An Experimental Study On Rapamycin In Preventing Artery Restenosis After Stent Implantation

BackgroundNow, percutaneous coronary intervention (PCI) has become a common therapy against CVD day by day, and drug-eluting stent (DES) used in PCI is proved by a large number of clinical tests that it can obviously decrease the restenosis rate. Rapamycin-eluting stent is one of them which can reduce the restenosis rate of inner stent vessel and stent located vessel. Recent studies show that as dominant-negative inhibitor of mTOR, rapamycin (RAPA) can promote autophagy of tumor cells to restrain tumor happen and grow. People pay close attention to the effect of RAPA in AS therapy again, because mTOR goes core position in PI3K/AKT/mTOR/p70sK signal pathway. A devil of clinical trials show RAPA has this kind of effect that it can well restrain neointimal ultra-hyperplasia, so RAPA-eluting stent can topical drug release to prevent stent restenosis. Researches show RAPA has unique pharmacologic actions mechanism that it can restrain karyokinesis revulsive P27kip1 expression. P27kip1 is the inhibitor of cyclin dependence kinase, Diez-Juan, etc study found P27kipl removed cyclin dependent kinase inhibitors can promote the grew of AS. Researches indicate RAPA restrain monocyte/macrophage related cytokinec product, especially MPC-1 and IL-6. Suzuki, etc study show that RAPA can decrease initiative MCP-1 mRNA level. Basso, etc researches found using RAPA for 12 weeks blood CH content had no change but at the part of aortic arch it reduced 36% compared with control group. Recently studies found that RAPA can work against hyperplasia and decrease inflammatory reaction is related to selectively restraining to PI3K/Akt/mTOR signaling pathway. Chen, etc researches found oral rapamycin can obviously reduce plaque burden of atherosclerosis rabbits, and it can decrease the macrophage soakage, reduce inflammation, and increase smooth muscles in fibrous cap to stable the plaque by restraining mTOR, but this had no influence to lipid levels.Recent years, intravascular ultrasound imaging technology (IVUS) is more and more valuable in PCI as a golden standards in CHD diagnosis, because it can accurately display the plaque and vessel wall situation, observe the stent stick the vessel wall or not and restenosis situation.Given the recognized adverse effects of RAPA, we use IVUS to evaluate the effect of oral rapamycin with BMS in reducing restenosis, hope to prove this therapy to be a more excellent PCI treatment. We theoretically conjecture:this treatment can decrease stent restenosis to similarity level as DES, at the same time it can reduce stent thrombosis incidence, besides above roles it can reduce anticoagulant drugs dosing time to achieve purpose that reduce intracerebral hemorrhage, gastrointestinal hemorrhage risk. At present, both home and abroad this report is found rarely.Objective1. Compare the effect of oral rapamycin with metal stent and rapamycin eluting stent in reducing restenosis.2. To study if oral rapamycin with BMS is a more valuable therapy in coronary intervention.MethodsAll 40 male purebred New Zealand big white rabbits after one week acclimatization period, every of them was exsanguinated 4ml blood specimens from ear margin veins to exam TC, LDL-L, HDL-L etc lipid changes. Balloons injure all rabbits' abdominal aorta:3% sodium pentobarbital 30mg/kg anesthesia all animals, puncture right femoral artery, send 3.5mm diameter and 15mm length balloon catheter into descending aorta about 20cm. Fill balloon to 14 ATM and pull back and forth 3 times to injure abdominal aorta endomentrial, then ligature femoral artery at the far end, suture wound. Give each rabbit intramuscular injection antibiotics to prevent infarction. Each animal was given a diet of 1% cholesterol 120-140g per day for 8 weeks.8 weeks end, each animal was exsanguinated 4ml blood specimens from ear margin veins to exam blood lipid indexes. Then all the rabbits were randomly divided into five groups (n=8 equally):control group, oral rapamycin group (RAPA group), bare metal stent group(BMS group), bare metal stent with oral rapamycin group(BMS+RAPA group) and rapamycin eluting stent group (DES group).3% sodium pentobarbital 30mg/kg anesthesia all animals, make right femoral artery exposure, do abdominal artery imaging, observe plaque situation this part. And with the help of guide wire and expansion pipe we give each rabbit heparin (100u/kg) to anticoagulant, after that we insert IVUS detector along the wire guided in imaging guidance to narrow distal part vessel. Then pull back IVUS detector to measure each index of prepare stent implantation site and reference vessel, such as the minimal luminal diameter (MLD), external elastic membrane (EEM) area, lumen area (LA), plaque area (PA), plaque burden (PB), max diameter (Dmax) and min diameter of plaque. According to formula calculate eccentricity index (EI) and reconstruction index (RI):EI= Dmax/Dmin, RI=EEMA (pathologic part)/EEMA (average proximal and distal reference vessel). All stent group animals were lay 3.0*12mm stent at below renal artery lmm,16-20 ATM released. Then use IVUS to measure indexes, save the imaging video, and ligature femoral artery at the far end, suture wound. Give each rabbit intramuscular injection antibiotics to prevent infarction two days, and 2500u/d heparin one week. Continue to give a diet of 1% cholesterol to each animal for 4 weeks. The same time RAPA group and BMS+RAPA group rabbits were given 0.5mg/kg/d RAPA oral for 4weeks.4 weeks end, after exsanguinating each animal 4ml venous blood to exam lipid levels, IVUS measured those indexes and calculated vessel lumen lost of stent groups. Then put whole animals to death, keep whole stent section vessel as well as proximal and distal it. First we get 1mm plaque part vessel to search macrophagus by electron microscope, left sample transected into two same parts. One part was doused in formaldehyde for HE drum dyeing etc and the other part was -80℃cryopreservation for molecular biology study. Group data were analyzed with the use of SPSS13.0 (SPSS Inc., Chicago, IL, USA) for windows. All values are expressed as mean±SD. Data were analyzed using an ANOVA procedure. A value of P<0.05 was considered statistical analysis.Results1. Compared with control group, RAPA group, BMS+RAPA group and DES group have no significant difference regarding the serum lipid levels (P >0.05).2. PA and PB of RAPA, BMS+RAPA and DES groups are much less than control group and BMS group (P<0.01)3. MLD of BMS+RAPA group and DES group are obviously more than BMS group (P<0.05)4. BMS+RAPA group and DES group have less lumen reduction than BMS group (P<0.05).5. There is no significant difference between BMS+RAPA and DES groups of IVUS indexes (P>0.05).6. EEMA and LA reduction of oral RAPA group are all less than control group.7. Electron microscope scanning:we can find macrophagocytes aggregation in plaque of RAPA, BMS+RAPA and DES groups is much less than control group.ConclusionsIn this study, we got stent placement animal models successfully. We studied IVUS imaging characteristics of plaques section vessel, found that EEM and lumen reduction in oral RAPA group are all less than control group. We also found oral rapamycin matching up with BMS had similar effect with DES in decreasing lumen reduction and plaque burden. And compared with BMS group, RAPA+BMS is indeed more effective in reducing plaque area and lumen reduction. Therefore, we conclude that oral administration of rapamycin can effectively make plaque growing slow, and RAPA+BMS has the same effects on reduction of plaque burden and stent restenosis with the rapamycin eluting stent.