(Journal Article): Enriched Human Pancreatic Ductal Cultures Obtained from Selective Death of Acinar Cells Express Pancreatic and Duodenal Homeobox Gene-1 Age-Dependently
Street CN, Lakey JR, Rajotte RV, Shapiro AM, Kieffer TJ, Lyon JG, Kin T, Korbutt GS (Surgical Medical Research Institute, University of Alberta, Edmonton, AB, Canada T6G 2R3,
korbutt@ualberta.ca
)
IN:
Rev Diabetic Stud
2004; 1(2):66-79
Impact Factor(s) of Rev Diabetic Stud: 0.125 (2006)
ABSTRACT: Adult pancreatic ductal cells are believed to be islet precursors. Our aim was to obtain an enriched human ductal cell population in defined culture conditions, and to characterize these cultures for the presence of pancreatic developmental transcription factors. Non-endocrine adult human pancreatic digest was cultured for 4 days in serum-containing and serum-free media. During this time, analysis was done for phenotypic changes, cell death, and expression of islet and islet precursor markers. Culture in serum-supplemented and serum-free media gave similar recoveries of an enriched ductal population after 4 days. Extensive cell death due to apoptosis and necrosis was also observed over this time period. A donor-age dependent expression of pancreatic and duodenal homeobox gene-1 (PDX-1) in ductal cells was seen at 4 days whereby donors <25 yr expressed significantly more than donors >25 yr. Analysis of gene expression by RT-PCR showed the presence of islet developmental transcription factors neuroD, Nkx6.1, and PDX-1, as well as mature islet hormones. While acinar-ductal transdifferentiation of some cells cannot be ruled out, we provide evidence that the predominant mechanism for the derivation of enriched human ductal cultures in our culture conditions is selective acinar cell death. Furthermore, we have shown that ductal cultures from younger donors exhibit greater plasticity through expression of PDX-1, and may be of greater value in attempts to induce islet neogenesis. The presence, however, of insulin and glucagon mRNA indicates that contaminating endocrine cells remain in these cultures and underscores the need to use caution when assessing differentiation potential.
TYPE OF PUBLICATION: Original article
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(Journal Article): Modulation of rat pancreatic acinoductal transdifferentiation and expression of PDX-1 in vitro.
Rooman I, Heremans Y, Heimberg H, Bouwens L (Department of Experimental Pathology, Free University of Brussels (VUB), Belgium.)
IN:
Diabetologia
2000; 43(7):907-910
Impact Factor(s) of Diabetologia: 5.583 (2004), 5.689 (2003), 5.136 (2002), 6.299 (2001)
ABSTRACT: AIMS/HYPOTHESIS: In adult pancreatic regeneration models exocrine acini are found to transdifferentiate to duct-like complexes. This has also been associated with the formation of new endocrine islet cells. We aimed to establish an in vitro model in which this transdifferentiation process is characterised and can be modulated. METHODS: Purified rat pancreatic acini were cultured in suspension. Differentiation was analysed by immunocytochemistry, electron microscopy, western blotting and RT-PCR. RESULTS: During culture acinar cells directly transdifferentiated without dividing, the cells lost their acinar phenotype and started to express cytokeratins 20 and 7 and fetal liver kinase-1 (Flk-1) receptors for vascular endothelial growth factor. Expression of the acinar pancreatic exocrine transcription factor (PTF-1) remained and the pancreatic duodenal homeobox-containing transcription factor (PDX-1) was induced. When transdifferentiation was completed, the cells started to express protein gene product 9.5, a panneuroendocrine marker. By combining these features, the transdifferentiated cells show similar characteristics to precursor cells during active beta-cell neogenesis. We were able to modulate the differentiation state by addition of nicotinamide or sodium butyrate, agents which are known to stimulate endocrine differentiation in other models. CONCLUSION/INTERPRETATION: Here, we present an in vitro system in which the cellular differentiation of putative pancreatic endocrine precursor cells and their PDX-1 expression can be modulated, thereby providing a possible model for the study of beta-cell transdifferentiation.
TYPE OF PUBLICATION: Original article
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(Journal Article): Pancreatic duodenal homeobox-1, PDX-1, a major regulator of beta-cell identity and function.
Mckinnon CM, Docherty K (Department of Molecular and Cell Biology, University of Aberdeen, Institute of Medical Sciences, Foresterhill, Aberdeen, UK.)
IN:
Diabetologia
2001; 44(10):1203-1214
Impact Factor(s) of Diabetologia: 5.583 (2004), 5.689 (2003), 5.136 (2002), 6.299 (2001)
ABSTRACT: Pancreatic duodenal homeobox -1 is a transcription factor that is expressed in beta and delta cells of the islets of Langerhans and in dispersed endocrine cells of the duodenum. It is involved in regulating the expression of a number of key beta-cell genes as well as somatostatin. It also plays a pivotal part in the development of the pancreas and islet cell ontogeny. Thus homozygous disruption of the gene in mice and humans results in pancreatic agenesis. Heterozygous mutations in the gene result in impaired glucose tolerance and symptoms of diabetes as seen in MODY4 and late-onset Type II (non-insulin-dependent) diabetes mellitus. In adults pancreatic duodenal homeobox-1 expression is increased in duct cells of the pancreas that have been induced to proliferate and differentiate to form new islets. Defects in pancreatic duodenal homeobox-1 could therefore contribute to Type II diabetes by affecting compensatory mechanisms that increase the rate of beta-cell neogenesis to meet the increased insulin secretory demand. It could also be a pharmacological target for beta-cell defects in Type II diabetes, while its role as a regulator of islet stem cell activity is being exploited to produce a replenishable source of islet tissue for transplantation in Type I (insulin-dependent) diabetes mellitus.
TYPE OF PUBLICATION: Review
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(Journal Article): Pancreatic duodenal homeobox-1 and islet neogenesis associated protein: a possible combined marker of activateable pancreatic cell precursors.
Gagliardino JJ, Del Zotto H, Massa L, Flores LE, Borelli MI (CENEXA - Centre of Experimental and Applied Endocrinology (UNLP-CONICET, PAHO/WHO Collaborating Centre), National University of La Plata School of Medicine, La Plata, Argentina.)
IN:
J Endocrinol
2003; 177(2):249-259
Impact Factor(s) of J Endocrinol: 3.319 (2004), 3.023 (2003), 2.834 (2001)
ABSTRACT: The aim of this work was to study the possible relationship between pancreatic duodenal homeobox-1 (Pdx-1) and islet neogenesis-associated protein (INGAP) during induced islet neogenesis. Pregnant hamsters were fed with (S) and without (C) sucrose, and glycemia, insulin secretion in vitro, and pancreas immunomorphometric parameters were measured in their 7-day-old offspring. S offspring had significantly lower glycemic levels than C animals. Insulin release in response to increasing glucose concentrations in the incubation medium (2-16 mM glucose) did not increase in pancreata from either C or S offspring. However, pancreata from S offspring released more insulin than those from C animals. In S offspring, beta-cell mass, beta-cell replication rate and islet neogenesis increased significantly, with a simultaneous decrease in beta-cell apoptotic rate. INGAP- and Pdx-1-positive cell mass also increased in the islets and among acinar and duct cells. We found two subpopulations of Pdx-1 cells: INGAP-positive and INGAP-negative. Pdx-1/INGAP-positive cells did not stain with insulin, glucagon, somatostatin, pancreatic polypeptide, or neurogenin 3 antibodies. The increment of Pdx-1/INGAP-positive cells represented the major contribution to the Pdx-1 cell mass increase. Such increments varied among pancreas subsectors: ductal>insular>extrainsular. Our results suggested that INGAP participates in the regulation of islet neogenesis, and Pdx-1/INGAP-positive cells represent a new stem cell subpopulation at an early stage of development, highly activateable in neogenesis.
TYPE OF PUBLICATION: Original article
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(Journal Article): Expansion of PDX-1 expressing pancreatic epithelium and islet neogenesis in transgenic mice overexpressing transforming growth factor-alpha.
Young-Song S, Gannon M, Washington MK, Scoggins CR, Meszoely IM, Goldenring JR, Marino CR, Sandgren EP, Coffey RJ, Wright CV, Leach SD (Department of Surgery, Vanderbilt-Ingram Cancer Center, Vanderbilt University Medical Center and Nashville VAMC, Nashville, TN 37232-2736, USA.)
IN:
Gastroenterology
1999; 117(6):1416-1426
Impact Factor(s) of Gastroenterology: 13.092 (2004), 12.718 (2003), 13.44 (2002), 13.02 (2001)
ABSTRACT: BACKGROUND & AIMS: The progenitor cells responsible for transforming growth factor (TGF)-alpha-induced pancreatic ductal metaplasia and neoplasia remain uncharacterized. During pancreatic development, differentiated cell types arise from ductal progenitor cells expressing the Pdx1 homeodomain transcription factor. The aims of this study were, first, to evaluate the role of Pdx1-expressing stem cells in MT-TGFalpha transgenic mice, and second, to further characterize cell proliferation and differentiation in this model. METHODS: To assess Pdx1 gene expression in normal and metaplastic epithelium, we performed in vivo reporter gene analysis using heterozygous Pdx1(lacZ/+) and bigenic Pdx1(lacZ/+)/MT-TGFalpha mice. RESULTS: Pdx1(lacZ/+)/MT-TGFalpha bigenics showed up-regulated Pdx1 expression in premalignant metaplastic ductal epithelium. In addition to Pdx1 gene activation, TGF-alpha-induced metaplastic epithelium demonstrated a pluripotent differentiation capacity, as evidenced by focal expression of Pax6 and initiation of islet cell neogenesis. The majority of Pdx1-positive epithelial cells showed no expression of insulin, similar to the pattern observed during embryonic development. CONCLUSIONS: Overexpression of TGF-alpha induces expansion of a Pdx1-expressing epithelium characterized by focal expression of Pax6 and initiation of islet neogenesis. These findings suggest that premalignant events induced by TGF-alpha in mouse pancreas may recapitulate a developmental program active during embryogenesis.
TYPE OF PUBLICATION: Original article
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(Journal Article): Improved glucose tolerance and acinar dysmorphogenesis by targeted expression of transcription factor PDX-1 to the exocrine pancreas.
Heller RS, Stoffers DA, Bock T, Svenstrup K, Jensen J, Horn T, Miller CP, Habener JF, Madsen OD, Serup P (Department of Developmental Biology, Hagedorn Research Institute, DK 2820, Gentofte, Denmark.,
shll@hagedorn.dk
)
IN:
Diabetes
2001; 50(7):1553-1561
Impact Factor(s) of Diabetes: 8.848 (2004), 8.298 (2003), 8.256 (2002), 7.7 (2001)
ABSTRACT: The homeodomain protein PDX-1 is critical for pancreas development and is a key regulator of insulin gene expression. PDX-1 nullizygosity and haploinsufficiency in mice and humans results in pancreatic agenesis and diabetes, respectively. At embryonic day (e) 10.5, PDX-1 is expressed in all pluripotential gut-derived epithelial cells destined to differentiate into the exocrine and endocrine pancreas. At e15, PDX-1 expression is downregulated in exocrine cells, but remains high in endocrine cells. The aim of this study was to determine whether targeted overexpression of PDX-1 to the exocrine compartment of the developing pancreas at e15 would allow for respecification of the exocrine cells. Transgenic (TG) mice were generated in which PDX-1 was expressed in the exocrine pancreas using the exocrine-specific elastase-1 promoter. These mice exhibited a marked dysmorphogenesis of the exocrine pancreas, manifested by increased rates of replication and apoptosis in acinar cells and a progressive fatty infiltration of the exocrine pancreas with age. Interestingly, the TG mice exhibited improved glucose tolerance, but absolute beta-cell mass was not increased. These findings indicate that downregulation of PDX-1 is required for the proper maintenance of the exocrine cell phenotype and that upregulation of PDX-1 in acinar cells affects beta-cell function. The mechanisms underlying these observations remain to be elucidated.
TYPE OF PUBLICATION: Original article
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(Journal Article): Pancreatic and duodenal homeobox gene 1 induces expression of insulin genes in liver and ameliorates streptozotocin induced hyperglycemia.
Ferber S, Halkin A, Cohen H, Ber I, Einav Y, Goldberg I, Barshack I, Seijffers R, Kopolovic J, Kaiser N, Karasik A (Endocrine Institute, Sheba Medical Center, Tel-Hashomer 52621, Israel.,
berezin@post.tau.ac.il
)
IN:
Nat Med
2000; 6(5):568-572
Impact Factor(s) of Nat Med: 28.878 (2005), 31.223 (2004), 30.55 (2003), 28.74 (2002), 27.906 (2001)
ABSTRACT: Insulin gene expression is restricted to islet beta cells of the mammalian pancreas through specific control mechanisms mediated in part by specific transcription factors. The protein encoded by the pancreatic and duodenal homeobox gene 1 (PDX-1) is central in regulating pancreatic development and islet cell function. PDX-1 regulates insulin gene expression and is involved in islet cell-specific expression of various genes. Involvement of PDX-1 in islet-cell differentiation and function has been demonstrated mainly by 'loss-of-function' studies. We used a 'gain-of-function' approach to test whether PDX-1 could endow a non-islet tissue with pancreatic beta-cell characteristics in vivo. Recombinant-adenovirus-mediated gene transfer of PDX-1 to the livers of BALB/C and C57BL/6 mice activated expression of the endogenous, otherwise silent, genes for mouse insulin 1 and 2 and prohormone convertase 1/3 (PC 1/3). Expression of PDX-1 resulted in a substantial increase in hepatic immunoreactive insulin content and an increase of 300% in plasma immunoreactive insulin levels, compared with that in mice treated with control adenovirus. Hepatic immunoreactive insulin induced by PDX-1 was processed to mature mouse insulin 1 and 2 and was biologically active; it ameliorated hyperglycemia in diabetic mice treated with streptozotocin. These data indicate the capacity of PDX-1 to reprogram extrapancreatic tissue towards a beta-cell phenotype, may provide a valuable approach for generating 'self' surrogate beta cells, suitable for replacing impaired islet-cell function in diabetics.
TYPE OF PUBLICATION: Original article
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(Journal Article): Adult human pancreatic duct and islet cells exhibit similarities in expression and differences in phosphorylation and complex formation of the homeodomain protein IPF-1.
Heimberg H, Bouwens L, Heremans Y, Van den Casteele M, Lefebvre V, Pipeleers D (Diabetes Research Center, Vrije Universiteit Brussel, Belgium.,
hheimber@vub.vub.ac.be
)
IN:
Diabetes
2000; 49(4):571-579
Impact Factor(s) of Diabetes: 8.848 (2004), 8.298 (2003), 8.256 (2002), 7.7 (2001)
ABSTRACT: The homeodomain transcription factor encoded by the pancreatic and duodenal homeobox gene-1 (Ipf-1) is essential for pancreatic ontogenesis. Whether Ipf-1 is also involved in the neogenesis of beta-cells in the adult pancreas is unknown. We examined whether Ipf-1 is expressed in adult human pancreatic ducts, which are thought to generate new beta-cells. In tissue sections, virtually all duct cells were immunopositive for Ipf-1, as were the islet beta-cells but not the acinar cells. After isolation and culture, both duct and islet cell preparations contained the Ipf-1 immunoreactive proteins p42 and p45 (42 and 45 kDa, respectively) in similar proportions, but the expression levels were twofold lower in duct cells. After 4 h of labeling, the endocrine cells exhibited a sevenfold higher phosphorylation of p42 than the duct cells, whereas p45 was phosphorylated only in endocrine cells. Homeobox binding transcription factor complexes with Ipf-1 in duct cells differed from those in endocrine cells in terms of gel mobility, sequence specificity, and affinity. The observed similarities in Ipf-1 expression by adult human pancreatic duct cells and endocrine cells may reflect their common ontogenic origin, whereas the differences in Ipf-1 phosphorylation and complex formation may correlate with their divergent differentiation.
TYPE OF PUBLICATION: Original article
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