Jaffa Aneese A, Jaffa Miran A, Moussa Mayssam, Ahmed Ibrahim A, Karam Mia, Aldeen Kawthar Sharaf, Al Sayegh Rola, El-Achkar Ghewa A, Nasrallah Leila, Yehya Yara, Habib Aida, Ziyadeh Fuad N, Eid Ali H, Kobeissy Firas H, Jaffa Ayad A
Department of Biology, Faculty of Arts and Sciences, American University of Beirut, Beirut, Lebanon.
Epidemiology and Population Health Department, Faculty of Health Sciences, American University of Beirut, Beirut, Lebanon.
Front Pharmacol. 2021 Nov 15;12:743059. doi: 10.3389/fphar.2021.743059. eCollection 2021.
Microglia, the resident phagocytes of the central nervous system and one of the key modulators of the innate immune system, have been shown to play a major role in brain insults. Upon activation in response to neuroinflammation, microglia promote the release of inflammatory mediators as well as promote phagocytosis. Plasma prekallikrein (PKall) has been recently implicated as a mediator of neuroinflammation; nevertheless, its role in mediating microglial activation has not been investigated yet. In the current study, we evaluate the mechanisms through which PKall contributes to microglial activation and release of inflammatory cytokines assessing PKall-related receptors and their dynamics. Murine N9-microglial cells were exposed to PKall (2.5 ng/ml), lipopolysaccharide (100 ng/ml), bradykinin (BK, 0.1 μM), and neuronal cell debris (16.5 μg protein/ml). Gene expression of bradykinin 2 receptor (BKR), protease-activated receptor 2 (PAR-2), along with cytokines and fibrotic mediators were studied. Bioinformatic analysis was conducted to correlate altered protein changes with microglial activation. To assess receptor dynamics, HOE-140 (1 μM) and GB-83 (2 μM) were used to antagonize the BKR and PAR-2 receptors, respectively. Also, the role of autophagy in modulating microglial response was evaluated. Data from our work indicate that PKall, LPS, BK, and neuronal cell debris resulted in the activation of microglia and enhanced expression/secretion of inflammatory mediators. Elevated increase in inflammatory mediators was attenuated in the presence of HOE-140 and GB-83, implicating the engagement of these receptors in the activation process coupled with an increase in the expression of BKR and PAR-2. Finally, the inhibition of autophagy significantly enhanced the release of the cytokine IL-6 which were validated via bioinformatics analysis demonstrating the role of PKall in systematic and brain inflammatory processes. Taken together, we demonstrated that PKall can modulate microglial activation via the engagement of PAR-2 and BKR where PKall acts as a neuromodulator of inflammatory processes.
小胶质细胞是中枢神经系统的常驻吞噬细胞，也是先天免疫系统的关键调节因子之一，已被证明在脑损伤中起主要作用。在响应神经炎症而被激活后，小胶质细胞会促进炎症介质的释放并促进吞噬作用。血浆前激肽释放酶（PKall）最近被认为是神经炎症的介质；然而，其在介导小胶质细胞激活中的作用尚未得到研究。在本研究中，我们评估了PKall通过评估PKall相关受体及其动态变化来促进小胶质细胞激活和炎性细胞因子释放的机制。将小鼠N9小胶质细胞暴露于PKall（2.5 ng/ml）、脂多糖（100 ng/ml）、缓激肽（BK，0.1 μM）和神经元细胞碎片（16.5 μg蛋白/ml）。研究了缓激肽2受体（BKR）、蛋白酶激活受体2（PAR-2）以及细胞因子和纤维化介质的基因表达。进行生物信息学分析以将改变的蛋白质变化与小胶质细胞激活相关联。为了评估受体动态变化，分别使用HOE-140（1 μM）和GB-83（2 μM）拮抗BKR和PAR-2受体。此外，还评估了自噬在调节小胶质细胞反应中的作用。我们的工作数据表明，PKall、LPS、BK和神经元细胞碎片导致小胶质细胞激活，并增强了炎症介质的表达/分泌。在存在HOE-140和GB-83的情况下，炎症介质的升高增加有所减弱，这表明这些受体参与了激活过程，同时BKR和PAR-2的表达增加。最后，自噬的抑制显著增强了细胞因子IL-6的释放，这通过生物信息学分析得到验证，证明了PKall在全身和脑部炎症过程中的作用。综上所述，我们证明PKall可以通过PAR-2和BKR的参与来调节小胶质细胞激活，其中PKall作为炎症过程的神经调节剂。
