CAJ | 학술논문

目的探讨携带IL-18基因的溶瘤腺病毒(ZD55-IL-18)对裸鼠恶性黑素瘤移植瘤的抗肿瘤效果.方法取A375细胞2×106个接种于BALB/c裸鼠腋下组织内,建立裸鼠黑素瘤模型,当移植瘤体积达100～150 mm3时将其随机分为3组,每组8只,分别于瘤体内注射ZD55-IL-18、Ad-IL-18和磷酸盐缓冲液(PBS).通过定期测量肿瘤体积观察肿瘤生长抑制情况,并切取肿瘤进行HE染色,观察肿瘤细胞生长形态;激光共聚焦显微镜下观察移植瘤组织IL-18表达水平、EIA蛋白的表达情况.免疫组化检测荷瘤裸鼠肿瘤组织CD34染色标记测定的肿瘤组织微血管密度;TUNEL法检测移植瘤组织细胞凋亡情况.结果肿瘤生长曲线表明,ZD55-IL-18处理组肿瘤生长速度较PBS组明显减慢,接种44 d后ZD55-IL-18组肿瘤平均体积为(1039.378±29.67)mm3,Ad-IL-18组为(2900.46±62.65)mm3,PBS组为(3980.24±63.78)mm3,各组间差异有统计学意义(P＜0.01).HE染色发现,ZD55-IL-18处理组细胞核深染,很少见核仁,显示肿瘤组织生长抑制.激光共聚焦结果表明,ZD55-IL-18处理组IL-18的表达阳性率比Ad-IL-18处理组明显增强(P＜0.01),并可以在肿瘤组织中表达E1A蛋白;免疫组化检测显示,ZD55-IL-18治疗组肿瘤微血管密度明显低于PBS组(P＜0.05).不同处理组移植瘤细胞凋亡阳性率ZD55-IL-18处理组(86.28%±3.25%)＞Ad-IL-18处理组(43.67%±3.46%)＞PBS处理组(10.73%±2.48%).各组之间差异有统计学意义(P＜0.05).结论携带IL-18基因的溶瘤腺病毒ZD55-IL-18对裸鼠恶性黑素瘤移植瘤的生长及转移有抑制作用,并探讨了其可能的作用环节.
목적 탐토휴대IL-18기인적용류선병독(ZD55-IL-18)대라서악성흑소류이식류적항종류효과.방법 취A375세포2×106개접충우BALB/c라서액하조직내,건립라서흑소류모형,당이식류체적체100～150 mm3시장기수궤분위3조,매조8지,분별우류체내주사ZD55-IL-18、Ad-IL-18화린산염완충액(PBS).통과정기측량종류체적관찰종류생장억제정황,병절취종류진행HE염색,관찰종류세포생장형태;격광공취초현미경하관찰이식류조직IL-18표체수평、EIA단백적표체정황.면역조화검측하류라서종류조직CD34염색표기측정적종류조직미혈관밀도;TUNEL법검측이식류조직세포조망정황.결과 종류생장곡선표명,ZD55-IL-18처리조종류생장속도교PBS조명현감만,접충44 d후ZD55-IL-18조종류평균체적위(1039.378±29.67)mm3,Ad-IL-18조위(2900.46±62.65)mm3,PBS조위(3980.24±63.78)mm3,각조간차이유통계학의의(P＜0.01).HE염색발현,ZD55-IL-18처리조세포핵심염,흔소견핵인,현시종류조직생장억제.격광공취초결과표명,ZD55-IL-18처리조IL-18적표체양성솔비Ad-IL-18처리조명현증강(P＜0.01),병가이재종류조직중표체E1A단백;면역조화검측현시,ZD55-IL-18치료조종류미혈관밀도명현저우PBS조(P＜0.05).불동처리조이식류세포조망양성솔ZD55-IL-18처리조(86.28%±3.25%)＞Ad-IL-18처리조(43.67%±3.46%)＞PBS처리조(10.73%±2.48%).각조지간차이유통계학의의(P＜0.05).결론 휴대IL-18기인적용류선병독ZD55-IL-18대라서악성흑소류이식류적생장급전이유억제작용,병탐토료기가능적작용배절.
Objective To evaluate the antitumor effect of oncolytic adenovirus expressing human IL-18 gene on malignant melanoma implanted in nude mice.Methods BALB/c nude mice were subcutaneously inoculated with A375 cells to establish a model of malignant melanoma.When the volume of implanted tumor reached 100-150 mm3,murine models were randomly divided into three groups to receive a 3-day intratumoral injection of IL-18 gene-expressing human oncolytic adenovirus named ZD55-IL-18,IL-18 gene-expressing adenovirus named Ad-IL-18,phosphate buffer saline(PBS),respectively.The tumor size was measured at an interval of 4 days for 9 weeks.Hematoxylin and eosin (HE)staining was performed to observe the morphological changes of tumor cells.The protein expression of IL-18 and E1A.microvessel density in tumor tissue,and apoptosis of tumor xenografts were detected by immuno fluorescence assay,immunohistochemistry and in situ end labeling technique (TUNEL).respectively.Results The treatment with ZD55-IL-18 significantly inhibited the growth of tumor.Forty-four days after the treatment,the mean tumor volume was 1039.378±29.67 mm3 in ZD55-IL-18-treated mice.significantly smaller than that in Ad-IL-18 treated mice(2900.46±62.65 mm3)and PBS-treated mice(3980.24±63.78 mm3).HE staining showed that the nuclei of tumor cells were heavily stained with few nucleoli in ZD55-IL-18-treated mice.Increased positivity rate of IL-18 was noticed in ZD55-IL-18-treated mice vs.AD55-IL-18-treated mice(83.4%±3.2%vs 24.4%±2.1%.P＜0.01).Moreover,immunofluorescence assay revealed the presence of E1A protein in tumor tissue.A decrease was found in the microvessel density in ZD55-IL-18-treated mice compared with the PBS-treated mice(P＜0.01).The apoptosis rate in tumor cells from high to low was 86.28%±3.25%in ZD55-IL-18-treated mice,43.67%±3.46%in Ad-IL-18-treated mice,and 10.73%±2.48%in PBS-treated mice;there was a significant difference between the three groups(all P＜0.05).Conclusion The oncolytic adenovirus expressing human IL-18 gene,ZD55-IL-18,has a significant inhibitory effect on the growth and metastasis of malignant melanoma implanted in nude mice.