Study On Intima Endothelialization Of Vascular Prothesis Seeded By CD34~+ Stem Cells

Objective:To investigate endothelialization and medium-long term patency rate of ePTFE (expanded polytera fluoro ethylene) and Dacron artificail blood vessel planted by canine bone marrow CD34+ stem cells. And study the relationship between the 6-Keto-PGF1a, TXB2 and the endotheliosis and thrombosis of the new endomembrance on the prostheses seeded by CD34+ stem cells.Methods:CD34+ stem cells were isolated from canine bone marrow and purified by immunomagnetic beads system, which were seeded on the surface of ePTFE and Dacron blood vessel prostheses. Dogs were separate to ePTFE and Dacron experimental group and control groups. Implant the same sort of artificial blood vessel into the abdominal aorta artery(AAA) and inferior vena cava(IVC) of the same dog. Blood was collected in preoperative and 3,7,14,30,60 days postoperative from femoral vein. PGFla and TXB2 concentrations were detected by EIA assay. Obtaining the specimens of the experimental groups and the control groups to compare the patency rate in the 10,30,60,100 days after transplantation. Compare the patency rate of the experimental group and control group. Blade the specimens, identify the endothelial cell by immunohistochemistry method. Observe the characteristic of tunica intima yasorum and intima endothelial of related time section by optical and scanning electronic microscope, observe the new born intima thickness, and compare the endothelialization of the artery and venous system of those implanted artificial blood vessel described above. Results:(1) The isolated cells from canine bone marrow were identified to be CD34+ stem cells by flow cytometry. Those CD34+ cells were isolated and purified by immunomagnetic beads system, the concentration of cells were measured by trypan blue dyeing method, is 2.6(±0.3)×10'/ml.(2) The PLT concentration in the experimental group increased at first, then decreased and maintained at a certain level, while the control group, after a marked increase, remained at a high level;The serum PGF1αin experimental group reduced firstly and then increased while the control group decreased significantly and then maintained at a lower level; TXB2 concentration in the experimental group increased at first, then decreased and maintained at a certain level, while the control group, after a marked increase, remained at a high level; P/T ratio in experimental group reduced firstly and then increased while the control group decreased significantly and then maintained at a lower level.(3) For the experimental group:in the artery case, the occlusion rate is O, for ePTFE group the stenosis rate is 25%, for the Dacron group the stenosis rate is 25% too. In the vein case, for ePTFE group, the occlusion rate is 12.5%, the stenosis rate is 37.5%, and occlusion rate and stenosis rate is 0 and 37.5% for the Dacron group. For the control group, the stenosis rate is 0 and 100% for the artery and vein implanted blood vessels.(4)Observe by the optical microscope, the new born inner membrane is dense and complete for the HE staining Dacron blood vessel, while the intima of the ePTFE staining vessel is partly separately. The new intima thickness of those two experimental group is smaller than the control group, while there are some difference between those two experimental group.(5)Observe by SEM:for the 30 days specimens of the experimental group, the endothelial cell density decrease gradually from the stoma to the middle of the artificial blood vessel; for the 100 days specimens, the endothelial cells ranked regularly and completely. For the Dacron artificial blood vessel, the endothelial cells form a complete inner membrane which follow the long axis of the original artificial blood vessel. For the ePTFE blood vessel, the new intima is less complete, there are some area lack of endothelial cell, while some blood cell and cellulose attach instead. There are some necrotic and senile endothelial cell between. For the control group, the situation is simple, there are only some cellulose, no endothelial cell is found to cover the inner wall.Conclusion:(1)The CD34+ stem cells isolated by immuno-magnetic beads system can be purified, and can have an appropriate increment capability, the quantity of the cells is enough to implant in one artificial blood vessel with length 20-30cm and diameter 6-8mm.(2) Purified CD34+ cells were implanted into ePTFE and Dacron artificial blood vessels, the intima reach expected endothelialization and patency rate. Such a method is better for Dacron than ePTFE artificial blood vessel.(3) The endothelialization and patency rate of the implanted artery blood vessels is better than those implanted venous blood vessels.(4) It can inhibit the adhesion, aggregation and activation of platelet, also inhibit the production of bioactivators through displacing blood vessel with prostheses seeded with CD34+ stem cell.(5)It makes endothelial cell grow faster, which can produce prostacyclin, adenosine, nitric oxide, VEGF and other bioactivators through displacing blood vessel with prostheses seeded with CD34+ stem cell. And then they inhibit PGF1αand P/ T ratio of over-reduced effectively, restrain TXB2 and platelet excessive rise, finnally prevent intimal hyperplasia and thrombosis...