Abstract
Immobilization of discrete cell clusters within a partially crosslinked matrix prevents reaggregation of primary tissues and may provide a means for long-term maintenance of encapsulated cells. Dissociated bovine adrenal chromaffin (BAC) cells were suspended throughout crosslinked polyanionic microspheres previously shown to be selectively permeable. Microcapsules approximately 500 µm in diameter were seeded with: 1) three different densities of BAC cells; and 2) BAC cells suspended in Matrigel® or coseeded with a genetically modified nerve growth factor (NGF)- releasing fibroblast cell line. Each group was analyzed in vitro at 1, 4 and 8 weeks for spontaneous and potassium-evoked release of catecholamines, and maintained in vitro for up to 12 weeks for morphological observations. Over time, release of norepinephrine (NE) and epinephrine (EPI) diminished, while dopamine (DA) remained constant from the monoseeded capsules. In the coseeded group, an increase in potassium-evoked release of DA was observed from 1 to 4 weeks, and remained at that level up to 8 weeks. Encapsulated chromaffin cells retained a rounded morphology typical of undifferentiated cells. Intact chromaffin cells with well preserved and abundant secretory granules were observed ultrastructurally after 4 weeks in vitro. Small neurites from the chromaffin cells in the coseeded group were observed at 4 weeks with light microscopy, and up to 12 weeks with electron microscopy. Under static incubation conditions, 1 mM D-amphetamine resulted in a significant increase in the output of NE and DA from the coseeded capsules 8 weeks postimplantation, as compared to microcapsules loaded with chromaffin cells alone. Encapsulation within an immobilization matrix allows manipulation of the internal environment, thereby providing the ability to pre-treat cells with various factors in a non-invasive manner, which may enhance long-term cellular viability.
Highlights
Neural implantation of polymer-encapsulated tissues has been utilized as a model system for analyzing the effectiveness of diffusive release of neuroactive agents in denervated regions of the central nervous system /1,26,32/
There was no significant difference in the average diameters of microcapsules loaded with low, medium and high densities of chromaffin
The monoseeded microcapsules contained an average of 600 chromaffin cells, while the coseeded group had 450 chromaffin cells and 200 R208N.8 fibroblasts/capsule
Summary
Neural implantation of polymer-encapsulated tissues has been utilized as a model system for analyzing the effectiveness of diffusive release of neuroactive agents in denervated regions of the central nervous system /1,26,32/. Extensive neuritic outgrowths from chromaffin cells are dependent on the presence of differentiating factors, such as NGF, when maintained in culture /13,25,29/ Both adrenal medulla/6,9,15,22/and dissociated chromaffin cell/3,4,17/grafts show poor survival when transplanted into striatal parenchyma. NGF infused into the region of the adrenal medullary cell transplants has been reported to enhance survival, induce neuritic extensions, and increase the effectiveness of behavioral recovery/23/. This raises the question of whether low concentrations of NGF in a lesioned striatum /11/ contribute to the inability of the chromaffin cells to maintain a neuronal phenotype which may limit their survival, or are other factors contributing to poor intrastriatal viability? This raises the question of whether low concentrations of NGF in a lesioned striatum /11/ contribute to the inability of the chromaffin cells to maintain a neuronal phenotype which may limit their survival, or are other factors contributing to poor intrastriatal viability? The ability to deliver a chronic supply of NGF to chromaffin cells from a living system may improve graft survival and avoid the problems and limitations inherent in pumps or polymer systems/30/
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