Drug Resistance Group, Centre for Cancer Research and Cell Biology, Queen's University Belfast, Belfast, Northern Ireland.
PLoS One. 2011;6(5):e20276. doi: 10.1371/journal.pone.0020276. Epub 2011 May 24.
The role of the calcium binding protein, Calbindin 2 (CALB2), in regulating the response of colorectal cancer (CRC) cells to 5-Fluorouracil (5-FU) was investigated. Real-time RT-PCR and Western blot analysis revealed that CALB2 mRNA and protein expression were down-regulated in p53 wild-type and p53 null isogenic HCT116 CRC cell lines following 48 h and 72 h 5-FU treatment. Moreover, 5-FU-induced apoptosis was significantly reduced in HCT116 and LS174T CRC cell lines in which CALB2 expression had been silenced. Further investigation revealed that CALB2 translocated to the mitochondria following 5-FU treatment and that 5-FU-induced loss of mitochondrial membrane potential (Δψ(m)) was abrogated in CALB2-silenced cells. Furthermore, CALB2 silencing decreased 5-FU-induced cytochrome c and smac release from the mitochondria and also decreased 5-FU-induced activation of caspases 9 and 3/7. Of note, co-silencing of XIAP overcame 5-FU resistance in CALB2-silenced cells. Collectively, these results suggest that following 5-FU treatment in CRC cell lines, CALB2 is involved in apoptosis induction through the intrinsic mitochondrial pathway. This indicates that CALB2 may be an important mediator of 5-FU-induced cell death. Moreover, down-regulation of CALB2 in response to 5-FU may represent an intrinsic mechanism of resistance to this anti-cancer drug.
钙结合蛋白 Calbindin 2（CALB2）在调节结直肠癌细胞（CRC）对 5-氟尿嘧啶（5-FU）的反应中的作用进行了研究。实时 RT-PCR 和 Western blot 分析显示，在 p53 野生型和 p53 缺失同基因 HCT116 CRC 细胞系中，CALB2 mRNA 和蛋白表达在 5-FU 处理 48 h 和 72 h 后下调。此外，在 CALB2 表达沉默的 HCT116 和 LS174T CRC 细胞系中，5-FU 诱导的细胞凋亡明显减少。进一步研究表明，CALB2 在 5-FU 处理后转移到线粒体，并且在 CALB2 沉默的细胞中，5-FU 诱导的线粒体膜电位（Δψ(m)）丧失被阻断。此外，CALB2 沉默减少了 5-FU 诱导的细胞色素 c 和 smac 从线粒体释放，并减少了 5-FU 诱导的 caspase 9 和 3/7 的激活。值得注意的是，XIAP 的共沉默克服了 CALB2 沉默细胞对 5-FU 的耐药性。总之，这些结果表明，在 CRC 细胞系中经 5-FU 处理后，CALB2 通过内在的线粒体途径参与凋亡诱导。这表明 CALB2 可能是 5-FU 诱导细胞死亡的重要介质。此外，CALB2 对 5-FU 的下调可能代表对这种抗癌药物的内在耐药机制。
