无血清培养法分离并鉴定胰腺癌细胞系BxPC-3中肿瘤干细胞

Isolation and Identification of Pancreatic Cancer Stem Cells from BxPC-3 Cell Line Cultured in Serum-Free Medium

[目的]从人胰腺癌细胞系BxPC-3中分离并鉴定胰腺癌干细胞。[方法]用无血清培养基培养人胰腺癌BxPC-3得到肿瘤细胞球。将肿瘤细胞球传代扩增,观察各传代后细胞成球能力;并用含血清培养基诱导促使其分化;蛋白印迹法测定肿瘤干细胞特异性转录因子OCT-4蛋白表达情况,并将肿瘤细胞球细胞植入NOD/SCID小鼠皮下,观察移植瘤的形成。[结果]在无血清培养基中,BxPC-3细胞能形成少量肿瘤细胞球,具有很强的自我更新能力,成球能力随着球体细胞传代次数的增加而增加。在含血清环境中,球体细胞逐渐分化而呈贴壁生长。肿瘤细胞球OCT-4蛋白表达明显高于BxPC-3贴壁细胞。1×104个球体细胞即能在NOD/SCID小鼠皮下成瘤。[结论]无血清培养能够有效的分离和扩增人胰腺癌细胞系BxPC-3的肿瘤干细胞。

[Purpose] To isolate and identify pancreatic cancer stem cells from human pancreatic cancer cell line BxPC-3. [Method]BxPC-3 cells were cultured in serum-free medium to derive tumor spheres. Tumor spheres were then expanded and passage to estimate the potency of sphere-forming,then cultured in serumcontaining medium to induce their differentiation. The protein expression of stem cell related transcript factor OCT-4 was evaluated by western blot. The tumorigenesis of the sphere-forming cells was observed in subcutaneously implanted NOD/SCID mice models. [Results] A few of floating tumor spheres were isolated and expanded in serum-free medium. These tumor spheres attached to the bottom of culture plates and began to differentiate when cultured in serum-containing medium. The protein expression of OCT-4 was significantly higher in sphere-forming cells of BxPC-3 than that attached cells. Only one thousand spheroid cells can form tumor in NOD/SCID mice. [Conclusion] Serum-free medium culture is an effective way to isolate and expand the pancreatic cancer stem cell from BxPC-3.