The Study On A New Kind Of Low Nickel And High Nitrogen Stainless Steel Coronary Stent

BackgroundCoronary heart disease,referred to as CHD,is a common cardiac disease caused by coronary artery stenosis and insufficient blood supply.It has become the leading cause of human death.At present,there are three main treatments for coronary heart disease:drug therapy,surgery and interventional therapy.The treatment principle is mainly to improve the blood supply of coronary artery,reduce myocardial oxygen consumption and prevent the development of atherosclerosis.Percutaneous coronary intervention(PCI)is widely used in the treatment of cardiovascular stenosis at present.However,due to the mechanical dilation of coronary artery during stent implantation,which results in deep injury of vascular wall,inflammation and vascular smooth muscle cell hyperplasia leading to intimal hyperplasia and thrombosis,13%to 18%restenosis still occurs 3-6 months after stent implantation.Therefore,the study of in-stent restenosis(ISR)mechanism is still one of the hotspots in the field of cardiovascular research,which has great theoretical and practical significance.A large number of studies have found that when medical metal materials are corroded or abraded in human body,wear and corrosion will cause metal ions to dissolve and affect the metabolism of human tissues.Nickel(Ni)ions released from stent are a well-known harmful element.Nickel ions can cause cytotoxicity,activate inflammatory processes and increase the production of inflammatory cytokines,including Interleukin-1B(IL-1b).Interleukin-6(IL-6)and tumor necrosis factor(TNF).On the other hand,metal stents can continuously release nickel ions,leading to excessive accumulation of vascular smooth muscle cells(SMC),which is related to restenosis.The reason of nickel ion-induced restenosis is related to vascular endothelial growth factor(VEGF).Although nickel allergy is associated with a higher risk of restenosis after stent implantation,it is necessary to add nickel to the stent to increase its flexibility.Therefore,the development of a low nickel and high nitrogen stent to reduce restenosis after stenting has become a research hotspot.We studied a 180-day long-term observation to evaluate the effect of high-nitrogen and low-nickel austenitic stainless steel(HNS)stents on endothelialization.In vitro experiments included assessing the proliferation of human umbilical vein endothelial cells(HUVEC)and the response of vascular endothelial cells,and studying the mechanism of the effect of low-nickel and high-nitrogen stainless steel materials on neointimal proliferation.The purpose of this study is to provide a scientific basis for the clinical application of high nitrogen and low nickel austenitic stainless steel stents.PurposeIn this study,low nickel and high nitrogen stainless steel metal stent(HNS)was used to evaluate the blood compatibility and cytocompatibility of the new low nickel and high nitrogen scaffold in vitro.The in vivo safety of the new stent was evaluated.Then,the occurrence of in stent restenosis after 1 months and 6 months of the new low nickel and high nitrogen coronary stent implantation was observed by histological imaging.Finally,the effects of HNS stent on the proliferation of HUVECs and the possible mechanism were observed by molecular biology and other experimental methods.Method1.Blood compatibility test:The blood compatibility of 316L stent and HNS stent were evaluated by hemolysis test,protein adsorption test and platelet adsorption test.2.Evaluation of histocompatibility:HUVEC cells were cultured on 316L stent and HNS stent.The cytocompatibility of HUVEC cells in vitro was evaluated by 2 methyl thiazole diphenyl tetrazolium bromide salt(MTT)assay.3.In vivo study:316L stainless steel stents and HNS stents were implanted into miniature pig coronary arteries.Angiography and OCT were used to detect stenosis degree at 1 and 6 months after stent implantation.Histological examination was performed on the coronary artery of stent implanted segment.Finally,Q-PCR was used to detect the expression of vascular endothelial VEGF-A and b-FGF mRNA in stent implantation.4.In vitro experiment:HUVECs were co-cultured with 316L stent and HNS stent,and the content of IL-6 released by HUVEC in culture medium was detected by ELISA on 1 and 3 days.The proliferation of HUVEC cells was observed by fluorescence microscopy on 1,3 and 5 days.The expression of p-STAT3 and erk1/2in HUVEC cells cultured for 7 days was detected by Western blot.Result1.316L group and HNS group both had less than 5%hemolysis rate,and there was no difference between the two groups.Compared with 316L stainless steel stent group,the amount of adsorbed albumin in HNS group was less,but there was no statistical difference(P>0.05);316L group and HNS group had no significant difference in the amount of adsorbed fibrin(P>0.05);compared with 316L group,the number of platelets adsorbed on the surface of the HNS group significantly decreased,and the statistical analysis showed significant difference(P<0.05).2.MTT assay showed that there was no significant difference in HUVEC cell activity between HNS group and 316L group.The results suggest that the HNS stent has similar cytotoxicity to 316L stent and good cytocompatibility.3.In vivo experimental results:(1)The angiographic results of 316L stainless steel stent and HNS stent implantation for 1 month showed that there was no significant difference in diameter stenosis percentage and LLL between the two groups,while the angiographic results at 6 months showed that HNS stent group was superior to 316L stainless steel stent in diameter stenosis percentage(35.9±10.2%vs 53.5±20.8%,P<0.05)and LLL(1.08±0.27mm vs 1.61±0.68mm,P<0.05).Group.(2)The stents in HNS group and 316L group were covered by intima at 1 month,either in general specimens or by scanning electron microscopy.(3)After 6 months of HE staining,the corresponding data of two groups of scaffolds were compared and the results were the same as those of angiography.In HNS group,the lumen area(3.46±1.49mm~2 vs.1.91±1.04mm~2,P<0.05)and the percentage of area narrowing(54.0±17.8%vs.73.6±14.5%,P<0.05)were better than 316L stainless steel stent group.(4)The inflammatory index of HNS stent group was significantly better than 316L stainless steel group in reflecting the inflammation around the stent(1.71±0.43 vs2.68±0.57,P<0.05).(5)Whether in 1month ot 6 months observation,no animal death or ST occurred in HNS group and 316L group,indicating that the safety of HNS material was at least consistent with 316L.(6)After 28 days of stent implantation,the expression of VEGF-A mRNA in neointima was significantly higher in 316L group(1.08±0.09)than that in HNS group(0.99±0.16,P>0.05),but there was no statistical difference.After 6 months of stent implantation,the expression of VEGF-A in 316L group(2.86±0.8)was significantly higher than that in HNS group(1.66±0.22,P<0.01).There was no difference in the expression of b-FGF mRNA between316L group and HNS group both after 28 days and 6 months stetns implantation.4.In vitro experiment results:after co-culture of HUVEC with 316L stent or co-culture of HUVEC with HNS stent for 1,3 and 5 days,it was observed that the number of HNS surface cells was less than 316L group;the level of IL-6 released by316L group(0.31±0.054pg/ml)was significantly higher than that of HNS group(0.23±0.02 pg/ml,P<0.01)after 1day co-culture;and the level of IL-6 released by316L group(0.35±0.087pg/ml)was also higher than that of HNS group(0.26±0.02pg/ml,P<0.05)after 3 days of HUVEC culture.Western blot results showed that the expression of p-STAT3 in HUVECs grown on HNS group after 7 days of culture was significantly lower than that of HUVECs grown on 316L group(0.2485±0.006,P<0.05);the expression of ERK1/2 in HUVECs grown on HNS(0.671±0.016)was also significantly lower than that of 316Lgroup(0.829±0.054,P<0.05).ConclusionThe HNS stent has good blood and biocompatibility.In this study,we found that arteries implanted with HNS stents exhibited fewer neointimal formation and inflammation after 180 days than those implanted with 316L stents.The reason maybe due to the low expression of STAT3 and the low release of IL-6 by HUVEC which was on the surface of HNS stents and the low expression of VEGF.Therefore,HNS stent is a promising alternative stent for coronary intervention.