不同接种部位肿瘤进行激光共聚焦显微镜观察的可行性

Feasibility of laser confocal microscope observation of tumors at different inoculation sites

目的比较用小鼠腹部皮下瘤与耳部皮下瘤进行激光共聚焦显微镜原位观察药物瘤内微观行为方法的可行性与适用性,为研究原位观察肿瘤内药物渗透提供更好的方法。方法体外培养转染绿色荧光蛋白(GFP)的小鼠乳腺癌细胞4T1(4T1-GFP)。将4T1-GFP细胞以皮下注射的方式分别接种到6~8周龄SPF级雌性ICR小鼠的腹部皮下静脉旁或耳部静脉旁。待肿瘤长到4~7mm3大小即对小鼠实施手术,将肿瘤粘贴于载玻片上。通过尾静脉将带有Cy5荧光标记的肿瘤纳米药物注射到小鼠体内,用激光共聚焦显微镜对肿瘤药物渗透过程进行程序性图像采集。结果耳部肿瘤细胞最适接种浓度在(2~3)×10^4/μl,从第2天开始可用于成像;腹部肿瘤细胞适宜的接种浓度在(1~2)×10^4/μl,从第3天开始可用于成像。呈圆盘状包绕血管生长、血管分支较少的肿瘤更有利于进行肿瘤药物血管渗透成像。耳部和腹部皮下瘤模型均可较好呈现药物的渗透过程。结论耳部与腹部皮下瘤模型在药物血管渗透效果上各有优劣,这两种模型都为肿瘤药物血管渗透过程的研究提供了新方法。

Objective To compare the feasibility and applicability of laser confocal microscoe for in situ observation of drug intratumoral micro-behavior in mice with abdominal subcutaneous tumors and ear subcutaneous tumors, so as to provide a better method for in situ observation of drug penetration in tumors. Methods The murine breast cancer cell line 4 T1 transfected with green fluorescent protein(GFP)(4 T1-GFP) was cultured in vitro. 4 T1-GFP cells were subcutaneously injected into the abdominal subcutaneous vein or ear vein of female ICR mice aged 6-8 weeks. When the tumors grow to 4-7 mm^3, the mice would be operated on and the tumors will be pasted on the slide. Cy5-labeled nanodrugs were injected into mice via tail vein, and programmed image acquisition was performed by confocal laser microscope. Results The optimal concentration of tumor cells was(2-3)×10~4/μl for ear tumor inoculation, the tumors could be used for experiments from the second day. And the optimal concentration of tumor cells was(1-2) ×10~4/μl for abdominal subcutaneous inoculation, the tumors could be used for experiments from the third day. Tumors with less vascular branches and disc-like growth were more conducive to drug-mediated angiosmosis imaging. The model of ear and abdominal subcutaneous tumors could well show the process of drug penetration. Conclusion The models of ear and abdominal subcutaneous tumors have their own advantages and disadvantages in drug vascularization. Both models provide a new method for the study of drug vascularization in tumors.