Abstract
Previous studies reported that secreted frizzled-related protein-5 (Sfrp5) decreases beta cell proliferation and increases fasting insulin levels, but studies on direct effects of Sfrp5 on insulin secretion and its underlying mechanisms are missing. This study examined effects of Sfrp5 on (i) beta cell viability and proliferation, (ii) basal and glucose-stimulated insulin secretion and (iii) canonical and non-canonical Wnt signalling pathways. We incubated rat INS-1E cells with 0.1, 1 or 5 μg/ml recombinant Sfrp5 for 24h. We measured basal and glucose-stimulated insulin secretion at glucose concentrations of 2.5 and 20 mmol/l. Phosphorylated and total protein content as well as mRNA levels of markers of cell proliferation, canonical and non-canonical Wnt signalling pathways were examined using Western blotting and real-time PCR. Differences between treatments were analysed by repeated measurement one-way ANOVA or Friedman’s test followed by correction for multiple testing using the Benjamini-Hochberg procedure. At 5 μg/ml, Sfrp5 reduced mRNA levels of cyclin-B1 by 25% (p<0.05). At 1 and 5 μg/ml, Sfrp5 increased glucose-stimulated insulin secretion by 24% and by 34% (both p<0.05), respectively, but had no impact on basal insulin secretion. Sfrp5 reduced the phosphorylation of the splicing forms p46 and p54 of JNK by 39% (p<0.01) and 49% (p<0.05), respectively. In conclusion, Sfrp5 reduced markers of cell proliferation, but increased in parallel dose-dependently glucose-stimulated insulin secretion in INS-1E cells. This effect is likely mediated by reduced JNK activity, an important component of the non-canonical Wnt signalling pathway.
Highlights
The secreted frizzled-related protein (Sfrp)5 belongs to the Sfrp family, the largest group of WNT inhibitors [1]
The treatment of INS-1E cells with 0.1, 1 or 5 μg/ml secreted frizzled-related protein-5 (Sfrp5) for 24h had no impact on the ATP content of these cells as marker for cell viability compared to untreated cells (S2 Fig, S1 Data)
We did not find any differences between untreated INS-1E cells and those treated with the different concentrations of Sfrp5
Summary
The secreted frizzled-related protein (Sfrp) belongs to the Sfrp family, the largest group of WNT inhibitors [1]. Sfrp is a secreted protein which is produced by several human tissues such as visceral and subcutaneous adipose tissue, liver, mononuclear blood cells and pancreatic islets [2,3,4,5]. The impact of Sfrp on the non-canonical Wnt signalling pathway has not been investigated in these cells. Two epidemiological studies investigated the association between Sfrp and markers for insulin secretion. Sfrp is downregulated in pancreatic islets of obese rodents and humans [4]. The administration of Sfrp reduced the proliferation in primary islet cells [4] and the overexpression of Sfrp led to higher serum fasting insulin levels in mice [9]
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