Figure 4
From: Dysfunctional MDR-1 disrupts mitochondrial homeostasis in the oocyte and ovary
mdr1a mutant oocytes have increased ROS and perturbed mitochondrial membrane potential. (A) Images of oocytes from mdr1a mutant and wild type mice stained with MitoSOX Red Mitochondrial Superoxide Indicator imaged imaged on a confocal microscope (60x oil magnification) to show MitoTracker Green, MitoSOX, and merge. (B) Quantification of average fluorescence intensity (wild type n = 4, mdr1a mutant n = 8, positive control n = 4). Significantly more (p < 0.05) mitochondrial superoxide signaling was detected in the mdr1a mutant oocytes compared to wild type oocytes at baseline without a toxic challenge. (C) Images of wild type, mdr1a mutant, and wild type H2O2 treated oocytes as a positive control stained in CM-H2DCFDA imaged on a confocal microscope (40x oil magnification) in brightfield, GFP, and merge. (D) Quantification of average fluorescence intensity (wild type n = 5, mdr1a mutant n = 9, positive control n = 5). Significantly more (p < 0.05) cellular superoxide signaling was detected in the mdr1a mutant oocytes compared to wild type oocytes. (E) Images of wild type, mdr1a mutant (60x oil magnification), and wild type CCCP treated oocytes as a positive control (30x magnification) stained with JC-1. MitoPT JC-1 accumulates in negatively charged (normal) mitochondria and aggregates, fluorescing red upon excitation. When mitochondrial membrane potential is disturbed (abnormal), MitoPT is distributed in its monomeric form throughout the cell, fluorescing green upon excitation. (F) Ratio of red to green fluorescence signal intensities across 10 sections of individual oocytes (wild type n = 6, mdr1a mutant n = 3, positive control n = 10) in random fluorescence units (RFUs). Wild type oocytes exhibit higher red to green fluorescence ratios, indicating that the cells contain more mitochondria with normal negative membrane potential than mutant oocytes. Quantification performed using ImageJ.
