636. Enhanced Skeletal Muscle Regeneration in Mice After Multiple Pregnancies
Authors: Alex C. Scibetta, Aiping Lu, Johnny Huard
Journal: Molecular Therapy (2016)

Introduction: It has been recently shown that the reduction in the regenerative potential of various tissues during aging is potentially attributed to the depletion of stem cells1Conboy et al., 2003.Google Scholar. Interestingly, aged progenitor cells can be rejuvenated when exposed to a young systemic environment. Pregnant women transiently acquire a population of cells from their fetuses2Lapaire et al., 2007.Google Scholar, and recent studies have shown that fetal cells persist in maternal blood and organs decades after delivery3Bianchi et al., 1996.Google Scholar, 4Khosrotehrani et al., 2004.Google Scholar. Pregnancy is believed to represent a natural form of parabiosis, which joins two organisms so that they share a circulatory system and a systemic environment5Slavin, 2002.Google Scholar. Fetal cells have the potential to differentiate and repair injured tissues and organs in the mother, including skeletal muscle6Nguyen et al., 2007.Google Scholar, 7Zeng et al., 2010.Google Scholar. Previous studies have demonstrated that pregnant mice have improved musculoskeletal tissue healing after injury compared to non-pregnant controls2Lapaire et al., 2007.Google Scholar. In this study, we wanted to determine if the effect of pregnancy on the regenerative potential of skeletal muscle is only transient during the pregnancy period or can persist after pregnancy. Methods: This study used 5 retired breeders and 5 virgin heterozygous Zmpste24 mice (normal phenotype) that we received from Dr. Carlos López Otín. Mice were 8 months of age at the time of cardiotoxin (CTX) injury, and all breeders had been pregnant at least 5 times. To create muscle injury, 4uM of CTX was injected intramuscularly into the gastrocnemius muscle of mice. The mice were sacrificed 7 days after injury and muscles were harvested, flash frozen, and sectioned. H&E staining was performed according to the manufacturer's instructions. Cryosections from the gastrocnemius were stained for mouse IgG using a Mouse on Mouse (M.O.M) kit to analyze necrosis. Muscle regeneration was evaluated using an eMyHC antibody, and the extent of muscle tissue inflammation was analyzed using an F4/80 antibody (macrophage marker). Results: (1) H&E showed histology improvement in the muscles of the breeder mice 7 days post-injury compared to virgin mice. Damaged areas and mononuclear infiltration are two indicators of muscle damaged. We observed that virgin mice had significantly larger damaged areas (p=0.016) and more mononuclear infiltration (p=0.016) compared to breeder mice, which suggested a slower muscle regeneration in the virgin mice. There were no differences in centrally nucleated myofibers, an indicator of early regenerating muscle, between the two groups (p=0.069). (2) Based on immunostaining there was no difference in macrophage infiltration between the two groups. We also did not find significant differences for eMyHC and mouse IgG between breeder and virgin mice. Conclusion: During pregnancy, the fetus may supply the mother with rejuvenating factors and cells that alter the maternal microenvironment. This study demonstrated that mice that have experienced multiple pregnancies repair their muscle faster than virgin mice do, and the rejuvenating effects of the fetus on maternal muscle repair may continue after childbirth. Future studies comparing breeder and virgin mice will increase the number of animals used. These studies may provide insight into the long-lasting effects of pregnancy and have applications for gene and cell therapy. The ultimate goal is to identify beneficial factors from the fetus that can be purified and eventually used to delay aging related disease and conditions.