M. M. Hodges1, C. Zgheib1, J. Xu1, J. Hu1, K. W. Liechty1 1University Of Colorado Denver,Department Of General Surgery, Laboratory For Fetal And Regenerative Biology,Aurora, CO, USA
Introduction: Ischemic heart disease remains the leading cause of death worldwide. Despite tremendous advances in the medical management of ischemic cardiomyopathy, the 5-year mortality rate following a diagnosis of heart failure exceeds 50%. In our novel, ovine model of myocardial regeneration after myocardial infarction (MI), we have previously shown that the fetal response to MI is regenerative, whereas the adult response to MI is reparative and is associated with decreased angiogenesis, increased inflammation, and increased fibrosis. Macrophages have been shown to play key roles in regulation of angiogenesis and inflammation; however, the impact that macrophages have on the development of fibrosis and scar formation after MI has yet to be described. We hypothesize that macrophages differentially regulate gene expression of collagen, matrix metalloproteinases (MMP-2 and MMP-9), and transforming growth factors (TGFβ-1 and TGFβ-3) in fetal and adult cardiac fibroblasts.
Methods: To test this hypothesis, cardiac fibroblasts were isolated from the left ventricle of adult and fetal sheep. After isolation, these cardiac fibroblasts were co-cultured for 24 hours with macrophages from the RAW 264.7 cell line. Quantitative polymerase chain reaction was used to quantify the gene expression of collagen 1α2, collagen 3α1, MMP-2, MMP-9, TGFβ-1, and TGFβ-3.
Results: When compared to adult cardiac fibroblasts, fetal cardiac fibroblasts have significantly upregulated expression of both col1α2 and col3α1, as well as significantly upregulated expression of MMP-2, MMP-9, TGFβ-1, and TGFβ-3. After co-culture with macrophages, adult cardiac fibroblasts demonstrated significant upregulation of collagen 1α2, which was associated with a significant upregulation in TGFβ-3 and downregulation in MMP-9 gene expression. Conversely, after co-culture with macrophages, fetal cardiac fibroblasts demonstrate a significant downregulation in both col1α2 and col3α1, as well as a significant downregulation in MMP-2, MMP-9, TGFβ-1, and TGFβ-3. These results suggest a differential response of fetal and adult cardiac fibroblasts to co-culture with macrophages.
Conclusion: Our results suggest that macrophages may play an integral role in regulating the differential responses of the fetus and adult following MI. Additionally, our results suggest that fetal cardiac fibroblasts and adult cardiac fibroblasts are characterized by baseline phenotypic differences that may further contribute to the regenerative response observed in the fetus after MI, compared to the reparative response observed in the adult after MI. A more in-depth understanding of both the phenotypic differences between adult and fetal cardiac fibroblasts, as well as the varying responses following co-culture with macrophages will enable a more complete understanding of the role macrophages play in regulating fibrosis after MI.