Dual Roles for Glucokinase in Glucose Homeostasis as Determined by Liver and Pancreatic β Cell-specific Gene Knock-outs Using Cre Recombinase

Abstract

Glucokinase (GK) gene mutations cause diabetes mellitus in both humans and mouse models, but the pathophysiological basis is only partially defined. We have used cre-loxP technology in combination with gene targeting to perform global, ␤ cell-, and hepatocyte-specific gene knock-outs of this enzyme in mice. Gene targeting was used to create a triple-loxed gk allele, which was converted by partial or total Cre-mediated recombination to a conditional allele lacking neomycin resistance, or to a null allele, respectively. ␤ cell-and hepatocytespecific expression of Cre was achieved using transgenes that contain either insulin or albumin promoter/ enhancer sequences. By intercrossing the transgenic mice that express Cre in a cell-specific manner with mice containing a conditional gk allele, we obtained animals with either a ␤ cell or hepatocyte-specific knock-out of GK. Animals either globally deficient in GK, or lacking GK just in ␤ cells, die within a few days of birth from severe diabetes. Mice that are heterozygous null for GK, either globally or just in the ␤ cell, survive but are moderately hyperglycemic. Mice that lack GK only in the liver are only mildly hyperglycemic but display pronounced defects in both glycogen synthesis and glucose turnover rates during a hyperglycemic clamp. Interestingly, hepatic GK knock-out mice also have impaired insulin secretion in response to glucose. These studies indicate that deficiencies in both ␤ cell and hepatic GK contribute to the hyperglycemia of MODY-2.

Liver glucokinase gene expression is controlled by the onecut transcription factor hepatocyte nuclear factor-6

“…These mutations decrease the affinity of glucokinase for glucose and MODY2 seems to result from decreased glucose sensitivity and insulin secretion by the beta cell and from decreased liver glucose metabolism [4,5]. The hnf6-/-phenotype is not fully penetrant since 20 to 25% of the mice are not diabetic.…”

“…In these mice, liver glucokinase gene expression is reduced to barely detectable levels, presumably because the lack of stimulation of the liver glucokinase promoter by HNF-6 is combined with the lack of insulin stimulation of the promoter. This liver defect in turn is expected to contribute to glucose intolerance [5], so that the diabetes mellitus of hnf6-/-mice would result from a defective function of the pancreas as well as of the liver.…”

“…tions in the glucokinase gene are associated with a subtype of MODY, MODY2 [2,3] and inactivation of the glucokinase gene causes Type II (non-insulindependent) diabetes mellitus in the mouse [4,5].…”

Pancreatic deletion of insulin receptor substrate 2 reduces beta and alpha cell mass and impairs glucose homeostasis in mice

“…Deletion of the insulin-like growth factor 1 receptor gene (Igf1r) likewise impairs insulin synthesis and secretion and combined deletion of the insulin receptor gene (Insr) and Igf1r causes marked beta cell failure [6,7]. Using similar techniques, we and others have recently generated 'beta cell-specific' Irs2-null mice that have utilised a rat insulin promoter 2 Cre recombinase transgenic mouse (RIPCre or B6.Cg-tg(Ins2-cre)25Mgn/J) to delete the floxed alleles of Irs2 [8][9][10][11]. Our studies and those of others have demonstrated that beta cell deletion of Irs2 reduces beta cell mass and impairs glucose tolerance.…”

Impact of whole body irradiation and vascular endothelial growth factor-A on increased beta cell mass after bone marrow transplantation in a mouse model of diabetes induced by streptozotocin

“…Green fluorescent protein (GFP) transgenic mice, in which enhanced GFP production was under the control of the cytomegalovirus enhancer and the chicken β-actin promoter, were generously provided by M. Okabe (Osaka University, Osaka, Japan) [24]. The generation and characterisation of RIP-CRE transgenic mice [25] and vascular endothelial growth factor (VEGF fl/fl ) mice (VEGF fl/fl [CRE − ] mice) [26] has been reported previously [27,28].…”