细胞膜富含脯氨酸和谷氨酰胺的剪切因子维持急性髓系白血病HL-60细胞对多柔比星的敏感性

Splicing factor proline/glutamine-rich contributes to the sensitivity of acute myeloid leukemia HL-60 cells to doxorubicin

目的探讨细胞膜抗原富含脯氨酸和谷氨酰胺的剪切因子（SFPQ）在多柔比星（ADR）耐药的白血病细胞中的作用。方法以急性髓系白血病细胞株HL-60及其ADR耐药株HL．60／ADR细胞为研究对象，通过免疫荧光法比较HL。60和HL-60／ADR细胞膜SFPQ表达丰度；应用单抗5D12封闭细胞膜SFPQ，通过MTr法测定5D12对HL-60、HL-60／ADR细胞增殖的影响，计算ADR的半数抑制浓度（Jc钔）和5D12对细胞增殖的促进率；流式细胞术检测5D12对罗丹明在细胞内蓄积的影响。结果SFPQ在HL-60细胞膜表达高于HL-60／ADR细胞。5D12封闭膜SFPQ作用后，ADR作用HL-60细胞48h的‰由（0．19±0．03）μg／ml升高至（0．95±0．13）μg／ml，HL-60／ADR细胞的IC50（4．41±2．42）μg／ml升高至（21．33±4．26）μg／m1。l、3、5、8、12μg／ml的5D12作用细胞96h后，HL-60细胞增殖促进率分别为（9．12±2．02）％、（16．63±0．92）％、（19．04±0．25）％、（24．17±0．53）％、（34．04±3．20）％，HL-60／ADR细胞分别为（7．40±2．23）％、（8．72±2．38）％、（10．47±3．78）％、（11．57±1．49）％、（13．97±0．91）％。结论核蛋白SFPQ在HL-60细胞膜表达丰度高于HL-60／ADR细胞；该蛋白在细胞膜易位表达不改变药物在细胞内的蓄积，通过抑制细胞增殖维持HL-60细胞对ADR的敏感性。

Objective To investigate the anti-leukemia adriamyein （ADR） effect of membrane protein Splicing factor proline/glutamine-rich （SFPQ）. Methods HL-60 and its adriamycin-resistant cells HL-60/ADR were cultured in vitro. Expression of SFPQ on HL-60 and HL-60/ADR cell membranes were examined by immunofluorescence. McAb 5D12 was used to block membrane SFPQ protein activity. ADR susceptibility and cell proliferation were analyzed by MTF assay, lCso values of ADR in HL-60 and HL-60/ADR cell lines and up-regulations in cell proliferation induced by 5D12 were calculated. Intracellular accumulation of rhodamine in HL-60 and HL-60/ADR cells were measured using fluorescence-activated cell sorting. Results Expression of SFPQ on cell membrane was higher in HL-60 ceils compared to the HL-60/ADR cell line. After membrane-blocking with 5D12, ADR sensitivity was decreased in vitro compared with the untreated cells [the 48 h IC50 value, HL-60 cell line （0.19±0.03）μg/ml vs （0.95±0.13）μg/ml, HL-60/ADR cell line （14.41±2.42）μg/ml vs （21.33±4.26） μg/ml]. Blocking of membrane SFPQ by 5D12 did not affect the intracellular accumulation of rhodamine in these cells, however, 5D12 induced HL-60 and HL-60/ADR cell proliferation, following 1, 3, 5, 8 and 12 μg/ml 5D12 treatment for 96 h were （9.12±2.02） %, （16.63±0.92） %, （19.04±0.25） %, （24.17± 0.53） %, （34.04±3.20） % （HE-60）, and （7.40±2.23） %, （8.72±2.38） %, （10.47±3.78） %, （11.57±1.49） %, （13.97±±0.91） % （HL-60/ADR）, respectively. Conclusion Nuclear protein-SFPQ contributes to HL-60＇ s sensitivity to adriamyein by increasing its surface expression and promoting cell proliferation, but the protein has no significant effect on the intracellular accumulation of the drug.