β1受体阻滞剂通过调节G蛋白信号通路对感染性休克免疫失衡的作用及机制研究

Extensive pro-inflammatory immune responses and immune suppression result in organ injury in septic shock (SS). Sympathetic nervous system (SNS) is persistently over activated in septic shock. Our previous results found that using β1 blocker (BB1) to modulate SNS at the level of receptors could decrease simultaneously the levels of pro- and anti-inflammatory cytokines, reduce immune imbalance and improve cardiovascular function as well as survival in SS. However, the mechanisms of immune re-balance by BB1 remain unclear. It was reported that β1 receptor can modulate inflammatory response through G-protein pathway, Furthermore, β1 receptors are expressed on immune cells. So, we hypothesize that BB1 reduces immune imbalance by regulating monocyte secretion, lymphocyte apoptosis and Th0 cell differentiation through G-protein pathway. This project is to determine, by using wild type and β1 receptor knock out models, if different doses of BB1 inhibit over secretion of pro-inflammatory cytokines by monocytes through decreasing NF-κB activation, decrease lymphocyte apoptosis by increasing Bcl-2 activation, and reduce differentiation of Th0 to Th2 by decreasing STAT6 activation from animal, cell and protein levels, thereby reducing extensive pro-inflammatory immune responses and immune suppression in SS. Thereafter a clinical trial will be performed to confirm the results. The results of this study will provide a theoretical basis and dose recommendation for using β1-blocker as an adjuvant treatment in SS.

在感染性休克（SS）中，过度促炎反应和免疫抑制导致器官功能受损。在SS中交感神经系统（SNS）持续过度激活。我们前期研究发现用β1受体阻滞剂（BB1）在受体水平调节SNS，可同时降低SS中促炎和抑炎因子水平，减轻免疫失衡，改善心血管功能及预后。但BB1调节免疫失衡的机制尚不清楚。已有报道β1受体可通过G蛋白信号途径调节炎症反应，且免疫细胞均表达β1受体。因此，我们假设BB1通过调控G蛋白信号途径调节单核细胞分泌功能、淋巴细胞凋亡和Th0分化来减轻免疫失衡。我们拟采用正常和β1受体基因敲除模型，从动物、细胞和蛋白水平探究不同剂量的BB1是否通过抑制G蛋白活性，减少NF-κB激活抑制单核细胞过度分泌促炎因子、增加Bcl-2活性减少淋巴细胞凋亡、减少STAT6激活抑制Th0向Th2分化，从而改善SS中过度促炎反应和免疫抑制。最后进行临床试验，为BB1辅助治疗感染性休克提供理论依据及剂量参考。

持续免疫抑制是感染性休克（SS）患者多器官功能障碍甚至死亡的主要原因。我们研究证实，感染性休克大鼠给予低剂量艾司洛尔治疗后，其心血管功能得到明显改善，全身及心血管组织炎症反应降低，提示艾司洛尔可能改善感染性休克免疫失衡。我们前期研究发现，（1）感染性休克大鼠外周血淋巴细胞和单核细胞大量凋亡，Th2反应偏倚加剧；（2）低剂量艾司洛尔治疗可改善感染性休克大鼠外周血T淋巴细胞凋亡和Th2/Th1失衡。另有研究发现β1受体可通过G蛋白信号途径调节炎症反应，且免疫细胞表达β1受体。据此假设，β1受体阻滞剂通过G蛋白信号途径参与改善感染性休克免疫失衡。本项目研究证实，（1）低剂量艾司洛尔治疗可改善感染性休克大鼠平均动脉压和多器官损伤；（2）感染性休克大鼠T淋巴细胞AKT磷酸化表达水平较假手术组（Sham）大鼠下降，线粒体凋亡关键分子Bcl-2表达减少，Cleaved Caspase-3表达增加，ERK1/2磷酸化水平表达升高。（3）与感染性休克大鼠比较，低剂量艾司洛尔治疗的感染性休克大鼠T淋巴细胞AKT磷酸化表达水平升高，ERK1/2磷酸化水平表达降低，而Bcl-2表达增加，Cleaved Caspase-3表达减少。据此推论，艾司洛尔通过PI3K/AKT和Erk1/2信号通路参与改善感染性休克T淋巴细胞凋亡。上述研究结果为今后临床使用β1受体阻滞剂辅助治疗感染性休克进一步提供理论依据。