Shao Fangyu, Lv Mei, Zheng Yuanyuan, Jiang Junshu, Wang Yue, Lv Li, Wang Jihong
Department of Pharmacology, Dalian Medical University, Dalian, Liaoning Province, 116044, China.
Department of Otorhinolaryngology and Head and Neck Surgery, The First Affiliated Hospital of Dalian Medical University, Dalian, Liaoning, 116011, China.
Biochimie. 2015 Dec;119:183-91. doi: 10.1016/j.biochi.2015.11.004. Epub 2015 Nov 6.
The objective of this study is to investigate the antiproliferative activity and mechanism of integrin-binding rLj-RGD4 in a Hep-2 human laryngeal carcinoma-bearing nude mouse model.
Human laryngeal squamous carcinoma cells (Hep-2) were inoculated subcutaneously into the axilla of nude mice to generate a Hep-2 human laryngeal carcinoma-bearing nude mouse model. When the Hep-2 xenograft model was successfully established, the animals were randomly separated into five groups. Three groups were treated with different dosages of rLj-RGD4. Cisplatin was administered to the positive control group, and normal saline (NaCl) was administered to the negative control group for 3 weeks. The body weights and the survival of the nude mice were evaluated, and the volumes and weights of the solid tumours were measured. The mechanism underlying rLj-RGD4 inhibition of tumour growth in transplanted Hep-2 human laryngeal carcinoma-bearing nude mice was evaluated by haematoxylin-eosin (HE) staining, terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labelling (TUNEL), measurement of intratumoural microvessel density (MVD), Western blotting, and quantitative reverse transcription-polymerase chain reaction (qRT-PCR).
The tumour volumes and weights of the treatment groups were reduced compared with the model group, and survival times were improved by rLj-RGD4 treatment in Hep-2 human laryngeal carcinoma-bearing nude mice. The number of apoptotic Hep-2 human cells and intratumoural MVD significantly decreased after the administration of rLj-RGD4. In the xenograft tissue of animals treated with rLj-RGD4, FAK, PI3K, and Akt expression was unaltered, whereas P-FAK, P-PI3K, Bcl-2, P-Akt, and VEGF levels were down-regulated. In addition, activated caspase-3, activated caspase-9, and Bax levels were up-regulated.
rLj-RGD4 exhibits potent in vivo activity and inhibits the growth of transplanted Hep-2 human laryngeal carcinoma cells in a nude mouse model. Thus, these results indicate that the recombinant RGD toxin protein rLj-RGD4 may serve as a potent clinical therapy for human laryngeal squamous carcinoma.
本研究旨在探讨整合素结合型rLj-RGD4在荷人喉癌Hep-2裸鼠模型中的抗增殖活性及作用机制。
将人喉鳞状癌细胞（Hep-2）皮下接种于裸鼠腋窝，建立荷人喉癌Hep-2裸鼠模型。待Hep-2异种移植模型成功建立后，将动物随机分为五组。三组分别给予不同剂量的rLj-RGD4进行治疗。阳性对照组给予顺铂，阴性对照组给予生理盐水（NaCl），持续3周。评估裸鼠的体重和生存期，并测量实体瘤的体积和重量。通过苏木精-伊红（HE）染色、末端脱氧核苷酸转移酶介导的dUTP-生物素缺口末端标记（TUNEL）、肿瘤内微血管密度（MVD）测定、蛋白质免疫印迹法及定量逆转录-聚合酶链反应（qRT-PCR），评估rLj-RGD4抑制荷人喉癌Hep-2裸鼠移植瘤生长的机制。
与模型组相比，各治疗组的肿瘤体积和重量均减小，rLj-RGD4治疗可延长荷人喉癌Hep-2裸鼠的生存期。给予rLj-RGD4后，凋亡的人Hep-2细胞数量及肿瘤内MVD显著降低。在接受rLj-RGD4治疗的动物异种移植组织中，黏着斑激酶（FAK）、磷脂酰肌醇-3激酶（PI3K）和蛋白激酶B（Akt）的表达未发生改变，而磷酸化FAK（P-FAK）、磷酸化PI3K（P-PI3K）、B细胞淋巴瘤-2（Bcl-2）、磷酸化Akt（P-Akt）和血管内皮生长因子（VEGF）水平下调。此外，活化的半胱天冬酶-3、活化的半胱天冬酶-9及Bax水平上调。
rLj-RGD4在体内具有显著活性，可抑制荷人喉癌Hep-2细胞在裸鼠模型中的生长。因此，这些结果表明重组RGD毒素蛋白rLj-RGD4可能成为治疗人喉鳞状细胞癌的有效临床疗法。
