| Objective1. To evaluate the effect of different dose of early X-ray externalradiation on the restenosis of rabbit iliac artery after percutaneoustransluminal angioplasty2. To measure the level of Cyt c、PCNA、CD34、-SMA in rabbit iliacartery with restenosis cause by percutaneous transluminal angioplastytreated by different dose of early X-ray external radiation, and to investigatethe mechanism of X-ray external radiation preventing the restenosis afterpercutaneous transluminal angioplasty.Methods1. Animals and dietTwenty four healthy adult New Zealand white rabbits, weighing2.5±0.5kg, were randomly divided into control group (four rabbits) andexperimental group (twenty rabbits). The control group was fed with normaldiet, while the experimental group was fed with hypercholesterol diet.Rabbits in the experimental group were randomly divided into five groups: the first group was no radiation group, the second group was3Gy group, thethird group was6Gy group, the fourth group was9Gy group, the fifth groupwas12Gy group, and each group had four rabbits. Rabbits in experimentalgroup underwent balloon dilating injury four weeks later, and fed withhypercholesterol diet.2. Construction of restenosis modelRabbits in experimental group fed with hypercholesterol diet for fourweeks, then underwent balloon dilating injury. After anesthesia anddisinfection, a litter cut was given at the femoral artery, and a3F ballooncatheter was inserted into the femoral artery for8cm. The iliac artery wasinjured by engorged balloons for7cm. The rabbits in3Gy group,6Gy group,9Gy group,12Gy group received3Gy,6Gy,9Gy,12Gy of X-ray externalradiation on the first and second day after the balloon dilating injury.3. The animals were sacrificed four weeks later. The iliac arteries werestained by HE staining and immunohistochemical staining. Pathologicalchanges were observed. Histomorphometric parameters were performed bycalculation of the intimal area, medial area and degree of stenosis. Theexpression levels of-SMA, cd34, PCNA and Cyt c were evaluated byimmunohistochemical analysis. The level of Cyt c was evaluated by Westernblot.Results1. Pathological change The control group: The demarcation of intima, media and extima iniliac artery were clear. The intima was smooth. The endothelial cells wereflattened and were not droped off. The media was constituted with smoothmuscle cells and ring-shaped elastic fibers. The extima was constituted withthin connective tissue. The thickness of vascular wall was homogeneous.The0Gy group: The vascular stenosis was obvious. The intima wasapparently thicker. The thickness of vascular wall was not homogeneous.The hyperplasia of smooth muscle cell was observed in intima, and thearrangement of smooth muscle cell was irregular.3Gy group–12Gy group: The vascular stenosis was slightly. Theintima was slightly thicker. A little hyperplasia of smooth muscle cell wasobserved in intima. The levels of intimal thickening and vascular stenosiswere lower than the0Gy group.2. HistomorphometryIntimal area: The intimal areas of3Gy group,6Gy group，9Gy group，12Gy group were significantly decreased compared with that of0Gy group（p<0.05）.Statistic differences werefound among3Gygroup,6Gygroup，9Gy group，12Gy group（p<0.05）.The intimal area gradually decreased asthe radiation dose increased. The intimal area of control group wassignificantly decreased compared with that of0Gy group,3Gy group,6Gygroup，9Gy group（p<0.05）.No statistic difference of was found betweencontrol group and12Gy group（p>0.05）. Medial area: The medial area of3Gy group,6Gy group，9Gy group，12Gy group were significantly decreased compared with that of0Gy group（p<0.05）. The medial area of control group was significantly decreasedcompared with that of0Gy group（p<0.05）.Degree of stenosis: The stenosis degree of6Gy group，9Gy group，12Gy group were significantly decreased compared with that of0Gy group（p<0.05）. The stenosis degree of9Gy group，12Gy group weresignificantly decreased compared with that of3Gy group（p<0.05）. Thestenosis degree of12Gy group was significantly decreased compared withthat of9Gy group（p<0.05）. The stenosis degree of control group wassignificantly decreased compared with that of0Gy group,3Gy group,6Gygroup，9Gy group（p<0.05）. The stenosis degree of control group and12Gy group had no statistic difference（p>0.05）.3. ImmunohistochemistryThe positive-SMA cells with claybank endochylema were observedin intima and media. The levels of-SMA in3Gy group,6Gy group，9Gygroup,12Gy group were significantly decreased compared with that of0Gygroup（p<0.05）. The level of-SMA in12Gy group was significantlydecreased compared with that of3Gy group,6Gy group，9Gy group（p<0.05）. The levels of-SMA in0Gy group,3Gy group,6Gy group，9Gy group were significantly increased compared with that of control group（p<0.05）. No statistic difference was found between control group and 12Gy group（p>0.05）.The positive CD34cells with claybank endochylema can be seen inendothelial cell in intima. No statistic difference was found among all groups(p>0.05).The positive Cyt c cells with claybank endochylema can be seen inintima，media and extima，especially the intima and media. The levels of Cytc in9Gy group and12Gy group were significantly increased compared withthat of0Gy group（p<0.05）. The levels of Cyt c in9Gy group and12Gygroup were significantly increased compared with that of3Gy group（p<0.05）.Thelevels ofCytc betweencontrolgroup and0Gygroup had nostatistic difference (p>0.05). The levels of Cyt c in3Gy group,6Gy group,9Gy group,12Gy group were significantly increased compared with that ofcontrol group（p<0.05）.The positive PCNA cells with claybank karyon can be seen in intimaand media. The levels of PCNA in6Gy group,9Gy group and12Gy groupwere significantly decreased compared with that of0Gy group（p<0.05）.The levels of PCNA in6Gy group,9Gy group and12Gy group weresignificantly decreased compared with that of3Gy group（p<0.05）. Thelevel of PCNA in control group was significantly decreased compared withthat of0Gy group,3Gy group,6Gy group,9Gy group,12Gy group（p<0.05）.4. Western blot: The levels of Cyt c in3Gy group,6Gy group,9Gy group and12Gy group were significantly increased compared with that of0Gy group（P<0.05）. The levels of Cyt c in6Gy group,9Gy group and12Gygroup were significantly increased compared with that of3Gy group（p<0.05）.Thelevels ofCytcbetweencontrolgroup and0Gygroup had nostatistic difference (p>0.05). The levels of Cyt c in3Gy group,6Gy group,9Gy group and12Gy group were significantly increased compared with thatof control group（p<0.05）.Conclusion1. With6-24Gy of X-ray external radiation, the intimal area, medialarea, degree of stenosis and the level of PCNA and-SMA significantlydecreased, and the level of Cyt c significantly increased.6-24Gy of X-rayexternal radiation can inhibit the hyperplasia of smooth muscle cell, andinduce the apoptosis of cells in intima and media, and prevent the restenosisafter PTA. And the prevention of24Gy of X-ray external radiation is moreevident.2. With6-24Gy of X-ray external radiation, the expression level ofCD34had no decrease.6-24Gy of X-ray external radiation didn’tsignificantly cause the decrease of endothelial cell and won’t break theendothelial integrity.3.6-24Gy of X-ray external radiation can cause the increase ofmitochondria releasing Cyt c, may induce apoptosis of cells in intima andmedia through the mitochondrial pathway to prevent restenosis after PTA. 18-24Gy dose of radiation is more evident in causing the release of Cyt c. |
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