Significance of transforming growth factors for ocular malignant tumors and their application in gene therapy.

本研究的目的是阐明转化生长因子（TGF）在眼部恶性肿瘤中的作用，并将其应用于基因治疗。TGF信号传导是引发Sonic hedgehog（Shh）效应所必需的。环巴胺是一种天然化合物，能够抑制Smoothened活性，这在Shh/Gli信号激活中至关重要。1.将9，10-二甲基-1，2-苯并蒽和佛波醇-12-肉豆蔻酸酯-13-乙酸酯的化学致癌物混合物局部施加于XPC-裸小鼠的眼睑，每两天持续2至8个月，并且在眼睑中产生上皮肿瘤。每天皮下摄入环巴胺治疗这些肿瘤，持续7天。组织学显示，环磷酰胺治疗抑制了BrdU掺入，并诱导了这些肿瘤中大部分细胞的凋亡.使用从人烧伤瘢痕衍生的鳞状细胞癌建立的鳞状细胞癌细胞系。将细胞维持在Dulbecco的MEM中。通过添加重组Shh来增强细胞，并且通过添加环巴胺来消除这种作用。然而，在重组Shh的作用下，加入cyclopamine对SCC细胞系的增殖没有影响。用注射针将SCC细胞悬液接种于裸鼠背部皮下。植入后10天，动物每两天接受一次环巴胺全身治疗，共三次。SCC细胞在裸鼠体内形成的肿瘤中表达Shh，环巴胺抑制Shh信号通路可诱导肿瘤细胞凋亡并抑制肿瘤细胞增殖。环巴胺在体内抑制上皮性肿瘤细胞增殖并诱导凋亡。Hedgehog通路可能是眼睑肿瘤患者潜在的治疗靶点。

The purpose of this study is to clarify the role of transforming growth factor (TGF) for ocular malignant tumors and to apply them in gene therapy. TGF signaling is required for eliciting the effects of Sonic hedgehog (Shh). Cyclopamine is a natural compound that is capable of suppression of Smoothened activity which is critical in Shh/Gli signal activation.1. Chemical carcinogen cocktail of 9,10-dimethyl-1,2-benzanthracene and phorbol-12-myristate-13-acetate was topically applied to eyelid of XPC-null mice every two days for 2 to 8 months, and epithelial tumors developed in eyelid. These tumors were treated daily with subcutaneous ingestion of cyclopamine for 7 days. Histology showed that cylopamine treatment suppressed BrdU incorporation and induced apoptosis in the majority of cells in these tumors.2. Squamous cell carcinoma cell line established from human burned-scar-derived SCC was used. The cells were maintained in Dulbecco's MEM. The cells were enhanced by adding recombinant Shh and this effect was abolished by adding cyclopamine. However, the proliferation of SCC cell line was not affected by adding cylopamine in the abscence of recombinant Shh.3. Cell suspension of SCC cell line was implanted subcutaneously by using a injection needle in the back of nude mice. Ten days after implantation, the animals were treated systemic cyclopamine at every two days for three times. The SCC cells express Shh in vivo in tumor developed in nude mice and inhibition of Shh signaling pathway by cyclopamine induced apoptosis and suppressed proliferation in the tumor. Cyclopamine inhibits proliferation and induces apoptosis in epithelial tumor cells in vivo. The hedgehog pathway may be a potential therapeutic target for patients with eyelid tumors.