively. The 5-year overall survival and recurrence-free rates were 93.7%/99.77% and 90.49%/98.90% in the absolute and the expanded indication groups, respectively, with no significant differences between the groups for either measure. Multivariate analyses revealed that affected horizontal margin and tumor location were independent predictive factors for recurrence. CONCLUSION: The expanded indication group showed excellent post-endoscopic submucosal dissection short-term and long-term outcomes compared with the absolute indications group, demonstrating that expanded indications are suitable for endoscopic submucosal dissection for early gastric cancer.

SMAD2 disruption in mouse pancreatic beta cells leads to islet hyperplasia and impaired insulin secretion due to the attenuation of ATP-sensitive K+ channel activity.

AIMS/HYPOTHESIS: The TGF-ß superfamily of ligands provides important signals for the development of pancreas islets. However, it is not yet known whether the TGF-ß family signalling pathway is required for essential islet functions in the adult pancreas. METHODS: To identify distinct roles for the downstream components of the canonical TGF-ß signalling pathway, a Cre-loxP system was used to disrupt SMAD2, an intracellular transducer of TGF-ß signals, in pancreatic beta cells (i.e. Smad2ß knockout [KO] mice). The activity of ATP-sensitive K(+) channels (KATP channels) was recorded in mutant beta cells using patch-clamp techniques. RESULTS: The Smad2ßKO mice exhibited defective insulin secretion in response to glucose and overt diabetes. Interestingly, disruption of SMAD2 in beta cells was associated with a striking islet hyperplasia and increased pancreatic insulin content, together with defective glucose-responsive insulin secretion. The activity of KATP channels was decreased in mutant beta cells. CONCLUSIONS/INTERPRETATION: These results suggest that in the adult pancreas, TGF-ß signalling through SMAD2 is crucial for not only the determination of beta cell mass but also the maintenance of defining features of mature pancreatic beta cells, and that this involves modulation of KATP channel activity.

Prolyl hydroxylase domain protein 2 plays a critical role in diet-induced obesity and glucose intolerance.

BACKGROUND: Recent studies suggest that the oxygen-sensing pathway consisting of transcription factor hypoxia-inducible factor and prolyl hydroxylase domain proteins (PHDs) plays a critical role in glucose metabolism. However, the role of adipocyte PHD in the development of obesity has not been clarified. We examined whether deletion of PHD2, the main oxygen sensor, in adipocytes affects diet-induced obesity and associated metabolic abnormalities. METHODS AND RESULTS: To delete PHD2 in adipocyte, PHD2-floxed mice were crossed with aP2-Cre transgenic mice (Phd2(f/f)/aP2-Cre). Phd2(f/f)/aP2-Cre mice were resistant to high-fat diet-induced obesity (36.7±1.7 versus 44.3±2.0 g in control; P<0.01) and showed better glucose tolerance and homeostasis model assessment-insulin resistance index than control mice (3.6±1.0 versus 11.1±2.1; P<0.01). The weight of white adipose tissue was lighter (epididymal fat, 758±35 versus 1208±507 mg in control; P<0.01) with a reduction in adipocyte size. Macrophage infiltration into white adipose tissue was also alleviated in Phd2(f/f)/aP2-Cre mice. Target genes of hypoxia-inducible factor, including glycolytic enzymes and adiponectin, were upregulated in adipocytes of Phd2(f/f)/aP2-Cre mice. Lipid content was decreased and uncoupling protein-1 expression was increased in brown adipose tissue of Phd2(f/f)/aP2-Cre mice. Knockdown of PHD2 in 3T3L1 adipocytes induced a decrease in the glucose level and an increase in the lactate level in the supernatant with upregulation of glycolytic enzymes and reduced lipid accumulation. CONCLUSIONS: PHD2 in adipose tissue plays a critical role in the development of diet-induced obesity and glucose intolerance. PHD2 might be a novel target molecule for the treatment of obesity and associated metabolic abnormalities.

Maternal high-fat diet induces insulin resistance and deterioration of pancreatic ß-cell function in adult offspring with sex differences in mice.

Intrauterine environment may influence the health of postnatal offspring. There have been many studies on the effects of maternal high-fat diet (HFD) on diabetes and glucose metabolism in offspring. Here, we investigated the effects in male and female offspring. C57/BL6J mice were bred and fed either control diet (CD) or HFD from conception to weaning, and offspring were fed CD or HFD from 6 to 20 wk. At 20 wk, maternal HFD induced glucose intolerance and insulin resistance in offspring. Additionally, liver triacylglycerol content, adipose tissue mass, and inflammation increased in maternal HFD. In contrast, extending previous observations, insulin secretion at glucose tolerance test, islet area, insulin content, and PDX-1 mRNA levels in isolated islets were lower in maternal HFD in males, whereas they were higher in females. Oxidative stress in islets increased in maternal HFD in males, whereas there were no differences in females. Plasma estradiol levels were lower in males than in females and decreased in offspring fed HFD and also decreased by maternal HFD, suggesting that females may be protected from insulin deficiency by inhibiting oxidative stress. In conclusion, maternal HFD induced insulin resistance and deterioration of pancreatic ß-cell function, with marked sex differences in adult offspring accompanied by adipose tissue inflammation and liver steatosis. Additionally, our results demonstrate that potential mechanisms underlying sex differences in pancreatic ß-cell function may be related partially to increases in oxidative stress in male islets and decreased plasma estradiol levels in males.

PSCs and GLP-1R: occurrence in normal pancreas, acute/chronic pancreatitis and effect of their activation by a GLP-1R agonist.

There is increasing concern about the development of pancreatitis in patients with diabetes mellitus who received long-term glucagon-like peptide-1 (GLP-1) analog treatment. Its pathogenesis is unknown. The effects of GLP-1 agonists on pancreatic endocrine cells are well studied; however, there is little information on effects on other pancreatic tissues that might be involved in inflammatory processes. Pancreatic stellate cells (PSCs) can have an important role in pancreatitis, secreting various inflammatory cytokines/chemokines, as well as collagen. In this study, we investigated GLP-1R occurrence in normal pancreas, acute pancreatitis (AP)/chronic pancreatitis (CP), and the effects of GLP-1 analog on normal PSCs, their ability to stimulate inflammatory mediator secretion or proliferation. GLP-1 receptor (GLP-1R) expression/localization in normal pancreas and pancreatitis (AP/CP) tissues were evaluated with histological/immunohistochemical analysis. PSCs were isolated from male Wistar rats. GLP-1R expression and effects of GLP-1 analog on activated PSCs was examined with real-time PCR, MTS assays and western blotting. In normal pancreas, pancreatic ß cells expressed GLP-1R, with only low expression in acinar cells, whereas in AP or CP, acinar cells, ductal cells and activated PSCs expressed GLP-1R. With activation of normal PSCs, GLP-1R is markedly increased, as is multiple other incretin-related receptors. The GLP-1 analog, liraglutide, did not induce inflammatory genes expression in activated PSCs, but induced proliferation. Liraglutide activated multiple signaling cascades in PSCs, and the extracellular signal-regulated kinase pathway mediated the PSCs proliferation. GLP-1Rs are expressed in normal pancreas and there is marked enhanced expression in AP/CP. GLP-1-agonist induced cell proliferation of activated PSCs without increasing release of inflammatory mediators. These results suggest chronic treatment with GLP-1R agonists could lead to proliferation/chronic activation of PSCs, which may lead to important effects in the pancreas.

Serum chromogranin A is a useful marker for Japanese patients with pancreatic neuroendocrine tumors.

Although chromogranin A (CGA) is a useful marker for pancreatic neuroendocrine tumors (pNET) in the West, its usefulness in Japanese populations is unclear. To assess this, we evaluated the serum CGA levels in 189 patients with various pancreatic diseases, including proven pNET (n = 69), pancreatic cancer (PC) (n = 50), chronic pancreatitis (CP) (n = 50) and autoimmune pancreatitis (AIP) (n = 20), and 112 normal controls (controls) using an ELISA kit. The mean CGA level of patients with pNET was significantly higher than any of the other groups (407.8 ± 984.6 ng/mL [pNET] vs 91.8 ± 101.8 ng/mL [PC], 93.6 ± 57.5 ng/mL [CP], 69.9 ± 52.4 ng/mL [AIP] and 62.5 ± 48.3 ng/mL [controls]). Limiting the analysis to patients not using proton pump inhibitors (PPI), the CGA level of patients with PC or CP was not significantly different compared with the controls. Discriminant analysis revealed that the best cut-off value of CGA to distinguish patients with pNET from the controls was 78.7 ng/mL, with a sensitivity and specificity of 53.6% and 78.6%, respectively. In patients with pNET, significant factors associating with elevated CGA levels were tumor classification, tumor size, and the presence of liver metastases in univariate analysis as well as PPI use and the presence of liver metastases in multivariate analysis. We show that CGA is a useful marker for diagnosing pNET in Japanese populations and for distinguishing patients with pNET from patients with other pancreatic diseases. The increased use of CGA in Japan will likely be a helpful tool in managing these patients, as found in the West.