Evidence for the presence of stem cell-like progenitor cells in human adult pancreas

The origin of cells replacing ageing beta-cells in adult life is unknown. This study assessed the expression of classic stem cell markers: Oct4, Sox2 and CD34 in islet-enriched fractions versus exocrine cell-enriched fractions from 25 adult human pancreases following human islet isolation. Expression of Oct4, Sox2 and CD34 mRNAs was found in all cell samples, with no significant differences between endocrine and exocrine cell fractions. Immunohistochemical staining for Oct4, Sox2, CD133, CD34, CK19, insulin and nestin on human pancreas sections showed that the majority of Oct4(+ve) cells were found in the walls of small ducts. Similar localisations were observed for Sox2(+ve) cells. The majority of Sox2(+ve) cells were found to co-express Oct4 proteins, but not vice versa. Cells positive for Oct4 and Sox2 appeared to be a unique cell population in the adult human pancreases without co-expression for CK19, CD34, CD133, insulin and nestin proteins. The numbers of Oct4(+ve) and Sox2(+ve) cells varied among donors and were approximately 1-200 and 1-30 per 100 000 pancreatic cells respectively.     

Ductal and acinar differentiation in pancreatic endocrine tumors

Rare pancreatic endocrine tumors consisting of both exocrine and endocrine components have been reported sporadically. We investigated the ductal and acinar differentiation in pancreatic endocrine tumors. In immunohistochemical studies of 28 pancreatic endocrine tumors, staining with anti-carcinoembryonic antigen (CEA) or CA19-9 antibody indicated ductal differentiation, while staining with anti-amylase or anti-trypsin antibody indicated acinar differentiation. K-ras gene mutations and p53 gene alterations also were studied. Ten tumors were immunoreactive for CEA or CA19-9. Five tumors diffusely immunoreactive for CEA or CA19-9, in addition to endocrine markers, were diagnosed as duct–endocrine cell tumors of the pancreas. Two tumors diffusely immunoreactive for CEA or CA19-9 and also for pancreaticogut hormones as well as endocrine markers were diagnosed as duct–acinar–endocrine cell tumors. These tumors showed uniform histologic features and synchronous ductal, acinar, and endocrine differentiation, distinct from the coexisting different cellular populations seen in collision tumors. All tumors were malignant. These duct–endocrine cell tumors or duct–acinar–endocrine cell tumors of the pancreas may be derived from a stem cell that retains the capability of expressing either an exocrine or endocrine phenotype. Only one K-ras gene mutation and no p53 gene alterations were detected in these tumors, which suggests that they constitute an entity with a different origin than ductal carcinomas.     

Hepatocyte turnover and regeneration: virtually a virtuoso performance

The liver and exocrine pancreas share a common structure, with functioning units (hepatic plates and pancreatic acini) connected to the ductal tree. Here we show that Sox9 is expressed throughout the biliary and pancreatic ductal epithelia, which are connected to the intestinal stem-cell zone. Cre-based lineage tracing showed that adult intestinal cells, hepatocytes and pancreatic acinar cells are supplied physiologically from Sox9-expressing progenitors. Combination of lineage analysis and hepatic injury experiments showed involvement of Sox9-positive precursors in liver regeneration. Embryonic pancreatic Sox9-expressing cells differentiate into all types of mature cells, but their capacity for endocrine differentiation diminishes shortly after birth, when endocrine cells detach from the epithelial lining of the ducts and form the islets of Langerhans. We observed a developmental switch in the hepatic progenitor cell type from Sox9-negative to Sox9-positive progenitors as the biliary tree develops. These results suggest interdependence between the structure and homeostasis of endodermal organs, with Sox9 expression being linked to progenitor status.     

Non-islet origin of pancreatic islet cell tumors

The histogenesis of pancreatic islet cell tumors was investigated by morphological identification of putative precursor lesions in pancreatic tissue from patients with multiple endocrine neoplasia type 1 (MEN1), tissue microdissection, and genetic analysis. MEN1 mutation and absence of the MEN1 wild-type allele in different precursor lesions strongly suggest that pancreatic islet cell tumors are derived from the ductal/acinar system but not from pancreatic islet tissue. Pluripotent cells within the exocrine pancreas appear capable of formation into small atypical accumulations of MEN1-deficient cells with both exocrine and endocrine phenotype. The findings suggest presence of multiple developmental aberrations in MEN1 pancreas that potentially serve as precursor material for neuroendocrine tumors.     

Characterization of pancreatic endocrine cells of the European common frog Rana temporaria

To characterize the endocrine cell types of the pancreas of Rana temporaria, conventional staining, silver impregnation, and immunocytochemical methods for light and electron microscopy have been applied to paraffin, thin and semithin sections, many of them serial pairs. Quantitative data on the frequency and distribution (insular, extrainsular among the exocrine cells, or within the pancreatic ducts) of each endocrine cell type are also reported. Four distinct endocrine cell types have been identified: insulin (B) cells, which are also immunoreactive for [Met]enkephalin; glucagon/PP (A/PP) cells, also immunoreactive for GLP1; somatostatin (D) cells; and a fourth endocrine-like cell type (X cells) of unknown content and function. X cells display characteristic ultrastructure and tinctorial traits but are nonimmunoreactive for all of the 37 antisera tested. The presence of [Met]enkephalin in amphibian pancreatic endocrine cells is now reported for the first time. Almost half (44.9 +/- 7.9) of the total endocrine cell population lies outside the islets, mainly spread among the exocrine cells. Approximately 37.2 +/- 4.6% of the total endocrine cell population was immunoreactive for insulin, 48.8 +/- 6.9% was immunoreactive for glucagon/PP, and 14.0 +/- 4.9% was immunoreactive for somatostatin; 79.2 +/- 6.4% of glucagon/PP cells are found within the exocrine parenchyma, representing the majority (86.4 +/- 4.3%) of extrainsular endocrine component. On the contrary, most B cells (94.2 +/- 2.1%) are located within the islets; 30.8 +/- 12.9% of D cells are found outside the islets.     

Hedgehog signaling is required for effective regeneration of exocrine pancreas

Although both endocrine and the exocrine pancreas display a significant capacity for tissue regeneration and renewal, the existence of progenitor cells in the adult pancreas remains uncertain. Using a model of cerulein-mediated injury and repair, we demonstrate that mature exocrine cells, defined by expression of an Elastase1 promoter, actively contribute to regenerating pancreatic epithelium through formation of metaplastic ductal intermediates. Acinar cell regeneration is associated with activation of Hedgehog (Hh) signaling, as assessed by up-regulated expression of multiple pathway components, as well as activation of a Ptch-lacZ reporter allele. Using both pharmacologic and genetic techniques, we also show that the ability of mature exocrine cells to accomplish pancreatic regeneration is impaired by blockade of Hh signaling. Specifically, attenuated regeneration in the absence of an intact Hh pathway is characterized by persistence of metaplastic epithelium expressing markers of pancreatic progenitor cells, suggesting an inhibition of redifferentiation into mature exocrine cells. Given the known role of Hh signaling in exocrine pancreatic cancer, these findings may provide a mechanistic link between injury-induced activation of pancreatic progenitors and subsequent pancreatic neoplasia.     

Nestin-positive cells in adult pancreas express amylase and endocrine precursor Cells

The neural precursor cell-specific marker nestin is expressed in fetal and adult pancreas, but its role is not fully understood. Using nestin-enhanced green fluorescent protein (EGFP) transgenic mice and fluorescence activated cell sorter, we characterized nestin-positive cells in adult mice pancreas. EGFP mRNA- and protein-positive cells expressed amylase, a pancreatic exocrine marker. Interestingly, EGFP mRNA-negative and protein-positive cells expressed insulin, glucagon, somatostatin and pancreatic polypeptide, pancreatic endocrine markers. These findings demonstrate that nestin-positive cells comprise a portion of pancreatic exocrine cells and suggest that they can be differentiated into pancreatic endocrine cells.     

Betacellulin inhibits amylase and glucagon production and promotes beta cell differentiation in mouse embryonic pancreas

Aims/hypothesis Betacellulin, a member of the epidermal growth factor family, is expressed in the pancreas and is thought to regulate differentiation of beta cells during development. The aim of the present study was to investigate the effects of exogenous betacellulin on the development of the mouse embryonic pancreas. Materials and methods We used an in vitro culture model system based on the isolation and culture of the dorsal embryonic pancreas from day 11.5 embryos. Cultures were treated for up to 10 days with 10 ng/ml betacellulin and then analysed for changes in the expression of pancreatic exocrine, endocrine and ductal markers. Results Pancreases developed in culture and expressed the full complement of exocrine (both acinar and ductal) and endocrine cell types. Betacellulin enhanced branching morphogenesis and the proliferation of mesenchyme, increased Pdx1 and insulin production and inhibited the production of the exocrine cell marker amylase and the endocrine hormone glucagon. Conclusions/interpretation These results suggest betacellulin has distinct and separate effects on exocrine, endocrine and ductal differentiation. In the future, betacellulin could perhaps be utilised to increase the production of beta cells from embryonic pancreatic tissue for therapeutic transplantation. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorised users.     

Peanut-like 1 (septin 5) gene expression in normal and neoplastic human endocrine pancreas

Peanut-like 1 (PNUTL1) is a septin gene which is expressed at high levels in human brain. There it plays a role in the process of membrane fusion during exocytosis by interacting with syntaxin and synaptophysin. As the secretory apparatus of pancreatic islet cells closely resembles that of neurons, we decided to study the expression of PNUTL1 in the human endocrine pancreas, both in normal islets and in pancreatic endocrine tumors (PETs). Normal pancreatic tissue, purified islets, 11 PETs and two cell lines were used to evaluate the presence of PNUTL1 by RT-PCR and Western blot. The expression of the PNUTL1 protein was also evaluated by immunohistochemistry on normal pancreas, additional 26 PETs, eight pancreatic adenocarcinomas, one mixed endocrine-exocrine pancreatic neoplasm, a specimen of solid papillary pseudomucinous tumor, an adult islet cell hyperplasia and a case of neonatal nesidioblastosis. In addition, a tissue array (LandMark High Density Cancer Tissue MicroArray) comprising 280 various tumor and matched normal specimens was utilized. In PETs, the expression of pancreatic hormones, chromogranin-A, synaptophysin and Ki-67 were also evaluated. In the normal pancreas PNUTL1 expression is almost exclusively confined to the islet cells, weak expression was occasionally seen in some acinar cells, while immunoreactivity was completely absent in the ductal epithelia. PNUTL1 expression is maintained at similar high levels in hyperplastic and neoplastic islet cells, but this did not correlate with any of the clinicopathological data nor with proliferation status in PETs. Weak immunoreactivity was also noted in a proportion of exocrine neoplasms. Our findings describe for the first time the high expression levels of PNUTL1 in human pancreatic endocrine cells that suggests a similar role of this protein in islet cells to that demonstrated in neuronal tissues, and warrants further functional studies of this protein.     

胰腺内分泌肿瘤细胞成分的免疫组织化学观察及其对肿瘤起源的探讨

目的探讨胰腺内分泌肿瘤的生物学起源。方法应用链菌素亲生物素－过氧化物酶标免疫组织化学技术（Ｓ－Ｐ法）对５２例胰腺内分泌肿瘤（胰岛素瘤３２例，非功能性胰腺内分泌肿瘤２０例）瘤细胞内胰岛素、胰高血糖素、舒血管肠肽、生长抑素、胃泌素、Ｐ一物质、促肾上腺皮质激素、绒毛膜促性腺激素及５－羟色胺等９种激素或激素类物质的分布进行了观察。结果６５。５％（１３／２０）的非功能性胰腺内分泌肿瘤的细胞具有合成与低存激素的功能．３２．７％（１７／５２）胰腺内分泌肿瘤可含有分泌异位激素的细胞，６３．５％（３３／５２）的胰腺内分泌肿瘤是由多种内分泌细胞所构成。结论支持胰腺内分泌肿瘤是由胰腺内分泌于细胞起源的理论。     

胎儿胰腺组织中巢蛋白阳性细胞的分离和体外增殖以及诱导分化

目的 研究从胎儿胰腺组织中分离巢蛋白(Nestin)阳性细胞以及该细胞的体外扩增与向胰岛内分泌细胞分化的能力。方法 采用胶原酶消化法，从胎儿胰腺组织中分离获得胰岛样细胞簇(islet-like cell clusters，ICCs)，ICCs经手工挑拣后接种，待形成单层上皮样细胞后，进行传代培养和诱导分化。利用逆转录-聚合酶链反应(RT-PCR)、免疫荧光染色及放射免疫分析(RIA)等方法，检测该细胞中分子标志物的表达，并对其向胰岛内分泌细胞分化的能力进行鉴定。结果 (1)上述单层上皮样细胞具有很强的增殖能力，可至少连续传16代；(2)RT-PCR、免疫荧光染色分析显示，该细胞可表达干细胞的标志分子巢蛋白和ABCG2；(3)RT-PCR分析显示，在多种细胞因子和无血清的条件下，巢蛋白阳性细胞经诱导后可出现胰岛素、胰升糖素和胰十二指肠同源盒基因-1(PDX-1)mRNA的表达，而巢蛋白和Neurogenin3(Ngn3)mRNA表达消失。RIA分析也可检测到诱导后的细胞内有胰岛素产生。结论 从胎儿胰腺中分离得到的巢蛋白阳性细胞具有胰腺前体细胞的特性，在体外具有很强的增殖能力，并可诱导分化为胰岛内分泌细胞。该细胞有望为胰岛移植提供一种新的细胞来源。     

Insulin-like growth factor II in human fetal pancreas and its co-localization with the major islet hormones: comparison with adult pancreas

Insulin-like growth factor II (IGF-II) appears to play an important role during fetal life in cell growth and differentiation in several organs, including the pancreas. In the present study we investigated the cellular localization of IGF-II in human fetal pancreas at 16, 18 and 22 embryonic weeks and compared it with adult pancreas. Single and double immunofluorescence methods were used to study co-localization of IGF-II with the four major islet hormones - insulin, glucagon, somatostatin, pancreatic polypeptide - and with islet amyloid polypeptide (IAPP). Distinct IGF-II immunoreactive (IR) cells were found in the endocrine, but not in the exocrine, pancreas. The intensity of IGF-II immunoreactivity was more pronounced in the fetal than in the adult pancreas. In fetal pancreas IGF-II immunoreactivity was observed in virtually all insulin-IR cells and in subsets of the glucagon, somatostatin and IAPP cells. In the adult pancreas, IGF-II immunoreactivity was found in insulin/IAPP cells only. Our results suggest a broader effect of IGF-II in fetal endocrine pancreatic cells than in the adult.