Human fetal membranes (amniochorion) that line the intrauterine cavity consist of two distinct cell layers; single-layer amnion epithelial cells (AEC) and multilayer chorion trophoblast cells (CTC). These layers are connected through a collagen-rich extracellular matrix. Cellular remodeling helps support membrane growth and integrity during gestation and helps to maintain pregnancy. Preterm prelabor rupture of the human amniochorionic (fetal) membrane (pPROM) is antecedent to 40% of all spontaneous preterm birth. Oxidative stress (OS) induced activation of the p38 MAPK due to various maternal risk exposures and the amniochorion cells' senescence are reported pathological features of pPROM. Our transcriptomics analysis implicated dysregulated autophagy and epithelial-mesenchymal transition (EMT) in fetal membranes from pPROM. The molecular interplay between OS-induced p38 MAPK activation, autophagy, and EMT was investigated in AECs and CTCs to better understand the involvement of autophagy and EMT. We report the differential impact of OS on the autophagic machinery in AECs and CTCs, resulting in distinct cell fates. In AECs, OS-induced p38 MAPK activation causes autophagosome accumulation and reduced autophagic flux mediated by decreased ULK1 activity and kinase activity, leading to senescence. In CTCs, induction of autophagy has a limited effect; however, inhibition of autophagy led to SQSTM1-mediated EMT of trophoblast cells. Autophagy, EMT, and senescence were associated with proinflammatory changes. Thus, AECs and CTCs respond differently to OS via differential autophagy response, partly mediated via p38 MAPK. Besides senescence, OS-induced autophagy dysregulation in amniochorion cells may play a mechanistic role in pPROM pathophysiology.

• Klíčová slova
• Membrane rupture, Oxidative stress, amnion, autophagy, chorion, epithelial-mesenchymal transition, p38 MAPK, preterm birth, preterm premature rupture of membranes, senescence, trophoblast,
• Publikační typ
• časopisecké články MeSH

In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis updated guidelines for monitoring autophagy in different organisms. Despite numerous reviews, there continues to be confusion regarding acceptable methods to evaluate autophagy, especially in multicellular eukaryotes. Here, we present a set of guidelines for investigators to select and interpret methods to examine autophagy and related processes, and for reviewers to provide realistic and reasonable critiques of reports that are focused on these processes. These guidelines are not meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being asked and the system being used. Moreover, no individual assay is perfect for every situation, calling for the use of multiple techniques to properly monitor autophagy in each experimental setting. Finally, several core components of the autophagy machinery have been implicated in distinct autophagic processes (canonical and noncanonical autophagy), implying that genetic approaches to block autophagy should rely on targeting two or more autophagy-related genes that ideally participate in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation in the field.

• Klíčová slova
• Autophagosome, LC3, cancer, flux, lysosome, macroautophagy, neurodegeneration, phagophore, stress, vacuole,
• MeSH
• autofagie * fyziologie   MeSH
• autofagozomy MeSH
• biologické markery MeSH
• biotest normy   MeSH
• lidé MeSH
• lyzozomy MeSH
• proteiny spojené s autofagií metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH
• směrnice MeSH
• Názvy látek
• biologické markery MeSH
• proteiny spojené s autofagií MeSH

Autophagy is an evolutionarily conserved process that captures aberrant intracellular proteins and/or damaged organelles for delivery to lysosomes, with implications for cellular and organismal homeostasis, aging and diverse pathologies, including cancer. During cancer development, autophagy may play both tumour-supporting and tumour-suppressing roles. Any relationships of autophagy to the established oncogene-induced replication stress (RS) and the ensuing DNA damage response (DDR)-mediated anti-cancer barrier in early tumorigenesis remain to be elucidated. Here, assessing potential links between autophagy, RS and DDR, we found that autophagy is enhanced in both early and advanced stages of human urinary bladder and prostate tumorigenesis. Furthermore, a high-content, single-cell-level microscopy analysis of human cellular models exposed to diverse genotoxic insults showed that autophagy is enhanced in cells that experienced robust DNA damage, independently of the cell-cycle position. Oncogene- and drug-induced RS triggered first DDR and later autophagy. Unexpectedly, genetic inactivation of autophagy resulted in RS, despite cellular retention of functional mitochondria and normal ROS levels. Moreover, recovery from experimentally induced RS required autophagy to support DNA synthesis. Consistently, RS due to the absence of autophagy could be partly alleviated by exogenous supply of deoxynucleosides. Our results highlight the importance of autophagy for DNA synthesis, suggesting that autophagy may support cancer progression, at least in part, by facilitating tumour cell survival and fitness under replication stress, a feature shared by most malignancies. These findings have implications for better understanding of the role of autophagy in tumorigenesis, as well as for attempts to manipulate autophagy as an anti-tumour therapeutic strategy.

• MeSH
• autofagie * MeSH
• autofagozomy účinky léků   metabolismus   MeSH
• biologické modely MeSH
• fyziologický stres * účinky léků   MeSH
• kamptothecin farmakologie   MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• onkogeny * MeSH
• replikace DNA * účinky léků   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• kamptothecin MeSH

• Klíčová slova
• LC3, autolysosome, autophagosome, chaperone-mediated autophagy, flux, lysosome, macroautophagy, phagophore, stress, vacuole,
• MeSH
• autofagie * fyziologie   MeSH
• biotest metody   normy   MeSH
• lidé MeSH
• počítačová simulace MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Research Support, N.I.H., Extramural MeSH
• směrnice MeSH