title: Tunable Volumetric Density and Porous Structure of Spherical Poly-ε-caprolactone Microcarriers, as Applied in Human Mesenchymal Stem Cell Expansion.
abstract: Polymeric microspheres may serve as microcarrier (MC) matrices, for the expansion of anchorage-dependent stem cells. They require surface properties that promote both initial cell adhesion and the subsequent spreading of cells, which is a prerequisite for successful expansion. When implemented in a three-dimensional culture environment, under agitation, their suspension under low shear rates depends on the MCs having a modest negative buoyancy, with a density of 1.02-1.05 g/cm3. Bioresorbable poly-ε-caprolactone (PCL), with a density of 1.14 g/cm3, requires a reduction in volumetric density, for the microspheres to achieve high cell viability and yields. Uniform-sized droplets, from solutions of PCL dissolved in dichloromethane (DCM), were generated by coaxial microfluidic geometry. Subsequent exposure to ethanol rapidly extracted the DCM solvent, solidifying the droplets and yielding monodisperse microspheres with a porous structure, which was demonstrated to have tunable porosity and a hollow inner core. The variation in process parameters, including the molecular weight of PCL, its concentration in DCM, and the ethanol concentration, served to effectively alter the diffusion flux between ethanol and DCM, resulting in a broad spectrum of volumetric densities of 1.04-1.11 g/cm3. The solidified microspheres are generally covered by a smooth thin skin, which provides a uniform cell culture surface and masks their internal porous structure. When coated with a cationic polyelectrolyte and extracellular matrix protein, monodisperse microspheres with a diameter of approximately 150 μm and densities ranging from 1.05-1.11 g/cm3are capable of supporting the expansion of human mesenchymal stem cells (hMSCs). Validation of hMSC expansion was carried out with a positive control of commercial Cytodex 3 MCs and a negative control of uncoated low-density PCL MCs. Static culture conditions generated more than 70% cell attachment and similar yields of sixfold cell expansion on all coated MCs, with poor cell attachment and growth on the negative control. Under agitation, coated porous microspheres, with a low density of 1.05 g/cm3, achieved robust cell attachment and resulted in high cell yields of ninefold cell expansion, comparable with those generated by commercial Cytodex 3 MCs.
authors.affiliation: Institute of Materials Research and Engineering, A*STAR (Agency for Science, Technology and Research) , 2 Fusionopolis Way, Innovis, #08-03, 138634, Singapore.
authors.forename: Jian
authors.identifier: 0000-0002-6681-458X
authors.initials: J
authors.lastname: Li
authors.affiliation: Bioprocessing Technology Institute, A*STAR (Agency for Science, Technology and Research) , 20 Biopolis Way, #06-01, 138668, Singapore.
authors.forename: Alan Tin-Lun
authors.initials: AT
authors.lastname: Lam
authors.affiliation: Institute of Materials Research and Engineering, A*STAR (Agency for Science, Technology and Research) , 2 Fusionopolis Way, Innovis, #08-03, 138634, Singapore.
authors.forename: Jessica Pei Wen
authors.initials: JP
authors.lastname: Toh
authors.affiliation: Bioprocessing Technology Institute, A*STAR (Agency for Science, Technology and Research) , 20 Biopolis Way, #06-01, 138668, Singapore.
authors.forename: Shaul
authors.initials: S
authors.lastname: Reuveny
authors.affiliation: Bioprocessing Technology Institute, A*STAR (Agency for Science, Technology and Research) , 20 Biopolis Way, #06-01, 138668, Singapore.
authors.forename: Steve Kah-Weng
authors.initials: SK
authors.lastname: Oh
authors.affiliation: Institute of Materials Research and Engineering, A*STAR (Agency for Science, Technology and Research) , 2 Fusionopolis Way, Innovis, #08-03, 138634, Singapore.
authors.forename: William R
authors.identifier: 0000-0001-8150-9033
authors.initials: WR
authors.lastname: Birch
mesh_terms: D002470:Cell Survival; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000166:cytology*; D008752:Methylene Chloride / Q000737:chemistry; D008863:Microspheres; D015394:Molecular Structure; D010316:Particle Size; D011091:Polyesters / Q000737:chemistry*; D016062:Porosity; D013499:Surface Properties
chemical_list: D011091:Polyesters; C016240:polycaprolactone; D008752:Methylene Chloride