CAJ | 학술논문

目的探讨311 nm中波紫外线通过Rho家族小GTP酶对B16小鼠黑素瘤细胞树突生成的影响.方法用相差显微镜和激光共聚焦显微镜分别观察311 nm中波紫外线照射后对B16黑素瘤细胞的树突变化和细胞骨架蛋白F-肌动蛋白的影响;用Pull down方法检测311 nm中波紫外线照射前及照射后不同时间点(15、30、60和120 min)B16黑素瘤细胞GTP-RhoA和GTP-Racl蛋白的表达情况.结果 311 nm中波紫外线100 mJ/cm2照射后24 h,B16黑素瘤细胞呈现胞体增大近似球状,树突明显增多似树枝,与未照光细胞比较树突数量明显增多(P＜0.01);而未照光细胞的树突仅表现为双极或三极,细胞分叉不明显.激光共聚焦显微镜显示,311 nm中波紫外线100 mJ/cm2照射前,B16黑素瘤细胞内张力纤维纹理清晰,照射后30 min和60 min细胞内张力纤维解聚、断裂,纹理欠清晰,在照射后6 h时张力纤维纹理模糊,呈团块状.Pull down结果显示,311 nm中波紫外线100 mJ/cm2照射后15 min和30 min,B16黑素瘤细胞GTP-Racl蛋白的表达逐渐升高,在30 min时表达是照射前的2倍多,随后略有下降,但在照射后60 min和120 min时表达仍高于对照组;而GTP-RhoA蛋白在照射后略有下降,而后逐渐升高,在照射后120 min时升高到照射前的1.6倍.结论 311 nm中波紫外线可通过活化Racl促B16黑素瘤细胞树突生成.
목적 탐토311 nm중파자외선통과Rho가족소GTP매대B16소서흑소류세포수돌생성적영향.방법 용상차현미경화격광공취초현미경분별관찰311 nm중파자외선조사후대B16흑소류세포적수돌변화화세포골가단백F-기동단백적영향;용Pull down방법검측311 nm중파자외선조사전급조사후불동시간점(15、30、60화120 min)B16흑소류세포GTP-RhoA화GTP-Racl단백적표체정황.결과 311 nm중파자외선100 mJ/cm2조사후24 h,B16흑소류세포정현포체증대근사구상,수돌명현증다사수지,여미조광세포비교수돌수량명현증다(P＜0.01);이미조광세포적수돌부표현위쌍겁혹삼겁,세포분차불명현.격광공취초현미경현시,311 nm중파자외선100 mJ/cm2조사전,B16흑소류세포내장력섬유문리청석,조사후30 min화60 min세포내장력섬유해취、단렬,문리흠청석,재조사후6 h시장력섬유문리모호,정단괴상.Pull down결과현시,311 nm중파자외선100 mJ/cm2조사후15 min화30 min,B16흑소류세포GTP-Racl단백적표체축점승고,재30 min시표체시조사전적2배다,수후략유하강,단재조사후60 min화120 min시표체잉고우대조조;이GTP-RhoA단백재조사후략유하강,이후축점승고,재조사후120 min시승고도조사전적1.6배.결론 311 nm중파자외선가통과활화Racl촉B16흑소류세포수돌생성.
Objective To investigate the effect ofnarrow-band ultraviolet-B(NB-UVB)(311 nm)on dendrite formation in B16 melanoma cells.Methods B16 melanoma cells were irradiated with various doses of NB-UVB(0,25,50,100,200,300 mJ/cm2).Atier additional culture of varying durations,irradiated cells were harvested and subjected to the observation of morphological changes and cell cytoskeleton F-actin microfilaments by phase contrast microscopy and laser scanning confocal microscopy(LSCM).respectively,and to the detection of cell proliferation bv MTT colorimetric assay.Pull down assay was performed to detect the activity of GTP-RhoAA and GTP-Rac1 in B16 cells before and after UVB irradiation.Results Twenty-four hours after irradiation with UVB of 100 mJ/cm2.an increase was observed in the cell body of B16 cells which appeared in sphericity,as well as in the number of dendrites(P＜0.01)which showed a branch-like appearance.compared with non-irradiated cells which had 2-3 dendrites and obscure branches.LSCM revealed that F-actin microfilaments in B16 cells were well organized with clear textures before irradiation;after irradiation wim NB-UVB of 100 mJ/cm2.stress fibers were disassembled and disrupted and the texture became unclear,which was observed as early as 30 minutes and became more and more evident,and at 6 hours the stress fibers displayed a clumping appearance with obscure textures.Following the irradiation with NB-UVB of 100 mJ/cm2,the expression level of GTP-Rac 1 protein increased at l 5 minutes,and.at 30minutes,reached 2 times of that observed in nonirradiated cells,then decreased a liale,but still remained elevated at 60 minutes and 120 minutes,compared to unirradiated cells;meanwhile.the level of GTP-RhoA dropped a little at 30 minutes,then gradually increased and,at 120 minutes.reached 1.6 times of that observed in unirradiated cells.Conclusion Narrow-band UVB(311 nm)can promote dendrite formation.likely via regulating the expression of GTP-Racl and GTP-RhoA in B16 melanoma cells.