Isolation of apoptotic mouse fetal oocytes by AnnexinV assay

Int J Dev Biol. 2007;51(2):157-60. doi: 10.1387/ijdb.062203al.

Abstract

Expression of phosphotidylserine by fetal oocytes in culture renders significant numbers of such cells able to bind AnnexinV-coated microbeads and allows their separation from Annexin V-negative oocytes on a Magnetic Cell Separation (MACS) column in a magnetic field. The majority of oocytes (> or =75%) which bound Annexin V-coated microbeads were viable, as indicated by their propidium iodine (PI) negativity. However, they showed apoptotic morphologies and were found to be TUNEL-positive. On the other hand, AnnexinV-negative oocytes, besides being PI negative, appeared morphologically healthy and TUNEL negative. Moreover, AnnexinV-positive oocytes showed a marked lower ratio of Bcl-xL/Bax transcripts in comparison to AnnexinV-negative oocytes. We conclude that the present method is able to separate fetal oocytes in two distinct populations: AnnexinV-positive oocytes showing features typical of apoptotic cells and AnnexinV-negative oocytes comprising for the most part viable non-apoptotic cells. This procedure should greatly facilitate studies aimed to identify the currently poorly understood molecular pathways governing apoptosis in mammalian fetal oocytes.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Annexin A5 / analysis*
  • Biomarkers / analysis
  • Cell Culture Techniques
  • Cell Separation / methods
  • Female
  • In Situ Nick-End Labeling
  • Mice
  • Oocytes / cytology*
  • Oocytes / physiology*
  • Protein Binding
  • Reverse Transcriptase Polymerase Chain Reaction
  • Transcription, Genetic
  • bcl-2-Associated X Protein / genetics
  • bcl-X Protein / genetics

Substances

  • Annexin A5
  • Biomarkers
  • bcl-2-Associated X Protein
  • bcl-X Protein