中华肿瘤杂志
中華腫瘤雜誌
중화종류잡지
CHINESE JOURNAL OF ONCOLOGY
2014年
1期
11-16
,共6页
周永春%王熙才%陈艳%黄尤光%刘馨%金从国%陈晓群%李佳%黄云超
週永春%王熙纔%陳豔%黃尤光%劉馨%金從國%陳曉群%李佳%黃雲超
주영춘%왕희재%진염%황우광%류형%금종국%진효군%리가%황운초
肺肿瘤%肿瘤移植%Sox4,基因%小鼠
肺腫瘤%腫瘤移植%Sox4,基因%小鼠
폐종류%종류이식%Sox4,기인%소서
Lung neoplasms%Neoplasms transplantation%Sox4,Gene%Mice
目的 探讨抑制宣威地区女性肺癌细胞系XWLC-05中转录因子Sox4的表达对裸鼠皮下移植瘤生长的影响.方法 构建靶向抑制Sox4基因的shRNA干扰质粒pGFP-V-RS-Sox4 shRNA,并转染XWLC-05细胞;采用实时荧光定量PCR和Western blot检测Sox4的表达水平;将稳定转染重组质粒的XWLC-05细胞接种于裸小鼠皮下,建立移植瘤模型;观测裸小鼠一般情况、成瘤情况和肿瘤生长情况,计算肿瘤抑制率;对小鼠行CT扫描以观察肿瘤的转移情况;免疫组化染色检测移植瘤中Sox4和ki-67的表达.结果 成功构建了高效抑制Sox4基因表达的pGFP-V-RS-Sox4 shRNA重组干扰质粒,并筛选出稳定转染的XWLC-05细胞.除接种生理盐水组外,接种稳定转染干扰质粒pGFP-V-RS-Sox4 shRNA的XWLC-05细胞组、接种稳定转染pGFP-V-RS-scram shRNA的XWLC-05细胞组和接种亲本XWLC-05细胞组小鼠的成瘤率均为100％.空白对照组(不加任何试剂的亲本细胞)、阴性对照组(转染质粒pGFP-V-RS-scram shRNA)和实验组(转染质粒pGFP-V-RS-Sox4 shRNA)小鼠的体重均有不同程度的增长,但差异无统计学意义(P＞0.05).与空白对照组和阴性对照组比较,抑制Sox4基因表达后的实验组小鼠移植瘤的增长趋势明显受到抑制,差异均有统计学意义(均P＜0.05).实验组离体移植瘤体积为(2.30 ±0.34)cm3,与阴性对照组[(3.99±0.45)cm3]和空白对照组[(4.03±0.42)cm3]比较,差异均有统计学意义(均P＜0.05).实验组离体移植瘤重量为(0.86±0.14)g,低于阴性对照组[(1.84±0.27)g]和空白对照组[(1.86±0.22)g],差异均有统计学意义(均P＜0.05).Sox4和ki-67蛋白主要表达于细胞核,实验组移植瘤细胞中Sox4蛋白的表达低于阴性对照组和空白对照组,差异均有统计学意义(均P ＜0.05).影像学和病理学检查结果显示,在实验期内成瘤小鼠的肿瘤均未发生远处转移.结论 成功建立了宣威女性肺癌细胞系XWLC-05的裸鼠移植瘤模型,干扰Sox4基因的表达可抑制移植瘤的生长.
目的 探討抑製宣威地區女性肺癌細胞繫XWLC-05中轉錄因子Sox4的錶達對裸鼠皮下移植瘤生長的影響.方法 構建靶嚮抑製Sox4基因的shRNA榦擾質粒pGFP-V-RS-Sox4 shRNA,併轉染XWLC-05細胞;採用實時熒光定量PCR和Western blot檢測Sox4的錶達水平;將穩定轉染重組質粒的XWLC-05細胞接種于裸小鼠皮下,建立移植瘤模型;觀測裸小鼠一般情況、成瘤情況和腫瘤生長情況,計算腫瘤抑製率;對小鼠行CT掃描以觀察腫瘤的轉移情況;免疫組化染色檢測移植瘤中Sox4和ki-67的錶達.結果 成功構建瞭高效抑製Sox4基因錶達的pGFP-V-RS-Sox4 shRNA重組榦擾質粒,併篩選齣穩定轉染的XWLC-05細胞.除接種生理鹽水組外,接種穩定轉染榦擾質粒pGFP-V-RS-Sox4 shRNA的XWLC-05細胞組、接種穩定轉染pGFP-V-RS-scram shRNA的XWLC-05細胞組和接種親本XWLC-05細胞組小鼠的成瘤率均為100％.空白對照組(不加任何試劑的親本細胞)、陰性對照組(轉染質粒pGFP-V-RS-scram shRNA)和實驗組(轉染質粒pGFP-V-RS-Sox4 shRNA)小鼠的體重均有不同程度的增長,但差異無統計學意義(P＞0.05).與空白對照組和陰性對照組比較,抑製Sox4基因錶達後的實驗組小鼠移植瘤的增長趨勢明顯受到抑製,差異均有統計學意義(均P＜0.05).實驗組離體移植瘤體積為(2.30 ±0.34)cm3,與陰性對照組[(3.99±0.45)cm3]和空白對照組[(4.03±0.42)cm3]比較,差異均有統計學意義(均P＜0.05).實驗組離體移植瘤重量為(0.86±0.14)g,低于陰性對照組[(1.84±0.27)g]和空白對照組[(1.86±0.22)g],差異均有統計學意義(均P＜0.05).Sox4和ki-67蛋白主要錶達于細胞覈,實驗組移植瘤細胞中Sox4蛋白的錶達低于陰性對照組和空白對照組,差異均有統計學意義(均P ＜0.05).影像學和病理學檢查結果顯示,在實驗期內成瘤小鼠的腫瘤均未髮生遠處轉移.結論 成功建立瞭宣威女性肺癌細胞繫XWLC-05的裸鼠移植瘤模型,榦擾Sox4基因的錶達可抑製移植瘤的生長.
목적 탐토억제선위지구녀성폐암세포계XWLC-05중전록인자Sox4적표체대라서피하이식류생장적영향.방법 구건파향억제Sox4기인적shRNA간우질립pGFP-V-RS-Sox4 shRNA,병전염XWLC-05세포;채용실시형광정량PCR화Western blot검측Sox4적표체수평;장은정전염중조질립적XWLC-05세포접충우라소서피하,건립이식류모형;관측라소서일반정황、성류정황화종류생장정황,계산종류억제솔;대소서행CT소묘이관찰종류적전이정황;면역조화염색검측이식류중Sox4화ki-67적표체.결과 성공구건료고효억제Sox4기인표체적pGFP-V-RS-Sox4 shRNA중조간우질립,병사선출은정전염적XWLC-05세포.제접충생리염수조외,접충은정전염간우질립pGFP-V-RS-Sox4 shRNA적XWLC-05세포조、접충은정전염pGFP-V-RS-scram shRNA적XWLC-05세포조화접충친본XWLC-05세포조소서적성류솔균위100％.공백대조조(불가임하시제적친본세포)、음성대조조(전염질립pGFP-V-RS-scram shRNA)화실험조(전염질립pGFP-V-RS-Sox4 shRNA)소서적체중균유불동정도적증장,단차이무통계학의의(P＞0.05).여공백대조조화음성대조조비교,억제Sox4기인표체후적실험조소서이식류적증장추세명현수도억제,차이균유통계학의의(균P＜0.05).실험조리체이식류체적위(2.30 ±0.34)cm3,여음성대조조[(3.99±0.45)cm3]화공백대조조[(4.03±0.42)cm3]비교,차이균유통계학의의(균P＜0.05).실험조리체이식류중량위(0.86±0.14)g,저우음성대조조[(1.84±0.27)g]화공백대조조[(1.86±0.22)g],차이균유통계학의의(균P＜0.05).Sox4화ki-67단백주요표체우세포핵,실험조이식류세포중Sox4단백적표체저우음성대조조화공백대조조,차이균유통계학의의(균P ＜0.05).영상학화병이학검사결과현시,재실험기내성류소서적종류균미발생원처전이.결론 성공건립료선위녀성폐암세포계XWLC-05적라서이식류모형,간우Sox4기인적표체가억제이식류적생장.
Objective To study the effect of targeted Sox4 gene-knock-down on the growth of xenografts of Xuanwei female lung cancer cell line XWLC-05 cells in nude mice.Methods Recombinant plasmid pGFP-V-RS-Sox4 shRNA was constructed and transfected into XWLC-05 cells.Real-time quantitative PCR and Western blot were applied to confirm the effect of Sox4 gene-knock-down.XWLC-05 cells stably transfected with the plasmids were inoculated into nude mice to establish the xenograft model.The nude mouse status,tumor formation and tumor growth were observed,and the tumor inhibition rate was calculated.CT scan was performed to assess the metastasis of xenografts.Immunohistochemical staining was applied to detect Sox4 and ki-67 protein expression.Results Recombinant plasmid pGFP-V-RS-A-Sox4 shRNA which can effectively knocking-down Sox4 gene was successfully constructed and the stable transfected cells were selected by puromycin-screening.The success rate of tumor cell inoculation was 100％ in the mice of all groups except those inoculated with saline.The body weight of all mice inoculated with parental XWLC-05 cells (blank control),pGFP-V-RS-scram shRNA trsfected XWLC-05 cells (negative control),and pGFP-V-RS-Sox4 shRNA transfected XWLC-05 cells was increased to a varying degree,but there was no significant difference among the groups (P ＞ 0.05).The growth of xenografts was significantly inhibited after silencing the Sox4 gene expression when compared with that of the blank and negative controls (P ＜ 0.05).The volume of removed tumors of the Sox4 gene-inhibited mice was (2.30 ± 0.34) cm3,significantly smaller than that of the negative control (3.99 ± 0.45) cm3 and the blank control (4.03 ± 0.42) cm3(P ＜ 0.05).The weight of removed tumors of Sox4 gene-inhibited mice was (0.86 ± 0.14)g,significantly lower than that of the negative control (1.84 ± 0.27) g and blank control (1.86 ± 0.22) g,(P ＜0.05).Immunohistochemical staining showed that Sox4 and ki-67 proteins mainly expressed in cell nuclei.The staining was significantly decreased in xenografts of Sox4-inhibited mice when compared with the negative and blank controls (P ＜ 0.05).No distant metastasis was found in any mouse by CT imaging and pathological examination during the observation period.Conclusions The xenograft model of Xuanwei female lung cancer cell line XWLC-05 cells in nude mice is successfully established.Knocking-down of Sox4 gene can suppress the xenograft tumor growth.
