目的 探讨舒尼替尼(Sunitinib)、顺铂(CDDP)及两者联合用药对小儿睾丸卵黄囊瘤(TYST)异种移植荷瘤鼠模型的抗肿瘤作用及相关作用机制.方法 肿瘤标本来自本实验室的小儿睾丸卵黄囊瘤裸鼠第17代模型,并接种在雄性裸鼠单侧腹股沟皮下区,成瘤后随机分成4组(n=5):对照组、CDDP组、Sunitinib组和Sunitinib+CDDP组.绘制肿瘤体积和裸鼠体重变化曲线图,计算肿瘤消退率;HE染色观察肿瘤组织形态学变化;免疫组织化学法检测AFP、Ki-67、Glypican-3、CD105在各组肿瘤中的表达:CD105测定微血管密度(MVD),Ki-6表示细胞增殖率(PI);TUNEL法检测肿瘤细胞凋亡率(AI);实时荧光定量PCR(RT-qPCR)验证靶向因子的mRNA表达变化.结果 各治疗组均能显著抑制肿瘤生长,并能消退肿瘤体积.在治疗后肿瘤体积上,除顺铂组与舒尼替尼组间无统计学差异外(41.61±7.61比67.15±5.39,P＞0.05),其余各组间都有统计学差异:对照组与顺铂(651.72±121.16比41.61±7.61,P＜0.05),对照组与舒尼替尼组(651.72±121.16比67.15±5.39,P＜0.05),对照组联合舒尼替尼±顺铂组(651.72±121.16比23.03±2.37,P＜0.05),舒尼替尼组与联合舒尼替尼+顺铂组(67.15±5.39比23.03±2.37,P＜0.05),顺铂组与联合舒尼替尼+顺铂组(41.61±7.61比23.03±2.37,P＜0.05);在裸鼠体重上,相比对照组,除舒尼替尼组无统计学差异外(25.90±0.75比26.66±0.65,P＞0.05),其余各组间差异均有统计学意义:对照组与顺铂组(25.90±0.75比18.90±0.63,P＜0.05),对照组与联合舒尼替尼+顺铂组(25.90±0.75比18.26±1.20,P＜0.05);AFP、Glypican-3在各治疗组阳性表达面积(Pixels)均少于对照组(AFP:对照组与顺铂组,1.26×106土1.48×105比5.54×105±8.14×104,P＜0.05;对照组与舒尼替尼组,1.26×106±1.48×105比7.09×105±6.64×104,P＜0.05;对照组与联合舒尼替尼+顺铂组,1.26×106±1.48×105比3.62×105±4.83×104,P＜0.05.Glypican-3:对照组与顺铂组,9.68×105±7.63×104比4.04×105±5.04×104,P＜0.05;对照组与舒尼替尼组,9.68×105±7.63×104比4.59×105±2.32×104,P＜0.05;对照组与联合舒尼替尼+顺铂组,9.68×105±7.63×104比1.89×105±2.55×104,P＜0.05).两者在顺铂组与舒尼替尼组的比较中差异无统计学意义(P＞0.05);PI和AI在各治疗组中相比对照组,差异都具有统计学意义(PI和AI:对照组与顺铂组,58.97土1.38比42.36±1.28和1.69±0.20比54.62±2.49,P＜0.01;对照组与舒尼替尼组,58.97±1.38比43.48±1.00和1.69±0.20比47.32±2.00,P＜0.01;对照组与联合舒尼替尼+顺铂组,58.97±1.38比33.34±1.19和1.69±0.20比63.41土2.23,P＜0.01).顺铂组相比联合舒尼替尼+顺铂组,PI:P=0.001,AI:P=0.002;舒尼替尼组相比联合舒尼替尼+顺铂组,PI和AI:P＜0.001;顺铂组相比舒尼替尼组,PI.P=0.597,AI:P=0.059;RT-qPCR证实M-CSFR、PDGFR-β、RET、VEGFR-2的mRNA表达受到抑制.结论 Sunitinib能显著抑制小儿睾丸卵黄囊瘤的生长,并消退肿瘤体积:主要通过直接抑制肿瘤细胞的生长和肿瘤血管的新生,从而诱导肿瘤细胞凋亡,同时伴有直接细胞毒作用引起坏死;Sunitinib相比CDDP具有更轻的毒副作用,且联合CDDP具有增强抗肿瘤作用.
目的 探討舒尼替尼(Sunitinib)、順鉑(CDDP)及兩者聯閤用藥對小兒睪汍卵黃囊瘤(TYST)異種移植荷瘤鼠模型的抗腫瘤作用及相關作用機製.方法 腫瘤標本來自本實驗室的小兒睪汍卵黃囊瘤裸鼠第17代模型,併接種在雄性裸鼠單側腹股溝皮下區,成瘤後隨機分成4組(n=5):對照組、CDDP組、Sunitinib組和Sunitinib+CDDP組.繪製腫瘤體積和裸鼠體重變化麯線圖,計算腫瘤消退率;HE染色觀察腫瘤組織形態學變化;免疫組織化學法檢測AFP、Ki-67、Glypican-3、CD105在各組腫瘤中的錶達:CD105測定微血管密度(MVD),Ki-6錶示細胞增殖率(PI);TUNEL法檢測腫瘤細胞凋亡率(AI);實時熒光定量PCR(RT-qPCR)驗證靶嚮因子的mRNA錶達變化.結果 各治療組均能顯著抑製腫瘤生長,併能消退腫瘤體積.在治療後腫瘤體積上,除順鉑組與舒尼替尼組間無統計學差異外(41.61±7.61比67.15±5.39,P＞0.05),其餘各組間都有統計學差異:對照組與順鉑(651.72±121.16比41.61±7.61,P＜0.05),對照組與舒尼替尼組(651.72±121.16比67.15±5.39,P＜0.05),對照組聯閤舒尼替尼±順鉑組(651.72±121.16比23.03±2.37,P＜0.05),舒尼替尼組與聯閤舒尼替尼+順鉑組(67.15±5.39比23.03±2.37,P＜0.05),順鉑組與聯閤舒尼替尼+順鉑組(41.61±7.61比23.03±2.37,P＜0.05);在裸鼠體重上,相比對照組,除舒尼替尼組無統計學差異外(25.90±0.75比26.66±0.65,P＞0.05),其餘各組間差異均有統計學意義:對照組與順鉑組(25.90±0.75比18.90±0.63,P＜0.05),對照組與聯閤舒尼替尼+順鉑組(25.90±0.75比18.26±1.20,P＜0.05);AFP、Glypican-3在各治療組暘性錶達麵積(Pixels)均少于對照組(AFP:對照組與順鉑組,1.26×106土1.48×105比5.54×105±8.14×104,P＜0.05;對照組與舒尼替尼組,1.26×106±1.48×105比7.09×105±6.64×104,P＜0.05;對照組與聯閤舒尼替尼+順鉑組,1.26×106±1.48×105比3.62×105±4.83×104,P＜0.05.Glypican-3:對照組與順鉑組,9.68×105±7.63×104比4.04×105±5.04×104,P＜0.05;對照組與舒尼替尼組,9.68×105±7.63×104比4.59×105±2.32×104,P＜0.05;對照組與聯閤舒尼替尼+順鉑組,9.68×105±7.63×104比1.89×105±2.55×104,P＜0.05).兩者在順鉑組與舒尼替尼組的比較中差異無統計學意義(P＞0.05);PI和AI在各治療組中相比對照組,差異都具有統計學意義(PI和AI:對照組與順鉑組,58.97土1.38比42.36±1.28和1.69±0.20比54.62±2.49,P＜0.01;對照組與舒尼替尼組,58.97±1.38比43.48±1.00和1.69±0.20比47.32±2.00,P＜0.01;對照組與聯閤舒尼替尼+順鉑組,58.97±1.38比33.34±1.19和1.69±0.20比63.41土2.23,P＜0.01).順鉑組相比聯閤舒尼替尼+順鉑組,PI:P=0.001,AI:P=0.002;舒尼替尼組相比聯閤舒尼替尼+順鉑組,PI和AI:P＜0.001;順鉑組相比舒尼替尼組,PI.P=0.597,AI:P=0.059;RT-qPCR證實M-CSFR、PDGFR-β、RET、VEGFR-2的mRNA錶達受到抑製.結論 Sunitinib能顯著抑製小兒睪汍卵黃囊瘤的生長,併消退腫瘤體積:主要通過直接抑製腫瘤細胞的生長和腫瘤血管的新生,從而誘導腫瘤細胞凋亡,同時伴有直接細胞毒作用引起壞死;Sunitinib相比CDDP具有更輕的毒副作用,且聯閤CDDP具有增彊抗腫瘤作用.
목적 탐토서니체니(Sunitinib)、순박(CDDP)급량자연합용약대소인고환란황낭류(TYST)이충이식하류서모형적항종류작용급상관작용궤제.방법 종류표본래자본실험실적소인고환란황낭류라서제17대모형,병접충재웅성라서단측복고구피하구,성류후수궤분성4조(n=5):대조조、CDDP조、Sunitinib조화Sunitinib+CDDP조.회제종류체적화라서체중변화곡선도,계산종류소퇴솔;HE염색관찰종류조직형태학변화;면역조직화학법검측AFP、Ki-67、Glypican-3、CD105재각조종류중적표체:CD105측정미혈관밀도(MVD),Ki-6표시세포증식솔(PI);TUNEL법검측종류세포조망솔(AI);실시형광정량PCR(RT-qPCR)험증파향인자적mRNA표체변화.결과 각치료조균능현저억제종류생장,병능소퇴종류체적.재치료후종류체적상,제순박조여서니체니조간무통계학차이외(41.61±7.61비67.15±5.39,P＞0.05),기여각조간도유통계학차이:대조조여순박(651.72±121.16비41.61±7.61,P＜0.05),대조조여서니체니조(651.72±121.16비67.15±5.39,P＜0.05),대조조연합서니체니±순박조(651.72±121.16비23.03±2.37,P＜0.05),서니체니조여연합서니체니+순박조(67.15±5.39비23.03±2.37,P＜0.05),순박조여연합서니체니+순박조(41.61±7.61비23.03±2.37,P＜0.05);재라서체중상,상비대조조,제서니체니조무통계학차이외(25.90±0.75비26.66±0.65,P＞0.05),기여각조간차이균유통계학의의:대조조여순박조(25.90±0.75비18.90±0.63,P＜0.05),대조조여연합서니체니+순박조(25.90±0.75비18.26±1.20,P＜0.05);AFP、Glypican-3재각치료조양성표체면적(Pixels)균소우대조조(AFP:대조조여순박조,1.26×106토1.48×105비5.54×105±8.14×104,P＜0.05;대조조여서니체니조,1.26×106±1.48×105비7.09×105±6.64×104,P＜0.05;대조조여연합서니체니+순박조,1.26×106±1.48×105비3.62×105±4.83×104,P＜0.05.Glypican-3:대조조여순박조,9.68×105±7.63×104비4.04×105±5.04×104,P＜0.05;대조조여서니체니조,9.68×105±7.63×104비4.59×105±2.32×104,P＜0.05;대조조여연합서니체니+순박조,9.68×105±7.63×104비1.89×105±2.55×104,P＜0.05).량자재순박조여서니체니조적비교중차이무통계학의의(P＞0.05);PI화AI재각치료조중상비대조조,차이도구유통계학의의(PI화AI:대조조여순박조,58.97토1.38비42.36±1.28화1.69±0.20비54.62±2.49,P＜0.01;대조조여서니체니조,58.97±1.38비43.48±1.00화1.69±0.20비47.32±2.00,P＜0.01;대조조여연합서니체니+순박조,58.97±1.38비33.34±1.19화1.69±0.20비63.41토2.23,P＜0.01).순박조상비연합서니체니+순박조,PI:P=0.001,AI:P=0.002;서니체니조상비연합서니체니+순박조,PI화AI:P＜0.001;순박조상비서니체니조,PI.P=0.597,AI:P=0.059;RT-qPCR증실M-CSFR、PDGFR-β、RET、VEGFR-2적mRNA표체수도억제.결론 Sunitinib능현저억제소인고환란황낭류적생장,병소퇴종류체적:주요통과직접억제종류세포적생장화종류혈관적신생,종이유도종류세포조망,동시반유직접세포독작용인기배사;Sunitinib상비CDDP구유경경적독부작용,차연합CDDP구유증강항종류작용.
Objective To study the antitumor effects of Sunitinib or Sunitinib combind with cis - diamminedichloroplatinum (CDDP) on an athymic mouse human testicular yolk sac tumor xenograft model. Methods The athymic mouse human testicular yolk sac tumor xenograft model was established by subcutaneous injection of 17th passage pediatric testicular yolk sac tumor cells in the unilateral inguinal region of the male nude mice. The mice of control group didn't receive any treatment. The tumor bearing mice were treated with either CDDP, or Sunitinib group, or Sunitinib combined with CDDP. The tumor-bearing nude mice were divided into 4 groups (5 in each) according the treatment they underwent. The tumor volumes and mice weight were measured to calculate the regression rate of tumor. The tumor was collected for H&E staining and immunohistochemical staining of AFP, Ki-67,Glypican-3 and CD105. Microvessel density (MVD) was measured by analyzing the CD105 expression. The tumors' proliferation index (PI) was studied by analyzing Ki-67 expression. The apoptosis of the tumor was quantitated using TUNEL staining. The mRNA expressions of cytokines were determined by quantitative real-time PCR (RT-qPCR). Results The tumor volumes were significantly decreased after chemotherapy. No difference of tumor volume was found between CDDP group and Sunitinib group (41.61 ± 7. 61 vs. 67. 15 ± 5. 39, P＞0. 05). Significant differences of tumor volumes were found between CDDP group and CDDP+ Sunitinib group (41.61 ± 7. 61 vs. 23. 03 ± 2. 37, P＜0. 05), and between Sunitinib group and CDDP+ Sunitinib group (67. 15 ± 5. 39 vs. 23.03 ± 2. 37, P＜0. 05). Of the body weight of tumor-bearing mice, no difference was found between controls and Sunitinib group (25. 90 ± 0. 75 vs. 26. 66 ± 0. 65, P＞0. 05). And significant differences of the body weight were noted between controls and CDDP group (25.90 ± 0. 75 vs. 18. 90 ± 0. 63, P＜0. 05),controls and CDDP+ Sunitinib group (25. 90 ± 0. 75 vs. 18. 26 ± 1.20,P＜0. 05). The positive areas (pixels) of AFP in the mice with chemotherapy were less than those of control mice (Control vs. CDDP: 1.26 × 106±1.48 × 105 vs. 5. 54 × 105 ± 8. 14 × 104 , P＜0. 05. Control vs. Sunitinib: 1.26 × 106 ± 1.48 × 105 vs. 7. 09 × 105 ± 6. 64 × 104, P＜0. 05. Control vs. CDDP + Sunitinib: 1.26 × 106 ± 1.48 × 105 vs. 3. 62 × 105 ± 4. 83 × 104, P＜0. 05). The positive areas (pixels) of Glypican-3 in the mice with chemotherapy were less than those of control mice (Control vs. CDDP: 9. 68 × 105 ± 7. 63 × 104 vs. 4. 04 × 105 ± 5. 04 × 104 , P＜0. 05. Control vs. Sunitinib: 9. 68 × 105 ± 7. 63 × 104 vs. 4. 59 × 105 ± 2. 32 × 104 , P＜0. 05. Control vs. CDDP + Sunitinib: 9. 68 × 105 ± 7. 63 × 104 vs. 1.89 × 105 ± 2. 55 × 104, P＜0. 05). However, there were no statistical differences of the positive areas (pixels) of AFP and Glypican-3 between CDDP and Sunitinib groups (P＞0. 05). The PI was significantly decreased after chemotherapy (Control vs. CDDP: 58. 97 ± 1.38 vs. 42. 36 ± 1.28, P＜ 0. 01. Control vs.Sunitinib: 58. 97 ± 1.38 vs. 43. 48 ± 1.00, P＜0. 01. Control vs. CDDP+ Sunitinib: 58. 97 ± 1.38 vs.33. 34 ± 1.19, P＜0. 01 ). The apoptosis index (AI) was also significantly increased after chemotherapy (Control vs. CDDP: 1.69 ± 0. 20 vs. 54. 62 ± 2. 49, P＜0. 01. Control vs. Sunitinib: 1.69 ± 0. 20vs. 47. 32 ± 2. 00, P＜0. 01. Control vs. CDDP + Sunitinib: 1.69 ± 0. 20 vs. 63. 41 ± 2. 23, P＜0. 01). Significantly differences of PI and AI were found between CDDP and CDDP + Sunitinib groups (P＜0. 01 ), and Sunitinib and CDDP + Sunitinib (P＜0. 01 ). RT-qPCR study confirmed that the mRNA expressions of M-CSFR, PDGFR-β, RET and VEGFR-2 were decreased in the mice underwent chemotherapy. Conclusions Sunitinib is effective to suppress the pediatric testicular yolk sac tumor growth, and reduce the tumor volume. Sunitinib can inhibit the angiogenesis in tumor, induce apoptosis of tumor cells, and kill the tumor cells directly. Sunitinib is less toxic than CDDP, and synergistic with the antitumor effect of CDDP.
