Clec4e deletion in cardiac resident cells leads to smaller infarct size and better cardiac remodeling following myocardial ischemia-reperfusion injury

Abstract Introduction Clec4e-signalling in cardiac resident cells and circulating immune cells stimulates recruitment of proinflammatory leukocytes to infarcted myocardium following ischemia reperfusion injury (I/R) and contributes to ischemic myocardial damage. Clec4e deletion is associated with reduced cardiac injury and better left ventricular structural and functional remodeling. The mechanisms of this cardioprotective effect remain unknown. Research question: We investigated whether Clec4e gene function in cardiac resident cells vs infiltrating immune cells mediates cardioprotection during myocardial ischemia-reperfusion injury. Methods We generated chimeric mice by performing bone marrow transfer in wild type (WT) C57Bl6/J and C57Bl6/J Clec4e-/- mice. Following total body irradiation using 9.5 Gy in 8w-old WT and Clec4e-/- receptor mice, 5 x 10^6 bone marrow cells originating from Clec4e-/- and WT donor mice, respectively, were injected IV. Four weeks later, mice were anesthetized and subjected to ischemia reperfusion injury (I/R) by transient occlusion of the LAD for 60 min. Troponin levels were measured at 90 min post I/R and 7T-magnetic resonance imaging with late gadolinium enhancement (LGE) for infarct size evaluation was performed at 24 hours and 28 days post I/R without LGE. Mice were euthanized at 28 days post I/R and the heart and long shaft bones were processed for postmortem immunohistochemistry and analysis of BM chimerism. Results Immunohistochemical analysis of CLEC4E expression on a bone marrow smear 8 w post bone marrow transfer confirmed successful BM chimerism (7% vs 99%, p<0.01). TnI levels at 90 minutes and LGE assessed infarct size on MRI at 24 hours were not significantly different between groups (n=17 in both groups) (42.1 vs 48.4 ng/ml, and 25.2 vs 31.5% of LVmass in Clec4e-/- mice with WT bone marrow versus in WT mice with Clec4e-/- BM, respectively) suggesting similar myocardial damage . After 28 days, mean LV end diastolic volumes (43µL vs 55µL), and mean end systolic volumes (13µL vs 25µL) were significantly (p<0,05) smaller in Clec4e-/- mice with WT bone marrow resulting in better preserved global LV function (LVEF 62% vs 47%, p<0,05, fig A). Infarct size, semiquantitatively assessed using planimetry on Sirius red stains showed a smaller infarct size in the Clec4e-/- mice with WT bone marrow (6.9% vs 19.8%, p<0.01). Figure B shows a representative example of LGE infarct size to scar tissue on MRI and Sirius red staining. Conclusion Clec4e gene deletion in cardiac resident cells, but not in infiltrating hematopoietic immune cells is associated with better functional and structural remodeling after myocardial I/R. Inhibition of Clec4e signaling via blocking antibodies or small molecule inhibitors in cardiac myocytes might be a promising cardioprotective strategy.