Prenatal alcohol exposure impairs autophagy in neonatal brain cortical microvessels.
Brain developmental lesions are a devastating consequence of prenatal alcohol exposure (PAE). We recently showed that PAE affects cortical vascular development with major effects on angiogenesis and endothelial cell survival. The underlying molecular mechanisms of these effects remain poorly understood. This study aimed at characterizing the ethanol exposure impact on the autophagic process in brain microvessels in human fetuses with fetal alcohol syndrome (FAS) and in a PAE mouse model. Our results indicate that PAE induces an increase of autophagic vacuole number in human fetal and neonatal mouse brain cortical microvessels. Subsequently, ex vivo studies using green fluorescent protein (GFP)-LC3 mouse microvessel preparations revealed that ethanol treatment alters autophagy in endothelial cells. Primary cultures of mouse brain microvascular endothelial cells were used to characterize the underlying molecular mechanisms. LC3 and p62 protein levels were significantly increased in endothelial cells treated with 50 mM ethanol. The increase of autophagic vacuole number may be due to excessive autophagosome formation associated with the partial inhibition of the mammalian target of rapamycin pathway upon ethanol exposure. In addition, the progression from autophagosomes to autolysosomes, which was monitored using autophagic flux inhibitors and mRFP-EGFP vector, showed a decrease in the autolysosome number. Besides, a decrease in the Rab7 protein level was observed that may underlie the impairment of autophagosome-lysosome fusion. In addition, our results showed that ethanol-induced cell death is likely to be mediated by decreased mitochondrial integrity and release of apoptosis-inducing factor. Interestingly, incubation of cultured cells with rapamycin prevented ethanol effects on autophagic flux, ethanol-induced cell death and vascular plasticity. Taken together, these results are consistent with autophagy dysregulation in cortical microvessels upon ethanol exposure, which could contribute to the defects in angiogenesis observed in patients with FAS. Moreover, our results suggest that rapamycin represents a potential therapeutic strategy to reduce PAE-related brain developmental disorders.
Normandie University, UNIROUEN, INSERM U1245, NeoVasc Team, Rouen, France.
Virginie
V
Girault
Normandie University, UNIROUEN, INSERM U1245, NeoVasc Team, Rouen, France.|Department of Neurosurgery, Rouen University Hospital, Rouen, France.
Vianney
V
Gilard
Normandie University, UNIROUEN, INSERM U1245, NeoVasc Team, Rouen, France.|Pathology Laboratory, Rouen University Hospital, Rouen, France.
Florent
F
Marguet
Normandie University, UNIROUEN, INSERM U1245, NeoVasc Team, Rouen, France.|Department of Metabolic Biochemistry, Rouen University Hospital, Rouen, France.
Céline
C
Lesueur
Normandie University, UNIROUEN, INSERM U1245, NeoVasc Team, Rouen, France.
Michelle
M
Hauchecorne
Normandie University, UNIROUEN, INSERM U1245, NeoVasc Team, Rouen, France.
Yasmina
Y
Ramdani
Normandie University, UNIROUEN, INSERM U1245, NeoVasc Team, Rouen, France.|Pathology Laboratory, Rouen University Hospital, Rouen, France.
Annie
A
Laquerrière
Normandie University, UNIROUEN, INSERM U1245, NeoVasc Team, Rouen, France.|Department of Neonatal Pediatrics and Intensive Care and Neuropediatrics, Rouen University Hospital, Rouen, France.
Stéphane
S
Marret
Normandie University, UNIROUEN, INSERM U1245, NeoVasc Team, Rouen, France.
Sylvie
S
Jégou
Normandie University, UNIROUEN, INSERM U1245, NeoVasc Team, Rouen, France.
Bruno Jose
BJ
Gonzalez
Normandie University, UNIROUEN, INSERM U1245, NeoVasc Team, Rouen, France.
Carole
C
Brasse-Lagnel
Normandie University, UNIROUEN, INSERM U1245, NeoVasc Team, Rouen, France.|Department of Metabolic Biochemistry, Rouen University Hospital, Rouen, France.
Soumeya
S
Bekri
D000818:Animals; D000071182:Autophagosomes / Q000187:drug effects / Q000378:metabolism; D001343:Autophagy / Q000187:drug effects*; D001921:Brain / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D000431:Ethanol / Q000009:adverse effects*; D005260:Female; D063647:Fetal Alcohol Spectrum Disorders / Q000378:metabolism / Q000473:pathology; D006801:Humans; D008247:Lysosomes / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D055806:Microvessels / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D023421:Models, Animal; D011247:Pregnancy; D011297:Prenatal Exposure Delayed Effects / Q000139:chemically induced* / Q000378:metabolism / Q000473:pathology*; D016601:RNA-Binding Proteins / Q000378:metabolism; D020123:Sirolimus / Q000494:pharmacology
D008869:Microtubule-Associated Proteins; D016601:RNA-Binding Proteins; D000431:Ethanol; D020123:Sirolimus