Abstract: In this study, three fusion proteins were constructed by linking Fc fragment, membrane penetrating peptide R11 and membrane penetrating peptide TAT to cFGF-21 respectively, aiming to investigate the long-lasting hypoglycemic effect of cFGF-21 on type 1 diabetes. A mouse model of type 1 diabetes was established with streptozotocin (STZ) and subsequently treated with cFGF-21, R11-cFGF-21, Fc-cFGF-21 and TAT-cFGF-21, respectively. Treatment was divided into three phases: 1) daily dosing (1-14 days); 2) every 3 days (15-29 days); and 3) every 5 days (30-44 days). During the treatment, mice were tested for blood glucose every three days, and at the end of each phase, experimental mice were tested for blood glucose fluctuations over 12 h. Before the end of the experiment, oral glucose tolerance test was performed on all mice. The expression levels of inflammation-related factors IL-10, IL-1β, IL-17A and anti-inflammatory factor IL-10 in mice serum were measured by ELISA. The transcript levels of glucose metabolism-related factors in liver, fat, intestine and kidney tissues were detected by Real-time PCR. At the end of the experiment, all mice were tested for glycosylated hemoglobin (HbA1c), lipid quadruple and liver function, serum insulin and HbA1c and pathological analysis of mouse pancreatic tissues were performed. In the first phase of treatment, the hypoglycemic effect of TAT-cFGF-21 was significantly lower compared with the cFGF-21 group, and there was no significant difference between the R11-cFGF-21 and Fc-cFGF-21 groups; in the second and third phases of treatment, compared with the cFGF-21 group, the blood glucose in the TAT-cFGF-21 group increased significantly, and the R11-cFGF-21 and Fc-cFGF-21 groups could control blood glucose at a lower level within 3-5 days of dosing. After 8 weeks of treatment, oral glucose tolerance (OGTT) and dyslipidemia were significantly improved in the R11-cFGF-21 and Fc-cFGF-21 groups compared to the cFGF-21 group, glycosylated hemoglobin (HbA1c) was near normal levels, and liver damage was significantly repaired, while there was no significant difference between the TAT-cFGF-21 and cFGF-21 groups. In addition, ELISA and HE staining results showed that oxidative stress and inflammatory response were significantly improved in the R11-cFGF-21 and Fc-cFGF-21 groups compared with the cFGF-21 and TAT-cFGF-21 groups, and the islet β-cell repair effect was obvious. The significant increase of long-term hypoglycemic capacity of cFGF-21 modified by R11 and Fc, while the long-term hypoglycemic capacity of TAT-cFGF-21 was weaker. In addition, cFGF21 modified by R11 and Fc significantly improved oxidative stress, pancreatic inflammation and lipid metabolism, and repaired liver injury and pancreatic β-cells, with significantly better therapeutic effects than cFGF21.

Key words: canine diabetes mellitus, fibroblast growth factor 21, cFGF-21, Fc, membrane penetrating peptide, long-acting