Application of cyclopeptide COX52-69 in inhibition of proliferation of smooth muscle cells induced by high glucose

Abstract

The vasculopathy of hyperglycemia is a common and highly harmful complication of diabetes mellitus, and is characterized by abnormal accumulation of vascular smooth muscle cells (VSMCs) in vascular intima and media due to excessive proliferation and reduction of apoptosis of the VSMCs. This proliferation and apoptosis imbalance of VSMCs is considered to be likely to play a key role in the pathogenesis and progression of diabetic vascular complications. According to the invention, the cyclopeptide COX52-69 is found to be capable of directly inhibiting proliferation and migration of smooth muscle cells induced by high glucose. The evidence of the cyclopeptide is that in CCK8 cell proliferation detection and cell migration experiments, we find that the cyclopeptide can significantly inhibit abnormal proliferation and migration of high-glucose-induced smooth muscle cells, while the cyclopeptide with the same concentration acts on high-glucose-induced HEK293T cells to perform CCK8 cell proliferation detection. However, the cyclopeptide is found to have no influence on proliferation of HEK293T cells in high glucose (no promotion or inhibition effect). Therefore, the peptide has targeting specificity on the inhibition effect on proliferation and migration of high-glucose-induced smooth muscle cells, and also has certain biological safety. High glucose-induced proliferation and migration of smooth muscle cells are main causes of diabetic vasculopathy, so that the peptide can be used for complications caused by the diabetic vasculopathy. A new hope is brought to intervention of diabetic complications.

Description

Technical Field

[0001] This invention belongs to the field of biomedicine, specifically relating to a novel cyclic peptide COX with activity that inhibits the abnormal proliferation of vascular smooth muscle cells induced by high glucose. 52 - 69 Use in the preparation of medicines for the prevention and / or treatment of complications caused by diabetic vascular disease. Background Technology

[0002] Diabetes mellitus, a common chronic metabolic disease, has a high incidence rate worldwide. Hyperglycemia is the core characteristic of diabetes, and vascular complications caused by hyperglycemia are a common and extremely harmful complication. Vascular smooth muscle cells (VSMCs) play a crucial role in the development and progression of diabetic vascular complications. Under normal circumstances, VSMCs maintain the structural and functional stability of blood vessels. However, in a hyperglycemic environment, VSMCs undergo abnormal changes, specifically manifested as abnormal accumulation in the endothelium and media of blood vessels. This accumulation is mainly attributed to excessive proliferation and reduced apoptosis of VSMCs, leading to an imbalance between VSMC proliferation and apoptosis. This imbalance is considered to play a key role in the pathogenesis and progression of diabetic vascular complications. As the disease progresses, hyperglycemia-induced vascular complications lead to a series of serious consequences. For example, it can cause thickening of the vascular endothelium, narrowing the lumen of blood vessels and impairing blood flow. Obstructed blood flow further leads to increased blood pressure, increasing the burden on the heart, and also affecting the blood supply to various organs of the body. Prolonged hyperglycemia can also lead to various diabetic vascular complications such as atherosclerosis, calcification of the endothelial media, and peripheral artery disease, seriously threatening patients' health and quality of life, and even endangering their lives. Currently, there are no effective treatments or drugs for complications caused by diabetic vascular complications. Therefore, finding new therapeutic targets and drugs is of great clinical significance. Summary of the Invention

[0003] To address the problems existing in the prior art, this invention discovers the cyclic peptide COX. 52 - 69 (This cyclic peptide was discovered by our laboratory and a patent application has been filed previously.) It has significant effects in preventing and / or treating complications caused by diabetic vascular disease. Specifically, it can inhibit the proliferation and migration of vascular smooth muscle cells (VSMCs) induced by high glucose, thereby inhibiting adverse conditions such as vascular intimal thickening, poor blood flow, and high blood pressure caused by hyperglycemia. It can be used to prevent and treat diabetic vascular disease, including atherosclerosis, vascular intimal thickening, medial calcification, or peripheral artery disease. Technical solution

[0004] It significantly inhibits the proliferation of VSMCs (as confirmed by CCK-8 assay).

[0005] Significantly inhibits VSMC migration (as confirmed by scratch / Transwell experiments).

[0006] It exhibits certain biosafety: at the same concentration, it has no inhibitory or promoting effect on the proliferation of HEK293T cells treated with high glucose. Function

[0007] Inhibition of VSMC proliferation and migration: Through relevant experimental studies, including CCK8 cell proliferation assays and cell migration experiments, the cyclic peptide COX was found to inhibit VSMC proliferation and migration. 52 - 69 It can directly act on vascular smooth muscle cells (VSMCs), significantly inhibiting high glucose-induced abnormal proliferation and migration of VSMCs. Under high glucose conditions, the proliferation and migration activities of VSMCs are enhanced, while the cyclic peptide COX... 52 - 69 This can interfere with the abnormal process, regulate the physiological state of VSMCs, restore their normal function, and thus reduce the abnormal accumulation of VSMCs in the vascular intima and media.

[0008] Inhibits symptoms related to vascular lesions: due to the cyclic peptide COX 52 - 69 It inhibits the abnormal proliferation and migration of VSMCs, thereby suppressing vascular intimal thickening caused by hyperglycemia. Vascular intimal thickening is a key characteristic of diabetic vascular complications, narrowing the blood vessel lumen and affecting normal blood flow. Cyclic peptide COX 52 - 69 Its function is to maintain the normal diameter of blood vessels and ensure smooth blood flow. Once poor blood flow is improved, elevated blood pressure will also be alleviated, reducing the risk of cardiovascular diseases and other complications caused by hypertension.

[0009] Prevention and treatment of various diabetic vascular complications: The diabetic vascular complications covered in this invention encompass a variety of types, including atherosclerosis, intimal thickening, medial calcification, and peripheral artery disease. Cyclic peptide COX 52 - 69 By regulating the function of VSMCs, the physiological state of blood vessels can be improved, thereby preventing and inhibiting the occurrence and development of these vascular lesions at their root. For example, in the formation of atherosclerosis, the abnormal proliferation and migration of VSMCs are crucial steps, and the cyclic peptide COX... 52 - 69 Its inhibitory effect can reduce the formation of atherosclerotic plaques; for medial calcification, the cyclic peptide COX 52 - 69 It can regulate calcium metabolism-related signaling pathways and inhibit abnormal calcium deposition in the vascular media, thereby preventing the occurrence of media calcification. Attached Figure Description

[0010] Figure 1 This represents the proliferation rate of aortic vascular smooth muscle cells under high glucose conditions compared to the normal glucose group in this embodiment.

[0011] Figure 2 The figures represent the inhibition rates of different concentrations of cyclic peptides on high glucose-induced abnormal proliferation of aortic vascular smooth muscle cells in the embodiments of the present invention.

[0012] Figure 3 This example illustrates the effect of different concentrations of cyclic peptides on the proliferation of HEK293T cells under high glucose conditions.

[0013] Figure 4 , Figure 5 This is a comparison diagram showing the inhibition of high glucose-induced aortic vascular smooth muscle cell migration by different concentrations of cyclic peptides in embodiments of the present invention. Detailed Implementation

[0014] The present invention will now be described in detail with reference to specific embodiments, but the scope of protection of the present invention is not limited to the following embodiments.

[0015] Example 1: Male Wistar rats weighing between 120g and 150g were selected. After isoflurane anesthesia, the rats were quickly euthanized by dislocation. Following thorough alcohol disinfection, the aorta was isolated in a sterile laminar flow hood. Peripheral fat was removed, the adventitia was peeled off, the intima was gently scraped away, and the media was preserved and minced. Primary vascular smooth muscle cells were obtained using the tissue block adhesion method. When the cell density reached above 80%, the cells were passaged. The obtained vascular smooth muscle cells were seeded into a 96% blank medium and cultured for 24 hours in serum-free DMEM containing 5.5mM, 25mM, and 50mM glucose, respectively. CCK8 proliferation assays were then performed. The results showed that the cell survival rate of vascular smooth muscle cells significantly increased with increasing glucose content, indicating cell proliferation.

[0016] Example 2: Vascular smooth muscle cells in the logarithmic growth phase were seeded into a 96 blank, and COX containing 1 μM, 500 nM, and 100 nM were added respectively. 52 - 69 The polypeptide was dissolved in dmembrane containing 25 mM glucose and cultured for 24 hours. CCK8 proliferation assay was performed. The results showed that different concentrations of the polypeptide could significantly inhibit the abnormal proliferation of vascular smooth muscle cells induced by high glucose, and the highest concentration of 1 μM cyclic peptide had the most significant inhibitory effect.

[0017] Example 3: HEK293T cells in the logarithmic growth phase were seeded into a 96% blank, and COX was added at concentrations of 1 μM, 500 nM, and 100 nM, respectively. 52 - 69 This polypeptide was dissolved in dMEM containing 25 mM glucose. After culturing for 24 hours, CCK8 proliferation was detected. The results showed that different concentrations of the polypeptide had no effect on the high glucose-induced proliferation of HEK293T cells, demonstrating that concentrations of 1 μM, 500 nM, and 100 nM COX... 52 - 69Cyclic peptides have a certain degree of biocompatibility.

[0018] Example 4: Primary smooth muscle cells in the logarithmic growth phase were seeded into 6-well plates. When the cell density reached approximately 90%, cell scratching was performed using the same white pipette tip. After scratching, cell debris was thoroughly washed away with PBS, and the cells were divided into a normal glucose group, a high glucose group, and a high glucose plus COX group. 52 - 69 In the treatment group, cell migration area was recorded at 0h, 24h, and 48h. The experimental results indicate that high glucose plus COX... 52 - 69 Cell migration in the group was significantly inhibited.

Claims

1. A cyclic peptide COX 52 - 69 Use in the preparation of medicines for the prevention and / or treatment of complications caused by diabetic vascular disease.

2. The application according to claim 1, characterized in that, The cyclic peptide COX 52 - 69 It can inhibit the proliferation and migration of high glucose-induced vascular smooth muscle cells (VSMCs).

3. The application according to claims 1 and 2, characterized in that, The cyclic peptide COX 52 - 69 It can inhibit the thickening of the vascular endothelium, poor blood flow, and high blood pressure caused by high blood sugar.

4. The use according to any one of claims 1-3, wherein the diabetic vascular lesions include atherosclerosis, intimal thickening, medial calcification, or peripheral artery disease.

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