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. Author manuscript; available in PMC: 2016 Apr 8.
Published in final edited form as: Cell Stem Cell. 2015 Oct 1;17(4):486–498. doi: 10.1016/j.stem.2015.07.010

Figure 5. Treatment of long-standing T1D with SPK replenishes CoSCs and restores the CoSC signature profile and mini-gut development through restoration of circulating IGF-I and IGFBP3.

Figure 5

A, B, C. Bar graphs depict results of flow cytometric analysis of EphB2hi+, EphB2hi+LGR5+, EphB2+h-TERT+ cells obtained from isolated crypts in long-standing T1D (Baseline), T1D+ESRD who underwent kidney pancreas (SPK) or kidney alone (K+T1D) transplantation at 8 years of follow-up. n=10 subjects per group were evaluated. D, E, F. Bar graphs depict normalized mRNA expression of intestinal stem cell markers EphB2, LGR5, h-TERT, measured by quantitative RT-PCR on isolated intestinal crypts obtained from long-standing T1D (Baseline), T1D+ESRD who underwent kidney pancreas (SPK) or kidney alone (K+T1D) transplantation at 8 years of follow-up. All samples were run in triplicate and normalized to expression of the housekeeping gene ACTB using the ΔΔCt method. n=10 subjects per group were evaluated. G. Western blot analysis depicts the expression of EphB2, LGR5, h-TERT in isolated intestinal crypts of the four groups at 8 years of follow-up. n=5 subjects per group were evaluated. H. Bar graph depicts the % of developed mini-guts of the total at 8 days of culture of freshly isolated intestinal crypts obtained from long-standing T1D individuals (Baseline), SPK and K+T1D subjects at 8 years of follow-up. n=10 subjects per group were evaluated. I. Heat map represents the CoSC signature marker transcriptomic profiling examined in freshly isolated intestinal crypts of CTRL, long-standing T1D individuals (T1D+ESRD), SPK and K+T1D subjects at 8 years of follow-up. n=10 subjects per group were evaluated. J. Bar graph represents IGF-I levels measured by ELISA in serum of the four groups of subjects at 8 years of follow-up. n=10 subjects per group were evaluated. K. Bar graph depicts IGFBP3 levels measured by ELISA in serum of the four groups of subjects. n=20 subjects per group were evaluated. L, M Correlation between IGFBP3 serum levels and intestinal symptoms assessed using the GSRS questionnaire (0-7) in n=20 subjects of K+T1D (L) and SPK (M) group. Analysis was conducted using ANOVA (p<0.05) in comparing all groups. Data are expressed as mean ± standard error of the mean (SEM) unless differently reported. *p<0.01; **p<0.001; ***p<0.0001. See also Figure S5 and S6.

Abbreviations: CoSC, colonic stem cell; T1D, type 1 diabetes; ESRD, end stage renal disease; CTRL, healthy subjects; SPK, simultaneous kidney-pancreas transplantation; EphB2, Ephrin B receptor 2; LGR5, leucine-rich repeat containing G protein-coupled receptor 5; RT-PCR, real-time polymerase chain reaction; ACTB, beta actin; K+T1D, kidney transplantation alone in type 1 diabetes; IGF-I, insulin-like growth factor 1; IGFBP3, insulin-like growth factor binding protein 3; SEM, standard error of the mean.

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