Microparticles supporting cells and active substances
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example 1
General Presentation of MPAs for Cell Grafts
[0044] The non-limiting examples below pertain to the field of neurotransplantation (grafts of nerve cells in the central nervous system). Biocompatible, biodegradable MPAs of a diameter of 60 μm were prepared. They were constituted of PLGA and coated with a fine layer of synthetic adherence molecules (poly-D-lysine and fibronectin-like agent) and release NGF, a neurotrophic factor, on a continuous basis (for at least 15 days). Cells responding to NGF, such as the PC12 cell line capable of secreting dopamine and of differentiating itself into sympathetic-like neurons under the effect of this factor were used. The efficacy of these MPAs was evaluated in vivo successfully using an animal model of Parkinson's disease.
[0045] Neurotransplantations began clinically in the 1980s essentially in the context of Parkinson's disease (Menei et al., 1991a, 1991b). They continue at present in the form of clinical research and remain essentially limited...
example 2
Preparation of PC12 / NGF MPAs
1) Preparation of the Microparticles
[0052] The microparticles were produced by double emulsion (H / L / H) evaporation / extraction of solvent.
[0053] The aqueous phase was constituted of 60 μl of citrate buffer (16 mM, pH 6), 2.5 mg of HSA (or other molecule having a surface-active power), 90 μl of PEG 400 and 100 μg of NGF. 50 mg-100 mg of PLGA 37.5 / 25 (poly D,L-lactide-co-glycolide, Mw 21,000 Da, I=1.7) or another biocompatible, biodegradable polymer were dissolved in 2 ml of an organic solution constituted of 75% of dichloromethane and 25% acetone. The primary emulsion was made from these two phases by sonication at 0-4° C. (15 s, 5-6 W). This primary emulsion was added under mechanical agitation (200 rpm) to 30-150 ml of a polyvinyl alcohol solution (0.8-4.5%, 4-8° C.) containing 10% (W / V) of NaCl containing 0 to 2% of dichloromethane. Agitation was maintained for 1 to 7 minutes. The secondary emulsion was then poured into 400 ml of a 10% aqueous soluti...
example 3
Results of the NGF / PC12 MPAs on Animals
[0061] The MPAs release NGF in a controlled, prolonged manner. In fact, the initial results of the kinetic of release in vitro showed that for 200 μg of encapsulated NGF, 15% was released in a continuous manner during the first two weeks. The implantation of 0.5 mg of MPAs thus leads to the release of 5-10 ng of NGF per day which is in agreement with the quantities necessary for action of NGF on the cells.
[0062] After implementation in the denervated striatum of parkinsonian rats, the PC12 cells remained strongly adhered on the microparticles. The transported cells continued to express tyrosine hydroxylase and were thus capable of producing dopamine. The microparticles still had not degraded and still functioned as cell supports two weeks after implantation. In fact, the microparticles were still spherical and had a rather smooth appearance without vacuolization pores. Generally speaking, the cells adhered to the microparticles with or withou...
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