Studies on the cellular modulation in cryopreservation focusing the allogenicity of thoracic tissues

背景：冷冻保存人类同种异体心脏瓣膜和血管已广泛应用于心胸外科。收获的组织通常被冷冻在一个程序化的冰箱里，这样可以缓慢冷冻，避免组织被冰点附近的外来热破坏。冷冻保存的气管比新鲜的气管更容易植入。冷冻保存过程似乎降低了同种异体免疫原性。本研究主要关注低温保存对气管组织中细胞异体性的影响，即气道上皮细胞、血管内皮细胞和成纤维细胞。方法：采用比色MTT法测定细胞活力。同种异体活性通过流式细胞术测定干扰素- γ (IFN)异体分子表达增加的程度来评估。单向混合淋巴细胞刺激细胞培养（MLC）。结果：各细胞系均能很好地保存程序冷冻细胞的活力。IFN可诱导细胞表面表达hla - II类，除气道上皮细胞外，经低温保存后其表达无明显恶化。低温保存的气道细胞系对IFN表达hlaⅱ类抗原的敏感性较低。成纤维细胞和淋巴细胞的MLC在成纤维细胞低温保存和解冻后均无明显下降。结论：低温保存对人气管组织细胞的异体免疫原性有不同程度的调节作用。低温保存是否有利于同种异体移植物的保存，目前还存在争议。细胞活力的保存对于同种异体移植物的耐久性至关重要，然而，它也与移植物的同种异体性有关。进一步研究抗原提呈细胞，如同种异体移植物中的树突状细胞，将揭示在冷冻保存组织中对同种异体的调节。

Background : Cryopreserved human allogenic heart valves and vessels have come to be widely utilized in cardiothoracic surgey. The harvested tissues are usually frozen in a programmed freezer, which enables slow freezing and avoids tissue destruction by the exotic fever around the freezing point. Cryopreserved tracheas have been found to be more easily implanted than fresh tracheas. The cryopreservation procedure seems to decrease alloimmunogenicity. In this study focused the influence of cryopreservation on the allogenicity of cells included in the tracheal tissue, i.e. airway epithelial cells, vascular endothelial cells and, fibroblasts.Methods : Viability of the cells was determined by colorimetric MTT assay. Allogenic activity was assessed by flow-cytometric assay determining the degree of increase in expression of allogeneic molecules with interferon-gamma (IFN). One-way mixed lymphocyte-stimulator cell culture (MLC) was also made up.Results : Viability of program-frozen cells was well preserved in each cell line. Expression of HLA-class II on cell surface was evoked by IFN, which ms not deteriorated by cryopreservation except for the airway epithelial cell. The cryopreserved airway cell line was less sensitive to IFN in expressing HLA-class II antigen. MLC of fibroblast and lymphocyte showed no decreased reaction even after the fibroblast was cryopreserved and thawed.Conclusions : Process of cryopreservation variously modulated the allogeneic immunogenicity of the cells included in the human tracheal tissue. It is controversial whether the cryopreservation is favorable for duration of the allograft. Preservation of the cell viability is essential for durability of the allograft, however, it is also associated with the allogenicity of the graft. Further studies concerning the antigen presenting cells, such as the dendritic cells included in the allograft, would reveal the modulation of allogenicity in cryopreserved tissues.