Department of Hepatobiliary Surgery, Chongqing Medical University, Second Affiliated Hospital, Chongqing, China.
J Surg Res. 2012 Jun 1;175(1):88-100. doi: 10.1016/j.jss.2011.02.026. Epub 2011 Mar 17.
High mobility group protein B1 (HMGB1) is an important late inflammatory mediator in sepsis. Understanding the mechanisms that regulate HMGB1 release from cells and their downstream signal transduction pathways may lead to the ability to develop anti-HMGB1 therapies to treat inflammation.
We stimulated murine macrophage-like RAW 264.7 cells with lipopolysaccharide (LPS) and LPS+ ethylpyruvate (EP) and examined the resulting HMGB1 expression and release. We also studied the expression of related signal transduction factors (NF-κB, p38 MAPK, and CBP).
Gene expression of HMGB1 mRNA in RAW264.7 cell showed no significant change at 0-18 h after stimulation with LPS, but increased significantly at 24, 36, and 48 h. HMGB1 mRNA expression in the LPS+EP group was significantly lower than in LPS alone. HMGB1 was distributed mainly in the nucleus; the cytoplasmic level was low before LPS stimulation. After stimulation with LPS, cytoplasmic HMGB1 increased gradually and plateaued at a high level at 12-48 h. Nuclear HMGB1 decreased gradually at 12-24 h, then increased, maintaining a comparatively high level at 36-48 h. EP prevented this pattern significantly. LPS induced p38 MAPK activation and NF-κB signal pathways first, followed by CBP activation. Activated CBP acetylated HMGB1 was stored in a crino-lysosome and secreted activated NF-κB resulted in increased transcription and synthesis of HMGB1, but the expression of up-regulated HMGB1 mRNA was delayed. Extracellular HMGB1 originated from early synthetic reserves present in the nucleus. New HMGB1 protein was synthesized in the nucleus and transferred into the cytoplasm, causing an increase in HMGB1 in the nucleus and cytoplasm. EP inhibits HMGB1 mRNA up-regulation and release from LPS- stimulated macrophages. The molecular function of EP is to attenuate the activation p38 MAPK, NF-κB, and CBP signaling pathways.
高迁移率族蛋白 B1（HMGB1）是脓毒症中一种重要的晚期炎症介质。了解调节细胞内 HMGB1 释放及其下游信号转导途径的机制，可能有助于开发抗 HMGB1 治疗炎症的方法。
我们用脂多糖（LPS）和 LPS+乙基丙酮酸（EP）刺激鼠巨噬样 RAW 264.7 细胞，检测 HMGB1 的表达和释放。同时，我们还研究了相关信号转导因子（NF-κB、p38MAPK 和 CBP）的表达。
LPS 刺激 RAW264.7 细胞 0-18 h 时，HMGB1mRNA 基因表达无明显变化，但在 24、36 和 48 h 时明显增加。LPS+EP 组的 HMGB1mRNA 表达明显低于 LPS 组。HMGB1 主要分布在细胞核内；LPS 刺激前细胞质中 HMGB1 水平较低。LPS 刺激后，细胞质 HMGB1 逐渐增加，在 12-48 h 时达到较高水平并保持稳定。核内 HMGB1 在 12-24 h 逐渐减少，然后增加，在 36-48 h 时保持较高水平。EP 可显著抑制这种模式。LPS 首先诱导 p38MAPK 激活和 NF-κB 信号通路，然后激活 CBP。激活的 CBP 乙酰化 HMGB1 被储存于皱粒-溶酶体中，激活的 NF-κB 导致 HMGB1 转录和合成增加，但上调的 HMGB1mRNA 表达延迟。细胞外 HMGB1 来源于核内早期合成的储备。新的 HMGB1 蛋白在核内合成并转移到细胞质中，导致核内和细胞质内 HMGB1 增加。EP 抑制 LPS 刺激的巨噬细胞中 HMGB1mRNA 的上调和释放。EP 的分子功能是减弱 p38MAPK、NF-κB 和 CBP 信号通路的激活。
