Mitochondrial CaMKII drives smooth muscle migration and neointimal hyperplasia

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Project Summary / Abstract Vascular occlusive disease remains a critical cardiovascular health issue with about 500,000 percutaneous coronary and 50,000 peripheral balloon angioplasties performed annually in the US alone. Hemodynamically relevant restenoses through neointimal hyperplasia occur in 10 to 30% of patients. Vascular smooth muscle cell (VSMC) migration significantly contributes to neointimal hyperplasia after vascular injury. VSMC migration is a Ca2+-dependent process; migrating cells must maintain cytosolic Ca2+ gradients and create local Ca2+ pulses near the leading edge likely to accomplish dynamic cytoskeletal turnover. Mitochondria are one of the major buffers of intracellular Ca2+ in VSMC and participate in localized Ca2+ responses. In other cell types, cell migration is dependent upon mitochondrial localization to the leading edge. Thus, we propose that mitochondria provide localized control of Ca2+ transients, serving to facilitate VSMC migration. Previous work from our group established that the multifunctional Ca2+/calmodulin-dependent protein kinase II (CaMKII) is a key regulator of VSMC cell migration and proliferation. In more recent studies, we discovered that CaMKII is present and active in the mitochondrial matrix, where it is believed to promote mitochondrial matrix Ca2+ influx. In preliminary studies, mitoCaMKII inhibition in VSMC in a novel transgenic model developed in our laboratory blocks neointimal hyperplasia in vivo. Moreover, mitoCaMKII inhibition blocks mitochondrial Ca2+ uptake, mitochondrial mobility and VSMC migration. These data position mitochondrial CaMKII (mitoCaMKII) as a gatekeeper of mitochondrial function and of key VSMC phenotypes relevant for neointimal hyperplasia. In this this application, we will directly test our working hypothesis that mitoCaMKII inhibition blocks mitochondrial matrix Ca2+ uptake, thereby affecting mitochondrial mobility and VSMC migration and ultimately neointimal hyperplasia. We will test our working hypothesis in two aims: 1. Determine whether mitoCaMKII in VSMC controls neointimal formation through regulation of mitochondrial Ca2+ uptake and 2. Dissect the pathways and mechanisms by which mitoCaMKII controls VSMC migration. Studies will include in vivo analysis of neointimal formation using novel transgenic models of mitoCaMKII inhibition or overexpression and in vitro imaging of localized Ca2+ dysregulation, including ER/mitochondrial Ca2+ transition, Ca2+ waves and flickers as well as analysis of cytoskeleton and focal adhesion turnover. Moreover, novel pathways that link mitochondrial matrix Ca2+ uptake to VSMC migration will be tested. It is anticipated that the successful completion of the proposed studies will provide mechanistic insight into how changes in mitochondrial function lead to VSMC migration and neointimal formation. Such knowledge could lead to first-in-class, mitochondria- targeted therapies for vascular disease, in particular neointimal hyperplasia.