Rat NaV1.7 loss-of-function genetic model: Deficient nociceptive and neuropathic pain behavior with retained olfactory function and intra-epidermal nerve fibers
Rat NaV1.7 loss-of-function genetic model: Deficient nociceptive and neuropathic pain behavior with retained olfactory function and intra-epidermal nerve fibers
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DOI：
10.1177/1744806919881846
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发表时间：
2019-10-01
期刊：
影响因子：
3.3
通讯作者：
Gingras, J.
中科院分区：
文献类型：
--
作者：
Grubinska, B.;Chen, L.;Gingras, J.
使用遗传动物模型重述人类疾病病理生理学是一种强有力的方法，使机制的理解基因型-表型的关系，用于药物开发。Na（V）1.7是在外周神经系统中表达的钠通道，具有作为疼痛靶点的强人类遗传验证。努力确定新的镇痛药，是非成瘾性导致行业探索一类磺酰胺化合物结合的第四个电压传感器域的Na（V）1.7。由于该区域的序列差异，磺胺类阻滞剂通常对人Na（V）1.7通道有效，但对大鼠Na（V）1.7通道无效。为了在大鼠疼痛模型中测试基于磺酰胺的化学物质，我们产生了表达嵌合Na（V）1.7蛋白的人源化Na（V）1.7大鼠，所述嵌合Na（V）1.7蛋白含有人基因序列的磺酰胺结合位点作为等效大鼠序列的替代。出乎意料的是，在转录时，人类插入片段被剪接掉，导致过早的终止密码子。使用经验证的抗体，证实Na（V）1.7蛋白在脑干、背根神经节、坐骨神经和胃肠道组织中丢失，但在敲入等位基因（HOM-KI）纯合大鼠的鼻甲或嗅球中未丢失。HOM-KI大鼠表现出正常的表皮内神经纤维密度，小直径背根神经节神经元的河豚毒素敏感电流密度和动作电位放电降低。HOM-KI大鼠在热板或辣椒素诱导的退缩试验中没有表现出伤害性疼痛反应，并且在脊神经结扎后没有表现出神经性疼痛反应。与嗅觉组织中嵌合Na（V）1.7的表达一致，HOM-KI大鼠保留了嗅觉功能。这种新的遗传模型强调了Na（V）1.7对大鼠疼痛行为的必要性，并表明在人类中充分抑制Na（V）1.7可能会减少神经性疾病的疼痛。由于保留了嗅觉功能，该大鼠模型代表了在出生后早期发育期间需要时间密集型人工喂养的全局Na（V）1.7敲除小鼠的替代方案。
Recapitulating human disease pathophysiology using genetic animal models is a powerful approach to enable mechanistic understanding of genotype-phenotype relationships for drug development. Na(V)1.7 is a sodium channel expressed in the peripheral nervous system with strong human genetic validation as a pain target. Efforts to identify novel analgesics that are nonaddictive resulted in industry exploration of a class of sulfonamide compounds that bind to the fourth voltage-sensor domain of Na(V)1.7. Due to sequence differences in this region, sulfonamide blockers generally are potent on human but not rat Na(V)1.7 channels. To test sulfonamide-based chemical matter in rat models of pain, we generated a humanized Na(V)1.7 rat expressing a chimeric Na(V)1.7 protein containing the sulfonamide-binding site of the human gene sequence as a replacement for the equivalent rat sequence. Unexpectedly, upon transcription, the human insert was spliced out, resulting in a premature stop codon. Using a validated antibody, Na(V)1.7 protein was confirmed to be lost in the brainstem, dorsal root ganglia, sciatic nerve, and gastrointestinal tissue but not in nasal turbinates or olfactory bulb in rats homozygous for the knock-in allele (HOM-KI). HOM-KI rats exhibited normal intraepidermal nerve fiber density with reduced tetrodotoxin-sensitive current density and action potential firing in small diameter dorsal root ganglia neurons. HOM-KI rats did not exhibit nociceptive pain responses in hot plate or capsaicin-induced flinching assays and did not exhibit neuropathic pain responses following spinal nerve ligation. Consistent with expression of chimeric Na(V)1.7 in olfactory tissue, HOM-KI rats retained olfactory function. This new genetic model highlights the necessity of Na(V)1.7 for pain behavior in rats and indicates that sufficient inhibition of Na(V)1.7 in humans may reduce pain in neuropathic conditions. Due to preserved olfactory function, this rat model represents an alternative to global Na(V)1.7 knockout mice that require time-intensive hand feeding during early postnatal development.