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目的:制备不具有促红细胞生成作用的氨甲酰化促红细胞生成素(CEPO)。方法:用氰酸钾使促红细胞生成素(EPO)氨甲酰化,对CEPO的蛋白酶(endoproteinase Lys-C)酶切产物进行电泳鉴定。分别给予成年KM小鼠腹腔注射EPO、CEPO和生理盐水,每周3次,于3、7和14 d测定小鼠红细胞(RBC)、网织红细胞(Ret)数及Ret百分数,观察其动态变化。另外,给予不同种属的成年C57/B6小鼠腹腔注射增加1倍剂量的EPO、CEPO和生理盐水,隔日1次,30 d后测定各组小鼠上述的血细胞指标,以证明CEPO不具有促红细胞生成作用。结果:经SDS-PAGE电泳鉴定,EPO已成功地氨甲酰化为CEPO,并且有较高的纯度。KM小鼠和C57/B6小鼠分别接受EPO注射后,RBC数、Ret数、Ret百分数均随给药次数的增多而显著增加,而接受CEPO和生理盐水组均未见有明显的改变。结论:经氨甲酰化反应后,EPO成功地转化为CEPO,并消除了促RBC生成的作用。
Objective: To prepare carbamated erythropoietin (CEPO) that does not have erythropoietic activity. Methods: Erythropoietin (EPO) was carbamylated with potassium cyanate, and the digestion products of CEPO endoproteinase Lys-C were identified by electrophoresis. Adult KM mice were given intraperitoneal injection of EPO, CEPO and normal saline three times a week. The red blood cells (RBCs), reticulocytes (Ret) and Ret percentage were measured at 3, 7 and 14 days. The dynamic changes . In addition, adult C57 / B6 mice of different species were intraperitoneally injected with 1 time dose of EPO, CEPO and normal saline once a day. After 30 days, the above-mentioned blood cell indexes of mice in each group were determined to prove that CEPO did not promote Erythropoiesis. Results: It was identified by SDS-PAGE electrophoresis that EPO has been successfully carbamoylated to CEPO and has high purity. After receiving EPO injection, KM mice and C57 / B6 mice, respectively, the number of RBC, Ret, Ret percentage increased significantly with the increase in the number of administration, but no significant changes were seen in both CEPO and saline groups. CONCLUSION: After carbamoylation, EPO was successfully converted to CEPO and the effect of promoting RBC production was abolished.