高压氧对鼻咽癌裸鼠移植瘤生长的抑制作用及其机制

Inhibitory effect of hyperbaric oxygen on growth of transplanted nasopharyngeal carcinoma in nude mice and possible mechanism involved

目的探讨高压氧(HBO)对鼻咽癌裸鼠移植瘤生长的抑制作用及其机制。方法建立人鼻咽癌CNE-2细胞裸鼠移植瘤模型,取荷瘤成功裸鼠20只,按照随机数字表法分为对照组和HBO组,每组10只。HBO组裸鼠每天接受HBO暴露1次,每次1 h,连续处理2周。处理2周后,将2组裸鼠全部处死并测量肿瘤质量和体积,计算抑瘤率。采用RT-PCR实验检测移植瘤血管内皮细胞生长因子(VEGF)和碱性成纤维细胞生长因子(bFGF)mRNA的表达水平。采用免疫组化法检测移植瘤VEGF、bFGF的蛋白表达和肿瘤病理微血管密度(MVD)。采用Spearman相关性分析评估VEGF、bFGF蛋白表达与MVD的相关性。结果HBO连续处理14 d,HBO组移植瘤裸鼠肿瘤体质量抑制率和肿瘤体积抑制率分别为43.12%和43.06%,均明显低于对照组(P<0.01)。HBO组移植瘤组织中VEGF、bFGF mRNA的表达量均显著低于对照组(P<0.01)。免疫组化结果显示,VEGF、bFGF蛋白在肿瘤细胞胞质内表达,阳性细胞细胞质染色呈棕黄色,细胞核染色为淡蓝色。HBO组移植瘤组织中VEGF、bFGF蛋白表达均显著低于对照组(P<0.01)。免疫组化结果显示,CD-34蛋白在血管内皮细胞内表达,染色呈棕黄色,微血管形态各异、大小差别大,尚未形成规则血管腔。HBO组MVD(33.16±4.89)显著低于对照组(45.69±5.26),差异有统计学意义(P<0.01)。Spearman相关性分析结果显示,VEGF蛋白表达与MVD呈正相关(r=0.862,P=0.001),bFGF蛋白表达与MVD亦呈正相关(r=0.745,P=0.013)。结论HBO对鼻咽癌裸鼠移植瘤具有显著的生长抑制作用,其作用机制可能与降低VEGF、bFGF的表达水平有关。

Objective To investigate the inhibitory effect of hyperbaric oxygen(HBO)on the growth of transplanted nasopharyngeal carcinoma(NPC)in nude mice and its mechanism.Methods After the transplanted tumor model of human NPC cell line(CNE-2)in nude mice was established,twenty tumor-bearing nude mice were divided into control and HBO group by using a random number table method,each with 10 mice.The nude mice in the HBO group were exposed to one-hour-session of HBO a day for two consecutive weeks.After two weeks treatments,the mice in both groups were sacrificed,and tumor mass and volume were measured to calculate the tumor growth inhibition rate.Real-time polymerase chain reaction(RT-PCR)was used to detect the mRNA expression levels of vascular endothelial growth factor(VEGF)and basic fibroblast growth factor(bFGF)in transplanted tumor cells.Immunohistochemistry analysis was used to detect the expression levels of VEGF and bFGF proteins and tumor microvessel density(MVD).Spearman correlation analysis was used to assess the correlation between VEGF/bFGF expressions and MVD.Results After 14 consecutive days of HBO treatment,the inhibition rates of tumor mass and tumor volume in the transplanted tumor model mice of the HBO group were respectively 43.12%and 43.06%,both were significantly lower than those of the control group(P<0.01).The expression levels of VEGF mRNA and bFGF mRNA in the transplanted tumor tissue of the HBO group were significantly lower than those of the control group(P<0.01).Immunohistochemistry detection results showed that,regarding the VEGF and bFCG expressions in the cytoplasm of tumor cells,the cytoplasmic staining of positive cells was brownish yellow,and the nucleus staining was light blue.The expression levels of VEGF and bFGF in the HBO group were significantly lower than those in the control group(P<0.01).The immunohistochemistry detection results also indicated that,regarding the CD-34 protein expression in vascular endothelial cells,the staining was brownish yellow.The microvessels were of different shapes and sizes,and no regular vascular lumen was formed.The MVD value of the HBO group(33.16±4.89)was significantly lower than that of the control group(45.69±5.26),with statistical significance(P<0.01).Spearman correlation analysis demonstrated that VEGF protein expression was positively correlated with MVD expression(r=0.862,P=0.001),and bFGF protein expression was also positively correlated with MVD expression(r=0.745,P=0.013).Conclusion HBO has a significant growth inhibitory effect on the transplanted NPC in nude mice,the mechanism of which might be related to the reduction of the expression levels of VEGF and bFGF.