LW-AFC对环磷酰胺处理小鼠免疫功能的影响

目的探讨LW-AFC对免疫功能低下小鼠是否有改善作用。方法昆明小鼠ip给予环磷酰胺(CTX)0.02g·kg-1,连续10d,制备免疫功能低下小鼠模型。六味地黄(LW)浓缩丸(1.4g·kg-1)和LW-AFC(0.2,0.4和0.8g·kg-1)组小鼠在每次给予CTX后ig给予相应药物,连续10d。监测小鼠体重,以及胸腺和脾脏重量变化,并计算胸腺和脾脏系数;3H]TdR掺入法检测脾淋巴细胞增殖反应,MTT法检测脾脏自然杀伤(NK)细胞杀伤活性,流式细胞术检测脾淋巴细胞中CD4+CD8-和CD4-CD8+T细胞亚群百分率,并计算CD4+/CD8+T细胞亚群比值。结果与正常对照组比较,模型组小鼠胸腺和脾重量及其系数明显降低,体重增长显著缓慢;LW和LW-AFC对小鼠体重增长缓慢、胸腺重量及其系数降低无明显改善作用;LW-AFC0.2g·kg-1对CTX导致的脾重及系数降低有明显改善(P<0.05)。模型组小鼠脾淋巴细胞自发增殖反应、刀豆蛋白A(ConA)和脂多糖(LPS)诱导的脾细胞增殖反应与正常对照组比较均明显降低,3H]TdR掺入值由正常对照组的6115±441,19432±1778和(23345±7296)cpm分别降低为2741±340,9210±1387和(3983±263)cpm;与模型组比较,LW-AFC0.8g·kg-1对脾淋巴细胞自发增殖反应有明显的改善作用;LW浓缩丸和LW-AFC对ConA诱导的脾细胞增殖反应均具有明显的促进作用,3H]TdR掺入值分别为13996±5161,27550±2356,15427±1444和(27333±1701)cpm;对LPS诱导的脾细胞增殖降低无改善作用,甚至低于CTX模型组(P<0.01)。模型组小鼠NK细胞杀伤活性由正常对照组的(39.5±0.5)%降低为(37.0±1.0)%,LW和LW-AFC明显促进小鼠NK细胞杀伤活性,杀伤率分别为(40.9±0.6)%,(39.7±0.8)%,(42.4±0.5)%和(39.8±0.9)%。与正常对照组比较,模型组小鼠脾脏CD3+,CD4+CD8-和CD4-CD8+T细胞百分率明显增加,CD4+/CD8+比值无明显变化;LW浓缩丸和LW-AFC可使CTX导致的升高的CD3+,CD4+CD8-和CD4-CD8+T细胞百分率明显降低,并可明显降低CD4+/CD8+比值。结论 LW-AFC对CTX所致小鼠免疫功能低下具有一定的改善作用。

Effect of LW-AFC on immune function in cyclophosphamide treated mice

OBJECTIVE To investigate the improvement of LW-AFC on immunodeficiency. METHODS An immunodeficiency model of Kunming mice was established after mice were ip given CTX 0.02 g·kg^-1, once a day, for 10 d. Liuwei Dihuang concentrated pills (LW) 1.4 g·kg^-1 and LW-AFC 0.2, 0.4 and 0.8 g·kg^-1 were ig given in LW and LW-AFC groups, respectively, followed by daily administration of CTX for 10 d. The body weight of mice was measured, once every 2 d. On the 11th day the mice were sacrificed, the thymus and spleen were taken and weighed, and their indexes were calculated. The splenocyte proliferation was measured by ［³H］TdR incorporation assay, the natural killer (NK) cell activity was measured by MTT assay, and CD4⁺CD8^- and CD4^-CD8⁺ T lymphocyte subsets were measured by flow cytometry. RESULTS Compared with normal control group, the weight and index of the thymus, and weight gain in CTX treated mice were significantly reduced. LW and LW-AFC had no obvious effect on these changes. The weight index of the spleen in LW-AFC 0.2 g·kg^-1 group was significantly improved. The spontaneous, Con A induced and LPS induced proliferation of splenocytes of CTX treated mice was significantly decreased compared with normal control group. The ［³H］TdR incorporation decreased from 6115±441, 19 432±1778 and (23 345±7296) cpm to 2741±340, 9210±1387 and (3983±263)cpm, respectively. LW-AFC 0.8 g·kg^-1 could significantly improve the spontaneous proliferation of splenocytes while LW and LW-AFC 0.2, 0.4 and 0.8 g·kg^-1 could significantly promote the Con A induced splenocyte proliferation. The ［³H］TdR incorporation was 13 996±5161, 27 550±2356, 15 427±1444 and (27 333±1701) cpm, respectively. LW and LW-AFC had no significant effect on the LPS induced splenocyte proliferation. The NK cell activity in CTX treated mice was decreased from (39.5±0.5)％ of normal control group to (37.0±1.0)％, but could be greatly improved by LW and LW-AFC 0.2, 0.4 and 0.8 g·kg^-1. The NK cell activity was (40.9±0.6)％, (39.7±0.8)％, (42.4±0.5)％ and (39.8±0.9)％, respectively. The percentages of CD3⁺, CD4⁺CD8^- and CD4^-CD8⁺ T cells of CTX treated mice was significantly increased, compared with normal control group, but CD4⁺/CD8⁺ ratio was not obviously changed. LW and LW-AFC could significantly decrease the percentage of CD3⁺, CD4⁺CD8^- and CD4^-CD8⁺ T cells and CD4⁺/CD8⁺ ratio. CONCLUSION LW-AFC can make remarkable improvement on immunodeficiency induced by CTX.