Rahman Mohd Akhlakur, Mitra Srabani, Sarkar Anasuya, Wewers Mark D
Dorothy M. Davis Heart and Lung Research Institute, Department of Internal Medicine, Division of Pulmonary, Allergy, Critical Care and Sleep Medicine, Wexner Medical Center, Ohio State University, Columbus, OH, United States of America.
PLoS One. 2015 Feb 6;10(2):e0117330. doi: 10.1371/journal.pone.0117330. eCollection 2015.
Alpha 1-antitrypsin (A1AT) is a 52 kDa serine protease inhibitor produced largely by hepatocytes but also by mononuclear phagocytes. A1AT chiefly inhibits neutrophil elastase and proteinase-3 but has also been reported to have immune modulatory functions including the ability to inhibit caspases. Its clinical availability for infusion suggests that A1AT therapy might modulate caspase related inflammation. Here we tested the ability of A1AT to modulate caspase-1 function in human mononuclear phagocytes.
Purified plasma derived A1AT was added to active caspase-1 in a cell-free system (THP-1 lysates) as well as added exogenously to cell-culture models and human whole blood models of caspase-1 activation. Functional caspase-1 activity was quantified by the cleavage of the caspase-1 specific fluorogenic tetrapeptide substrate (WEHD-afc) and the release of processed IL-18 and IL-1β.
THP-1 cell lysates generated spontaneous activation of caspase-1 both by WEHD-afc cleavage and the generation of p20 caspase-1. A1AT added to this cell free system was unable to inhibit caspase-1 activity. Release of processed IL-18 by THP-1 cells was also unaffected by the addition of exogenous A1AT prior to stimulation with LPS/ATP, a standard caspase-1 activating signal. Importantly, the A1AT exhibited potent neutrophil elastase inhibitory capacity. Furthermore, A1AT complexed to NE (and hence conformationally modified) also did not affect THP-1 cell caspase-1 activation. Finally, exogenous A1AT did not inhibit the ability of human whole blood samples to process and release IL-1β.
A1AT does not inhibit human monocyte caspase-1.
α1-抗胰蛋白酶（A1AT）是一种52 kDa的丝氨酸蛋白酶抑制剂，主要由肝细胞产生，但也可由单核吞噬细胞产生。A1AT主要抑制中性粒细胞弹性蛋白酶和蛋白酶-3，但也有报道称其具有免疫调节功能，包括抑制半胱天冬酶的能力。其可用于输注的临床可用性表明，A1AT治疗可能调节与半胱天冬酶相关的炎症。在此，我们测试了A1AT调节人单核吞噬细胞中半胱天冬酶-1功能的能力。
将纯化的血浆来源的A1AT添加到无细胞系统（THP-1裂解物）中的活性半胱天冬酶-1中，并外源性添加到半胱天冬酶-1激活的细胞培养模型和人全血模型中。通过半胱天冬酶-1特异性荧光四肽底物（WEHD-afc）的裂解以及加工后的IL-18和IL-1β的释放来定量功能性半胱天冬酶-1活性。
THP-1细胞裂解物通过WEHD-afc裂解和p20半胱天冬酶-1的产生自发激活半胱天冬酶-1。添加到该无细胞系统中的A1AT无法抑制半胱天冬酶-1活性。在用LPS/ATP（一种标准的半胱天冬酶-1激活信号）刺激之前，外源性添加A1AT也不会影响THP-1细胞加工后的IL-18的释放。重要的是，A1AT表现出强大的中性粒细胞弹性蛋白酶抑制能力。此外，与NE复合（因此构象发生改变）的A1AT也不影响THP-1细胞半胱天冬酶-1的激活。最后，外源性A1AT不会抑制人全血样本加工和释放IL-1β的能力。
A1AT不抑制人单核细胞半胱天冬酶-1。
