Differentiation of bone marrow mesenchymal stem cells into insulin producing cells by liraglutide and effect on type 1 diabetes rats

Objective To investigate the differentiation effect of liraglutide on bone marrow mesenchymal stem cells (BM-MSCs) into insulin producing cells (IPCs) in vitro, and further to observe the therapeutic action of IPCs on type 1 diabetes(T1DM) rats. Methods BM-MSCs were separated and purified by density gradient centrifugation combined with attachment culture method in vitro, and they were divided into control group, high glucose with nicotinamide induction group, glucagon-like peptide 1 (GLP-1) induction group and liraglutide induction group. The morphological change of cells was observed by inverted microscope during induction, and they were confirmed by dithizone (DTZ) staining. Gene measurement of Nestin, pancreatic and duodenal homeobox 1(PDX-1), glucokinase(GK), glucose transporter-2(Glut-2), insulin and glucagon were tested by Real-time PCR. Protein expression of insulin and glucagon were detected by cell immunofluorescence. Thirty SD rats (weight 180-220 g) were injected 60 mg/kg of streptozotocin to establish the type 1 diabetes model, then they were randomly divided into control group (group T1DM, n=8), BM-MSCs transplantation group (group BM-MSCs, n=9)and BM-MSCs induced by liraglutide transplantation group (group LIRA+ BM-MSCs, n=9). Random blood glucose was monitored regularly during the treatment for 8 weeks. In addition, the intraperitoneal injection of glucose tolerance test (IPGTT) was carried out after the blood glucose was stable, by which another 4 normal rats were selected as control. Results (1) In vitro, BM-MSCs showed distinct tendency to form clumps at confluence when liraglutide was added, and dithizone staining was positive. Compared to high glucose with nicotinamide induction group, the mRNA expression of Nestin was lower in liraglutide induction group (0.003 8±0.000 4 vs 0.007 5±0.003 0), while the level of insulin(0.000 20±0.000 03 vs 0.000 08±0.000 02)and glucagon(0.001 1±0.000 4 vs 0.000 7±0.000 1, F=7.26, 10.06, 4.92, all P<0.05) was higher. The mRNA expression of PDX-1, Glut-2, GK was also increased. The expressions of insulin and glucagon protein were positive in liraglutide and GLP-1 induction group. (2) In vivo, compared to group T1DM, the concentration of randomly blood glucose was significantly decreased both in group LIRA+ BM-MSCs and group BM-MSCs((28.0±1.2), (8.9±1.1), (14.5±0.9)mmol/L, respectively, F=719.61, all P<0.05). IPGTT showed blood glucose reached to a peak at 30th minutes and dropped to fasting glucose at 150th minutes in group LIRA+ BM-MSCs, which was similar to normal. Conclusions BM-MSCs could be enhanced to differentiate into IPCs by liraglutide in vitro, and further to make effect by IPCs in vivo. Key words: Diabetes mellitus, type 1; Liraglutide; Bone marrow mesenchymal stem cells; Differentiation; Insulin producing cells