Saito Aya, Motomura Noboru, Kakimi Kazuhiro, Ono Minoru, Takai Daiya, Sumida Sajio, Takamoto Shinichi
Department of Cardiothoracic Surgery, Graduate School of Medicine, The University of Tokyo, Japan.
Cryobiology. 2006 Apr;52(2):251-60. doi: 10.1016/j.cryobiol.2005.12.002. Epub 2006 Jan 25.
Cryopreservation is a valuable technique for storing heart valve and vascular allografts. However, the biological ramifications of cryopreservation are still unclear; therefore, using animal experiments we assessed how 'cryopreservation' influences graft allogenicity and cell viability.
Thoracic aortas of Lewis rats were prepared as fresh (F) or cryopreserved (CP) grafts, and implanted into the infrarenal aorta of Lewis or Brown Norway rats (BNs). The grafts and spleens were harvested at post-operative day 7 and 28 (POD7, POD28) for analyses.
First, the systemic immune response to transplantation was estimated by mixed lymphocyte reaction analyses using spleen cells from naïve or recipient BNs. The alloreactivity of the recipients increased to 1.5 times that of the naïve BNs at POD7 and POD28, when stimulated by mitomycin C-treated Lewis spleen cells. Second, local immune response was estimated by TNFalpha, IFNgamma, and iNOS mRNA expression in the grafts by quantitative PCR, which revealed 20- to 40-fold increases at POD28 after allotransplantation. Third, endothelial cell viability was estimated by endothelial NOS mRNA expression level: it was similar and highest in F and CP grafts before transplantation then significantly decreased after both syngeneic and allogeneic transplantation. Finally, intimal hyperplasia, expressed by I/M ratio, developed over time after allotransplantation, reaching 2.5 times the thickness of F grafts before transplantation. The results of these experiments revealed no difference between F and CP grafts before and after transplantation.
Cryopreservation did not modify the allogenicity of vascular allografts and had minimal adverse impacts on graft cell viability.
冷冻保存是一种用于储存心脏瓣膜和血管同种异体移植物的重要技术。然而，冷冻保存的生物学影响仍不明确；因此，我们通过动物实验评估了“冷冻保存”如何影响移植物的同种异体性和细胞活力。
将Lewis大鼠的胸主动脉制备成新鲜（F）或冷冻保存（CP）移植物，并植入Lewis或棕色挪威大鼠（BNs）的肾下腹主动脉。在术后第7天和第28天（POD7、POD28）采集移植物和脾脏进行分析。
首先，通过使用来自未免疫或受体BNs的脾细胞进行混合淋巴细胞反应分析来估计对移植的全身免疫反应。当用丝裂霉素C处理的Lewis脾细胞刺激时，受体的同种异体反应性在POD7和POD28时增加到未免疫BNs的1.5倍。其次，通过定量PCR评估移植物中TNFα、IFNγ和iNOS mRNA的表达来估计局部免疫反应，结果显示同种异体移植后POD28时增加了20至40倍。第三，通过内皮型一氧化氮合酶mRNA表达水平评估内皮细胞活力：在移植前F和CP移植物中相似且最高，然后在同基因和异基因移植后均显著降低。最后，以I/M比值表示的内膜增生在同种异体移植后随时间发展，达到移植前F移植物厚度的2.5倍。这些实验结果显示F和CP移植物在移植前后没有差异。
冷冻保存并未改变血管同种异体移植物的同种异体性，并且对移植物细胞活力的不利影响最小。
