Modulation of autophagy influences development and apoptosis in mouse embryos developing in vitro

Mol Reprod Dev. 2011 Jul;78(7):498-509. doi: 10.1002/mrd.21331. Epub 2011 Jun 16.

Abstract

Autophagyis, the bulk degradation of proteins and organelles, is essential for cellular maintenance, cell viability, and development, and is often involved in type II programmed cell death in mammals. This study investigated the expression levels of autophagy-related genes and the effect of 3-methyladenine (3-MA, an autophagy inhibitor) or rapamycin (an autophagy inducer) on the in vitro development and apoptosis of mouse embryos. LC3, which is essential for the formation of autophagosomes, was widely expressed in mouse embryos, and high levels of transcript were present from 1 to 4 cells but gradually decreased through the morula and blastocyst stages. 3-MA-treated embryos exhibited significantly reduced developmental rates and total cell numbers, but increased rates of apoptosis. Furthermore, both the expression of Lc3, Gabarap, Atg4A, and Atg4B, and the synthesis of LC3 were significantly reduced at the blastocyst stage. Although rapamycin treatment did not affect developmental rates, cell numbers decreased, and the apoptosis rate increased. Expression of Lc3, Gabarap, Atg4A, and Atg4B, and synthesis of LC3 increased as well. Modulation of Lc3 mRNA and LC3 protein levels using 3-MA or rapamycin significantly increased apoptotic cell death through the disruption of mitochondrial morphology and reduction of mtDNA copy number at the blastocyst stage. Interestingly, the inner cell mass, detected by immunostaining with POU5F1 (OCT3/4) after 3-MA or rapamycin treatment of embryos, was significantly increased compared to controls. These results suggest that autophagy influences developmental patterning and apoptosis, and may play a role in early mouse embryogenesis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenine / analogs & derivatives
  • Adenine / pharmacology
  • Animals
  • Apoptosis / drug effects
  • Apoptosis / physiology*
  • Apoptosis Regulatory Proteins / genetics
  • Apoptosis Regulatory Proteins / metabolism
  • Autophagy / drug effects
  • Autophagy / physiology*
  • Blastocyst / metabolism
  • Cell Nucleus / metabolism
  • Embryo, Mammalian
  • Female
  • Gene Expression Regulation, Developmental
  • Histocytochemistry
  • Male
  • Mice / embryology*
  • Mice, Inbred ICR
  • Microscopy, Confocal
  • Mitochondria / metabolism
  • Morula / metabolism
  • Octamer Transcription Factor-3 / genetics
  • Octamer Transcription Factor-3 / metabolism
  • Sirolimus / pharmacology

Substances

  • Apoptosis Regulatory Proteins
  • Octamer Transcription Factor-3
  • Pou5f1 protein, mouse
  • 3-methyladenine
  • Adenine
  • Sirolimus