Spinal cord genes enriched in rat dorsal horn and induced by noxious stimulation identified by subtraction cloning and differential hybridization.
Spinal cord genes enriched in rat dorsal horn and induced by noxious stimulation identified by subtraction cloning and differential hybridization.
复制标题
通过消减克隆和差异杂交鉴定出大鼠背角中富含并由有害刺激诱导的脊髓基因。
DOI：
10.1016/s0306-4522(00)00573-x
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发表时间：
2001
期刊：
影响因子：
3.3
通讯作者：
Iadarola,MJ
中科院分区：
文献类型：
--
作者：
Yang,HY;Wilkening,S;Iadarola,MJ
持续的伤害性输入会增加神经元的兴奋性并诱导背侧脊髓的基因表达程序。基因表达的改变从立即早期基因的磷酸化和诱导开始，并继续到靶基因。目前仅确定了少数靶基因。本报告使用基于聚合酶链反应的消减克隆程序来获得“解剖学聚焦”的互补DNA文库，该文库富含与感觉脊髓（大鼠背角减去腹角）相关的转录本。筛选该文库中的克隆子集 (n=158)，以验证背角富集并鉴定受角叉菜胶诱导的外周炎症调节的克隆。显示出丰富表达的分子类别包括以前与感觉过程不相关的原癌基因、控制细胞形状的 Rho/Rac 途径的两个调节因子以及涉及细胞骨架调节和支架的三个基因。编码参与细胞间通讯或细胞内功能的蛋白质的其他转录本。在 158 个转录本中，持续的有害输入诱导了一个已知基因和两个未知基因。已知的基因编码分泌型半胱氨酸蛋白酶抑制剂胱抑素 C，表明细胞外蛋白水解活性发生调节。由于胱抑素 C 是分泌性的，因此它还可以为持续性疼痛状态提供脑脊液生物标志物。利用解剖学和功能相结合的方法，我们在几个新的方向上扩展了二阶神经元或支持神经胶质细胞中表达和诱导的基因的分子库，特别强调细胞形态和质膜动力学的调节。其中一些蛋白质揭示了脊髓感觉半部信息信号传导的新途径，需要进一步研究以了解它们在成人脊髓中的作用。诱导的基因可能为治疗开发提供新的分子靶标，并为研究持续疼痛状态期间发生的细胞活动的动态状态提供新的探针。
Persistent nociceptive input increases neuronal excitability and induces a program of gene expression in the dorsal spinal cord. The alteration in gene expression commences with phosphorylation and induction of immediate early genes and proceeds to target genes. Only a few target genes have been identified as yet. The present report uses a polymerase chain reaction-based subtraction cloning procedure to obtain an “anatomically focused” complementary DNA library enriched in transcripts related to sensory spinal cord (rat dorsal horn minus ventral horn). A subset of clones from this library (n=158) was screened to verify dorsal horn enrichment and to identify those regulated by carrageenan-induced peripheral inflammation. Molecular classes which displayed enriched expression included a proto-oncogene not previously associated with sensory processes, two regulators of the Rho/Rac pathway which controls cell shape, and three genes involved in cytoskeletal regulation and scaffolding. Additional transcripts coded for proteins involved in intercellular communication or intracellular function. Within the set of 158 transcripts, one known and two unknown genes were induced by persistent noxious input. The known gene codes for the secreted cysteine proteinase inhibitor, cystatin C, suggesting that modulation of extracellular proteolytic activity occurs. Since it is secreted, cystatin C may also provide a cerebrospinal fluid bio-marker for persistent pain states. Using a combined anatomical and functional approach, we have extended the molecular repertoire of genes expressed and induced in second-order neurons or supporting glial cells in several new directions, with particular emphasis on regulation of cell morphology and plasma membrane dynamics. Some of these proteins reveal new pathways for information signaling in the sensory half of the spinal cord and require further research to understand their role in the adult spinal cord. The induced genes may provide new molecular targets for therapeutic development and provide new probes for investigating the dynamic state of cellular activity that occurs during persistent pain states.