Temporal monitoring of pO2 in stem cell cultures : a potential tool for directed differentiation

Problem The objective of this in-vivo study is to investigate the histologic effects of MSC and PRP therapy on ligament healing in a rat medial collateral ligament (MCL) injury model. Background Cell therapy for cartilage and ligament injuries has become a widely debated topic in the orthopaedic literature. Adult mesenchymal stem cells (MSCs) are of interest given their high capacity for self-renewal and multipotency to differentiate into chondrocytes and tenocytes. Platelet rich plasma (PRP) has been reported to promote collagen synthesis and cell proliferation, influencing the healing of ligaments and cartilage. Hypothesis We hypothesize that the addition of MSCs to an injured Rat MCL will show increase collagen regeneration when compared to controls and those MCLs treated with PRP. Research MCLs of 20 female Sprague rats were bilaterally transected and treated with either saline or 1 of 3 treatment groups; (1) MSCs (10^5cells), (2) MSCs + PRP, and (3) PRP. Ratswere sacrificed 16 days post-surgery and the MCLs harvested, fixed in formalin, processed routinely and stained with H&E and Alcian blue. Histological analysis was performed by a pathologist blinded to the treatment groups, and the tissue specimens were graded based on the degree of cellularity (0-+3), change in collagen representing new or regenerative collagen fibers (0-+3), vascularity (0-+2) and inflammation (0-+3). Statistical analysis was performed using Analysis of variance (ANOVA) and Fischers exact test. Observations The degree of cellularity and change in collagen fibers were statistically significantly increased between the four groups (p<.0032). Paired comparisons between groups demonstrated that all treatment groups had increased cellularity and collagen changes when compared to controls but not between the experimental groups. Alcian blue staining revealed increased extracellular matrix within treatment groups (MSCs= avg. grade of 1.33, MSCs & PRP = avg. grade of 2.67 and PRP = avg. grade of 2) when compared to the control (avg. grade of 1). The addition of MSCs and/or PRP to an injured MCL increases cellularity and the regeneration of collagen fibers in the setting of an acute MCL injury when compared to controls. While not significant, there was a trend towards even higher cellularity and collagen fiber regeneration in the treatment group treated with MSC and PRP, indicating a possible potentiation effect between the two treatments. This study suggests improved ligament healing with MSC and PRP treatment and that biomechanical testing is warranted to explore whether this treatment will result in improved biomechanical properties of MCL injuries

Single and double encapsulation of cell and cell clusters for regenerative medicine and cell transplantation

Problem Conventional alginate microencapsulation has been tested in pre-clinical animal models, including non-human primates (NHPs) and in humans with unsatisfactory results. This is likely because of the large (600-1000\u3bcm) diameter of standard microcapsules that impairs oxygen and nutrients exchange and causes delays in secretion of trophic factors following cell stimulation by the host; and limited choice for transplantation sites due to the high total volume of the encapsulation product; finally long-term applications might fail due to the poor mechanical properties and poor in vivo stability of conventional encapsulation materials. Background Successful encapsulation of autologous, allogeneic or xenogeneic cells with a semipermeable barrier can be beneficial for many therapeutic applications, including improving preservation and shipment of coated cells and guaranteeing immunoprotection after transplantation for regenerative medicine. Especially in cell transplantation for treatment of autoimmune diseases, including type-1 diabetes, immunoisolating properties of capsules might allow transplantation in absence of or with reduced systemic immunosuppression. Hypothesis We hypothesize that by developing novel encapsulation technologies that address most concerns associated with conventional cell microencapsulation we can turn cell encapsulation into an effective and safe therapy and that by designing the procedure to be versatile, reproducible, and scalable we can facilitate its translatability to clinical applications. Research As an alternative to conventional alginate microencapsulation, we developed two novel encapsulation technologies: a \u201cconformal coating\u201d microencapsulation and a \u201cdouble conformal coating\u201d encapsulation and compared to alginate micrencapsulation. Conformal coating is achieved by exploiting a fluid dynamic principle that allows to wrap cell clusters with a 10-20\u3bcm layer of polymer, minimizing diffusion barriers and size of encapsulated cell product and allowing a wider choice of transplantation sites. The double coating technology confers a conformal coating as an extra layer to improve mechanical, permselectivity and immunoisolating properties of enclosed alginate microcapsules (400-600\u3bcm in diameter) that are obtained with a conventional micro droplet generator. Our combined encapsulation methods are versatile because they can be applied to any coating materials and to any cell type and are reproducible and scalable for clinical translation. Observations We have shown that both the conformal coating and alginate microencapsulation technologies do not to affect cell viability and function (i.e. secretion of trophic factor upon stimulation) of cell clusters of different origins and we are currently optimizing the double coating technology by combining the two encapsulation methods. We strongly believe that our platform for cell encapsulation will allow us to compare different encapsulation technology and tailor them for different therapeutic applications as a valid and may be superior alternative to standard alginate microencapsulatio