The landscape of implantation and placentation: deciphering the function of dynamic RNA methylation at the maternal-fetal interface.

The maternal-fetal interface is defined as the interface between maternal tissue and sections of the fetus in close contact. RNA methylation modifications are the most frequent kind of RNA alterations. It is effective throughout both normal and pathological implantation and placentation during pregnancy. By influencing early embryo development, embryo implantation, endometrium receptivity, immune microenvironment, as well as some implantation and placentation-related disorders like miscarriage and preeclampsia, it is essential for the establishment of the maternal-fetal interface. Our review focuses on the role of dynamic RNA methylation at the maternal-fetal interface, which has received little attention thus far. It has given the mechanistic underpinnings for both normal and abnormal implantation and placentation and could eventually provide an entirely novel approach to treating related complications.

ER-phagy Is Involved in the Degradation of Collagen I by IL-1ß in Human Amnion in Parturition.

The process of parturition is associated with inflammation within the uterine tissues, and IL-1ß is a key proinflammatory cytokine involved. Autophagy is emerging as an important pathway to remove redundant cellular components. However, it is not known whether IL-1ß employs the autophagy pathway to degrade collagen, thereby participating in membrane rupture at parturition. In this study, we investigated this issue in human amnion. Results showed that IL-1ß levels were significantly increased in human amnion obtained from deliveries with spontaneous labor and membrane rupture, which was accompanied by decreased abundance of COL1A1 and COL1A2 protein but not their mRNA, the two components of collagen I. Consistently, IL-1ß treatment of cultured primary human amnion fibroblasts reduced COL1A1 and COL1A2 protein but not their mRNA abundance along with increased abundance of autophagy activation markers, including the microtubule-associated protein L chain 3ß II/I ratio and autophagy-related 7 (ATG7) in the cells. The reduction in COL1A1 and COL1A2 protein abundance induced by IL-1ß could be blocked by the lysosome inhibitor chloroquine or small interfering RNA-mediated knockdown of ATG7 or ER-phagy receptor FAM134C, suggesting that FAM134C-mediated ER-phagy was involved in IL-1ß-induced reduction in COL1A1 and COL1A2 protein in amnion fibroblasts. Consistently, levels of L chain 3ß II/I ratio, ATG7, and FAM134C were significantly increased in human amnion obtained from deliveries with spontaneous labor and membrane rupture. Conclusively, increased IL-1ß abundance in human amnion may stimulate ER-phagy-mediated COL1A1 and COL1A2 protein degradation in amnion fibroblasts, thereby participating in membrane rupture at parturition.

ER-phagy Is Involved in the Degradation of Collagen I by IL-1ß in Human Amnion in Parturition.

The process of parturition is associated with inflammation within the uterine tissues, and IL-1ß is a key proinflammatory cytokine involved. Autophagy is emerging as an important pathway to remove redundant cellular components. However, it is not known whether IL-1ß employs the autophagy pathway to degrade collagen, thereby participating in membrane rupture at parturition. In this study, we investigated this issue in human amnion. Results showed that IL-1ß levels were significantly increased in human amnion obtained from deliveries with spontaneous labor and membrane rupture, which was accompanied by decreased abundance of COL1A1 and COL1A2 protein but not their mRNA, the two components of collagen I. Consistently, IL-1ß treatment of cultured primary human amnion fibroblasts reduced COL1A1 and COL1A2 protein but not their mRNA abundance along with increased abundance of autophagy activation markers, including the microtubule-associated protein L chain 3ß II/I ratio and autophagy-related 7 (ATG7) in the cells. The reduction in COL1A1 and COL1A2 protein abundance induced by IL-1ß could be blocked by the lysosome inhibitor chloroquine or small interfering RNA-mediated knockdown of ATG7 or ER-phagy receptor FAM134C, suggesting that FAM134C-mediated ER-phagy was involved in IL-1ß-induced reduction in COL1A1 and COL1A2 protein in amnion fibroblasts. Consistently, levels of L chain 3ß II/I ratio, ATG7, and FAM134C were significantly increased in human amnion obtained from deliveries with spontaneous labor and membrane rupture. Conclusively, increased IL-1ß abundance in human amnion may stimulate ER-phagy-mediated COL1A1 and COL1A2 protein degradation in amnion fibroblasts, thereby participating in membrane rupture at parturition.

Involvement of serum amyloid A1 in the rupture of fetal membranes through induction of collagen I degradation.

The de novo synthesis of serum amyloid A1 (SAA1) is augmented in human fetal membranes at parturition. However, its role in parturition remains largely unknown. Here, we investigated whether SAA1 was involved in the rupture of fetal membranes, a crucial event in parturition accompanied with extensive degradation of collagens. Results showed that SAA1 decreased both intracellular and extracellular COL1A1 and COL1A2 abundance, the two subunits of collagen I, without affecting their mRNA levels in human amnion fibroblasts. These reductions were completely blocked only with inhibition of both matrix metalloproteases (MMPs) and autophagy. Consistently, SAA1 increased MMP-2/9 abundance and the markers for autophagic activation including autophagy related (ATG) 7 (ATG7) and the microtubule-associated protein light chain 3 ß (LC3B) II/I ratio with the formation of LC3 punctas and autophagic vacuoles in the fibroblasts. Moreover, the autophagic degradation of COL1A1/COL1A2 and activation of MMP-2/9 by SAA1 were blocked by inhibitors for the toll-like receptors 2/4 (TLR2/4) or NF-κB. Finally, reciprocal corresponding changes of SAA1 and collagen I were observed in the amnion following spontaneous rupture of membranes (ROM) at parturition. Conclusively, SAA1 may participate in membrane rupture at parturition by degradating collagen I via both autophagic and MMP pathways. These effects of SAA1 appear to be mediated by the TLR2/4 receptors and the NF-κB pathway.

Proteasome-mediated degradation of collagen III by cortisol in amnion fibroblasts.

Rupture of fetal membranes (ROM) can initiate parturition at both term and preterm. Collagen III in the compact layer of the amnion contributes to the tensile strength of fetal membranes. However, the upstream signals triggering collagen III degradation remain mostly elusive. In this study, we investigated the role of cortisol regenerated by 11ß-hydroxysteroid dehydrogenase 1 (11ß-HSD1) in collagen III degradation in human amnion fibroblasts with an aim to seek novel targets for the prevention of preterm premature ROM (pPROM)-elicited preterm birth. Human amnion tissue and cultured amnion tissue explants and amnion fibroblasts were used to study the regulation of collagen III, which is composed of three identical 3α 1 chains (COL3A1), by cortisol. Cortisol decreased COL3A1 protein but not mRNA abundance in a concentration-dependent manner. Cortisone also decreased COL3A1 protein, which was blocked by 11ß-HSD1 inhibition. The reduction in COL3A1 protein by cortisol was not affected by a transcription inhibitor but was further enhanced by a translation inhibitor. Autophagic pathway inhibitor chloroquine or siRNA-mediated knock-down of ATG7, an essential protein for autophagy, failed to block cortisol-induced reduction in COL3A1 protein abundance, whereas proteasome pathway inhibitors MG132 and bortezomib significantly attenuated cortisol-induced reduction in COL3A1 protein abundance. Moreover, cortisol increased COL3A1 ubiquitination and the reduction of COL3A1 protein by cortisol was blocked by PYR-41, a ubiquitin-activating enzyme inhibitor. Conclusively, cortisol regenerated in amnion fibroblasts may be associated with ROM at parturition by reducing collagen III protein abundance through a ubiquitin-proteasome pathway.

AKAP95-mediated nuclear anchoring of PKA mediates cortisol-induced PTGS2 expression in human amnion fibroblasts.

Phosphorylation of the transcription factors cyclic adenosine monophosphate response element-binding protein (CREB) and signal transducer and activator of transcription 3 (STAT3) by protein kinase A (PKA) is required for the cortisol-induced production of cyclooxygenase-2 (COX-2) and prostaglandin E2 (PGE2) in human amnion fibroblasts, which critically mediates human parturition (labor). We found that PKA was confined in the nucleus by A-kinase-anchoring protein 95 (AKAP95) in amnion fibroblasts and that this localization was key to the cortisol-induced expression of PTGS2, the gene encoding COX-2. Cortisol increased the abundance of nuclear PKA by stimulating the expression of the gene encoding AKAP95. Knockdown of AKAP95 not only reduced the amounts of nuclear PKA and phosphorylated CREB but also attenuated the induction of PTGS2 expression in primary human amnion fibroblasts treated with cortisol, whereas the phosphorylation of STAT3 in response to cortisol was not affected. The abundances of AKAP95, phosphorylated CREB, and COX-2 were markedly increased in human amnion tissue after labor compared to those in amnion tissues from cesarean sections without labor. These results highlight an essential role for PKA that is anchored in the nucleus by AKAP95 in the phosphorylation of CREB and the consequent induction of COX-2 expression by cortisol in amnion fibroblasts, which may be important in human parturition.

Autophagic Degradation of Collagen 1A1 by Cortisol in Human Amnion Fibroblasts.

Rupture of fetal membranes can initiate parturition at both term and preterm. Collagen is the crucial factor determining the tensile strength of the membranes. Toward the end of gestation, a feed-forward regeneration of cortisol via 11ß-hydroxysteroid dehydrogenase 1 exists in fetal membranes. It remains undetermined whether cortisol contributes to collagen reduction in fetal membranes. In this study, we have examined whether cortisol accumulation is a causative factor for collagen reduction in human amnion fibroblasts, the major source of collagens in the membranes. Cortisol had no effect on collagen 1A1 (COL1A1) and 1A2 (COL1A2) messenger RNA (mRNA) abundance but decreased their protein abundance. The latter effect was affected by neither mRNA transcription inhibitor nor protein translation inhibitor. Mechanistic studies revealed that the reduction in COL1A1 but not COL1A2 protein by cortisol was blocked by lysosome inhibitor chloroquine or small interfering RNA (siRNA)-mediated knockdown of autophagy-related protein 7, an essential protein for autophagy, whereas the proteasome inhibitors MG132 and bortezomib were ineffective. Further analysis showed that cortisol dose dependently increased the ratio of LC3II/LC3I, a marker of lysosome activation, an effect blocked by the glucocorticoid receptor (GR) antagonist RU486 and siRNA-mediated knockdown of GR. Consistently, cortisol decreased COL1A1 and COL1A2 protein abundance in amnion tissue explants, and decreased COL1A1 and COL1A2 protein abundance was observed at parturition in the amnion tissue. Conclusively, cortisol regeneration in fetal membranes may contribute to rupture of fetal membranes at parturition by reducing collagen protein abundance. Lysosome-mediated autophagy accounts for the reduction in COL1A1 by cortisol, but the mechanism underlying the reduction in COL1A2 awaits further investigation.

Inhibition of Lysyl Oxidase by Cortisol Regeneration in Human Amnion: Implications for Rupture of Fetal Membranes.

The mechanisms underlying human parturition are still not understood, yet we need this knowledge to combat preterm birth. Fetal membranes express abundant 11ß-hydroxysteroid dehydrogenase 1 (11ß-HSD1), which converts inert cortisone to active cortisol. We examined whether cortisol regeneration in the amnion might play a role in human parturition through regulation of lysyl oxidase (LOX), a collagen cross-linking enzyme, thereby contributing to the rupture of fetal membranes. By using cultured human primary amnion fibroblasts, we demonstrated that, in addition to the induction of the key enzymes involved in prostaglandin E2 (PGE2) synthesis, cortisol stimulated 11ß-HSD1 and inhibited LOX reciprocally. These results were reproduced in human amnion tissue explants after cortisol treatment. Cortisone also inhibited LOX expression, which was abolished by the inhibition of 11ß-HSD1. Despite the inhibition of LOX by PGE2, inhibition of the PGE2 pathway failed to block the inhibition of LOX by cortisol. However, inhibition of glucocorticoid receptor and mutation of a negative glucocorticoid response element in LOX promoter abolished the inhibition of LOX by cortisol. Chromatin immunoprecipitation assay revealed that cortisol increased GR binding to the LOX promoter. Moreover, increased cortisol and 11ß-HSD1 abundance and decreased LOX abundance were observed in human amnion tissue after the labor-initiated spontaneous rupture of membranes. These data highlight a crucial role for local cortisol regeneration by 11ß-HSD1 in the down-regulation of LOX expression via glucocorticoid receptor binding to a negative glucocorticoid response element to its promoter in the amnion, which may contribute to rupture of fetal membranes at parturition.