Human dental stem cells of the apical papilla associated to BDNF-loaded pharmacologically active microcarriers (PAMs) enhance locomotor function after spinal cord injury.

There is no treatment for spinal cord injury (SCI) that fully repairs the damages. One strategy is to inject mesenchymal stem cells around the lesion to benefit from their immunomodulatory properties and neuroprotective effect. Our hypothesis was that the combination of dental stem cells from the apical papilla (SCAP) with pharmacologically active microcarriers (PAMs) releasing brain-derived neurotrophic factor (BDNF) would improve rat locomotor function by immunomodulation and neuroprotection. BDNF-PAMs were prepared by solid/oil/water emulsion of poly(L-lactide-co-glycolide) and nanoprecipitated BDNF and subsequent coating with fibronectin. SCAP were then seeded on BDNF-PAMs. SCAP expression of neuronal and immunomodulatory factors was evaluated in vitro. SCAP BDNF-PAMs were injected in a rat spinal cord contusion model and their locomotor function was evaluated by Basso, Beattie, and Bresnahan (BBB) scoring. Impact on inflammation and neuroprotection/axonal growth was evaluated by immunofluorescence. Culture on PAMs induced the overexpression of immunomodulatory molecules and neural/neuronal markers. Injection of SCAP BDNF-PAMs at the lesion site improved rat BBB scoring, reduced the expression of inducible nitric oxide synthase and increased the expression of ßIII tubulin, GAP43, and 5-HT. These results confirm the suitability and versatility of PAMs as combined drug and cell delivery system for regenerative medicine applications but also that BDNF-PAMs potentialize the very promising therapeutic potential of SCAP in the scope of SCI.

A Combinatorial Cell and Drug Delivery Strategy for Huntington's Disease Using Pharmacologically Active Microcarriers and RNAi Neuronally-Committed Mesenchymal Stromal Cells.

For Huntington's disease (HD) cell-based therapy, the transplanted cells are required to be committed to a neuronal cell lineage, survive and maintain this phenotype to ensure their safe transplantation in the brain. We first investigated the role of RE-1 silencing transcription factor (REST) inhibition using siRNA in the GABAergic differentiation of marrow-isolated adult multilineage inducible (MIAMI) cells, a subpopulation of MSCs. We further combined these cells to laminin-coated poly(lactic-co-glycolic acid) PLGA pharmacologically active microcarriers (PAMs) delivering BDNF in a controlled fashion to stimulate the survival and maintain the differentiation of the cells. The PAMs/cells complexes were then transplanted in an ex vivo model of HD. Using Sonic Hedgehog (SHH) and siREST, we obtained GABAergic progenitors/neuronal-like cells, which were able to secrete HGF, SDF1 VEGFa and BDNF, of importance for HD. GABA-like progenitors adhered to PAMs increased their mRNA expression of NGF/VEGFa as well as their secretion of PIGF-1, which can enhance reparative angiogenesis. In our ex vivo model of HD, they were successfully transplanted while attached to PAMs and were able to survive and maintain this GABAergic neuronal phenotype. Together, our results may pave the way for future research that could improve the success of cell-based therapy for HDs.

Pharmacologically active microcarriers delivering BDNF within a hydrogel: Novel strategy for human bone marrow-derived stem cells neural/neuronal differentiation guidance and therapeutic secretome enhancement.

Stem cells combined with biodegradable injectable scaffolds releasing growth factors hold great promises in regenerative medicine, particularly in the treatment of neurological disorders. We here integrated human marrow-isolated adult multilineage-inducible (MIAMI) stem cells and pharmacologically active microcarriers (PAMs) into an injectable non-toxic silanized-hydroxypropyl methylcellulose (Si-HPMC) hydrogel. The goal is to obtain an injectable non-toxic cell and growth factor delivery device. It should direct the survival and/or neuronal differentiation of the grafted cells, to safely transplant them in the central nervous system, and enhance their tissue repair properties. A model protein was used to optimize the nanoprecipitation conditions of the neuroprotective brain-derived neurotrophic factor (BDNF). BDNF nanoprecipitate was encapsulated in fibronectin-coated (FN) PAMs and the in vitro release profile evaluated. It showed a prolonged, bi-phasic, release of bioactive BDNF, without burst effect. We demonstrated that PAMs and the Si-HPMC hydrogel increased the expression of neural/neuronal differentiation markers of MIAMI cells after 1week. Moreover, the 3D environment (PAMs or hydrogel) increased MIAMI cells secretion of growth factors (b-NGF, SCF, HGF, LIF, PlGF-1, SDF-1α, VEGF-A & D) and chemokines (MIP-1α & ß, RANTES, IL-8). These results show that PAMs delivering BDNF combined with Si-HPMC hydrogel represent a useful novel local delivery tool in the context of neurological disorders. It not only provides neuroprotective BDNF but also bone marrow-derived stem cells that benefit from that environment by displaying neural commitment and an improved neuroprotective/reparative secretome. It provides preliminary evidence of a promising pro-angiogenic, neuroprotective and axonal growth-promoting device for the nervous system. STATEMENT OF SIGNIFICANCE: Combinatorial tissue engineering strategies for the central nervous system are scarce. We developed and characterized a novel injectable non-toxic stem cell and protein delivery system providing regenerative cues for central nervous system disorders. BDNF, a neurotrophic factor with a wide-range effect, was nanoprecipitated to maintain its structure and released in a sustained manner from novel polymeric microcarriers. The combinatorial 3D support, provided by fibronectin-microcarriers and the hydrogel, to the mesenchymal stem cells guided the cells towards a neuronal differentiation and enhanced their tissue repair properties by promoting growth factors and cytokine secretion. The long-term release of physiological doses of bioactive BDNF, combined to the enhanced secretion of tissue repair factors from the stem cells, constitute a promising therapeutic approach.