The research aims at developing implants (hydrogels, polymeric scaffolds, microcarriers) delivering growth factors, drugs and cells that provide sustained delivery of bioactive molecules, support survival, infiltration and proliferation of cells for tissue engineering, and in particular spinal cord injury.
Our group has gained expertise in drug delivery to the spinal cord that we combined with transplantation of adult mesenchymal stem cells; more particularly human dental stem cells. Indeed, human dental stem cells display superior neural stem cell properties than bone marrow-derived mesenchymal stem cells since they originate from the neural crest.
We have first evaluated the impact of growth factor delivery encapsulated in micro- and nanoparticles from injectable hydrogels. Then, we decided to explore the therapeutic potential of stem cells from the apical papilla (SCAP) for spinal cord injury. We tested different ways of administration in rat spinal cord injury models.
We evaluated the effect of VEGF and GDNF delivery, free or encapsulated, from an alginate:fibrinogen hydrogel injected in a rat spinal cord hemisection model. Local VEGF delivery from alginate:fibrinogen hydrogel gelifying in situ induced angiogenesis and neurite growth but no functional improvement. However, local GDNF delivery significantly improved functional recovery of rats. Indeed, the animals treated with free GDNF-loaded hydrogel experienced superior functional recovery compared to the animals treated with GDNF microsphere-loaded hydrogels and non-treated animals (in collaboration with Prof. Blanco-Prieto, Navarra University, Spain, Drs Schakman and Deumens, UCL, IoNS).
As a source of human mesenchymal stem cells, we selected human dental stem cells of the apical papilla (SCAP) due to their neural crest origin but also because they are easily accessible (obtained from extracted wisdom tooth roots). They express numerous neuronal markers, display enhanced neural stem cell properties compared to bone marrow-derived mesenchymal stem cells and possess higher proliferation and differentiation rates compared to dental pulp stem cells. The studies performed on SCAP have been done in collaboration with Prof. Diogenes (San Antonio University, San Antonio, USA).
We used 3 strategies to deliver SCAP: in their original niche (apical papilla), incorporated in hydrogels or seeded on PLGA microcarriers (Figure 1).
Figure 1: SCAP delivery strategies.
a. Implantation of a whole papilla in aspinal cord lesion
We hypothesized that isolating and expanding SCAP would change their properties and characteristics while keeping them in their niche would not. When rats were treated with a human apical papilla implanted as a whole, we observed a significant improvement of motor function compared to the control groups (lateral hemisection model) (collaboration with Prof. Leprince, LDRI, UCL) (Figure 9). This might be explained by injury stabilization (papilla still in place after 6 weeks) and by the action of the cells present in the papilla (cells positive for human mitochondria in the papilla after 6 weeks).
b. Incorporation of SCAP in hydrogels
To deliver and maintain SCAP at the lesion site, we have selected injectable hydrogels (fibrin, alginates and Corgel®). No study previously compared the impact of hydrogel properties on SCAP (collaboration with Prof. Dupont, IMCN, UCL). We observed that fibrinogen concentration in fibrin hydrogel impacted SCAP neurodifferentiation in vitro, but also proliferation and angiogenesis in vivo. When comparing different alginates and Corgel®, not a single property, but the appropriate combination of surface and mechanical characteristics dictates SCAP fate.
We also studied the influence of decellularized extracellular matrix-based hydrogels (dECMh) originating from different organs (bone, dentin and spinal cord) (Erasmus Mundus NanoFar, collaboration with Prof. Shakesheff and Dr. White, University of Nottingham, UK). dECMh are thermosensitive (gelation at 37°C), contain preserved cell adhesion sites and active molecules specific of the organ of origin. We demonstrated that dECMh origin impacted hydrogel properties and SCAP viability and neuronal gene expression, spinal cord dECMh being the most favorable for neural differentiation.
c. Development of growth factor loaded microcarriers for SCAP delivery
Another strategy to deliver cells is to seed them on microcarriers designed to support cell adhesion and viability and to deliver growth factors. In the scope of an Erasmus Mundus project, we co-supervised a PhD thesis with Prof. Montero-Menei (Angers University, FR) that aimed to optimize the formulation of BDNF-loaded pharmacologically active microcarriers (PAM). We demonstrated that PAM supported the viability of mesenchymal stem cells and impacted their secretome and proteome. BDNF-PAM and SCAP were then combined and injected in a spinal cord contusion model. An improvement of rat locomotor function, a decrease of inflammation and neuroprotection were observed when SCAP where implanted associated with BDNF-PAM.