神经母细胞瘤细胞分化过程中细胞周期及端粒酶基因的变化

目的：观察神经母细胞瘤细胞在被维甲类化合物诱导分化过程中细胞形态、细胞周期及端粒酶基因表达方面的变化，探讨其作用机理及临床意义。方法：查耳酮酸（R9158）及丁羟胺酸（R8605）均为新一代合成维甲类化合物，采用工作浓度10^-6mol/L，加入两个神经母细胞瘤细胞系SH-SY-5Y和SK-N-SH的培养基，作为观察组；与观察组同浓度的DMSO分别加入两个细胞系的培养基，作为溶剂对照组。分别观察用药前后细胞形态学变化、流式细胞术检测细胞周期分布及凋亡、原位杂交细胞化学（ISHH）方法检测端粒酶基因（hTR )的表达。结果：1.SH-SY-5Y细胞在药物处理后第5天明显的神经突起形成，形态类似成熟神经元。SK-N-SN细胞在药物处理后7天内均无明显变化。2.流式细胞分析发现，SH-SY-5Y细胞在药物处理后第5天明显地阻滞于G1期，S期比例显著减少，G2/M期比例基本不变，并有少量细胞凋亡。SK-N-SH细胞在药物处理第3天及第5天均无明显差别，仅第5天有个别细胞凋亡。3.显微镜下观察，SH-SY-5Y细胞于药物作用5天后，hTR阳性细胞显著减少，SK-N-SH细胞则无明显变化。结论：新合成维甲类化合物R9158及R8605诱导维甲酸敏感细胞SH-SY-5Y发生形态学分化，细胞周期阻滞于G1期，端粒酶基因（hTR）表达显著减少，而维甲酸耐受细胞SK-N-SH则无此变化。端粒酶与其他基因相互关联，是维甲类化合物诱导神经母细胞瘤细胞分化过程中重要的调控靶点。

Variation of Cell Cycle and Telomerase Gene Expression in the Differentiation of Neuroblastoma Cells Induced by Retinoids

Objective:The current study was designed to observe and discuss the variation of neu roblastoma cells in morphology,cell cycle and telome rase gene(hTR)expression induced by retinoids.Methods:Two newly-synthesized retinoids,R9158and R8605,were supplemented into the media of neurobla stoma cells,SH-SY-5Y and SK-N-SH,with the concentration of 10 -6 mol /L as the treatment group.Meanwhile the same supplements of DMSO were as the control group.Pre-and post-supplement cells were observed for morphological changes,cell cycle was analyzed by flow cyto metry(FCM)and hTR expression was analyzed by in situ hybridization chemistry(ISHH).Results:1)The shape of SH-SY-5Y cells changed significantly at the 5th day of R9158and R8605treatment,having multiple neurite extensions just as matureneu r on.Ho wevernoshapechange ofS K2 N2 SH ce lls was f ound in obse r va ti on f or 7 days.2 For S H2 S Y2 5Y ce lls t he pe rcen t age ofS t age G1 S G2 /M d iff ered much be t ween t he tr eat mentgroup and t hecon tr olgroupat5th day when t he ce llcyc le oft he tr ea t mentgroup wasretar ded atS t age G1 and t he ratio ofS t age decreased significantly.For S K2 N2 SH ce lls ho wever t he re waslittle var i a ti on ofce llcyc le stat usin t he tr ea t mentgroup.3 The S H2 S Y2 5Y ce ll nu mbe rsof hTR pos iti veexp ressionint hetr ea t mentgroupsdecreasedsignificantlyat t he5 thday.Nodecreasewasf oundin t he R9158 and R8605 groupsofS K2 N2 SH ce lls in t he tr ea t mentgroups.Conc l usi ons Du ri ng t he d iff eren ti a ti on of RA2sens iti ve ce lls i nduced by r e ti no i ds t he t e l o merase exp ression was dep ressed and t he ce llcyc le r e t arded whe reas t he RA2resista nt ce llschanged little in morpho l ogy t e l o merase exp ressionand ce llcyc le aswe ll.The t e l o merase gene assoc i a t edwitho t hergeneswou ldbeani mpo rt antregu l a t edt arget int hed iff eren ti a ti on i nducedbyreti no i d s.