CAJ | 학술논문

目的探讨不同体外培养条件与血管平滑肌细胞(VSMCs)表型、增殖和细胞骨架蛋白表达之间的差异及生物学意义.方法体外培养大鼠主动脉血管平滑肌细胞,实验分为6组:2代对照组、2代血清饥饿组、4代对照组、4代血清饥饿组、6代对照组和6代血清饥饿组;用5-Brdu标记增殖细胞;RT-PCR检测表型基因SM22ɑ mRNA的表达;免疫细胞化学检测细胞骨架蛋白SMɑ-actin、β-Tubulin和Desmin的表达. 结果　体外培养的VSMCs,随培养代数的增多,细胞骨架蛋白表达减少,电镜超微结构可见胞质中内质网、高尔基体等细胞器增多,有大量的分泌小泡;血清饥饿胞中线粒体增多并电子密度增高. 血清培养条件下SMɑ-actin随培养代数的增加其表达减少,血清饥饿培养可使SMɑ-actin表达可逆性上调;β-Tubulin和Desmin随培养代数的增加表达减少更明显,而血清饥饿上调作用也不明显,至第6代基本不表达. 结论血清培养和血清饥饿培养在不同代中对VSMCs表型转化、增殖和细胞骨架有着不同的影响,它们之间存在着内在的联系,在维持细胞形态、收缩功能和血管重构方面起了重要作用.β-Tubulin和Desmin表达消失可能具有重要生物学意义.
목적 탐토불동체외배양조건여혈관평활기세포(VSMCs)표형、증식화세포골가단백표체지간적차이급생물학의의.방법 체외배양대서주동맥혈관평활기세포,실험분위6조:2대대조조、2대혈청기아조、4대대조조、4대혈청기아조、6대대조조화6대혈청기아조;용5-Brdu표기증식세포;RT-PCR검측표형기인SM22ɑ mRNA적표체;면역세포화학검측세포골가단백SMɑ-actin、β-Tubulin화Desmin적표체. 결과　체외배양적VSMCs,수배양대수적증다,세포골가단백표체감소,전경초미결구가견포질중내질망、고이기체등세포기증다,유대량적분비소포;혈청기아포중선립체증다병전자밀도증고. 혈청배양조건하SMɑ-actin수배양대수적증가기표체감소,혈청기아배양가사SMɑ-actin표체가역성상조;β-Tubulin화Desmin수배양대수적증가표체감소경명현,이혈청기아상조작용야불명현,지제6대기본불표체. 결론 혈청배양화혈청기아배양재불동대중대VSMCs표형전화、증식화세포골가유착불동적영향,타문지간존재착내재적련계,재유지세포형태、수축공능화혈관중구방면기료중요작용.β-Tubulin화Desmin표체소실가능구유중요생물학의의.
Objective To study the influence of different culture conditions in vitro on phenotype, proliferation and cytoskeletal proteins expression of vascular smooth muscle cells(VSMCs). Methods The cultured VSMCs from rat aorta were divided into six groups: P2 control,P2 starvation,P4 control,P4 starvation,P6 control and P6 starvation. The proliferating cells were labeled by 5-bromodeoxyuridine (5-BrdU); The mRNA expression of smooth muscle 22 alpha (SM22α) was detected by reverse transcription-polymerase chain reaction method (RT-PCR); The cytoskeletal proteins including SMα-actin,β-Tubulin and Desmin were observed through immunohistochemical staining. Results With the increase of cell passage, cytoskeletal proteins expression of VSMCs decreased,cellular organs increased and secretory vesicles were abundant; in serum-free cultured cells mitochondria increased and electron density enhanced in cytoplasm of VSMCs.On the contrary the expression of SMα-actin decreased, and the expression of SMα-actin increased. The expression of β-Tublin and Desmin decreased more obviously, and at 6 passages failed to express. Conclusion The conditioned medium and serum-free had the different effects on the phenotype,proliferation and cytoskeleton of VSMCs in different passage, and there was internal relationship among them. The internal relationship played an important role in the maintaining of cell morphology, contractile function and vascular remodeling. The disappearance of expression of β-Tubulin and desmin might have important biological significance.