Mice harboring the human SLC30A8 R138X loss-of-function mutation have increased insulin secretory capacity

Abstract

SLC30A8 encodes a zinc transporter that is primarily expressed in the pancreatic islets of Langerhans. In β-cells it transports zinc into insulin-containing secretory granules. Loss-of-function (LOF) mutations in SLC30A8 protect against type 2 diabetes in humans. In this study, we generated a knockin mouse model carrying one of the most common human LOF mutations for SLC30A8, R138X. The R138X mice had normal body weight, glucose tolerance, and pancreatic β-cell mass. Interestingly, in hyperglycemic conditions induced by the insulin receptor antagonist S961, the R138X mice showed a 50% increase in insulin secretion. This effect was not associated with enhanced β-cell proliferation or mass. Our data suggest that the SLC30A8 R138X LOF mutation may exert beneficial effects on glucose metabolism by increasing the capacity of β-cells to secrete insulin under hyperglycemic conditions.

Keywords: SLC30A8; genetic mutation; insulin secretion; pancreatic beta cell; zinc transporter.

Fig. 1.

Analysis of Slc30a8 RNA and protein in islets from male R138X mice on chow diet. (A) Slc30a8 RNA in situ hybridization of pancreatic islets isolated from wild-type, knockout, and R138X mice. KO islets were used as negative control. Red, glucagon RNA; green, insulin RNA; white, Slc30a8 RNA. (B) Quantification of islet Slc30a8 RNA levels using qPCR analysis. n.d., not detected. (C) Western blot of islets isolated from chow-fed WT, KO, and R138X mice. KO islets were used as negative control. The arrow indicates SLC30A8 protein; asterisks denote unspecific bands. (D) Dithizone staining of pancreatic islets isolated from WT, KO, and R138X mice.

Fig. 2.

Metabolic phenotype of male R138X mice on chow diet. (A–C) Body weight (A), blood glucose (B), and plasma insulin (C) in fed and fasted WT and R138X mice (n = 18 to 20 per genotype). (D) Plasma insulin levels in WT or R138X mice after an overnight fast (time 0) and at the indicated times after an i.p. injection of glucose. Data are displayed as blood insulin levels over time (n = 6 per genotype). (E) Oral glucose tolerance test in overnight-fasted (time 0) WT and R138X mice. Data are displayed as blood glucose over time (n = 6 per genotype). (F) Insulin tolerance test in 4-h-fasted WT and R138X mice. Data are displayed as blood glucose levels over time (n = 6 per genotype). (G) Histology for insulin in pancreas isolated from WT and R138X mice. (H) Quantification of pancreatic insulin staining (n = 6 per genotype). Values represent the means ± SEM. Data were analyzed by two-way ANOVA or Student’s t test. No significance was reached.

Fig. 3.

R138X mice secrete more insulin under chronic hyperglycemia caused by the insulin receptor antagonist S961. (A) Nonfasted plasma glucose levels in R138X and WT mice continuously treated with the insulin receptor antagonist S961 (20 nmol/wk) or PBS for 22 d. (B) Nonfasted plasma insulin on day 0, 4, 19, and 22 R138X and WT mice treated with S961 (20 nmol/wk) or PBS. (C) Plasma active GLP-1 levels in WT and R138X mice after 22 d of treatment. (D and E) Fed and fasted glucose (D) and insulin (E) levels measured day 19 and 20 after initiation of treatment. (F) Immunohistochemistry for Ki-67 (white), insulin (green), and glucagon (red). (G) Quantification of Ki-67 and insulin double-positive cells. (H) Quantification of pancreatic insulin staining shown in I. (I) Histology for insulin in pancreas isolated from WT and R138X mice after 22 d of treatment. Values represent the means ± SEM (n = 5 to 7 mice per treatment and genotype). *P < 0.5, **P < 0.01, ****P < 0.0001.

Fig. 4.

Gene expression in islets from WT versus R138X mice. (A) RPKM values for insulin 2 and insulin 1. (B) RPKM values for β-cell regulators. (C) RPKM values for Hvcn1. **P < 0.01 defined by DESeq2 (Wald test) (24).