Bambang L S, Mazzucotelli J P, Moczar M, Beaujean F, Loisance D
Centre de Recherches Chirurgicales Henri Mondor, CNRS URA 1431, Créteil, France.
J Thorac Cardiovasc Surg. 1995 Oct;110(4 Pt 1):998-1004. doi: 10.1016/s0022-5223(05)80167-9.
Human saphenous veins were cryopreserved in 4% human albumin and 10% dimethyl sulfoxide. The effect of cryopreservation on endothelial cells was studied in terms of the anticoagulant activity of thrombomodulin and in terms of cell proliferation. After storage for 2 weeks at -150 degrees C, 0.45 +/- 0.07 x 10(5) endothelial cells/cm2 were detected in cryopreserved veins and 1.03 +/- 0.04 x 10(5) endothelial cells/cm2 in fresh veins (p < 0.01). The thrombin-catalyzed activation of protein C decreased after cryopreservation, indicating altered thrombomodulin activity in the endothelial cells. On a cell number basis, the release of soluble thrombomodulin was three times higher from the cryopreserved endothelium than from the fresh endothelium (p < 0.05). The amount of spontaneous release of von Willebrand factor from the endothelial surface was not significantly different between fresh and cryopreserved veins. Endothelial cells were cultured from fresh veins and from their cryopreserved counterparts. On plating of endothelial cells in primary culture, the number of adhered cells was 0.9 +/- 0.09 x 10(3) cells/cm2 from fresh veins and 0.25 +/- 0.03 x 10(3) cells/cm2 from cryopreserved veins (p < 0.01). The positive immunohistochemical stain for von Willebrand factor indicated that the endothelial cell character was maintained after cryopreservation. The endothelial desquamation with loss of anticoagulant function and the slow proliferation of surviving cells in vitro suggest an impaired endothelial healing in vivo. The loss of anticoagulant activity complicates the problems of the exposure of thrombogenic subendothelial matrix to blood in implanted cryopreserved veins.
人隐静脉在4%人白蛋白和10%二甲基亚砜中进行冷冻保存。从血栓调节蛋白的抗凝活性和细胞增殖方面研究了冷冻保存对内皮细胞的影响。在-150℃储存2周后，冷冻保存静脉中检测到0.45±0.07×10⁵个内皮细胞/cm²，新鲜静脉中为1.03±0.04×10⁵个内皮细胞/cm²（p<0.01）。冷冻保存后凝血酶催化的蛋白C活化降低，表明内皮细胞中血栓调节蛋白活性改变。以细胞数量为基础，冷冻保存的内皮细胞释放的可溶性血栓调节蛋白比新鲜内皮细胞高3倍（p<0.05）。新鲜静脉和冷冻保存静脉之间内皮表面血管性血友病因子的自发释放量无显著差异。从新鲜静脉及其冷冻保存的对应物中培养内皮细胞。在原代培养中接种内皮细胞时，新鲜静脉中贴壁细胞数量为0.9±0.09×10³个细胞/cm²，冷冻保存静脉中为0.25±0.03×10³个细胞/cm²（p<0.01）。血管性血友病因子的阳性免疫组化染色表明冷冻保存后内皮细胞特征得以维持。抗凝功能丧失导致内皮剥脱以及体外存活细胞增殖缓慢提示体内内皮修复受损。抗凝活性丧失使植入的冷冻保存静脉中促血栓形成的内皮下基质暴露于血液的问题更加复杂。
