CIBZ, a Novel BTB Domain-Containing Protein, Is Involved in Mouse Spinal Cord Injury via Mitochondrial Pathway Independent of p53 Gene

Spinal cord injury (SCI) induces both primary uncontrollable mechanical injury and secondary controllable degeneration, which further results in the activation of cell death cascades that mediate delayed tissue damage. To alleviate its impairments and seek for an effective remedy, mRNA differential display was used to investigate gene mRNA expression profiling in mice following SCI. A specific Zinc finger and BTB domain-containing protein, CIBZ, was discovered to implicate in the SCI process for the first time. Further researches indicated that CIBZ was extensively distributed in various tissues, and the expression level was highest in muscle, followed by spinal cord, large intestine, kidney, spleen, thymus, lung, cerebrum, stomach, ovary and heart, respectively. After injury, the CIBZ expression decreased dramatically and reached the lowest level at 8 h, but it gradually increased to the maximal level at 7 d. Caspase-3 and C-terminal-binding protein (CtBP), two CIBZ-related proteins, showed similar tendency. Interestingly, p53 expression remained constant in all groups. Via flow cytometry (FCM) analysis, it was found that the cell death rate in SCI group markedly increased and reached the highest value 1 d after surgery and the mitochondrial transmembrane potential (ΔΨm) at 1 d was the lowest in all groups. Taken together, it is suggested that: (i) in the presence of CtBP, CIBZ gene is involved in secondary injury process and trigger the activation of apoptotic caspase-3 and bax genes independent of p53; (ii) abrupt down-regulation of CtBP at 8 h is a sign of mitochondria dysfunction and the onset of cell death; (iii) it could be used as an inhibitor or target drug of caspase-3 gene to improve spinal cord function.

脊髓损伤（SCI）可同时引发原发性不可控机械损伤与继发性可控退行性变，后者进一步激活介导迟发性组织损伤的细胞死亡级联反应。为缓解该病症的损伤效应并探寻有效治疗策略，本研究采用mRNA差异显示技术（mRNA differential display），探究了小鼠脊髓损伤后基因的mRNA表达谱。首次发现一种特定的含锌指和BTB结构域蛋白（Zinc finger and BTB domain-containing protein）CIBZ参与脊髓损伤进程。后续研究表明，CIBZ广泛分布于多种组织，其表达水平在肌肉中最高，随后依次为脊髓、大肠、肾脏、脾脏、胸腺、肺、大脑、胃、卵巢与心脏。损伤发生后，CIBZ的表达水平显著下调，并于损伤后8小时降至最低值，随后逐渐升高，至损伤后7天达到峰值。与CIBZ相关的两种蛋白——半胱天冬酶-3（caspase-3）与羧基末端结合蛋白（CtBP），呈现出相似的表达趋势。值得注意的是，所有组别中p53的表达水平均保持稳定。通过流式细胞术（FCM）分析发现，脊髓损伤组的细胞死亡率显著升高，并于术后1天达到峰值；同时，损伤后1天的线粒体跨膜电位（ΔΨm）为所有组别中最低。综上，本研究提示：（1）在羧基末端结合蛋白（CtBP）存在的条件下，CIBZ基因参与脊髓继发性损伤进程，且可不依赖p53通路激活凋亡相关的半胱天冬酶-3（caspase-3）与bax基因（bax）；（2）损伤后8小时羧基末端结合蛋白（CtBP）的急剧下调可作为线粒体功能障碍与细胞死亡启动的标志；（3）CIBZ可作为半胱天冬酶-3（caspase-3）基因的抑制剂或靶向药物，用于改善脊髓功能。