A new method for encapsulation of living cells: Preliminary results with PC12 cell line

Société Kappa Biotech, Montauban, France.
Journal of Microencapsulation (Impact Factor: 1.59). 01/2001; 18(3):323-34. DOI: 10.1080/02652040010018092
Source: PubMed


A new method is described for encapsulation of living cells. PC12 rat adrenal pheochromocytoma cells, which have been shown to synthesize, store and release dopamine were employed. The particles are made first and the cells then incorporated in a gentle mechanical procedure. The morphology (by light and electron microscopic observation), stability, rheology, texture and permeability of these microcapsules provided by Kappa Biotech were investigated. Membrane permeability studies demonstrated exclusion of 69,000 Da human serum albumin, but equilibrium of D-glucose and inulin was within 24h, indicating a molecular weight cut-off in the 5000-70,000 Da range. The viability and the function of the encapsulated cells were evaluated by measuring the spontaneous release of dopamine by high performance liquid chromatography with electrochemical detection. The results show that dopamine-secreting cells can be sequestered in a semi-permeable capsule and still display good viability and proliferation for at least 1 month.

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    • "Complex coacervation of water-soluble polyelectrolytes allows the formation of microcapsules in aqueous solutions without the involvement of any organic solvents. Both polyanionic and polycationic water-soluble polymers have been used to form the positivelycharged or negatively-charged core of the microcapsules containing living cells (Chia et al., 2000, 2002; Gray, 1997; Hagihara et al., 1997; Hasan et al., 2000; Hunkeler et al., 1997; Mercier et al., 2001; Miyoshi et al., 1996). Recently, positively-charged 3D microenvironments such as those formed with chitosan have attracted much attention because of their ability to support cell proliferation/functions (Li et al., 2003; Park et al., 2003; Wang et al., 2003; Zuekubski and Aebischer, 1994). "
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    ABSTRACT: A new class of microcapsules was prepared under physiological conditions by polyelectrolyte complexation between two oppositely-charged, water-soluble polymers. The microcapsules consisted of an inner core of half N-acetylated chitosan and an outer shell of methacrylic acid (MAA) (20.4%)-hydroxyethyl methacrylate (HEMA) (27.4%)-methyl methacrylate (MMA) (52.2%) (MAA-HEMA-MMA) terpolymer. Both 400 and 150 kDa half N-acetylated chitosans maintained good water solubility and supplied enough protonated amino groups to coacervate with terpolymer at pH 7.0-7.4, in contrast to other chitosan-based microcapsules which must be prepared at pH <6.5. The viscosity of half N-acetylated chitosan solutions between 80 and 3000 cPas allowed the formation of microcapsules with spherical shape. Molar mass, pH and concentration of half N-acetylated chitosan, and reaction time, influenced the morphology, thickness and porosity of the microcapsules. Microcapsules formed with high concentration of half N-acetylated chitosan exhibited improved mechanical stability, whereas microcapsules formed with low concentration of half N-acetylated chitosan exhibited good permeability. This 3D microenvironment has been configured to cultivate sensitive anchorage-dependent cells such as hepatocytes to maintain high level of functions.
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    • "The amounts of catecholamines released by the PCC and BCC cultures were determined by measuring of epinephrine or norepinephrine with electrochemical detection after extraction by ion-exchange column and separation by reversephase HPLC as previously described (Duplan et al., 2000; Mercier et al., 2001). Secreted methionine-enkephalin levels were measured with a commercially available Met-enkephalin radioimmu- noassay 125 I RIA kit (Amersham Bioscience Europe, Orsay, France). "
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    ABSTRACT: Adrenal medullary chromaffin cells synthetize and secrete a combination of pain-reducing neuroactive compounds including catecholamines and opioid peptides. Previous reports have shown that implantation of chromaffin cells into the spinal subarachnoid space can reduce both acute and chronic pain in several animal models. We recently demonstrated that human chromaffin cell grafts in the cerebrospinal fluid (CSF) could alleviate intractable cancer pain after failure of systemic opiates. However, wider application of this approach was limited by the limited availability of allogeneic donor material. Alternatively, chromaffin cells from xenogeneic sources such as bovine adrenal medulla were successful in the experimental treatment of pain, but recent concern over risk of prion transmission precluded use of bovine grafts in human clinical trials. The objective of the present study was to investigate the possibility of developing a new xenogeneic porcine source of therapeutic chromaffin cells because this strategy is currently considered the safest for transplantation in man. In the present study, we report the isolation and the characterization of primary porcine chromaffin cells (PCC) compared to bovine cells. We show, for the first time, that these cells grafted in the rat subarachnoid space can attenuate pain-related behaviors as assessed by the formalin test, a model of tonic pain. Moreover, in addition to behavioral studies, immunohistochemical analysis revealed robust survival of chromaffin cells 35 days after transplantation. Taken together, these results support the concept that porcine chromaffin cells may offer an alternative xenogeneic cell source for transplants delivering pain-reducing neuroactive substances.
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    ABSTRACT: A copolymer of N-isopropylacrylamide (98 mole% in the feed) and acrylic acid poly(N-isopropylacrylamide-co-acrylic acid) [p(NiPAAm-co-AAc)], and the adhesion molecule, an Arg-Gly-Asp (RGD)-incorporated hydrogel, were used to entrap pheochromocytoma cells (PC12). In a 28-d culture period, the PC12 cells in the RGD-conjugated gel maintained higher viability and produced dopamine at constant rates, while there was lower cell viability and less dopamine secretion by PC12 cells in p(NiPAAm-co-AAc). PC12 cells cultured in the RGD-conjugated gel would constitute a potentially useful three-dimensional cell system for application in nerve regeneration.
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