Evaluation of sex differences on mitochondrial bioenergetics and apoptosis in mice

Abstract

It has been postulated that the differences in longevity observed between organisms of different sexes within a species can be attributed to differences in oxidative stress. It is generally accepted that differences are due to the higher female estrogen levels. However, in some species males live the same or longer despite their lower estrogen values. Therefore, in the present study, we analyze key parameters of mitochondrial bioenergetics, oxidative stress and apoptosis in the B6 (C57Bl/6J) mouse strain. There are no differences in longevity between males and females in this mouse strain, although estrogen levels are higher in females. We did not find any differences in heart, skeletal muscle and liver mitochondrial oxygen consumption (State 3 and State 4) and ATP content between male and female mice. Moreover, mitochondrial H(2)O(2) generation and oxidative stress levels determined by cytosolic protein carbonyls and concentration of 8-hydroxy-2'-deoxyguanosine in mitochondrial DNA were similar in both sexes. In addition, markers of apoptosis (caspase-3, caspase-9 and mono- and oligonucleosomes: the apoptosis index) were not different between male and female mice. These data show that there are no differences in mitochondrial bioenergetics, oxidative stress and apoptosis due to gender in this mouse strain according with the lack of differences in longevity. These results support the Mitochondrial Free Radical Theory of Aging, and indicate that oxidative stress generation independent of estrogen levels determines aging rate.

Fig. 1

Heart, skeletal muscle and liver mouse cytosolic protein carbonyls determined by ELISA (see Methods). Data are shown as means ± SEM. For heart & skeletal muscle, n=6; for liver, n=5.

Fig. 2

Levels of mitochondrial oxidized DNA (8-oxodGuo/10⁶ dGuo) in liver and skeletal muscle. Data are shown as means ± SEM. For heart & skeletal muscle, n=6; for liver, n=5.

Fig. 3

Caspase-3 and caspase-9 activity in heart, skeletal muscle and liver mouse cytosol. Results are reported as arbitrary fluorescence units. Data are shown as means ± SEM. For heart & skeletal muscle, n = 6; for liver, n=5.

Fig. 4

Heart, skeletal muscle and liver cytosolic cleaved caspase-3 content. Results are reported as arbitrary optical density (OD) units/mm². Data are shown as means ± SEM. For heart & muscle groups, n = 6; for liver groups, n = 5.

Fig. 5

Quantification of mono- and oligonucleosomes (apoptotic index) in the heart, skeletal muscle and liver of males and females mice using a quantitative ELISA. Results are reported as arbitrary optical density (OD) units/mg protein. Data are shown as means ± SEM. For heart & muscle groups, n = 6; for liver groups, n = 5.