积雪草提取物对脾淋巴细胞细胞周期及细胞内游离钙的影响

目的: 观察积雪草提取物对小鼠脾淋巴细胞细胞周期及细胞内游离Ca²⁺的影响,并对其免疫调节作用机制进行初步探讨。方法: 采用乙醇浸提、极性萃取分离得到积雪草粗提物,利用Sephadex-LH20凝胶柱层析进一步分离纯化获得积雪草各组份提取物,通过观察各组份抑制1,1-二苯基-2-苦基苯肼(DPPH)自由基和羟自由基的抗氧化作用及对脾淋巴细胞增殖的影响,筛选出其活性成份。采用流式细胞术观察此活性成份对脾淋巴细胞细胞周期影响,并用Fluo-3/AM荧光探针结合流式细胞术分析积雪草提取物对脾淋巴细胞内游离Ca²⁺影响。结果: (1)获得进一步纯化并具有抗氧化活性的积雪草提取物D;(2)积雪草提取物D对脾淋巴细胞增殖抑制作用显著;(3)积雪草提取物D可降低脾淋巴细胞内游离Ca²⁺浓度,且呈量效关系;(4)积雪草提取物D能阻滞脾淋巴细胞进入S期和G₂/M期。结论: 积雪草提取物抑制小鼠脾淋巴细胞增殖,其作用机制可能是通过降低细胞内游离Ca²⁺浓度从而抑制脾淋巴细胞进入细胞分裂期而发挥作用。

Effects of Centella asiatica extracts on cell cycle and intracellular free calcium in spleen lymphocytes

AIM: To investigate the effects of Centella asiatica extracts on the cell cycle and intracellular free calcium ( _i) in spleen lymphocytes. METHODS: Centella asiatica was extracted with alcohol and different organic solvents. Sephadex-LH20 gel column chromatography was applied for further purification. To screen the active constituents, the scavenging ability of the fractions generated by elution of chromatography against radicals of OH·, DPPH· and their anti-proliferative effects on spleen lymphocytes were measured. To assess the cell cycle and _i in spleen lymphocytes, flow cytometry was performed. RESULTS: The D fraction eluted from the crude extract of Centella asiatica was obtained, which had strong suppressive effect on lymphocyte proliferation and decreased the _i of the cells in a dose-dependent manner. The cell cycle of the spleen lymphocytes arrested at S and G₂/M stages when treated with the D fraction. CONCLUSION: The effects of Centella asiatica for lowing _i and arresting cell cycle at S and G₂/M stages of spleen lymphocytes are responsible for its anti-proliferative activitiy.