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Gene expression profiling of a mouse model of pancreatic islet dysmorphogenesis.

Wilding Crawford L, Tweedie Ables E, Oh YA, Boone B, Levy S, Gannon M - PLoS ONE (2008)

Bottom Line: Despite this success, many of the factors necessary for proper islet morphogenesis and function remain uncharacterized.Genome-wide microarray analysis was used to identify differences in the gene expression profiles of late gestation and early postnatal total pancreas tissue from wild type and Hnf6 transgenic animals.This study provides a unique dataset that can act as a starting point for other investigators to explore the role of the identified genes in pancreatogenesis, islet morphogenesis and mature beta cell function.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Physiology and Biophysics, Vanderbilt University Medical Center, Nashville, Tennessee, USA.

ABSTRACT

Background: In the past decade, several transcription factors critical for pancreas organogenesis have been identified. Despite this success, many of the factors necessary for proper islet morphogenesis and function remain uncharacterized. Previous studies have shown that transgenic over-expression of the transcription factor Hnf6 specifically in the pancreatic endocrine cell lineage resulted in disruptions in islet morphogenesis, including dysfunctional endocrine cell sorting, increased individual islet size, increased number of peripheral endocrine cell types, and failure of islets to migrate away from the ductal epithelium. The mechanisms whereby maintained Hnf6 causes defects in islet morphogenesis have yet to be elucidated.

Methodology/principal findings: We exploited the dysmorphic islets in Hnf6 transgenic animals as a tool to identify factors important for islet morphogenesis. Genome-wide microarray analysis was used to identify differences in the gene expression profiles of late gestation and early postnatal total pancreas tissue from wild type and Hnf6 transgenic animals. Here we report the identification of genes with an altered expression in Hnf6 transgenic animals and highlight factors with potential importance in islet morphogenesis. Importantly, gene products involved in cell adhesion, cell migration, ECM remodeling and proliferation were found to be altered in Hnf6 transgenic pancreata, revealing specific candidates that can now be analyzed directly for their role in these processes during islet development.

Conclusions/significance: This study provides a unique dataset that can act as a starting point for other investigators to explore the role of the identified genes in pancreatogenesis, islet morphogenesis and mature beta cell function.

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Hnf6 Tg animals exhibit abnormal islet morphogenesis.(A) Islet from a four week old WT animal showing insulin-producing β cells at the islet core (green) and glucagon-producing α cells at the periphery (red). (B) At the same age, individual Hnf6 Tg islets are larger, have a mixed islet phenotype, and are closely apposed to the ductal epithelium. (A*, B*) At e18.5, WT endocrine cells have begun to adopt the stereotypic architecture; however, Hnf6 Tg α cells are often clustered together in islet-like structures that have few, if any, β cells (arrows in B*). (C) Morphometric analysis demonstrates increased glucagon+ cell area (red) as early as e15.5 in Hnf6 Tg islets (dark red). This relative increase over WT islets (solid bars) persists into postnatal stages. Total insulin+ area decreases in transgenic animals (dark green) at postnatal stages, but does not appear to be altered during embryogenesis. e15.5, n = 4; e18.5, n = 4; P1, n = 2; 2 weeks, n = 3. Error bars were determined by 95% confidence interval. *p<0.05, **p<0.005, as determined by Student's t-test. d, duct; bv, blood vessel.
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pone-0001611-g001: Hnf6 Tg animals exhibit abnormal islet morphogenesis.(A) Islet from a four week old WT animal showing insulin-producing β cells at the islet core (green) and glucagon-producing α cells at the periphery (red). (B) At the same age, individual Hnf6 Tg islets are larger, have a mixed islet phenotype, and are closely apposed to the ductal epithelium. (A*, B*) At e18.5, WT endocrine cells have begun to adopt the stereotypic architecture; however, Hnf6 Tg α cells are often clustered together in islet-like structures that have few, if any, β cells (arrows in B*). (C) Morphometric analysis demonstrates increased glucagon+ cell area (red) as early as e15.5 in Hnf6 Tg islets (dark red). This relative increase over WT islets (solid bars) persists into postnatal stages. Total insulin+ area decreases in transgenic animals (dark green) at postnatal stages, but does not appear to be altered during embryogenesis. e15.5, n = 4; e18.5, n = 4; P1, n = 2; 2 weeks, n = 3. Error bars were determined by 95% confidence interval. *p<0.05, **p<0.005, as determined by Student's t-test. d, duct; bv, blood vessel.

Mentions: We previously described a transgenic mouse model which displayed alterations in islet morphogenesis [15], [22]. In this model, a 1.0 kb endocrine-specific enhancer (PB; [23]) from the 5′ regulatory region of the pancreas and duodenal homeobox 1 (Pdx1) gene was used to drive the expression of Hnf6 specifically to the pancreatic endocrine lineage. Pdx1 is normally expressed in the antral stomach, pancreas, and proximal duodenum and is essential for pancreas development and mature β cell function [24], [25], [26], [27]. As Hnf6 is normally down-regulated in the endocrine lineage prior to birth, the Pdx1PBHnf6 transgene (Hnf6 Tg) allowed for the maintenance of Hnf6 expression in endocrine tissue during late embryogenesis and continuing into adulthood [15]. We found that maintaining Hnf6 expression in islets resulted in a host of morphological and functional deficiencies, including a scattering of cell types within the islets, abnormal islet size, decreased insulin secretion, and a block to β cell maturation (Figure 1; [15], [22]). Additional morphometric analysis during embryonic and perinatal development showed that islet composition is also altered in Hnf6 Tg mice. As seen in Figure 1C, glucagon+ cell area was increased in Hnf6 Tg embryos as early as e15.5, and this increase was maintained postnatally (Figure 1C and data not shown). The increase in glucagon+ cell area was manifested by larger glucagon+ cell clusters that appeared to be segregated from insulin+ cells (Figure 1B*). Insulin+ area was not affected by the maintenance of Hnf6 until postnatal stages, at which point a significant decrease in insulin+ area as compared to WT controls is observed (Figure 1C). As we have previously shown, however, this mild decrease in the number of β cells does not lead to the diabetes observed in this transgenic line [22]. Rather, defects in insulin granule biosynthesis and vesicle trafficking cause the dramatic decrease in glucose stimulated insulin secretion and ultimately, diabetes [22]. The time period of embryonic glucagon+ cell population expansion coincides with previous reports of a dramatic increase in hormone-expressing cells in normal pancreas tissue known as the secondary transition, in which pro-endocrine cells that will mature to form the functional units of the adult pancreas are thought to undergo substantial differentiation and proliferation (for review, see [2]).


Gene expression profiling of a mouse model of pancreatic islet dysmorphogenesis.

Wilding Crawford L, Tweedie Ables E, Oh YA, Boone B, Levy S, Gannon M - PLoS ONE (2008)

Hnf6 Tg animals exhibit abnormal islet morphogenesis.(A) Islet from a four week old WT animal showing insulin-producing β cells at the islet core (green) and glucagon-producing α cells at the periphery (red). (B) At the same age, individual Hnf6 Tg islets are larger, have a mixed islet phenotype, and are closely apposed to the ductal epithelium. (A*, B*) At e18.5, WT endocrine cells have begun to adopt the stereotypic architecture; however, Hnf6 Tg α cells are often clustered together in islet-like structures that have few, if any, β cells (arrows in B*). (C) Morphometric analysis demonstrates increased glucagon+ cell area (red) as early as e15.5 in Hnf6 Tg islets (dark red). This relative increase over WT islets (solid bars) persists into postnatal stages. Total insulin+ area decreases in transgenic animals (dark green) at postnatal stages, but does not appear to be altered during embryogenesis. e15.5, n = 4; e18.5, n = 4; P1, n = 2; 2 weeks, n = 3. Error bars were determined by 95% confidence interval. *p<0.05, **p<0.005, as determined by Student's t-test. d, duct; bv, blood vessel.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2249940&req=5

pone-0001611-g001: Hnf6 Tg animals exhibit abnormal islet morphogenesis.(A) Islet from a four week old WT animal showing insulin-producing β cells at the islet core (green) and glucagon-producing α cells at the periphery (red). (B) At the same age, individual Hnf6 Tg islets are larger, have a mixed islet phenotype, and are closely apposed to the ductal epithelium. (A*, B*) At e18.5, WT endocrine cells have begun to adopt the stereotypic architecture; however, Hnf6 Tg α cells are often clustered together in islet-like structures that have few, if any, β cells (arrows in B*). (C) Morphometric analysis demonstrates increased glucagon+ cell area (red) as early as e15.5 in Hnf6 Tg islets (dark red). This relative increase over WT islets (solid bars) persists into postnatal stages. Total insulin+ area decreases in transgenic animals (dark green) at postnatal stages, but does not appear to be altered during embryogenesis. e15.5, n = 4; e18.5, n = 4; P1, n = 2; 2 weeks, n = 3. Error bars were determined by 95% confidence interval. *p<0.05, **p<0.005, as determined by Student's t-test. d, duct; bv, blood vessel.
Mentions: We previously described a transgenic mouse model which displayed alterations in islet morphogenesis [15], [22]. In this model, a 1.0 kb endocrine-specific enhancer (PB; [23]) from the 5′ regulatory region of the pancreas and duodenal homeobox 1 (Pdx1) gene was used to drive the expression of Hnf6 specifically to the pancreatic endocrine lineage. Pdx1 is normally expressed in the antral stomach, pancreas, and proximal duodenum and is essential for pancreas development and mature β cell function [24], [25], [26], [27]. As Hnf6 is normally down-regulated in the endocrine lineage prior to birth, the Pdx1PBHnf6 transgene (Hnf6 Tg) allowed for the maintenance of Hnf6 expression in endocrine tissue during late embryogenesis and continuing into adulthood [15]. We found that maintaining Hnf6 expression in islets resulted in a host of morphological and functional deficiencies, including a scattering of cell types within the islets, abnormal islet size, decreased insulin secretion, and a block to β cell maturation (Figure 1; [15], [22]). Additional morphometric analysis during embryonic and perinatal development showed that islet composition is also altered in Hnf6 Tg mice. As seen in Figure 1C, glucagon+ cell area was increased in Hnf6 Tg embryos as early as e15.5, and this increase was maintained postnatally (Figure 1C and data not shown). The increase in glucagon+ cell area was manifested by larger glucagon+ cell clusters that appeared to be segregated from insulin+ cells (Figure 1B*). Insulin+ area was not affected by the maintenance of Hnf6 until postnatal stages, at which point a significant decrease in insulin+ area as compared to WT controls is observed (Figure 1C). As we have previously shown, however, this mild decrease in the number of β cells does not lead to the diabetes observed in this transgenic line [22]. Rather, defects in insulin granule biosynthesis and vesicle trafficking cause the dramatic decrease in glucose stimulated insulin secretion and ultimately, diabetes [22]. The time period of embryonic glucagon+ cell population expansion coincides with previous reports of a dramatic increase in hormone-expressing cells in normal pancreas tissue known as the secondary transition, in which pro-endocrine cells that will mature to form the functional units of the adult pancreas are thought to undergo substantial differentiation and proliferation (for review, see [2]).

Bottom Line: Despite this success, many of the factors necessary for proper islet morphogenesis and function remain uncharacterized.Genome-wide microarray analysis was used to identify differences in the gene expression profiles of late gestation and early postnatal total pancreas tissue from wild type and Hnf6 transgenic animals.This study provides a unique dataset that can act as a starting point for other investigators to explore the role of the identified genes in pancreatogenesis, islet morphogenesis and mature beta cell function.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Physiology and Biophysics, Vanderbilt University Medical Center, Nashville, Tennessee, USA.

ABSTRACT

Background: In the past decade, several transcription factors critical for pancreas organogenesis have been identified. Despite this success, many of the factors necessary for proper islet morphogenesis and function remain uncharacterized. Previous studies have shown that transgenic over-expression of the transcription factor Hnf6 specifically in the pancreatic endocrine cell lineage resulted in disruptions in islet morphogenesis, including dysfunctional endocrine cell sorting, increased individual islet size, increased number of peripheral endocrine cell types, and failure of islets to migrate away from the ductal epithelium. The mechanisms whereby maintained Hnf6 causes defects in islet morphogenesis have yet to be elucidated.

Methodology/principal findings: We exploited the dysmorphic islets in Hnf6 transgenic animals as a tool to identify factors important for islet morphogenesis. Genome-wide microarray analysis was used to identify differences in the gene expression profiles of late gestation and early postnatal total pancreas tissue from wild type and Hnf6 transgenic animals. Here we report the identification of genes with an altered expression in Hnf6 transgenic animals and highlight factors with potential importance in islet morphogenesis. Importantly, gene products involved in cell adhesion, cell migration, ECM remodeling and proliferation were found to be altered in Hnf6 transgenic pancreata, revealing specific candidates that can now be analyzed directly for their role in these processes during islet development.

Conclusions/significance: This study provides a unique dataset that can act as a starting point for other investigators to explore the role of the identified genes in pancreatogenesis, islet morphogenesis and mature beta cell function.

Show MeSH
Related in: MedlinePlus