Destruction of pancreatic β-cells by transgenic induction of prostaglandin e2 in the islets

金沢大学がん研究所附属がん幹細胞研究センター　Type 2 diabetes mellitus is characterized by insulin resistance of peripheral tissues and dysfunction of pancreatic β-cells. Furthermore, the number of pancreatic β-cells decreases as a secondary effect of advanced type 2 diabetes, although the molecular mechanism has not been elucidated. Recently, it has been shown that hyperglycemic conditions induce the expression of cyclooxygenase-2 in pancreatic islets and increase the downstream product prostaglandin E2 (PGE2). To investigate whether high glucose-induced PGE2 has an adverse effect on pancreatic β-cells, we generated transgenic mice (RIP-C2mE) that express cyclooxygenase-2 and microsomal prostaglandin E synthase-1 in their β-cells using the rat insulin-2 gene promoter (RIP). The homozygous RIP-C2mE (Tg/Tg) mice showed severe hyperglycemia from six weeks of age. Although the heterozygous RIP-C2mE (Tg/-) mice showed normal blood glucose levels throughout their lifetime, this level increased significantly compared with that of wild-type mice when glucose was loaded. The relative number of β-cells to the total islet cell number was reduced to 54 and 14% in the RIP-C2mE (Tg/-) and (Tg/Tg) mice, respectively, whereas that in the wild-type mice was 84%. Importantly, the proliferation rate in the islets of the RIP-C2mE (Tg/Tg) mice at four weeks of age decreased significantly in comparison to that in the wild-type mice. Because β-cells replicate not only during the postnatal period but also in the adult pancreas at a basal level, it is possible that increased PGE2 signaling thus contributes to the reduction of the pancreatic β-cell mass through inhibition of proliferation, thereby aggravating diabetes further. © 2006 by The American Society for Biochemistry and Molecular Biology, Inc

Hypergravity induces expression of cyclooxygenase-2 in the heart vessels

金沢大学がん研究所Cyclooxygenase-2 (COX-2), a rate-limiting enzyme for prostaglandin biosynthesis, is induced by various stimuli including mechanical stress and plays important roles in pathophysiological conditions. For example, gravitational stress has been shown to induce expression of COX-2 in bone tissues, which is essential for bone homeostasis. To investigate whether COX-2 is induced by gravitational loading in other tissues than bone, we exposed mice to hypergravity at 2G and 3G for 4 h. We demonstrate here that COX-2 is induced in the mouse heart vessels by hypergravity. Moreover, hypoxia-inducible factor (HIF)-1α and its downstream genes such as inducible nitric oxide synthase, vascular endothelial growth factor, and heme oxygenase-1 were induced in the heart simultaneously, while none of these genes were induced in the COX-2(−/−) mouse heart. Therefore, COX-2 induced in the heart helps protect the heart function against hypoxia under hypergravity condition through HIF-1α induction

Loss of pancreas upon activated Wnt signaling is concomitant with emergence of gastrointestinal identity

Organ formation is achieved through the complex interplay between signaling pathways and transcriptional cascades. The canonical Wnt signaling pathway plays multiple roles during embryonic development including patterning, proliferation and differentiation in distinct tissues. Previous studies have established the importance of this pathway at multiple stages of pancreas formation as well as in postnatal organ function and homeostasis. In mice, gain-of-function experiments have demonstrated that activation of the canonical Wnt pathway results in pancreatic hypoplasia, a phenomenon whose underlying mechanisms remains to be elucidated. Here, we show that ectopic activation of epithelial canonical Wnt signaling causes aberrant induction of gastric and intestinal markers both in the pancreatic epithelium and mesenchyme, leading to the development of gut-like features. Furthermore, we provide evidence that β -catenin-induced impairment of pancreas formation depends on Hedgehog signaling. Together, our data emphasize the developmental plasticity of pancreatic progenitors and further underscore the key role of precise regulation of signaling pathways to maintain appropriate organ boundaries

Induction of prostaglandin E2 pathway promotes gastric hamartoma development with suppression of bone morphogenetic protein signaling.

金沢大学がん研究所がん幹細胞研究センターMutations in bone morphogenetic protein (BMP) receptor IA (BMPRlA) are responsible for a subset of cases of juvenile polyposis(JF) syndrome that develops hamartomatous tumors in the gastrointestinal tract. Mouse genetic studies have shown thatsuppression of BMP signaling in the intestines causes JP-type hamartoma development. Here, we generated K19-Nog transgenic miceexpressing noggin, a BMP antagonist, in gastric epithelium. However, inhibition of BMP signaling did not cause gastric phenotypes.We thus crossed K19-Nog with K19-C2mE mice that expressed Ptgs2 and Ptges in the stomach to generate compound transgenic mice. Expression of Ptgs2 and Ptges results in prostaglandin E2 (PGE2) biosynthesis, and both enzymes are induced in most human gastrointestinal tumors. Importantly, K19-Nog/ C2mE compound mice developed gastric hamartomas that were morphologicallysimilar to those found in JP with mucin-containing dilated cysts and inflammatory infiltration. Notably, treatment of K19-Nog/C2mE mice with a cyclo-oxygenase-2 inhibitor, celecoxib, significantly reduced tumor size with suppression of angiogenesis, suggesting that induction of the FGE2 pathway together with inhibition of BMP signaling is required for gastric hamartoma development.Moreover, microarray analyses revealed that canonical Wnt signaling target genes were not induced in K19-Nog/ C2mE hamartomas, indicating that BMP inhibition and PGE2 induction lead to gastric hamartoma development indepen- dent of the Wnt/ß-catenin pathway. These results, taken together, suggest that the FGE2 pathway is an effective preventive target against BMP-suppressed gastric hamartomas, as well as for Wnt/ß-catenin-activated adenocarcinomas. ©2009 American Association for Cancer Research

Carcinogenesis in Mouse Stomach by Simultaneous Activation of the Wnt Signaling and Prostaglandin E2 Pathway

金沢大学がん研究所附属がん幹細胞研究センター　Background & Aims: Accumulating evidence indicates that prostaglandin E2 (PGE2), a downstream product of cyclooxygenase 2 (COX-2), plays a key role in gastric tumorigenesis. The Wnt pathway is also suggested to play a causal role in gastric carcinogenesis. However, the molecular mechanism remains poorly understood of how the Wnt and PGE2 pathways contribute to gastric tumorigenesis. To investigate the role of Wnt and PGE2 in gastric cancer, we have generated transgenic mice that activate both pathways and examined their phenotypes. Methods: We constructed K19-Wnt1 transgenic mice expressing Wnt1 in the gastric mucosa using the keratin 19 promoter. We then crossed K19-Wnt1 mice with another transgenic line, K19-C2mE, to obtain K19-Wnt1/C2mE compound transgenic mice. The K19-C2mE mice express COX-2 and microsomal prostaglandin E synthase-1 (mPGES-1) in the stomach, showing an increased gastric PGE2 level. We examined the gastric phenotypes of both K19-Wnt1 and K19-Wnt1/C2mE mice. Results: K19-Wnt1 mice had a significant suppression of epithelial differentiation and developed small preneoplastic lesions consisting of undifferentiated epithelial cells with macrophage accumulation. Importantly, additional expression of COX-2 and mPGES-1 converted the preneoplastic lesions in the K19-Wnt1 mice into dysplastic gastric tumors by 20 weeks of age. Notably, we found mucous cell metaplasia in the glandular stomach of the K19-Wnt1/C2mE mice as early as 5 weeks of age, before the dysplastic tumor development. Conclusions: Wnt signaling keeps the gastric progenitor cells undifferentiated. Simultaneous activation of both Wnt and PGE2 pathways causes dysplastic gastric tumors through the metaplasia-carcinoma sequence. © 2006 American Gastroenterological Association (AGA) Institute

Wnt/beta-catenin signaling controls development of the blood–brain barrier

The blood–brain barrier (BBB) is confined to the endothelium of brain capillaries and is indispensable for fluid homeostasis and neuronal function. In this study, we show that endothelial Wnt/beta-catenin (beta-cat) signaling regulates induction and maintenance of BBB characteristics during embryonic and postnatal development. Endothelial specific stabilization of beta-cat in vivo enhances barrier maturation, whereas inactivation of beta-cat causes significant down-regulation of claudin3 (Cldn3), up-regulation of plamalemma vesicle-associated protein, and BBB breakdown. Stabilization of beta-cat in primary brain endothelial cells (ECs) in vitro by N-terminal truncation or Wnt3a treatment increases Cldn3 expression, BBB-type tight junction formation, and a BBB characteristic gene signature. Loss of beta-cat or inhibition of its signaling abrogates this effect. Furthermore, stabilization of beta-cat also increased Cldn3 and barrier properties in nonbrain-derived ECs. These findings may open new therapeutic avenues to modulate endothelial barrier function and to limit the devastating effects of BBB breakdown

Adenomatous polyposis coli heterozygous knockout mice display hypoactivity and age-dependent working memory deficits

A tumor suppressor gene, Adenomatous polyposis coli (Apc), is expressed in the nervous system from embryonic to adulthood stages, and transmits the Wnt signaling pathway in which schizophrenia susceptibility genes, including T-cell factor 4 (TCF4) and calcineurin (CN), are involved. However, the functions of Apc in the nervous system are largely unknown. In this study, as the first evaluation of Apc function in the nervous system, we have investigated the behavioral significance of the Apc gene, applying a battery of behavioral tests to Apc heterozygous knockout (Apc+/−) mice. Apc+/− mice showed no significant impairment in neurological reflexes or sensory and motor abilities. In various tests, including light/dark transition, open-field, social interaction, eight-arm radial maze, and fear conditioning tests, Apc+/− mice exhibited hypoactivity. In the eight-arm radial maze, Apc+/− mice 6–7 weeks of age displayed almost normal performance, whereas those 11–12 weeks of age showed a severe performance deficit in working memory, suggesting that Apc is involved in working memory performance in an age-dependent manner. The possibility that anemia, which Apc+/− mice develop by 17 weeks of age, impairs working memory performance, however, cannot be excluded. Our results suggest that Apc plays a role in the regulation of locomotor activity and presumably working memory performance

Suppressing TGFβ signaling in regenerating epithelia in an inflammatory microenvironment is sufficient to cause invasive intestinal cancer

Genetic alterations in the TGFβ signaling pathway in combination with oncogenic alterations lead to cancer development in the intestines. However, the mechanisms of TGFβ signaling suppression in malignant progression of intestinal tumors have not yet been fully understood. We have examined ApcΔ716 TGFβr2ΔIEC compound mutant mice that carry mutations in Apc and TGFβr2 genes in the intestinal epithelial cells. We found inflammatory microenvironment only in the invasive intestinal adenocarcinomas but not in noninvasive benign polyps of the same mice. We thus treated simple TGFβr2ΔIEC mice with dextran sodium sulfate (DSS) that causes ulcerative colitis. Importantly, these TGFβr2ΔIEC mice developed invasive colon cancer associated with chronic inflammation.Wealso found that TGFβ signaling is suppressed in human colitis-associated colon cancer cells. In the mouse invasive tumors, macrophages infiltrated and expressed MT1-MMP, causingMMP2activation. These results suggest that inflammatory microenvironment contributes to submucosal invasion of TGFβ signaling-repressed epithelial cells through activation of MMP2. We further found that regeneration was impaired in TGFβr2ΔIEC mice for intestinal mucosa damaged by DSS treatment or X-ray irradiation, resulting in the expansion of undifferentiated epithelial cell population. Moreover, organoids of intestinal epithelial cells cultured from irradiated TGFβr2ΔIEC mice formed "long crypts" in Matrigel, suggesting acquisition of an invasive phenotype into the extracellular matrix. These results, taken together, indicate that a simple genetic alteration in the TGFβ signaling pathway in the inflamed and regenerating intestinal mucosa can cause invasive intestinal tumors. Such a mechanism may play a role in the colon carcinogenesis associated with inflammatory bowel disease in humans