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Results: 8

1.
Fig. 5.

Fig. 5. From: Rgs16 and Rgs8 in embryonic endocrine pancreas and mouse models of diabetes.

Rgs16::GFP+ cells are associated with ducts and blood vessels during postnatal islet formation. (A–E) Whole-mount staining for GFP in P15 BAC transgenic neonates demonstrates the close association of Rgs16-expressing VDACs (arrows) with blood vessels (PECAM) (A), ducts (DBA lectin) (B), endocrine cells (synaptophysin) (C), late endocrine cells (insulin) (D) and lymphatic vessels (Lyve-1) (E). Note the rare Rgs16::GFP+ cells observed within islets (yellow arrow in D). At higher resolution, a portion of VDACs (5%) can be seen to express Sox9 (F). (A–E) Confocal micrographs of whole-mount preparations, 30× (A–E). Confocal micrographs of sectioned tissue, 100× (F).

Alethia Villasenor, et al. Dis Model Mech. 2010 Sep-Oct;3(9-10):567-580.
2.
Fig. 6.

Fig. 6. From: Rgs16 and Rgs8 in embryonic endocrine pancreas and mouse models of diabetes.

Rgs16::GFP is expressed in regenerating islets of hyperglycemic PANIC-ATTAC mice. Rgs16::GFP and insulin are co-expressed in a few cells within islets of hyperglycemic PANIC-ATTAC;Rgs16::GFP transgenic mice during pancreatic β-cell proliferation (recovery) following ablation. (A,B) Expression is detected in severely hyperglycemic (A) or moderate glycemic (B) mice after 2 weeks of islet recovery (arrows mark Rgs16::GFP+;insulin+ double-positive cells). (C) Rgs16::GFP is not found in glucagon-expressing α-cells in the recovering islet (6 weeks of islet recovery). (D) No background GFP expression is detected in hyperglycemic PANIC-ATTAC mice. (E) Rgs16::GFP is never detected in normoglycemic mice PANIC-ATTAC;Rgs16::GFP mice. (F) Rgs16::GFP is expressed in Glut2+ β-cells in the recovering islet (6 weeks of β-cell recovery). Dotted white lines depict islet margin. Bars, 50 μm.

Alethia Villasenor, et al. Dis Model Mech. 2010 Sep-Oct;3(9-10):567-580.
3.
Fig. 1.

Fig. 1. From: Rgs16 and Rgs8 in embryonic endocrine pancreas and mouse models of diabetes.

Rgs8::GFP and Rgs16::GFP are expressed in the pancreatic bud. Rgs8 and Rgs16 are tandemly duplicated genes that are separated by 42,682 bp (43 kb) of intragenic DNA (Sierra et al., 2002). (A,B) Transgenic mice containing a BAC with enhanced GFP (eGFP) inserted at the translation initiation site of either Rgs8::GFP (GENSAT ID: BX478, constructed from RP23-184B11) (A) or Rgs16::GFP (GENSAT ID: BX843, constructed from RP23-101N8) (B) (Gong et al., 2003). (C,D) Expression of Rgs8::GFP (C) and Rgs16::GFP (D) in the dorsal pancreas during initial budding (arrows). Embryos were collected at the indicated stages. dp, dorsal pancreatic bud; nt, neural tube. Bar, 250 μm.

Alethia Villasenor, et al. Dis Model Mech. 2010 Sep-Oct;3(9-10):567-580.
4.
Fig. 8.

Fig. 8. From: Rgs16 and Rgs8 in embryonic endocrine pancreas and mouse models of diabetes.

Exendin-4 induces Rgs16::GFP expression in adult pancreas. Normoglycemic Rgs16::GFP BAC transgenic male mice (8–12 weeks) were injected (intraperitoneally) twice daily for 6 days with PBS, glucose, exendin-4 plus glucose (Ex+Glc), or exendin-4 alone (Ex). (A,B) After 3 days, Rgs16::GFP+ VDACs were observed but only in Ex+Glu-treated mice; (B) bright field of panel A (n=3/group). (C–F) After 6 days, Rgs16::GFP+ VDACs and/or islets were observed in Ex+Glu (C,D) or Ex-treated (E,F) mice; (D,F) bright field of panels C and E. (G) An islet from a PBS-injected mouse at day 3; GFP expression was not observed in any PBS-treated mice (n=2). (H) An islet of an Ex+Glc-injected mouse at day 5; GFP expression was observed in the islets of Ex+Glu-injected mice (n=2) (see supplementary material Fig. S11C). Dotted white lines depict islet margin. Bars, 100 μm (A–F); 50 μm (G–H).

Alethia Villasenor, et al. Dis Model Mech. 2010 Sep-Oct;3(9-10):567-580.
5.
Fig. 7.

Fig. 7. From: Rgs16 and Rgs8 in embryonic endocrine pancreas and mouse models of diabetes.

Rgs16::GFP is expressed in expanding islets of hyperglycemic ob/ob mice. (A) Rgs16::GFP is expressed in expanding islets of hyperglycemic ob/ob adult mice, but not in compound heterozygous mice (A), or in normoglycemic Rgs16::GFP;ob/ob or non-GFP ob/ob adults (supplementary material Fig. S7). (B,C) The number of islets expressing Rgs16::GFP generally increases with co-ordinate increases in glucose and insulin levels in Rgs16::GFP;ob/ob mice (see supplementary material Fig. S7). (C) Note the close association of Rgs16::GFP+ islets with large blood vessels (vessel, red arrowhead). (D–H) Double immunostaining for Rgs16::GFP and insulin (D), Glut-2 (E) and glucagon (F) demonstrates that Rgs16+ cells are found within islet β-cells but not α-cells, and are closely associated with blood vessels (PECAM) (G) and ducts (DBA) (H). (I) Cells with proliferative capacity (Ki67+, red arrows) are rare and equally distributed between cells that express Rgs16::GFP (white arrows) and those that do not express Rgs16::GFP. Dotted white lines depict islet margin. Bars, 100 μm (A–C,G,H); 50 μm (D–F,I).

Alethia Villasenor, et al. Dis Model Mech. 2010 Sep-Oct;3(9-10):567-580.
6.
Fig. 3.

Fig. 3. From: Rgs16 and Rgs8 in embryonic endocrine pancreas and mouse models of diabetes.

Rgs16::GFP is expressed in early pancreatic progenitors and in delaminated cells of the endocrine lineage at E15.5. Rgs16::GFP is initially expressed in a subset of cells within the early pancreatic epithelium, at E8.75, and is co-expressed with Sox9 (A) and Pdx1(B). (C) A subset of Rgs16::GFP cells can be seen within the pancreatic bud epithelium, as shown by E-cadherin staining, at E10.0. (D) Later, Rgs16::GFP expression becomes mutually exclusive with Sox9 and (E) overlaps with the endocrine marker glucagon. By E15.5, Rgs16::GFP is primarily not co-expressed with E-cadherin (F) or Sox9 (G) in the pre-ductal epithelium, or with Ngn3 in endocrine cells prior to their delamination (H) (note that, in panel F, E-cadherin is expressed at low levels in budding endocrine cells that express high levels of Rgs16::GFP, green arrow). A few (2%) Rgs16::GFP-expressing cells, however, still reside in the epithelium (see inset in F). At E15.5, Rgs16::GFP is co-expressed (white arrows) with markers of differentiating endocrine cells: Pdx1(I), nkx6.1(J), synaptophysin (K), insulin (L) and glucagon (M). It is almost completely co-expressed with pooled antibodies to all differentiated endocrine cells (anti-ins, -gluc, -somatostatin, -ghrelin). Rgs16::GFP is not co-expressed with amylase (N) in exocrine cells or with PECAM (O) in vascular cells. (G–O) Nuclei are identified by DAPI staining of DNA (blue). The insets in F–M display either co-expression (F,I–M) or lack of co-expression (G,H) of markers, as indicated. Confocal micrographs, 50× (A,B); 40× (C–O).

Alethia Villasenor, et al. Dis Model Mech. 2010 Sep-Oct;3(9-10):567-580.
7.
Fig. 4.

Fig. 4. From: Rgs16 and Rgs8 in embryonic endocrine pancreas and mouse models of diabetes.

Rgs16::GFP+ cells are associated with blood vessels during neonatal islet formation. Neonatal Rgs16::GFP expression is observed in coalescing islets throughout the postnatal pancreas (at postnatal day indicated). (A) Rgs expression is evident in endocrine cells as they aggregate into clusters along blood vessels at P0. (B) A forming islet (white arrow) can be seen overlying a blood vessel (white arrowhead). (C) Expression continues in islets until approximately P11 (short red arrows), but expression remains in scattered cells, or VDACs, along the axes of lateral branches (short white arrows). (D–F) At weaning (P16), the expression of Rgs16::GFP is no longer detectable in islets (D); however, expression continues in VDACs lining axial blood vessels at the center of lateral pancreatic branches (short white arrows) (E,F). (F) GFP+ cells (arrow) are tightly associated with tracts of blood vessels and associated pancreatic ducts, and are especially enriched at vascular branch points (red arrowhead; the red dotted line delineates a mature islet). (G) Expression vanishes by P27 (12 days past weaning). (C–E) The dotted white lines depict margins of lateral pancreatic branches in postnatal pancreas. (H) Schematic of VDACs location along the ‘triad’ composed of artery (red), vein (blue) and duct (yellow). The cross section shows the approximate location of VDACs (green) at the interface between ducts and blood vessels. Scale bars, 100 μm.

Alethia Villasenor, et al. Dis Model Mech. 2010 Sep-Oct;3(9-10):567-580.
8.
Fig. 2.

Fig. 2. From: Rgs16 and Rgs8 in embryonic endocrine pancreas and mouse models of diabetes.

Rgs16::GFP, Rgs8::GFP and Ngn3::GFP expression in the developing pancreatic bud. (A) Rgs16::GFP is broadly expressed in the endoderm at E8.5 but resolves (B) to the dorsal pancreas and liver bud by E9.5. (C) E10.5, expression in pancreatic endoderm, but not in mesoderm. (D,E) Expression in groups of cells along the central axis of the pancreatic tree, as branching elaborates. (F) In the postnatal pancreas, expression becomes restricted to forming islets (arrows). By contrast, (G) Rgs8::GFP is first expressed in a more localized domain within the foregut (arrowhead). (H–L) Similar to Rgs16, Rgs8 expression continues in scattered cells within the pancreatic endoderm between E9.5–10.5 (H,I), in the central region of the branching pancreatic bud between E12.5–E16.5 (J,K), and strongly in forming islets after birth (L). (G,M) Note that Rgs8::GFP is expressed in the dorsal pancreas prior to Ngn3::GFP (arrowheads), which is first detected in the dorsal pancreas at about E9.5 (compare G,H with M,N). (C–F,I–L,O–R) All three genes are expressed in the endocrine pancreas from E9.5 in a very similar pattern. (F,L,R) Arrows mark coalescing islets during postnatal development. The expression of Ngn3-eGFP was lower, but detectable, up to a few weeks post-birth. (D,E,J,K,P,Q) Dotted lines define the limit of the developing pancreas at each stage. Top row: schematics of the embryonic structures displayed in the rows below. aip, anterior intestinal portal; d, duodenum; dp, dorsal pancreatic bud; gte, gut tube endoderm; h, heart; l, liver; m, mesoderm; nf, neural folds; nt, neural tube; s, somite; vp, ventral pancreatic bud; ys, yolk sac. Bars, 100 μm (A–E,G–K,M–Q); 50 μm (F,L,R).

Alethia Villasenor, et al. Dis Model Mech. 2010 Sep-Oct;3(9-10):567-580.

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