CAJ | 학술논문

目的：探讨膜结合型巨噬细胞集落刺激因子（ mM-CSF）及其剪切体（ mM-CSF-Δ）对淋巴细胞白血病Ramos细胞增殖的抑制作用。方法采用Overlap PCR法构建带有mM-CSF的真核表达质粒pTARGET-mM-CSF，再进一步构建胞内区截短30个氨基酸的表达质粒pTARGET-mM-CSF-Δ，并进行PCR及DNA双向测序鉴定。将空载体pTARGET、pTARGET-mM-CSF、pTARGET-mM-CSF-Δ质粒分别转染Ramos细胞，经G418筛选稳定表达细胞株，并用RT-PCR、Western blotting进行鉴定；MTT法检测细胞增殖，流式细胞仪检测细胞周期。结果成功构建了mM-CSF和 mM-CSF-Δ的真核表达载体，获得了稳定转染细胞株 Ramos-V、Ramos-M 和 Ramos-Δ。 Ramos-M、Ramos-Δ、Ramos-V细胞增殖能力的OD值分别为0．413±0．014、0．384±0．019、0．463±0．037，Ramos-M细胞与Ramos-Δ细胞比较，Ramos-M、Ramos-Δ细胞分别与Ramos-V细胞比较，P＜0．05或＜0．01。 Ramos-M、Ramos-Δ、Ramos-V细胞处于G0／G1期的比例分别为41．54％±1．22％、45．60％±1．09％、39．20％±1．53％，Ramos-M细胞与Ramos-Δ细胞比较， Ramos-M、Ramos-Δ细胞分别与 Ramos-V 细胞比较， P ＜0．05或＜0．01。结论 mM-CSF、mM-CSF-Δ均能抑制淋巴细胞白血病Ramos细胞增殖，且后者抑制作用更强。
목적：탐토막결합형거서세포집락자격인자（ mM-CSF）급기전절체（ mM-CSF-Δ）대림파세포백혈병Ramos세포증식적억제작용。방법채용Overlap PCR법구건대유mM-CSF적진핵표체질립pTARGET-mM-CSF，재진일보구건포내구절단30개안기산적표체질립pTARGET-mM-CSF-Δ，병진행PCR급DNA쌍향측서감정。장공재체pTARGET、pTARGET-mM-CSF、pTARGET-mM-CSF-Δ질립분별전염Ramos세포，경G418사선은정표체세포주，병용RT-PCR、Western blotting진행감정；MTT법검측세포증식，류식세포의검측세포주기。결과성공구건료mM-CSF화 mM-CSF-Δ적진핵표체재체，획득료은정전염세포주 Ramos-V、Ramos-M 화 Ramos-Δ。 Ramos-M、Ramos-Δ、Ramos-V세포증식능력적OD치분별위0．413±0．014、0．384±0．019、0．463±0．037，Ramos-M세포여Ramos-Δ세포비교，Ramos-M、Ramos-Δ세포분별여Ramos-V세포비교，P＜0．05혹＜0．01。 Ramos-M、Ramos-Δ、Ramos-V세포처우G0／G1기적비례분별위41．54％±1．22％、45．60％±1．09％、39．20％±1．53％，Ramos-M세포여Ramos-Δ세포비교， Ramos-M、Ramos-Δ세포분별여 Ramos-V 세포비교， P ＜0．05혹＜0．01。결론 mM-CSF、mM-CSF-Δ균능억제림파세포백혈병Ramos세포증식，차후자억제작용경강。
Objective To investigate the inhibitory effect of membrane-bound macrophage colony-stimulating factor ( mM-CSF) and its spliceosome ( mM-CSF-Δ) on proliferation of lymphocytic leukemia cell line Ramos.Methods The eukaryotic expression plasmid of pTARGET-mM-CSF with mM-CSF was constructed with Overlap PCR, and then pTAR-GET-mM-CSF-Δof 30 amino acide located in the intracellular region of brachytmema mutation was obtained; and mean-while, they were identified by PCR and DNA sequencing.The empty vector pTARGET, pTARGET-mM-CSF and pTAR-GET-mM-CSF-Δplasmid were transfected into Ramos cells, the cell line with stable expression was screened by G418 and was detected by RT-PCR and Western blotting.Proliferation and cell cycle of these stably transfected cells were detected by MTT and flow cytometry.Results The mM-CSF and mM-CSF-Δeukaryotic expression vectors were successfully construc-ted.The stable transfectants Ramos-V, Ramos-M and Ramos-Δwere obtained.The OD of proliferation ability of Ramos-M, Ramos-Δand Ramos-V were 0.413 ±0.014, 0.384 ±0.019 and 0.463 ±0.037, respectively.Significant difference was found between the cell lines of Ramos-M and Ramos-Δ, among the cell lines of Ramos-M, Ramos-Δand Ramos-V, re-spectively (P<0.05 or P <0.01).The proportion of G0/G1 phases of Ramos-M, Ramos-Δand Ramos-V cells were 41.54%±1.22%, 45.60%±1.09% and 39.20% ±1.53%, respectively.Significant difference was found between <br> Ramos-M and Ramos-Δcells, among Ramos-M, Ramos-Δand Ramos-V cells (P<0.05 or P<0.01).Conclusion mM-CSF and mM-CSF-Δmay inhibit the proliferation of lymphocytic leukemia cell line Ramos and the inhibitory effect of the latter is stronger.