| BackgroundWith the change of life-style, enhancement of social stress, increase of risk factors such as hypertension, dyslipidemia, diabetes mellitus and smoking, and aging of population, recent decades have seen that cardiovascular disease has been becoming the first killer of people in China. Regardless of a great progress of medicinal and intervational therapies, peripheral and coronary artery bypass grafing (CABG) is still in an indispensible position of treating complex and multiple vascular lesions because of demand of complete revascularization. However, compared with the arterial conduit, especially, internal mammary artery graft, saphenous graft (SVG) is more prone to neointimal hyperplasia and atherosclerosis, subsequently, easy to restenosis. Thus, a lower long-term patency of SVG than that of the arterial graft greatly limits its therapeutic property in arterial bypass grafting.Growing evidence from clinical and experimental studies indicates that endothelial cell (EC) injury resulting in endothelial dysfuction due to ischemia/repefusion (I/R) induced injury, increased stretch stress under arterialization initiates the thrombosis and neointimal hyperplasia in autologous vein grafts. Additionally, the death of vascular smooth muscle cells (VSMCs) can provoke migration of another VSMCs into tunica intima and infiltration of inflammatory cells into the vein graft wall, and cytokines and growth factors released from the inflammatory cells, in turn, inducing the migration and proliferation of VSMCs, resultly, enhancing neointimal hyperplasia.Considerable studies have demonstrated that glucose-insulin-potassium (GIK) /insulin remarkably improves the prognosis of pateints undergoing CABG; furthermore, a recent clinical trial indicated that insulin (GIK) infusion significantly reduces reccurent ischemic episodes in post-CABG patients with diabetes. Our previous studies have demonstrated that insulin, as the key compound of GIK, plays a pivotal role in inhibiting the apoptosis of both I/R cardiac myocytes and coronary arterial endothelial cells via PI3K/Akt-eNOS -NO cell survival signaling pathway. Importantly, recent studies indicated that insulin markedly inhibits neointimal hyperplasia induced by balloon injury in the rat carotid artery.Nevertheless, it is still to be evaluated whether GIK/insulin plays a protective role in vein grafts and inhibits neointimal hyperplasia in autologous vein grafts.AimsThe purposes of this study were (1) to verify whether apoptotic phenomenon of ECs and VSMCs exists in vein grafts at the earlier stage of postoperation; (2) to assess whether the administration of GIK/ insulin plays a protective role in I/R vein grafts; and if so, (3) to elucidate the invovled mechanisms; (4) to investigate influence of a short-term continuous insulin administration on neointimal hyperplasia in autologous vein grafts.Materials and methodsThirty-two mongrel dogs were subjected to external jugular-carotid interposition bypass grafting, and underwent infusions of vehicle (saline), GIK (glucose, 250 g/L; insulin, 60 U/L; potassium, 80 mmol/L), GK or GIK plus wortmannin (i.v., 2.0?g/kg/min for 1 hour immediately after reperfusion) 5 minutes before and 4 hours after reperfusion, respectively (n = 8, in each group). Other 8 sham I/R dogs underwent the same operation except the excision of the external jugular vein. All dogs were euthanized at 4 hours postoperationally to harvest vein grafts. Active caspase-3 immunohistochemistry and TUNEL stainings were used to detect apoptosis of ECs and VSMCs in vein grafts, and Western blotting was used to determine Akt/phosphorylated Akt (pAkt), eNOS/phosphorylated eNOS (peNOS) and active caspase-3 levels in vein grafts. In a separate study, 16 dogs undergoing the same operation were randomly administered with subcutaneous injection of 0.5-mL saline or 4-U insulin two times per day for 30 days postoperatively(n = 8, in each group).At 1 month postoperationally, dogs were sacrificed for sampling vein grafts. BrdU incorporation immunohistochemistry and Verhoeff-Van Gieson stainings were used for cell proliferation and intimal/medial thickness assays. Western blotting of alpha-smooth muscle actin was used to assess the phenotype of VSMCs, and immunohistochemistry staining of eNOS was utilized to evaluate endothelial function of vein grafts.Results 1. Much apoptosis of ECs and VSMCs existed in canine vein grafts at the early stage after reperfusion in vivo;2. Treatment with GK aggravated apoptosis of both ECs and VSMCs (P < 0.05 vs vehicle ); however, treatment with GIK significantly reduced the apoptosis of both ECs and VSMCs, evidenced by decrease of apoptotic index (P < 0.001 vs vehicle and GK) and inhibition of caspase-3 activation in canine vein grafts (P < 0.01 vs vehicle and GK);3. Treatment with GK attenuated the phosphorylation of Akt and eNOS in canine vein grafts (P < 0.05, respectively vs vehicle); meanwhile, treatment with GIK enhanced the phosphorylation of Akt and eNOS nearly by 2 and 2.5 times, respectively (P < 0.001, respectively vs vehicle). However, wortmannin largely abolished the GIK-elicited effects;4. At 1 month postoperatively, administration of insulin greatly increased alpha-smooth muscle actin expression in canine vein grafts (P < 0.001 vs normal control and vehicle groups, respectively );5. Administration of insulin inhibited the cell proliferation in canine vein grafts (11.15%±0.93% vs 25.55%±2.02%, P < 0.001 vs vehicle);6. Administration of insulin increased the eNOS expression in intima of canine vein grafts (P < 0.05, respectively vs vehicle);7. Administration of insulin inhibited significantly neointimal hyperplasia of canine vein grafts (intimal/medial thickness 136.51±8.97?m vs 185.92±8.34?m, P < 0.05 vs vehicle).Conclusions1. Hyperglycemia aggravates the injury to ischemic/reperfused canine autologous vein grafts through provoking the apoptosis of ECs and VSMCs.2. Insulin exerts its anti-apoptotic effect on ECs and VSMCs via PI3K/Akt pathway in canine autologous vein grafts.3. Administration of insulin at the early stage of postoperation plays an active role in preventing vein grafts from early degeneration.4. Postoperational insulin administration exerts an preventive effect on neointimal hyperplasia in autologous vein grafts probably through improvement of endothelial function and inhibition of the proliferation of VSMCs. |
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