基于“肺肾同源”理论探讨肾IRI时HGMB1/TLR4信号调控AQP-1的机制及发酵虫草菌粉的干预作用

Acute kidney injury (AKI) is critical disease in nephropathy department. The theory of same source both lung and kidney in Chinese Traditional medicine demonstrates the regulation of Lung and kidney with mother-child relationship on both the "water" and "gas", which is tha same as the understanding on the across-talking in AKI-induced ALI.And complicated acute lung injury (ALI) responses to the bad prognosis in AKI. Therefore, understanding the pathophysiological mechanisn of AKI-induced ALI is vital. Our previous study have difined that IRI could induce inflammation, Cordyceps sinensis provide a role of blocking the Notch signaling , inhibiting inflammtion meditor ,supressing oxide and reducing apoptosis. In this study, based the breakthrough points including - High-mobility group box 1/ Toll-like receptor 4 (HGMB1/TLR-4) siganling,AQP-1,and the material basis on the theory of same source both lung and kidney, we observe the role of later inflammation mediator HGMB1/TLR-4 siganling on the regulation of Aquaporin-1 expression in the across-talking between kidney and lung following IRI by means of Terminal-deoxynucleoitidyl Transferase Mediated Nick End Labeling, Immune confocal laser, real-timequantitativepolymerasechainreaction and western blot, and the nephroprotective mechanism of Cordyceps sinensis, trying to demonstrating the signaling mechanism of across-talking between kidney and lung and the material basis in the theory of same source both lung and kidney in Chinese Traditional medicine to improve the prognosis of AKI , and to provide the theory evidence for the clinical practice in Cordyceps sinensis.

急性肾损伤（AKI）是肾内科危急重症。中医学 "肺肾同源"理论阐述肺肾母子对"水"和"气"的调节，这与AKI肾肺存在复杂across-talking认识一致,并发急性肺损伤（ALI）为AKI预后差的关键。因此，理解AKI诱发ALI的信号机制至关重要。前期研究证实肾缺血再灌注损伤（IRI）诱发炎症反应，虫草菌粉通过阻断Notch信号通路、抗炎、抗氧化和抗凋亡发挥肾保护作用。本课题以HGMB1/TLR-4信号、水通道蛋白-1（AQP-1）和"肺肾同源"物质基础为切入点，通过TUNEL、免疫激光共聚焦、实时荧光定量PCR、western blot观察IRI时晚期炎症信号HGMB1/TLR-4调控AQP-1表达在肾肺across-talking的作用，以及发酵虫草菌粉肺肾保护作用的机制，以阐明AKI肾肺同病的信号机制及"肺肾同源"理论的物质基础，以期改善AKI预后，为虫草菌粉临床应用提供理论依据。

急性肾损伤（AKI）是肾内科危急重症。中医学“肺肾同源”理论阐述肺肾母子对“水”和“气”的调节，这与 AKI 肾肺存在复杂 across-talking 认识一致,并发急性肺损伤（ALI）为 AKI 预后差的关键。因此，理解 AKI 诱发 ALI 的信号机制至关重要。本课题以缺血-再灌注损伤大鼠和体外缺氧-复氧的肾小管上皮细胞模型为研究对象。我们的体内外的研究结果表明，肾IRI后大鼠血清BUN和Scr水平升高，肾小管出现坏死，肺组织的湿干重比升高，肺组织出现气管上皮细胞变性坏死，肺间质炎细胞浸润，肺泡间质水肿形成，而且可见肺泡出血，ALI发生时间晚于AKI，提示肾IRI导致的AKI可诱发ALI。进一步的研究发现，肾IRI可诱导大鼠血清、肺泡灌洗液以及细胞上清液的TNF-α、IL-1β、HMGB1水平增高，且肺泡灌洗液水平升高时间晚于血清，提示早期和晚期炎症反应，尤其是晚期炎症反应，可能是AKI诱发ALI发生的分子机制；肾IRI可诱导大鼠肾和肺组织、体外培养的肾小管上皮细胞HGMB1、TLR4、MyD88、p-TRAF-6、p-IRAK1、F-κB蛋白表达上调，AQP-1表达下调，提示肾IRI诱导了肾和肺组织依赖于MyD88的HGMB1/TLR4信号通路活化，之后调控TNF-α、IL-1β的转录增加，继而通过炎症因子抑制AQP-1的转录，参与AKI诱导ALI的发生发展，这可能是其重要的分子信号机制；进一步的研究发现，冬虫夏草可降低大鼠血清BUN和Scr水平，改善肾和肺组织的病理改变，降低肺组织的湿干重比，体内证明冬虫夏草对AKI诱发的ALI均有保护作用，且具有剂量依赖性。而且冬虫夏草可以降低血清、肺泡灌洗液以及细胞上清液的TNF-α、IL-1β、HMGB1水平，下调大鼠肾组织和肺组织、体外肾小管上皮细胞的HGMB1、TLR4、MyD88、p-TRAF-6、p-IRAK1、NF-κB蛋白，上调肾和肺的AQP-1表达，提示冬虫夏草可通过阻断依赖于MyD88的HGMB1/TLR4信号通路活化，抑制TNF-α、IL-1β的表达，介导AQP-1的转录，通过抑制I/R诱导的肺肾晚期炎症，发挥肾和肺的保护作用，且具有剂量依赖性，晚期炎症反应、AQP-1和HGMB1/TLR4信号通路可能是肾IRI病理生理过程中肺肾同源的物质基础。