去磷酸化stathmin1促进微管解聚在LPS诱导血管内皮细胞损伤中的作用研究

Severe sepsis or septic shock is a common and serious consequence of gram-negative infection, which accounts for high mortality in severe burned patients. And endothelial cells (ECs) injury is an important process in sepsis pathogenesis. However, the mechanisms behind LPS-induced vascular ECs injury are still not well clear. The Rho guanine nucleotide exchange factor H1(GEF-H1) is uniquely regulated by microtubule binding and is essential in coupling microtubule dynamics to Rho-GTPase activation in a variety of normal biological situations. Published studies identified that microtubule depolymerization releases GEF-H1 to catalyze nucleotide exchange on RhoA, and that re-locatization to microtubules has an inhibitory effect on the enzymatic activity of GEF-H1. Nevertheless, microtubule dynamics changes are regulated by several microtubule binding proteins (MBPs). As the important member of MBPs, stathmin1 is a key regulator of microtubule dynamics, and dephosphorylated stathmin1 is known to destabilize microtubules in special ways. Phosphorylation of stathmin1 on one or more of its four serine residues (Ser16, Ser25, Ser38, and Ser63) reduces its microtubule-destabilizing activity. Our previous studies demonstrated that GEF-H1/RhoA signaling is involved in LPS-induced overexpression of inflammatory cytokines and vascular leakage dysfunction in vascular ECs. Additionally, a series of preliminary trials for present study showed that dephosphorylated stathmin1 is closely associated with microtubule destabilization in LPS-challenged ECs. Moreover, depleted stathmin1 using small interfering RNA (siRNA) partially attenuated transendothelial electrical resistance (TER) decline induced by LPS stimulation, whereas treatment with nonspecific RNA duplex oligonucleotide did not affect LPS-induced TER decline. These results showed direct involvement of dephsophorylated stathmin1 in LPS-induced vascular barrier compromise. Taken together, we assume that dephosphorylated stathmin1 plays an important role in LPS-induced several inflammatory cytokine expression and vascular barrier leakage in ECs. The purpose of present study not only intends to fully elucidate the underlying regulatory mechanisms and pathological significance of dephosphorylated stathmin1 in LPS-induced ECs injury under septic conditions, but to provide novel treatments for sepsis or septic shock.

严重烧伤后脓毒症的发病率及死亡率一直居高不下，其内在分子机制仍不完全清楚。鸟嘌呤核交换因子H1（GEF-H1）异常活化在脓毒症LPS诱导血管内皮细胞损伤过程中起着重要作用，而GEF-H1活化首先需要微管发生解聚。Stathmin1是调控微管结构的关键蛋白，去磷酸化状态的Stathmin1促进微管解聚。本课题预实验结果显示，LPS能显著提高人肺微血管内皮细胞（HPMECs）内stathmin1去磷酸化水平，促使微管解聚；而抑制stathmin1表达能改善LPS诱导的HPMECs通透性变化。因此，本课题拟通过构建特异性siRNA和stathmin持续磷酸化突变体抑制或下调stathmin活性证实去磷酸化stathmin1促进微管解聚，引起GEF-H1的释放异常增加，调控LPS诱导的HPMECs炎症反应及通透性变化，探讨LPS诱导HPMECs损伤的内在分子机制，以期为临床防治脓毒症提供理论依据。

严重烧伤后脓毒症的发病率及死亡率一直居高不下，其内在分子机制仍不完全清楚。鸟嘌呤核交换因子H1（GEF-H1）异常活化在内毒素(LPS)诱导血管内皮细胞损伤过程中起着重要作用，而GEF-H1活化首先需要微管发生解聚。Stathmin1是调控微管结构的关键蛋白，去磷酸化状态的Stathmin1促进微管解聚。LPS能显著提高人肺微血管内皮细胞（HPMECs）内stathmin1去磷酸化水平，促使微管解聚；而抑制stathmin1表达能改善LPS诱导的HPMECs通透性变化。探讨LPS诱导HPMECs损伤的内在分子机制，以期为临床防治脓毒症提供理论依据。