Oxidative stress-induced TGF-beta/TAB1-mediated p38MAPK activation in human amnion epithelial cells†

Abstract

Term and preterm parturition are associated with oxidative stress (OS)-induced p38 mitogen-activated protein kinase (MAPK)-mediated fetal tissue (amniochorion) senescence. p38MAPK activation is a complex cell and stimulant-dependent process. Two independent pathways of OS-induced p38MAPK activation were investigated in amnion epithelial cells (AECs) in response to cigarette smoke extract (CSE: a validated OS inducer in fetal cells: 1) The OS-mediated oxidation of apoptosis-signaling kinase (ASK)-1 bound Thioredoxin (Trx[SH]2 dissociates this complex, creating free and activated ASK1-signalosome and 2) transforming growth factor mediated activation of (TGF)-beta-activated kinase (TAK)1 and TGF-beta-activated kinase 1-binding protein (TAB)1. AECs isolated from normal term, not-in-labor fetal membranes increased p38MAPK in response to CSE and downregulated it in response to antioxidant N-acetylcysteine. In AECs, both Trx and ASK1 were localized; however, they remained dissociated and not complexed, regardless of conditions. Silencing either ASK1 or its downstream effectors (MKK3/6) did not affect OS-induced p38MAPK activation. Conversely, OS increased TGF-beta's release from AECs and increased phosphorylation of both p38MAPK and TAB1. Silencing of TAB1, but not TAK1, prevented p38MAPK activation, which is indicative of TAB1-mediated autophosphorylation of p38MAPK, an activation mechanism seldom seen. OS-induced p38MAPK activation in AECs is ASK1-Trx signalosome-independent and is mediated by the TGF-beta pathway. This knowledge will help to design strategies to reduce p38MAPK activation-associated pregnancy risks.

Amnion membrane organ‐on‐chip: an innovative approach to study cellular interactions

“…During gestation, AECs and AMCs undergo cyclic cellular remodeling to heal gaps and MFs in the membranes, a mechanism required to maintain membrane homeostasis. Membrane remodeling at a cellular level is achieved by EMT of AECs and MET of AMCs, aided by redox radicals, growth factors (e.g., TGF-b), and endocrine mediators (e.g., progesterone) (17,19,33,45). Cellular gaps are created (8) when AECs are shed from the membrane because of cellular senescence, mechanical disruption caused by fetal and amniotic fluid shear stress, or both.…”

“…Increased OS at term has been shown to induce laborassociated changes, such as cellular senescence, matrix metallopeptidase 9 up-regulation, and increased proinflammatory cytokine production in fetal membrane cells, including AECs and AMCs (2,23). CSE, a potent and reliable OS inducer (2,17,23,(31)(32)(33)(34)(35), has been shown to recreate the labor phenotype (OS experienced at term labor in amnion membranes) in vitro and to induce a static state of EMT in AECs (17,33). EMT contributes to sustained inflammation that promotes the labor-related cascade of events.…”

Interleukin (IL)-6: A Friend or Foe of Pregnancy and Parturition? Evidence From Functional Studies in Fetal Membrane Cells

“…Highly elastic amnion layer membranes maintain homeostasis through cyclic cellular transitions of amnion epithelial cells (AEC) to amnion mesenchymal cell (AMC) (EMT) and mesenchymal back to epithelial (MET) cells ( Richardson et al, 2020 ). A terminal state of EMT occurs at term where MET is stalled due to oxidative stress (OS)-induced senescence ( Richardson et al, 2020 ) and accumulation of pro-EMT factors like TGFβ in membrane cells and in the amniotic fluid ( Richardson et al, 2018 ). This leads to the accumulation of AMCs in the matrix and promotes localized inflammation and collagenolysis, leading to mechanical weakening ( Richardson et al, 2020 ).…”

Changes in mediators of pro‐cell growth, senescence, and inflammation during murine gestation

“…Fetal membranes collected on different gestational days were lysed with RIPA lysis buffer (50 mmol/L Tris pH 8.0, 150 mmol/L NaCl, 1% Triton X‐100, and 1.0 mmol/L EDTA pH 8.0, 0.1% SDS) supplemented with protease and phosphatase inhibitor cocktail and phenylmethylsulfonyl fluoride as described previously by our laboratory . A Pierce BCA Protein Assay Kit (Thermo Scientific) was utilized to determine protein concentration of samples, and 45 µg of tissue lysate was used for loading the gels for Western blotting, which was performed as previously described by our laboratory . Briefly, samples were run on gradient (4%‐15%) SDS‐PAGE Mini‐PROTEAN TGX pre‐cast gels (Bio‐Rad) and transferred to the membrane using a Bio‐Rad gel transfer device (Bio‐Rad).…”