isolation-and-culture-of-mouse-primary-pancreatic-acinar-cells

Isolation and Culture of Mouse Primary Pancreatic Acinar Cells

This protocol permits rapid isolation (in less than 1 hr) of murine pancreatic acini, making it possible to maintain them in culture for more than one ...

Transcriptional intermediary factor 1Î³ (TIF1Î³; alias, TRIM33/RFG7/PTC7/ectodermin) belongs to an evolutionarily conserved family of nuclear factors that have been implicated in stem cell pluripotency, embryonic development, and tumor suppression. TIF1Î³ expression is markedly down-regulated in human pancreatic tumors, and Pdx1-driven Tif1Î³ inactivation cooperates with the Kras(G12D) oncogene in the mouse pancreas to induce intraductal papillary mucinous neoplasms. In this study, we report that aged Pdx1-Cre; LSL-Kras(G12D); Tif1Î³(lox/lox) mice develop pancreatic ductal adenocarcinomas (PDACs), an aggressive and always fatal neoplasm, demonstrating a Tif1Î³ tumor-suppressive function in the development of pancreatic carcinogenesis. Deletion of SMAD4/DPC4 (deleted in pancreatic carcinoma locus 4) occurs in approximately 50% of human cases of PDAC. We, therefore, assessed the genetic relationship between Tif1Î³ and Smad4 signaling in pancreatic tumors and found that Pdx1-Cre; LSL-Kras(G12D); Smad4(lox/lox); Tif1Î³(lox/lox) (alias, KSSTT) mutant mice exhibit accelerated tumor progression. Consequently, Tif1Î³ tumor-suppressor effects during progression from a premalignant to a malignant state in our mouse model of pancreatic cancer are independent of Smad4. These findings establish, for the first time to our knowledge, that Tif1Î³ and Smad4 both regulate an intraductal papillary mucinous neoplasm-to-PDAC sequence through distinct tumor-suppressor programs.

Tif1Î³ suppresses murine pancreatic tumoral transformation by a Smad4-independent pathway.

NUPR1 (nuclear protein 1), also called P8 (molecular mass 8 kDa) or COM1 (candidate of metastasis 1), is involved in the stress response and in cancer progression. In the present study, we investigated whether human NUPR1 expression was regulated by TGFÎ² (transforming growth factor Î²), a secreted polypeptide largely involved in tumorigenesis. We demonstrate that the expression of NUPR1 was activated by TGFÎ² at the transcriptional level. We show that this activation is mediated by the SMAD proteins, which are transcription factors specifically involved in the signalling of TGFÎ² superfamily members. NUPR1 promoter analysis reveals the presence of a functional TGFÎ²-response element binding the SMAD proteins located in the genomic DNA region corresponding to the 5'-UTR (5'-untranslated region). Altogether, the molecular results of the present study, which demonstrate the existence of a TGFÎ²/SMAD/NUPR1 activation cascade, open the way to consider and investigate further a new mechanism enabling TGFÎ² to promote tumorigenesis by inducing stress resistance.

The human NUPR1/P8 gene is transcriptionally activated by transforming growth factor Î² via the SMAD signalling pathway.

Alk4 is a type I receptor that belongs to the transforming growth factor-beta (TGF-Î²) family. It takes part in the signaling of TGF-Î² ligands such as Activins, Gdfs, and Nodal that had been demonstrated to participate in numerous mechanisms ranging from early embryonic development to adult-tissue homeostasis. Evidences indicate that Alk4 is a key regulator of many embryonic processes, but little is known about its signaling in adult tissues and in pathological conditions where Alk4 mutations had been reported. Conventional deletion of Alk4 gene (Acvr1b) results in early embryonic lethality prior gastrulation, which has precluded study of Alk4 functions in postnatal and adult mice. To circumvent this problem, we have generated a conditional Acvr1b floxed-allele by flanking the fifth and sixth exons of the Acvr1b gene with loxP sites. Cre-mediated deletion of the floxed allele generates a deleted allele, which behaves as an Acvr1b null allele leading to embryonic lethality in homozygous mutant animals. A tamoxifen-inducible approach to target disruption of Acvr1b specifically in adult tissues was used and proved to be efficient for studying Alk4 functions in various organs. We report, therefore, a novel conditional model allowing investigation of biological role played by Alk4 in a variety of tissue-specific contexts.

Generation of a conditional mouse model to target Acvr1b disruption in adult tissues.

Pdx1-Cre; LSL-KRAS(G12D) mice develop premalignant pancreatic ductal lesions that can possibly progress spontaneously to pancreatic ductal adenocarcinoma (PDAC). Although Pdx1-Cre is expressed in the embryonic endoderm, which gives rise to all pancreatic lineages, the possible consequences of KRAS(G12D) expression in the endocrine compartment have never been finely explored.

Article	Year
Tif1Î³ suppresses murine pancreatic tumoral transformation by a Smad4-independent pathway. The American journal of pathology\| PubMed ID: 22469842	2012
The human NUPR1/P8 gene is transcriptionally activated by transforming growth factor Î² via the SMAD signalling pathway. The Biochemical journal\| PubMed ID: 22738338	2012
Generation of a conditional mouse model to target Acvr1b disruption in adult tissues. Genesis (New York, N.Y. : 2000)\| PubMed ID: 23109354	2013
The conditional expression of KRAS G12D in mouse pancreas induces disorganization of endocrine islets prior the onset of ductal pre-cancerous lesions. Pancreatology : official journal of the International Association of Pancreatology (IAP) ... [et al.]\| PubMed ID: 23719586	2013

Roxane M. Pommier

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