Abstract：
Toll-like receptors (TLRs) are activated by endogenous molecules released from damaged cells and contribute to neuroinflammation following traumatic brain injury (TBI) and epilepsy. TLR1/2 agonist tri-palmitoyl-S-glyceryl-cysteine (Pam3cys) is a vaccine adjuvant with confirmed safety in humans. We assessed impact of TLR1/2 postconditioning by Pam3cys on epileptogenesis and neuroinflammation in male rats, 6, 24, and 48 h after mild-to-moderate TBI. Pam3cys was injected into cerebral ventricles 30 min after controlled cortical impact (CCI) injury. After 24 h, rats underwent chemical kindling by once every other day injections of pentylenetetrazole (PTZ) 35 mg/kg until development of generalized seizures. Number of intact neurons, brain expression of proinflammatory cytokine TNF-α, anti-inflammatory cytokine IL-10, and marker of anti-inflammatory microglia arginase1 (Arg1) were determined by immunoblotting. Astrocytes and macrophage/microglia activation/polarization at the contused area was assessed by double immunostaining with Iba1/Arg1, Iba1/iNOS and GFAP/iNOS, specific antibodies. The CCI-injured rats became kindled by less number of PTZ injections than sham-operated rats (9 versus 14 injections, p < 0.0001). Pam3cys treatment returned the accelerated rate of epileptogenesis in TBI state to the sham level. Pam3cys decreased neural death 48 h after TBI. It decreased TNF-α (6 h post-TBI, p < 0.01), and up-regulated IL-10 (p < 0.01) and Arg1 (p < 0.05) 48 h after TBI. The iNOS-positive cells decreased (p < 0.001) whereas Iba1/Arg1-positive cells enhanced (p < 0.01) after Pam3cys treatment. Pam3cys inhibits TBI-accelerated acquisition of seizures. Pam3cys reprograms microglia and up-regulates anti-inflammatory cytokines during the first few days after TBI. This capacity along with the clinical safety, makes Pam3cys a potential candidate for development of effective medications against posttraumatic epilepsy.
摘要：
Toll样受体（TLR）被受损细胞释放的内源性分子激活，并导致创伤性脑损伤（TBI）和癫痫后的神经炎症。TLR1/2激动剂三棕榈酰-S-甘油基-半胱氨酸（Pam3cys）是一种在人体中具有确认安全性的疫苗佐剂。我们评估了通过Pam3cys进行TLR1/2后处理对雄性大鼠癫痫发生和神经炎症的影响，轻度至中度TBI后6、24和48小时。控制性皮质撞击（CCI）损伤后30分钟，将Pam3cys注入脑室。24小时后，通过每隔一天一次注射戊四唑（PTZ）35mg/kg，对大鼠进行化学点燃，直到出现全身性癫痫发作。完整神经元的数量，脑前炎症细胞因子TNF-α的表达，通过免疫印迹测定抗炎细胞因子IL-10和抗炎小胶质细胞精氨酸酶1（Arg1）的标志物。通过Iba1/Arg1，Iba1/iNOS和GFAP/iNOS的双重免疫染色评估了挫伤区域的星形胶质细胞和巨噬细胞/小胶质细胞活化/极化，特异性抗体。与假手术大鼠相比，受CCI损伤的大鼠被较少的PTZ注射点燃（9对14次注射，p<0.0001)。Pam3cys治疗使TBI状态下癫痫发生的加速速率恢复到假手术水平。Pam3cys减少TBI后48小时的神经死亡。它降低了TNF-α（TBI后6小时，p<0.01),并在TBI后48小时上调IL-10(p<0.01)和Arg1(p<0.05)。Pam3cys处理后，iNOS阳性细胞减少（p<0.001），而Iba1/Arg1阳性细胞增强（p<0.01）。Pam3cys抑制TBI加速的癫痫发作。Pam3cys在TBI后的最初几天重新编程小胶质细胞并上调抗炎细胞因子。这种能力以及临床安全性，使Pam3cys成为开发针对创伤后癫痫的有效药物的潜在候选者。
