Abstract
The combination of protein-coated graphene oxide (GO) and microencapsulation technology has moved a step forward in the challenge of improving long-term alginate encapsulated cell survival and sustainable therapeutic protein release, bringing closer its translation from bench to the clinic. Although this new approach in cell microencapsulation represents a great promise for long-term drug delivery, previous studies have been performed only with encapsulated murine C2C12 myoblasts genetically engineered to secrete murine erythropoietin (C2C12-EPO) within 160 µm diameter hybrid alginate protein-coated GO microcapsules implanted into syngeneic mice. Here, we show that encapsulated C2C12-EPO myoblasts survive longer and release more therapeutic protein by doubling the micron diameter of hybrid alginate-protein-coated GO microcapsules to 380 µm range. Encapsulated mesenchymal stem cells (MSC) genetically modified to secrete erythropoietin (D1-MSCs-EPO) within 380 µm-diameter hybrid alginate-protein-coated GO microcapsules confirmed this improvement in survival and sustained protein release in vitro. This improved behavior is reflected in the hematocrit increase of allogeneic mice implanted with both encapsulated cell types within 380 µm diameter hybrid alginate-protein-coated GO microcapsules, showing lower immune response with encapsulated MSCs. These results provide a new relevant step for the future clinical application of protein-coated GO on cell microencapsulation.
Highlights
Alginate microencapsulated cell-based therapies can provide the sustained release of therapeutic products in the treatment of a variety of diseases
We aimed to study the behavior of encapsulated genetically modified C2C12 myoblasts within hybrid alginate-protein-coated graphene oxide (GO) microcapsules with diameters higher than 300 mm, trying to determine if an increment in size would affect the benefits observed with 160 mm diameter hybrid microcapsules (Saenz Del Burgo et al, 2017)
Metabolic activity, cell membrane integrity and protein release of encapsulated C2C12-EPO cells in 160 and 380 mm diameter hybrid microcapsules, performed in a pneumatic (Bioencapsulation portable platform CellenaVR ) and an electrostatic (NiscoVR ) atomization generator, respectively (Supplementary Figure 1). We generated both diameter hybrid microcapsules at a concentration of 50 mg/ml of GO, comparing them with microcapsules without GO, since this GO concentration within alginate microcapsules has previously shown to be the optimal concentration for myoblasts survival enhancement (Ciriza et al, 2015)
Summary
Alginate microencapsulated cell-based therapies can provide the sustained release of therapeutic products in the treatment of a variety of diseases. Several challenges in cell therapy using microencapsulated cells need to be overcome, such as the decrease of the dying cell number inside the microcapsules or the enhancement of the sustained protein release from encapsulated cells, achieving long-lasting treatments. In this regard, we combined cell microencapsulation technology with graphene oxide (GO), a highly oxidized form of chemically modified graphene, consisting of a single atom thick layer of graphene sheets with carboxylic acid, epoxide, and hydroxyl groups (Goenka et al, 2013). We incorporated different concentrations of GO into 160 mm diameter alginate microcapsules, showing that GO concentrations between 25 and 50 mg/ ml increased the viability, metabolic activity, membrane integrity, and erythropoietin (EPO) release of encapsulated murine C2C12 myoblasts genetically engineered to secrete murine erythropoietin (C2C12-EPO) (Ciriza et al, 2015), improving even more by the formation of a protein bio-corona with fetal bovine serum (FBS) (Saenz Del Burgo et al, 2017)
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