海马体C/EBPβ在糖尿病神经病理性疼痛中的作用及机制研究*

Role and Mechanism of Hippocampal C/EBPβ in Diabetic Neuropathic Pain*

摘要:

目的：探讨海马体中C/EBPβ在小鼠糖尿病神经病理性疼痛(DPNP)中的作用及机制。方法：本实验将小鼠随机分为4组：对照组(Control)、模型组 (DPNP)、注射阴性对照腺相关病毒组(DPNP+IE-control)、注射抑制C/EBPβ的腺相关病毒组(DPNP+IE-CEBP/β)。通过腹腔注射链脲佐菌素(STZ)建立小鼠的糖尿病周围神经病理性疼痛模型,并通过血糖升高及机械性痛觉过敏确认模型的成功建立。利用脑立体定位技术,向DPNP模型小鼠海马体注射携带C/EBPβ shRNA的腺相关病毒(AAV),特异性抑制C/EBPβ表达。检测机械性异常疼痛变化,使用免疫印迹检测海马体中C/EBPβ及NLRP3、NF-κB、Caspase-1、GSDMD和IL-1β等蛋白表达水平,利用RT-qPCR检测海马体中炎症因子IL-6、IL-1β和TNF-α的mRNA水平,以评估炎症反应。结果：注射AAV-shRNA后,海马体中C/EBPβ表达显著降低,机械性异常疼痛明显缓解,NLRP3、NF-κB、Caspase-1、GSDMD和IL-1β的蛋白表达水平下降,IL-6、IL-1β和TNF-α的mRNA水平显著减少,神经炎症减轻。结论：C/EBPβ在DPNP模型海马体中的高表达,通过调控炎症信号通路及神经炎症,参与DPNP的发生和维持。

Abstract:

Objective: This study aimed to explore the role and mechanism of C/EBPβ in the hippocampus in diabetic peripheral neuropathic pain (DPNP). Specifically, the study focused on how the suppression of C/EBPβ expression in the hippocampus could influence pain behaviors in a DPNP mouse model. Methods: In this experiment, mice were randomly divided into four groups: Control, DPNP model, DPNP+IE-Control (negative control adenovirus injection), and DPNP+IE-C/EBPβ (adenovirus injection to inhibit C/EBPβ expression). The diabetic peripheral neuropathic pain (DPNP) model was established by intraperitoneal injection of streptozotocin (STZ), and the model"s success was confirmed through elevated blood glucose levels and mechanical allodynia. Stereotaxic brain injection was used to administer an adeno-associated virus (AAV) carrying C/EBPβ shRNA into the hippocampus of DPNP mice, specifically inhibiting C/EBPβ expression. Mechanical allodynia was measured to evaluate changes in pain sensitivity. Western blotting was used to assess the expression levels of C/EBPβ and key inflammatory pathway proteins such as NLRP3, NF-κB, Caspase-1, GSDMD, and IL-1β in the hippocampus. Additionally, RT-qPCR was employed to quantify the mRNA levels of inflammatory cytokines IL-6, IL-1β, and TNF-α in the hippocampus, thereby evaluating the inflammatory response. Results: The results demonstrated that after AAV-shRNA injection, C/EBPβ expression in the hippocampus was significantly reduced, leading to a marked alleviation of mechanical pain in DPNP mice. Furthermore, the expression levels of NLRP3, NF-κB, Caspase-1, GSDMD, and IL-1β were significantly decreased in the hippocampus following AAV-shRNA treatment. A corresponding reduction in the mRNA levels of IL-6, IL-1β, and TNF-α was also observed, indicating a decrease in neuroinflammation. Conclusion: C/EBPβ was highly expressed in the hippocampus of a DPNP model and was involved in the development and maintenance of DPNP by modulating inflammatory signaling pathways and neuroinflammation.