| Objective: To establish a rabbit model of autologous vein grafting and investigate the apoptosis level in the remodeling process of vein grafting and the time characteristics of proliferating cell nuclear antigen (PCNA) expression by using various research methods including Morphometric analyses,Western Blot, immunohistochemistry, TUNEL method and Transmission electron microscopy.Methods:42healthy purebred New Zealand big-eared white rabbits were used to establish the rabbit model of common carotid artery bypass grafting via the external jugular vein. They were divided into seven groups and their vein grafts were respectively removed before surgery, at1d,3d,7d,14d,28d and90d postoperatively for Morphometric analyses,Western Blot, immunohistochemistry,apoptosis-related detection and Transmission electron microscopy. The time characteristics of apoptosis level and PCNA expression were explored.Results:1. Morphometric analyses showed that the thickness of the intima was (4.66μm±0.6μm) in the normal vein group it began to slightly decline at1d postoperatively (1.47μm±0.3μm) and was significantly different from the normal vein group (P<0.05) it began to increase in the postoperative3d group (3.41μm±0.2μm) and was significantly different from the normal vein group (P<0.05) at7d postoperatively (18.1μm±2.5μm)14d postoperatively (35.7μm±5.1μm)28d postoperatively (65.5μm±5.0μm)90d postoperatively (93.5μm±6.4μm),higher than normal vein group, and the difference was statistically significant (P<0.5). The thickness of the media+adventitia was (6.4μm±0.9μm) in the normal vein group began to increase in the postoperative1d group (8.5μm±1.1μm) at3d postoperatively (15.6μm±1.9μm) at7d postoperatively (45.6μm±5.0μm) at14d postoperatively (62.3μm±2.7μm) at28d postoperatively (123.9μm±5.6μm) at90d postoperatively (130.6μm±10.0μm),higher than normal vein group, and the difference was statistically significant (P<0.5).2. Western Blot showed that the percentage of MOD of PCNA target band to internal reference band was (40.6%±2.4%) in the normal vein group the postoperative1d group (36.4%±5.1%) was lower than the normal vein group, and the difference was statistically significant (P<0.05) it was then raised, so that the postoperative3d group (43.6%±4.4%) was higher than the normal vein group, and the difference was statistically significant (P<0.05) it reached the peak in the postoperative7d group (77.9%±7.1%), higher than the normal vein group and the postoperative1d group, and the difference was statistically significant (P<0.05) it was then slowly declined.3. Immunohistochemistry showed that the percentage of PCNA-positive cells was (6.07%±1.1%) in the normal vein group it began to slightly decline at1d postoperatively (5.05%±0.9%) and was significantly different from the normal vein group (P<0.05) it began to increase in the postoperative3d group (11.26%±1.6%) and was significantly different from the normal vein group (P<0.05) it reached the peak at7d postoperatively (31.2%±3.4%), higher than normal vein group and the postoperative1d group, and the difference was statistically significant (P<0.05) it was then slowly declined.4. TUNEL method showed that the apoptotic cells were rarely expressed in the normal vein group (1.49%±0.2%), which reached the peak (18.02%±1.9%) at1d postoperatively and was significantly different from the normal vein group (P<0.05) it began to decline in the postoperative3d group (7.29%±0.7%) and significantly different from the normal vein group (P<0.05) it was declined (2.09%±0.5%) at7d postoperatively and significantly different from the normal vein group (P<0.05) it then continued to maintain at a low level.5. The electron microscope showed that after the first day the nucleolus disappeared when group by electron microscopy, chromatin edge set Postoperative day3set of visible synthetic smooth muscle cells, contain a large number of golgi apparatus and rough endoplasmic reticulum. The iconic apoptotic body and late apoptosis.7days is a lot of contractive phenotype of smooth muscle cells, and the surrounding of elastic fibers. And a large number of collagen fibers. After14days,28days still a small amount of synthetic phenotype of smooth muscle cells and a lot of contractive phenotype of smooth muscle cells, as well as a large number of collagen fibers.Conclusion:1. The proliferation coexists with apoptosis after vein grafting, suggesting that proliferation and apoptosis are involved in the remodeling process of vein grafting.2. The apoptosis level is higher in the early stage after vein grafting (1d to3d postoperatively) the proliferation level is relatively higher after surgery for7d after vein grafting. |
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