Project Abstract
Critical limb threatening ischemia (CLTI), the end stage of peripheral arterial disease (PAD), is diagnosed in
500,000 patients each year, often results in amputation, and has a ~50% 5-year mortality rate. Diabetic CLTI
patients experience especially high morbidity and mortality, and no effective non-surgical therapy exists for this
population. Data from our Phase II MOBILE trial demonstrated that autologous bone marrow nucleated cells
(ABMNC) were refractory in preventing amputations in diabetic patients. Thus, a more effective cell preparation
is needed to successfully treat diabetic CLTI. Recent data from our Phase I CHAMP trial shows that allogeneic
bone marrow-derived mesenchymal stromal cells (BMD-MSC) are more robust than ABMNC for stimulating
angiogenesis in ischemic muscle, specifically in diabetics. Our long-term goal is to develop a next generation
cell-based regenerative therapy to improve muscle function and prevent amputations in diabetic patients.
Preliminary data from a polygenic diabetic mouse model of CLTI indicate that human BMD-MSC stimulate limb
perfusion as well as promoting muscle function and regeneration. Furthermore, our preliminary data suggest
that alginate hydrogel-encapsulated MSC, including a novel vertebral bone adherent (vBA) MSC, are
significantly superior to unassociated MSC for tissue regeneration presumably due to protection from host
immune clearance and production of an exosome profile associated with angio- and myogenesis. Based on
preliminary data and published studies, our central hypothesis is that allogeneic encapsulated vBA-MSC will
promote limb perfusion and muscle regeneration to the greatest extent possible, and thus ameliorate diabetic
limb ischemia to a greater degree than conventional allogeneic or autologous MSC. We will test the central
hypothesis by addressing the following two specific aims: Aim 1. Determine the optimal hydrogel type, MSC
preparation, dose, and route of injection for limb perfusion and muscle regeneration using polygenic diabetic
and diet-induced obese (DIO) mouse models of CLTI. Studies will compare human allogeneic BMD-MSC to a
vBA-MSC preparation in both unassociated and alginate hydrogel encapsulated forms. Aim 2. Determine the
molecular mechanisms of encapsulated MSC-mediated ischemic muscle regeneration. The working hypothesis
is that phenotypic alterations induced by alginate encapsulation will result in upregulating expression of MSC
molecules, such as IL-10 and IL-33, that stimulate Treg function and promote the M1 to M2-biased
macrophage phenotype switch which in turn stimulates muscle regeneration via progenitor cell function. This
aim will utilize gain and loss of function of cytokine production to determine the role of MSC-stimulated muscle
regeneration. The successful completion of this study will identify the most effective preparation of MSC and
determine primary mechanisms by which exogenous MSC mediate muscle regeneration in diabetics.
Identifying the impact of MSC on the molecular pathways regulating muscle regeneration will allow
development of alternative non-cell-based therapies to treat diabetic CLTI patients.
