Leptin, a hormone produced mainly by adipose tissue, is believed to balance energy levels in vivo by regulation of food intake and body weight. Higher concentrations of leptin are observed in obese people, who are thought to be “leptin resistant.” Leptin resistance is common in patients with diabetes and studies show leptin can exhibit glucoregulatory effects on patients with type 1 and type 2 diabetes. In spite of this, obese diabetic patients do not exhibit a strong response to exogenous leptin. Thus, a detailed understanding of the glucose regulatory function of leptin is of crucial importance in order to overcome the shortcomings of leptin therapy and its use as a potential therapeutic for humans. We have provided a novel aspect on leptin function by investigating it in the blood stream. Previously, our group has reported that C-peptide, a 31 amino acid peptide secreted from pancreatic beta cells, binds to red blood cells (RBCs) and has cellular energetic effects. Here, we show that leptin actually amplifies the effects of C-peptide. ATP release from RBCs was measured in the presence of leptin, C-peptide, zinc, and combinations thereof, while also monitoring the translocation of GLUT-1 to the RBC membrane. In the presence of C-peptide and zinc, a 30% increase in RBC-derived ATP is measured; this signal is enhanced by another 20% in the presence of leptin. This effect was also amplified under high-glucose conditions, as seen in diabetes. however, leptin alone showed no effect on RBCs. Interestingly, leptin in the presence of zinc and C-peptide increases GLUT-1 translocation by 20% compared to the control and samples including zinc and C-peptide only. ATP measurements were performed using Luciferin/ Luciferase chemiluminescence assay and GLUT-1 translocation was monitored by SDS- PAGE gel and western blot techniques. We have also measured an increase in the amount of C-peptide bound to RBCs in the presence of leptin, and a saturable dose dependent response was observed between C-peptide and RBCs. Binding studies were performed using C-peptide and leptin enzyme linked immunosorbent assay (ELISA) respectively. Our studies show that levels of leptin in the bloodstream may be correlated with glucose concentrations, and the ability of RBCs to use glucose. Therefore, leptin along with zinc and C-peptide can potentially be targeted as a therapeutic for diabetes.
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