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PubMedDate not supplied

Alginate-encapsulated producer cells: a potential new approach for the treatment of malignant brain tumors.

In recent years gene therapy has evolved as a new treatment for brain tumors, where genetically engineered cells can be used to deliver specific substances to target cells. However, clinical success has been limited due to insufficient gene transfer, lack of prolonged gene expression, and immunorejection of producer cells. These obstacles may be overcome by encapsulating producer cells into immunoisolating substances such as alginate. This may provide a stable in situ delivery system of specific proteins, which can interfere with tumor growth and differentiation. This article represents a fundamental study describing the in vitro and the in vivo behavior of alginate-encapsulated producer cells. The viability and cell cycle distribution of encapsulated NIH 3T3 cells was studied by confocal laser scanning microscopy (CLSM) and by flow cytometry. The CLSM study showed a high viability of the encapsulated NIH 3T3 cells during 9 weeks in culture. The flow cytometric analysis revealed a change in cellular ploidy after 1 week in culture, with normalization in ploidy after 3 and 9 weeks. The production of the bacterial E. coli beta-galactosidase in alginate-encapsulated BT4CnVlacZ cells was studied by x-gal staining, and the cells expressed prolonged beta-galactosidase activity. H528 hybridoma cells producing monoclonal antibodies (mAbs) against the human epidermal growth factor receptor (EGFR) were encapsulated in alginate, and the mAb release was determined. The release of mAbs stabilized around 400 ng/ml/h after 12 days in vitro. To actually demonstrate that alginate-encapsulated H528 cells potentially inhibit a heterogeneous glioma cell population, cell migration from human GaMg glioma spheroids was studied during stimulation with EGF in the presence of encapsulated H528 cells. The migration in vitro was totally inhibited in the presence of H528 encapsulated cells. Alginate beads with H528 cells were also implanted into rat brains, and after 9 weeks the distribution of mAbs within the brain was studied by immunohistochemistry. It is shown that the alginate entrapped H528 cells produce mAbs inside the brain for prolonged periods and that the mAbs are distributed within all CSF compartments. Encapsulated producer cells represent a potential delivery system for specific proteins to brain tumors. Different producer cells may be encapsulated in alginate to target phenotypic features and microenvironmental factors, which may influence the progressive growth of brain tumors.

3T3 Cells↗

PubMed2005

Calcium alginate microencapsulation of ovarian follicles impacts FSH delivery and follicle morphology.

BACKGROUND: We have previously shown that suspension culture prevents follicle flattening and maintains three-dimensional follicle architecture better than culture on flat plates. However, many of the follicles cultured in suspension do eventually rupture, as basement membrane integrity is lost and the three-dimensional structure of the follicle is altered. Therefore, the objective of this study is to support three-dimensional follicle architecture during in vitro growth of ovarian follicles through encapsulation in calcium alginate, while maintaining responsiveness to FSH stimulation. METHODS: Preantral follicles (150-160 micrometers in diameter) were isolated from the ovaries of juvenile rats and grown in culture tubes or encapsulated in calcium alginate and grown in culture tubes. Previous studies revealed that follicles maintained structural integrity but did not grow as well when encapsulated in calcium alginate. In these studies, we evaluated the effect of calcium alginate on FSH-stimulated follicle growth, survival, and morphology in suspension culture. Follicles were grown under 5 culture conditions: 1) not encapsulated; with FSH in the medium, 2) encapsulated in the absence of FSH, grown in medium without FSH, 3) encapsulated with calcium alginate containing FSH but grown in medium without FSH, 4) encapsulated without FSH but grown in medium containing FSH and 5) encapsulated with calcium alginate containing FSH and in medium containing FSH. To assess growth rates, follicles were cultured for 72 hours and analyzed for follicle size increase and DNA content. Survival analysis for encapsulated and unencapsulated follicles was performed by constructing a Kaplan Meier survival curve of daily observations of intact follicle survival. Three-dimensional architecture was assessed histologically and by analysis of the pattern of connexin 43 expression in the cultured follicles. RESULTS: In the absence of FSH, follicle diameter increased by only 6.4%. When FSH was included in the alginate bead alone or the media alone, the follicle diameter increased by 13.5% and 19.9% respectively. This was greater than follicles cultured in the absence of FSH (p < 0.05), but less than that of the FSH-treated unencapsulated follicles (p < 0.05). However, when follicles were cultured with FSH included in both the media and the bead, a 32.6% increase in follicle diameter was observed, statistically no different than the growth rate of the unencapsulated follicles grown with FSH. CONCLUSION: Microencapsulation supports three-dimensional follicle growth, but may limit access to hormones in the medium resulting in altered development compared to unencapsulated follicles. Inclusion of FSH in the alginate bead restores the follicle growth response to FSH, while also providing a scaffold of support for three-dimensional growth. The application of tissue engineering principles to the problems of follicle culture in vitro may provide advances applicable to fertility preservation in women and endangered species.

Alginates↗

PubMed2005

Calcium alginate gel as a biocompatible material for endovascular arteriovenous malformation embolization: six-month results in an animal model.

OBJECTIVE: We sought to expand our assessment of calcium alginate as an embolic agent in an animal model of a cerebral arteriovenous malformation (AVM). The objective of this study was to assess the long-term biocompatibility and stability of calcium alginate in AVM swine models that survived from 1 to 6 months. METHODS: The swine model included a carotid-jugular anastomosis to redirect flow to the rete mirabile (RM), thereby simulating flow to an AVM. Alginate and the reactive component, calcium chloride, were injected from double-lumen or concentric-tube microcatheters to form an occlusion of the RM feeding vessel and the inferior portion of the RM. RESULTS: Angiography and histology verified complete occlusion of the RM feeding vessel for up to 6 months in eight of nine swine. Blood flow remained open to the superior portion of the RM and the circle of Willis. No evidence of downstream calcium alginate gel was seen in the follow-up angiograms or the histological preparations of the circle of Willis. A minor bioactive response to the alginate gel was noted at 1 month, yet no degenerative or inflammatory response was seen. At 6 months, there was moderate fibrous tissue around the alginate, which further sealed off flow to the embolized areas of the RM. CONCLUSION: Over a period of 6 months, calcium alginate was an effective endovascular occlusion material that blocked blood flow to the inferior portion of the RM. The chronic AVM model verified the long-term stability and biocompatibility of calcium alginate.

Alginates↗

PubMed2005

Method for quantitative determination of spatial polymer distribution in alginate beads using Raman spectroscopy.

A new method based on Raman spectroscopy is presented for non-invasive, quantitative determination of the spatial polymer distribution in alginate beads of approximately 4 mm diameter. With the experimental setup, a two-dimensional image is created along a thin measuring line through the bead comprising one spatial and one spectral dimension. For quantitative analysis of the Raman spectra, the method of indirect hard modeling was applied to make use of the information contained in the entire recorded spectra. For quantification of the alginate signals from within the beads, a calibration curve acquired from sodium alginate solutions was used after it was shown that only negligible differences occur between signals from alginate solutions and alginate gels. The distribution of alginate over the bead gel matrix was acquired with high spatial (51 microm) and time (12 s) resolution. The inhomogeneous distribution obtained using the new measuring technique is qualitatively in excellent agreement with data from the literature. In contrast to known measuring techniques, correct quantitative information about the spatial polymer distribution within the matrix was derived. It gave an alginate mass fraction of approximately 0.045 g/g at the edges and 0.02 g/g in the center of the beads. Next to the determination of mere polymer concentrations, the excellent time resolution of the presented method will enable investigation of the dynamic process of gel formation and it will also serve as a basis for investigation of mass transfer of small diffusing molecules in alginate matrices.

Alginates↗

PubMed2003

Structure of dried cellular alginate matrix containing fillers provides extra protection for microorganisms against UVC radiation.

Soil microorganisms in general and biocontrol agents in particular are very sensitive to UV light. The packaging of biocontrol microorganisms into cellular solids has been developed as a means of reducing loss caused by exposure to environmental UV radiation. The bacterial and fungal biocontrol agents Pantoea agglomerans and Trichoderma harzianum were immobilized in freeze-dried alginate beads containing fillers and subjected to 254 nm UV radiation (UVC). Immobilization of cells in freeze-dried alginate-glycerol beads resulted in greater survival after UV irradiation than for a free cell suspension. Adding chitin, bentonite or kaolin as fillers to the alginate-glycerol formulation significantly increased bacterial survival. Immobilization in alginate-glycerol-kaolin beads resulted in the highest levels of survival. The transmissive properties of the dried hydrocolloid cellular solid had a major influence on the amount of protection by the cell carrier. Dried alginate matrix (control) transmitted an average of 7.2% of the radiation. Filler incorporation into the matrix significantly reduced UV transmission: Alginate with kaolin, bentonite and chitin transmitted an average of 0.15, 0.38 and 3.4% of the radiation, respectively. In addition, the filler inclusion had a considerable effect on the bead's average wall thickness, resulting in a approximately 1.5- to threefold increase relative to beads based solely on alginate. These results suggest that the degree of protection of entrapped microorganisms against UVC radiation is determined by the UV-transmission properties of the dried matrix and the cellular solid's structure. It is concluded that for maximum protection against UV-radiation-induced cell loss, biocontrol microorganisms should be immobilized in alginate-glycerol beads containing kaolin.

Alginates↗

PubMedDate not supplied

Liposome-encapsulated alginate: controlled hydrogel particle formation and release.

Large unilamellar liposomes of dipalmitoyl phosphatidylcholine have been employed as reaction sites for calcium alginate gelation. Encapsulation of sodium alginate was accomplished by extrusion of phospholipid dispersions through polycarbonate filter of uniform pore size followed by incubation with high concentrations of calcium chloride. The diffusion of calcium into the liposome interior resulted in alginate gelatin within the liposome. Detergent treatment of the liposomes resulted in solubilization of the lipid bilayer with subsequent release of the alginate beads which were measured by laser light scattering and electron microscopy. The release profiles of both the liposomes with entrapped alginate beads and the alginate beads (released by detergent treatment of the liposomes) were determined using cytochrome-c as a marker for release. The release profiles show a rapid release of cytochrome-c over the first 2 for both preparations with slower subsequent release rates for the liposomes with encapsulated alginate beads. The similar early profile may be due to the release of cytochrome-c from bound calcium alginate adsorbed to the outer leaflet of the liposome. This is also supported by calorimetric results which indicate a marked reduction in the enthalpy of the main gel to liquid crystalline phase transition of multilamellar and large unilamellar dispersions of the lipid. This method results in the fabrication of easily controlled, unimodal submicron hydrogel particles which may be used for controlled-release applications.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed1997

A highly sensitive model for quantification of in vivo tumor angiogenesis induced by alginate-encapsulated tumor cells.

A remarkable approach to a specific tumor angiogenesis model in vivo is the use of alginate implants encapsulating tumor cells. However, this previously reported approach has often been questioned because of doubts regarding the relevance of hemoglobin at the alginate implant as a parameter of vascularization. In the present investigation, we examined whether or not the use of the blood pool agents FITC-dextran of high molecular weight would significantly improve the determination of vascularization at the alginate implant. In our experiments, we found a rapid distribution of FITC-dextran within the blood circulation of mice after i.v. bolus injection. The amount of FITC-dextran within alginate implants strongly correlated with the number of LL2 carcinoma cells or B16/F10 cells encapsulated. Even a low number of 10(3) cells per alginate implant led to a significantly increased accumulation of FITC-dextran. A more than 10-fold stimulation above that of controls was found with alginate implants containing 10(4) LL2 or B16/F10 tumor cells. Using the investigational compound AGM-1470 in different treatment schedules, we found that quantification of alginate implant anglogenesis with FITC-dextran is a sensitive method for the determination of angiogenesis inhibition. In conclusion, our results demonstrated that the use of FITC-dextran enables highly sensitive, quantitative measurement of blood vessel formation by alginate implants.

Alginates↗

PubMedDate not supplied

Evaluation of a collagen-alginate wound dressing in the management of diabetic foot ulcers.

Efficacy and safety of a collagen-alginate topical wound dressing (FIBRACOL Collagen-Alginate Wound Dressing) in the treatment of diabetic foot ulcers was compared with that of regular gauze moistened with normal saline. Seventy-five patients with foot ulcers were assigned randomly in a 2:1 ratio to the collagen-alginate test dressing or the gauze dressing. At the end of the study, the mean percent reduction of the wound area was 80.6% +/- 6% in the collagen-alginate dressing group and 61.1% +/- 26% in the gauze dressing group (p = .4692). Thirty-nine (78%) patients treated with the collagen-alginate dressing achieved > or = 75% wound area reduction, compared with 15 (60%) of gauze-treated patients. Complete healing was achieved in 24 (48%) of the collagen-alginate dressing group and 9 (36%) of the gauze dressing group. Wound size, when averaged over the 8-week period and with the duration of the ulcer taken into account, was reduced significantly in the collagen-alginate dressing group, as compared with the gauze dressing group (df = 1, p = .0049). It is concluded that the collagen-alginate test dressing is as or more effective and safe as the currently used treatment.

Adult↗

PubMed2005

Preparation of alkaline earth phosphates with sol containing sodium alginate and sodium diphosphate.

Magnesium hydrogen phosphate, calcium hydroxyapatite, and strontium hydroxyapatite were successfully prepared from sol consisting of sodium alginate and Na4P2O7 with Mg2+, Ca2+, and Sr2+ in the corresponding nitrates, respectively. It is revealed that the order of the addition of those substrates and the role of sodium alginate are important factors for the preparation of desired phosphate compounds. According to the previous paper on the preparation of calcium hydroxyapatite, sodium alginate was mixed with aqueous Na4P2O7, followed by the addition of the aqueous divalent cations, resulting in the poor formation of the target phosphates. However, as a revised sol-gel technique, sodium alginate was added to the mixture of Na4P2O7 and aqueous Mg2+ and Sr2+, resulting in a rather favorable formation of MgHPO4 and strontium hydroxyapatite, respectively, while the sol thus obtained was stable within a few days. However for aqueous Ca2+, calcium hydroxyapatite could not be obtained through the revised sol-gel technique. In the preparation of magnesium hydrogen phosphate, sodium alginate contributes mainly to the sol formation of the precursor. The ion exchange between Na+ in sodium alginate and aqueous Ca2+ was important for the preparation of calcium hydroxyapatite. In contrast, the reaction of sodium alginate with the mixture of Na4P2O7 and aqueous Sr2+ afforded strontium hydroxyapatite at the specific ratio of those three substrates. The structure of calcium and strontium phosphates prepared from the revised sol-gel process evidently depended on the amount of sodium alginate introduced into the mixture of Na4P2O7 and the corresponding divalent cations.

Journal Article↗

PubMed2006

Critical exponents and self-similarity for sol-gel transition in aqueous alginate systems induced by in situ release of calcium cations.

The sol-gel transition in aqueous alginate systems induced by in situ released calcium cations was monitored with rheology methods. Four alginate samples with different molecular weights and M/G ratios were used over the concentration C(Alg) of 2 approximately 6 wt % with different mole ratios f of Ca2+ to the alginate repeat unit. The scaling for the zero shear viscosity eta(0) before the gel point and the equilibrium modulus Ge after the gel point was described as eta(0) approximately epsilon(-k) and Ge approximately epsilon(z), respectively, where the relative distance to the gel point f(gel) was epsilon = (/f-f(gel)/)/f(gel). The relaxation critical exponent n was determined with Winter's criterion, and the critical exponents k and z estimated respectively from independent measurements of eta(0) and Ge gave n from z/(k + z). Before the gel point, the storage and loss moduli G' and G'' obtained at various epsilon can be superposed fairly well to form the master curve. The critical exponents n, k, and z were also evaluated from the shift factors and the structure self-similarity was found in the critical gel. The critical exponents evaluated with different methods agreed well with each other, suggesting two categories of the gelation as growth and cross-link. For the alginate with lower molecular weight, the critical exponents were almost independent of alginate concentration and close to the percolation prediction. For the alginate with higher molecular weight, the critical exponents, however, changed with alginate sample and concentration. The relative alginate concentration C(Alg)/C(Alg)* was found to serve as a criterion to divide these two transitions.

Journal Article↗

PubMed2004

Structure and function of a hypothetical Pseudomonas aeruginosa protein PA1167 classified into family PL-7: a novel alginate lyase with a beta-sandwich fold.

Structural and functional analyses of alginate lyases are important in the clarification of the biofilm-dependent ecosystem in Pseudomonas aeruginosa and in the development of therapeutic agents for bacterial disease. Most alginate lyases are classified into polysaccharide lyase (PL) family-5 and -7 based on their primary structures. Family PL-7 enzymes are still poorly characterized especially in structural properties. Among family PL-7, a gene coding for a hypothetical protein (PA1167) homologous to Sphingomonas alginate lyase A1-II was found to be present in the P. aeruginosa genome. PA1167 overexpressed in Escherichia coli cleaved glycosidic bonds in alginate and released unsaturated saccharides, indicating that PA1167 is an alginate lyase catalyzing a beta-elimination reaction. The enzyme acted preferably on heteropolymeric regions endolytically and worked most efficiently at pH 8.5 and 40 degrees C. The specific activity of PA1167, however, was much weaker than that of the known alginate lyase AlgL, suggesting that AlgL plays a main role in alginate depolymerization in P. aeruginosa. In addition to this specific activity, differences were found between PA1167 and AlgL in enzyme properties such as molecular mass, optimum pH, salt effect, and substrate specificity. The first crystal structure of the family PL-7 alginate lyase was determined at 2.0 A resolution. PA1167 was found to form a glove-like beta-sandwich composed of 15 beta-strands and 3 alpha-helices. The structural difference between the beta-sandwich PA1167 of family PL-7 and alpha/alpha-barrel AlgL of family PL-5 may be responsible for the enzyme characteristics. Crystal structures of polysaccharide lyases determined so far indicate that they can be assigned to three folding groups having parallel beta-helix, alpha/alpha-barrel, and alpha/alpha-barrel + antiparallel beta-sheet structures as basic frames. PA1167 is the fourth novel folding structure found among polysaccharide lyases.

Amino Acid Sequence↗

PubMed1998

Alginate lyase from Pseudomonas aeruginosa CF1/M1 prefers the hexameric oligomannuronate as substrate.

In order to investigate the catalytic properties of alginate lyase from Pseudomonas aeruginosa CF1/M1, a clinical isolate, regarding the capability to perform beta-elimination on oligomannuronates of defined length (2-9), the alginate lyase was purified from periplasmic extracts. A purification method for unsaturated and saturated oligomannuronates applying anionic exchange chromatography on a FPLC apparatus was established. The alginate lyase showed the highest activity, when hexamers were provided as substrate. This indicated that the alginate lyase best accommodates a chain of six alginate residues in the active center. As a minimum chain length, the pentameric oligomannuronate was still accepted as substrate. Mannuronate oligomers shorter than the pentamer were not accepted as substrate for alginate lyase. Furthermore, oligomer pattern analysis of polymannuronate which was subjected to beta-elimination by alginate lyase revealed that the trimer is the most abundant oligomer. These data indicated that beta-elimination and cleavage occurred at mannuronic acid residue no. 3 of the accommodated hexameric alginate chain.

Chromatography, Ion Exchange↗

PubMed2026

Azotobacter vinelandii AmrZ is a global regulator linking alginate production and c-di-GMP homeostasis.

Azotobacter vinelandii, a member of the Pseudomonadaceae, produces the exopolysaccharide alginate during vegetative growth; however, the circuitry linking alginate biosynthesis to lifestyle transitions remains poorly defined. Here, we show that the Ribbon-Helix-Helix transcription factor AmrZ coordinates alginate production, intracellular c-di-GMP levels and motility. Deletion of amrZ abolished alginate synthesis, whereas chromosomal complementation restored it. A PalgD-gusA fusion and RT-qPCR demonstrated that algD, the first gene in the alginate biosynthetic cluster, depends on AmrZ for expression. Motif analysis identified multiple AmrZ sites upstream of algD, and electrophoretic mobility-shift assays (EMSAs) confirmed specific binding to these regions. AmrZ also positively autoregulates: PamrZ-gusA activity decreased in ΔamrZ, and purified AmrZ bound the amrZ promoter in EMSA. Moreover, PamrZ activity required the sigma factor AlgU, consistent with the presence of an AlgU promoter; this positive, AlgU-dependent feedback may stabilize AmrZ under alginate-inducing conditions. To probe AmrZ control of c-di-GMP, we implemented a riboswitch-based biosensor in A. vinelandii. The ΔamrZ strain showed a markedly reduced signal, similar to a diguanylate cyclase (DGC) mutant, whereas a phosphodiesterase mutant displayed elevated output, validating the assay. RNA-seq and RT-qPCR identified two DGC genes, AVAEIV_RS11610 and AVAEIV_RS18795, as AmrZ-activated targets; EMSA verified direct binding at the RS11610 regulatory region. By contrast, transcription of the principal vegetative DGC AvGReg was not AmrZ-regulated. Lower c-di-GMP in ΔamrZ correlated with larger swimming halos. Collectively, these genetic, biochemical and transcriptomic data support a model in which AmrZ directly activates algD and elevates c-di-GMP via selected DGCs, thereby promoting alginate synthesis while reducing motility. RNA-seq data also indicate that AmrZ influences broader cellular programmes, including metabolism and iron homeostasis, positioning AmrZ as a central regulator that links c-di-GMP homeostasis to coordinated exopolysaccharide production in A. vinelandii. This work contributes to our understanding of the regulatory networks controlled by AmrZ outside the Pseudomonas genus and reveals important differences in its targets and regulatory mechanisms.

Azotobacter vinelandii↗

PubMed1996

Effects of calcium alginate on cellular wound healing processes modeled in vitro.

Although the clinical experience with calcium alginate has been generally good, well-controlled studies examining the effect of such dressings on the processes of wound healing have not been conducted. The healing of cutaneous ulcers requires the development of a vascularized granular tissue bed, filling of large tissue defects by dermal regeneration, and the restoration of a continuous epidermal keratinocyte layer. These processes were modeled in vitro in the present study, utilizing human dermal fibroblast, microvascular endothelial cell (HMEC), and keratinocyte cultures to examine the effect of calcium alginate on the proliferation and motility of these cultures, and the formation of capillarylike structures by HMEC. This study demonstrates that the calcium alginate tested increased the proliferation of fibroblasts but decreased the proliferation of HMEC and keratinocytes. In contrast, the calcium alginate decreased fibroblast motility but had no effect on keratinocyte motility. There was no significant effect of calcium alginate on the formation of capillarylike structures by HMEC. The effects of calcium alginate on cell proliferation and migration may have been mediated by released calcium ions. These results suggest that the calcium alginate tested may improve some cellular aspects of normal wound healing, but not others.

Alginates↗

PubMed1991

A versatile alginate droplet generator applicable for microencapsulation of pancreatic islets.

Alginate beads for immunoisolation of pancreatic islets by microencapsulation should be small, smooth, and spherical in order to ensure that around the islets a strong alginate-polylysine-alginate capsule will be formed with optimal biocompatibility and diffusion of nutrients and hormones. However, the preparation of small capsules around islets is difficult. Our newly designed air jet droplet generator allows for variations in the length and diameter of the alginate nozzle and the air jacket and is in this way adaptable to a required bead size. Alginate droplets are converted into rigid beads in a 100 mM CaCl 2 solution. Their size depends upon the diameter of the jacket, the air flow rate, and the outer diameter of the nozzle, whereas the production rate depends upon the pressure on the alginate, and on the diameter and the length of the nozzle. When the air flow or the alginate flow surpasses a certain rate, the droplets are fragmented. This study describes the mutual relationship of these variables and defines their optimal range for reproducible production of smooth and spherical beads for microencapsulation of islets at an acceptable production rate.

Alginates↗

PubMed2002

Alginate type and RGD density control myoblast phenotype.

Alginates are being increasingly used for cell encapsulation and tissue engineering applications; however, these materials cannot specifically interact with mammalian cells. We have covalently modified alginates of varying monomeric ratio with RGD-containing cell adhesion ligands using carbodiimide chemistry to initiate cell adhesion to these polymers. We hypothesized that we could control the function of cells adherent to RGD-modified alginate hydrogels by varying alginate polymer type and cell adhesion ligand density, and we have addressed this possibility by studying the proliferation and differentiation of C2C12 skeletal myoblasts adherent to these materials. RGD density on alginates of varying monomeric ratio could be controlled over several orders of magnitude, creating a range of surface densities from 1-100 fmol/cm(2). Myoblast adhesion to these materials was specific to the RGD ligand, because adhesion could be competed away with soluble RGD in a dose-dependent manner. Myoblast proliferation and differentiation could be regulated by varying the alginate monomeric ratio and the density of RGD ligands at the substrate surface, and specific combinations of alginate type and RGD density were required to obtain efficient myoblast differentiation on these materials.

Alginates↗

PubMed2003

Biocompatibility and surface structure of chemically modified immunoisolating alginate-PLL capsules.

Grafting of encapsulated living cells has the potential to cure a wide variety of diseases. Large-scale application of the technique, however, is hampered by insufficient biocompatibility of the capsules. A major factor in the biocompatibility of capsules is inadequate covering of the inflammatory poly-L-lysine (PLL) on the capsules' surface. In the present study, we investigate whether tissue responses against alginate-PLL capsules can be reduced by crosslinking the surface of the capsules with heparin or polyacrylic acid. Our transplant study in rats shows a tissue response composed of fibroblasts and macrophages on alginate-PLL-alginate and alginate-PLL-heparin capsules that was completely absent on alginate-PLL-polyacrylic acid capsules. Atomic force microscopy analyses of the capsules demonstrates that the improved biocompatibility of alginate-PLL-capsules by polyacrylic acid coating should not only be explained by a more adequate binding of PLL but also by the induction of a smoother surface. This study shows for the first time that biologic responses against capsules can be successfully deleted by chemically crosslinking biocompatible molecules on the surface of alginate-PLL capsules.

Acrylic Resins↗

PubMed2003

Chondrogenesis of human mesenchymal stem cells encapsulated in alginate beads.

Mesenchymal stem cells (MSCs) have the capacity for self-renewal and can form bone, fat, and cartilage. Alginate forms a viscous solution when dissolved in 0.9% saline and gels on contact with divalent cations. The viability and phenotype maintenance of chondrocytes in alginate beads have been well documented. However, little is known about the effect of microencapsulation in alginate on chondrogenesis of MSCs. In this study, human MSCs encapsulated in alginate beads were cultured in serum-free medium with the addition of transforming growth factor (TGF)beta1 (10 ng/mL), dexamethasone (10(-7) M), and ascorbate 2-phosphate (50 microg/mL). The MSCs in alginate assumed a rounded morphology with lacunae around them after 1 week in culture. Cell aggregates were observed at 2 weeks or longer in culture. Histological findings agreed with the clinical determination of hyaline cartilage, characterized by isolated cells with ground substance positive in Safranin-O staining and immunohistochemistry for collagen type II at the periphery of cells. Reverse transcriptase-polymerase chain reaction (RT-PCR) revealed the expression of COL2A1 and COL10A1, marker of chondrocytes and hypertrophy chondrocytes, respectively. These results indicate MSCs in alginate can form cartilage and the MSCs-alginate system represents a relevant model for the study of the molecular mechanisms involved in the chondrogenesis and endochondral ossification.

Alginates↗