FIG. 1.

Upregulation of HES-1 in cultured human islet cells and eGFP⁺ cells derived from β-cells correlates with downregulation of insulin. A: qPCR analysis of RNA extracted from islet cells derived from nine donors. P indicates passage number and weeks in culture. RQ indicates relative quantification compared with P0, which represents islet cells at culture initiation. Data are means ± SD (n = 9). Asterisks indicate statistical significance, compared with P0 (P < 3.3 × 10⁻⁵ for insulin and P < 0.006 for HES1). The increase in HES1 mRNA levels at P1 was marginally significant (P = 0.06). B: Immunoblotting for HES-1 in protein extracted from islet cells at the indicated passage number. β-Actin served as a loading control. C: Immunoblotting for HES-1 in protein extracted from islet cells at P2, following a 17-h incubation with the γ-secretase inhibitor l-685458. HSC70 served as a loading control. D: Immunofluorescence analysis of islet cells (left panel) and eGFP⁺ cells derived from β-cells (right panel) following 10 days in culture. The left panel is merged with a phase contrast image. Arrow points to a β-cell that still expresses insulin and is not labeled for HES-1. eGFP is detected in both cytoplasm and nucleus. Bar = 20 μm. (Please see http://dx.doi.org/10.2337/db07-1323 for a high-quality digital representation of this figure.)