冻干同种异体骨粉与人淋巴细胞共培养时对细胞的影响

背景：同种异体骨材料一直存在着免疫原性方面的问题，目前关于其免疫原性的研究大多以临床疗效为依据，因此在实验室层面上对其免疫原性方面的研究及相关方法学的探讨有一定的意义。 目的：对冻干同种异体骨进行免疫原性分析。 设计、时间及地点：对比观察实验，于2008-05/12在南方医科大学附属南方医院临床医学实验研究中心完成。 材料：羟基磷灰石粉(200目)由北京京航生物有限公司提供；同种异体冻干骨粉(200目)由山西奥瑞生物材料有限公司提供。淋巴细胞来源10名健康青年志愿者，年龄23~26岁。 方法：实验分为阳性对照组（植物血球凝集素），阴性对照组(羟基磷灰石)及实验组(冻干骨粉2，1，0.5 g/L)，采用淋巴细胞转化实验Alamarblue法进行细胞与材料共培养，72 h后酶标仪570 nm，600 nm双波长下读取吸光度值计算各组淋巴细胞转化率。 主要观察指标：酶标仪检测淋巴细胞转化率。 结果：人冻干骨粉各组与羟基磷灰石组比较，差异无显著性意义(P > 0.05)；与植物血球凝集素组比较，差异有显著性意义(P < 0.001)；骨粉各组之间两两比较，差异无显著性意义(P > 0.05)。 结论：在本实验中冻干同种异体骨未出现对淋巴细胞刺激的免疫原性作用，不能对淋巴细胞的增殖产生影响。骨粉在0.5~ 2.0 g/L剂量内不存在量效关系。

Effect of freeze-dried bone allograft co-cultured with human lymphocytes on cells

BACKGROUND: Allogeneic bone implants materials have a long history in clinical application, but there has been a relevant immunogenicity related issues. Therefore, to explore the immunogenicity research and related methodologies has certain significance. OBJECTIVE: To analyzed immunogenicity of freeze-dried bone allograft. DESIGN, TIME AND SETTING: The contrast observation experiment was completed in the Research Center of Nanfang Hospital, Southern Medical University between May and December 2008. MATERIALS: Hydroxyapatite powder (200 meshes) was offered by Beijing Jing Air Biological Co., Ltd; Freeze-dried bone allograft (200 meshes) was supplied by Shan Xi Aorui Biological Material Co., Ltd. The lymphocytes were originated in 10 healthy young volunteers, aged 23-26 years old. METHODS: The experiment was randomly divided into positive control (phytohemmagglutinin), negative control (hydroxyapatite) and experimental (Freeze-dried bone meal 2, 1, 0.5 g/L) groups. Lymphocyte were separated and co-cultured with freeze-dried bone by lymphocyte transformation test, A value and lymphocyte transformation rate was calculated at 72 hours later with 570 nm, 600 nm dual-wavelength absorption Reader. MAIN OUTCOME MEASURES: The rate of lymphocyte transformation was detected by Labsystems. RESULTS: There was no significant difference between the experimental group and negative control group (P > 0.05); compared with the positive control group, the difference had statistically significant (P < 0.001); there was no obvious difference between each experimental group (P > 0.05). CONCLUSION: Freeze-dried bone allograft does not appear immunogenicity to lymphocyte stimulating, which can not affect the lymphocytes proliferation. There is no dose-effect relationship when the dosage of bone allograft is between 0.5 mg and 2 mg.