Epilepsy, one of the most common neurological disorders, is characterized by spontaneous recurrent seizures. Temporal lobe epilepsy (TLE) is one of the most common medically intractable seizure disorders. Traf2-and NcK-interacting kinase (TNIK) has recently attracted attention as a critical modulation target of many neurological and psychiatric disorders, but its role in epilepsy remains unclear. In this study, we hypothesized the involvement of TNIK in epilepsy and investigated TNIK expression in patients with intractable TLE and in a pilocarpine-induced rat model of epilepsy by western blotting, immunofluorescence, and immunohistochemistry. A pentylenetetrazole (PTZ)-induced epilepsy rat model was used to determine the effect of the TNIK inhibitor NCB-0846 on behavioral manifestations of epilepsy. Coimmunoprecipitation (Co-IP)/mass spectrometry (MS) was used to identify the potential mechanism. Through Co-IP, we detected and confirmed the main potential TNIK interactors. Subcellular fractionation was used to establish the effect of NCB-0846 on the expression of the main interactors in postsynaptic density (PSD) fractions. We found that TNIK was primarily located in neurons and decreased significantly in epilepsy model rats and TLE patients compared with controls. NCB-0846 delayed kindling progression and decreased seizure severity. Co-IP/MS identified 63 candidate TNIK interactors in rat hippocampi, notably CaMKII. Co-IP showed that TNIK might correlate with endogenous GRIA1, SYN2, PSD-95, CaMKIV, GABRG1, and GABRG2. In addition, the significant decrease in GRIA1 in hippocampal total lysate and PSDs after NCB-0846 treatment might help modify the progression of PTZ kindling. Our results suggest that TNIK contributes to epileptic pathology and is a potential antiepileptic drug target.

癫痫是最常见的神经系统疾病之一，其特征是自发性反复发作。颞叶癫痫 （TLE） 是最常见的医学难治性癫痫发作疾病之一。Traf2 和 NcK 相互作用激酶 （TNIK） 作为许多神经和精神疾病的关键调节靶点最近引起了人们的关注，但其在癫痫中的作用仍不清楚。在这项研究中，我们假设了 TNIK 与癫痫的关系，并通过蛋白质印迹、免疫荧光和免疫组织化学研究了 TNIK 在难治性 TLE 患者和毛果芸香碱诱导的大鼠癫痫模型中的表达。采用戊四唑 （PTZ） 诱导的癫痫大鼠模型测定 TNIK 抑制剂 NCB-0846 对癫痫行为表现的影响。采用免疫共沉淀 （Co-IP）/质谱 （MS） 确定潜在机制。通过 Co-IP，我们检测并确认了主要的潜在 TNIK 交互者。亚细胞组分分离用于确定 NCB-0846 对突触后致密 （PSD） 组分中主要相互作用物表达的影响。我们发现 TNIK 主要位于神经元中，与对照组相比，癫痫模型大鼠和 TLE 患者显著降低。NCB-0846 延缓了点燃进展并降低了癫痫发作的严重程度。Co-IP/MS 在大鼠海马体中鉴定了 63 个候选 TNIK 相互作用器，尤其是 CaMKII。Co-IP 显示 TNIK 可能与内源性 GRIA1 、 SYN2 、 PSD-95 、 CaMKIV 、 GABRG1 和 GABRG2 相关。此外，NCB-0846 处理后海马总裂解物和 PSD 中 GRIA1 的显着降低可能有助于改变 PTZ 点燃的进展。 我们的结果表明，TNIK 有助于癫痫病理学，并且是潜在的抗癫痫药物靶点。