Decreased blood vessel density and endothelial cell subset dynamics during ageing of the endocrine system (2020)
Figure 7

Figure 7. Endothelial Gja1 regulates β-cell expansion

A Representative tile scan 3D images with α-SMA, Isolectin, FABP4 and Emcn in the pancreas from 17-weeks-old Gja1i∆EC mutant and control mice. Insets show higher magnifications of regions.

B Box whiskers, and scatter plot (left) shows the quantification of CD31+ Emcnhi vessel density in Gja1i∆EC mutant and control pancreatic islets. (n=7), P-value, and two-tailed unpaired t-test. Combo plot (right) shows the quantification of P-Histone H3+ EC numbers per islet (f.u) counted on thick murine pancreatic sections. (n=5), P-value, and two-tailed unpaired t-test.

C 3D images show CD31 and Emcn immunostaining in the Gja1i∆EC mutant and control pancreas. The scale bars are 50 µm.

D Combined violin, scatter and sample distribution plot shows the quantification of β-cell mass (%) in pancreatic islets from Gja1i∆EC and control mice. Data represent mean ± s.d. (n=140 islets from 7 pancreas), P-value, and two-tailed unpaired t-test.

E Representative 3D images with immunostaining of α-SMA, Insulin and VE-Cadherin in the pancreatic islets from Gja1i∆EC and control mice. The scale bars are 50 µm.

F Combo plot (left) shows insulin measurement in lysate of pancreas by ELISA. Combo plot (right) shows the percentage of Insulin+ β-cells per islet in Gja1i∆EC mutant and control mice. (n=5), P-values, and two-tailed unpaired t-tests.

G Representative 3D images of ESM-1 and Caveolin-1 in pancreatic islets from Gja1i∆EC and control mice. Insets (* and #) show higher magnifications with surface rendering for ESM-1 and Caveolin-1 immunostaining. Combo plot with the quantification of ESM-1+ cell numbers per islet (f.u). (n=5), P-value, and two-tailed unpaired t-test. The scale bars for the 3D images are 50 µm, and 5 µm for the higher magnifications with single-cell resolution images.

H 3D images with CXCR4 and Emcn immunostaining in Gja1i∆EC and control pancreas. The scale bars are 50 µm.

I Representative 3D images show CXCR4, Ki67 and VE-Cadherin immunostaining in Gja1i∆EC and control pancreas. The scale bars are 50 µm.

J Combo plots show the percentage of CXCR4+ β-cells per islet and Ki67+ β-cells normalized to the total number of β-cells per islet in Gja1i∆EC mutant versus the littermate control mice. (n=5), P-values, and two-tailed unpaired t-tests.

K Box whiskers, and scatter plot shows the quantification of CD31+ Emcnhi vessel density in aged Gja1i∆EC mutant and control pancreatic islets. Data represent mean±s.d. (n=4), P-value, and two-tailed unpaired t-test. Combo plot (below) shows the quantification of β-cells on thick murine pancreatic sections. Data represent mean±s.d. (n=4), P-value, and two-tailed unpaired t-test.

Data information: The dashed lines in panel A represent the outlines of pancreas. The dashed lines in panel C, E, G, H and I represent the outlines of pancreatic islets. Two-tailed Student's t-test were performed for the statistical analysis. **: P < 0.01; ****: P < 0.0001. Nuclei: DAPI or TO-PRO-3 as indicated. Is: pancreatic islet. f.u: fluorescence unit. The line on the right side of the combo plots represent the sample distribution. Scale bars are 200 µm for tile scan images of longitudinal thick sections across the entire glands, and 50 µm for the higher magnification insets images.

Legends to Expanded View Figures

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