The mechanism of Qingre Huoxue formula regulating PKC/IP3R/GLUT4 signaling pathway on diabetic lower limb vascular disease model
JIN Mu-yang,MA Lu-bo,LIU Jian-gang
Abstract:
Objective To through the rat model of chronic ischemia of lower limbs caused by diabetes combined with arteriosclerosis, to study the mechanism of Qingre Huoxue formula regulating PKC signaling pathway in lower limb angiopathy of diabetes. Methods Fifty SPF-grade rats were selected, and type 2 diabetes was induced by feeding them a high-fat diet and intraperitoneal injection of streptozotocin. An animal model of chronic lower limb ischemia caused by diabetes complicated with arteriosclerosis was established by intravascular balloon injury to the femoral artery endothelium. The rats were randomly divided into 5 groups with 10 rats in each group: model group (equal volume of purified water), simvastatin group[3.6 mg/(kg·d)], small-dose Qingre Huoxue formula group [6.3 g/(kg·d)], large-dose Qingre Huoxue formula group [12.6 g/(kg·d)], and sham operation group (equal volume of purified water). Gastric administration was performed regularly for 4 consecutive weeks. The sham operation control group only received vascular puncture and guidewire exchange without femoral artery injury, no streptozotocin injection, and was fed with a normal diet. Pathological sections were prepared from the affected blood vessels of the rats. HE and Masson staining were used to observe the changes in vascular structure, as well as the distribution of collagen, smooth muscle cells, and endothelial cells in the vascular wall. Routine biochemical methods were used to determine the levels of glucose (GLU), total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C), and high-density lipoprotein cholesterol (HDL-C). ELISA was used to detect the content of advanced glycation end products (AGEs). Western blotting was used to detect the expressions of protein kinase C(PKC), inositol 1,4,5-trisphosphate receptor (IP3R), vascular endothelial growth factor (VEGF), glucose transporter-4 (GLUT-4), and angiopoietin-1 (ANG-1). Results Vascular pathology showed that compared with the sham surgery group, the model group had uneven vascular thickness, with hyperplasia of the intima and media, increased number and uneven arrangement of VSMCs; All treatment groups showed improvement. Compared with the sham surgery group, the levels of GLU, TC, TG, AGEs, GLUT4, PKC, and VEGF in the model group increased (P <0.05), and there was an increasing trend in HDL and LDL, but there was no statistical difference (P >0.05); Compared with the model group, the simvastatin group showed a significant increase in GLU levels (P <0.05), while other indicators showed varying degrees of improvement (P >0.05); Compared with the model group, the levels of TG, TC, and AGES in the low-dose group were significantly reduced (P <0.05), while the levels of PKC, VEGF, IP3R, ANG-1, and GLUT4 were significantly increased (P <0.05). The levels of HDL and LDL improved, but there was no statistical difference (P >0.05); High dose group GLU TC、TG, significantly decreased (P <0.05), while the levels of PKC, ANG-1, VEGF, and IP3R significantly increased (P <0.05). Conclusion Qingre Huoxue formula can significantly improve the metabolism of glucose and lipid in rats, promote the formation of new blood vessels, reduce the deposition of vascular collagen fibers, and inhibit the development of atherosclerosis after intravascular injury in lower limbs of diabetes rats. Its pathological mechanism is related to the expression of related proteins regulating PKC/IP3R/GLUT4 signaling pathway.
