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Sample records for alginate cell encapsulation

  1. Process engineering of high voltage alginate encapsulation of mesenchymal stem cells.

    Science.gov (United States)

    Gryshkov, Oleksandr; Pogozhykh, Denys; Zernetsch, Holger; Hofmann, Nicola; Mueller, Thomas; Glasmacher, Birgit

    2014-03-01

    Encapsulation of stem cells in alginate beads is promising as a sophisticated drug delivery system in treatment of a wide range of acute and chronic diseases. However, common use of air flow encapsulation of cells in alginate beads fails to produce beads with narrow size distribution, intact spherical structure and controllable sizes that can be scaled up. Here we show that high voltage encapsulation (≥ 15 kV) can be used to reproducibly generate spherical alginate beads (200-400 μm) with narrow size distribution (± 5-7%) in a controlled manner under optimized process parameters. Flow rate of alginate solution ranged from 0.5 to 10 ml/h allowed producing alginate beads with a size of 320 and 350 μm respectively, suggesting that this approach can be scaled up. Moreover, we found that applied voltages (15-25 kV) did not alter the viability and proliferation of encapsulated mesenchymal stem cells post-encapsulation and cryopreservation as compared to air flow. We are the first who employed a comparative analysis of electro-spraying and air flow encapsulation to study the effect of high voltage on alginate encapsulated cells. This report provides background in application of high voltage to encapsulate living cells for further medical purposes. Long-term comparison and work on alginate-cell interaction within these structures will be forthcoming.

  2. Process engineering of high voltage alginate encapsulation of mesenchymal stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Gryshkov, Oleksandr, E-mail: gryshkov@imp.uni-hannover.de [Institute for Multiphase Processes, Leibniz University Hannover, D-30167 Hannover (Germany); Pogozhykh, Denys, E-mail: pogozhykh@imp.uni-hannover.de [Institute for Multiphase Processes, Leibniz University Hannover, D-30167 Hannover (Germany); Zernetsch, Holger, E-mail: zernetsch@imp.uni-hannover.de [Institute for Multiphase Processes, Leibniz University Hannover, D-30167 Hannover (Germany); Hofmann, Nicola, E-mail: hofmann@imp.uni-hannover.de [Institute for Multiphase Processes, Leibniz University Hannover, D-30167 Hannover (Germany); Mueller, Thomas, E-mail: mueller.thomas@mh-hannover.de [Institute for Transfusion Medicine, Medical School Hannover, D-30625 Hannover (Germany); Glasmacher, Birgit, E-mail: glasmacher@imp.uni-hannover.de [Institute for Multiphase Processes, Leibniz University Hannover, D-30167 Hannover (Germany)

    2014-03-01

    Encapsulation of stem cells in alginate beads is promising as a sophisticated drug delivery system in treatment of a wide range of acute and chronic diseases. However, common use of air flow encapsulation of cells in alginate beads fails to produce beads with narrow size distribution, intact spherical structure and controllable sizes that can be scaled up. Here we show that high voltage encapsulation (≥ 15 kV) can be used to reproducibly generate spherical alginate beads (200–400 μm) with narrow size distribution (± 5–7%) in a controlled manner under optimized process parameters. Flow rate of alginate solution ranged from 0.5 to 10 ml/h allowed producing alginate beads with a size of 320 and 350 μm respectively, suggesting that this approach can be scaled up. Moreover, we found that applied voltages (15–25 kV) did not alter the viability and proliferation of encapsulated mesenchymal stem cells post-encapsulation and cryopreservation as compared to air flow. We are the first who employed a comparative analysis of electro-spraying and air flow encapsulation to study the effect of high voltage on alginate encapsulated cells. This report provides background in application of high voltage to encapsulate living cells for further medical purposes. Long-term comparison and work on alginate–cell interaction within these structures will be forthcoming. - Highlights: • High voltage alginate encapsulation of mesenchymal stem cells (MSCs) was designed. • Reproducible and spherical alginate beads were generated via high voltage. • Air flow encapsulation was utilized as a comparative approach to high voltage. • High voltage did not alter the viability and proliferation of encapsulated MSCs. • High voltage encapsulation can be scaled up and applied in cell-based therapy.

  3. Alginate encapsulation parameters influence the differentiation of microencapsulated embryonic stem cell aggregates.

    Science.gov (United States)

    Wilson, Jenna L; Najia, Mohamad Ali; Saeed, Rabbia; McDevitt, Todd C

    2014-03-01

    Pluripotent embryonic stem cells (ESCs) have tremendous potential as tools for regenerative medicine and drug discovery, yet the lack of processes to manufacture viable and homogenous cell populations of sufficient numbers limits the clinical translation of current and future cell therapies. Microencapsulation of ESCs within microbeads can shield cells from hydrodynamic shear forces found in bioreactor environments while allowing for sufficient diffusion of nutrients and oxygen through the encapsulation material. Despite initial studies examining alginate microbeads as a platform for stem cell expansion and directed differentiation, the impact of alginate encapsulation parameters on stem cell phenotype has not been thoroughly investigated. Therefore, the objective of this study was to systematically examine the effects of varying alginate compositions on microencapsulated ESC expansion and phenotype. Pre-formed aggregates of murine ESCs were encapsulated in alginate microbeads composed of a high or low ratio of guluronic to mannuronic acid residues (High G and High M, respectively), with and without a poly-L-lysine (PLL) coating, thereby providing four distinct alginate bead compositions for analysis. Encapsulation in all alginate compositions was found to delay differentiation, with encapsulation within High G alginate yielding the least differentiated cell population. The addition of a PLL coating to the High G alginate prevented cell escape from beads for up to 14 days. Furthermore, encapsulation within High M alginate promoted differentiation toward a primitive endoderm phenotype. Taken together, the findings of this study suggest that distinct ESC expansion capacities and differentiation trajectories emerge depending on the alginate composition employed, indicating that encapsulation material physical properties can be used to control stem cell fate.

  4. Alginate Encapsulation Parameters Influence the Differentiation of Microencapsulated Embryonic Stem Cell Aggregates

    Science.gov (United States)

    Wilson, Jenna L.; Najia, Mohamad Ali; Saeed, Rabbia; McDevitt, Todd C.

    2014-01-01

    Pluripotent embryonic stem cells (ESCs) have tremendous potential as tools for regenerative medicine and drug discovery, yet the lack of processes to manufacture viable and homogenous cell populations of sufficient numbers limits the clinical translation of current and future cell therapies. Microencapsulation of ESCs within microbeads can shield cells from hydrodynamic shear forces found in bioreactor environments while allowing for sufficient diffusion of nutrients and oxygen through the encapsulation material. Despite initial studies examining alginate microbeads as a platform for stem cell expansion and directed differentiation, the impact of alginate encapsulation parameters on stem cell phenotype has not been thoroughly investigated. Therefore, the objective of this study was to systematically examine the effects of varying alginate compositions on microencapsulated ESC expansion and phenotype. Pre-formed aggregates of murine ESCs were encapsulated in alginate microbeads composed of a high or low ratio of guluronic to mannuronic acid residues (High G and High M, respectively), with and without a poly-l-lysine (PLL) coating, thereby providing four distinct alginate bead compositions for analysis. Encapsulation in all alginate compositions was found to delay differentiation, with encapsulation within High G alginate yielding the least differentiated cell population. The addition of a PLL coating to the High G alginate prevented cell escape from beads for up to 14 days. Furthermore, encapsulation within High M alginate promoted differentiation toward a primitive endoderm phenotype. Taken together, the findings of this study suggest that distinct ESC expansion capacities and differentiation trajectories emerge depending on the alginate composition employed, indicating that encapsulation material physical properties can be used to control stem cell fate. PMID:24166004

  5. Bioluminescence tracking of alginate micro-encapsulated cell transplants.

    Science.gov (United States)

    Tiernan, Aubrey R; Sambanis, Athanassios

    2017-02-01

    Cell-based therapies to treat loss-of-function hormonal disorders such as diabetes and Parkinson's disease are routinely coupled with encapsulation strategies, but an understanding of when and why grafts fail in vivo is lacking. Consequently, investigators cannot clearly define the key factors that influence graft success. Although bioluminescence is a popular method to track the survival of free cells transplanted in preclinical models, little is known of the ability to use bioluminescence for real-time tracking of microencapsulated cells. Furthermore, the impact that dynamic imaging distances may have, due to freely-floating microcapsules in vivo, on cell survival monitoring is unknown. This work addresses these questions by applying bioluminescence to a pancreatic substitute based on microencapsulated cells. Recombinant insulin-secreting cells were transduced with a luciferase lentivirus and microencapsulated in Ba(2+) crosslinked alginate for in vitro and in vivo studies. In vitro quantitative bioluminescence monitoring was possible and viable microencapsulated cells were followed in real time under both normoxic and anoxic conditions. Although in vivo dispersion of freely-floating microcapsules in the peritoneal cavity limited the analysis to a qualitative bioluminescence evaluation, signals consistently four orders of magnitude above background were clear indicators of temporal cell survival. Strong agreement between in vivo and in vitro cell proliferation over time was discovered by making direct bioluminescence comparisons between explanted microcapsules and parallel in vitro cultures. Broader application of this bioluminescence approach to retrievable transplants, in supplement to currently used end-point physiological tests, could improve understanding and accelerate development of cell-based therapies for critical clinical applications. Copyright © 2014 John Wiley & Sons, Ltd.

  6. Cell-matrix Interactions of Factor IX (FIX)-engineered human mesenchymal stromal cells encapsulated in RGD-alginate vs. fibrinogen-alginate microcapsules.

    Science.gov (United States)

    Sayyar, Bahareh; Dodd, Megan; Marquez-Curtis, Leah; Janowska-Wieczorek, Anna; Hortelano, Gonzalo

    2014-04-01

    The success of cell microencapsulation technology in tissue engineering and protein delivery applications depends on the viability and functionality of the encapsulated cells, which in turn are dependent upon cell/matrix interactions. In this work, we compared the viability of cord blood-derived mesenchymal stromal cells (CB MSCs), engineered to secrete factor IX (FIX) for hemophilia treatment, and encapsulated in arginine-glycine-aspartate (RGD)-alginate versus fibrinogen-alginate microcapsules. We evaluated the effect of the biomimetic matrix on cell attachment, proliferation, and secretion of FIX. Compared with nonsupplemented alginate matrix, RGD-alginate significantly enhanced the viability of the encapsulated MSCs. Further, cells in RGD-alginate displayed distinct attachment morphology, thus suggesting that RGD-alginate can potentially be used for the encapsulation of MSCs in tissue engineering applications that require enhanced cell attachment and viability. However, our data also showed that RGD-alginate microcapsules, in contrast to fibrinogen-alginate microcapsules, did not significantly improve cell proliferation of or FIX secretion by encapsulated MSCs. Our findings suggest that evidence of cell attachment alone may not accurately predict the functionality of cells in biomimetic microcapsules.

  7. RGDS-fuctionalized alginates improve the survival rate of encapsulated embryonic stem cells during cryopreservation.

    Science.gov (United States)

    Sambu, S; Xu, X; Schiffer, H A; Cui, Z F; Ye, H

    2011-01-01

    Cryopreservation of stem cells, especially embryonic stem cells, is problematic because of low post-thaw cell survival rates and spontaneous differentiation following recovery. In this investigation, mouse embryonic stem cells (mESCs) were encapsulated in arginine-glycine-aspartic acid-serine (RGDS)-coupled calcium alginates (1.2 percent, w/v), allowed to attach to the substratum and then cryopreserved in 10 percent (v/v) dimethyl sulfoxide (DMSO) solution at a slow cooling rate of 1 C per min. RGDS coupling to alginate was confirmed by Transmission Fourier Transform Infra-Red spectroscopy (T-FTIR) and quantified by using ninhydrin-Ultraviolet/Visible light (ninhydrin-UV/VIS) assay. Flow cytometry data showed that mESCs cryopreserved in RGDS-alginate beads had a higher expression of stem cell markers compared with cells cryopreserved in suspension or cells cryopreserved in unmodified alginates. Cell viability after thawing was assessed using trypan blue exclusion assay and monitored using Alamar blue assay for 6 hours. It was shown that post-thaw cell survival rate was significantly higher for cells encapsulated in RGDS-modified alginate (93 ± 2 percent, mean and standard error) than those in suspension (52 ± 2 percent) or in unmodified alginates (62 ± 3 percent). These results showed that cells encapsulated and attached to a substratum have better survival rate and stem cell marker expression 24 hours after cryopreservation than those in suspension. Encapsulation in RGDS-alginate was optimized for peptide concentration, cryoprotective agent loading time and cooling rate. The best result was obtained when using 12.5 mg peptide per g alginate, 30 minutes loading time and 1 C per min cooling rate.

  8. Impact of residual contamination on the biofunctional properties of purified alginates used for cell encapsulation.

    Science.gov (United States)

    Tam, Susan K; Dusseault, Julie; Polizu, Stefania; Ménard, Martin; Hallé, Jean-Pierre; Yahia, L'Hocine

    2006-03-01

    Alginate is frequently used for cell encapsulation, but its biocompatibility is neither optimal nor reproducible. Purifying the alginate is critical for achieving a suitable biocompatibility. However, published purification methods vary in efficiency and may induce changes in polymer biofunctionality. Applying X-ray photoelectron spectroscopy, we showed that commercial alginates, purified by in-house and industrial methods, contained elemental impurities that contributed 0.41-1.73% of their atomic composition. Residual contaminants were identified to be proteins (nitrogen/COOH), endotoxins (phosphorus), and fucoidans (sulphur). Studies using attenuated total reflectance Fourier transform infrared spectroscopy suggested that trace contamination did not alter the alginate molecular structure. Alginate hydrophilicity increased by 19-40% after purification, in correlation with a reduction in protein and polyphenol content. Solution viscosity of the alginate increased by 28-108% after purification, in correlation with a reduction in protein content. These results demonstrate that commercial alginates contain potentially immunogenic contaminants that are not completely eliminated by current purification methods. Moreover, these contaminants alter the functional properties of the alginate in a manner that may compromise biocompatibility: Hydrophilicity may affect protein adsorption and solution viscosity influences the morphology of alginate-based microcapsules. These findings highlight the need to improve and better control alginate purity to ensure a reproducible biofunctionality and optimal biocompatibility of alginate and microcapsules.

  9. Encapsulated feeder cells within alginate beads for ex vivo expansion of cord blood-derived CD34(+) cells

    National Research Council Canada - National Science Library

    Pan, Xiuwei; Sun, Qiong; Cai, Haibo; Gao, Yun; Tan, Wensong; Zhang, Weian

    2016-01-01

    A co-culture system based on encapsulated feeder cells within alginate beads was developed through optimizing the detailed aspects of the cell culture system to expand CD34-positive (CD34(+)) cells ex vivo...

  10. Release of tissue inhibitor of metalloproteinase-2 from alginate microcapsule encapsulating genetically engineered cells

    Directory of Open Access Journals (Sweden)

    Kim YS

    2013-11-01

    Full Text Available Yeon Seong Kim,1,* Young-Il Jeong,2,* Shu-Guang Jin,2 Jian Pei,2 Min Wen,2 In-Young Kim,1 Kyung-Sub Moon,1 Tae-Young Jung,1 Hyang-Hwa Ryu2, Shin Jung1–3 1Department of Neurosurgery, 2Brain Tumor Research Laboratory, 3Chonnam National University Research Institute of Medical Sciences, Chonnam National University Hwasun Hospital and Medical School, Jeollanam-do, Korea *These authors contributed equally to this work Background: In this study, 293T cells were genetically engineered to secrete tissue inhibitor of metalloproteinase-2 (TIMP2 and encapsulated into alginate microcapsules to continuously release TIMP2 protein. Methods: The anti-invasive potential of the microcapsules was studied in vitro using brain tumor cells. The TIMP2 gene was transfected to 293T cells, and genetically engineered 293TIMP2 cells were encapsulated into alginate microcapsules. Release of TIMP2 protein was detected with Western blot analysis and the anti-invasive potential against U87MG cells was tested using gelatin zymography and a Matrigel assay. Results: Cell viability within the alginate microcapsules was maintained at a cell density of 5 × 106. Because polycationic polymers are helpful for maintaining the mechanical strength of microcapsules with good cell viability, the alginate microcapsules were reinforced with chitosan (0.1% w/v. Expression of TIMP2 protein in cell lysates and secretion of TIMP2 into the conditioned medium was confirmed by Western blot analysis. Alginate microcapsules encapsulating 293TIMP2 cells released TIMP2 protein into the medium efficiently, where the TIMP2 protein participated in degradation of the matrix metalloproteinase-2 enzyme and inhibited invasion of U87MG cells. Conclusion: Alginate microcapsules encapsulating 293TIMP2 cells are promising candidates for anti-invasive treatment of glioma. Keywords: 293T cells, tissue inhibitor of metalloproteinase-2, alginate microcapsule, therapeutic protein

  11. Production and in vitro evaluation of macroporous, cell-encapsulating alginate fibres for nerve repair.

    Science.gov (United States)

    Lin, Sharon Chien-Yu; Wang, Yiwei; Wertheim, David F; Coombes, Allan G A

    2017-04-01

    The prospects for successful peripheral nerve repair using fibre guides are considered to be enhanced by the use of a scaffold material, which promotes attachment and proliferation of glial cells and axonal regeneration. Macroporous alginate fibres were produced by extraction of gelatin particle porogens from wet spun fibres produced using a suspension of gelatin particles in 1.5% w/v alginate solution. Gelatin loading of the starting suspension of 40.0, 57.0, and 62.5% w/w resulted in gelatin loading of the dried alginate fibres of 16, 21, and 24% w/w respectively. Between 45 and 60% of the gelatin content of hydrated fibres was released in 1h in distilled water at 37°C, leading to rapid formation of a macroporous structure. Confocal laser scanning microscopy (CLSM) and image processing provided qualitative and quantitative analysis of mean equivalent macropore diameter (48-69μm), pore size distribution, estimates of maximum porosity (14.6%) and pore connectivity. CLSM also revealed that gelatin residues lined the macropore cavities and infiltrated into the body of the alginate scaffolds, thus, providing cell adhesion molecules, which are potentially advantageous for promoting growth of glial cells and axonal extension. Macroporous alginate fibres encapsulating nerve cells [primary rat dorsal root ganglia (DRGs)] were produced by wet spinning alginate solution containing dispersed gelatin particles and DRGs. Marked outgrowth was evident over a distance of 150μm at day 11 in cell culture, indicating that pores and channels created within the alginate hydrogel were providing a favourable environment for neurite development. These findings indicate that macroporous alginate fibres encapsulating nerve cells may provide the basis of a useful strategy for nerve repair. Copyright © 2016 Elsevier B.V. All rights reserved.

  12. Synthesis of Thermal Polymerizable Alginate-GMA Hydrogel for Cell Encapsulation

    Directory of Open Access Journals (Sweden)

    Xiaokun Wang

    2015-01-01

    Full Text Available Alginate is a negative ionic polysaccharide that is found abundantly in nature. Calcium is usually used as a cross-linker for alginate. However, calcium cross-linked alginate is used only for in vitro culture. In the present work, alginate was modified with glycidyl methacrylate (GMA to produce a thermal polymerizable alginate-GMA (AA-GMA macromonomer. The molecular structure and methacrylation (%DM of the macromonomer were determined by 1H NMR. After mixing with the correct amount of initiator, the AA-GMA aqueous solution can be polymerized at physiological temperature. The AA-GMA hydrogels exhibited a three-dimensional porous structure with an average pore size ranging from 50 to 200 μm, directly depending on the macromonomer concentration. Biocompatibility of the AA-GMA hydrogel was determined by in vivo muscle injection and cell encapsulation. Muscle injection in vivo showed that the AA-GMA solution mixed with initiator could form a hydrogel in situ and had a mild inflammatory effect. Human umbilical vein endothelial cells (HUVECs were encapsulated in the AA-GMA hydrogels in situ at 37°C. Cell viability and proliferation were unaffected by macromonomer concentrations, which suggests that AA-GMA has a potential application in the field of tissue engineering, especially for myocardial repair.

  13. Encapsulation of Huh-7 cells within alginate-poly(ethylene glycol) hybrid microspheres.

    Science.gov (United States)

    Mahou, Redouan; Tran, Nhu Mai; Dufresne, Murielle; Legallais, Cécile; Wandrey, Christine

    2012-01-01

    Novel calcium alginate poly(ethylene glycol) hybrid microspheres (Ca-alg-PEG) were developed and evaluated as potentially suitable materials for cell microencapsulation. Grafting 5-13% of the backbone units of sodium alginate (Na-alg) with α-amine-ω-thiol PEG maintained the gelling capacity in presence of calcium ions, while thiol end groups allowed for preparing chemically crosslinked hydrogel via spontaneous disulfide bond formation. The combination of these two gelling mechanisms yielded Ca-alg-PEG. Human hepatocellular carcinoma cells (Huh-7) were encapsulated in Ca-alg-PEG and calcium alginate beads (Ca-alg), and cultured for 2 weeks under agitation conditions. Immediately after completion of the microencapsulation, the cell viability was 60% and similar in Ca-alg-PEG and Ca-alg. The proliferation of Huh-7 encapsulated in Ca-alg-PEG was slightly higher than in Ca-alg. Accelerated proliferation after 2 weeks was observed for the encapsulation in Ca-alg-PEG. The production of albumin confirmed the functionality of the encapsulated Huh-7 cells. The study confirms the suitability of Ca-alg-PEG and the one-step technology for cell microencapsulation.

  14. Enrichment of committed human nucleus pulposus cells expressing chondroitin sulfate proteoglycans under alginate encapsulation.

    Science.gov (United States)

    Sun, Y; Lv, M; Zhou, L; Tam, V; Lv, F; Chan, D; Wang, H; Zheng, Z; Cheung, K M C; Leung, V Y L

    2015-07-01

    Intervertebral disc (IVD) degeneration is associated with a malfunction of the nucleus pulposus (NP). Alginate culturing provides a favorable microenvironment for the phenotypic maintenance of chondrocyte-like NP cells. However, NP cells are recently evidenced to present heterogeneous populations, including progenitors, fibroblastic cells and primitive NP cells. The aim of this study is to profile the phenotypic changes of distinct human NP cells populations and describe the dynamic expression of chondroitin sulfate glycosaminoglycans (CS-GAGs) in extended alginate encapsulation. Non-degenerated (ND-NPC) and degenerated (D-NPC) NP cells were expanded in monolayers, and subject to 28-day culture in alginate after serial passaging. CS-GAG compositional expression in monolayer-/alginate-cultured NP cells was evaluated by carbohydrate electrophoresis. Cellular phenotypic changes were assessed by immunologic detection and gene expression analysis. Relative to D-NPC, ND-NPC displayed remarkably higher expression levels of chondroitin-4-sulfate GAGs over the 28-day culture. Compared with monolayer culture, ND-NPC showed increased NP marker expression of KRT18, KRT19, and CDH2, as well as chondrocyte markers SOX9 and MIA in alginate culture. In contrast, expression of fibroblastic marker COL1A1, COL3A1, and FN1 were reduced. Interestingly, ND-NPC showed a loss of Tie2+ but gain in KRT19+/CD24+ population during alginate culture. In contrast, D-NPC showed more consistent expression levels of NP surface markers during culture. We demonstrate for the first time that extended alginate culture selectively enriches the committed NP cells and favors chondroitin-4-sulfate proteoglycan production. These findings suggest its validity as a model to investigate IVD cell function. Copyright © 2015 Osteoarthritis Research Society International. Published by Elsevier Ltd. All rights reserved.

  15. Alginate hydrogel protects encapsulated hepatic HuH-7 cells against hepatitis C virus and other viral infections.

    Directory of Open Access Journals (Sweden)

    Nhu-Mai Tran

    Full Text Available Cell microencapsulation in alginate hydrogel has shown interesting applications in regenerative medicine and the biomedical field through implantation of encapsulated tissue or for bioartificial organ development. Although alginate solution is known to have low antiviral activity, the same property regarding alginate gel has not yet been studied. The aim of this work is to investigate the potential protective effect of alginate encapsulation against hepatitis C virus (HCV infection for a hepatic cell line (HuH-7 normally permissive to the virus. Our results showed that alginate hydrogel protects HuH-7 cells against HCV when the supernatant was loaded with HCV. In addition, alginate hydrogel blocked HCV particle release out of the beads when the HuH-7 cells were previously infected and encapsulated. There was evidence of interaction between the molecules of alginate hydrogel and HCV, which was dose- and incubation time-dependent. The protective efficiency of alginate hydrogel towards HCV infection was confirmed against a variety of viruses, whether or not they were enveloped. This promising interaction between an alginate matrix and viruses, whose chemical mechanisms are discussed, is of great interest for further medical therapeutic applications based on tissue engineering.

  16. An animal model study for bone repair with encapsulated differentiated osteoblasts from adipose-derived stem cells in alginate

    OpenAIRE

    Hashemibeni, Batool; Esfandiari, Ebrahim; Sadeghi, Farzaneh; Heidary, Fariba; Roshankhah, Shiva; Mardani, Mohammad; Goharian, Vahid

    2014-01-01

    Objective(s): Adipose derived stem cells (ADSCs) can be engineered to express bone specific markers. The aim of this study is to evaluate repairing tibia in animal model with differentiated osteoblasts from autologous ADSCs in alginate scaffold. Materials and Methods: In this study, 6 canine's ADSCs were encapsulated in alginate and differentiated into osteoblasts. Alkaline phosphatase assay (ALP) and RT-PCR method were applied to confirm the osteogenic induction. Then, encapsulated different...

  17. An animal model study for bone repair with encapsulated differentiated osteoblasts from adipose-derived stem cells in alginate

    OpenAIRE

    Shiva Roshankhah; Mohammad Mardani; Vahid Goharian

    2014-01-01

    Objective(s): Adipose derived stem cells (ADSCs) can be engineered to express bone specific markers. The aim of this study is to evaluate repairing tibia in animal model with differentiated osteoblasts from autologous ADSCs in alginate scaffold. Materials and Methods: In this study, 6 canine’s ADSCs were encapsulated in alginate and differentiated into osteoblasts. Alkaline phosphatase assay (ALP) and RT-PCR method were applied to confirm the osteogenic induction. Then, encapsulated differ...

  18. Fibronectin-Alginate microcapsules improve cell viability and protein secretion of encapsulated Factor IX-engineered human mesenchymal stromal cells.

    Science.gov (United States)

    Sayyar, Bahareh; Dodd, Megan; Marquez-Curtis, Leah; Janowska-Wieczorek, Anna; Hortelano, Gonzalo

    2015-01-01

    Continuous delivery of proteins by engineered cells encapsu-lated in biocompatible polymeric microcapsules is of considerable therapeutic potential. However, this technology has not lived up to expectations due to inadequate cell--matrix interactions and subsequent cell death. In this study we hypoth-esize that the presence of fibronectin in an alginate matrix may enhance the viability and functionality of encapsulated human cord blood-derived mesenchymal stromal cells (MSCs) expressing the human Factor IX (FIX) gene. MSCs were encapsulated in alginate-PLL microcapsules containing 10, 100, or 500 μg/ml fibronectin to ameliorate cell survival. MSCs in microcapsules with 100 and 500 μg/ml fibronectin demonstrated improved cell viability and proliferation and higher FIX secretion compared to MSCs in non-supplemented microcapsules. In contrast, 10 μg/ml fibronectin did not significantly affect the viability and protein secretion from the encapsulated cells. Differentiation studies demonstrated osteogenic (but not chondrogenic or adipogenic) differentiation capability and efficient FIX secretion of the enclosed MSCs in the fibronectin-alginate suspension culture. Thus, the use of recombinant MSCs encapsulated in fibronectin-alginate microcapsules in basal or osteogenic cultures may be of practical use in the treatment of hemophilia B.

  19. Gel Microstructure Regulates Proliferation and Differentiation of MC3T3-E1 Cells Encapsulated in Alginate Beads

    Science.gov (United States)

    Lee, Baek-Hee; Li, Bing; Guelcher, Scott A.

    2012-01-01

    For cell transplantation into damaged tissues, viable cells must be delivered to the defect site in a suitable carrier. However, the hypoxic and nutrient-limited environment in the carrier can induce massive cell death. The aims of this study were to increase the viability and regulate the behavior of osteoprogenitor cells encapsulated in alginate hydrogels through control of the gel microstructure. Cell survivability in alginate beads was improved through the use of α-MEM as the solvent for alginic acid sodium salt and CaCl2 solutions, which supplied additional nutrients for the cells compared to water or buffer. The mesh size and shear modulus of the hydrogel were hypothesized to regulate proliferation and differentiation of osteoprogenitor cells. MC3T3-E1 cells demonstrated enhanced osteoblast differentiation when encapsulated in high-density alginate with smaller mesh size and more rigid mechanical properties, as confirmed by increased alkaline phosphatase activity and osteocalcin secretion. However, MC3T3-E1 cells encapsulated in low-density alginate beads with a larger mesh size and more compliant mechanical properties exhibited increased proliferation. These results demonstrate that the microstructure of alginate hydrogels can regulate the behavior of osteoprogenitor cells, thus suggesting that the tuning the properties of the gel may be a useful approach for enhancing new bone formation. PMID:22306825

  20. Light-Addressable Electrodeposition of Magnetically-Guided Cells Encapsulated in Alginate Hydrogels for Three-Dimensional Cell Patterning

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    Shih-Hao Huang

    2014-11-01

    Full Text Available This paper describes a light-addressable electrolytic system used to perform an electrodeposition of magnetically-guided cells encapsulated in alginate hydrogels using a digital micromirror device (DMD for three-dimensional cell patterning. In this system, the magnetically-labeled cells were first manipulated into a specific arrangement by changing the orientation of the magnetic field, and then a patterned light illumination was projected onto a photoconductive substrate serving as a photo-anode to cause gelation of calcium alginate through sol-gel transition. By controlling the illumination pattern on the DMD, we first successfully produced cell-encapsulated multilayer alginate hydrogels with different shapes and sizes in each layer via performing multiplexed micropatterning. By combining the magnetically-labeled cells, light-addressable electrodeposition, and orientation of the magnetic fields, we have successfully demonstrated to fabricate two layers of the cell-encapsulated alginate hydrogels, where cells in each layer can be manipulated into cross-directional arrangements that mimic natural tissue. Our proposed method provides a programmable method for the spatiotemporally controllable assembly of cell populations into three-dimensional cell patterning and could have a wide range of biological applications in tissue engineering, toxicology, and drug discovery.

  1. Muscle Tissue Engineering Using Gingival Mesenchymal Stem Cells Encapsulated in Alginate Hydrogels Containing Multiple Growth Factors.

    Science.gov (United States)

    Ansari, Sahar; Chen, Chider; Xu, Xingtian; Annabi, Nasim; Zadeh, Homayoun H; Wu, Benjamin M; Khademhosseini, Ali; Shi, Songtao; Moshaverinia, Alireza

    2016-06-01

    Repair and regeneration of muscle tissue following traumatic injuries or muscle diseases often presents a challenging clinical situation. If a significant amount of tissue is lost the native regenerative potential of skeletal muscle will not be able to grow to fill the defect site completely. Dental-derived mesenchymal stem cells (MSCs) in combination with appropriate scaffold material, present an advantageous alternative therapeutic option for muscle tissue engineering in comparison to current treatment modalities available. To date, there has been no report on application of gingival mesenchymal stem cells (GMSCs) in three-dimensional scaffolds for muscle tissue engineering. The objectives of the current study were to develop an injectable 3D RGD-coupled alginate scaffold with multiple growth factor delivery capacity for encapsulating GMSCs, and to evaluate the capacity of encapsulated GMSCs to differentiate into myogenic tissue in vitro and in vivo where encapsulated GMSCs were transplanted subcutaneously into immunocompromised mice. The results demonstrate that after 4 weeks of differentiation in vitro, GMSCs as well as the positive control human bone marrow mesenchymal stem cells (hBMMSCs) exhibited muscle cell-like morphology with high levels of mRNA expression for gene markers related to muscle regeneration (MyoD, Myf5, and MyoG) via qPCR measurement. Our quantitative PCR analyzes revealed that the stiffness of the RGD-coupled alginate regulates the myogenic differentiation of encapsulated GMSCs. Histological and immunohistochemical/fluorescence staining for protein markers specific for myogenic tissue confirmed muscle regeneration in subcutaneous transplantation in our in vivo animal model. GMSCs showed significantly greater capacity for myogenic regeneration in comparison to hBMMSCs (p engineering.

  2. Encapsulation of factor IX-engineered mesenchymal stem cells in fibrinogen-alginate microcapsules enhances their viability and transgene secretion.

    Science.gov (United States)

    Sayyar, Bahareh; Dodd, Megan; Wen, Jianping; Ma, Shirley; Marquez-Curtis, Leah; Janowska-Wieczorek, Anna; Hortelano, Gonzalo

    2012-01-01

    Cell microencapsulation holds significant promise as a strategy for cellular therapies; however, inadequate survival and functionality of the enclosed cells limit its application in hemophilia treatment. Here, we evaluated the use of alginate-based microcapsules to enhance the viability and transgene secretion of human cord blood-derived mesenchymal stem cells in three-dimensional cultures. Given the positive effects of extracellular matrix molecules on mesenchymal stem cell growth, we tested whether fibrinogen-supplemented alginate microcapsules can improve the efficiency of encapsulated factor IX-engineered mesenchymal stem cells as a treatment of hemophilia B. We found that fibrinogen-supplemented alginate microcapsules (a) significantly enhanced the viability and proliferation of factor IX-engineered mesenchymal stem cells and (b) increased factor IX secretion by mesenchymal stem cells compared to mesenchymal stem cells in nonsupplemented microcapsules. Moreover, we observed the osteogenic, but not chondrogenic or adipogenic, differentiation capability of factor IX-engineered cord blood mesenchymal stem cells and their efficient factor IX secretion while encapsulated in fibrinogen-supplemented alginate microcapsules. Thus, the use of engineered mesenchymal stem cells encapsulated in fibrinogen-modified microcapsules may have potential application in the treatment of hemophilia or other protein deficiency diseases.

  3. Encapsulation of factor IX–engineered mesenchymal stem cells in fibrinogen–alginate microcapsules enhances their viability and transgene secretion

    Directory of Open Access Journals (Sweden)

    Bahareh Sayyar

    2012-12-01

    Full Text Available Cell microencapsulation holds significant promise as a strategy for cellular therapies; however, inadequate survival and functionality of the enclosed cells limit its application in hemophilia treatment. Here, we evaluated the use of alginate-based microcapsules to enhance the viability and transgene secretion of human cord blood–derived mesenchymal stem cells in three-dimensional cultures. Given the positive effects of extracellular matrix molecules on mesenchymal stem cell growth, we tested whether fibrinogen-supplemented alginate microcapsules can improve the efficiency of encapsulated factor IX–engineered mesenchymal stem cells as a treatment of hemophilia B. We found that fibrinogen-supplemented alginate microcapsules (a significantly enhanced the viability and proliferation of factor IX–engineered mesenchymal stem cells and (b increased factor IX secretion by mesenchymal stem cells compared to mesenchymal stem cells in nonsupplemented microcapsules. Moreover, we observed the osteogenic, but not chondrogenic or adipogenic, differentiation capability of factor IX–engineered cord blood mesenchymal stem cells and their efficient factor IX secretion while encapsulated in fibrinogen-supplemented alginate microcapsules. Thus, the use of engineered mesenchymal stem cells encapsulated in fibrinogen-modified microcapsules may have potential application in the treatment of hemophilia or other protein deficiency diseases.

  4. Reversal of diabetes by βTC3 cells encapsulated in alginate beads generated by emulsion and internal gelation.

    Science.gov (United States)

    Hoesli, Corinne A; Kiang, Roger L J; Mocinecová, Dušana; Speck, Madeleine; Mošková, Daniela Jochec; Donald-Hague, Christine; Lacík, Igor; Kieffer, Timothy J; Piret, James M

    2012-05-01

    Encapsulation of insulin-producing cells in alginate beads could improve the treatment of type 1 diabetes by reducing or eliminating the need for immunosuppression. We have recently adapted an emulsion and internal gelation process to β-cell encapsulation. This process has the advantages of being well suited for m(3)/h production rates and allowing the use of increased alginate concentrations. Compared with 1.5% alginate beads generated by a standard extrusion process, 5% alginate emulsion-generated beads demonstrated greater in vitro stability and greater volumetric exclusion of antibody-sized pullulan. When βTC3 cells were transplanted into streptozotocin-induced allogeneic diabetic mice, a significant decrease in the blood glucose levels was seen within 2 days with the 5% emulsion-generated beads but not until >16 days with the 1.5% extrusion-generated beads. This was correlated with higher cell survival and lower graft-specific plasma immunoglobulin levels. These results suggest that higher-concentration alginate beads generated by emulsion and internal gelation have improved graft immunoprotection. The emulsion process is a promising and scalable technology for cellular therapies requiring immune isolation.

  5. Oral delivery of probiotic expressing M cell homing peptide conjugated BmpB vaccine encapsulated into alginate/chitosan/alginate microcapsules.

    Science.gov (United States)

    Jiang, Tao; Singh, Bijay; Maharjan, Sushila; Li, Hui-Shan; Kang, Sang-Kee; Bok, Jin-Duck; Cho, Chong-Su; Choi, Yun-Jaie

    2014-11-01

    Oral administration of live probiotics as antigen delivery vectors is a promising approach in vaccine development. However, the low survival of probiotics in the gastrointestinal tract limits this approach. Therefore, the aim of this study was the encapsulation of probiotic expressing vaccine into alginate/chitosan/alginate (ACA) microcapsules (MCs) for efficient oral vaccine delivery. Here, recombinant Lactobacillus plantarum 25 (LP25) expressing M cell homing peptide fused BmpB protein was used as a model probiotic. The viability of LP25 in ACA MCs was more than 65% in simulated gastric fluid (SGF, pH 2.0) and 75% in simulated small intestinal fluid (SIF, pH 7.2) up to 2h. Encapsulated LP25 was completely released from ACA MCs in SIF within 12h. When stored at room temperature (RT) or 4°C, the viability of LP25 in ACA MCs was higher than free LP25. Interestingly, the viability of LP25 in ACA MCs at 4°C for 5weeks was above 58%, whereas viability of free LP25 stored at RT up to 5weeks was zero. After 4weeks from the first immunization, LP25-M-BmpB-loaded ACA MCs induced a stronger BmpB-specific IgG and IgA production in mice. Collectively, these findings suggest that encapsulation of probiotic by ACA MCs is a promising delivery system for oral administration of probiotic expressing vaccine. Copyright © 2014 Elsevier B.V. All rights reserved.

  6. Quantitative Assessment of Islets of Langerhans Encapsulated in Alginate

    Science.gov (United States)

    Johnson, Amy S.; O'Sullivan, Esther; D'Aoust, Laura N.; Omer, Abdulkadir; Bonner-Weir, Susan; Fisher, Robert J.; Weir, Gordon C.

    2011-01-01

    Improved methods have recently been developed for assessing islet viability and quantity in human islet preparations for transplantation, and these measurements have proven useful for predicting transplantation outcome. The objectives of this study were to adapt these methods for use with microencapsulated islets, to verify that they provide meaningful quantitative measurements, and to test them with two model systems: (1) barium alginate and (2) barium alginate containing a 70% (w/v) perfluorocarbon (PFC) emulsion, which presents challenges to use of these assays and is of interest in its own right as a means for reducing oxygen supply limitations to encapsulated tissue. Mitochondrial function was assessed by oxygen consumption rate measurements, and the analysis of data was modified to account for the increased solubility of oxygen in the PFC-alginate capsules. Capsules were dissolved and tissue recovered for nuclei counting to measure the number of cells. Capsule volume was determined from alginate or PFC content and used to normalize measurements. After low oxygen culture for 2 days, islets in normal alginate lost substantial viable tissue and displayed necrotic cores, whereas most of the original oxygen consumption rate was recovered with PFC alginate, and little necrosis was observed. All nuclei were recovered with normal alginate, but some nuclei from nonrespiring cells were lost with PFC alginate. Biocompatibility tests revealed toxicity at the islet periphery associated with the lipid emulsion used to provide surfactants during the emulsification process. We conclude that these new assay methods can be applied to islets encapsulated in materials as complex as PFC-alginate. Measurements made with these materials revealed that enhancement of oxygen permeability of the encapsulating material with a concentrated PFC emulsion improves survival of encapsulated islets under hypoxic conditions, but reformulation of the PFC emulsion is needed to reduce toxicity

  7. Magnetization transfer contrast MRI for non-invasive assessment of innate and adaptive immune responses against alginate-encapsulated cells.

    Science.gov (United States)

    Chan, Kannie W Y; Liu, Guanshu; van Zijl, Peter C M; Bulte, Jeff W M; McMahon, Michael T

    2014-09-01

    By means of physical isolation of cells inside semi-permeable hydrogels, encapsulation has been widely used to immunoprotect transplanted cells. While spherical alginate microcapsules are now being used clinically, there still is little known about the patient's immune system response unless biopsies are obtained. We investigated the use of Magnetization Transfer (MT) imaging to non-invasively detect host immune responses against alginate capsules containing xenografted human hepatocytes in four groups of animals, including transplanted empty capsules (-Cells/-IS), capsules with live cells with (+LiveCells/+IS) and without immunosuppression (+LiveCells/-IS), and capsules with apoptotic cells in non-immunosuppressed animals (+DeadCells/-IS). The highest MT ratio (MTR) was found in +LiveCells/-IS, which increased from day 0 by 38% and 53% on days 7 and 14 after transplantation respectively, and corresponded to a distinctive increase in cell infiltration on histology. Furthermore, we show that macromolecular ratio maps based on MT data are more sensitive to cell infiltration and fibrosis than conventional MTR maps. Such maps showed a significant difference between +LiveCells/-IS (0.18 ± 0.02) and +DeadCells/-IS (0.13 ± 0.02) on day 7 (P < 0.01) existed, which was not observed on MTR imaging. We conclude that MT imaging, which is clinically available, can be applied for non-invasive monitoring of the occurrence of a host immune response against encapsulated cells. Copyright © 2014 Elsevier Ltd. All rights reserved.

  8. Encapsulated human hepatocellular carcinoma cells by alginate gel beads as an in vitro metastasis model

    Energy Technology Data Exchange (ETDEWEB)

    Xu, Xiao-xi; Liu, Chang [Laboratory of Biomedical Material Engineering, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, 457 Zhongshan Road, Dalian 116023 (China); University of Chinese Academy of Sciences, 19A Yuquanlu, Beijing 100049 (China); Liu, Yang [Laboratory of Biomedical Material Engineering, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, 457 Zhongshan Road, Dalian 116023 (China); Li, Nan [Laboratory of Biomedical Material Engineering, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, 457 Zhongshan Road, Dalian 116023 (China); University of Chinese Academy of Sciences, 19A Yuquanlu, Beijing 100049 (China); Guo, Xin [Laboratory of Biomedical Material Engineering, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, 457 Zhongshan Road, Dalian 116023 (China); Wang, Shu-jun [Laboratory of Biomedical Material Engineering, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, 457 Zhongshan Road, Dalian 116023 (China); School of Life Science and Biotechnology, Dalian University of Technology, 2 Linggong Road, Dalian 116024 (China); Sun, Guang-wei, E-mail: sungw@dicp.ac.cn [Laboratory of Biomedical Material Engineering, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, 457 Zhongshan Road, Dalian 116023 (China); Wang, Wei [Laboratory of Biomedical Material Engineering, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, 457 Zhongshan Road, Dalian 116023 (China); Ma, Xiao-jun, E-mail: maxj@dicp.ac.cn [Laboratory of Biomedical Material Engineering, Dalian Institute of Chemical Physics, Chinese Academy of Sciences, 457 Zhongshan Road, Dalian 116023 (China)

    2013-08-15

    Hepatocellular carcinoma (HCC) is the most common primary liver cancer and often forms metastases, which are the most important prognostic factors. For further elucidation of the mechanism underlying the progression and metastasis of HCC, a culture system mimicking the in vivo tumor microenvironment is needed. In this study, we investigated the metastatic ability of HCC cells cultured within alginate gel (ALG) beads. In the culture system, HCC cells formed spheroids by proliferation and maintained in nuclear abnormalities. The gene and protein expression of metastasis-related molecules was increased in ALG beads, compared with the traditional adhesion culture. Furthermore, several gene expression levels in ALG bead culture system were even closer to liver cancer tissues. More importantly, in vitro invasion assay showed that the invasion cells derived from ALG beads was 7.8-fold higher than adhesion cells. Our results indicated that the in vitro three-dimensional (3D) model based on ALG beads increased metastatic ability compared with adhesion culture, even partly mimicked the in vivo tumor tissues. Moreover, due to the controllable preparation conditions, steady characteristics and production at large-scale, the 3D ALG bead model would become an important tool used in the high-throughput screening of anti-metastasis drugs and the metastatic mechanism research. -- Highlights: •We established a 3D metastasis model mimicking the metastatic ability in vivo. •The invasion ability of cells derived from our model was increased significantly. •The model is easy to reproduce, convenient to handle, and amenable for large-scale.

  9. Sustained Delivery of Bioactive GDNF from Collagen and Alginate-Based Cell-Encapsulating Gel Promoted Photoreceptor Survival in an Inherited Retinal Degeneration Model.

    Directory of Open Access Journals (Sweden)

    Francisca S Y Wong

    Full Text Available Encapsulated-cell therapy (ECT is an attractive approach for continuously delivering freshly synthesized therapeutics to treat sight-threatening posterior eye diseases, circumventing repeated invasive intravitreal injections and improving local drug availability clinically. Composite collagen-alginate (CAC scaffold contains an interpenetrating network that integrates the physical and biological merits of its constituents, including biocompatibility, mild gelling properties and availability. However, CAC ECT properties and performance in the eye are not well-understood. Previously, we reported a cultured 3D CAC system that supported the growth of GDNF-secreting HEK293 cells with sustainable GDNF delivery. Here, the system was further developed into an intravitreally injectable gel with 1x104 or 2x105 cells encapsulated in 2mg/ml type I collagen and 1% alginate. Gels with lower alginate concentration yielded higher initial cell viability but faster spheroid formation while increasing initial cell density encouraged cell growth. Continuous GDNF delivery was detected in culture and in healthy rat eyes for at least 14 days. The gels were well-tolerated with no host tissue attachment and contained living cell colonies. Most importantly, gel-implanted in dystrophic Royal College of Surgeons rat eyes for 28 days retained photoreceptors while those containing higher initial cell number yielded better photoreceptor survival. CAC ECT gels offers flexible system design and is a potential treatment option for posterior eye diseases.

  10. 3D Cell Culture in Alginate Hydrogels

    Directory of Open Access Journals (Sweden)

    Therese Andersen

    2015-03-01

    Full Text Available This review compiles information regarding the use of alginate, and in particular alginate hydrogels, in culturing cells in 3D. Knowledge of alginate chemical structure and functionality are shown to be important parameters in design of alginate-based matrices for cell culture. Gel elasticity as well as hydrogel stability can be impacted by the type of alginate used, its concentration, the choice of gelation technique (ionic or covalent, and divalent cation chosen as the gel inducing ion. The use of peptide-coupled alginate can control cell–matrix interactions. Gelation of alginate with concomitant immobilization of cells can take various forms. Droplets or beads have been utilized since the 1980s for immobilizing cells. Newer matrices such as macroporous scaffolds are now entering the 3D cell culture product market. Finally, delayed gelling, injectable, alginate systems show utility in the translation of in vitro cell culture to in vivo tissue engineering applications. Alginate has a history and a future in 3D cell culture. Historically, cells were encapsulated in alginate droplets cross-linked with calcium for the development of artificial organs. Now, several commercial products based on alginate are being used as 3D cell culture systems that also demonstrate the possibility of replacing or regenerating tissue.

  11. Characterization of viability and proliferation of alginate-poly-L-lysine-alginate encapsulated myoblasts using flow cytometry.

    Science.gov (United States)

    Thakur, Ajit; Sengupta, Ruchira; Matsui, Hideto; Lillicrap, David; Jones, Kim; Hortelano, Gonzalo

    2010-08-01

    Genetically modified cells encapsulated in alginate-poly-L-lysine-alginate (APA) are being developed to deliver therapeutic products to treat a variety of diseases. The characterization of the encapsulated cells thus becomes paramount. This study reports a novel method to assess the viability, granularity and proliferation of encapsulated cells based on flow cytometry. The in vitro viability of encapsulated G8 murine myoblasts secreting canine FVIII (cFVIII) measured by flow cytometry was comparable to the traditional trypan blue exclusion method and both correlated with cFVIII secretion levels. In contrast, after implantation into mice, only viability measured by flow cytometry correlated with cFVIII secretion. Further, flow cytometry analysis of encapsulated cells maintained in vitro and in vivo revealed a greater fraction of granular cells compared to free cells, suggesting that encapsulation influences the morphology (cytoplasmic composition) of cells within APA microcapsules. Interestingly, the proliferation study showed that encapsulated cells proliferate faster, on average, and were more heterogeneous in vivo compared to in vitro culture conditions, suggesting that encapsulated cell proliferation is complex and environment-dependent. In conclusion, we show that flow cytometry analysis allows for a more consistent and comprehensive examination of encapsulated cells to aid in the development of cell therapy protocols.

  12. GOLD NANOPARTICLES ENCAPSULATED IN A POLYMERIC MATRIX OF SODIUM ALGINATE

    Directory of Open Access Journals (Sweden)

    Oana Lelia POP

    2016-11-01

    Full Text Available Plasmonic nanoparticles can be used as building blocks for the design of multifunctional systems based on polymeric capsules. The use of functionalised particles in therapeutics and imaging and understanding their effect on the cell functions are among the current challenges in nanobiotechnology and nanomedicine. The aim of the study was to manufacture and characterize polymeric microstructures by encapsulating plasmonic gold nanoparticles in biocompatible matrix of sodium alginate. The gold nanoparticles were obtained by reduction of tetracluoroauric acid with sodium citrate. To characterize the microcapsules, UV-Vis and FTIR spectroscopy, optical and confocal microscopy experiments were performed. In vitro cytotoxicity tests on HFL-1 cells were also performed. The capsules have spherical shape and 120 μm diameter. The presence of encapsulated gold nanoparticles is also shown by confocal microscopy. In vitro tests show that the microcapsules are not cytotoxic upon 24 h of cells exposure to microcapsules concentrations ranging from 2.5 to 25 capsules per cell. The obtained microcapsules of sodium alginate loaded with plasmonic gold nanoparticles could potentially be considered as release systems for biologically relevant molecules.

  13. Live encapsulated Lactobacillus acidophilus cells in yogurt for therapeutic oral delivery: preparation and in vitro analysis of alginate-chitosan microcapsules.

    Science.gov (United States)

    Urbanska, Aleksandra Malgorzata; Bhathena, Jasmine; Prakash, Satya

    2007-09-01

    Targeted delivery of live microencapsulated bacterial cells has strong potential for application in treating various diseases, including diarrhea, kidney failure, liver failure, and high cholesterol, among others. This study investigates the potential of microcapsules composed of two natural polymers, alginate and chitosan (AC), and the use of these artificial cells in yogurt for delivery of probiotic Lactobacillus acidophilus bacterial live cells. Results show that the integrity of AC microcapsules was preserved after 76 h of mechanical shaking in MRS broth and after 12 h and 24 h in simulated gastric and intestinal fluids. Using an in vitro computer-controlled simulated human gastrointestinal (GI) model, we found 8.37 log CFU/mL of viable bacterial cells were present after 120 min of gastric exposure and 7.96 log CFU/mL after 360 min of intestinal exposure. In addition, AC microcapsules composed of chitosan 10 and 100 at various concentrations were subjected to 4-week storage in 2% milk fat yogurt or 0.85% physiological solution. It was found that 9.37 log CFU/mL of cells encapsulated with chitosan 10 and 8.24 log CFU/mL of cells encapsulated with chitosan 100 were alive after 4 weeks. The AC capsule composed of 0.5% chitosan 10 provided the highest bacterial survival of 9.11 log CFU/mL after 4 weeks. Finally, an investigation of bacterial viability over 72 h in different pH buffers yielded highest survival of 6.34 log CFU/mL and 10.34 log CFU/mL at pH 8 for free and AC-encapsulated cells, respectively. We conclude from these findings that encapsulation allows delivery of a higher number of bacteria to desired targets in the GI tract and that microcapsules containing bacterial cells are good candidates for oral artificial cells for bacterial cell therapy.

  14. Alginate: A Versatile Biomaterial to Encapsulate Isolated Ovarian Follicles.

    Science.gov (United States)

    Vanacker, Julie; Amorim, Christiani A

    2017-02-28

    In vitro culture of ovarian follicles isolated or enclosed in ovarian tissue fragments and grafting of isolated ovarian follicles represent a potential alternative to restore fertility in cancer patients who cannot undergo cryopreservation of embryos or oocytes or transplantation of frozen-thawed ovarian tissue. In this regard, respecting the three-dimensional (3D) architecture of isolated follicles is crucial to maintaining their proper follicular physiology. To this end, alginate hydrogel has been widely investigated using follicles from numerous animal species, yielding promising results. The goal of this review is therefore to provide an overview of alginate applications utilizing the biomaterial as a scaffold for 3D encapsulation of isolated ovarian follicles. Different methods of isolated follicle encapsulation in alginate are discussed in this review, as its use of 3D alginate culture systems as a tool for in vitro follicle analysis. Possible improvements of this matrix, namely modification with arginine-glycine-aspartic acid peptide or combination with fibrin, are also summarized. Encouraging results have been obtained in different animal models, and particularly with isolated follicles encapsulated in alginate matrices and grafted to mice. This summary is designed to guide the reader towards development of next-generation alginate scaffolds, with enhanced properties for follicle encapsulation.

  15. In vivo selection of biocompatible alginates for islet encapsulation and subcutaneous transplantation.

    Science.gov (United States)

    Vériter, Sophie; Mergen, Julien; Goebbels, Rose-Marie; Aouassar, Najima; Grégoire, Charles; Jordan, Bénédicte; Levêque, Philippe; Gallez, Bernard; Gianello, Pierre; Dufrane, Denis

    2010-05-01

    Islet encapsulation requires several properties including (1) biocompatibility, (2) immunoprotection, and (3) oxygen diffusion for islet survival and diabetes correction. New chemical alginates were tested in vivo and compared with traditional high-mannuronate and -guluronate alginates. New alginates with coupled peptide sequence (sterile lyophilized high mannuronate [SLM]-RGD3% and sterile lyophilized high guluronate [SLG]-RGD3%), to improve encapsulated cell adherence in the matrix, and alginates with a very low viscosity (VLDM7% and VLDG7%), to reduce implant size by loading a higher number of islets per volume of polymer, were implanted subcutaneously in 70 Wistar rats for comparison with alginates of high viscosity and high content of mannuronic (SLM3%) or guluronic acids (SLG3%). Permeability of alginates to 36-, 75-, and 150-kDa lectins coupled to fluorescein isothiocynate was quantified before implantation and at 2, 4, and 12 weeks after implantation. Biocompatibility (fibrosis, graft stability, immunologic infiltration by CD3/CD68 cells, and neovascularization) was assessed at each explantation time. Permeability to small molecules was found for all alginates. Impermeability to 150-kDa molecules, such as IgG, was observed only for SLM3% before implantation and was maintained up to 12 weeks after implantation. SLM3% and SLG3% demonstrated better graft stability with lower CD3/CD68 recruitment and fibrosis than the other alginates. SLM3% induced a significantly higher angiogenesis and maintained oxygen pressure at approximately 40 mm Hg for up to 4 weeks after implantation as measured by in vivo electronic paramagnetic resonance oximetry. SLM-encapsulated pig islets implanted subcutaneously in rats demonstrated no inflammatory/immunologic reactions and islets functioned for up to 60 days without immunosuppression. A traditional alginate made of high mannuronic content (SLM3%) is an adapted material to immunoprotect islets in subcutaneous tissue. No

  16. Efficiencies in alginate encapsulation of vegetative explants

    Science.gov (United States)

    The goal of this study was to improve a non-mechanized bulk encapsulation technique to standardize encapsulation procedures and reduce the labor time compared to encapsulating individual nodes. Four mm-long nodal segments from Stage II cultures of Hibiscus moscheutos L. ‘Lord Baltimore’ were encapsu...

  17. Time-Dependent Effect of Encapsulating Alginate Hydrogel on Neurogenic Potential

    Science.gov (United States)

    Razavi, Shahnaz; Khosravizadeh, Zahra; Bahramian, Hamid; Kazemi, Mohammad

    2015-01-01

    Objective Due to the restricted potential of neural stem cells for regeneration of central nervous system (CNS) after injury, providing an alternative source for neural stem cells is essential. Adipose derived stem cells (ADSCs) are multipotent cells with properties suitable for tissue engineering. In addition, alginate hydrogel is a biocompatible polysaccharide polymer that has been used to encapsulate many types of cells. The aim of this study was to assess the proliferation rate and level of expression of neural markers; NESTIN, glial fibrillary acidic protein (GFAP) and microtubule-associated protein 2 (MAP2) in encapsulated human ADSCs (hADSCs) 10 and14 days after neural induction. Materials and Methods In this experimental study, ADSCs isolated from human were cultured in neural induction media and seeded into alginate hydrogel. The rate of proliferation and differentiation of encapsulated cells were evaluated by 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl tetrazolium bromide (MTT) assay, immunocytoflourescent and realtime reverse transcriptase polymerase chain reaction (RT-PCR) analyzes 10 and 14 days after induction. Results The rate of proliferation of encapsulated cells was not significantly changed with time passage. The expression of NESTIN and GFAP significantly decreased on day 14 relative to day 10 (P<0.001) but MAP2 expression was increased. Conclusion Alginate hydrogel can promote the neural differentiation of encapsulated hADSCs with time passage. PMID:26199909

  18. Encapsulation of liquid smoke flavoring in ca-alginate and ca-alginate-chitosan beads.

    Science.gov (United States)

    Petzold, Guillermo; Gianelli, María Pia; Bugueño, Graciela; Celan, Raymond; Pavez, Constanza; Orellana, Patricio

    2014-01-01

    Encapsulation is a technique used in foods that may protect some compounds with sensory impact, in particular flavoring as liquid smoke. We used the dripping method, obtaining two different layers for encapsulation of liquid smoke: calcium alginate and calcium alginate-chitosan. The results show that the load capacity of liquid smoke encapsulation reached values above 96 %. The beads exhibit syneresis at room temperature, but in opposite side, refrigeration temperature stabilizes the hydrogel of beads, allowing the samples loss weight less than 3 % after 72 h. Heated capsules with liquid smoke released several volatile compounds in the headspace and may identify 66 compounds. Among these volatile compounds, phenols derivatives can be considered sensory descriptors to contribute to the specific flavor of smoke. We conclude that the dripping method is highly efficient to encapsulate liquid smoke and released several volatile compounds, although it is necessary to minimize syneresis at room temperature.

  19. Identification of IL-1β and LPS as optimal activators of monolayer and alginate-encapsulated mesenchymal stromal cell immunomodulation using design of experiments and statistical methods.

    Science.gov (United States)

    Gray, Andrea; Maguire, Timothy; Schloss, Rene; Yarmush, Martin L

    2015-01-01

    Induction of therapeutic mesenchymal stromal cell (MSC) function is dependent upon activating factors present in diseased or injured tissue microenvironments. These functions include modulation of macrophage phenotype via secreted molecules including prostaglandin E2 (PGE2). Many approaches aim to optimize MSC-based therapies, including preconditioning using soluble factors and cell immobilization in biomaterials. However, optimization of MSC function is usually inefficient as only a few factors are manipulated in parallel. We utilized fractional factorial design of experiments to screen a panel of 6 molecules (lipopolysaccharide [LPS], polyinosinic-polycytidylic acid [poly(I:C)], interleukin [IL]-6, IL-1β, interferon [IFN]-β, and IFN-γ), individually and in combinations, for the upregulation of MSC PGE2 secretion and attenuation of macrophage secretion of tumor necrosis factor (TNF)-α, a pro-inflammatory molecule, by activated-MSC conditioned medium (CM). We used multivariable linear regression (MLR) and analysis of covariance to determine differences in functions of optimal factors on monolayer MSCs and alginate-encapsulated MSCs (eMSCs). The screen revealed that LPS and IL-1β potently activated monolayer MSCs to enhance PGE2 production and attenuate macrophage TNF-α. Activation by LPS and IL-1β together synergistically increased MSC PGE2, but did not synergistically reduce macrophage TNF-α. MLR and covariate analysis revealed that macrophage TNF-α was strongly dependent on the MSC activation factor, PGE2 level, and macrophage donor but not MSC culture format (monolayer versus encapsulated). The results demonstrate the feasibility and utility of using statistical approaches for higher throughput cell analysis. This approach can be extended to develop activation schemes to maximize MSC and MSC-biomaterial functions prior to transplantation to improve MSC therapies. © 2015 American Institute of Chemical Engineers.

  20. Rapid one-step purification of single-cells encapsulated in alginate microcapsules from oil to aqueous phase using a hydrophobic filter paper: implications for single-cell experiments.

    Science.gov (United States)

    Lee, Do-Hyun; Jang, Miran; Park, Je-Kyun

    2014-10-01

    By virtue of the biocompatibility and physical properties of hydrogel, picoliter-sized hydrogel microcapsules have been considered to be a biometric signature containing several features similar to that of encapsulated single cells, including phenotype, viability, and intracellular content. To maximize the experimental potential of encapsulating cells in hydrogel microcapsules, a method that enables efficient hydrogel microcapsule purification from oil is necessary. Current methods based on centrifugation for the conventional stepwise rinsing of oil, are slow and laborious and decrease the monodispersity and yield of the recovered hydrogel microcapsules. To remedy these shortcomings we have developed a simple one-step method to purify alginate microcapsules, containing a single live cell, from oil to aqueous phase. This method employs oil impregnation using a commercially available hydrophobic filter paper without multistep centrifugal purification and complicated microchannel networks. The oil-suspended alginate microcapsules encapsulating single cells from mammalian cancer cell lines (MCF-7, HepG2, and U937) and microorganisms (Chlorella vulgaris) were successfully exchanged to cell culture media by quick (~10 min) depletion of the surrounding oil phase without coalescence of neighboring microcapsules. Cell proliferation and high integrity of the microcapsules were also demonstrated by long-term incubation of microcapsules containing a single live cell. We expect that this method for the simple and rapid purification of encapsulated single-cell microcapsules will attain widespread adoption, assisting cell biologists and clinicians in the development of single-cell experiments. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Enhancement of survival of alginate-encapsulated Lactobacillus casei NCDC 298.

    Science.gov (United States)

    Mandal, Surajit; Hati, Subrota; Puniya, Anil Kumar; Khamrui, Kaushik; Singh, Kishan

    2014-08-01

    Micro-encapsulation of hydrocolloids improves the survival of sensitive probiotic bacteria in the harsh conditions that prevail in foods and during gastrointestinal passage by segregating them from environments. Incorporation of additives in encapsulating hydrocolloids and coatings of microcapsules further improves the survival of the probiotics. In this study, the effect of incorporation of resistant-maize starch in alginate for micro-encapsulation and coating of microcapsules with poly-l-lysine, stearic acid and bees wax on the survival of encapsulated Lactobacillus casei NCDC 298 at pH 1.5, 2% high bile salt, 65 °C for 20 min and release of viable lactobacilli cells from the capsule matrix in simulated aqueous solutions of colonic pH were assessed. Addition of resistant maize starch (2%) improved the survival of encapsulated L. casei NCDC 298. Coating of microcapsules with poly-L-lysine did not further improve the protection of encapsulated cells from the harsh conditions; however, bees wax and stearic acid (2%) improved the survival under similar conditions. Incorporation of maize starch (2%) in alginate followed by coating of beads with stearic acid (2%) led to better protection and complete release of entrapped lactobacilli in simulated colonic pH solution was observed. Additional treatments improve the survival of alginate-encapsulated lactobacilli cells without hindering the release of active cells from the capsule matrix and hence, the resulting encapsulated probiotics can be exploited in the development of probiotic functional foods with better survival of sensitive probiotic organisms. © 2013 Society of Chemical Industry.

  2. The in vitro and in vivo capacity of culture-expanded human cells from several sources encapsulated in alginate to form cartilage

    Directory of Open Access Journals (Sweden)

    MM Pleumeekers

    2014-04-01

    Full Text Available Cartilage has limited self-regenerative capacity. Tissue engineering can offer promising solutions for reconstruction of missing or damaged cartilage. A major challenge herein is to define an appropriate cell source that is capable of generating a stable and functional matrix. This study evaluated the performance of culture-expanded human chondrocytes from ear (EC, nose (NC and articular joint (AC, as well as bone-marrow-derived and adipose-tissue-derived mesenchymal stem cells both in vitro and in vivo. All cells (≥ 3 donors per source were culture-expanded, encapsulated in alginate and cultured for 5 weeks. Subsequently, constructs were implanted subcutaneously for 8 additional weeks. Before and after implantation, glycosaminoglycan (GAG and collagen content were measured using biochemical assays. Mechanical properties were determined using stress-strain-indentation tests. Hypertrophic differentiation was evaluated with qRT-PCR and subsequent endochondral ossification with histology. ACs had higher chondrogenic potential in vitro than the other cell sources, as assessed by gene expression and GAG content (p < 0.001. However, after implantation, ACs did not further increase their matrix. In contrast, ECs and NCs continued producing matrix in vivo leading to higher GAG content (p < 0.001 and elastic modulus. For NC-constructs, matrix-deposition was associated with the elastic modulus (R2 = 0.477, p = 0.039. Although all cells – except ACs – expressed markers for hypertrophic differentiation in vitro, there was no bone formed in vivo. Our work shows that cartilage formation and functionality depends on the cell source used. ACs possess the highest chondrogenic capacity in vitro, while ECs and NCs are most potent in vivo, making them attractive cell sources for cartilage repair.

  3. Properties of Lactobacillus reuteri chitosan-calcium-alginate encapsulation under simulated gastrointestinal conditions.

    Science.gov (United States)

    Huang, Hui-Ying; Tang, Yi-Ju; King, V An-Erl; Chou, Jen-Wei; Tsen, Jen-Horng

    2015-03-01

    The protective effects of encapsulation on the survival of Lactobacillus reuteri and the retention of the bacterium's probiotic properties under simulated gastrointestinal conditions were investigated. Viable counts and the remaining probiotic properties of calcium (Ca)-alginate encapsulated (A group), chitosan-Ca-alginate encapsulated (CA group), and unencapsulated, free L. reuteri (F group) were determined. Encapsulation improved the survival of L. reuteri subjected to simulated gastrointestinal conditions, with the greatest protective effect achieved in the CA group. The degree of cell membrane injury increased with increasing bile salt concentrations at constant pH, but the extent of injury was less in the encapsulated than in the free cells. Adherence rates were, in descending order: CA (0.524%)>A (0.360%)>F (0.275%). Lactobacillus reuteri cells retained their antagonistic activity toward Listeria monocytogenes even after incubation of the lactobacilli under simulated gastrointestinal conditions. Displacement of the pathogen by cells released from either of the encapsulation matrices was higher than that by free cells. The safety of L. reuteri was demonstrated in an in vitro invasion assay. Copyright© by the Spanish Society for Microbiology and Institute for Catalan Studies.

  4. Cytoprotective alginate/polydopamine core/shell microcapsules in microbial encapsulation.

    Science.gov (United States)

    Kim, Beom Jin; Park, Taegyun; Moon, Hee Chul; Park, So-Young; Hong, Daewha; Ko, Eun Hyea; Kim, Ji Yup; Hong, Jong Wook; Han, Sang Woo; Kim, Yang-Gyun; Choi, Insung S

    2014-12-22

    Chemical encapsulation of microbes in threedimensional polymeric microcapsules promises various applications, such as cell therapy and biosensors, and provides a basic platform for studying microbial communications. However, the cytoprotection of microbes in the microcapsules against external aggressors has been a major challenge in the field of microbial microencapsulation, because ionotropic hydrogels widely used for microencapsulation swell uncontrollably, and are physicochemically labile. Herein, we developed a simple polydopamine coating for obtaining cytoprotective capability of the alginate capsule that encapsulated Saccharomyces cerevisiae. The resulting alginate/ polydopamine core/shell capsule was mechanically tough, prevented gel swelling and cell leakage, and increased resistance against enzymatic attack and UV-C irradiation. We believe that this multifunctional core/shell structure will provide a practical tool for manipulating microorganisms inside the microcapsules. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  5. Encapsulation of urease enzyme in xanthan-alginate spheres.

    Science.gov (United States)

    Elçin, Y M

    1995-10-01

    Urease-containing xanthan-alginate spheres were prepared by a two-step process which involved the Ca2+ coupling of the polysaccharides, followed by gentle glutaraldehyde cross-linking with amine groups of gelatin present in the initial mixture. This second step caused a slight decrease in the enzymatic activity but increased the stability. The water content and size distribution of the spheres were examined together with the sphere morphology. The effect of polymer ratio and enzyme loading on urease activity was investigated. An increase in xanthan content was found to affect the water uptake of the spheres. Temperature and pH stability of encapsulated urease was found to be higher than the free form. The xanthan-alginate spheres showed 75% of maximum urease activity even after 20 repeated uses under optimal conditions.

  6. Boar sperm changes after sorting and encapsulation in barium alginate membranes.

    Science.gov (United States)

    Spinaci, M; Bucci, D; Chlapanidas, T; Vallorani, C; Perteghella, S; Communod, R; Vigo, D; Tamanini, C; Galeati, G; Faustini, M; Torre, M L

    2013-09-15

    A routine use of boar-sexed semen is limited by the long sorting time necessary to obtain an adequate number of sexed spermatozoa for artificial insemination and by the high susceptibility of spermatozoa of this species to damages induced by sorting procedure and subsequent cryopreservation. The aim of this work was to study the impact of encapsulation in barium alginate membrane on sorted boar spermatozoa by evaluating membrane integrity, chlortetracycline staining patterns, protein tyrosine phosphorylation, and Hsp70 immunolocalization during storage over 72 hours in liquid or encapsulated form. The encapsulation procedure significantly (P < 0.05) decreased the overall membrane integrity of control unsorted semen (81.8 vs. 57.4, CTR vs. CPS), but did not negatively affect the overall viability and the chlortetracycline staining patterns of sorted encapsulated cells. Moreover, encapsulation significantly decreased (P < 0.05) the overall phosphotyrosin A pattern cell percentage in unsorted (98.4 vs. 92.6, CTR vs. CPS) but not in sorted semen (64.0 vs. 74.2; SORT CTR vs. SORT CPS). As for Hsp70, the overall percentage of cells displaying the different patterns was significantly influenced (P < 0.05) by treatment but not by storage time. The sorting procedure seems to induce the major changes, whereas encapsulation tends to exert a protective effect on sorted semen by increasing the percentage of spermatozoa displaying the T pattern (2.8 vs. 24.3; SORT CTR vs. SORT CPS). In conclusion, our data confirm that the damaging impact of the encapsulation in barium alginate capsules seems to be limited when compared with that of the sorting procedure and, moreover, the association of the two procedures does not result in an algebraic sum of the negative effects. These results suggest the possibility of a future utilization of the encapsulation technology in order to store sorted spermatozoa and permit their controlled release in the female genital tract.

  7. Pulsed-low intensity ultrasound enhances extracellular matrix production by fibroblasts encapsulated in alginate

    Directory of Open Access Journals (Sweden)

    Siti PM Bohari

    2012-12-01

    Full Text Available In this study, the effect of pulsed-low intensity ultrasound on cell proliferation, collagen production and glycosaminoglycan deposition by 3T3 fibroblasts encapsulated in alginate was evaluated. Hoechst 33258 assay for cell number, hydroxyproline assay for collagen content and dimethylamine blue assay for glycosaminoglycan content were performed on samples from cell cultures treated with pulsed-low intensity ultrasound and a control group. Pulsed-low intensity ultrasound shows no effect on cell proliferation, while collagen and glycosaminoglycan contents were consistently higher in the samples treated with pulsed-low intensity ultrasound, showing a statistically significant difference (p < 0.05 on day 10. Alcian blue staining showed that glycosaminoglycans were deposited around the cells in both groups. These results suggest that pulsed-low intensity ultrasound shows no effect on cell proliferation but has potential for inducing collagen and glycosaminoglycan production in cells cultured in alginate gels.

  8. Encapsulation of phycocyanin-alginate for high stability and antioxidant activity

    Science.gov (United States)

    Hadiyanto; Suzery, Meiny; Setyawan, Deny; Majid, Dian; Sutanto, Heri

    2017-02-01

    The aim of this study was to obtain optimal condition of phycocyanin-alginate encapsulation, encapsulation efficiency and phycocyanin load, physicochemical properties of beads, in vitro release study, stability and antioxidant activity. The result product with alginate content 1,5% (w/v) and 2% (w/v) produced were in spherical shape than product with alginate content 2,5% (w/v) by ratio of phycocyanin 1:1. Increasing alginate content on encapsulation process will increase of encapsulation efficiency and phycocyanin load. In vitro released study showed that phycocyanin-alginate beads were more resistant in simulated gastric fluid, while rapidly release in simulated intestinal fluid. The antioxidant activity showed that phycocyanin antioxidant activity decreased after encapsulation process due to duration of storage and the possibility of a cracking which will cause reduced stability of phycocyanin.

  9. Encapsulation of probiotic bacteria with alginate-starch and evaluation of survival in simulated gastrointestinal conditions and in yoghurt.

    Science.gov (United States)

    Sultana, K; Godward, G; Reynolds, N; Arumugaswamy, R; Peiris, P; Kailasapathy, K

    2000-12-05

    A modified method using calcium alginate for the microencapsulation of probiotic bacteria is reported in this study. Incorporation of Hi-Maize starch (a prebiotic) improved encapsulation of viable bacteria as compared to when the bacteria were encapsulated without the starch. Inclusion of glycerol (a cryo-protectant) with alginate mix increased the survival of bacteria when frozen at -20 degrees C. The acidification kinetics of encapsulated bacteria showed that the rate of acid produced was lower than that of free cultures. The encapsulated bacteria, however, did not demonstrate a significant increase in survival when subjected to in vitro high acid and bile salt conditions. A preliminary study was carried out in order to monitor the effects of encapsulation on the survival of Lactobacillus acidophilus and Bifidobacterium spp. in yoghurt over a period of 8 weeks. This study showed that the survival of encapsulated cultures of L. acidophilus and Bifidobacterium spp. showed a decline in viable count of about 0.5 log over a period of 8 weeks while there was a decline of about 1 log in cultures which were incorporated as free cells in yoghurt. The encapsulation method used in this study did not result in uniform bead size, and hence additional experiments need to be designed using uniform bead size in order to assess the role of different encapsulation parameters, such as bead size and alginate concentration, in providing protection to the bacteria.

  10. Encapsulation in alginate and alginate coated-chitosan improved the survival of newly probiotic in oxgall and gastric juice.

    Science.gov (United States)

    Trabelsi, Imen; Bejar, Wacim; Ayadi, Dorra; Chouayekh, Hichem; Kammoun, Radhouane; Bejar, Samir; Ben Salah, Riadh

    2013-10-01

    This study was undertaken to develop an optimum composition model for the microencapsulation of a newly probiotic on sodium alginate using response surface methodology. The individual and interactive effects of three independent variables, namely sodium alginate concentration, biomass concentration, and hardening time, were investigated using Box-Behnken design experiments. A second ordered polynomial model was fitted and optimum conditions were estimated. The optimal conditions identified were 2% for sodium alginate, 10(10)UFC/ml for biomass, and 30 min for hardening time. The experimental value obtained for immobilized cells under these conditions was about 80.98%, which was in close agreement with the predicted value of 82.6%. Viability of microspheres (96%) was enhanced with chitosan as coating materials. The survival rates of free and microencapsulated Lactobacillus plantarum TN8 during exposure to artificial gastrointestinal conditions were compared. The results revealed that the encapsulated cells exhibited significantly higher resistances to artificial intestinal juice (AIJ) and artificial gastric juice (AGJ). Microencapsulation was also noted to effectively protect the strain from heating at 65 °C and refrigerating at 4 °C. Taken together, the findings indicated that microencapsulation conferred important protective effects to L. plantarum against the gastrointestinal conditions encountered during the transit of food. Copyright © 2013 Elsevier B.V. All rights reserved.

  11. Encapsulation of volatiles by homogenized partially-cross linked alginates.

    Science.gov (United States)

    Inguva, Pavan K; Ooi, Shing Ming; Desai, Parind M; Heng, Paul W S

    2015-12-30

    Cross-linked calcium alginate gels are too viscous to be efficaciously incorporated into spray dried formulations. Thus, viscosity reduction is essential to ensure the processability of calcium alginate gels to be sprayed. Viscosity reduction by high pressure homogenization can open new formulation possibilities. Presently, testing of microcapsule integrity is also limited because either single particle tests neglect collective particle behaviours in bulk or bulk testing methods are often associated with single compressions which may not fully characterize individual particle strengths. The aim of this study was sub-divided into three objectives. First objective was to evaluate the impact of high pressure homogenization on gel viscosity. Second objective was to explore the use of the homogenized gels with modified starch for microencapsulation by spray drying. The final objective was to develop a stamping system as microcapsule strength tester that can assess microcapsules in bulk and evaluate the impact of multiple compressions. Collectively, this study would lead towards developing a pressure-activated patch of microcapsules with encapsulated volatiles and the method to assess the patch efficacy. The alginate gels largely experienced an exponential decay in viscosity when homogenized. Furthermore, the homogenized gels were successfully incorporated in spray drying formulations for microencapsulation. The custom-designed microcapsule strength tester was successfully used and shown to possess the required sensitivity to discern batches of microcapsules containing volatiles to have different release profiles. Addition of homogenized gels strengthened the microcapsules only at high wall to core ratios with low mass-load alginate gels. High mass-load gels weaken the microcapsules, exhibiting a higher release at low stamping pressures and wrinkling on the microcapsules surface.

  12. Highly Efficient Malolactic Fermentation of Red Wine Using Encapsulated Bacteria in a Robust Biocomposite of Silica-Alginate.

    Science.gov (United States)

    Simó, Guillermo; Vila-Crespo, Josefina; Fernández-Fernández, Encarnación; Ruipérez, Violeta; Rodríguez-Nogales, José Manuel

    2017-06-28

    Bacteria encapsulation to develop malolactic fermentation emerges as a biotechnological strategy that provides significant advantages over the use of free cells. Two encapsulation methods have been proposed embedding Oenococcus oeni, (i) interpenetrated polymer networks of silica and Ca-alginate and (ii) Ca-alginate capsules coated with hydrolyzed 3-aminopropyltriethoxysilane (hAPTES). On the basis of our results, only the first method was suitable for bacteria encapsulation. The optimized silica-alginate capsules exhibited a negligible bacteria release and an increase of 328% and 65% in L-malic acid consumption and mechanical robustness, respectively, compared to untreated alginate capsules. Moreover, studies of capsule stability at different pH and ethanol concentrations in water solutions and in wine indicated a better behavior of silica-alginate capsules than untreated ones. The inclusion of silicates and colloidal silica in alginate capsules containing O. oeni improved markedly their capacity to deplete the levels of L-malic acid in red wines and their mechanical robustness and stability.

  13. Encapsulation of sorbitan ester-based organogels in alginate microparticles.

    Science.gov (United States)

    Sagiri, Sai S; Pal, Kunal; Basak, Piyali; Rana, Usman Ali; Shakir, Imran; Anis, Arfat

    2014-10-01

    Leaching of the internal apolar phase from the biopolymeric microparticles during storage is a great concern as it undoes the beneficial effects of encapsulation. In this paper, a novel formulation was prepared by encapsulating the sunflower oil-based organogels in alginate microparticles. Salicylic acid and metronidazole were used as the model drugs. The microparticles were prepared by double emulsion methodology. Physico-chemical characterization of the microparticles was done by microscopy, FTIR, XRD, and DSC studies. Oil leaching studies, biocompatibility, mucoadhesivity, in vitro drug release, and the antimicrobial efficiency of the microparticles were also performed. The microparticles were found to be spherical in shape. Gelation of the sunflower oil prevented leaching of the internal phase from the microparticles. Release of drugs from the microparticles followed Fickian kinetics and non-Fickian kinetics in gastric and intestinal environments, respectively. Microparticles showed good antimicrobial activity against both Gram-positive (Bacillus subtilis) and Gram-negative (Escherichia coli) bacteria. The results suggested that the developed formulations hold promise to carry oils without leakage of the internal phase. Encapsulation of organogels within the microparticles has improved the drug entrapment efficiency and improved characteristics for controlled delivery applications.

  14. Survivability of probiotics encapsulated in alginate gel microbeads using a novel impinging aerosols method.

    Science.gov (United States)

    Sohail, Asma; Turner, Mark S; Coombes, Allan; Bostrom, Thor; Bhandari, Bhesh

    2011-01-31

    Encapsulation of probiotic bacteria in cross-linked alginate beads is of major interest for improving the survivability in harsh acid and bile environment and also in food matrices. Alginate micro beads (10-40 μm) containing the probiotics Lactobacillus rhamnosus GG and Lactobacillus acidophilus NCFM were produced by a novel technique based on dual aerosols of alginate solution and CaCl(2) cross linking solution. Extruded macro beads (approximately 2mm diameter) produced by the conventional method and micro beads produced by novel aerosols technique offered comparable protection to L. rhamnosus in high acid and bile environment. Chitosan coating of micro beads resulted in a significant increase in survival time of L. rhamnosus from 40 to 120 min in acid condition and the reduction in cell numbers was confined to 0.94 log over this time. Alginate macro beads are more effective than micro beads in protecting L. acidophilus against high acid and bile. Chitosan coating of micro beads resulted in similar protection to L. acidophilus in macro beads in acid and extended the survival time from 90 to at least 120 min. Viability of this organism in micro beads was 3.5 log after 120 min. The continuous processing capability and scale-up potential of the dual aerosol technique offers potential for an efficient encapsulation of probiotics in very small alginate micro beads below sensorial detection limits while still being able to confer effective protection in acid and bile environment. Copyright © 2010. Published by Elsevier B.V.

  15. Preparation, characterisation and viability of encapsulated Trichoderma harzianum UPM40 in alginate-montmorillonite clay.

    Science.gov (United States)

    Adzmi, Fariz; Meon, Sariah; Musa, Mohamed Hanafi; Yusuf, Nor Azah

    2012-01-01

    Microencapsulation is a process by which tiny parcels of an active ingredient are packaged within a second material for the purpose of shielding the active ingredient from the surrounding environment. This study aims to determine the ability of the microencapsulation technique to improve the viability of Trichoderma harzianum UPM40 originally isolated from healthy groundnut roots as effective biological control agents (BCAs). Alginate was used as the carrier for controlled release, and montmorillonite clay (MMT) served as the filler. The encapsulated Ca-alginate-MMT beads were characterised using Fourier transform infrared spectroscopy (FTIR), thermogravimetric analysis (TGA) and scanning electron microscopy (SEM). The FTIR results showed the interaction between the functional groups of alginate and MMT in the Ca-alginate-MMT beads. Peaks at 1595, 1420 and 1020 cm(-1) characterised alginate, and peaks at 1028 and 453 cm(-1) characterised MMT; both sets of peaks appeared in the Ca-alginate-MMT FTIR spectrum. The TGA analysis showed an improvement in the thermal stability of the Ca-alginate-MMT beads compared with the alginate beads alone. SEM analysis revealed a homogeneous distribution of the MMT particles throughout the alginate matrix. T. harzianum UPM40 was successfully encapsulated in the Ca-alginate-MMT beads. Storage analysis of the encapsulated T. harzianum UPM40 showed that the low storage temperature of 5°C resulted in significantly (p < 0.05) better storage compared with room temperature (30°C).

  16. Coating of alginate capsules

    OpenAIRE

    Hadjialirezaei, Soosan

    2013-01-01

    Alginate is a popular candidate for encapsulation of cells due to the formation of gels with divalent ions under physiological conditions. Stable alginate gels can be formed by the selection of alginates with a high content of guluronic acid (G) and gelling in a mixture of calcium and barium. These alginate gels have been proposed as immune protective barriers for the transplantation of human pancreatic islets (insulin producing cells) for the treatment of type 1 diabetes where the alginate g...

  17. Demonstrating Encapsulation and Release: A New Take on Alginate Complexation and the Nylon Rope Trick

    Science.gov (United States)

    Friedli, Andrienne C.; Schlager, Inge R.; Wright, Stephen W.

    2005-01-01

    Three variations on a classroom demonstration of the encapsulation of droplets and evidence for release of the interior solution are described. The first two demonstrations mimic biocompatible applications of encapsulation. Reversible formation of capsules from aqueous solutions of sodium alginate, a negatively charged polysaccharide derived from…

  18. Exploring Catalase and Invertase Activity Using Sodium Alginate-Encapsulated Yeast (Yeast Spheres

    Directory of Open Access Journals (Sweden)

    Pamela J. Bryer

    2016-12-01

    Full Text Available Finding the right enzyme experiment can be problematic, depending what one is trying to show, what supplies and equipment are available, and the time one can devote to the topic.  I’ve developed simple and inexpensive labs for looking at catalase and invertase activity using yeast encapsulated in sodium alginate.  Single-celled yeast, Saccharomyces cerevisiae, are encapsulated in sodium alginate, a readily available extract from brown algae that, when it comes in contact with calcium chloride (CaCl2, forms a sphere or “bead.”  These spheres may then be put into a solution containing substrate to test for enzyme activity.  The spheres are easy to manipulate, one doesn’t have the variability and mess of a yeast solution, and since there are no cells in solution, there is nothing to interfere with the various assay methods one might want to use to test for product.  The graduated cylinder method for testing catalase activity introduced here is especially good for collecting large amounts of data that enable students to use statistics and, unlike similar yeast catalase experiments using paper disks and a yeast solution, the yeast spheres are easy to manipulate and there is very little variability.  I have used this procedure with students in class and with teachers in workshops with positive results and comments.

  19. Proteomic Analysis of the Increased Stress Tolerance of Saccharomyces cerevisiae Encapsulated in Liquid Core Alginate-Chitosan Capsules

    Science.gov (United States)

    Westman, Johan O.; Taherzadeh, Mohammad J.; Franzén, Carl Johan

    2012-01-01

    Saccharomyces cerevisiae CBS8066 encapsulated in semi-permeable alginate or alginate-chitosan liquid core capsules have been shown to have an enhanced tolerance towards complex dilute-acid lignocellulose hydrolysates and the lignocellulose-derived inhibitor furfural, as well as towards high temperatures. The underlying molecular reasons for these effects have however not been elucidated. In this study we have investigated the response of the encapsulation on the proteome level in the yeast cells, in comparison with cells grown freely in suspension under otherwise similar conditions. The proteomic analysis was performed on whole cell protein extracts using nLC-MS/MS with TMT® labelling and 2-D DIGE. 842 and 52 proteins were identified using each method, respectively. The abundances of 213 proteins were significantly different between encapsulated and suspended cells, with good correlation between the fold change ratios obtained by the two methods for proteins identified in both. Encapsulation of the yeast caused an up-regulation of glucose-repressed proteins and of both general and starvation-specific stress responses, such as the trehalose biosynthesis pathway, and down-regulation of proteins linked to growth and protein synthesis. The encapsulation leads to a lack of nutrients for cells close to the core of the capsule due to mass transfer limitations. The triggering of the stress response may be beneficial for the cells in certain conditions, for example leading to the increased tolerance towards high temperatures and certain inhibitors. PMID:23152898

  20. Proteomic analysis of the increased stress tolerance of saccharomyces cerevisiae encapsulated in liquid core alginate-chitosan capsules.

    Directory of Open Access Journals (Sweden)

    Johan O Westman

    Full Text Available Saccharomyces cerevisiae CBS8066 encapsulated in semi-permeable alginate or alginate-chitosan liquid core capsules have been shown to have an enhanced tolerance towards complex dilute-acid lignocellulose hydrolysates and the lignocellulose-derived inhibitor furfural, as well as towards high temperatures. The underlying molecular reasons for these effects have however not been elucidated. In this study we have investigated the response of the encapsulation on the proteome level in the yeast cells, in comparison with cells grown freely in suspension under otherwise similar conditions. The proteomic analysis was performed on whole cell protein extracts using nLC-MS/MS with TMT® labelling and 2-D DIGE. 842 and 52 proteins were identified using each method, respectively. The abundances of 213 proteins were significantly different between encapsulated and suspended cells, with good correlation between the fold change ratios obtained by the two methods for proteins identified in both. Encapsulation of the yeast caused an up-regulation of glucose-repressed proteins and of both general and starvation-specific stress responses, such as the trehalose biosynthesis pathway, and down-regulation of proteins linked to growth and protein synthesis. The encapsulation leads to a lack of nutrients for cells close to the core of the capsule due to mass transfer limitations. The triggering of the stress response may be beneficial for the cells in certain conditions, for example leading to the increased tolerance towards high temperatures and certain inhibitors.

  1. Dodecenyl succinylated alginate as a novel material for encapsulation and hyperactivation of lipases.

    Science.gov (United States)

    Falkeborg, Mia; Paitaid, Pattarapon; Shu, Allen Ndonwi; Pérez, Bianca; Guo, Zheng

    2015-11-20

    Alginate was modified with dodecenyl succinic anhydride (SAC12) in an aqueous reaction medium at neutral pH. The highest degree of succinylation (33.9±3.5%) was obtained after 4h at 30°C, using four mole SAC12 per mol alginate monomer. Alginate was modified with succinic anhydride (SAC0) for comparison, and the structures and thermal properties of alg-SAC0 and alg-SAC12 were evaluated using FTIR, (1)H NMR, and DSC. Calcium-hydrogel beads were formed from native and modified alginates, in which lipases were encapsulated with a load of averagely 76μg lipase per mg alginate, irrespective of the type of alginate. Lipases with a "lid", which usually are dependent on interfacial activation, showed a 3-fold increase in specific activity toward water-soluble substrates when encapsulated in alg-SAC12, compared to the free lipase. Such hyperactivation was not observed for lipases independent of interfacial activation, or for lipases encapsulated in native alginate or alg-SAC0 hydrogels.

  2. Encapsulated Escherichia coli in alginate beads capable of secreting a heterologous pectin lyase

    Directory of Open Access Journals (Sweden)

    Trikka Fotini A

    2005-12-01

    Full Text Available Abstract Background Production of heterologous proteins in the E. coli periplasm, or into the extracellular fluid has many advantages; therefore naturally occurring signal peptides are selected for proteins translocation. The aim of this study was the production in high yields of a recombinant pectin lyase that is efficiently secreted and the encapsulation of transformed E. coli cells for pectin degradation in a biotechnological process. Results The nucleotide sequence of Bacillus subtilis α-amylase's signal peptide was fused to the N-terminal of an heterologously expressed pectin lyase in E. coli BL21 [DE3]. Thus pectin lyase secretion was achieved into the extracellular growth medium. E. coli cells harboring the recombinant plasmid heterologously express pectin lyase to around 22% of the total cellular proteins, as it was estimated by SDS-PAGE and image analysis. IPTG induces the heterologously expressed enzyme, which is initially distributed extracellularly (7 hour and later on at the periplasmic (9 hours or cytosolic fraction (20 hours. No pectin lyase activity was found in the membranes fraction and in the inclusion bodies. Encapsulation of the recombinant strains of E. coli in alginate or alginate/silica beads 1:5 showed that pectin lyase could degrade effectively its substrate, for at least ten operational cycles. Conclusion Secretion of an heterologously overexpressed pectin lyase in E. coli BL21 [DE3] was achieved in this study. For this purpose the signal peptide of α-amylase from B. subtilis was fused to the N-terminal domain of pectin lyase. Encapsulated E. coli BL21 [DE3] cells harboring pET29c/exPNL were used successfully for pectin degradation up to ten operational cycles indicating that under special conditions this might have biotechnological implementations.

  3. HAp granules encapsulated oxidized alginate-gelatin-biphasic calcium phosphate hydrogel for bone regeneration.

    Science.gov (United States)

    Sarker, Avik; Amirian, Jhaleh; Min, Young Ki; Lee, Byong Taek

    2015-11-01

    Bone repair in the critical size defect zone using 3D hydrogel scaffold is still a challenge in tissue engineering field. A novel type of hydrogel scaffold combining ceramic and polymer materials, therefore, was fabricated to meet this challenge. In this study, oxidized alginate-gelatin-biphasic calcium phosphate (OxAlg-Gel-BCP) and spherical hydroxyapatite (HAp) granules encapsulated OxAlg-Gel-BCP hydrogel complex were fabricated using freeze-drying method. Detailed morphological and material characterizations of OxAlg-Gel-BCP hydrogel (OGB00), 25wt% and 35wt% granules encapsulated hydrogel (OGB25 and OGB35) were carried out for micro-structure, porosity, chemical constituents, and compressive stress analysis. Cell viability, cell attachment, proliferation and differentiation behavior of rat bone marrow-derived stem cell (BMSC) on OGB00, OGB25 and OGB35 scaffolds were confirmed by MTT assay, Live-Dead assay, and confocal imaging in vitro experiments. Finally, OGB00 and OGB25 hydrogel scaffolds were implanted in the critical size defect of rabbit femoral chondyle for 4 and 8 weeks. The micro-CT analysis and histological studies conducted by H&E and Masson's trichrome demonstrated that a significantly higher (***phydrogel than in OxAlg-Gel-BCP complex alone. All results taken together, HAp granules encapsulated OxAlg-Gel-BCP system can be a promising 3D hydrogel scaffold for the healing of a critical bone defect.

  4. Alginate as a cell culture substrate for growth and differentiation of human retinal pigment epithelial cells.

    Science.gov (United States)

    Heidari, Razeih; Soheili, Zahra-Soheila; Samiei, Shahram; Ahmadieh, Hamid; Davari, Maliheh; Nazemroaya, Fatemeh; Bagheri, Abouzar; Deezagi, Abdolkhalegh

    2015-03-01

    The purpose of this study was to evaluate retinal pigment epithelium (RPE) cells' behavior in alginate beads that establish 3D environment for cellular growth and mimic extracellular matrix versus the conventional 2D monolayer culture. RPE cells were encapsulated in alginate beads by dripping alginate cell suspension into CaCl2 solution. Beads were suspended in three different media including Dulbecco's modified Eagle's medium (DMEM)/F12 alone, DMEM/F12 supplemented with 10 % fetal bovine serum (FBS), and DMEM/F12 supplemented with 30 % human amniotic fluid (HAF). RPE cells were cultivated on polystyrene under the same conditions as controls. Cell phenotype, cell proliferation, cell death, and MTT assay, immunocytochemistry, and real-time RT-PCR were performed to evaluate the effect of alginate on RPE cells characteristics and integrity. RPE cells can survive and proliferate in alginate matrixes. Immunocytochemistry analysis exhibited Nestin, RPE65, and cytokeratin expressions in a reasonable number of cultured cells in alginate beads. Real-time PCR data demonstrated high levels of Nestin, CHX10, RPE65, and tyrosinase gene expressions in RPE cells immobilized in alginate when compared to 2D monolayer culture systems. The results suggest that alginate can be used as a reliable scaffold for maintenance of RPE cells' integrity and in vitro propagation of human retinal progenitor cells for cell replacement therapies in retinal diseases.

  5. Removal of Cr(VI) by zero-valent, iron-encapsulated alginate beads

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Xue Song; Tang, Yu Jun; Chen, Li Fang; Li, Fei Yan; Wan, Wen Ya; Tan, Ye Bin [Department of Chemical Engineering, Huaihai Institute of Technology, Jiangsu (China)

    2010-03-15

    Zero-valent, iron-encapsulated alginate beads were synthesized and were applied for the removal of Cr(VI) from aqueous solutions. The effects of several important parameters including solution pH, contact time, initial concentration and reaction temperature on Cr(VI) removal levels were investigated in batch studies. An initial solution pH of 1.0 was seen to be most favorable for Cr(VI) removal. The removal process was quick and almost 80% of the removal was attained within 60 min. The kinetic data followed the second-order equation well. The Cr(VI) removal was almost reaction temperature-independent and decreased with an increase in Cr(VI) initial concentration. The removal of Cr(VI) by iron-encapsulated alginate beads was found to be significantly higher than that of non-encapsulated alginate beads. (Abstract Copyright [2010], Wiley Periodicals, Inc.)

  6. The effect of alginate and chitosan concentrations on some properties of chitosan-coated alginate beads and survivability of encapsulated Lactobacillus rhamnosus in simulated gastrointestinal conditions and during heat processing.

    Science.gov (United States)

    Abbaszadeh, Sepideh; Gandomi, Hassan; Misaghi, Ali; Bokaei, Saeid; Noori, Negin

    2014-08-01

    In this study, chitosan-coated alginate beads were produced with different concentrations of chitosan and alginate to evaluate the survival of encapsulated Lactobacillus rhamnosus GG during exposure to adverse conditions in gastrointestinal simulated juice and heat processing. The encapsulation yield of different encapsulation treatments was between 25 and 53.2%. Although there was a drastic decrease in pH within 48 h of incubation in MRS medium inoculated with free and encapsulated bacteria, no significant changes (P > 0.05) in bacterial count were observed among different encapsulation treatments. Moreover, the survival rate after gastrointestinal juice exposure of all prepared beads was 10-87 times greater than that of free cells and was significantly enhanced by increasing chitosan and alginate concentrations. The encapsulated bacteria survived significantly (P encapsulated L. rhamnosus was reduced by only 2.55 log cycles. Encapsulation effectively protected L. rhamnosus against heat treatment and gastrointestinal conditions, and this effect is important in delivering sufficient numbers of viable probiotic bacteria to the gastrointestinal tract. © 2013 Society of Chemical Industry.

  7. Noninvasive evaluation of the vascular response to transplantation of alginate encapsulated islets using the dorsal skin-fold model.

    Science.gov (United States)

    Krishnan, Rahul; Arora, Rajan P; Alexander, Michael; White, Sean M; Lamb, Morgan W; Foster, Clarence E; Choi, Bernard; Lakey, Jonathan R T

    2014-01-01

    Alginate encapsulation reduces the risk of transplant rejection by evading immune-mediated cell injury and rejection; however, poor vascular perfusion results in graft failure. Since existing imaging models are incapable of quantifying the vascular response to biomaterial implants after transplantation, in this study, we demonstrate the use of in vivo laser speckle imaging (LSI) and wide-field functional imaging (WiFI) to monitor the microvascular environment surrounding biomaterial implants. The vascular response to two islet-containing biomaterial encapsulation devices, alginate microcapsules and a high-guluronate alginate sheet, was studied and compared after implantation into the mouse dorsal window chamber (N = 4 per implant group). Images obtained over a 14-day period using LSI and WiFI were analyzed using algorithms to quantify blood flow, hemoglobin oxygen saturation and vascular density. Using our method, we were able to monitor the changes in the peri-implant microvasculature noninvasively without the use of fluorescent dyes. Significant changes in blood flow, hemoglobin oxygen saturation and vascular density were noted as early as the first week post-transplant. The dorsal window chamber model enables comparison of host responses to transplanted biomaterials. Future experiments will study the effect of changes in alginate composition on the vascular and immune responses.

  8. Effect of acidification on the protection of alginate-encapsulated probiotic based on emulsification/internal gelation.

    Science.gov (United States)

    Qu, Fangning; Zhao, Meng; Fang, Yapeng; Nishinari, Katsuyoshi; Phillips, Glyn O; Wu, Zhengjun; Chen, Chen

    2016-10-01

    The method of emulsification/internal gelation is commonly used to prepare alginate microspheres for lactic acid bacteria (LAB). This paper focused on the influence of acidification parameters, i.e. acid/Ca molar ratio and acidification time, on the physical properties and cell protection efficiency of microspheres and their correlations. With increasing acid/Ca molar ratio and acidification time, the average diameter of microspheres decreased and their mechanical strength increased. Interestingly, wet alginate microspheres shrank in simulated gastric juice (SGJ) while they swelled in bile salts solution (BS). The shrinkage or swelling ratio decreased with increasing mechanical strength. Correlation analysis showed that the encapsulated cell survivals in both SGJ and BS were positively correlated with the mechanical strength of microspheres but negatively with the shrinkage or swelling ratio. BacLight LIVE/DEAD assay suggested that the viability of encapsulated cells in fresh, SGJ-treated and BS-treated microspheres was closely related to cell membrane integrity. Acidification is a key step during microsphere preparation, which strongly affected the physical properties of alginate microspheres, resulting in different cell protection efficiency. The resulting well-protected LAB can be applied in probiotics foods. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.

  9. A First Step in De Novo Synthesis of a Living Pulp Tissue Replacement Using Dental Pulp MSCs and Tissue Growth Factors, Encapsulated within a Bioinspired Alginate Hydrogel.

    Science.gov (United States)

    Bhoj, Manasi; Zhang, Chengfei; Green, David W

    2015-07-01

    A living, self-supporting pulp tissue replacement in vitro and for transplantation is an attractive yet unmet bioengineering challenge. Our aim is to create 3-dimensional alginate-based microenvironments that replicate the shape of gutta-percha and comprise key elements for the proliferation of progenitor cells and the release of growth factors. An RGD-bearing alginate framework was used to encapsulate dental pulp stem cells and human umbilical vein endothelial cells in a ratio of 1:1. The alginate hydrogel also retained and delivered 2 key growth factors, vascular endothelial growth factor-121 and fibroblast growth factor, in a sufficient amount to induce proliferation. A method was then devised to replicate the shape of gutta-percha using RGD alginate within a custom-made mold of thermoresponsive N-isopropylacrylamide. Plugs of alginate containing different permutations of growth factor-based encapsulates were tested and evaluated for viability, proliferation, and release kinetics between 1 and 14 days. According to scanning electron microscopic and confocal microscopic observations, the encapsulated human endothelial cells and dental pulp stem cell distribution were frequent and extensive throughout the length of the construct. There were also high levels of viability in all test environments. Furthermore, cell proliferation was higher in the growth factor-based groups. Growth factor release kinetics also showed significant differences between them. Interestingly, the combination of vascular endothelial growth factor and fibroblast growth factor synergize to significantly up-regulate cell proliferation. RGD-alginate scaffolds can be fabricated into shapes to fill the pulp space by simple templating. The addition of dual growth factors to cocultures of stem cells within RGD-alginate scaffolds led to the creation of microenvironments that significantly enhance the proliferation of dental pulp stem cell/human umbilical vein endothelial cell combinations. Copyright

  10. The efficacy of alginate encapsulated CHO-K1 single chain-TRAIL producer cells in the treatment of brain tumors

    NARCIS (Netherlands)

    Kuijlen, JMA; de Haan, BJ; Helfrich, W; de Boer, JF; Samplonius, D; Mooij, JJA; de Vos, P

    Objective: Patients with astrocytic tumors in the central nervous system (CNS) have low survival rates despite surgery and radiotherapy. Innovative therapies and strategies must be developed to prolong survival of these patients. The alginate microencapsulation method, used to continuously release a

  11. Protein extraction from Ca-alginate encapsulated plant material for comparative proteomic analysis.

    Science.gov (United States)

    Domżalska, Lucyna; Mikuła, Anna; Rybczyński, Jan J

    2016-10-01

    The extensive use of encapsulation material in biotechnology drove the need to develop analytical techniques for this type of material. This study focuses on the specific problems of protein extraction from Ca-alginate encapsulated plant material. Proteomics is one of the fast-developing analysis categories, specifically for stress resistance and developmental changes in plant material. Sample preparation is a critical step in a two-dimensional gel electrophoresis proteome approach and is essential for good results. The aim was to avoid preliminary manipulations and get good quality material for comparative proteome analysis technique 2DE. The phenol extraction method and the complex method with preliminary TCA precipitation, SDS buffer and phenol phase were compared with respect to the efficiency and quality of the resulting 2DE gel. The most appropriate method turned out to be the TCA/phenol method with the phenol fractioning technique adapted to the gentian cell suspension. It resulted in a high protein concentration and good quality sample that could be analyzed using the standard separation procedures of 2DE and spectrometric identification with high efficiency. The work presented here confirms the possibility of obtaining a sufficient protein sample for effective proteomic analysis from a small number of capsules.

  12. Influence of Flow Behavior of Alginate-Cell Suspensions on Cell Viability and Proliferation.

    Science.gov (United States)

    Ning, Liqun; Guillemot, Arthur; Zhao, Jingxuan; Kipouros, Georges; Chen, Xiongbiao

    2016-07-01

    Tissue scaffolds with living cells fabricated by three-dimensional bioprinting/plotting techniques are becoming more prevalent in tissue repair and regeneration. In the bioprinting process, cells are subject to process-induced forces (such as shear force) that can result in cell damage and loss of cell function. The flow behavior of the biomaterial solutions that encapsulate living cells in this process plays an important role. This study used a rheometer to examine the flow behavior of alginate solution and alginate-Schwann cell (RSC96), alginate-fibroblast cell (NIH-3T3), and alginate-skeletal muscle cell (L8) suspensions during shearing with respect to effects on cell viability and proliferation. The flow behavior of all the alginate-cell suspensions varied with alginate concentration and cell density and had a significant influence on the viability and proliferation of the cells once sheared as well as on the recovery of the sheared cells. These findings provide a mean to preserve cell viability and/or retain cell proliferation function in the bioprinting process by regulating the flow behavior of cell-biomaterial suspensions and process parameters.

  13. Alginate-Poly(ethylene glycol) Hybrid Microspheres for Primary Cell Microencapsulation

    OpenAIRE

    Redouan Mahou; Meier, Raphael P H; Bühler, Léo H.; Christine Wandrey

    2014-01-01

    The progress of medical therapies, which rely on the transplantation of microencapsulated living cells, depends on the quality of the encapsulating material. Such material has to be biocompatible, and the microencapsulation process must be simple and not harm the cells. Alginate-poly(ethylene glycol) hybrid microspheres (alg-PEG-M) were produced by combining ionotropic gelation of sodium alginate (Na-alg) using calcium ions with covalent crosslinking of vinyl sulfone-terminated multi-arm poly...

  14. Surface modified alginate microcapsules for 3D cell culture

    Science.gov (United States)

    Chen, Yi-Wen; Kuo, Chiung Wen; Chueh, Di-Yen; Chen, Peilin

    2016-06-01

    Culture as three dimensional cell aggregates or spheroids can offer an ideal platform for tissue engineering applications and for pharmaceutical screening. Such 3D culture models, however, may suffer from the problems such as immune response and ineffective and cumbersome culture. This paper describes a simple method for producing microcapsules with alginate cores and a thin shell of poly(L-lysine)-graft-poly(ethylene glycol) (PLL-g-PEG) to encapsulate mouse induced pluripotent stem (miPS) cells, generating a non-fouling surface as an effective immunoisolation barrier. We demonstrated the trapping of the alginate microcapsules in a microwell array for the continuous observation and culture of a large number of encapsulated miPS cells in parallel. miPS cells cultured in the microcapsules survived well and proliferated to form a single cell aggregate. Droplet formation of monodisperse microcapsules with controlled size combined with flow cytometry provided an efficient way to quantitatively analyze the growth of encapsulated cells in a high-throughput manner. The simple and cost-effective coating technique employed to produce the core-shell microcapsules could be used in the emerging field of cell therapy. The microwell array would provide a convenient, user friendly and high-throughput platform for long-term cell culture and monitoring.

  15. Studies on the PEO-PPO-PEO Block Copolymer Release from Alginate Hydrogel

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    @@ Introduction Alginate hydrogel is one of the most widely used carriers for the immobilization of micro bial cells. If surfactants are encapsulated with alginate hydrogel, increasing temperature or concentration can make the encapsulated surfactants aggregate and form micelle.

  16. Mussel-inspired alginate gel promoting the osteogenic differentiation of mesenchymal stem cells and anti-infection.

    Science.gov (United States)

    Zhang, Shiwen; Xu, Kaige; Darabi, Mohammad Ali; Yuan, Quan; Xing, Malcolm

    2016-12-01

    Alginate hydrogels have been used in cell encapsulation for many years but a prevalent issue with pure alginates is that they are unable to provide enough bioactive properties to interact with mammalian cells. This paper discusses the modification of alginate with mussel-inspired dopamine for cell loading and anti-infection. Mouse bone marrow stem cells were immobilized into alginate and alginate-dopamine beads and fibers. Through live-dead and MTT assay, alginates modified by dopamine promoted cell viability and proliferation. In vitro cell differentiation results showed that such an alginate-dopamine gel can promote the osteogenic differentiation of mesenchymal stem cell after PCR and ALP assays. In addition to that, the adhesive prosperities of dopamine allowed for coating the surface of alginate-dopamine gel with silver nanoparticles, which provided the gel with significant antibacterial characteristics. Overall, these results demonstrate that a dopamine-modified alginate gel can be a great tool for cell encapsulation to promote cell proliferation and can be applied to bone regeneration, especially in contaminated bone defects. Copyright © 2016. Published by Elsevier B.V.

  17. Alginate-encapsulation of shoot tips of jojoba [Simmondsia chinensis (Link) Schneider] for germplasm exchange and distribution.

    Science.gov (United States)

    Kumar, Sunil; Rai, Manoj K; Singh, Narender; Mangal, Manisha

    2010-12-01

    Shoot tips excised from in vitro proliferated shoots derived from nodal explants of jojoba [Simmondsia chinensis (Link) Schneider] were encapsulated in calcium alginate beads for germplasm exchange and distribution. A gelling matrix of 3 % sodium alginate and 100 mM calcium chloride was found most suitable for formation of ideal calcium alginate beads. Best response for shoot sprouting from encapsulated shoot tips was recorded on 0.8 % agar-solidified full-strength MS medium. Rooting was induced upon transfer of sprouted shoots to 0.8 % agar-solidified MS medium containing 1 mg l(-1) IBA. About 70 % of encapsulated shoot tips were rooted and converted into plantlets. Plants regenerated from encapsulated shoot tips were acclimatized successfully. The present encapsulation approach could also be applied as an alternative method of propagation of desirable elite genotype of jojoba.

  18. Activity of encapsulated Lactobacillus bulgaricus in alginate-whey protein microspheres

    Directory of Open Access Journals (Sweden)

    Meng-Yan Chen

    2014-10-01

    Full Text Available In this work, alginate-whey protein was used as wall materials for encapsulating Lactobacillus delbrueckii subsp. bulgaricus (L. bulgaricus. The characteristics of encapsulated and free L. bulgaricus showed that the free L. bulgaricus lost viability after 1 min exposure to simulated gastric fluid (SGF at pH 2.0 and 2.5. However, the viability of encapsulated L. bulgaricus did not decrease in SGF at pH 2.5 for 2 h incubation. The viable numbers of encapsulated L. bulgaricus decreased less than 1.0 log unit for 2 h incubation in SGF at pH 2.0. For bile stability, only 1.2 log units and 2.0 log units viability of the encapsulated L. bulgaricus was lost in 1 and 2% bile for 1 h exposure, respectively, compared with no survival of free L. bulgaricus under the same conditions. Encapsulated L. bulgaricus was completely released from the microspheres in simulated intestinal fluid (SIF, pH 6.8 in 3 h. The viability of the encapsulated L. bulgaricus retained more 8.0 log CFU/g after stored at 4°C for four weeks. However, for free L. bulgaricus, only around 3.0 log CFU/mL was found at the same storage conditions. Results showed that the encapsulation could improve the stability of L. bulgaricus.

  19. Alginate-hydroxypropylcellulose hydrogel microbeads for alkaline phosphatase encapsulation

    NARCIS (Netherlands)

    Karewicz, A.; Zasada, K.; Bielska, D.; Douglas, T.E.L.; Jansen, J.A.; Leeuwenburgh, S.C.G.; Nowakowska, M.

    2014-01-01

    There is a growing interest in using proteins as therapeutics agents. Unfortunately, they suffer from limited stability and bioavailability. We aimed to develop a new delivery system for proteins. ALP, a model protein, was successfully encapsulated in the physically cross-linked sodium

  20. Alginate/chitosan nanoparticles for encapsulation and controlled release of vitamin B2.

    Science.gov (United States)

    Azevedo, Maria A; Bourbon, Ana I; Vicente, António A; Cerqueira, Miguel A

    2014-11-01

    This work aims at evaluating encapsulation and controlled release of vitamin B2 from alginate/chitosan nanoparticles. Ionotropic polyelectrolyte pre-gelation was used as production method being chitosan and alginate used as main materials. Nanoparticles were characterized in terms of average size, polydispersity index (PDI), zeta potential and vitamin entrapment efficiency. The average size for alginate/chitosan nanoparticles was 119.5±49.9nm for samples without vitamin B2 and 104.0±67.2nm with the encapsulation of vitamin B2, presenting a PDI of 0.454±0.066 and 0.319±0.068, respectively. The nanoparticles showed encapsulation efficiency and loading capacity values of 55.9±5.6% and 2.2±0.6%, respectively. Release profiles were evaluated at different conditions showing that the polymeric relaxation was the most influent phenomenon in vitamin B2 release. In order to study their stability nanoparticles were stored at 4°C being particles sizes and PDI evaluated during 5 months showing the results that vitamin B2-loaded nanoparticles are more stable (in terms of size and PDI) than nanoparticles without vitamin B2.

  1. Encapsulation of protease from Aspergillus oryzae and lipase from Thermomyces lanuginoseus using alginate and different copolymer types

    Directory of Open Access Journals (Sweden)

    Truong Thi Mong Thu

    2016-05-01

    Full Text Available Although the application of enzymes in food as a food processing aid and enzyme supplement is of interest and widely used, the enzymes can be easily deactivated or lose their activity due to many causes such as pH and moisture as well as through the introduction of incompatible ingredients during food processing and storage. These problems can be solved by the encapsulation technique, especially in a gel matrix. The influences were studied of the alginate concentration, types of copolymer and their concentrations on the bead size, encapsulation yield (EY, encapsulation efficiency (EE, leakage and the retention of enzyme activity during storage period of encapsulated protease from Aspergillus oryzae and lipase from Thermomyces lanuginosus beads. A solution of purified protease or lipase was encapsulated in calcium alginate-chitosan beads (CACB, calcium alginate-xanthan gum beads (CAXB and calcium alginate-maltodextrin beads (CAMB using the extrusion method. Increasing the alginate and copolymer concentrations in the solution increased the bead size, EY, EE and the retention of enzyme activity during the storage period and reduced leakage of both the encapsulated protease and lipase. In addition, different types of copolymer significantly (p ≤ 0.05 affected these properties of both encapsulated enzymes. Furthermore, protease encapsulated using 2.0% alginate and 0.2% chitosan provided the highest EY (81.7% and EE (77.2% with a bead size of 1.85 mm and 8.1% leakage. The retention of encapsulated protease activity and the shelf-life of encapsulated enzyme which was expressed as half-life, the time required for the enzyme activity to decrease by half (thalf life were 75.8% and 27.2 wk, respectively after storage at 4 °C for 10 wk. For lipase, encapsulation using 2.0% alginate and 0.4% xanthan gum provided the highest EY (42.5% and EE (43.9% and the bead size and leakage were 1.81 mm and 6.2%, respectively. The retention of encapsulated

  2. Oil encapsulation in core-shell alginate capsules by inverse gelation. I: dripping methodology.

    Science.gov (United States)

    Martins, Evandro; Renard, Denis; Adiwijaya, Zenia; Karaoglan, Emre; Poncelet, Denis

    2017-02-01

    The production of capsules by inverse gelation consists of adding dropwise oil containing calcium dispersion into an alginate bath. A dripping technique to produce capsules from oil-in-water (O/W) emulsions was proposed by Abang. However, little is known about the oil encapsulation using water-in-oil (W/O) emulsions. This work aims to develop a new method of W/O emulsions encapsulation by inverse gelation. The success of the W/O emulsion encapsulation is due to three factors: 1) use of an emulsion with moderate stability (50 min); 2) production of an emulsion with at least 90 g/L of CaCl2 and 3) addition of ethanol (20% v/v) into the alginate bath. Both wet and dry capsules were obtained with a spherical shape with diameters of 7 and 3.6 mm, respectively. All volume of oil was encapsulated and the oil loading in the wet and dry capsules was of 23 and 68% v/v, respectively.

  3. Influence of mechanical properties of alginate-based substrates on the performance of Schwann cells in culture.

    Science.gov (United States)

    Ning, Liqun; Xu, Yitong; Chen, Xiongbiao; Schreyer, David J

    2016-06-01

    In tissue engineering, artificial tissue scaffolds containing living cells have been studied for tissue repair and regeneration. Notably, the performance of these encapsulated-in-scaffolds cells in terms of cell viability, proliferation, and expression of function during and after the scaffold fabrication process, has not been well documented because of the influence of mechanical, chemical, and physical properties of the scaffold substrate materials. This paper presents our study on the influence of mechanical properties of alginate-based substrates on the performance of Schwann cells, which are the major glial cells of peripheral nervous system. Given the fact that alginate polysaccharide hydrogel has poor cell adhesion properties, in this study, we examined several types of cell-adhesion supplements and found that alginate covalently modified with RGD peptide provided improved cell proliferation and adhesion. We prepared alginate-based substrates for cell culture using varying alginate concentrations for altering their mechanical properties, which were confirmed by compression testing. Then, we examined the viability, proliferation, morphology, and expression of the extracellular matrix protein laminin of Schwann cells that were seeded on the surface of alginate-based substrates (or 2D culture) or encapsulated within alginate-based substrates (3D cultures), and correlated the examined cell performance to the alginate concentration (or mechanical properties) of hydrogel substrates. Our findings suggest that covalent attachment of RGD peptide can improve the success of Schwann cell encapsulation within alginate-based scaffolds, and provide guidance for regulating the mechanical properties of alginate-based scaffolds containing Schwann cells for applications in peripheral nervous system regeneration and repair.

  4. Deterministic encapsulation of single cells in thin tunable microgels for niche modelling and therapeutic delivery

    Science.gov (United States)

    Mao, Angelo S.; Shin, Jae-Won; Utech, Stefanie; Wang, Huanan; Uzun, Oktay; Li, Weiwei; Cooper, Madeline; Hu, Yuebi; Zhang, Liyuan; Weitz, David A.; Mooney, David J.

    2016-10-01

    Existing techniques to encapsulate cells into microscale hydrogels generally yield high polymer-to-cell ratios and lack control over the hydrogel's mechanical properties. Here, we report a microfluidic-based method for encapsulating single cells in an approximately six-micrometre layer of alginate that increases the proportion of cell-containing microgels by a factor of ten, with encapsulation efficiencies over 90%. We show that in vitro cell viability was maintained over a three-day period, that the microgels are mechanically tractable, and that, for microscale cell assemblages of encapsulated marrow stromal cells cultured in microwells, osteogenic differentiation of encapsulated cells depends on gel stiffness and cell density. We also show that intravenous injection of singly encapsulated marrow stromal cells into mice delays clearance kinetics and sustains donor-derived soluble factors in vivo. The encapsulation of single cells in tunable hydrogels should find use in a variety of tissue engineering and regenerative medicine applications.

  5. Effect of Experimental Parameters on Alginate/Chitosan Microparticles for BCG Encapsulation

    Science.gov (United States)

    Caetano, Liliana A.; Almeida, António J.; Gonçalves, Lídia M.D.

    2016-01-01

    The aim of the present study was to develop novel Mycobacterium bovis bacille Calmette-Guérin (BCG)-loaded polymeric microparticles with optimized particle surface characteristics and biocompatibility, so that whole live attenuated bacteria could be further used for pre-exposure vaccination against Mycobacterium tuberculosis by the intranasal route. BCG was encapsulated in chitosan and alginate microparticles through three different polyionic complexation methods by high speed stirring. For comparison purposes, similar formulations were prepared with high shear homogenization and sonication. Additional optimization studies were conducted with polymers of different quality specifications in a wide range of pH values, and with three different cryoprotectors. Particle morphology, size distribution, encapsulation efficiency, surface charge, physicochemical properties and biocompatibility were assessed. Particles exhibited a micrometer size and a spherical morphology. Chitosan addition to BCG shifted the bacilli surface charge from negative zeta potential values to strongly positive ones. Chitosan of low molecular weight produced particle suspensions of lower size distribution and higher stability, allowing efficient BCG encapsulation and biocompatibility. Particle formulation consistency was improved when the availability of functional groups from alginate and chitosan was close to stoichiometric proportion. Thus, the herein described microparticulate system constitutes a promising strategy to deliver BCG vaccine by the intranasal route. PMID:27187418

  6. Alginate Encapsulation of Begonia Microshoots for Short-Term Storage and Distribution

    Directory of Open Access Journals (Sweden)

    Hamidou F. Sakhanokho

    2013-01-01

    Full Text Available Synthetic seeds were formed from shoot tips of two in vitro grown Begonia cultivars using 3% sodium alginate in Murashige and Skoog medium (MS salt solution as the gel matrix and 100 mM calcium chloride for complexation. Synthetic seed formation was achieved by releasing the sodium alginate/explant combination into 100 mM calcium chloride (CaCl2·H2O solution for 30 or 45 min. Both control and encapsulated shoots were transferred into sterile Petri dishes and stored at 4°C or 22°C for 0, 2, 4, 6, or 8 weeks. Conversion of synthetic seeds into plantlets for both storage environments was assessed in MS medium or peat-based substrate. No significant difference was found between the 30 and 45 min CaCl2·H2O treatments or the two cultivars. Encapsulation of explants improved survival rate over time irrespective of the medium type or storage environment. Survival rates of 88, 53, 28, and 11% for encapsulated microshoots versus 73, 13, 0, and 0% for control explants were achieved in microshoots stored for 2, 4, 6, and 8 weeks, respectively. The best results were obtained when synthetic seeds were stored at 4°C and germinated on MS medium. Regenerated plantlets were successfully established in potting soil.

  7. Alginate encapsulation of Begonia microshoots for short-term storage and distribution.

    Science.gov (United States)

    Sakhanokho, Hamidou F; Pounders, Cecil T; Blythe, Eugene K

    2013-01-01

    Synthetic seeds were formed from shoot tips of two in vitro grown Begonia cultivars using 3% sodium alginate in Murashige and Skoog medium (MS) salt solution as the gel matrix and 100 mM calcium chloride for complexation. Synthetic seed formation was achieved by releasing the sodium alginate/explant combination into 100 mM calcium chloride (CaCl₂ ·H₂O) solution for 30 or 45 min. Both control and encapsulated shoots were transferred into sterile Petri dishes and stored at 4°C or 22°C for 0, 2, 4, 6, or 8 weeks. Conversion of synthetic seeds into plantlets for both storage environments was assessed in MS medium or peat-based substrate. No significant difference was found between the 30 and 45 min CaCl₂ ·H₂O treatments or the two cultivars. Encapsulation of explants improved survival rate over time irrespective of the medium type or storage environment. Survival rates of 88, 53, 28, and 11% for encapsulated microshoots versus 73, 13, 0, and 0% for control explants were achieved in microshoots stored for 2, 4, 6, and 8 weeks, respectively. The best results were obtained when synthetic seeds were stored at 4°C and germinated on MS medium. Regenerated plantlets were successfully established in potting soil.

  8. The reinforcement and healing of asphalt mastic mixtures by rejuvenator encapsulation in alginate compartmented fibres

    Science.gov (United States)

    Tabaković, A.; Post, W.; Cantero, D.; Copuroglu, O.; Garcia, S. J.; Schlangen, E.

    2016-08-01

    This paper explores the potential use of compartmented alginate fibres as a new method of incorporating rejuvenators into asphalt pavement mixtures. The compartmented fibres are employed to locally distribute the rejuvenator and to overcome the problems associated with spherical capsules and hollow fibres. The work presents proof of concept of the encapsulation process which involved embedding the fibres into the asphalt mastic mixture and the survival rate of fibres in the asphalt mixture. To prove the effectiveness of the alginate as a rejuvenator encapsulating material and to demonstrate its ability survive asphalt production process, the fibres containing the rejuvenator were prepared and subjected to thermogravimetric analysis and uniaxial tensile test. The test results demonstrated that fibres have suitable thermal and mechanical strength to survive the asphalt mixing and compaction process. The CT scan of an asphalt mortar mix containing fibres demonstrated that fibres are present in the mix in their full length, undamaged, providing confirmation that the fibres survived the asphalt production process. In order to investigate the fibres physiological properties and ability to release the rejuvenator into cracks in the asphalt mastic, the environmental scanning electron microscope and optical microscope analysis were employed. To prove its success as an asphalt healing system, compartmented alginate fibres containing rejuvenator were embedded in asphalt mastic mix. The three point bend tests were performed on the asphalt mastic test samples and the degree to which the samples began to self-heal in response was measured and quantified. The research findings indicate that alginate fibres present a promising new approach for the development of self-healing asphalt pavement systems.

  9. The in vitro and in vivo capacity of culture-expanded human cells from several sources encapsulated in alginate to form cartilage

    NARCIS (Netherlands)

    M.M. Pleumeekers (Mieke); L. Nimeskern (Luc); J.L.M. Koevoet (Wendy); N. Kops (Nicole); R.M.L. Poublon (René); K.S. Stok (Kathryn); G.J.V.M. van Osch (Gerjo)

    2014-01-01

    textabstractAbstract Cartilage has limited self-regenerative capacity. Tissue engineering can offer promising solutions for reconstruction of missing or damaged cartilage. A major challenge herein is to define an appropriate cell source that is capable of generating a stable and functional matrix. T

  10. Alginate encapsulation supports the growth and differentiation of human primordial follicles within ovarian cortical tissue.

    Science.gov (United States)

    Laronda, Monica M; Duncan, Francesca E; Hornick, Jessica E; Xu, Min; Pahnke, Jennifer E; Whelan, Kelly A; Shea, Lonnie D; Woodruff, Teresa K

    2014-08-01

    In vitro follicle growth (IVFG) is an investigational fertility preservation technique in which immature follicles are grown in culture to produce mature eggs that can ultimately be fertilized. Although progress has been made in growing primate primary and secondary follicles in vitro, it has been a relatively greater challenge to isolate and culture primordial follicles. The purpose of this study was to develop methods to grow human primordial follicles in vitro using alginate hydrogels. We obtained human ovarian tissue for research purposes through the National Physicians Cooperative from nationwide sites and used it to test two methods for culturing primordial follicles. First, primordial follicles were isolated from the ovarian cortex and encapsulated in alginate hydrogels. Second, 1 mm × 1 mm pieces of 500 μm-thick human ovarian cortex containing primordial follicles were encapsulated in alginate hydrogels, and survival and follicle development within the tissue was assessed for up to 6 weeks. We found that human ovarian tissue could be kept at 4 °C for up to 24 h while still maintaining follicle viability. Primordial follicles isolated from ovarian tissue did not survive culture. However, encapsulation and culture of ovarian cortical pieces supported the survival, differentiation, and growth of primordial and primary follicles. Within several weeks of culture, many of the ovarian tissue pieces had formed a defined surface epithelium and contained growing preantral and antral follicles. The early stages of in vitro human follicle development require the support of the native ovarian cortex.

  11. Evaluation of alginate hydrogel cytotoxicity on three-dimensional culture of type A spermatogonial stem cells.

    Science.gov (United States)

    Jalayeri, Maryam; Pirnia, Afshin; Najafabad, Elaheh Poorazizi; Varzi, Ali Mohammad; Gholami, Mohammadreza

    2017-02-01

    The culture of spermatogonial cells for future transplantation, based on the specific biology of these cells is important and necessary. Recently, the use of scaffolds especially alginate for culturing stem cells has been the focus of many researchers. The aim of this study was to evaluate the cytotoxicity of alginate hydrogels to cultures of type A spermatogonial stem cells. Spermatogonial stem cells of 6day-old immature mice were isolated by surgery; thereafter, the cells were purified by MACS using antibodies against thy-1 and C-kit and cultured on a layer of laminin. After purification, spermatogonial stem cells were encapsulated in alginate hydrogels. After one month of encapsulation and culture in DMEM culture medium containing 10ng/ml GDNF, cells were removed from hydrogel and were examined for viability, cell morphology and structure, cytotoxicity and expression of apoptosis genes Fas, P53, Bax, Bcl2, Caspase3 by staining with trypan blue, scanning electron microscopy, LDH test, and Real time PCR, respectively. The encapsulation did not change the morphology and viability of spermatogonial stem cells. Investigations showed that spermatogonial stem cells preserve by the high viability (74.08%) and cytotoxicity of alginate hydrogel was estimated to be 5%. Expression of Fas gene increased in main group compared with the control group, and expression of Bax and P53 was reduced in main group compared with the control group. Expression of Bcl2 and Caspase3 genes did not show any significant difference between the main group and the control group. Considering the lack of cytotoxicity and antioxidant properties of alginate hydrogel scaffold and high viability of cells, this three-dimensional scaffold is applicable for culturing and encapsulation of spermatogonial stem cells. Copyright © 2016 Elsevier B.V. All rights reserved.

  12. Long-term graft function of cryostored alginate encapsulated rat islets

    Directory of Open Access Journals (Sweden)

    Schneider Stephan

    2011-09-01

    Full Text Available Abstract Microencapsulation of pancreatic islets before transplantation is a promising approach to enable graft function in an immunocompetent recipient without immunosuppression. However, the insufficient availability of allogenic islet tissue is a major problem. One concept to overcome these shortcomings is the cryopreservation of encapsulated allogenic islets. Recently, we reported a gentle cryopreservation protocol for rat islets encapsulated in an alginate-based microcapsule system. Here, we report for the first time long-term transplantation data of these cryopreserved microencapsulated islets. We detected a stable graft function for more than 12 month (experiments still continuing after transplantation of 2500 cryopreserved microencapsulated CD rat islets in streptozotocin-diabetic Wistar rats. Moreover, the glucose clearance rate during an IPGTT was well preserved up to 56 weeks after transplantation. In addition, hyperglycemic blood glucose levels after removal of rat islet grafts 12 and 56 weeks after transplantation confirmed the efficacy of the encapsulated islets. Finally, the retrieved encapsulated rat islets responded well with a 7-fold increase of insulin secretion to a glucose stimulus (12 and 56 weeks. In conclusion, our study demonstrates for the first time that cryopreservation of encapsulated rat islets is possible without substantial losses on graft function for a very long time.

  13. Effects of process variables on the encapsulation of oil in ca-alginate capsules using an inverse gelation technique.

    Science.gov (United States)

    Abang, Sariah; Chan, Eng-Seng; Poncelet, Denis

    2012-01-01

    The objective of this study was to investigate the effects of process variables on the encapsulation of oil in a calcium alginate membrane using an inverse gelation technique. A dispersion of calcium chloride solution in sunflower oil (water-in-oil emulsion) was added dropwise to the alginate solution. The migration of calcium ions to the alginate solution initiates the formation of a ca-alginate membrane around the emulsion droplets. The membrane thickness of wet capsules and the elastic modulus of dry capsules increased following first-order kinetics with an increasing curing time. An increase in the calcium chloride concentration increased the membrane thickness of wet capsules and the elastic modulus of dry capsules. An increase in the alginate concentration decreased the mean diameter of wet capsules but increased the elastic modulus of dry capsules.

  14. Hydrogels of sodium alginate in cationic surfactants: Surfactant dependent modulation of encapsulation/release toward Ibuprofen.

    Science.gov (United States)

    Jabeen, Suraya; Chat, Oyais Ahmad; Maswal, Masrat; Ashraf, Uzma; Rather, Ghulam Mohammad; Dar, Aijaz Ahmad

    2015-11-20

    The interaction of cetyltrimethylammoium bromide (CTAB) and its gemini homologue (butanediyl-1,4-bis (dimethylcetylammonium bromide), 16-4-16 with biocompatible polymer sodium alginate (SA) has been investigated in aqueous medium. Addition of K2CO3 influences viscoelastic properties of surfactant impregnated SA via competition between electrostatic and hydrophobic interactions. Viscosity of these polymer-surfactant systems increases with increase in concentration of K2CO3, and a cryogel is formed at about 0.5M K2CO3 concentration. The thermal stability of gel (5% SA+0.5M K2CO3) decreases with increase in surfactant concentration, a minimum is observed with increase in 16-4-16 concentration. The impact of surfactant addition on the alginate structure vis-à-vis its drug loading capability and release thereof was studied using Ibuprofen (IBU) as the model drug. The hydrogel with 16-4-16 exhibits higher IBU encapsulation and faster release in comparison to the one containing CTAB. This higher encapsulation-cum-faster release capability has been related to micelle mediated solubilization and greater porosity of the hydrogel with gemini surfactant.

  15. Effects of supplementation with L. plantarum TN8 encapsulated in alginate-chitosan in broiler chickens.

    Science.gov (United States)

    Trabelsi, Imen; Ktari, Naourez; Ben Slima, Sirine; Bouchaala, Kamel; Ben Salah, Riadh

    2016-08-01

    This study was undertaken to investigate the effects of supplementation of probiotic strain Lactobacillus plantarum TN8 encapsulated in sodium alginate-chitosan or a commercial blend of essential oils on total cholesterol, High Density Lipoprotein (HDL), Low Density Lipoprotein (LDL) and growth performance of broiler chickens. The results showed that the broiler chickens supplemented with encapsulated L. plantarum TN8 or essential oil has a higher growth than the control group. After 35days, the weight means were 1860 and 1880g respectively in dietary supplementation with probiotic or essential oil, while they are 1800g in the control group. The evolution of the feed consumption and feed conversion per week showed that the supplementation of encapsulated TN8 strain or essential oil in broiler chickens food has a positive influence on their appetite. Similarly, supplementation of the feed with this encapsulated strain significantly reduced the rate of cholesterol (HDL and LDL) as well as the contents of triglycerides in broiler chickens. Through our study, it appears that the use of the probiotic supplementation or essential oil to broilers were found to be better than the control group of chickens, resulting in a significant economic impact and promoting effect on health. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. Comparison of alginate and pectin based beads for production of poultry probiotic cells.

    Science.gov (United States)

    Voo, Wan-Ping; Ravindra, Pogaku; Tey, Beng-Ti; Chan, Eng-Seng

    2011-03-01

    A comparative study on the stability and potential of alginate and pectin based beads for production of poultry probiotic cells using MRS medium in repeated batch fermentation was conducted. The bead cores, made of three types of materials, i.e., ca-alginate, ca-pectinate and ca-alginate/pectinate, were compared. The effect of single and double layer coatings using chitosan and core material, respectively, on the bead stability and cell production were also studied. The pectin based beads were found to be more stable than that of the alginate beads and their stability was further improved by coating with chitosan. The cell concentration in pectin based beads was comparable to that in the alginate beads. On the other hand, pectin based beads gave significantly lower cell concentration in the growth medium for the initial fermentation cycles when compared to the alginate beads. In conclusion, pectin was found to be potential encapsulation material for probiotic cell production owing to its stability and favourable microenvironment for cell growth. Copyright © 2010 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

  17. Physical properties and heavy metal uptake of encapsulated Escherichia coli expressing a metal binding gene (NCP).

    Science.gov (United States)

    Bang, S S; Pazirandeh, M

    1999-01-01

    A recombinant Escherichia coli expressing the Neurospora crassa metallothionein gene (NCP) has previously been shown to remove low levels of Cd and other metals from solution. For further development as a biosorbent, the encapsulation of the NCP is investigated by various matrices. The NCP was encapsulated in alginate, chitosan-alginate or kappa-carrageenan, and its physical properties characterized. Results indicated that encapsulation in alginate resulted in fragile beads, whereas encapsulation in kappa-carrageenan or chitosan-alginate provided more physical and chemical integrity to the beads. Maximal heavy metal removal by cells encapsulated in carrageenan occurred within 3 h, while a gradual increase in removal was observed up to 24 h for cells encapsulated in chitosan-alginate. Metal removal by cells encapsulated in alginate beads was lower than those encapsulated in carrageenan or chitosan-alginate.

  18. A novel hybrid material: an inorganic silica aerogel core encapsulated with a tunable organic alginate aerogel layer

    OpenAIRE

    Ülker, Zeynep; Erkey, Can

    2014-01-01

    A novel layered material consisting of a silica aerogel core encapsulated by an alginate aerogel layer was developed. The components of the hybrid aerogel had the high surface area and high porosity of pure aerogels which should lead to development of new layered systems for a wide variety of applications.

  19. Effect of psyllium and gum Arabic biopolymers on the survival rate and storage stability in yogurt of Enterococcus duransIW3 encapsulated in alginate.

    Science.gov (United States)

    Nami, Yousef; Haghshenas, Babak; Yari Khosroushahi, Ahmad

    2017-05-01

    Different herbal biopolymers were used to encapsulate Enterococcus durans IW3 to enhance its storage stability in yogurt and subsequently its endurance in gastrointestinal condition. Nine formulations of encapsulation were performed using alginate (ALG), ALG-psyllium (PSY), and ALG-gum Arabic (GA) blends. The encapsulation efficiency of all formulations, tolerance of encapsulated E. durans IW3 against low pH/high bile salt concentration, storage lifetime, and release profile of cells in natural condition of yogurt were evaluated. Result revealed 98.6% encapsulation efficiency and 76% survival rate for all formulation compared with the unencapsulated formulation cells (43%). The ALG-PSY and ALG-GA formulations have slightly higher survival rates at low pH and bile salt condition (i.e., 76-93% and 81-95%, respectively) compared with the ALG formulation. All encapsulated E. durans IW3 was released from the prepared beads of ALG after 90 min, whereas both probiotics encapsulated in ALG-GA and ALG-PSY were released after 60 min. Enterococcus durans IW3 was successfully encapsulated in ALG, ALG-GA, and ALG-PSY beads prepared by extrusion method. ALG-GA and ALG-PSY beads are suitable delivery carriers for the oral administration of bioactive compounds like probiotics. The GA and PSY gels exhibited better potential for encapsulation of probiotic bacteria cells because of the amendment of ALG difficulties and utilization of therapeutic and prebiotic potentials of these herbal biopolymers.

  20. Technology of mammalian cell encapsulation

    NARCIS (Netherlands)

    Uludag, H; De Vos, P; Tresco, PA

    2000-01-01

    Entrapment of mammalian cells in physical membranes has been practiced since the early 1950s when it was originally introduced as a basic research tool. The method has since been developed based on the promise of its therapeutic usefulness in tissue transplantation. Encapsulation physically isolates

  1. Polymers in cell encapsulation from an enveloped cell perspective.

    Science.gov (United States)

    de Vos, Paul; Lazarjani, Hamideh Aghajani; Poncelet, Denis; Faas, Marijke M

    2014-04-01

    In the past two decades, many polymers have been proposed for producing immunoprotective capsules. Examples include the natural polymers alginate, agarose, chitosan, cellulose, collagen, and xanthan and synthetic polymers poly(ethylene glycol), polyvinyl alcohol, polyurethane, poly(ether-sulfone), polypropylene, sodium polystyrene sulfate, and polyacrylate poly(acrylonitrile-sodium methallylsulfonate). The biocompatibility of these polymers is discussed in terms of tissue responses in both the host and matrix to accommodate the functional survival of the cells. Cells should grow and function in the polymer network as adequately as in their natural environment. This is critical when therapeutic cells from scarce cadaveric donors are considered, such as pancreatic islets. Additionally, the cell mass in capsules is discussed from the perspective of emerging new insights into the release of so-called danger-associated molecular pattern molecules by clumps of necrotic therapeutic cells. We conclude that despite two decades of intensive research, drawing conclusions about which polymer is most adequate for clinical application is still difficult. This is because of the lack of documentation on critical information, such as the composition of the polymer, the presence or absence of confounding factors that induce immune responses, toxicity to enveloped cells, and the permeability of the polymer network. Only alginate has been studied extensively and currently qualifies for application. This review also discusses critical issues that are not directly related to polymers and are not discussed in the other reviews in this issue, such as the functional performance of encapsulated cells in vivo. Physiological endocrine responses may indeed not be expected because of the many barriers that the metabolites encounter when traveling from the blood stream to the enveloped cells and back to circulation. However, despite these diffusion barriers, many studies have shown optimal

  2. The efficacy of oral vaccination of mice with alginate encapsulated outer membrane proteins of Pasteurella haemolytica and One-Shot.

    Science.gov (United States)

    Kidane, A; Guimond, P; Ju, T R; Sanchez, M; Gibson, J; Bowersock, T L

    2001-03-21

    The goal of this study was to examine the efficacy of oral delivery of alginate encapsulated outer membrane proteins (OMP) of Pasteurella haemolytica and a commercial One-Shot vaccine in inducing protection in mice against lethal challenge with virulent P. haemolytica. We examined two alginate microsphere formulations and compared them with oral unencapsulated and subcutaneously administered vaccines. Alginate microspheres were made by the emulsion-cross-linking technique. They were examined for size, hydrophobicity, and antigen loading efficiency before they were used in the study. Mice were vaccinated by administering 200 microg of antigens in 200 microl of microspheres suspension orally or subcutaneously. One group of mice received blank microspheres and a second group was given unencapsulated antigen orally. A third and a fourth group received different formulations of alginate encapsulated antigens by oral administration. Three groups received subcutaneous inoculations (alginate encapsulated, non-adjuvanted and unencapsulated antigens, and adjuvanted One-Shot), and one group received water (naïve group). Mice were vaccinated orally for four consecutive days and challenged with P. haemolytica 5 weeks after the first vaccination. Weekly serum and feces samples were assayed for antigen specific antibodies. The number of dead mice in each group 4 days post challenge was used to compare the efficacy of the various vaccination groups. The mean volume sizes of blank alginate microsphere formulations A, and AA were 15.9, 16 and 9.2 microm, respectively. Hydrophobicity of the microspheres was evaluated by measuring contact angle on a glass slide coated with the microspheres. The contact angles on A and AA were 37.8 and 74.3 degrees, respectively. Antigen concentration in a 1:1 w/w suspension of microspheres in water was 0.9 mg/ml. Rate of death for the blank group was 42.8% whereas for groups vaccinated with antigens encapsulated in A and AA the death rates were 40

  3. Encapsulation Red Ginger Oleoresin (Zingiber officinale var. Rubrum With Chitosan-alginate as Wall Material Using Spray Drying

    Directory of Open Access Journals (Sweden)

    Jayanudin

    2015-08-01

    Full Text Available Encapsulation is the process of protecting the active ingredients that is susceptible to environmental influences by using a coating. Red ginger oleoresin contains bioactive components that can be used as natural antioxidants, but sensitive to environmental influences. Chitosan-alginate nanoparticle is used as the coating, because it is safe for consumption and also stable. The purpose of this research was to determine the effect of the Tripolyphosphate (TPP concentration against the emulsion droplet size and determine the encapsulation efficiency of red ginger oleoresin. Encapsulation method was done by mixing 2% chitosan solution with 1% acetic acid, 1% sodium alginate and 8 g of red ginger oleoresin. The mixture was stirred and added sodium tripolyphosphate (3.5, 4.5 and 5.5%, respectively, emulsion preparation process was done by adding tween 80 (3, 4 and 5%, respectively and then stirred using homogenizer with a speed of 22,000 rpm, emulsions formed were analyzed using nano-particle analyzer. Emulsion formed was flowed on to the spray dryer inlet temperature of 180°C to form a powder encapsulation. Powder products were analyzed for determining encapsulation efficiency and morphology of red ginger oleoresin powder using Scanning Electron Microscopy (SEM. Based on the research results, the smallest droplet size of the emulsion was obtained at 481.5 nm and the largest encapsulation efficiency was as high as 70.59%.

  4. Encapsulation Red Ginger Oleoresin (Zingiber officinale) var. Rubrum) With Chitosan-alginate as Wall Material Using Spray Drying

    OpenAIRE

    Jayanudin; Rochmadi,; Wiratni; Meri Yulvianti; Dhena Ria Barleany; Widya Ernayati

    2015-01-01

    Encapsulation is the process of protecting the active ingredients that is susceptible to environmental influences by using a coating. Red ginger oleoresin contains bioactive components that can be used as natural antioxidants, but sensitive to environmental influences. Chitosan-alginate nanoparticle is used as the coating, because it is safe for consumption and also stable. The purpose of this research was to determine the effect of the Tripolyphosphate (TPP) concentration against the emulsio...

  5. Factors influencing alginate gel biocompatibility.

    Science.gov (United States)

    Tam, Susan K; Dusseault, Julie; Bilodeau, Stéphanie; Langlois, Geneviève; Hallé, Jean-Pierre; Yahia, L'Hocine

    2011-07-01

    Alginate remains the most popular polymer used for cell encapsulation, yet its biocompatibility is inconsistent. Two commercially available alginates were compared, one with 71% guluronate (HiG), and the other with 44% (IntG). Both alginates were purified, and their purities were verified. After 2 days in the peritoneal cavity of C57BL/6J mice, barium (Ba)-gel and calcium (Ca)-gel beads of IntG alginate were clean, while host cells were adhered to beads of HiG alginate. IntG gel beads, however, showed fragmentation in vivo while HiG gel beads stayed firm. The physicochemical properties of the sodium alginates and their gels were thoroughly characterized. The intrinsic viscosity of IntG alginate was 2.5-fold higher than that of HiG alginate, suggesting a greater molecular mass. X-ray photoelectron spectroscopy indicated that both alginates were similar in elemental composition, including low levels of counterions in all gels. The wettabilities of the alginates and gels were also identical, as measured by contact angles of water on dry films. Ba-gel beads of HiG alginate resisted swelling and degradation when immersed in water, much more than the other gel beads. These results suggest that the main factors contributing to the biocompatibility of gels of purified alginate are the mannuronate/guluronate content and/or intrinsic viscosity.

  6. Polymeric microcapsules poduction from sodium alginic acid for cell therapy

    Directory of Open Access Journals (Sweden)

    Ana Carolina Vale Campos Lisboa

    2007-12-01

    Full Text Available Development of polymeric materials has been increasingly emphasized in Biomedicine. Here, we evaluate the use of microcapsules made of Biodritin®, a biocompatible polymer compound which contains sodium alginic acid, a natural polymer extracted from algae, and Cis-Chondroitin sulfate, a glycosaminoglycan from the extracellular matrix. Gelation of this polymer into microcapsules is achieved by dropping the compound into BaCl2 or CaCl2 gelling solutions. A functional microcapsule is dependent on its permeability, mechanical stability, immunoisolation capacity and biocompatibility. The mechanical stability of Biodritin-barium and Biodritin-calcium microcapsules was investigated after rotational stress upon in vitro culture and in vivo implantation. Viability studies of encapsulated cells were also performed to assess other functional parameters of the microcapsules. When subject to rotational stress, Biodritin-barium microcapsules exhibited breaks, whereas the Biodritin-calcium microcapsules did not. Both kinds of Biodritin® microcapsules proved to be mechanically resistant in in vitro and in vivo studies. However, the Biodritin-calcium material was found to be more elastic while the Biodritin-barium microcapsules displayed a more plastic behavior. These properties seem to be determinant for viability of the encapsulated cell’s, since the Biodritin-calcium microcapsules presented more viable cells than the Biodritin-barium microcapsules.

  7. Pancreatic cell immobilization in alginate beads produced by emulsion and internal gelation.

    Science.gov (United States)

    Hoesli, Corinne A; Raghuram, Kamini; Kiang, Roger L J; Mocinecová, Dušana; Hu, Xiaoke; Johnson, James D; Lacík, Igor; Kieffer, Timothy J; Piret, James M

    2011-02-01

    Alginate has been used to protect transplanted pancreatic islets from immune rejection and as a matrix to increase the insulin content of islet progenitor cells. The throughput of alginate bead generation by the standard extrusion and external gelation method is limited by the rate of droplet formation from nozzles. Alginate bead generation by emulsion and internal gelation is a scaleable alternative that has been used with biological molecules and microbial cells, but not mammalian cells. We describe the novel adaptation of this process to mammalian cell immobilization. After optimization, the emulsion process yielded 90 ± 2% mouse insulinoma 6 (MIN6) cell survival, similar to the extrusion process. The MIN6 cells expanded at the same rate in both bead types to form pseudo-islets with increased glucose stimulation index compared to cells in suspension. The emulsion process was suitable for primary pancreatic exocrine cell immobilization, leading to 67 ± 32 fold increased insulin expression after 10 days of immobilized culture. Due to the scaleability and broad availability of stirred mixers, the emulsion process represents an attractive option for laboratories that are not equipped with extrusion-based cell encapsulators, as well as for the production of immobilized or encapsulated cellular therapeutics on a clinical scale.

  8. Human adipose-derived stromal cells in a clinically applicable injectable alginate hydrogel

    DEFF Research Database (Denmark)

    Larsen, Bjarke Follin; Juhl, Morten; Cohen, Smadar

    2015-01-01

    . Hepatocyte growth factor mRNA was increased in ASCs cultivated in alginates compared with monolayer controls. Alginates and alginates containing ASCs did not induce dendritic cell maturation. ASCs in alginate responded like controls to interferon-gamma stimulation (licensing), and alginate culture increased...... is to inject the cells in an in situ cross-linked alginate hydrogel. METHODS: ASCs from abdominal human tissue were embedded in alginate hydrogel and alginate hydrogel modified with Arg-Gly-Asp motifs (RGD-alginate) and cultured for 1 week. Cell viability, phenotype, immunogenicity and paracrine activity were...... determined by confocal microscopy, dendritic cell co-culture, flow cytometry, reverse transcriptase quantitative polymerase chain reaction, Luminex multiplex, and lymphocyte proliferation experiments. RESULTS: ASCs performed equally well in alginate and RGD-alginate. After 1 week of alginate culture, cell...

  9. A novel multilayer immunoisolating encapsulation system overcoming protrusion of cells

    NARCIS (Netherlands)

    Bhujbal, Swapnil V.; de Haan, Bart; Niclou, Simone P.; de Vos, Paul

    2014-01-01

    Application of alginate-microencapsulated therapeutic cells is a promising approach for diseases that require a local and constant supply of therapeutic molecules. However most conventional alginate microencapsulation systems are associated with low mechanical stability and protrusion of cells which

  10. Promoting extracellular matrix remodeling via ascorbic acid enhances the survival of primary ovarian follicles encapsulated in alginate hydrogels.

    Science.gov (United States)

    Tagler, David; Makanji, Yogeshwar; Tu, Tao; Bernabé, Beatriz Peñalver; Lee, Raymond; Zhu, Jie; Kniazeva, Ekaterina; Hornick, Jessica E; Woodruff, Teresa K; Shea, Lonnie D

    2014-07-01

    The in vitro growth of ovarian follicles is an emerging technology for fertility preservation. Various strategies support the culture of secondary and multilayer follicles from various species including mice, non-human primate, and human; however, the culture of early stage (primary and primordial) follicles, which are more abundant in the ovary and survive cryopreservation, has been limited. Hydrogel-encapsulating follicle culture systems that employed feeder cells, such as mouse embryonic fibroblasts (MEFs), stimulated the growth of primary follicles (70-80 µm); yet, survival was low and smaller follicles (structure and degenerated. These morphologic changes were associated with a breakdown of the follicular basement membrane; hence, this study investigated ascorbic acid based on its role in extracellular matrix (ECM) deposition/remodeling for other applications. The selection of ascorbic acid was further supported by a microarray analysis that suggested a decrease in mRNA levels of enzymes within the ascorbate pathway between primordial, primary, and secondary follicles. The supplementation of ascorbic acid (50 µg/mL) significantly enhanced the survival of primary follicles (alginate hydrogels, which coincided with improved structural integrity. Follicles developed antral cavities and increased to diameters exceeding 250 µm. Consistent with improved structural integrity, the gene/protein expression of ECM and cell adhesion molecules was significantly changed. This research supports the notion that modifying the culture environment (medium components) can substantially enhance the survival and growth of early stage follicles. © 2013 Wiley Periodicals, Inc.

  11. Stability Comparison of Free and Encapsulated Lactobacilus casei ATCC 393 in Yoghurt for Long Time Storage

    OpenAIRE

    Oana Lelia POP; Vodnar, Dan Cristian; Ramona SUHAROSCHI; Socaciu, Carmen

    2016-01-01

    An innovative method of L. casei ATCC 393 encapsulation has been reported in the present study using pectin combined with alginate. The aim of this study was to investigate the effect of encapsulation on the survival of L. casei ATCC 393 in yoghurt during long time storage, free or encapsulated in alginate and alginate pectin microspheres, and influence over yoghurt properties, particularly acidification. Over 35 days of storage in yoghurt, the encapsulated probiotic cells proved a higher via...

  12. Propagation of human iPS cells in alginate-based microcapsules prepared using reactions catalyzed by horseradish peroxidase and catalase.

    Science.gov (United States)

    Ashida, Tomoaki; Sakai, Shinji; Taya, Masahito

    2016-09-01

    Cell encapsulation has been investigated as a bioproduction system in the biomedical and pharmaceutical fields. We encaps-ulated human induced pluripotent stem (hiPS) cells in duplex microcapsules prepared from an alginate derivative possessing phenolic hydroxyl moieties, in a single-step procedure based on two competing enzymatic reactions catalyzed by horseradish peroxidase (HRP) and catalase. The encapsulated cells maintained 91.4% viability and proliferated to fill the microcapsules following 19 days of culture. Encapsulated hiPS cells showed pluripotency comparable to that of unencapsulated cells during the cultures, as demonstrated by the expression of the SSEA-4 marker.

  13. Adsorption of ochratoxin A from grape juice by yeast cells immobilised in calcium alginate beads.

    Science.gov (United States)

    Farbo, Maria Grazia; Urgeghe, Pietro Paolo; Fiori, Stefano; Marceddu, Salvatore; Jaoua, Samir; Migheli, Quirico

    2016-01-18

    Grape juice can be easily contaminated with ochratoxin A (OTA), one of the known mycotoxins with the greatest public health significance. Among the different approaches to decontaminate juice from this mycotoxin, microbiological methods proved efficient, inexpensive and safe, particularly the use of yeast or yeast products. To ascertain whether immobilisation of the yeast biomass would lead to successful decontamination, alginate beads encapsulating Candida intermedia yeast cells were used in our experiments to evaluate their OTA-biosorption efficacy. Magnetic calcium alginate beads were also prepared by adding magnetite in the formulation to allow fast removal from the aqueous solution with a magnet. Calcium alginate beads were added to commercial grape juice spiked with 20 μg/kg OTA and after 48 h of incubation a significant reduction (>80%), of the total OTA content was achieved, while in the subsequent phases (72-120 h) OTA was slowly released into the grape juice by alginate beads. Biosorption properties of alginate-yeast beads were tested in a prototype bioreactor consisting in a glass chromatography column packed with beads, where juice amended with OTA was slowly flowed downstream. The adoption of an interconnected scaled-up bioreactor as an efficient and safe tool to remove traces of OTA from liquid matrices is discussed.

  14. Differentiation of embryoid-body cells derived from embryonic stem cells into hepatocytes in alginate microbeads in vitro

    Institute of Scientific and Technical Information of China (English)

    Sheng FANG; Yu-dong QIU; Liang MAO; Xiao-lei SHI; De-cai YU; Yi-tao DING

    2007-01-01

    Aim: Embryonic stem (ES) cells are being widely investigated as a promising source of hepatocytes with their proliferative, renewable, and pluripotent capacities. However, controlled and scalable ES cell differentiation culture into functional hepatocytes is challenging. In this study, we examined the differentiat- ing potential of embryoid-body cells derived from ES cells into hepatocytes in alginate microbeads containing exogenous growth factors in vitro. Methods: Embryoid bodies were formed from ES cells by suspension methods. Embryoid bodies cultured for 5 d were treated with trypsin-EDTA. The disaggregated cells were encapsulated in alginate microbeads and stimulated with exogenous growth factors to induce hepatic differentiation. In the course of cell differentiation, cell morphology and viability were observed, and the expression patterns of some genes of the hepatocyte were confirmed by RT-PCR. An immunofluorescence analysis revealed the expression of albumin (ALB) and cytokeratin-18 (CK18). Hepatocyte functional assays were confirmed by the secretion of ALB and urea. Results: We showed that embryoid-body cells could maintain cell viability in alginate microbeads in vitro. We also found that directed differentiated cells expressed several hepatocyte genes including α-fetoprotein (AFP), ALB, Cyp7al, CK18, transthyretin (TTR) and tyrosine aminotransferase (TAT) and produced ALB and urea in alginate microbeads. The directed differentiated cells expressed ALB and CK18 proteins on d 14. However, embryoid-body cells could not form hepatocytes without exogenous growth factors in alginate microbeads. Conclusion: The differentiation of embryoid-body cells into hepatocytes con- taining exogenous growth factors in alginate microbeads gives rise to functional hepatocytes and may develop scalable stem cell differentiation strategies for bioartificial livers and hepatocyte transplantation.

  15. The cell wall compound of Saccharomyces cerevisiae as a novel wall material for encapsulation of probiotics.

    Science.gov (United States)

    Mokhtari, Samira; Jafari, Seid Mahdi; Khomeiri, Morteza; Maghsoudlou, Yahya; Ghorbani, Mohammad

    2017-06-01

    Yeast cell wall is known as a food grade ingredient which is recently being used increasingly as a novel coating for encapsulation of different materials in the food industry. This application is limited to core materials smaller than yeast in size. In this study, we have tried to encapsulate larger particles by crushing yeast cells. Hence, probiotic bacteria of Lactobacillus acidophilus and Bifidobacterium bifidum were encapsulated firstly by calcium alginate using the emulsion method and these microbeads were coated again by Saccharomyces cerevisiae cell wall compound and another layer of calcium alginate. The average diameter of microcapsules for single layer microbeads (M), microbeads coated by two layers of alginate (MCA), and microbeads coated by a layer of yeast cell and two layers of alginate (MCYA) were 54.25±0.18, 77.43±8.24 and 103.66±13.33μm, respectively. In simulated gastrointestinal conditions, there was a significant (Pprobiotics and it can improve the survival of probiotics within food products. Copyright © 2017 Elsevier Ltd. All rights reserved.

  16. Growth and by-product profiles of Kluyveromyces marxianus cells immobilized in foamed alginate.

    Science.gov (United States)

    Wilkowska, Agnieszka; Kregiel, Dorota; Guneser, Onur; Karagul Yuceer, Yonca

    2015-01-01

    The aim of this research was to study how the yeast cell immobilization technique influences the growth and fermentation profiles of Kluyveromyces marxianus cultivated on apple/chokeberry and apple/cranberry pomaces. Encapsulation of the cells was performed by droplet formation from a foamed alginate solution. The growth and metabolic profiles were evaluated for both free and immobilized cells. Culture media with fruit waste produced good growth of free as well as immobilized yeast cells. The fermentation profiles of K. marxianus were different with each waste material. The most varied aroma profiles were noted for immobilized yeast cultivated on apple/chokeberry pomace.

  17. Development of sheep primordial follicles encapsulated in alginate or in ovarian tissue in fresh and vitrified samples.

    Science.gov (United States)

    Sadeghnia, Samaneh; Akhondi, Mohammad Mehdi; Hossein, Ghamartaj; Mobini, Sahba; Hosseini, Laleh; Naderi, Mohammad Mehdi; Boroujeni, Sara Borjian; Sarvari, Ali; Behzadi, Bahareh; Shirazi, Abolfazl

    2016-04-01

    In vitro follicle growth is a promising strategy for female fertility preservation. This study was conducted to compare the development of ovine follicles either isolated or in the context of ovarian cortical pieces after short term (8 days) three-dimensional culture in fresh and vitrified samples. Four different experiments were conducted; I) culture of ovarian cortical pieces encapsulated in 0.5% and 1% alginate and without alginate encapsulation (CP-0.5%, CP-1% and CP, respectively), II) culture of isolated primordial and primary follicles encapsulated in 1% and 2% alginate (IF-1% and IF-2%, respectively), III) culture of fresh and vitrified-warmed cortical pieces (F-CP and Vit-CP, respectively), and IV) culture of fresh and vitrified-warmed encapsulated isolated follicles (F-IF and Vit-IF, respectively). The number of secondary follicles after culture was negatively influenced by encapsulation of ovarian cortical pieces (6.3 ± 3.3 and 10.6 ± 0.9 vs 21.5 ± 2.3 in CP-0.5% and CP-1% vs CP, respectively). The diameter of follicles in IF-2% was higher than IF-1% (54.06 ± 2 vs 41.9 ± 1.5) and no significant difference in follicular viability was observed between the two groups. The proportions of different follicular types and their viability after culture in vitrified-warmed cortical pieces were comparable with fresh ones. The viability of vitrified-warmed isolated follicles was lower than fresh counterparts. The growth rate of fresh follicles was higher than vitrified-warmed follicles after culture (47.9 ± 1 vs 44.6 ± 1). In conclusion, while encapsulation of ovarian cortical pieces decreased the follicles' development, it could better support the growth of isolated follicles. Moreover, the viability and growth rate of isolated-encapsulated follicles was decreased by vitrification.

  18. In-situ formation of growth-factor-loaded coacervate microparticle-embedded hydrogels for directing encapsulated stem cell fate.

    Science.gov (United States)

    Jeon, Oju; Wolfson, David W; Alsberg, Eben

    2015-04-01

    The spontaneous formation of coacervate microdroplet-laden photo-crosslinked hydrogels derived from the simple mixing of oxidized, methacrylated alginate (OMA) and methacrylated gelatin (GelMA) enables simultaneous creation of drug-laden microdroplets and encapsulation of stem cells in photopolymerized coacervate hydrogels under physiological conditions. This can be utilized as a novel platform for in situ formation of localized, sustained bioactive molecule delivery to encapsulate stem cells for therapeutic applications.

  19. Interaction of Green Polymer Blend of Modified Sodium Alginate and Carboxylmethyl Cellulose Encapsulation of Turmeric Extract

    Directory of Open Access Journals (Sweden)

    Sa-Ad Riyajan

    2013-01-01

    Full Text Available Turmeric extract (tmr loaded nanoparticles were prepared by crosslinking modified carboxylmethyl cellulose (CMC and modified sodium alginate (SA with calcium ions, in a high pressure homogenizer. The FTIR spectra of CMC and SA were affected by blending due to hydrogen bonding. The negative zeta potential increased in magnitude with CMC content. The smallest nanoparticles were produced with a 10 : 5 SA/CMC blend. Also the release rates of the extract loading were measured, with model fits indicating that the loading level affected the release rate through nanoparticle structure. The 10 : 5 SA/CMC blend loading with tmr and pure tmr showed a good % growth inhibition of colon cancer cells which indicate that tmr in the presence of curcumin in tmr retains its anticancer activity even after being loaded into SA/CMC blend matrix.

  20. Long-term function of islets encapsulated in a re-designed alginate microcapsule construct in omentum pouches of immune-competent diabetic rats

    Science.gov (United States)

    Pareta, Rajesh; McQuilling, John P; Sittadjody, Sivanandane; Jenkins, Randy; Bowden, Stephen; Orlando, Giuseppe; Farney, Alan C; Brey, Eric M; Opara, Emmanuel C

    2014-01-01

    Objectives Our study aim was to determine encapsulated islet graft viability in an omentum pouch and the effect of FGF-1 released from our redesigned alginate microcapsules on the function of the graft. Methods Isolated rat islets were encapsulated in an inner core made with 1.5% low-viscosity high-mannuronic acid (LVM) alginate followed by an external layer made with 1.25% low-viscosity high-guluronic acid (LVG) alginate with or without FGF-1, in microcapsules measuring 300 – 400 μm in diameter. The two alginate layers were separated by a perm-selective membrane made with 0.1 % Poly-L-Ornithine (PLO), and the inner LVM core was partially chelated using 55 mM sodium citrate for 2 min. Results A marginal mass of encapsulated islet allografts (~2000 islets/kg) in Streptozotocin-diabetic Lewis rats caused significant reduction in blood glucose levels similar to the effect observed with encapsulated islet isografts. Transplantation of allo-islets co-encapsulated with FGF-1 did not result in better glycemic control, but induced greater body weight maintenance in transplant recipients compared to those that received only allo-islets. Histological examination of the retrieved tissue demonstrated morphologically and functionally intact islets in the microcapsules, with no signs of fibrosis. Conclusion We conclude that the omentum is a viable site for encapsulated islet transplantation. PMID:24681880

  1. Multipotent stromal cells derived from common marmoset Callithrix jacchus within alginate 3D environment: Effect of cryopreservation procedures.

    Science.gov (United States)

    Gryshkov, Oleksandr; Hofmann, Nicola; Lauterboeck, Lothar; Pogozhykh, Denys; Mueller, Thomas; Glasmacher, Birgit

    2015-08-01

    Multipotent stromal cells derived from the common marmoset monkey Callithrix jacchus (cjMSCs) possess high phylogenetic similarity to humans, with a great potential for preclinical studies in the field of regenerative medicine. Safe and effective long-term storage of cells is of great significance to clinical and research applications. Encapsulation of such cell types within alginate beads that can mimic an extra-cellular matrix and provide a supportive environment for cells during cryopreservation, has several advantages over freezing of cells in suspension. In this study we have analysed the effect of dimethyl sulfoxide (Me2SO, 2.5-10%, v/v) and pre-freeze loading time of alginate encapsulated cjMSCs in Me2SO (0-45 min) on the viability and metabolic activity of the cells after freezing using a slow cooling rate (-1°C/min). It was found that these parameters affect the stability and homogeneity of alginate beads after thawing. Moreover, the cjMSCs can be frozen in alginate beads with lower Me2SO concentration of 7.5% after 30 min of loading, while retaining high cryopreservation outcome. We demonstrated the maximum viability, membrane integrity and metabolic activity of the cells under optimized, less cytotoxic conditions. The results of this study are another step forward towards the application of cryopreservation for the long-term storage and subsequent applications of transplants in cell-based therapies.

  2. Application of modified-alginate encapsulated carbonate producing bacteria in concrete: a promising strategy for crack self-healing

    Directory of Open Access Journals (Sweden)

    Jianyun eWang

    2015-10-01

    Full Text Available Self-healing concrete holds promising benefits to reduce the cost for concrete maintenance and repair as cracks are autonomously repaired without any human intervention. In this study, the application of a carbonate precipitating bacterium Bacillus sphaericus was explored. Regarding the harsh condition in concrete, B. sphaericus spores were first encapsulated into a modified-alginate based hydrogel (AM-H which was proven to have a good compatibility with the bacteria and concrete regarding the influence on bacterial viability and concrete strength. Experimental results show that the spores were still viable after encapsulation. Encapsulated spores can precipitate a large amount of CaCO3 in/on the hydrogel matrix (around 70% by weight. Encapsulated B. sphaericus spores were added into mortar specimens and bacterial in-situ activity was demonstrated by the oxygen consumption on the mimicked crack surface. Specimens with free spores added showed no oxygen consumption. This indicates the efficient protection of the hydrogel for spores in concrete. To conclude, the AM-H encapsulated carbonate precipitating bacteria have great potential to be used for crack self-healing in concrete applications.

  3. Application of modified-alginate encapsulated carbonate producing bacteria in concrete: a promising strategy for crack self-healing.

    Science.gov (United States)

    Wang, Jianyun; Mignon, Arn; Snoeck, Didier; Wiktor, Virginie; Van Vliergerghe, Sandra; Boon, Nico; De Belie, Nele

    2015-01-01

    Self-healing concrete holds promising benefits to reduce the cost for concrete maintenance and repair as cracks are autonomously repaired without any human intervention. In this study, the application of a carbonate precipitating bacterium Bacillus sphaericus was explored. Regarding the harsh condition in concrete, B. sphaericus spores were first encapsulated into a modified-alginate based hydrogel (AM-H) which was proven to have a good compatibility with the bacteria and concrete regarding the influence on bacterial viability and concrete strength. Experimental results show that the spores were still viable after encapsulation. Encapsulated spores can precipitate a large amount of CaCO3 in/on the hydrogel matrix (around 70% by weight). Encapsulated B. sphaericus spores were added into mortar specimens and bacterial in situ activity was demonstrated by the oxygen consumption on the mimicked crack surface. While specimens with free spores added showed no oxygen consumption. This indicates the efficient protection of the hydrogel for spores in concrete. To conclude, the AM-H encapsulated carbonate precipitating bacteria have great potential to be used for crack self-healing in concrete applications.

  4. Cell-based delivery of glucagon-like peptide-1 using encapsulated mesenchymal stem cells

    DEFF Research Database (Denmark)

    Wallrapp, Christine; Thoenes, Eric; Thürmer, Frank

    2013-01-01

    Glucagon-like peptide-1 (GLP-1) CellBeads are cell-based implants for the sustained local delivery of bioactive factors. They consist of GLP-1 secreting mesenchymal stem cells encapsulated in a spherically shaped immuno-isolating alginate matrix. A highly standardized and reproducible encapsulation...... and quality control is performed in compliance with good manufacturing practice and fulfils all regulatory requirements for human clinical use. GLP-1 CellBeads combine the neuro- and cardioprotective properties of both GLP-1 and mesenchymal stem cells. First promising results were obtained from preclinical...... method is described for the manufacturing of homogeneous CellBeads. Viability and sustained secretion was shown for the recombinant GLP-1 and the cell endogenous bioactive factors like vascular endothelial growth factor, neurotrophin 3 (NT-3) and glial cell line-derived neurotrophic factor. Manufacturing...

  5. Preparation and characterization of novel polymeric microcapsules for live cell encapsulation and therapy.

    Science.gov (United States)

    Chen, Hongmei; Ouyang, Wei; Jones, Mitchell; Metz, Terrence; Martoni, Christopher; Haque, Tasima; Cohen, Rebecca; Lawuyi, Bisi; Prakash, Satya

    2007-01-01

    This article describes the preparation and in vitro characterization of novel genipin cross-linked alginate-chitosan (GCAC) microcapsules that have potential for live cell therapy applications. This microcapsule system, consisting of an alginate core with a covalently cross-linked chitosan membrane, was formed via ionotropic gelation between calcium ions and alginate, followed by chitosan coating by polyelectrolyte complexation and covalent cross-linking of chitosan by naturally derived genipin. Results showed that, using this design concept and the three-step procedure, spherical GCAC microcapsules with improved membrane strength, suppressed capsular swelling, and suitable permeability can be prepared. The suitability of this novel membrane formulation for live cell encapsulation was evaluated, using bacterial Lactobacillus plantarum 80 (pCBH1) (LP80) and mammalian HepG2 as model cells. Results showed that capsular integrity and bacterial cell viability were sustained 6 mo postencapsulation, suggesting the feasibility of using this microcapsule formulation for live bacterial cell encapsulation. The metabolic activity of the encapsulated HepG2 was also investigated. Results suggested the potential capacity of this GCAC microcapsule in cell therapy and the control of cell signaling; however, further research is required.

  6. Brown adipogenesis of mouse embryonic stem cells in alginate microstrands

    Science.gov (United States)

    Unser, Andrea Mannarino

    The ability of brown adipocytes (fat cells) to dissipate energy as heat shows great promise for the treatment of obesity and other metabolic disorders. Employing pluripotent stem cells, with an emphasis on directed differentiation, may overcome many issues currently associated with primary fat cell cultures. However, brown adipocytes are difficult to transplant in vivo due to the instability of fat, in terms of necrosis and neovascularization, once injected. Thus, 3D cell culture systems that have the potential to mimic adipogenic microenvironments are needed, not only to advance brown fat implantation, but also to better understand the role of brown adipocytes in treating obesity. To address this need, we created 3D "Brown-Fat-in-Microstrands" by microfluidic synthesis of alginate hydrogel microstrands that encapsulated cells and directly induced cell differentiation into brown adipocytes, using mouse embryonic stem cells (ESCs) as a model of pluripotent stem cells and brown preadipocytes as a positive control. The effect of hydrogel formation parameters on brown adipogenesis was studied, leading to the establishment of "Brown-Fat-in-Microstrands". Brown adipocyte differentiation within microstrands was confirmed by lipid droplet accumulation, immunocytochemistry and qPCR analysis of gene expression of brown adipocyte marker uncoupling protein 1 (UCP1) in addition to adipocyte marker expression. Compared to a 2D approach, 3D differentiated "Brown-Fat-in-Microstrands" exhibited higher level of brown adipocyte marker expression. The functional analysis of "Brown-Fat-in-Microstrands" was attempted by measuring the mitochondrial activity of ESC-differentiated brown adipocytes in 3D using Seahorse XF24 3 Extracellular Flux Analyzer. The ability to create "Brown-Fat-in-Microstrands" from pluripotent stem cells opens up a new arena to understanding brown adipogenesis and its implications in obesity and metabolic disorders.

  7. Tracking Hypoxic Signaling within Encapsulated Cell Aggregates

    Science.gov (United States)

    Skiles, Matthew L.; Sahai, Suchit; Blanchette, James O.

    2011-01-01

    In Diabetes mellitus type 1, autoimmune destruction of the pancreatic β-cells results in loss of insulin production and potentially lethal hyperglycemia. As an alternative treatment option to exogenous insulin injection, transplantation of functional pancreatic tissue has been explored1,2. This approach offers the promise of a more natural, long-term restoration of normoglycemia. Protection of the donor tissue from the host's immune system is required to prevent rejection and encapsulation is a method used to help achieve this aim. Biologically-derived materials, such as alginate3 and agarose4, have been the traditional choice for capsule construction but may induce inflammation or fibrotic overgrowth5 which can impede nutrient and oxygen transport. Alternatively, synthetic poly(ethylene glycol) (PEG)-based hydrogels are non-degrading, easily functionalized, available at high purity, have controllable pore size, and are extremely biocompatible,6,7,8. As an additional benefit, PEG hydrogels may be formed rapidly in a simple photo-crosslinking reaction that does not require application of non-physiological temperatures6,7. Such a procedure is described here. In the crosslinking reaction, UV degradation of the photoinitiator, 1-[4-(2-Hydroxyethoxy)-phenyl]-2-hydroxy-2-methyl-1-propane-1-one (Irgacure 2959), produces free radicals which attack the vinyl carbon-carbon double bonds of dimethacrylated PEG (PEGDM) inducing crosslinking at the chain ends. Crosslinking can be achieved within 10 minutes. PEG hydrogels constructed in such a manner have been shown to favorably support cells7,9, and the low photoinitiator concentration and brief exposure to UV irradiation is not detrimental to viability and function of the encapsulated tissue10. While we methacrylate our PEG with the method described below, PEGDM can also be directly purchased from vendors such as Sigma. An inherent consequence of encapsulation is isolation of the cells from a vascular network. Supply of

  8. Tracking hypoxic signaling within encapsulated cell aggregates.

    Science.gov (United States)

    Skiles, Matthew L; Sahai, Suchit; Blanchette, James O

    2011-12-16

    In Diabetes mellitus type 1, autoimmune destruction of the pancreatic β-cells results in loss of insulin production and potentially lethal hyperglycemia. As an alternative treatment option to exogenous insulin injection, transplantation of functional pancreatic tissue has been explored. This approach offers the promise of a more natural, long-term restoration of normoglycemia. Protection of the donor tissue from the host's immune system is required to prevent rejection and encapsulation is a method used to help achieve this aim. Biologically-derived materials, such as alginate and agarose, have been the traditional choice for capsule construction but may induce inflammation or fibrotic overgrowth which can impede nutrient and oxygen transport. Alternatively, synthetic poly(ethylene glycol) (PEG)-based hydrogels are non-degrading, easily functionalized, available at high purity, have controllable pore size, and are extremely biocompatible. As an additional benefit, PEG hydrogels may be formed rapidly in a simple photo-crosslinking reaction that does not require application of non-physiological temperatures. Such a procedure is described here. In the crosslinking reaction, UV degradation of the photoinitiator, 1-[4-(2-Hydroxyethoxy)-phenyl]-2-hydroxy-2-methyl-1-propane-1-one (Irgacure 2959), produces free radicals which attack the vinyl carbon-carbon double bonds of dimethacrylated PEG (PEGDM) inducing crosslinking at the chain ends. Crosslinking can be achieved within 10 minutes. PEG hydrogels constructed in such a manner have been shown to favorably support cells, and the low photoinitiator concentration and brief exposure to UV irradiation is not detrimental to viability and function of the encapsulated tissue. While we methacrylate our PEG with the method described below, PEGDM can also be directly purchased from vendors such as Sigma. An inherent consequence of encapsulation is isolation of the cells from a vascular network. Supply of nutrients, notably oxygen

  9. Alginate-Poly(ethylene glycol Hybrid Microspheres for Primary Cell Microencapsulation

    Directory of Open Access Journals (Sweden)

    Redouan Mahou

    2014-01-01

    Full Text Available The progress of medical therapies, which rely on the transplantation of microencapsulated living cells, depends on the quality of the encapsulating material. Such material has to be biocompatible, and the microencapsulation process must be simple and not harm the cells. Alginate-poly(ethylene glycol hybrid microspheres (alg-PEG-M were produced by combining ionotropic gelation of sodium alginate (Na-alg using calcium ions with covalent crosslinking of vinyl sulfone-terminated multi-arm poly(ethylene glycol (PEG-VS. In a one-step microsphere formation process, fast ionotropic gelation yields spherical calcium alginate gel beads, which serve as a matrix for simultaneously but slowly occurring covalent cross-linking of the PEG-VS molecules. The feasibility of cell microencapsulation was studied using primary human foreskin fibroblasts (EDX cells as a model. The use of cell culture media as polymer solvent, gelation bath, and storage medium did not negatively affect the alg-PEG-M properties. Microencapsulated EDX cells maintained their viability and proliferated. This study demonstrates the feasibility of primary cell microencapsulation within the novel microsphere type alg-PEG-M, serves as reference for future therapy development, and confirms the suitability of EDX cells as control model.

  10. Alginate-Poly(ethylene glycol) Hybrid Microspheres for Primary Cell Microencapsulation.

    Science.gov (United States)

    Mahou, Redouan; Meier, Raphael P H; Bühler, Léo H; Wandrey, Christine

    2014-01-09

    The progress of medical therapies, which rely on the transplantation of microencapsulated living cells, depends on the quality of the encapsulating material. Such material has to be biocompatible, and the microencapsulation process must be simple and not harm the cells. Alginate-poly(ethylene glycol) hybrid microspheres (alg-PEG-M) were produced by combining ionotropic gelation of sodium alginate (Na-alg) using calcium ions with covalent crosslinking of vinyl sulfone-terminated multi-arm poly(ethylene glycol) (PEG-VS). In a one-step microsphere formation process, fast ionotropic gelation yields spherical calcium alginate gel beads, which serve as a matrix for simultaneously but slowly occurring covalent cross-linking of the PEG-VS molecules. The feasibility of cell microencapsulation was studied using primary human foreskin fibroblasts (EDX cells) as a model. The use of cell culture media as polymer solvent, gelation bath, and storage medium did not negatively affect the alg-PEG-M properties. Microencapsulated EDX cells maintained their viability and proliferated. This study demonstrates the feasibility of primary cell microencapsulation within the novel microsphere type alg-PEG-M, serves as reference for future therapy development, and confirms the suitability of EDX cells as control model.

  11. Development and validation of a bioartificial liver device with fluidized bed bioreactors hosting alginate-encapsulated hepatocyte spheroids.

    Science.gov (United States)

    Figaro, S; Pereira, U; Rada, H; Semenzato, N; Pouchoulin, D; Legallais, C

    2015-08-01

    Acute and acute-on-chronic liver failure are associated to high mortality when transplantation is not possible. The lack of donors has resulted in an important demand for liver support devices. This paper describes the design and validation of a new bioartificial liver (BAL) device including fluidized bed bioreactors hosting alginate-encapsulated hepatocytes spheroids. To ensure the efficacy of the BAL and the safety of the patients, a complex extracorporeal circulation was designed to be compatible with a commercial medical device, the Prismaflex(®) monitor, already used in intensive care units. Preclinical studies on large animal show that the treatment was well tolerated in terms of hemodynamics considerations. A method using non adhesive coating in petri dish led to the production of large amount of viable spheroids in vitro that were further encapsulated to follow up bioartificial liver activity during four days.

  12. Flicking technique for microencapsulation of cells in calcium alginate leading to the microtissue formation.

    Science.gov (United States)

    Wong, Soon Chuan; Soon, Chin Fhong; Leong, Wai Yean; Tee, Kian Sek

    2016-01-01

    Microbeads have wide applications in biomedical engineering field that include drug delivery, encapsulation of biomolecules, tissue padding and tissue regeneration. In this paper, we report a simple, yet efficient, flicking technique to produce microcapsules of calcium alginate at a narrow distribution of size. The system consists of an infusion pump and a customised flicker that taps the syringe needle for dispersing microcapsules of sodium alginate that polymerised in the calcium chloride solution. The flow rate of the syringe pump and the velocity of the flicker were studied to achieve a well controlled and tunable size distribution of microbeads ranging from 200 to 400 μm. At a flow rate of 4 μl/min and flicking rate of 80 rpm, a narrow size distribution of microbeads were produced. Via this technique, HaCaT cells were encapsulated in calcium alginate microbeads that grown into microtissues with a size ranging from 100 to 300 μm after two weeks of culture. These microtissues could be potentially useful for pharmacological application.

  13. Enhanced lipid production by co-cultivation and co-encapsulation of oleaginous yeast Trichosporonoides spathulata with microalgae in alginate gel beads.

    Science.gov (United States)

    Kitcha, Suleeporn; Cheirsilp, Benjamas

    2014-05-01

    This study attempted to enhance biomass and lipid productivity of an oleaginous yeast Trichosporonoides spathulata by co-culturing with microalgae Chlorella spp., optimizing culture conditions, and encapsulating them in alginate gel beads. The co-culture of the yeast with microalgae Chlorella vulgaris var. vulgaris TISTR 8261 most enhanced overall biomass and lipid productivity by 1.6-fold of the yeast pure culture at 48 h and by 1.1-fold at 72 h. After optimization and scale-up in a bioreactor, this co-culture produced the highest biomass of 12.2 g/L with a high lipid content of 47 %. The dissolved oxygen monitoring system in the bioreactor showed that the microalgae worked well as an oxygen supplier to the yeast. This study also showed that the co-encapsulated yeast and microalgae could grow and produce lipid as same as their free cells did. Therefore, it is possible to apply this encapsulation technique for lipid production and simplification of downstream harvesting process. This co-culture system also produced the lipid with high content of saturated fatty acids, indicating its potential use as biodiesel feedstock with high oxidative stability.

  14. Gas-foaming calcium phosphate cement scaffold encapsulating human umbilical cord stem cells.

    Science.gov (United States)

    Chen, Wenchuan; Zhou, Hongzhi; Tang, Minghui; Weir, Michael D; Bao, Chongyun; Xu, Hockin H K

    2012-04-01

    Tissue engineering approaches are promising to meet the increasing need for bone regeneration. Calcium phosphate cement (CPC) can be injected and self-set to form a scaffold with excellent osteoconductivity. The objectives of this study were to develop a macroporous CPC-chitosan-fiber construct containing alginate-fibrin microbeads encapsulating human umbilical cord mesenchymal stem cells (hUCMSCs) and to investigate hUCMSC release from the degrading microbeads and proliferation inside the porous CPC construct. The hUCMSC-encapsulated microbeads were completely wrapped inside the CPC paste, with the gas-foaming porogen creating macropores in CPC to provide for access to culture media. Increasing the porogen content in CPC significantly increased the cell viability, from 49% of live cells in CPC with 0% porogen to 86% of live cells in CPC with 15% porogen. The alginate-fibrin microbeads started to degrade and release the cells inside CPC at 7 days. The released cells started to proliferate inside the macroporous CPC construct. The live cell number inside CPC increased from 270 cells/mm(2) at 1 day to 350 cells/mm(2) at 21 days. The pore volume fraction of CPC increased from 46.8% to 78.4% using the gas-foaming method, with macropore sizes of approximately 100 to 400 μm. The strength of the CPC-chitosan-fiber scaffold at 15% porogen was 3.8 MPa, which approximated the reported 3.5 MPa for cancellous bone. In conclusion, a novel gas-foaming macroporous CPC construct containing degradable alginate-fibrin microbeads was developed that encapsulated hUCMSCs. The cells had good viability while wrapped inside the porous CPC construct. The degradable microbeads in CPC quickly released the cells, which proliferated over time inside the porous CPC. Self-setting, strong CPC with alginate-fibrin microbeads for stem cell delivery is promising for bone tissue engineering applications.

  15. Biomimetic spinning of silk fibers and in situ cell encapsulation.

    Science.gov (United States)

    Cheng, Jie; Park, DoYeun; Jun, Yesl; Lee, JaeSeo; Hyun, Jinho; Lee, Sang-Hoon

    2016-07-01

    In situ embedding of sensitive materials (e.g., cells and proteins) in silk fibers without damage presents a significant challenge due to the lack of mild and efficient methods. Here, we report the development of a microfluidic chip-based method for preparation of meter-long silk fibroin (SF) hydrogel fibers by mimicking the silkworm-spinning process. For the spinning of SF fibers, alginate was used as a sericin-like material to induce SF phase separation and entrap liquid SFs, making it possible to shape the outline of SF-based fibers under mild physicochemical conditions. L929 fibroblasts were encapsulated in the fibric hydrogel and displayed excellent viability. Cell-laden SF fibric hydrogels prepared using our method offer a new type of SF-based biomedical device with potential utility in biomedicine.

  16. The collagen I mimetic peptide DGEA enhances an osteogenic phenotype in mesenchymal stem cells when presented from cell-encapsulating hydrogels.

    Science.gov (United States)

    Mehta, Manav; Madl, Christopher M; Lee, Shimwoo; Duda, Georg N; Mooney, David J

    2015-11-01

    Interactions between cells and the extracellular matrix (ECM) are known to play critical roles in regulating cell phenotype. The identity of ECM ligands presented to mesenchymal stem cells (MSCs) has previously been shown to direct the cell fate commitment of these cells. To enhance osteogenic differentiation of MSCs, alginate hydrogels were prepared that present the DGEA ligand derived from collagen I. When presented from hydrogel surfaces in 2D, the DGEA ligand did not facilitate cell adhesion, while hydrogels presenting the RGD ligand derived from fibronectin did encourage cell adhesion and spreading. However, the osteogenic differentiation of MSCs encapsulated within alginate hydrogels presenting the DGEA ligand was enhanced when compared with unmodified alginate hydrogels and hydrogels presenting the RGD ligand. MSCs cultured in DGEA-presenting gels exhibited increased levels of osteocalcin production and mineral deposition. These data suggest that the presentation of the collagen I-derived DGEA ligand is a feasible approach for selectively inducing an osteogenic phenotype in encapsulated MSCs.

  17. Encapsulation of α-lipoic acid intochitosan and alginate/gelatin hydrogel microparticles and its in vitro antioxidant activity

    Directory of Open Access Journals (Sweden)

    Vidović Bojana B.

    2016-01-01

    Full Text Available Alpha-lipoic acidis an organosulphur compound well-known for its therapeutic potential and antioxidant properties. However, the effective use of α-lipoic acid depends on biological plasma half-life and its preserving stability, which could be improved by encapsulation. In this study, α-lipoic acid was incorporated into chitosan microparticles obtained by reverse emulsion crosslinking technique, as well as into microparticles of alginate/gelatin crosslinked with zinc ions. Encapsulation of α-lipoic acid in both cases was carried out by swelling of synthesized dried microparticles by their dipping in a solution of the active substance under strictly controlled conditions. Encapsulation efficiency of α-lipoic acid obtained in this study was up to 53.9 %. The structural interaction of α-lipoic acid with the carriers was revealed by Fourier transform infrared spectroscopy. In vitro released studies showed that controlled release of α-lipoic acid was achieved through its encapsulation into chitosan microparticles. The results of in vitro antioxidative activity assays of released α-lipoic acid indicated that antioxidant activity was preserved at a satisfactory level. These obtained results suggested that chitosan microparticles could be suitable for modeling the controlled release of α-lipoic acid. [Projekat Ministartsva nauke Republike Srbije, br. III 46010 i br. III46001

  18. Mussel-inspired adhesive and transferable free-standing films by self-assembling dexamethasone encapsulated BSA nanoparticles and vancomycin immobilized oxidized alginate.

    Science.gov (United States)

    Han, Lu; Wang, Zhen-ming; Lu, Xiong; Dong, Li; Xie, Chao-ming; Wang, Ke-feng; Chen, Xiao-lang; Ding, Yong-hui; Weng, Lu-tao

    2015-02-01

    This study developed an adhesive and transferable free-standing (FS) film with dual function of osteoinductivity and antibacterial activity, which was obtained by sequentially assembling vancomycin immobilized oxidized sodium alginate and dexamethasone encapsulated chitosan coated BSA nanoparticles on a poly-dopamine layer. The FS films enabled the dual release of vancomycin and dexamethasone. The FS films had excellent osteoinductivity and antibacterial activity by cell culture and antibacterial assay. The FS film was detached from substrates and transferred to non-fouling surfaces by a wet transfer method, which demonstrated that the adhesive FS film is potential to modify biopolymers with non-fouling surfaces in mild and biocompatible conditions for biomedical applications.

  19. Oxidized alginate hydrogels as niche environments for corneal epithelial cells.

    Science.gov (United States)

    Wright, Bernice; De Bank, Paul A; Luetchford, Kim A; Acosta, Fernando R; Connon, Che J

    2014-10-01

    Chemical and biochemical modification of hydrogels is one strategy to create physiological constructs that maintain cell function. The aim of this study was to apply oxidised alginate hydrogels as a basis for development of a biomimetic niche for limbal epithelial stem cells that may be applied to treating corneal dysfunction. The stem phenotype of bovine limbal epithelial cells (LEC) and the viability of corneal epithelial cells (CEC) were examined in oxidised alginate gels containing collagen IV over a 3-day culture period. Oxidation increased cell viability (P ≤ 0.05) and this improved further with addition of collagen IV (P ≤ 0.01). Oxidised gels presented larger internal pores (diameter: 0.2-0.8 µm) than unmodified gels (pore diameter: 0.05-0.1 µm) and were significantly less stiff (P ≤ 0.001), indicating that an increase in pore size and a decrease in stiffness contributed to improved cell viability. The diffusion of collagen IV from oxidised alginate gels was similar to that of unmodified gels suggesting that oxidation may not affect the retention of extracellular matrix proteins in alginate gels. These data demonstrate that oxidised alginate gels containing corneal extracellular matrix proteins can influence corneal epithelial cell function in a manner that may impact beneficially on corneal wound healing therapy.

  20. Oxidized alginate hydrogels as niche environments for corneal epithelial cells

    Science.gov (United States)

    Wright, Bernice; De Bank, Paul A; Luetchford, Kim A; Acosta, Fernando R; Connon, Che J

    2014-01-01

    Chemical and biochemical modification of hydrogels is one strategy to create physiological constructs that maintain cell function. The aim of this study was to apply oxidised alginate hydrogels as a basis for development of a biomimetic niche for limbal epithelial stem cells that may be applied to treating corneal dysfunction. The stem phenotype of bovine limbal epithelial cells (LEC) and the viability of corneal epithelial cells (CEC) were examined in oxidised alginate gels containing collagen IV over a 3-day culture period. Oxidation increased cell viability (P ≤ 0.05) and this improved further with addition of collagen IV (P ≤ 0.01). Oxidised gels presented larger internal pores (diameter: 0.2–0.8 µm) than unmodified gels (pore diameter: 0.05–0.1 µm) and were significantly less stiff (P ≤ 0.001), indicating that an increase in pore size and a decrease in stiffness contributed to improved cell viability. The diffusion of collagen IV from oxidised alginate gels was similar to that of unmodified gels suggesting that oxidation may not affect the retention of extracellular matrix proteins in alginate gels. These data demonstrate that oxidised alginate gels containing corneal extracellular matrix proteins can influence corneal epithelial cell function in a manner that may impact beneficially on corneal wound healing therapy. © 2013 The Authors. Journal of Biomedical Materials Research Part A Published byWiley Periodicals, Inc. Part A: 102A: 3393–3400, 2014. PMID:24142706

  1. A novel alginate-encapsulated system to study biological response to critical-sized wear particles of UHMWPE loaded with alendronate sodium.

    Science.gov (United States)

    Liu, Yumei; Shi, Feng; Bo, Lin; Zhi, Wei; Weng, Jie; Qu, Shuxin

    2017-10-01

    The aim of this study was to develop a novel alginate-encapsulated system (Alg beads) to investigate the cell response to critical-sized wear particles of ultra-high molecular weight polyethylene loaded with alendronate sodium (UHMWPE-ALN), one of the most effective drugs to treat bone resorption in clinic. The extrusion method was used to prepare Alg beads encapsulating rat calvarial osteoblasts (RCOs) and critical-sized UHMWPE-ALN wear particles with spherical morphology and uniform size. The morphology, permeability and stability of Alg beads were characterized. The proliferation, ALP activity, cell apoptosis and distribution of live/dead RCOs co-cultured with wear particles in Alg beads were evaluated. RCOs and critical-sized UHMWPE-ALN wear particles distributed evenly and contacted efficiently in Alg beads. Alg beads were both permeable to trypsin and BSA, while the smaller the molecular was, the larger the diffuse was. The proliferation of RCOs in Alg beads increased with time, which indicated that Alg beads provided suitable conditions for cell culture. The long-term stability of Alg beads indicated the possibility for the longer time of co-cultured cells with wear particles. Critical-sized UHMWPE-ALN and UHMWPE wear particles both inhibited the proliferation and differentiation of RCOs, and induced the apoptosis of RCOs encapsulated in Alg beads. However, these effects could be significantly alleviated by the ALN released from the critical-sized UHMWPE-ALN wear particles. The present results suggested that this novel-developed co-culture system was feasible to evaluate the cell response to critical-sized UHMWPE-ALN wear particles for a longer time. Copyright © 2017 Elsevier B.V. All rights reserved.

  2. Stability Comparison of Free and Encapsulated Lactobacilus casei ATCC 393 in Yoghurt for Long Time Storage

    Directory of Open Access Journals (Sweden)

    Oana Lelia POP

    2016-11-01

    Full Text Available An innovative method of L. casei ATCC 393 encapsulation has been reported in the present study using pectin combined with alginate. The aim of this study was to investigate the effect of encapsulation on the survival of L. casei ATCC 393 in yoghurt during long time storage, free or encapsulated in alginate and alginate pectin microspheres, and influence over yoghurt properties, particularly acidification. Over 35 days of storage in yoghurt, the encapsulated probiotic cells proved a higher viability compared with free probiotic cells. An even higher viability and stability was observed for the samples where pectin was used. Pectin acts as prebiotic during encapsulation of L. casei ATCC 393.

  3. Alginate encapsulation of Begonia microshoots for short-term storage and distribution

    Science.gov (United States)

    Synthetic seeds were formed from in vitro grown Begonia cultivars (Sweetheart Mix and BabyWing White) shoot tips using 3% sodium alginate in Murashige and Skoog medium (MS) salt solution as the gel matrix and 100 mM calcium chloride for complexation. Synthetic seed formation was achieved by releasin...

  4. Versatile click alginate hydrogels crosslinked via tetrazine-norbornene chemistry.

    Science.gov (United States)

    Desai, Rajiv M; Koshy, Sandeep T; Hilderbrand, Scott A; Mooney, David J; Joshi, Neel S

    2015-05-01

    Alginate hydrogels are well-characterized, biologically inert materials that are used in many biomedical applications for the delivery of drugs, proteins, and cells. Unfortunately, canonical covalently crosslinked alginate hydrogels are formed using chemical strategies that can be biologically harmful due to their lack of chemoselectivity. In this work we introduce tetrazine and norbornene groups to alginate polymer chains and subsequently form covalently crosslinked click alginate hydrogels capable of encapsulating cells without damaging them. The rapid, bioorthogonal, and specific click reaction is irreversible and allows for easy incorporation of cells with high post-encapsulation viability. The swelling and mechanical properties of the click alginate hydrogel can be tuned via the total polymer concentration and the stoichiometric ratio of the complementary click functional groups. The click alginate hydrogel can be modified after gelation to display cell adhesion peptides for 2D cell culture using thiol-ene chemistry. Furthermore, click alginate hydrogels are minimally inflammatory, maintain structural integrity over several months, and reject cell infiltration when injected subcutaneously in mice. Click alginate hydrogels combine the numerous benefits of alginate hydrogels with powerful bioorthogonal click chemistry for use in tissue engineering applications involving the stable encapsulation or delivery of cells or bioactive molecules.

  5. Chondrocyte Culture in Three Dimensional Alginate Sulfate Hydrogels Promotes Proliferation While Maintaining Expression of Chondrogenic Markers

    Science.gov (United States)

    Mhanna, Rami; Kashyap, Aditya; Palazzolo, Gemma; Vallmajo-Martin, Queralt; Becher, Jana; Möller, Stephanie; Schnabelrauch, Matthias

    2014-01-01

    The loss of expression of chondrogenic markers during monolayer expansion remains a stumbling block for cell-based treatment of cartilage lesions. Here, we introduce sulfated alginate hydrogels as a cartilage biomimetic biomaterial that induces cell proliferation while maintaining the chondrogenic phenotype of encapsulated chondrocytes. Hydroxyl groups of alginate were converted to sulfates by incubation with sulfur trioxide–pyridine complex (SO3/pyridine), yielding a sulfated material cross-linkable with calcium chloride. Passage 3 bovine chondrocytes were encapsulated in alginate and alginate sulfate hydrogels for up to 35 days. Cell proliferation was five-fold higher in alginate sulfate compared with alginate (p=0.038). Blocking beta1 integrins in chondrocytes within alginate sulfate hydrogels significantly inhibited proliferation (p=0.002). Sulfated alginate increased the RhoA activity of chondrocytes compared with unmodified alginate, an increase that was blocked by β1 blocking antibodies (p=0.017). Expression and synthesis of type II collagen, type I collagen, and proteoglycan was not significantly affected by the encapsulation material evidenced by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and immunohistochemistry. Alginate sulfate constructs showed an opaque appearance in culture, whereas the unmodified alginate samples remained translucent. In conclusion, alginate sulfate provides a three dimensional microenvironment that promotes both chondrocyte proliferation and maintenance of the chondrogenic phenotype and represents an important advance for chondrocyte-based cartilage repair therapies providing a material in which cell expansion can be done in situ. PMID:24320935

  6. PLGA/alginate composite microspheres for hydrophilic protein delivery

    Energy Technology Data Exchange (ETDEWEB)

    Zhai, Peng [Department of Anatomy and Cell Biology, University of Saskatchewan, S7N5E5 (Canada); Division of Biomedical Engineering, University of Saskatchewan, S7N5A9 (Canada); Chen, X.B. [Department of Mechanical Engineering, University of Saskatchewan, S7N5A9 (Canada); Division of Biomedical Engineering, University of Saskatchewan, S7N5A9 (Canada); Schreyer, David J., E-mail: david.schreyer@usask.ca [Department of Anatomy and Cell Biology, University of Saskatchewan, S7N5E5 (Canada); Division of Biomedical Engineering, University of Saskatchewan, S7N5A9 (Canada)

    2015-11-01

    Poly(lactic-co-glycolic acid) (PLGA) microspheres and PLGA/alginate composite microspheres were prepared by a novel double emulsion and solvent evaporation technique and loaded with bovine serum albumin (BSA) or rabbit anti-laminin antibody protein. The addition of alginate and the use of a surfactant during microsphere preparation increased the encapsulation efficiency and reduced the initial burst release of hydrophilic BSA. Confocal laser scanning microcopy (CLSM) of BSA-loaded PLGA/alginate composite microspheres showed that PLGA, alginate, and BSA were distributed throughout the depths of microspheres; no core/shell structure was observed. Scanning electron microscopy revealed that PLGA microspheres erode and degrade more quickly than PLGA/alginate composite microspheres. When loaded with anti-laminin antibody, the function of released antibody was well preserved in both PLGA and PLGA/alginate composite microspheres. The biocompatibility of PLGA and PLGA/alginate microspheres were examined using four types of cultured cell lines, representing different tissue types. Cell survival was variably affected by the inclusion of alginate in composite microspheres, possibly due to the sensitivity of different cell types to excess calcium that may be released from the calcium cross-linked alginate. - Highlights: • A double emulsion technique is used to prepare protein-loaded PLGA or PLGA/alginate microspheres. • PLGA, alginate and protein are distributed evenly within microsphere structure. • Addition of alginate improves loading efficiency and slows degradation and protein release. • PLGA/alginate microspheres have favorable biocompatibility.

  7. Alginate encapsulated mesoporous silica nanospheres as a sustained drug delivery system for the poorly water-soluble drug indomethacin

    Directory of Open Access Journals (Sweden)

    Liang Hu

    2014-08-01

    Full Text Available We applied a combination of inorganic mesoporous silica material, frequently used as drug carriers, and a natural organic polymer alginate (ALG, to establish a sustained drug delivery system for the poorly water-soluble drug Indomethacin (IND. Mesoporous silica nanospheres (MSNs were synthesized using an organic template method and then functionalized with aminopropyl groups through postsynthesis. After drug loading into the pores of aninopropyl functionalized MSNs (AP-MSNs, IND loaded AP-MSNs (IND-AP-MSNs were encapsulated by ALG through the ionic interaction. The effects of surface chemical groups and ALG layer on IND release were systematically studied using scanning electron microscopy (SEM, transmission electron microscopy (TEM, nitrogen adsorption, zeta-potential analysis and TGA analysis. The surface structure and surface charge changes of the ALG encapsulated AP-MSNs (ALG-AP-MSNs were also investigated. The results showed that sustained release of IND from the designed drug delivery system was mainly due to the blockage effect from the coated ALG. We believe that this combination will help designing oral sustained drug delivery systems for poorly water-soluble drugs.

  8. Microencapsulation of bioactives in cross-linked alginate matrices by spray drying.

    Science.gov (United States)

    Santa-Maria, Monica; Scher, Herbert; Jeoh, Tina

    2012-01-01

    Microencapsulation of biomolecules, cells and chemicals is widely used in the food and pharmaceutical industries to improve stability, delivery and to control the release of encapsulated moieties. Among encapsulation matrices, alginate is preferred due to its low cost, biodegradability and biocompatibility. Current methods for producing stable alginate gels involve dropping alginate suspensions into divalent cation solutions. This procedure is difficult to scale-up and produces undesirably large alginate beads. In our novel encapsulation method, alginate gelation occurs during spray drying upon volatilisation of a base and rapid release of otherwise unavailable calcium ions. The resulting particles, with median particle sizes in the range 15-120 µm, are insoluble in solution. Cellulase and hemicellulase activities encapsulated by this method were not compromised during spray drying and remained stable over prolonged storage. The procedure described here offers a one-step alternative to other encapsulation methods that are costly and difficult to scale-up.

  9. Production and characterization of engineered alginate-based microparticles containing ECM powder for cell/tissue engineering applications.

    Science.gov (United States)

    Mazzitelli, Stefania; Luca, Giovanni; Mancuso, Francesca; Calvitti, Mario; Calafiore, Riccardo; Nastruzzi, Claudio; Johnson, Scott; Badylak, Stephen F

    2011-03-01

    A method for the production of engineered alginate-based microparticles, containing extracellular matrix and neonatal porcine Sertoli cells (SCs), is described. As a source for extracellular matrix, a powder form of isolated and purified urinary bladder matrix (UBM) was employed. We demonstrated that the incorporation of UBM does not significantly alter the morphological and dimensional characteristics of the microparticles. The alginate microparticles were used for SC encapsulation as an immunoprotective barrier for transplant purposes, while the co-entrapped UBM promoted retention of cell viability and function. These engineered microparticles could represent a novel approach to enhancing immunological acceptance and increasing the functional life-span of the entrapped cells for cell/tissue engineering applications. In this respect, it is noteworthy that isolated neonatal porcine SCs, administered alone in highly biocompatible microparticles, led to diabetes prevention and reversion in nonobese diabetic (NOD) mice.

  10. 3 dimensional cell cultures: a comparison between manually and automatically produced alginate beads.

    Science.gov (United States)

    Lehmann, R; Gallert, C; Roddelkopf, T; Junginger, S; Wree, A; Thurow, K

    2016-08-01

    Cancer diseases are a common problem of the population caused by age and increased harmful environmental influences. Herein, new therapeutic strategies and compound screenings are necessary. The regular 2D cultivation has to be replaced by three dimensional cell culturing (3D) for better simulation of in vivo conditions. The 3D cultivation with alginate matrix is an appropriate method for encapsulate cells to form cancer constructs. The automated manufacturing of alginate beads might be an ultimate method for large-scaled manufacturing constructs similar to cancer tissue. The aim of this study was the integration of full automated systems for the production, cultivation and screening of 3D cell cultures. We compared the automated methods with the regular manual processes. Furthermore, we investigated the influence of antibiotics on these 3D cell culture systems. The alginate beads were formed by automated and manual procedures. The automated steps were processes by the Biomek(®) Cell Workstation (celisca, Rostock, Germany). The proliferation and toxicity were manually and automatically evaluated at day 14 and 35 of cultivation. The results visualized an accumulation and expansion of cell aggregates over the period of incubation. However, the proliferation and toxicity were faintly and partly significantly decreased on day 35 compared to day 14. The comparison of the manual and automated methods displayed similar results. We conclude that the manual production process could be replaced by the automation. Using automation, 3D cell cultures can be produced in industrial scale and improve the drug development and screening to treat serious illnesses like cancer.

  11. Modulation of immunity and gut microbiota after dietary administration of alginate encapsulated Shewanella putrefaciens Pdp11 to gilthead seabream (Sparus aurata L.).

    Science.gov (United States)

    Cordero, Héctor; Guardiola, Francisco A; Tapia-Paniagua, Silvana Teresa; Cuesta, Alberto; Meseguer, José; Balebona, M Carmen; Moriñigo, M Ángel; Esteban, M Ángeles

    2015-08-01

    The potential benefits of probiotics when administering to fish could improve aquaculture production. The objective of this study was to examine the modulation of immune status and gut microbiota of gilthead seabream (Sparus aurata L.) specimens by a probiotic when administered encapsulated. Commercial diet was enriched with Shewanella putrefaciens Pdp11 (SpPdp11, at a concentration of 10(8) cfu g(-1)) before being encapsulated in calcium alginate beads. Fish were fed non-supplemented (control) or supplemented diet for 4 weeks. After 1, 2 and 4 weeks the main humoral and cellular immune parameters were determined. Furthermore, gene expression profile of five immune relevant genes (il1β, bd, mhcIIα, ighm and tcrβ) was studied by qPCR in head kidney. On the other hand, intestinal microbiota of fish was analysed at 7 and 30 days by DGGE. Results demonstrated that administration of alginate encapsulated SpPdp11 has immunostimulant properties on humoral parameters (IgM level and serum peroxidase activity). Although no immunostimulant effects were detected on leucocyte activities, significant increases were detected in the level of mRNA of head-kidney leucocytes for mhcIIα and tcrβ after 4 weeks of feeding the encapsulated-probiotic diet. The administration of SpPdp11 encapsulated in alginate beads produced important changes in the DGGE patterns corresponding to the intestinal microbiota. Predominant bands related to lactic acid bacteria, such as Lactococcus and Lactobacillus strains, were sequenced from the DGGE patterns of fish fed the probiotic diet, whereas they were not sequenced from fish receiving the control diet. The convenience or not of probiotic encapsulation is discussed. Copyright © 2015 Elsevier Ltd. All rights reserved.

  12. Ca2+ released from calcium alginate gels can promote inflammatory responses in vitro and in vivo

    Science.gov (United States)

    Chan, Gail; Mooney, David J.

    2013-01-01

    In general, alginate hydrogels are considered to be biologically inert and are commonly used for biomedical purposes that require minimum inflammation. However, Ca2+, which is commonly used to crosslink alginate, is a critical second messenger in immune cell signaling, and little has been done to understand its effect on immune cell fate when delivered as a component of alginate gels. We found that dendritic cells (DCs) encapsulated in Ca2+-crosslinked alginate (calcium alginate) secreted at least fivefold more of the inflammatory cytokine IL-1β when compared to DCs encapsulated in agarose and collagen gels, as well as DCs plated on tissue-culture polystyrene (TCPS). Plating cells on TCPS with the alginate polymer could not reproduce these results, whereas culturing DCs on TCPS with increasing concentrations of Ca2+ increased IL-1β, MHC class II and CD86 expression in a dose-dependent manner. In agreement with these findings, calcium alginate gels induced greater maturation of encapsulated DCs compared to barium alginate gels. When injected subcutaneously in mice, calcium alginate gels significantly upregulated IL-1β secretion from surrounding tissue relative to barium alginate gels, and similarly, the inflammatory effects of LPS were enhanced when it was delivered from calcium alginate gels rather than barium alginate gels. These results confirm that the Ca2+ used to crosslink alginate gels can be immunostimulatory and suggest that it is important to take into account Ca2+’s bioactive effects on all exposed cells (both immune and non-immune) when using calcium alginate gels for biomedical purposes. This work may strongly impact the way people use alginate gels in the future as well as provide insights into past work utilizing alginate gels. PMID:23938198

  13. Development of functionalized multi-walled carbon-nanotube-based alginate hydrogels for enabling biomimetic technologies

    Science.gov (United States)

    Joddar, Binata; Garcia, Eduardo; Casas, Atzimba; Stewart, Calvin M.

    2016-08-01

    Alginate is a hydrogel commonly used for cell culture by ionically crosslinking in the presence of divalent Ca2+ ions. However these alginate gels are mechanically unstable, not permitting their use as scaffolds to engineer robust biological bone, breast, cardiac or tumor tissues. This issue can be addressed via encapsulation of multi-walled carbon nanotubes (MWCNT) serving as a reinforcing phase while being dispersed in a continuous phase of alginate. We hypothesized that adding functionalized MWCNT to alginate, would yield composite gels with distinctively different mechanical, physical and biological characteristics in comparison to alginate alone. Resultant MWCNT-alginate gels were porous, and showed significantly less degradation after 14 days compared to alginate alone. In vitro cell-studies showed enhanced HeLa cell adhesion and proliferation on the MWCNT-alginate compared to alginate. The extent of cell proliferation was greater when cultured atop 1 and 3 mg/ml MWCNT-alginate; although all MWCNT-alginates lead to enhanced cell cluster formation compared to alginate alone. Among all the MWCNT-alginates, the 1 mg/ml gels showed significantly greater stiffness compared to all other cases. These results provide an important basis for the development of the MWCNT-alginates as novel substrates for cell culture applications, cell therapy and tissue engineering.

  14. Impact of alginate-producing Pseudomonas aeruginosa on alveolar macrophage apoptotic cell clearance.

    Science.gov (United States)

    McCaslin, Charles A; Petrusca, Daniela N; Poirier, Christophe; Serban, Karina A; Anderson, Gregory G; Petrache, Irina

    2015-01-01

    Pseudomonas aeruginosa infection is a hallmark of lung disease in cystic fibrosis. Acute infection with P. aeruginosa profoundly inhibits alveolar macrophage clearance of apoptotic cells (efferocytosis) via direct effect of virulence factors. During chronic infection, P. aeruginosa evades host defense by decreased virulence, which includes the production or, in the case of mucoidy, overproduction of alginate. The impact of alginate on innate immunity, in particular on macrophage clearance of apoptotic cells is not known. We hypothesized that P. aeruginosa strains that exhibit reduced virulence impair macrophage clearance of apoptotic cells and we investigated if the polysaccharide alginate produced by mucoid P. aeruginosa is sufficient to inhibit alveolar macrophage efferocytosis. Rat alveolar or human peripheral blood monocyte (THP-1)-derived macrophage cell lines were exposed in vitro to exogenous alginate or to wild type or alginate-overproducing mucoid P. aeruginosa prior to challenge with apoptotic human Jurkat T-lymphocytes. The importance of LPS contamination and that of structural integrity of alginate polymers was tested using alginate of different purities and alginate lyase, respectively. Alginate inhibited alveolar macrophage efferocytosis in a dose- and time-dependent manner. This effect was augmented but not exclusively attributed to lipopolysaccharide (LPS) present in alginates. Alginate-producing P. aeruginosa inhibited macrophage efferocytosis by more than 50%. A mannuronic-specific alginate lyase did not restore efferocytosis inhibited by exogenous guluronic-rich marine alginate, but had a marked beneficial effect on efferocytosis of alveolar macrophages exposed to mucoid P. aeruginosa. Despite decreased virulence, mucoid P. aeruginosa may contribute to chronic airway inflammation through significant inhibition of alveolar clearance of apoptotic cells and debris. The mechanism by which mucoid bacteria inhibit efferocytosis may involve alginate

  15. Engineered yeast whole-cell biocatalyst for direct degradation of alginate from macroalgae and production of non-commercialized useful monosaccharide from alginate.

    Science.gov (United States)

    Takagi, Toshiyuki; Yokoi, Takahiro; Shibata, Toshiyuki; Morisaka, Hironobu; Kuroda, Kouichi; Ueda, Mitsuyoshi

    2016-02-01

    Alginate is a major component of brown macroalgae. In macroalgae, an endolytic alginate lyase first degrades alginate into oligosaccharides. These oligosaccharides are further broken down into monosaccharides by an exolytic alginate lyase. In this study, genes encoding various alginate lyases derived from alginate-assimilating marine bacterium Saccharophagus degradans were isolated, and their enzymes were displayed using the yeast cell surface display system. Alg7A-, Alg7D-, and Alg18J-displaying yeasts showed endolytic alginate lyase activity. On the other hand, Alg7K-displaying yeast showed exolytic alginate lyase activity. Alg7A, Alg7D, Alg7K, and Alg18J, when displayed on yeast cell surface, demonstrated both polyguluronate lyase and polymannuronate lyase activities. Additionally, polyguluronic acid could be much easily degraded by Alg7A, Alg7K, and Alg7D than polymannuronic acid. In contrast, polymannuronic acid could be much easily degraded by Alg18J than polyguluronic acid. We further constructed yeasts co-displaying endolytic and exolytic alginate lyases. Degradation efficiency by the co-displaying yeasts were significantly higher than single alginate lyase-displaying yeasts. Alg7A/Alg7K co-displaying yeast had maximum alginate degrading activity, with production of 1.98 g/L of reducing sugars in a 60-min reaction. This system developed, along with our findings, will contribute to the efficient utilization and production of useful and non-commercialized monosaccharides from alginate by Saccharomyces cerevisiae.

  16. Encapsulation of probiotic Bifidobacterium longum BIOMA 5920 with alginate-human-like collagen and evaluation of survival in simulated gastrointestinal conditions.

    Science.gov (United States)

    Su, Ran; Zhu, Xiao-Li; Fan, Dai-Di; Mi, Yu; Yang, Chan-Yuan; Jia, Xin

    2011-12-01

    Alginate (ALg)-human-like collagen (HLC) microspheres were prepared by the technology of electrostatic droplet generation in order to develop a biocompatible vehicle for probiotic bacteria. Microparticles were spherical with mean particle size of 400μm. The encapsulation efficiency (EE) of ALg-HLC microspheres could reach 92-99.2%. Water-soluble and fibrous human-like collagen is combined with sodium alginate through intermolecular hydrogen bonding and electrostatic force which were investigated by Fourier transform infrared spectroscopy (FTIR) and differential scanning calorimetry (DSC), thus the matrix of ALg-HLC was very stable. Bifidobacterium longum BIOMA 5920, as a kind of probiotic bacteria, was encapsulated with alginate-human-like collagen to survive and function in simulated gastrointestinal juice. Microparticles were very easy to degradation in simulated intestinal juices. After incubation in simulated gastric (pH 2.0, 2h), the encapsulated B. longum BIOMA 5920 numbers were 4.81 ± 0.38 log cfu/g. Copyright © 2011 Elsevier B.V. All rights reserved.

  17. Inhibition of Tumor Growth in Mice by Endostatin Derived from Abdominal Transplanted Encapsulated Cells

    Institute of Scientific and Technical Information of China (English)

    Huaining TENG; Ying ZHANG; Wei WANG; Xiaojun MA; Jian FEI

    2007-01-01

    Endostatin, a C-terminal fragment of collagen 18a, inhibits the growth of established tumors and metastases in vivo by inhibiting angiogenesis. However, the purification procedures required for largescale production and the attendant cost of these processes, together with the low effectiveness in clinical tests, suggest that alternative delivery methods might be required for efficient therapeutic use of endostatin.In the present study, we transfected Chinese hamster ovary (CHO) cells with a human endostatin gene expression vector and encapsulated the CHO cells in alginate-poly-L-lysine microcapsules. The release of biologically active endostatin was confirmed using the chicken chorioallantoic membrane assay. The encapsulated endostatin-expressing CHO cells can inhibit the growth of primary tumors in a subcutaneous B16 tumor model when injected into the abdominal cavity of mouse. These results widen the clinical application of the microencapsulated cell endostatin delivery system in cancer treatment.

  18. Augmentation of the antibody response of Atlantic salmon by oral administration of alginate-encapsulated IPNV antigens.

    Directory of Open Access Journals (Sweden)

    Lihan Chen

    Full Text Available The objective of the present study was to assess the effect of alginate-encapsulated infectious pancreatic necrosis virus antigens in inducing the immune response of Atlantic salmon as booster vaccines. One year after intraperitoneal injection with an oil-adjuvanted vaccine, post-smolts were orally boosted either by 1 alginate-encapsulated IPNV antigens (ENCAP; 2 soluble antigens (UNENCAP or 3 untreated feed (control. This was done twice, seven weeks apart. Sampling was done twice, firstly at 7 weeks post 1st oral boost and the 2nd, at 4 weeks after the 2nd oral boost. Samples included serum, head kidney, spleen and hindgut. Serum antibodies were analyzed by ELISA while tissues were used to assess the expression of IgM, IgT, CD4, GATA3, FOXP3, TGF-β and IL-10 genes by quantitative PCR. Compared to controls, fish fed with ENCAP had a significant increase (p<0.04 in serum antibodies following the 1st boost but not after the 2nd boost. This coincided with significant up-regulation of CD4 and GATA3 genes. In contrast, serum antibodies in the UNENCAP group decreased both after the 1st and 2nd oral boosts. This was associated with significant up-regulation of FOXP3, TGF-β and IL-10 genes. The expression of IgT was not induced in the hindgut after the 1st oral boost but was significantly up-regulated following the 2nd one. CD4 and GATA3 mRNA expressions exhibited a similar pattern to IgT in the hindgut. IgM mRNA expression on the other hand was not differentially regulated at any of the times examined. Our findings suggest that 1 Parenteral prime with oil-adjuvanted vaccines followed by oral boost with ENCAP results in augmentation of the systemic immune response; 2 Symmetrical prime and boost (mucosal with ENCAP results in augmentation of mucosal immune response and 3 Symmetrical priming and boosting (mucosal with soluble antigens results in the induction of systemic immune tolerance.

  19. ENTRAPMENT OF FLUORESCENT E. COLI CELLS IN ALGINATE GEL

    Directory of Open Access Journals (Sweden)

    T. VINTILA

    2013-07-01

    Full Text Available By this experiment we will demonstrate the possibility to obtain genetically modified microbial strains that can be used as markers in different studies. The trait transferred in this study is the fluorescence in UV light expressed by a gene isolated from jellyfish. This gene was insered into a plasmid carrying ampiciline resistance and in the operon for arabinose fermentation. The plasmid was called pGLO. E coli HB101 K-12, ampicillin resistant colonies has been obtained. The colonies on the LB/amp/ara plate fluoresce green under UV light and the transformed colonies can grow on ampicillin. Transformation efficiency = 362 transformed colonies/ μg DNA. The cells where immobilized by entrapment in alginate gel to study the phenomenon involved in cells immobilization. After immobilization in alginate gel, 5x104 cells of E. coli pGLO / capsule and 1,4 x 105 cells of E. coli HB101/capsule has been found. Fluorescent microscopy revealed the presence of pGLO carrying cells into the capsules. After cultivation of alginate capsules containing E. coli in LB broth, and fluorescent microscopy of the capsule sections, several observations of the phenomenon involved in continuous fermentation using biocatalysts in has been made. These cells grow and migrate to the cortical part of the matrix where they are immobilized.

  20. ENTRAPMENT OF FLUORESCENT E. COLI CELLS IN ALGINATE GEL

    Directory of Open Access Journals (Sweden)

    V. IGNA

    2009-05-01

    Full Text Available By this experiment we will demonstrate the possibility to obtain genetically modifiedmicrobial strains that can be used as markers in different studies. The traittransferred in this study is the fluorescence in UV light expressed by a gene isolatedfrom jellyfish. This gene was insered into a plasmid carrying ampiciline resistanceand in the operon for arabinose fermentation. The plasmid was called pGLO. E coliHB101 K-12, ampicillin resistant colonies has been obtained. The colonies on theLB/amp/ara plate fluoresce green under UV light and the transformed colonies cangrow on ampicillin. Transformation efficiency = 362 transformed colonies/ μg DNA.The cells where immobilized by entrapment in alginate gel to study the phenomenoninvolved in cells immobilization. After immobilization in alginate gel, 5x104 cells ofE. coli pGLO / capsule and 1,4 x 105 cells of E. coli HB101/capsule has been found.Fluorescent microscopy revealed the presence of pGLO carrying cells into thecapsules. After cultivation of alginate capsules containing E. coli in LB broth, andfluorescent microscopy of the capsule sections, several observations of thephenomenon involved in continuous fermentation using biocatalysts in has beenmade. These cells grow and migrate to the cortical part of the matrix where they areimmobilized.

  1. Layer-shaped alginate hydrogels enhance the biological performance of human adipose-derived stem cells

    Directory of Open Access Journals (Sweden)

    Galateanu Bianca

    2012-06-01

    Full Text Available Abstract Background The reconstruction of adipose tissue defects is often challenged by the complications that may occur following plastic and reconstructive surgery, including donor-site morbidity, implant migration and foreign body reaction. To overcome these problems, adipose tissue engineering (ATE using stem cell-based regeneration strategies has been widely explored in the last years. Mounting evidence has shown that adipose-derived stem cells (ADSCs represent a promising cell source for ATE. In the context of a small number of reports concerning adipose tissue regeneration using three-dimensional (3-D systems, the present study was designed to evaluate the biological performance of a novel alginate matrix that incorporates human ADSCs (hADSCs. Results Culture-expanded cells isolated from the stromal vascular fraction (SVF, corresponding to the third passage which showed the expression of mesenchymal stem cell (MSC markers, were used in the 3-D culture systems. The latter represented a calcium alginate hydrogel, obtained by the diffusion of calcium gluconate (CGH matrix, and shaped as discoid-thin layer. For comparative purposes, a similar hADSC-laden alginate hydrogel cross-linked with calcium chloride was considered as reference hydrogel (RH matrix. Both hydrogels showed a porous structure under scanning electron microscopy (SEM and the hADSCs embedded displayed normal spherical morphologies, some of them showing signs of mitosis. More than 85% of the entrapped cells survived throughout the incubation period of 7 days. The percentage of viable cells was significantly higher within CGH matrix at 2 days post-seeding, and approximately similar within both hydrogels after 7 days of culture. Moreover, both alginate-based hydrogels stimulated cell proliferation. The number of hADSC within hydrogels has increased during the incubation period of 7 days and was higher in the case of CGH matrix. Cells grown under adipogenic conditions for

  2. Ultra-structural changes and expression of chondrogenic and hypertrophic genes during chondrogenic differentiation of mesenchymal stromal cells in alginate beads

    Directory of Open Access Journals (Sweden)

    Havva Dashtdar

    2016-03-01

    Full Text Available Chondrogenic differentiation of mesenchymal stromal cells (MSCs in the form of pellet culture and encapsulation in alginate beads has been widely used as conventional model for in vitro chondrogenesis. However, comparative characterization between differentiation, hypertrophic markers, cell adhesion molecule and ultrastructural changes during alginate and pellet culture has not been described. Hence, the present study was conducted comparing MSCs cultured in pellet and alginate beads with monolayer culture. qPCR was performed to assess the expression of chondrogenic, hypertrophic, and cell adhesion molecule genes, whereas transmission electron microscopy (TEM was used to assess the ultrastructural changes. In addition, immunocytochemistry for Collagen type II and aggrecan and glycosaminoglycan (GAG analysis were performed. Our results indicate that pellet and alginate bead cultures were necessary for chondrogenic differentiation of MSC. It also indicates that cultures using alginate bead demonstrated significantly higher (p < 0.05 chondrogenic but lower hypertrophic (p < 0.05 gene expressions as compared with pellet cultures. N-cadherin and N-CAM1 expression were up-regulated in second and third weeks of culture and were comparable between the alginate bead and pellet culture groups, respectively. TEM images demonstrated ultrastructural changes resembling cell death in pellet cultures. Our results indicate that using alginate beads, MSCs express higher chondrogenic but lower hypertrophic gene expression. Enhanced production of extracellular matrix and cell adhesion molecules was also observed in this group. These findings suggest that alginate bead culture may serve as a superior chondrogenic model, whereas pellet culture is more appropriate as a hypertrophic model of chondrogenesis.

  3. Effect of Glycerol, as Cryoprotectant in the Encapsulation and Freeze Drying of Microspheres Containing Probiotic Cells

    Directory of Open Access Journals (Sweden)

    Oana Lelia Pop

    2015-05-01

    Full Text Available It is reported that probiotics provide several health benefits as they help in maintaining a good balance and composition of intestinal flora, and increase the resistance against invasion of pathogens. Ensuring adequate dosages of probiotics at the time of consumption is a challenge, because several factors during processing and storage affect the viability of probiotic organisms. Major emphasis has been given to protect the microorganisms with the help of encapsulation technique, by addition of different protectants. In this study, probiotic cells (Bifidobacterium lactis 300B were entrapped in alginate/pullulan microspheres. In the encapsulation formula glycerol was used as cryoprotectant in the freeze drying process for long time storage. It was observed that the survival of Bifidobacterium lactis 300B when encapsulated without cryoprotectant was higher than the formula with glycerol in the fresh obtained microspheres. The addition of glycerol was in order to reduce the deep freezing and freeze drying damages. In the chosen formulations, glycerol did not proved protection for the entrapped probiotic cells in the freeze drying process, for which the use of glycerol as cryoprotectant for alginate/pullulan Bifidobacterium lactis 300B entrapment is not recommended.

  4. Evaluation of Three-Dimensional Porous Iron-Cross-Linked Alginate as a Scaffold for Cell Culture

    OpenAIRE

    Machida-Sano, Ikuko; Ogawa, Sakito; Hirakawa, Makoto; Namiki, Hideo

    2014-01-01

    We investigated the efficacy of three-dimensional porous ferric-ion-cross-linked alginate (Fe-alginate) gels as cell scaffolds, in comparison with calcium-ion-cross-linked alginate (Ca-alginate) gels. In a previous study, we had demonstrated that two-dimensional Fe-alginate film was an efficient material for use as a scaffold, allowing good cell adhesion and proliferation, unlike Ca-alginate film. In the present study, we fabricated three-dimensional porous Fe- and Ca-alginate gels by freeze-...

  5. Deletion of the tissue response against alginate-pll capsules by temporary release of co-encapsulated steroids

    NARCIS (Netherlands)

    Bunger, CM; Tiefenbach, B; Jahnke, A; Gerlach, C; Freier, T; Schmitz, KP; Hopt, UT; Schareck, W; Klar, E; de Vos, P

    Transplantation of encapsulated living cells is a promising approach for the treatment of a wide variety of diseases. Large-scale application of the technique, however, is hampered by inflammatory responses against the capsules. In the present study, we investigate whether tissue responses against

  6. Alginate gel particles-A review of production techniques and physical properties.

    Science.gov (United States)

    Ching, Su Hung; Bansal, Nidhi; Bhandari, Bhesh

    2017-04-13

    The application of hydrocolloid gel particles is potentially useful in food, chemical, and pharmaceutical industries. Alginate gel particles are one of the more commonly used hydrocolloid gel particles due to them being biocompatible, nontoxic, biodegradable, cheap, and simple to produce. They are particularly valued for their application in encapsulation. Encapsulation in alginate gel particles confers protective benefits to cells, DNA, nutrients, and microbes. Slow release of flavors, minerals, and drugs can also be achieved by encapsulation in gel particles. The particle size and shape of the gel particles are crucial for specific applications. In this review, current methods of producing alginate gel particles will be discussed, taking into account their advantages, disadvantages, scalability, and impact on particle size. The physical properties of alginate gel particles will determine the effectiveness in different application conditions. This review will cover the current understanding of the alginate biopolymer, gelation mechanisms and factors affecting release properties, gel strength, and rheology of the alginate gel particle systems.

  7. Enrichment of cancer stem cell-like cells by culture in alginate gel beads.

    Science.gov (United States)

    Xu, Xiao-xi; Liu, Chang; Liu, Yang; Yang, Li; Li, Nan; Guo, Xin; Sun, Guang-wei; Ma, Xiao-jun

    2014-05-10

    Cancer stem cells (CSCs) are most likely the reason of cancer reoccurrence and metastasis. For further elucidation of the mechanism underlying the characteristics of CSCs, it is necessary to develop efficient culture systems to culture and expand CSCs. In this study, a three-dimensional (3D) culture system based on alginate gel (ALG) beads was reported to enrich CSCs. Two cell lines derived from different histologic origins were encapsulated in ALG beads respectively and the expansion of CSCs was investigated. Compared with two-dimensional (2D) culture, the proportion of cells with CSC-like phenotypes was significantly increased in ALG beads. Expression levels of CSC-related genes were greater in ALG beads than in 2D culture. The increase of CSC proportion after being cultured within ALG beads was further confirmed by enhanced tumorigenicity in vivo. Moreover, increased metastasis ability and higher anti-cancer drug resistance were also observed in 3D-cultured cells. Furthermore, we found that it was hypoxia, through the upregulation of hypoxia-inducible factors (HIFs) that occurred in ALG beads to induce the increasing of CSC proportion. Therefore, ALG bead was an efficient culture system for CSC enrichment, which might provide a useful platform for CSC research and promote the development of new anti-cancer therapies targeting CSCs.

  8. Tracking Hypoxic Signaling within Encapsulated Cell Aggregates

    OpenAIRE

    Skiles, Matthew L.; Sahai, Suchit; Blanchette, James O.

    2011-01-01

    In Diabetes mellitus type 1, autoimmune destruction of the pancreatic β-cells results in loss of insulin production and potentially lethal hyperglycemia. As an alternative treatment option to exogenous insulin injection, transplantation of functional pancreatic tissue has been explored1,2. This approach offers the promise of a more natural, long-term restoration of normoglycemia. Protection of the donor tissue from the host's immune system is required to prevent rejection and encapsulation is...

  9. Biocompatible coating of encapsulated cells using ionotropic gelation.

    Directory of Open Access Journals (Sweden)

    Friederike Ehrhart

    Full Text Available The technique of immunoisolated transplantation has seen in the last twenty years improvements in biocompatibility, long term stability and methods for avoidance of fibrosis in alginate capsules. However, two major problems are not yet solved: living cellular material that is not centered in the capsule is not properly protected from the hosts' immune system and the total transplant volume needs to be reduced. To solve these problems, we present a method for applying fully biocompatible alginate multilayers to a barium-alginate core without the use of polycations. We report on the factors that influence layer formation and stability and can therefore provide data for full adjustability of the additional layer. Although known for yeast and plant cells, this technique has not previously been demonstrated with mammalian cells or ultra-high viscous alginates. Viability of murine insulinoma cells was investigated by live-dead staining and live cell imaging, for murine Langerhans' islets viability and insulin secretion have been measured. No hampering effects of the second alginate layer were found. This multi-layer technique therefore has great potential for clinical and in vitro use and is likely to be central in alginate matrix based immunoisolated cell therapy.

  10. Biocompatible Coating of Encapsulated Cells Using Ionotropic Gelation

    Science.gov (United States)

    Ehrhart, Friederike; Mettler, Esther; Böse, Thomas; Weber, Matthias Max; Vásquez, Julio Alberto; Zimmermann, Heiko

    2013-01-01

    The technique of immunoisolated transplantation has seen in the last twenty years improvements in biocompatibility, long term stability and methods for avoidance of fibrosis in alginate capsules. However, two major problems are not yet solved: living cellular material that is not centered in the capsule is not properly protected from the hosts’ immune system and the total transplant volume needs to be reduced. To solve these problems, we present a method for applying fully biocompatible alginate multilayers to a barium-alginate core without the use of polycations. We report on the factors that influence layer formation and stability and can therefore provide data for full adjustability of the additional layer. Although known for yeast and plant cells, this technique has not previously been demonstrated with mammalian cells or ultra-high viscous alginates. Viability of murine insulinoma cells was investigated by live-dead staining and live cell imaging, for murine Langerhans’ islets viability and insulin secretion have been measured. No hampering effects of the second alginate layer were found. This multi-layer technique therefore has great potential for clinical and in vitro use and is likely to be central in alginate matrix based immunoisolated cell therapy. PMID:24039964

  11. Encapsulation of brewing yeast in alginate/chitosan matrix: lab-scale optimization of lager beer fermentation.

    Science.gov (United States)

    Naydenova, Vessela; Badova, Mariyana; Vassilev, Stoyan; Iliev, Vasil; Kaneva, Maria; Kostov, Georgi

    2014-03-04

    Two mathematical models were developed for studying the effect of main fermentation temperature (TMF), immobilized cell mass (MIC) and original wort extract (OE) on beer fermentation with alginate-chitosan microcapsules with a liquid core. During the experiments, the investigated parameters were varied in order to find the optimal conditions for beer fermentation with immobilized cells. The basic beer characteristics, i.e. extract, ethanol, biomass concentration, pH and colour, as well as the concentration of aldehydes and vicinal diketones, were measured. The results suggested that the process parameters represented a powerful tool in controlling the fermentation time. Subsequently, the optimized process parameters were used to produce beer in laboratory batch fermentation. The system productivity was also investigated and the data were used for the development of another mathematical model.

  12. Encapsulation of brewing yeast in alginate/chitosan matrix: lab-scale optimization of lager beer fermentation

    Science.gov (United States)

    Naydenova, Vessela; Badova, Mariyana; Vassilev, Stoyan; Iliev, Vasil; Kaneva, Maria; Kostov, Georgi

    2014-01-01

    Two mathematical models were developed for studying the effect of main fermentation temperature (T MF), immobilized cell mass (M IC) and original wort extract (OE) on beer fermentation with alginate-chitosan microcapsules with a liquid core. During the experiments, the investigated parameters were varied in order to find the optimal conditions for beer fermentation with immobilized cells. The basic beer characteristics, i.e. extract, ethanol, biomass concentration, pH and colour, as well as the concentration of aldehydes and vicinal diketones, were measured. The results suggested that the process parameters represented a powerful tool in controlling the fermentation time. Subsequently, the optimized process parameters were used to produce beer in laboratory batch fermentation. The system productivity was also investigated and the data were used for the development of another mathematical model. PMID:26019512

  13. Structural insights into alginate binding by bacterial cell-surface protein.

    Science.gov (United States)

    Temtrirath, Kanate; Murata, Kousaku; Hashimoto, Wataru

    2015-03-02

    A gram-negative Sphingomonas sp. strain A1 inducibly forms a mouth-like pit on the cell surface in the presence of alginate and directly incorporates polymers into the cytoplasm via the pit and ABC transporter. Among the bacterial proteins involved in import of alginate, a cell-surface EfeO-like Algp7 shows an ability to bind alginate, suggesting its contribution to accumulate alginate in the pit. Here, we show identification of its positively charged cluster involved in alginate binding using X-ray crystallography, docking simulation, and site-directed mutagenesis. The tertiary structure of Algp7 was determined at a high resolution (1.99Å) by molecular replacement, although no alginates were included in the structure. Thus, an in silico model of Algp7/oligoalginate was constructed by docking simulation using atomic coordinates of Algp7 and alginate oligosaccharides, where some charged residues were found to be potential candidates for alginate binding. Site-directed mutagenesis was conducted and five purified mutants K68A, K69A, E194A, N221A, and K68A/K69A were subjected to a binding assay. UV absorption difference spectroscopy along with differential scanning fluorimetry analysis indicated that K68A/K69A exhibited a significant reduction in binding affinity with alginate than wild-type Algp7. Based on these data, Lys68/Lys69 residues of Algp7 probably play an important role in binding alginate.

  14. Three-dimensional Alginate-bead Culture of Human Pituitary Adenoma Cells.

    Science.gov (United States)

    Avila-Rodríguez, Dulce; Paisano-Cerón, Karina; Valdovinos-Ramírez, Irene; Solano-Agama, Carmen; Ortiz-Plata, Alma; Mendoza-Garrido, María E

    2016-02-18

    A three-dimensional culture method is described in which primary pituitary adenoma cells are grown in alginate beads. Alginate is a polymer derived from brown sea algae. Briefly, the tumor tissue is cut into small pieces and submitted to an enzymatic digestion with collagenase and trypsin. Next, a cell suspension is obtained. The tumor cell suspension is mixed with 1.2% sodium alginate and dropped into a CaCl2 solution, and the alginate/cell suspension is gelled on contact with the CaCl2 to form spherical beads. The cells embedded in the alginate beads are supplied with nutrients provided by the culture media enriched with 20% FBS. Three-dimensional culture in alginate beads maintains the viability of adenoma cells for long periods of time, up to four months. Moreover, the cells can be liberated from the alginate by washing the beads with sodium citrate and seeded on glass coverslips for further immunocytochemical analyses. The use of a cell culture model allows for the fixation and visualization of the actin cytoskeleton with minimal disorganization. In summary, alginate beads provide a reliable culture system for the maintenance of pituitary adenoma cells.

  15. [EXPERIMENTAL STUDY ON HUMAN UMBILICAL CORD MESENCHYMAL STEM CELLS-ALGINATE WOUND DRESSING].

    Science.gov (United States)

    Wang, Song; Su, Meilan; Yang, Huachao; Long, Gang; Tang, Zhenrui; Huang, Wen

    2015-09-01

    To observe the growth characteristics of human umbilical cord mesenchymal stem cells (hUCMSCs) cultured on the alginate gel scaffolds and to explore the feasibility of hUCMSCs-alginate dressing for wound healing. hUCMSCs were separated from human umbilical cords and cultured in vitro. After the 4th passage cells were co-cultured with alginate gel (experimental group), the cell growth characteristics were observed under the inverted phase contrast microscope. Vascular endothelial growth factor (VEGF) content was measured and the number of cells was counted at 0, 3, 6, and 9 days after culture; and the cell migration capacity was observed. The hUCMSCs were cultured without alginated gel as control. The model of full-thickness skin defects was established in 32 8-week-old Balb/c male mice and they were randomly divided into 4 groups (n=8): wounds were covered with hUCMSCs-alginate gel compound (MSC-gel group), cell supernatants-alginate gel compound (CS-gel group), 10% FBS-alginate gel compound (FBS-gel group), and 0.01 mol/L PBS-alginate compound (PBS-gel group), respectively. Wound healing rates at 5, 10, and 15 days were observed and calculated; and the wound tissues were harvested for histological and immunohistochemical staining to assess new skin conditions at 15 days after operation. hUCMSCs grew well with grape-like proliferation on the alginate gel, but no cell migration was observed at 7 days after cultivation. VEGF expression and cell number in experimental group were significantly less than those in control group at 3 days (P0.05). hUCMSCs can continuously express VEGF in alginate gel, which is necessary for wound healing. The hUCMSCs-alginate compound is probably a good wound dressing.

  16. Tracking Hypoxic Signaling in Encapsulated Stem Cells

    Science.gov (United States)

    Sahai, Suchit; McFarland, Rachel; Skiles, Mathew L.; Sullivan, Denise; Williams, Amanda

    2012-01-01

    Oxygen is not only a nutrient but also an important signaling molecule whose concentration can influence the fate of stem cells. This study details the development of a marker of hypoxic signaling for use with encapsulated cells. Testing of the marker was performed with adipose-derived stem cells (ADSCs) in two-dimensional (2D) and 3D culture conditions in varied oxygen environments. The cells were genetically modified with our hypoxia marker, which produces a red fluorescent protein (DsRed-DR), under the control of a hypoxia-responsive element (HRE) trimer. For 3D culture, ADSCs were encapsulated in poly(ethylene glycol)–based hydrogels. The hypoxia marker (termed HRE DsRed-DR) is built on a recombinant adenovirus and ADSCs infected with the marker will display red fluorescence when hypoxic signaling is active. This marker was not designed to measure local oxygen concentration but rather to show how a cell perceives its local oxygen concentration. ADSCs cultured in both 2D and 3D were exposed to 20% or 1% oxygen environments for 96 h. In 2D at 20% O2, the marker signal was not observed during the study period. In 1% O2, the fluorescent signal was first observed at 24 h, with maximum prevalence observed at 96 h as 59%±3% cells expressed the marker. In 3D, the signal was observed in both 1% and 20% O2. The onset of signal in 1% O2 was observed at 4 h, reaching maximum prevalence at 96 h with 76%±4% cells expressing the marker. Interestingly, hypoxic signal was also observed in 20% O2, with 13%±3% cells showing positive marker signal after 96 h. The transcription factor subunit hypoxia inducible factor-1α was tracked in these cells over the same time period by immunostaining and western blot analysis. Immunostaining results in 2D correlated well with our marker at 72 h and 96 h, but 3D results did not correlate well. The western blotting results in 2D and 3D correlated well with the fluorescent marker. The HRE DsRed-DR virus can be used to track

  17. Polygalacturonase production by calcium alginate immobilized Enterobacter aerogenes NBO2 cells.

    Science.gov (United States)

    Darah, I; Nisha, M; Lim, Sheh-Hong

    2015-03-01

    Bacterial cells of Enterobacter aerogenes NBO2 were entrapped in calcium alginate beads in order to enhance polygalacturonase production compared to free cells. The optimized condition of 5 % (w/v) sodium alginate concentration, agitation speed of 250 rpm, and 15 beads of calcium alginate with inoculum size of 4 % (v/v; 5.4 × 10(7) cells/ml) produced 23.48 U/mL of polygalacturonase compared to free cells of 18.54 U/ml. There was about 26.6 % increment in polygalaturonase production. However, in this study, there was 296.6 % of increment in polygalacturonase production after improvement parameters compared to before improvement parameters of calcium alginate bead immobilization cells (5.92 U/ml). This research has indicated that optimized physical parameters of calcium alginate bead immobilization cells have significantly enhanced the production of polygalacturonase.

  18. Osteogenic differentiation of human mesenchymal stem cells through alginate-graft-poly(ethylene glycol) microsphere-mediated intracellular growth factor delivery.

    Science.gov (United States)

    Miao, Tianxin; Rao, Krithika S; Spees, Jeffrey L; Oldinski, Rachael A

    2014-10-28

    The intracellular delivery of growth factors increases opportunities for controlling cell behavior and maintaining tissue homeostasis. Recently, VEGFA was reported to enhance osteogenic differentiation of mesenchymal stem cells (MSCs) through an intracrine mechanism, suggesting a new strategy to promote bone tissue formation in osteoporotic patients. The goal of this study was to design and fabricate ligand-conjugated alginate-graft-poly(ethylene glycol) microspheres for intracellular delivery and release of VEGFA in primary human MSCs to enhance osteogenic differentiation as a potential therapeutic. Three types of microspheres were synthesized and characterized by scanning electron microscopy, in vitro drug release kinetics, MSC uptake and internalization: alginate alone (Alg), alginate-graft-poly(ethylene glycol) (Alg-g-PEG) and alginate-graft-poly(ethylene glycol)-S-S-arginine-glycine-aspartic acid (Alg-g-RGD). Each of the different microsphere formulations successfully transported bioactive VEGFA into primary human MSCs within 48h of culture, and significantly enhanced osteogenic differentiation compared to control treatments with empty microspheres (intracellular control) or non-encapsulated VEGFA (extracellular control). Adipogenic differentiation was not affected by the presence of VEGFA intracellularly or extracellularly. These results demonstrating the internalization of alginate-based microspheres and intracellular delivery of VEGFA support the efficacy of using this drug delivery and intracrine mechanism to control the fate of human MSCs and enhance osteogenic differentiation. Copyright © 2014 Elsevier B.V. All rights reserved.

  19. Stabilization and encapsulation of photosensitive resveratrol within yeast cell.

    Science.gov (United States)

    Shi, Guorong; Rao, Liqun; Yu, Huazhong; Xiang, Hua; Yang, Hua; Ji, Runa

    2008-02-12

    The photosensitive resveratrol was successfully encapsulated in yeast cells for the first time, as characterized by FT-IR spectra, fluorescence and confocal micrographs of the yeast cells, resveratrol and microcapsules. The release characteristic of the obtained yeast-encapsulated resveratrol in simulated gastric fluid was evaluated, and its storage stability as a powder was investigated at 25 degrees C/75% relative humidity (RH), 25 degrees C/90% RH and 60 degrees C under the laboratory fluorescent lighting conditions (ca. 300 lx) or in the dark. Also, the scavenging capacity of yeast-encapsulated resveratrol on DPPH radical was compared with that of non-encapsulated resveratrol. It could be demonstrated clearly that no chemical changes occurred during the encapsulation. Besides, the DPPH radical-scavenging activity increased after the encapsulation. In addition, the yeast-encapsulated resveratrol exhibited good stability, and its bioavailability was enhanced as a result of increased solubility of resveratrol and sustained releasing.

  20. Predicting the survival rate of mouse embryonic stem cells cryopreserved in alginate beads.

    Science.gov (United States)

    Sambu, S; Xu, X; Ye, H; Cui, Z F

    2011-11-01

    Stem cell cryopreservation in three-dimensional (3D) scaffolds may offer better protection to cells leading to higher survival rates. However, it introduces heterogeneity in cryoprotective agent (CPA) concentrations, durations of exposure to CPA, and freezing and thawing rate within constructs. This paper applies a mathematical model which couples the mass transport of dimethyl sulphoxide (DMSO) in a cell-seeded spherical construct and cell membrane transport into mouse embryonic stem cells (mESCs) to predict overall cell survival rate (CSR) based on CPA equilibrium exposure times (t(E)) and concentrations. The effect of freeze-concentration is also considered. To enable such a prediction, a contour plot was constructed using experimental data obtained in cryopreservation of cell suspensions with DMSO at a cooling rate of 1 degrees C/min. Thereafter, the diffusion in the alginate bead and the membrane transport of CPA was numerically simulated. Results were mapped onto the survival rate contours yielding 'predicted' CSR. The effects of loading time, hindrance, construct radius, and CPA concentration on predicted CSR were examined. From these results, an operation window with upper and lower t(E) of 12-19 min (for 0.6 mm radius beads and 1.4 M DMSO) yielded an overall viability of 60 per cent. The model predictions and the best experimental cryopreservation results with encapsulated mESCs were in agreement. Hence, optimization based on post-thaw CSR can accelerate the identification of cryopreservation protocols and parameters for maximizing cell survival.

  1. Mechanically reinforced cell-laden scaffolds formed using alginate-based bioink printed onto the surface of a PCL/alginate mesh structure for regeneration of hard tissue.

    Science.gov (United States)

    Kim, Yong Bok; Lee, Hyeongjin; Yang, Gi-Hoon; Choi, Chang Hyun; Lee, DaeWeon; Hwang, Heon; Jung, Won-Kyo; Yoon, Hyeon; Kim, Geun Hyung

    2016-01-01

    Cell-printing technology has provided a new paradigm for biofabrication, with potential to overcome several shortcomings of conventional scaffold-based tissue regeneration strategies via controlled delivery of various cell types in well-defined target regions. Here we describe a cell-printing method to obtain mechanically reinforced multi-layered cell-embedded scaffolds, formed of micron-scale poly(ε-caprolactone) (PCL)/alginate struts coated with alginate-based bioink. To compare the physical and cellular activities, we used a scaffold composed of pure alginate (without cells) coated PCL/alginate struts as a control. We systematically varied the ratio of alginate cross-linking agent, and determined the optimal cell-coating conditions to form the PCL/alginate struts. Following fabrication of the cell (MG63)-laden PCL/alginate scaffold, the bioactivity was evaluated in vitro. The laden cells exhibited a substantially more developed cytoskeleton compared with those on a control scaffold consisting of the same material composition. Based on these results, the printed cells exhibited a significantly more homogenous distribution within the scaffold compared with the control. Cell proliferation was determined via MTT assays at 1, 3, 7, and 14 days of culture, and the proliferation of the cell-printed scaffold was substantially in excess (∼2.4-fold) of that on the control. Furthermore, the osteogenic activity such as ALP was measured, and the cell-laden scaffold exhibited significantly greater activity (∼3.2-fold) compared with the control scaffold.

  2. Alginate gelation-induced cell death during laser-assisted cell printing.

    Science.gov (United States)

    Gudapati, Hemanth; Yan, Jingyuan; Huang, Yong; Chrisey, Douglas B

    2014-09-01

    Modified laser-induced forward transfer has emerged as a promising bioprinting technique. Depending on the operating conditions and cell properties, laser cell printing may cause cell injury and even death, which should be carefully elucidated for it to be a viable technology. This study has investigated the effects of alginate gelation, gelation time, alginate concentration, and laser fluence on the post-transfer cell viability of NIH 3T3 fibroblasts. Sodium alginate and calcium chloride are used as the gel precursor and gel reactant solution to form cell-laden alginate microspheres. It is found that the effects of gelation depend on the duration of gelation. Two-minute gelation is observed to increase the cell viability after 24 h incubation, mainly due to the protective cushion effect of the forming gel membrane during droplet landing. Despite the cushion effect from 10 min gelation, it is observed that the cell viability decreases after 24 h incubation because of the forming thick gel membrane that reduces nutrient and oxygen diffusion from the culture medium. In addition, the cell viability after 24 h incubation decreases as the laser fluence or alginate concentration increases.

  3. Synthesis of multilayered alginate microcapsules for the sustained release of fibroblast growth factor-1

    Science.gov (United States)

    Khanna, Omaditya; Moya, Monica L; Opara, Emmanuel C; Brey, Eric M

    2010-01-01

    Alginate microcapsules coated with a permselective poly-L-ornithine (PLO) membrane have been investigated for the encapsulation and transplantation of islets as a treatment for type 1 diabetes. The therapeutic potential of this approach could be improved through local stimulation of microvascular networks in order to meet mass transport demands of the encapsulated cells. Fibroblast growth factor-1 (FGF-1) is a potent angiogenic factor with optimal effect occurring when it is delivered in a sustained manner. In this paper, a technique is described for the generation of multilayered alginate microcapsules with an outer alginate layer that can be used for the delivery of FGF-1. The influence of alginate concentration and composition (high mannuronic acid (M) or guluronic acid (G) content) on outer layer size and stability, protein encapsulation efficiency, and release kinetics was investigated. The technique results in a stable outer layer of alginate with a mean thickness between 113–164 µm, increasing with alginate concentration and G-content. The outer layer was able to encapsulate and release FGF-1 for up to thirty days, with 1.25% of high G alginate displaying the most sustained release. The released FGF-1 retained its biologic activity in the presence of heparin, and the addition of the outer layer did not alter the permselectivity of the PLO coat. This technique could be used to generate encapsulation systems that deliver proteins to stimulate local neovascularization around encapsulated islets. PMID:20725969

  4. Improving of catalase stability properties by encapsulation in alginate/Fe3O4 magnetic composite beads for enzymatic removal of H2O2.

    Science.gov (United States)

    Doğaç, Yasemin Ispirli; Çinar, Mürvet; Teke, Mustafa

    2015-01-01

    The aim of this study was enhancing of stability properties of catalase enzyme by encapsulation in alginate/nanomagnetic beads. Amounts of carrier (10-100 mg) and enzyme concentrations (0.25-1.5 mg/mL) were analyzed to optimize immobilization conditions. Also, the optimum temperature (25-50°C), optimum pH (3.0-8.0), kinetic parameters, thermal stability (20-70°C), pH stability (4.0-9.0) operational stability (0-390 min), and reusability were investigated for characterization of the immobilized catalase system. The optimum pH levels of both free and immobilized catalase were 7.0. At the thermal stability studies, the magnetic catalase beads protected 90% activity, while free catalase maintained only 10% activity at 70°C. The thermal profile of magnetic catalase beads was spread over a large area. Similarly, this system indicated the improving of the pH stability. The reusability, which is especially important for industrial applications, was also determined. Thus, the activity analysis was done 50 times in succession. Catalase encapsulated magnetic alginate beads protected 83% activity after 50 cycles.

  5. Biocompatibility of mannuronic acid-rich alginates.

    Science.gov (United States)

    Klöck, G; Pfeffermann, A; Ryser, C; Gröhn, P; Kuttler, B; Hahn, H J; Zimmermann, U

    1997-05-01

    Highly purified algin preparations free of adverse contaminants with endotoxins and other mitogens recently became available by a new purification process (Klöck et al., Appl. Microbiol. Biotechnol., 1994, 40, 638-643). An advantage of this purification protocol is that it can be applied to alginates with various ratios of mannuronic acid to guluronic acid. High mannuronic acid alginate capsules are of particular practical interest for cell transplantation and for biohybrid organs, because mannuronate-rich alginates are usually less viscous, allowing one to make gels with a higher alginate content. This will increase their stability and reduce the diffusion permeability and could therefore protect immobilized cells more efficiently against the host immune system. Here we report the biocompatibility of purified, mannuronic acid-rich alginate (68% mannuronate residues) in a series of in vitro, as well as in vivo, assays. In contrast to raw alginate extracts, the purified product showed no mitogenic activity towards murine lymphocytes in vitro. Its endotoxin content was reduced to the level of the solvent. Animal studies with these new, purified algin formulations revealed the absence of a mitogen-induced foreign body reaction, even when the purified material (after cross-linking with Ba2+ ions) is implanted into animal models with elevated macrophage activity (diabetes-prone BB/OK rat). Thus, alginate capsules with high mannuronic acid content become available for applications such as implantation. In addition to the utilization as implantable cell reactors in therapy and biotechnology, these purified algins have broad application potential as ocular fillings, tissue replacements, microencapsulated growth factors and/or interleukins or slow-release dosage forms of antibodies, surface coatings of sensors and other invasive medical devices, and in encapsulation of genetically engineered cells for gene therapy.

  6. Evaluation of the effect of CaCl2 and alginate concentrations and hardening time on the characteristics of Lactobacillus acidophilus loaded alginate beads using response surface analysis

    Science.gov (United States)

    Lotfipour, Farzaneh; Mirzaeei, Shahla; Maghsoodi, Maryam

    2012-01-01

    Purpose: This article describes preparation and characterization of beads of alginate containing probiotic bacteria of Lactobacillus acidophilus DMSZ20079. Methods: Fourteen formulations using different alginate (ALG) and CaCl2 concentrations as well as hardening times were prepared using extrusion technique. The prepared beads were characterized in terms of size, morphology, encapsulation efficiency and bacterial viabilities in acid (pH 1.8, 2 hours) condition. Results: The results showed that spherical beads with narrow size distribution ranging from 1.32±0.04 to 1.70±0.07 mm were achieved with encapsulation efficiency higher than 98%. Surface response analysis revealed that alginate concentration was the important factor for the size, shape and encapsulation efficiency of prepared beads. Furthermore, survived bacteria after acid exposure in all prepared beads (63-83%) were significantly higher than those of untreated cells (39%) and enhanced by increasing alginate concentration. Surface response analysis revealed that the effect of all three factors of alginate and CaCl2 concentrations as well as hardening times were significant in acid viability, however alginate concentration played the most important role according to its regression coefficient. Conclusion: Among alginate and CaCl2 concentrations as well as hardening times, alginate concentration was the most variable in the characteristics of Alginate beads. PMID:24312773

  7. Sustained and therapeutic levels of human factor IX in hemophilia B mice implanted with microcapsules: key role of encapsulated cells.

    Science.gov (United States)

    Wen, Jianping; Vargas, Andrew Gómez; Ofosu, Frederick A; Hortelano, Gonzalo

    2006-03-01

    A gene therapy delivery system based on microcapsules enclosing recombinant cells engineered to secrete a therapeutic protein was explored in this study. In order to prevent immune rejection of the delivered cells, they were enclosed in non-antigenic biocompatible alginate microcapsules prior to being implanted intraperitoneally into mice. We have shown that encapsulated C2C12 myoblasts can temporarily deliver therapeutic levels of factor IX (FIX) in mice, but the C2C12 myoblasts elicited an immune response to FIX. In this study we report the use of mouse fetal G8 myoblasts secreting hFIX in hemophilia mice. Mouse G8 myoblasts were transduced with MFG-FIX vector. A pool of recombinant G8 myoblasts secreting approximately 1500 ng hFIX/10(6) cells/24 h in vitro were enclosed in biocompatible alginate microcapsules and implanted intraperitoneally into immunocompetent C57BL/6 and hemophilic mice. Circulating levels of hFIX in treated mice reached approximately 400 ng/ml for at least 120 days (end of experiment). Interestingly, mice treated with encapsulated G8 myoblasts did not develop anti-hFIX antibodies. Activated partial thromboplastin time (APTT) of plasmas obtained from treated hemophilic mice was reduced from 107 to 82 sec on day 60 post-treatment, and whole blood clotting time (WBCT) was also corrected from 7-9 min before treatment to 3-5 min following microcapsule implantation. Further, mice were protected against bleeding following major trauma. Thus, the FIX delivery in vivo was biologically active. Our findings suggest that the type of cells encapsulated play a key role in the generation of immune responses against the transgene. Further, a judicious selection of encapsulated cells is critical for achieving sustained gene expression. Our findings support the feasibility of encapsulated G8 myoblasts as a gene therapy approach for hemophilia B.

  8. Immobilization of microalgae cells in alginate facilitates isolation of DNA and RNA.

    Science.gov (United States)

    Lopez, Blanca R; Hernandez, Juan-Pablo; Bashan, Yoav; de-Bashan, Luz E

    2017-04-01

    Isolation of nucleic acids from Chlorella is difficult, given the chemically complex nature of their cell walls and variable production of metabolites. Immobilization of microalgae in polymers adds additional difficulty. Here, we modified, amended, and standardized methods for isolation of nucleic acids and compared the yield of DNA and RNA from free-living and encapsulated microalgae C. sorokiniana. Isolation of nucleic acids from immobilized cells required two steps in dissolving the alginate matrix, releasing the cells, and mechanical disruption with glass beads. For DNA extraction, we used modified versions of a commercial kit along with the hexadecyltrimethylammonium bromide (CTAB) method. For RNA extraction, we used the commercial TRI reagent procedure and the CTAB-dithiotreitol method. Quantity and quality of nucleic acids in extracts varied with growth conditions, isolation procedures, and time of incubation of the original culture. There were consistently higher amounts of DNA and RNA in extracts from immobilized cells. Quantitatively, the modified procedure with the commercial Promega kit was the most reliable procedure for isolating DNA and a modified commercial TRI reagent procedure was the choice for isolating RNA. All four procedures eliminated proteins efficiently and had low levels of contamination from residual polysaccharides from the matrices and/or metabolites naturally produced by the microalgae. All DNA extracts under both growth conditions, time of incubation, and two isolation methods successfully amplified the 18S ribosomal RNA by PCR and quantitative reverse transcription (RT-qPCR).

  9. Effect of Eudragit S100 nanoparticles and alginate chitosan encapsulation on the viability of Lactobacillus acidophilus and Lactobacillus rhamnosus.

    Science.gov (United States)

    Ansari, Fereshteh; Pourjafar, Hadi; Jodat, Vahid; Sahebi, Javad; Ataei, Amir

    2017-12-01

    In this study, we examined a novel method of microencapsulation with calcium alginate-chitosan and Eudragit S100 nanoparticles for the improving viability of probiotic bacteria, Lactobacillus acidophilus and Lactobacillus rhamnosus. Extrusion technique was carried out in microencapsulation process. The viability of two probiotics in single coated beads (with only chitosan), double coated beads (with chitosan and Eudragit nanoparticles), and as free cells (unencapsulated) were conducted in simulated gastric juice (pH 1.55, without pepsin) followed by incubation in simulated intestinal juice (pH 7.5, with 1% bile salt). In case of single coated beads, presumably, lack of sufficient strength of chitosan under simulated gastric condition was the main reason of 4-log and 5-log reduction of the counts of the L. acidophilus and L. rhamnosus respectively. The results showed that with the second coat forming (Eudragit nanoparticles) over the first coat (chitosan), the strength of the beads and then viability rate of the bacteria were increased in comparison with the single coated beads.

  10. Encapsulated glucagon-like peptide-1-producing mesenchymal stem cells have a beneficial effect on failing pig hearts

    DEFF Research Database (Denmark)

    Wright, Elizabeth J; Farrell, Kelly A; Malik, Nadim;

    2012-01-01

    -life in vivo. The effects of prolonged GLP-1 delivery from stromal cells post-MI were evaluated in a porcine model. Human mesenchymal stem cells immortalized and engineered to produce a GLP-1 fusion protein were encapsulated in alginate (bead-GLP-1 MSC) and delivered to coronary artery branches. Control groups...... and showed decreased infarction area compared with controls. Histological analysis showed reduced inflammation and a trend toward reduced apoptosis in the infarct zone. Increased collagen but fewer myofibroblasts were observed in infarcts of the bead-GLP-1 MSC and bead-MSC groups, and significantly more...

  11. PLGA/alginate composite microspheres for hydrophilic protein delivery.

    Science.gov (United States)

    Zhai, Peng; Chen, X B; Schreyer, David J

    2015-11-01

    Poly(lactic-co-glycolic acid) (PLGA) microspheres and PLGA/alginate composite microspheres were prepared by a novel double emulsion and solvent evaporation technique and loaded with bovine serum albumin (BSA) or rabbit anti-laminin antibody protein. The addition of alginate and the use of a surfactant during microsphere preparation increased the encapsulation efficiency and reduced the initial burst release of hydrophilic BSA. Confocal laser scanning microcopy (CLSM) of BSA-loaded PLGA/alginate composite microspheres showed that PLGA, alginate, and BSA were distributed throughout the depths of microspheres; no core/shell structure was observed. Scanning electron microscopy revealed that PLGA microspheres erode and degrade more quickly than PLGA/alginate composite microspheres. When loaded with anti-laminin antibody, the function of released antibody was well preserved in both PLGA and PLGA/alginate composite microspheres. The biocompatibility of PLGA and PLGA/alginate microspheres were examined using four types of cultured cell lines, representing different tissue types. Cell survival was variably affected by the inclusion of alginate in composite microspheres, possibly due to the sensitivity of different cell types to excess calcium that may be released from the calcium cross-linked alginate.

  12. Plastic encapsulated, dye sensitised photovoltaic cells

    Energy Technology Data Exchange (ETDEWEB)

    Potter, R.J.; Otley, L.C.; Durrant, J.R.; Haque, S.; Xu, C. [Imperial College of Science, Technology and Medicine, London (United Kingdom); Holmes, A.B.; Park, T.; Schulte, N. [Cambridge Univ. (United Kingdom)

    2004-07-01

    The report presents the results of a collaborative project that aimed to demonstrate the technical feasibility of a plastic-encapsulated, solid state, dye-sensitised solar cell (DSSC) with an energy conversion efficiency (ECE) of at least 3%. DSSCs offer a possible 'step change' in photovoltaic technology resulting in lower costs compared with existing technologies. The project involved a series of eight main tasks: the development of first and second generation HTM electrolytes; the development of polymer-supported electrolytes; the development of low temperature electrode coating procedures; dye development; cell assembly and testing; component integration; and overall process development. A wide range of innovative HTMs have been synthesised, including materials incorporating both hole-transporting and ion-chelating functional groups. The ruthenium-based dye, N3, remained the preferred sensitising component. The project has produced a system that can routinely achieve over 5% ECE at 0.1 Sun illumination on 1 cm{sup 2} cells using polymer-supported electrolytes.

  13. Alginic acid sodium hydrogel co-transplantation with Schwann cells for rat spinal cord repair.

    Science.gov (United States)

    Wang, Haibao; Liu, Chibo; Ma, Xueqiang

    2012-07-04

    The aim of the study was investigating the influence of Schwann cells-alginic acid sodium hydrogel co-transplantation on a rat model of spinal cord injury. Sprague-Dawley (SD) rats were randomly assigned to 4 groups: control, injury, injury with Schwann cell transplantation, and injury with Schwann cells-alginic acid sodium hydrogel co-transplantation. Gelatin sponge blocks containing a Schwann cell suspension were transplanted into the injury site in the Schwann cell group; Schwann cells seeded in alginic acid sodium hydrogel were transplanted into the injury site in the Schwann cells-alginic acid sodium hydrogel group. At 12 h, 1, 3, 7, and 21 days after surgery, animals were assessed on the Basso, Beattie and Bresnahan (BBB) locomotor rating scale and then were sacrificed. In the injury group, Bcl-2 immunoreactive cells peaked at 3 days after surgery, and the expression level returned to normal level at 14 days. In the co-transplantation group, Bcl-2 immunoreactive cells in the spinal cord-transected segments were significantly increased until 7 days (p alginic acid sodium hydrogel transplantation group than in the simple injury and Schwann cell groups (p alginic acid sodium hydrogel co-transplantation could inhibit cellular apoptosis and enhance Bcl-2 expression in the spinal cord-transected segments, and thereby promote the recovery of locomotor function after spinal cord injury, although it did not reach full rehabilitation.

  14. Poly(ethylene glycol) hydrogel microstructures encapsulating living cells

    Science.gov (United States)

    Koh, Won-Gun; Revzin, Alexander; Pishko, Michael V.

    2002-01-01

    We present an easy and effective method for the encapsulation of cells inside PEG-based hydrogel microstructures fabricated using photolithography. High-density arrays of three-dimensional microstructures were created on substrates using this method. Mammalian cells were encapsulated in cylindrical hydrogel microstructures of 600 and 50 micrometers in diameter or in cubic hydrogel structures in microfluidic channels. Reducing lateral dimension of the individual hydrogel microstructure to 50 micrometers allowed us to isolate 1-3 cells per microstructure. Viability assays demonstrated that cells remained viable inside these hydrogels after encapsulation for up to 7 days.

  15. Electrochemical Hydrogel Lithography of Calcium-Alginate Hydrogels for Cell Culture

    Directory of Open Access Journals (Sweden)

    Fumisato Ozawa

    2016-08-01

    Full Text Available Here we propose a novel electrochemical lithography methodology for fabricating calcium-alginate hydrogels having controlled shapes. We separated the chambers for Ca2+ production and gel formation with alginate with a semipermeable membrane. Ca2+ formed in the production chamber permeated through the membrane to fabricate a gel structure on the membrane in the gel formation chamber. When the calcium-alginate hydrogels were modified with collagen, HepG2 cells proliferated on the hydrogels. These results show that electrochemical hydrogel lithography is useful for cell culture.

  16. A highly sensitive cell assay for validation of purification regimes of alginates.

    Science.gov (United States)

    Leinfelder, U; Brunnenmeier, F; Cramer, H; Schiller, J; Arnold, K; Vásquez, J A; Zimmermann, U

    2003-10-01

    Among the hydrogels used for microencapsulation of cells and tissues, alginate has been and will continue to be one of the most important biomaterials. A mandatory requirement for clinical immunoisolated transplantations is the reproducible production of biocompatible alginate. As shown here for alginates extracted from freshly collected algal stipes, the current assays used for validation of the quality of the alginate are not sufficient to screen for impurities arising from spores of gram-positive bacteria (and related contaminants). To assess the quality of alginate, we have developed a cell assay based on the induction of apoptosis in Jurkat cells. This assay allows in combination with the "modified mixed lymphocyte" assay a rapid and sensitive screening for any fibrosis-inducing impurities in alginate samples (even during the purification regime) as demonstrated by transplantation experiments performed in parallel with BB rats (exhibiting an elevated macrophage activity). The results clearly demonstrate that the quality of the input algal material is of key relevance for the production of transplantation-grade alginate.

  17. Three-dimensional bioprinting of complex cell laden alginate hydrogel structures.

    Science.gov (United States)

    Tabriz, Atabak Ghanizadeh; Hermida, Miguel A; Leslie, Nicholas R; Shu, Wenmiao

    2015-12-21

    Different bioprinting techniques have been used to produce cell-laden alginate hydrogel structures, however these approaches have been limited to 2D or simple three-dimension (3D) structures. In this study, a new extrusion based bioprinting technique was developed to produce more complex alginate hydrogel structures. This was achieved by dividing the alginate hydrogel cross-linking process into three stages: primary calcium ion cross-linking for printability of the gel, secondary calcium cross-linking for rigidity of the alginate hydrogel immediately after printing and tertiary barium ion cross-linking for long-term stability of the alginate hydrogel in culture medium. Simple 3D structures including tubes were first printed to ensure the feasibility of the bioprinting technique and then complex 3D structures such as branched vascular structures were successfully printed. The static stiffness of the alginate hydrogel after printing was 20.18 ± 1.62 KPa which was rigid enough to sustain the integrity of the complex 3D alginate hydrogel structure during the printing. The addition of 60 mM barium chloride was found to significantly extend the stability of the cross-linked alginate hydrogel from 3 d to beyond 11 d without compromising the cellular viability. The results based on cell bioprinting suggested that viability of U87-MG cells was 93 ± 0.9% immediately after bioprinting and cell viability maintained above 88% ± 4.3% in the alginate hydrogel over the period of 11 d.

  18. Antifibrotic effect of rapamycin containing polyethylene glycol-coated alginate microcapsule in islet xenotransplantation.

    Science.gov (United States)

    Park, Heon-Seok; Kim, Ji-Won; Lee, Seung-Hwan; Yang, Hae Kyung; Ham, Dong-Sik; Sun, Cheng-Lin; Hong, Tae Ho; Khang, Gilson; Park, Chung-Gyu; Yoon, Kun-Ho

    2017-04-01

    Islet microencapsulation is an attractive strategy for the minimization or avoidance of life-long immunosuppression after transplantation. However, the clinical implementation of this technique is currently limited by incomplete biocompatibility. Thus, the aim of the present study was to demonstrate the improved biocompatibility of rapamycin-containing polyethylene glycol (Rapa-PEG)-coating on alginate microcapsules containing xenogeneic islets. The Rapa-PEG-coating on the alginate layer was observed using scanning electron microscopy (SEM) and the molecular cut-off weight of the microcapsules was approximately 70 kDa. The viabilities of the alginate-encapsulated and Rapa-PEG-coated alginate-encapsulated islets were lower than the viability of the naked islets just after encapsulation, but these the differences diminished over time in culture dishes. Rapa-PEG-coating on the alginate capsules effectively decreased the proliferation of macrophage cells compared to the non-coating and alginate coating of xenogeneic pancreas tissues. Glucose-stimulated insulin secretion did not significantly differ among the groups prior to transplantation. The random blood glucose levels of diabetic mice significantly improved following the transplantation of alginate-encapsulated and Rapa-PEG-coated alginate-encapsulated islets, but there were no significant differences between these two groups. However, there was a significant decrease in the number of microcapsules with fibrotic cell infiltration in the Rapa-PEG-coated alginate microcapsule group compared to the alginate microcapsule group. In conclusion, Rapa-PEG-coating might be an effective technique with which to improve the biocompatibility of microcapsules containing xenogeneic islets. Copyright © 2015 John Wiley & Sons, Ltd. Copyright © 2015 John Wiley & Sons, Ltd.

  19. The relationship between the inflammatory response and cell adhesion on alginate-chitosan-alginate microcapsules after transplantation.

    Science.gov (United States)

    Li, Shen; Zhang, Ying; Chen, Li; Li, Na; Xie, Hongguo; Guo, Xin; Zhao, Shan; Yu, Weiting; Lv, Yan; Lv, Guojun; Wu, Huijian; Ma, Xiaojun

    2015-07-01

    Cell microencapsulation technology is a potential alternative therapy, but cell overgrowth and adhesion on the microcapsules after transplantation shortens their time of therapeutic efficacy. Inflammatory cells were the main cells that adhered to the microcapsules, so understanding the body's inflammatory processes would help to better identify the mechanisms of cell adhesion to the outer surface of the microcapsules. Our study measured the inflammatory cells and the cytokines and characterized the associated changes in the alginate-chitosan-alginate (ACA) microcapsules 1, 7, 14, and 28 days after implantation in the peritoneal cavity. Then the relationship between the inflammatory response and cell adhesion on the microcapsules was evaluated by multiple regression analysis. The results showed that the microcapsules did not evoke a systemic inflammatory response, but initiated a local inflammatory response in the peritoneal cavity. Furthermore, the correlation analysis showed that the level of cell adhesion on the microcapsules was related to the number of lymphocytes and macrophages, and the amount of IL-6, IL-10, and MCP-1 in the peritoneal cavity. Our results may provide a foundation for reducing the immune response to these microcapsules, prolonging graft survival and improving the efficacy of these treatments.

  20. An effective device for generating alginate microcapsules

    Directory of Open Access Journals (Sweden)

    Tatiana A.B. Bressel

    2008-01-01

    Full Text Available An alternative approach to somatic gene therapy is to deliver the therapeutic protein by implanting genetically modified cells that could overexpress the gene of interest. Microencapsulation devices were designed to protect cells from host rejection and prevent the foreign cells from spreading while allowing protein secretion. Alginate microcapsules form a semi-permeable structure that is suitable for in vivo injection. In this study, we report an effective laboratory protocol for producing calcium alginate microcapsules, following optimization of uniformly shaped and sized particles containing viable cells. Encapsulation of baby hamster kidney (BHK cells in alginate microcapsules was performed using a simple device consisting of a cylinder of compressed air and a peristaltic pump. A cell suspension flow of 100 mL h-1 and an air jet flow of 10 L min-1 produced the best uniformity of microcapsule size and shape. Cells maintained viability in culture for 4 weeks without any signs of necrosis, and protein diffusion was observed during this period. Our results clearly demonstrated that microisolation of BHK cells in alginate using a simple assembly device could provide an environment that is capable of preserving live cells. In addition, encapsulated cells under the conditions described were able to secrete an active enzyme even after four weeks, thus becoming potentially compatible with therapeutic protein delivery.

  1. Therapeutic levels of human factor VIII in mice implanted with encapsulated cells: potential for gene therapy of haemophilia A.

    Science.gov (United States)

    García-Martín, Carmen; Chuah, Marinee K L; Van Damme, An; Robinson, Kelly E; Vanzieleghem, Beatrijs; Saint-Remy, Jean-Marie; Gallardo, Dominique; Ofosu, Frederick A; Vandendriessche, Thierry; Hortelano, Gonzalo

    2002-01-01

    A gene therapy delivery system based on microcapsules enclosing recombinant cells engineered to secrete a therapeutic protein has been evaluated. The microcapsules are implanted intraperitoneally. In order to prevent cell immune rejection, cells are enclosed in non-antigenic biocompatible alginate microcapsules prior to their implantation into mice. It has been shown that encapsulated myoblasts can deliver therapeutic levels of Factor IX (FIX) in mice. The delivery of human Factor VIII (hFVIII) in mice using microcapsules was evaluated in this study. Mouse C2C12 myoblasts and canine MDCK epithelial kidney cells were transduced with MFG-FVIII (B-domain deleted) vector. Selected recombinant clones were enclosed in alginate microcapsules. Encapsulated recombinant clones were subsequently implanted intraperitoneally into C57BL/6 and immunodeficient SCID mice. Plasma of mice receiving C2C12 and encapsulated MDCK cells had transient therapeutic levels of FVIII in immunocompetent C57BL/6 mice (up to 20% and 7% of physiological levels, respectively). In addition, FVIII delivery in SCID mice was also transient, suggesting that a non-immune mechanism must have contributed to the decline of hFVIII in plasma. Quantitative RT-PCR analysis confirmed directly that the decline of hFVIII is due to a reduction in steady-state hFVIII mRNA, consistent with transcriptional repression. Furthermore, encapsulated cells retrieved from implanted mice were viable, but secreted FVIII ex vivo at three-fold lower levels than the pre-implantation levels. In addition, antibodies to hFVIII were detected in immunocompetent C57BL/6 mice. Implantable microcapsules can deliver therapeutic levels of FVIII in mice, suggesting the potential of this gene therapy approach for haemophilia A. The findings suggest vector down-regulation in vivo. Copyright 2002 John Wiley & Sons, Ltd.

  2. Culture of soybean mesophyll protoplasts in alginate beads.

    Science.gov (United States)

    Tricoli, D M; Hein, M B; Carnes, M G

    1986-10-01

    Mesophyll protoplasts were isolated from leaves of 10 day old aseptically grown soybean seedlings, or from surface disinfested leaves of 3 week old plants grown in environmental chambers. The protoplasts were encapsulated in 2mm diameter Ca alginate beads. Immobilized protoplasts were induced to divide by culturing in shaker flasks containing an actively growing soybean cell suspension. The feeder cell suspension supported the division of protoplasts independent of the protoplast density in the Ca alginate beads. At day 18 after encapsulation, the alginate matrix was dissolved, releasing viable callus colonies. The feeder cell suspension obviated plating of protoplasts at high density which is usually required for subsequent cell division and colony development. Since the protoplasts were embedded at low density, the cell colonies were derived from single cells.

  3. Double-Network Hydrogel with Tunable Mechanical Performance and Biocompatibility for the Fabrication of Stem Cells-Encapsulated Fibers and 3D Assemble

    Science.gov (United States)

    Liang, Zhe; Liu, Chenguang; Li, Lili; Xu, Peidi; Luo, Guoan; Ding, Mingyu; Liang, Qionglin

    2016-01-01

    Fabrication of cell-encapsulated fibers could greatly contribute to tissue engineering and regenerative medicine. However, existing methods suffered from not only unavoidability of cell damaging conditions and/or sophisticated equipment, but also unavailability of proper materials to satisfy both mechanical and biological expectations. In this work, a simple method is proposed to prepare cell-encapsulated fibers with tunable mechanical strength and stretching behavior as well as diameter and microstructure. The hydrogel fibers are made from optimal combination of alginate and poly(N-iso-propylacrylamide)-poly(ethylene glycol), characteristics of double-network hydrogel, with enough stiffness and flexibility to create a variety of three dimensional structures like parallel helical and different knots without crack. Furthermore, such hydrogel fibers exhibit better compatibility as indicated by the viability, proliferation and expression of pluripotency markers of embryonic stem cells encapsulated after 4-day culture. The double-network hydrogel possesses specific quick responses to either of alginate lyase, EDTA or lower environmental temperature which facilitate the optional degradation of fibers or fibrous assemblies to release the cells encapsulated for subsequent assay or treatment. PMID:27628933

  4. Combination of maghemite and titanium oxide nanoparticles in polyvinyl alcohol-alginate encapsulated beads for cadmium ions removal

    Energy Technology Data Exchange (ETDEWEB)

    Majidnia, Zohreh; Idris, Ani [Universiti Teknologi Malaysia, johor bahru (Malaysia)

    2015-06-15

    Both maghemite (γ-Fe{sub 2}O{sub 3}) and titanium oxide (TiO{sub 2}) nanoparticles were mixed at various ratios and embedded in polyvinyl alcohol (PVA)-alginate beads. These beads were tested for photocatalytic behavior in eliminating toxic Cd(Ⅱ) from the aqueous solution. The photocatalytic experiments were performed under sunlight irradiation at various pH, initial feed concentrations and γ-Fe{sub 2}O{sub 3}: TiO{sub 2} ratios. The recycling attribute of these beads was also investigated. The results revealed that 100% of the Cd(Ⅱ) was eliminated in 150 minutes at pH 7 under sunlight. It shows that maghemite and titania PVA-alginate beads can be readily isolated from the aqueous solution after the photocatalyst process and reused for at least six times without losing their initial properties.

  5. The Influence of Dopants on the Effectiveness of Alginate Beads in Immobilized Cell Reactors.

    Science.gov (United States)

    Nordmeier, Akira; Chidambaram, Dev

    2016-04-01

    Zymomonas mobilis immobilized in doped calcium alginate (Ca-alginate) was successfully employed for the production of ethanol in an immobilized cell reactor. Polyethylene oxide and F127 dimethacrylate were evaluated as potential dopants for Ca-alginate beads to decrease lag time and increase initial ethanol yield. The influence of the type and concentration of the dopant on the effectiveness of the microbe immobilized in Ca-alginate beads to produce ethanol was studied, and results were compared to the widely used 2 % Ca-alginate with no dopants, which acted as control. Immobilized cell reactors that were operated using beads doped with 0.25 % polyethylene oxide (PEO) reached an ethanol yield of ∼70 % in 24 h, which was significantly higher than an ethanol yield of 25 % obtained for the control reactor operated using undoped Ca-alginate beads. This study shows that the use of water-soluble dopants can potentially reduce the lag phase and thus improve the initial production yield of immobilized cell reactors, likely due to an increase in porosity and diffusion rate of the doped beads.

  6. Microencapsulation in alginate and chitosan microgels to enhance viability of Bifidobacterium longum for oral delivery

    Directory of Open Access Journals (Sweden)

    Timothy W. Yeung

    2016-04-01

    Full Text Available Probiotic microorganisms are incorporated into a wide variety of foods, supplements, and pharmaceuticals to promote human health and wellness. However, maintaining bacterial cell viability during storage and gastrointestinal transit remains a challenge. Encapsulation of bifidobacteria within food-grade hydrogel particles potentially mitigates their sensitivity to environmental stresses. In this study, Bifidobacterium longum subspecies and strains were encapsulated in core-shell microgels consisting of an alginate core and a microgel shell. Encapsulated obligate anaerobes Bifidobacterium longum subsp. infantis and Bifidobacterium longum subsp. longum exhibited differences in viability in a strain-dependent manner, without a discernable relationship to subspecies lineage. This includes viability under aerobic storage conditions and modeled gastrointestinal tract conditions. Coating alginate microgels with chitosan did not improve viability compared to cells encapsulated in alginate microgels alone, suggesting that modifying the surface charge alone does not enhance delivery. Thus hydrogel beads have great potential for improving the stability and efficacy of bifidobacterial probiotics in various nutritional interventions.

  7. Microencapsulation in Alginate and Chitosan Microgels to Enhance Viability of Bifidobacterium longum for Oral Delivery

    Science.gov (United States)

    Yeung, Timothy W.; Üçok, Elif F.; Tiani, Kendra A.; McClements, David J.; Sela, David A.

    2016-01-01

    Probiotic microorganisms are incorporated into a wide variety of foods, supplements, and pharmaceuticals to promote human health and wellness. However, maintaining bacterial cell viability during storage and gastrointestinal transit remains a challenge. Encapsulation of bifidobacteria within food-grade hydrogel particles potentially mitigates their sensitivity to environmental stresses. In this study, Bifidobacterium longum subspecies and strains were encapsulated in core-shell microgels consisting of an alginate core and a microgel shell. Encapsulated obligate anaerobes Bifidobacterium longum subsp. infantis and Bifidobacterium longum subsp. longum exhibited differences in viability in a strain-dependent manner, without a discernable relationship to subspecies lineage. This includes viability under aerobic storage conditions and modeled gastrointestinal tract conditions. Coating alginate microgels with chitosan did not improve viability compared to cells encapsulated in alginate microgels alone, suggesting that modifying the surface charge alone does not enhance delivery. Thus hydrogel beads have great potential for improving the stability and efficacy of bifidobacterial probiotics in various nutritional interventions. PMID:27148184

  8. Microscale Strategies for Generating Cell-Encapsulating Hydrogels

    Directory of Open Access Journals (Sweden)

    Ali Khademhosseini

    2012-09-01

    Full Text Available Hydrogels in which cells are encapsulated are of great potential interest for tissue engineering applications. These gels provide a structure inside which cells can spread and proliferate. Such structures benefit from controlled microarchitectures that can affect the behavior of the enclosed cells. Microfabrication-based techniques are emerging as powerful approaches to generate such cell-encapsulating hydrogel structures. In this paper we introduce common hydrogels and their crosslinking methods and review the latest microscale approaches for generation of cell containing gel particles. We specifically focus on microfluidics-based methods and on techniques such as micromolding and electrospinning.

  9. Differentiation of Wharton’s jelly mesenchymal stem cells into neurons in alginate scaffold

    Institute of Scientific and Technical Information of China (English)

    Seyed Mojtaba Hosseini; Attiyeh Vasaghi; Newsha Nakhlparvar; Reza Roshanravan; Tahereh Talaei-khozani; Zahra Razi

    2015-01-01

    Alginate scaffold has been considered as an appropriate biomaterial for promoting the differ-entiation of embryonic stem cells toward neuronal cell lineage. We hypothesized that alginate scaffold is suitable for culturing Wharton’s jelly mesenchymal stem cells (WJMSCs) and can pro-mote the differentiation of WJMSCs into neuron-like cells. In this study, we cultured WJMSCs in a three-dimensional scaffold fabricated by 0.25% alginate and 50 mM CaCl2 in the presence of neurogenic medium containing 10 µM retinoic acid and 20 ng/mL basic ifbroblast growth factor. These cells were also cultured in conventional two-dimensional culture condition in the presence of neurogenic medium as controls. After 10 days, immunolfuorescence staining was performed for detectingβ-tubulin (marker for WJMSCs-differentiated neuron) and CD271 (motor neuron marker).β-Tubulin and CD271 expression levels were significantly greater in the WJMSCs cultured in the three-dimensional alginate scaffold than in the conventional two-dimensional culture condition. These findings suggest that three-dimensional alginate scaffold cell culture system can induce neuronal differentiation of WJMSCs effectively.

  10. Microfabrication of proangiogenic cell-laden alginate-g-pyrrole hydrogels.

    Science.gov (United States)

    DeVolder, Ross J; Zill, Andrew T; Jeong, Jae H; Kong, Hyunjoon

    2012-11-01

    Cells have been extensively studied for their uses in various therapies because of their capacities to produce therapeutic proteins and recreate new tissues. It has often been suggested that the efficacy of cell therapies can greatly be improved through the ability to localize and regulate cellular activities at a transplantation site; however, the technologies for this control are lacking. Therefore, this study reports a cell-Laden hydrogel patch engineered to support the proliferation and angiogenic growth factor expression of cells adhered to their surfaces, and to further promote neovascularization. Hydrogels consisting of alginate chemically linked with pyrrole units, termed alginate-g-pyrrole, were prepared through an oxidative cross-linking reaction between pyrrole units. Fibroblasts adhered to the alginate-g-pyrrole hydrogels, and exhibited increased proliferation and overall vascular endothelial growth factor (VEGF) expression, compared to those on pyrrole-free hydrogels. Furthermore, the alginate-g-pyrrole hydrogel surfaces were modified to present microposts, subsequently increasing the amount of pyrrole units on their surfaces. Cells adhered to the microfabricated gel surfaces exhibited increased proliferation and overall VEGF expression proportional to the density of the microposts. The resulting micropatterned alginate-g-pyrrole hydrogels exhibited increases in the size and density of mature blood vessels when implanted on chick chorioallantoic membranes (CAMs). The hydrogel system developed in this study will be broadly useful for improving the efficacy of a wide array of cell-based wound healing and tissue regenerative therapies.

  11. In situ gelation for cell immobilization and culture in alginate foam scaffolds.

    Science.gov (United States)

    Andersen, Therese; Markussen, Christine; Dornish, Michael; Heier-Baardson, Helene; Melvik, Jan Egil; Alsberg, Eben; Christensen, Bjørn E

    2014-02-01

    Essential cellular functions are often lost under culture in traditional two-dimensional (2D) systems. Therefore, biologically more realistic three-dimensional (3D) cell culture systems are needed that provide mechanical and biochemical cues which may otherwise be unavailable in 2D. For the present study, an alginate-based hydrogel system was used in which cells in an alginate solution were seeded onto dried alginate foams. A uniform distribution of NIH:3T3 and NHIK 3025 cells entrapped within the foam was achieved by in situ gelation induced by calcium ions integrated in the foam. The seeding efficiency of the cells was about 100% for cells added in a seeding solution containing 0.1-1.0% alginate compared with 18% when seeded without alginate. The NHIK 3025 cells were allowed to proliferate and form multi-cellular structures inside the transparent gel that were later vital stained and evaluated by confocal microscopy. Gels were de-gelled at different time points to isolate the multi-cellular structures and to determine the spheroid growth rate. It was also demonstrated that the mechanical properties of the gel could largely be varied through selection of type and concentration of the applied alginate and by immersing the already gelled disks in solutions providing additional gel-forming ions. Cells can efficiently be incorporated into the gel, and single cells and multi-cellular structures that may be formed inside can be retrieved without influencing cell viability or contaminating the sample with enzymes. The data show that the current system may overcome some limitations of current 3D scaffolds such as cell retrieval and in situ cell staining and imaging.

  12. In Situ Gelation for Cell Immobilization and Culture in Alginate Foam Scaffolds

    Science.gov (United States)

    Markussen, Christine; Dornish, Michael; Heier-Baardson, Helene; Melvik, Jan Egil; Alsberg, Eben; Christensen, Bjørn E.

    2014-01-01

    Essential cellular functions are often lost under culture in traditional two-dimensional (2D) systems. Therefore, biologically more realistic three-dimensional (3D) cell culture systems are needed that provide mechanical and biochemical cues which may otherwise be unavailable in 2D. For the present study, an alginate-based hydrogel system was used in which cells in an alginate solution were seeded onto dried alginate foams. A uniform distribution of NIH:3T3 and NHIK 3025 cells entrapped within the foam was achieved by in situ gelation induced by calcium ions integrated in the foam. The seeding efficiency of the cells was about 100% for cells added in a seeding solution containing 0.1–1.0% alginate compared with 18% when seeded without alginate. The NHIK 3025 cells were allowed to proliferate and form multi-cellular structures inside the transparent gel that were later vital stained and evaluated by confocal microcopy. Gels were de-gelled at different time points to isolate the multi-cellular structures and to determine the spheroid growth rate. It was also demonstrated that the mechanical properties of the gel could largely be varied through selection of type and concentration of the applied alginate and by immersing the already gelled disks in solutions providing additional gel-forming ions. Cells can efficiently be incorporated into the gel, and single cells and multi-cellular structures that may be formed inside can be retrieved without influencing cell viability or contaminating the sample with enzymes. The data show that the current system may overcome some limitations of current 3D scaffolds such as cell retrieval and in situ cell staining and imaging. PMID:24125496

  13. MTS colorimetric assay in combination with a live-dead assay for testing encapsulated L929 fibroblasts in alginate poly-L-lysine microcapsules in vitro

    NARCIS (Netherlands)

    Bunger, CM; Jahnke, A; Stange, J; de Vos, P; Hopt, UT

    Biomaterials such as applied in microcapsules may have harmful effects on encapsulated cells. Up to now, there are no adequate assays available for testing the function and viability of cells in capsules. Therefore, we investigated whether the combination of MTS proliferation assay and live-dead

  14. MTS colorimetric assay in combination with a live-dead assay for testing encapsulated L929 fibroblasts in alginate poly-L-lysine microcapsules in vitro

    NARCIS (Netherlands)

    Bunger, CM; Jahnke, A; Stange, J; de Vos, P; Hopt, UT

    2002-01-01

    Biomaterials such as applied in microcapsules may have harmful effects on encapsulated cells. Up to now, there are no adequate assays available for testing the function and viability of cells in capsules. Therefore, we investigated whether the combination of MTS proliferation assay and live-dead via

  15. Design and performance of a sericin-alginate interpenetrating network hydrogel for cell and drug delivery

    Science.gov (United States)

    Zhang, Yeshun; Liu, Jia; Huang, Lei; Wang, Zheng; Wang, Lin

    2015-07-01

    Although alginate hydrogels have been extensively studied for tissue engineering applications, their utilization is limited by poor mechanical strength, rapid drug release, and a lack of cell adhesive ability. Aiming to improve these properties, we employ the interpenetrating hydrogel design rationale. Using alginate and sericin (a natural protein with many unique properties and a major component of silkworm silk), we develop an interpenetrating polymer network (IPN) hydrogel comprising interwoven sericin and alginate double networks. By adjusting the sericin-to-alginate ratios, IPNs’ mechanical strength can be adjusted to meet stiffness requirements for various tissue repairs. The IPNs with high sericin content show increased stability during degradation, avoiding pure alginate’s early collapse. These IPNs have high swelling ratios, benefiting various applications such as drug delivery. The IPNs sustain controlled drug release with the adjustable rates. Furthermore, these IPNs are adhesive to cells, supporting cell proliferation, long-term survival and migration. Notably, the IPNs inherit sericin’s photoluminescent property, enabling bioimaging in vivo. Together, our study indicates that the sericin-alginate IPN hydrogels may serve as a versatile platform for delivering cells and drugs, and suggests that sericin may be a building block broadly applicable for generating IPN networks with other biomaterials for diverse tissue engineering applications.

  16. Microencapsulation of bull spermatozoa: Its viability in alginate-egg yolk media

    Directory of Open Access Journals (Sweden)

    Kusumaningrum DA

    2015-03-01

    Full Text Available Microencapsulation of spermatozoa is a process to entrap a number of spermatozoa in microcapsule. Alginate, as a natural polymer polysaccharide is commonly used in cell microencapsulation. Tris Yolk Citrate buffer is a good buffer for spermatozoa dilution, therefore this experiment aimed to determine optimal concentration of alginate and egg yolk to sperm quality in bull spermatozoa microencapsulation. Concentration of egg yolk and alginate in media of encapsulation were determined in applications of sperm microencapsulation. Four bulls were used as semen source and only semen with good quality were used in this study. Poolled semen was diluted using the medium to get final concentration 100 x 106 cell/ ml. The first study was conducted to determine the effect of concentration of alginate (0, 1, and 1.5% on viability of spermatozoa. The second study to determine the effect of alginate concentration, egg yolk and its interaction was done by comparing two levels of alginate (1 and 1.5% with four levels of egg yolk (5, 10, 15 and 20%. Viability of spermatozoa, motility (M, live spermatozoa (L and Intact Apical Ridge (IAR were observed at 0, 1, 2 and 3 h incubation at room temperature. Results indicated that alginate concentration increased the osmolality and viscosity but did not affect pH of the medium. The osmolality and viscosity of medium were 275, 325, 425 and 1.12, 26.62, 47.98 for concentration of alginate 0, 1 and 1.5% respectively. Percentage of motility is significantly lower (P<0.05 in alginate medium than those of control, and 1.5% alginate could produce more uniform beads. Concentration of alginate, egg yolk and its interaction did not significantly affect viability of sperm. It is concluded that the combination of 1.5% alginate with 5, 10, 15 or 20% egg yolk can be used as media for sperm encapsulation.

  17. Therapeutic cell encapsulation: ten steps towards clinical translation.

    Science.gov (United States)

    Santos, Edorta; Pedraz, José Luis; Hernández, Rosa María; Orive, Gorka

    2013-08-28

    Since the conception of cell microencapsulation, many scientists bet on this biotechnology as they saw in it a promising alternative to protect transplanted cells from host immunoresponse. Some decades later, this initial enthusiasm is giving rise to a phase of certain conformism and lack of novel advances in the field. This perspective critically discusses current challenges needed to help this approach become a realistic clinical proposal. Alginate seems to be well established as the biomaterial of choice, but additional efforts are needed regarding current cross-linkers and coatings. Biofunctionalization of the matrices may provide the necessary biomimetic microenvironment to control cell behavior. Different alginate degradation rates would allow widening the applications of this biotechnology from drug delivery to cell delivery. In this sense, stem cells from stromal tissues could be the most suitable cell source due to their intrinsic hypoimmunogenicity, their immunomodulatory effects and their capacity to cell homing. The incorporation of suicide and reporter genes in the genome of enclosed cells may overcome some of the existing biosafety concerns. Administration and extraction by means of less invasive procedures also need to be developed to succeed in clinical translation. Finally, improving cost-effectiveness for the scale-up, together with establishing and fulfilling a series of strict regulatory aspects will be indispensable to make the final step to the clinic.

  18. Synthesis of "click" alginate hydrogel capsules and comparison of their stability, water swelling, and diffusion properties with that of Ca(+2) crosslinked alginate capsules.

    Science.gov (United States)

    Breger, Joyce C; Fisher, Benjamin; Samy, Raghu; Pollack, Steven; Wang, Nam Sun; Isayeva, Irada

    2015-07-01

    Ionically crosslinked alginate hydrogels have been extensively explored for encapsulation and immunoisolation of living cells/tissues to develop implantable cell therapies, such as islet encapsulation for bioartificial pancreas. Chemical instability of these hydrogels during long-term implantation hinders the development of viable cell therapy. The exchange between divalent crosslinking ions (e.g., Ca(+2) ) with monovalent ions from physiological environment causes alginate hydrogels to degrade, resulting in exposure of the donor tissue to the host's immune system and graft failure. The goal of this study was to improve stability of alginate hydrogels by utilizing covalent "click" crosslinking while preserving other biomedically viable hydrogel properties. Alginate was first functionalized to contain either pendant alkyne or azide functionalities, and subsequently reacted via "click" chemistry to form "click" gel capsules. Alginate functionalization was confirmed by NMR and gel permeation chromatography. When compared with Ca(+2) capsules, "click" capsules exhibited superior stability in ionic media, while showing higher permeability to small size diffusants and similar molecular weight cut-off and water swelling. Physicochemical properties of "click" alginate hydrogels demonstrate their potential utility for therapeutic cell encapsulation and other biomedical applications.

  19. Polymers in cell encapsulation from an enveloped cell perspective

    NARCIS (Netherlands)

    de Vos, Paul; Lazarjani, Hamideh Aghajani; Poncelet, Denis; Faas, Marijke M.

    2014-01-01

    In the past two decades, many polymers have been proposed for producing immunoprotective capsules. Examples include the natural polymers alginate, agarose, chitosan, cellulose, collagen, and xanthan and synthetic polymers poly(ethylene glycol), polyvinyl alcohol, polyurethane, poly(ether-sulfone),

  20. Investigation of cell viability and morphology in 3D bio-printed alginate constructs with tunable stiffness.

    Science.gov (United States)

    Shi, Pujiang; Laude, Augustinus; Yeong, Wai Yee

    2017-04-01

    In this article, mouse fibroblast cells (L929) were seeded on 2%, 5%, and 10% alginate hydrogels, and they were also bio-printed with 2%, 5%, and 10% alginate solutions individually to form constructs. The elastic and viscous moduli of alginate solutions, their interior structure and stiffness, interactions of cells and alginate, cell viability, migration and morphology were investigated by rheometer, MTT assay, scanning electron microscope (SEM), and fluorescent microscopy. The three types of bio-printed scaffolds of distinctive stiffness were prepared, and the seeded cells showed robust viability either on the alginate hydrogel surfaces or in the 3D bio-printed constructs. Majority of the proliferated cells in the 3D bio-printed constructs weakly attached to the surrounding alginate matrix. The concentration of alginate solution and hydrogel stiffness influenced cell migration and morphology, moreover the cells formed spheroids in the bio-printed 10% alginate hydrogel construct. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1009-1018, 2017.

  1. Effect of alginate and chitosan on viability and release behavior of Bifidobacterium pseudocatenulatum G4 in simulated gastrointestinal fluid.

    Science.gov (United States)

    Kamalian, Nikoo; Mirhosseini, Hamed; Mustafa, Shuhaimi; Manap, Mohd Yazid Abd

    2014-10-13

    The main aim of this study was to investigate the effect of different coating materials (i.e. Na-alginate and chitosan) on the viability and release behavior of Bifidobacterium pseudocatenulatum G4 in the simulated gastric fluid (SGF) and simulated intestinal fluid (SIF). This study reports the viability of encapsulated B. pseudocatenulatum G4 coated using different alginate (2-4 g/100mL) and chitosan (0.2-0.8 g/100mL) concentrations. The results indicated that the highest concentration of alginate (4.4142 g/100mL) along with 0.5578 g/100mL chitosan resulted in the highest viability of B. pseudocatenulatum G4. The release behavior of the encapsulated probiotics in SGF (pH 1.5) in 2h followed by 4h in SIF (pH 7.4) was also assessed. The resistance rate of alginate-chitosan capsule in SGF was higher than SIF. The alginate-chitosan encapsulated cells had also more resistance than alginate capsules. The current study revealed that alginate encapsulated B. Pseudocatenulatum G4 exhibited longer survival than its free cells (control).

  2. Effect of immobilized cells in calcium alginate beads in alcoholic fermentation.

    Science.gov (United States)

    Duarte, Juliana C; Rodrigues, J Augusto R; Moran, Paulo J S; Valença, Gustavo P; Nunhez, José R

    2013-05-30

    Saccharomyces cerevisiae cells were immobilized in calcium alginate and chitosan-covered calcium alginate beads and studied in the fermentation of glucose and sucrose for ethanol production. The batch fermentations were carried out in an orbital shaker and assessed by monitoring the concentration of substrate and product with HPLC. Cell immobilization in calcium alginate beads and chitosan-covered calcium alginate beads allowed reuse of the beads in eight sequential fermentation cycles of 10 h each. The final concentration of ethanol using free cells was 40 g L-1 and the yields using glucose and sucrose as carbon sources were 78% and 74.3%, respectively. For immobilized cells in calcium alginate beads, the final ethanol concentration from glucose was 32.9 ± 1.7 g L-1 with a 64.5 ± 3.4% yield, while the final ethanol concentration from sucrose was 33.5 ± 4.6 g L-1 with a 64.5 ± 8.6% yield. For immobilized cells in chitosan-covered calcium alginate beads, the ethanol concentration from glucose was 30.7 ± 1.4 g L-1 with a 61.1 ± 2.8% yield, while the final ethanol concentration from sucrose was 31.8 ± 6.9 g L-1 with a 62.1 ± 12.8% yield. The immobilized cells allowed eight 10 h sequential reuse cycles to be carried out with stable final ethanol concentrations. In addition, there was no need to use antibiotics and no contamination was observed. After the eighth cycle, there was a significant rupture of the beads making them inappropriate for reuse.

  3. [Marrow mesenchymal stem cell transplantation with sodium alginate gel for repair of spinal cord injury in mice].

    Science.gov (United States)

    Shi, Chen-yue; Ruan, Ling-qin; Feng, Yi-hui; Fang, Jia-lin; Song, Chen-jiao; Yuan, Zhang-gen; Ding, Yue-min

    2011-07-01

    To investigate the effects of sodium alginate gels on marrow mesenchymal stem cell transplantation for repair of spinal cord injury (SCI) in mice. In the present study, effects of different sterilization methods and concentrations of sodium alginate gels were examined. Marrow mesenchymal stem cells (mMSCs) were isolated from mice and cultured. Cells were cultured with sodium alginate gels and MTT assay was applied to determine the cell viability. Mice spinal cord injury was induced by spinal cord transection. mMSCs were transplanted into the cavity of injured spinal cord with sodium alginate gels. The effects of sodium alginate gel were assessed by BMS scales and immunofluorescence staining for NF-200. Compared with liquid form, solid form sodium alginate gel prepared with high pressure vapor sterilization had a better effect on cell viability. SCI mice treated with 10 % sodium alginate gel and mMSCs achieved higher score in BMS scale as well as higher expression of NF-200 compared with the untreated SCI group. Sodium alginate gel prepared with solid form sterilization induces mMSCs proliferation and thus can be used as the carrier of stem cell in treatment of SCI.

  4. Fabrication of large size alginate beads for three-dimensional cell-cluster culture

    Science.gov (United States)

    Zhang, Zhengtao; Ruan, Meilin; Liu, Hongni; Cao, Yiping; He, Rongxiang

    2017-08-01

    We fabricated large size alginate beads using a simple microfluidic device under a co-axial injection regime. This device was made by PDMS casting with a mold formed by small diameter metal and polytetrafluorothylene tubes. Droplets of 2% sodium alginate were generated in soybean oil through the device and then cross-linked in a 2% CaCl2 solution, which was mixed tween80 with at a concentration of 0.4 to 40% (w/v). Our results showed that the morphology of the produced alginate beads strongly depends on the tween80 concentration. With the increase of concentration of tween80, the shape of the alginate beads varied from semi-spherical to tailed-spherical, due to the decrease of interface tension between oil and cross-link solution. To access the biocompatibility of the approach, MCF-7 cells were cultured with the alginate beads, showing the formation of cancer cells clusters which might be useful for future studies.

  5. Fabrication of Freestanding Alginate Microfibers and Microstructures for Tissue Engineering Applications

    Science.gov (United States)

    Szymanski, John M; Feinberg, Adam W

    2014-01-01

    Natural biopolymers such as alginate have become important materials for a variety of biotechnology applications including drug delivery, cell encapsulation and tissue engineering. This expanding use has spurred the development of new approaches to engineer these materials at the nano- and microscales to better control cell interactions. Here we describe a method to fabricate freestanding alginate-based microfibers and microstructures with tunable geometries down to approximately 3 μm. To do this, a polydimethylsiloxane (PDMS) stamp is used to micromold alginate or alginate-fibrin blends onto a sacrificial layer of thermally-sensitive poly(N-isopropylacrylamide) (PIPAAm). A warm calcium chloride solution is then used to crosslink the alginate and upon cooling below the lower critical solution temperature (~32° C) the PIPAAm layer dissolves and releases the alginate or alginate-fibrin as freestanding microfibers and microstructures. Proof-of-concept experiments demonstrate that C2C12 myoblasts seeded onto the alginate-fibrin microfibers polarize along the fiber length forming interconnected cell strands. Thus, we have developed the ability to engineer alginate-based microstructured materials that can selectively bind cells and direct cellular assembly. PMID:24695323

  6. Improved probiotic viability in stress environments with post-culture of alginate-chitosan microencapsulated low density cells.

    Science.gov (United States)

    Song, Huiyi; Yu, Weiting; Liu, Xiudong; Ma, Xiaojun

    2014-08-08

    In this study, probiotics (Saccharomyces cerevisiae Y235) were entrapped in alginate-chitosan microcapsules by emulsification/internal gelation technique. Two different encapsulation patterns were established as directly entrapped high density cells (dEHDC) and entrapped low density cells with culture (ELDCwc). The performance of microencapsulated cells, with free cells (FC) as control, was investigated against sequential stress environments of freeze-drying, storage, and simulated gastrointestinal fluids. After being freeze-dried without cryoprotectant, the survival rate of ELDCwc (14.33%) was significantly higher than 10.00% of dEHDC, and 0.05% of FC. The lower temperature (-20°C) and ELDCwc pattern were beneficial for keeping viable cells at 7.00 logCFU g(-1) after 6 months. Furthermore, the ELDCwc microcapsule maintained viable cells of 6.29 logCFU g(-1) after incubation in SGF and SIF. These studies demonstrated that the pattern of entrapped low density cells with culture was an effective and superior technique of resisting harmful stress environments. Copyright © 2014 Elsevier Ltd. All rights reserved.

  7. Growth of Chlamydomonas reinhardtii in acetate-free medium when co-cultured with alginate-encapsulated, acetate-producing strains of Synechococcus sp. PCC 7002.

    Science.gov (United States)

    Therien, Jesse B; Zadvornyy, Oleg A; Posewitz, Matthew C; Bryant, Donald A; Peters, John W

    2014-01-01

    The model alga Chlamydomonas reinhardtii requires acetate as a co-substrate for optimal production of lipids, and the addition of acetate to culture media has practical and economic implications for algal biofuel production. Here we demonstrate the growth of C. reinhardtii on acetate provided by mutant strains of the cyanobacterium Synechococcus sp. PCC 7002. Optimal growth conditions for co-cultivation of C. reinhardtii with wild-type and mutant strains of Synechococcus sp. 7002 were established. In co-culture, acetate produced by a glycogen synthase knockout mutant of Synechococcus sp. PCC 7002 was able to support the growth of a lipid-accumulating mutant strain of C. reinhardtii defective in starch production. Encapsulation of Synechococcus sp. PCC 7002 using an alginate matrix was successfully employed in co-cultures to limit growth and maintain the stability. The ability of immobilized strains of the cyanobacterium Synechococcus sp. PCC 7002 to produce acetate at a level adequate to support the growth of lipid-accumulating strains of C. reinhartdii offers a potentially practical, photosynthetic alternative to providing exogenous acetate into growth media.

  8. Comparison of the activities of various alginates to induce TNF-alpha secretion in RAW264.7 cells.

    Science.gov (United States)

    Kurachi, Maki; Nakashima, Takuji; Miyajima, Chihiro; Iwamoto, Yoshiko; Muramatsu, Tsuyoshi; Yamaguchi, Kenichi; Oda, Tatsuya

    2005-08-01

    We compared the abilities of alginate polymers having different molecular sizes and compositions to induce the secretion of tumor necrosis factor (TNF)-alpha in RAW264.7 cells. The molecular sizes and alpha-L-guluronate/beta-D-mannuronate (M/G) ratios of highly purified alginate polymers used in this study were 9000-38 000 and 1.50-3.17, respectively. Among the alginates tested, I-S, which had the highest molecular weight, showed the most potent TNF-alpha-inducing activity. The M/G ratio also seemed to influence this activity, and, among alginates with similar molecular sizes, alginates with a higher M/G ratio tended to show higher activity. Interestingly, the enzymatic depolymerization of I-S with bacterial alginate lyase resulted in a dramatic increase in the TNF-alpha-inducing activity. Such an effect of enzymatic digestion was also observed in a relatively low-molecular-weight alginate (ULV-3), which originally had very low TNF-alpha-inducing activity. Furthermore, the inhibition profiles of the TNF-alpha-inducing activity of enzymatically digested I-S shown by three specific mitogen-activated protein (MAP) kinase inhibitors differed from those of intact I-S. These results suggest that the underlying mechanism of the TNF-alpha-inducing activity of enzymatically depolymerized alginate oligomers is not necessarily the same as that of original alginate polymer.

  9. Insights in Behavior of Variably Formulated Alginate-Based Microcapsules for Cell Transplantation

    Directory of Open Access Journals (Sweden)

    Pia Montanucci

    2015-01-01

    Full Text Available Alginate-based microencapsulation of live cells may offer the opportunity to treat chronic and degenerative disorders. So far, a thorough assessment of physical-chemical behavior of alginate-based microbeads remains cloudy. A disputed issue is which divalent cation to choose for a high performing alginate gelling process. Having selected, in our system, high mannuronic (M enriched alginates, we studied different gelling cations and their combinations to determine their eventual influence on physical-chemical properties of the final microcapsules preparation, in vitro and in vivo. We have shown that used of ultrapure alginate allows for high biocompatibility of the formed microcapsules, regardless of gelation agents, while use of different gelling cations is associated with corresponding variable effects on the capsules’ basic architecture, as originally reported in this work. However, only the final application which the capsules are destined to will ultimately guide the selection of the ideal, specific gelling divalent cations, since in principle there are no capsules that are better than others.

  10. Wound Dressing Model of Human Umbilical Cord Mesenchymal Stem Cells-Alginates Complex Promotes Skin Wound Healing by Paracrine Signaling

    Directory of Open Access Journals (Sweden)

    Song Wang

    2016-01-01

    Full Text Available Purpose. To probe growth characteristics of human umbilical cord mesenchymal stem cells (hUCMSCs cultured with alginate gel scaffolds, and to explore feasibility of wound dressing model of hUCMSCs-alginates compound. Methods. hUCMSCs were isolated, cultured, and identified in vitro. Then cells were cultivated in 100 mM calcium alginate gel, and the capacity of proliferation and migration and the expression of vascular endothelial growth factors (VEGF were investigated regularly. Wound dressing model of hUCMSCs-alginate gel mix was transplanted into Balb/c mice skin defects. Wound healing rate and immunohistochemistry were examined. Results. hUCMSCs grew well but with little migration ability in the alginate gel. Compared with control group, a significantly larger cell number and more VEGF expression were shown in the gel group after culturing for 3–6 days (P < 0.05. In addition, a faster skin wound healing rate with more neovascularization was observed in the hUCMSCs-alginate gel group than in control groups at 15th day after surgery (P < 0.05. Conclusion. hUCMSCs can proliferate well and express massive VEGF in calcium alginate gel porous scaffolds. Wound dressing model of hUCMSCs-alginate gel mix can promote wound healing through paracrine signaling.

  11. Degradation of synthetic pollutants in real wastewater using laccase encapsulated in core-shell magnetic copper alginate beads.

    Science.gov (United States)

    Le, Thao Thanh; Murugesan, Kumarasamy; Lee, Chung-Seop; Vu, Chi Huong; Chang, Yoon-Seok; Jeon, Jong-Rok

    2016-09-01

    Immobilization of laccase has been highlighted to enhance their stability and reusability in bioremediation. In this study, we provide a novel immobilization technique that is very suitable to real wastewater treatment. A perfect core-shell system composing copper alginate for the immobilization of laccase (Lac-beads) was produced. Additionally, nFe2O3 was incorporated for the bead recycling through magnetic force. The beads were proven to immobilize 85.5% of total laccase treated and also to be structurally stable in water, acetate buffer, and real wastewater. To test the Lac-beads reactivity, triclosan (TCS) and Remazol Brilliant Blue R (RBBR) were employed. The Lac-beads showed a high percentage of TCS removal (89.6%) after 8h and RBBR decolonization at a range from 54.2% to 75.8% after 4h. Remarkably, the pollutants removal efficacy of the Lac-beads was significantly maintained in real wastewater with the bead recyclability, whereas that of the corresponding free laccase was severely deteriorated. Copyright © 2016 Elsevier Ltd. All rights reserved.

  12. Copper(II) complexes encapsulated in human red blood cells.

    Science.gov (United States)

    Bonomo, R P; De Flora, A; Rizzarelli, E; Santoro, A M; Tabbí, G; Tonetti, M

    1995-09-01

    Copper(II) complexes were encapsulated in human red blood cells in order to test their possible use as antioxidant drugs by virtue of their labile character. ESR spectroscopy was used to verify whether encapsulation in red blood cells leads to the modification of such complexes. With copper(II) complexes bound to dipeptides or tripeptides, an interaction with hemoglobin was found to be present, the hemoglobin having a strong coordinative site formed by four nitrogen donor atoms. Instead, with copper(II) complexes with TAD or PheANN3, which have the greatest stability. ESR spectra always showed the original species. Only the copper(II) complex with GHL gave rise to a complicated behavior, which contained signals from iron(III) species probably coming from oxidative processes. Encapsulation of all copper(II) complexes in erythrocytes caused a slight oxidative stress, compared to the unloaded and to the native cells. However, no significant differences were observed in the major metabolic properties (GSH, glycolytic rate, hexose monophosphate shunt, Ca(2+)-ATPase) of erythrocytes loaded with different copper(II) complexes, with the exception of methemoglobin levels, which were markedly increased in the case of [Cu(GHL)H-1] compared to [Cu(TAD)]. This latter finding suggests that methemoglobin formation can be affected by the type of complex used for encapsulation, depending on the direct interaction of the copper(II) complex with hemoglobin.

  13. Factors influencing the mechanical stability of alginate beads applicable for immunoisolation of mammalian cells

    NARCIS (Netherlands)

    Bhujbal, Swapnil V.; Paredes, Genaro A.; Niclou, Simone P.; de Vos, Paul

    2014-01-01

    Transplantation of microencapsulated cells has been proposed as a cure for many types of endocrine disorders. Alginate-based microcapsules have been used in many of the feasibility studied addressing cure of the endocrine disorders, and different cancer types. Despite years of intensive research it

  14. Factors influencing the mechanical stability of alginate beads applicable for immunoisolation of mammalian cells

    NARCIS (Netherlands)

    Bhujbal, Swapnil V.; Paredes, Genaro A.; Niclou, Simone P.; de Vos, Paul

    Transplantation of microencapsulated cells has been proposed as a cure for many types of endocrine disorders. Alginate-based microcapsules have been used in many of the feasibility studied addressing cure of the endocrine disorders, and different cancer types. Despite years of intensive research it

  15. Fabrication and optimization of alginate hydrogel constructs for use in 3D neural cell culture

    Energy Technology Data Exchange (ETDEWEB)

    Frampton, J P; Hynd, M R; Shain, W [Department of Biomedical Sciences, School of Public Health, State University of New York at Albany, Albany, NY 12210 (United States); Shuler, M L, E-mail: jf7674@albany.edu [Department of Biomedical Engineering, 270 Olin Hall, Cornell University, Ithaca, NY 14850 (United States)

    2011-02-15

    Two-dimensional (2D) culture systems provide useful information about many biological processes. However, some applications including tissue engineering, drug transport studies, and analysis of cell growth and dynamics are better studied using three-dimensional (3D) culture systems. 3D culture systems can potentially offer higher degrees of organization and control of cell growth environments, more physiologically relevant diffusion characteristics, and permit the formation of more extensive 3D networks of cell-cell interactions. A 3D culture system has been developed using alginate as a cell scaffold, capable of maintaining the viability and function of a variety of neural cell types. Alginate was functionalized by the covalent attachment of a variety of whole proteins and peptide epitopes selected to provide sites for cell attachment. Alginate constructs were used to entrap a variety of neural cell types including astroglioma cells, astrocytes, microglia and neurons. Neural cells displayed process outgrowth over time in culture. Cell-seeded scaffolds were characterized in terms of their biochemical and biomechanical properties, effects on seeded neural cells, and suitability for use as 3D neural cell culture models.

  16. TRANSPLANTATION OF CRYOPRESERVED FETAL LIVER CELLS SEEDED INTO MACROPOROUS ALGINATE-GELATIN SCAFFOLDS IN RATS WITH LIVER FAILURE

    Directory of Open Access Journals (Sweden)

    D. V. Grizay

    2015-01-01

    Full Text Available Aim. To study the therapeutic potential of cryopreserved fetal liver cells seeded into macroporous alginategelatin scaffolds after implantation to omentum of rats with hepatic failure.Materials and methods.Hepatic failure was simulated by administration of 2-acetyl aminofl uorene followed partial hepatectomy. Macroporous alginate-gelatin scaffolds, seeded with allogenic cryopreserved fetal liver cells (FLCs were implanted into rat omentum. To prevent from colonization of host cells scaffolds were coated with alginate gel shell. Serum transaminase activity, levels of albumin and bilirubin as markers of hepatic function were determined during 4 weeks after failure model formation and scaffold implantation. Morphology of liver and scaffolds after implantation were examined histologically. Results. Macroporous alginate-gelatin scaffolds after implantation to healthy rats were colonized by host cells. Additional formation of alginate gel shell around scaffolds prevented the colonization. Implantation of macroporous scaffolds seeded with cryopreserved rat FLCs and additionally coated with alginate gel shell into omentum of rats with hepatic failure resulted in signifi cant improvement of hepatospecifi c parameters of the blood serum and positive changes of liver morphology. The presence of cells with their extracellular matrix within the scaffolds was confi rmed after 4 weeks post implantation.Conclusion. The data above indicate that macroporous alginate-gelatin scaffolds coated with alginate gel shell are promising cell carriers for the development of bioengineered liver equivalents.

  17. Salinomycin encapsulated nanoparticles as a targeting vehicle for glioblastoma cells.

    Science.gov (United States)

    Tığlı Aydın, R Seda; Kaynak, Gökçe; Gümüşderelioğlu, Menemşe

    2016-02-01

    Salinomycin has been introduced as a novel alternative to traditional anti-cancer drugs. The aim of this study was to test a strategy designed to deliver salinomycin to glioblastoma cells in vitro. Salinomycin-encapsulated polysorbate 80-coated poly(lactic-co-glycolic acid) nanoparticles (P80-SAL-PLGA) were prepared and characterized with respect to particle size, morphology, thermal properties, drug encapsulation efficiency and controlled salinomycin-release behaviour. The in vitro cellular uptake of P80-SAL-PLGA (5 and 10 µM) or uncoated nanoparticles was assessed in T98G human glioblastoma cells, and the cell viability was investigated with respect to anti-growth activities. SAL, which was successfully transported to T98G glioblastoma cells via P80 coated nanoparticles (∼14% within 60 min), greatly decreased (p salinomycin delivery system in the treatment of human glioblastoma.

  18. Biological activities of alginate.

    Science.gov (United States)

    Ueno, Mikinori; Oda, Tatsuya

    2014-01-01

    To gain insight into the structure-activity relationship of alginate, we examined the effect of alginates with varying molecular weights and M/G ratio on murine macrophage cell line, RAW264.7 cells in terms of induction of tumor necrosis factor-α (TNF-α) secretion. Among the alginates tested, alginate with the highest molecular weight (MW 38,000, M/G 2.24) showed the most potent TNF-α-inducing activity. Alginates having higher M/G ratio tended to show higher activity. These results suggest that molecular size and M/G ratio are important structural parameters influencing the TNF-α-inducing activity. Interestingly, enzymatic depolymerization of alginate with bacterial alginate lyase resulted in dramatic increase in the TNF-α-inducing activity. The higher activity of enzymatically digested alginate oligomers to induce nitric oxide production from RAW264.7 cells than alginate polymer was also observed. On the other hand, alginate polymer and oligomer showed nearly equal hydroxyl radical scavenging activities.

  19. A Transient Cell-shielding Method for Viable MSC Delivery Within Hydrophobic Scaffolds Polymerized in situ

    Science.gov (United States)

    2015-03-27

    macropores and cells within PUR scaffolds (Fig. 1B). Through encapsulation within partially oxidized sodium alginate (o-Alg) beads, cells were...2.1. Materials The sodium salt of alginic acid (Alg, viscosity ¼ 20e40 cPs) was supplied by Sigma Aldrich (St. Louis, MO). Acros Organics supplied...Fig. S1) to confirm their pluripotency. 2.3. Preparation of partially oxidized alginate Partially oxidized sodium alginate (o-Alg) degrades

  20. Investigating metal removal potential by Effective Microorganisms (EM) in alginate-immobilized and free-cell forms.

    Science.gov (United States)

    Ting, Adeline Su Yien; Rahman, Nurul Hidayah Abdul; Isa, Mohamed Ikmal Hafiz Mahamad; Tan, Wei Shang

    2013-11-01

    Metal removal potential of both alginate-immobilized and free-cells of Effective Microorganisms (EM-1™ Inoculant) was investigated in this study. Results revealed that removal of Cr(III), Cu(II) and Pb(II) followed a similar trend where alginate-immobilized EM were more efficient compared to free-cells of EM. For these metals, 0.940, 2.695 and 4.011 mg g(-1) of Cr(III), Cu(II) and Pb(II) were removed compared to only 0.160, 0.859 and 0.755 mg ml(-1) removed by free-cells, respectively. The higher efficiency of alginate-immobilized EM was primarily attributed to the alginate matrix. This was evident when both alginate-immobilized EM and plain alginate beads (without EM), were not significantly different in their removal efficacies. Presence of alginate also enhanced the use of the biosorbents as maximum metal sorption was achieved after 120 min as opposed to only 60 min for free-cells. EM per se in immobilized or free-cell forms did not enhance metal removal efficacy.

  1. Reduced liver cell death using an alginate scaffold bandage: a novel approach for liver reconstruction after extended partial hepatectomy.

    Science.gov (United States)

    Shteyer, Eyal; Ben Ya'acov, Ami; Zolotaryova, Lidia; Sinai, Avital; Lichtenstein, Yoav; Pappo, Orit; Kryukov, Olga; Elkayam, Tsiona; Cohen, Smadar; Ilan, Yaron

    2014-07-01

    Extended partial hepatectomy may be needed in cases of large hepatic mass, and can lead to fulminant hepatic failure. Macroporous alginate scaffold is a biocompatible matrix which promotes the growth, differentiation and long-term hepatocellular function of primary hepatocytes in vitro. Our aim was to explore the ability of implanted macroporous alginate scaffolds to protect liver remnants from acute hepatic failure after extended partial hepatectomy. An 87% partial hepatectomy (PH) was performed on C57BL/6 mice to compare non-treated mice to mice in which alginate or collagen scaffolds were implanted after PH. Mice were scarified 3, 6, 24 and 48 h and 6 days following scaffold implantation and the extent of liver injury and repair was examined. Alginate scaffolds significantly increased animal survival to 60% vs. 10% in non-treated and collagen-treated mice (log rank=0.001). Mice with implanted alginate scaffolds manifested normal and prolonged aspartate aminotransferases and alanine aminotransferases serum levels as compared with the 2- to 20-fold increase in control groups (Palginate-scaffold-treated mice 48 h after hepatectomy. Incorporation of BrdU-positive cells was 30% higher in the alginate-scaffold-treated group, compared with non-treated mice. Serum IL-6 levels were significantly decreased 3h post PH. Biotin-alginate scaffolds were quickly well integrated within the liver tissue. Collectively, implanted alginate scaffolds support liver remnants after extended partial hepatectomy, thus eliminating liver injury and leading to enhanced animal survival after extended partial hepatectomy.

  2. Terrestrial applications of FEP-encapsulated solar cell modules. [power systems using Fluorinated Ethylene Propylene encapsulation

    Science.gov (United States)

    Forestieri, A. F.; Ratajczak, A. F.

    1974-01-01

    The NASA-Lewis Research Center program of transferring the FEP-encapsulated solar cell technology developed for the space program to terrestrial applications is presented. The electrical power system design and the array mechanical design are described, and power systems being tested are discussed. The latter are located at NOAA-RAMOS weather stations at Sterling, Va., and Mammoth Mountain, Calif.; on the roof of the Lewis Research Center; on a NOAA-Coast Guard buoy in the Gulf of Mexico; in a U.S. Forest Service mountaintop voice repeater station in the Inyo National Forest, Calif., and in a backpack charger for portable transmitter/receivers being used in the same place. Preliminary results of testing are still incomplete, but show that rime ice can cause cracks in modular cells without damaging the FEP though, which keeps the grid lines intact, and that electrically active elements of the module must be completely sealed from salt water to prevent FEP delamination.

  3. On-chip Magnetic Separation and Cell Encapsulation in Droplets

    Science.gov (United States)

    Chen, A.; Byvank, T.; Bharde, A.; Miller, B. L.; Chalmers, J. J.; Sooryakumar, R.; Chang, W.-J.; Bashir, R.

    2012-02-01

    The demand for high-throughput single cell assays is gaining importance because of the heterogeneity of many cell suspensions, even after significant initial sorting. These suspensions may display cell-to-cell variability at the gene expression level that could impact single cell functional genomics, cancer, stem-cell research and drug screening. The on-chip monitoring of individual cells in an isolated environment could prevent cross-contamination, provide high recovery yield and ability to study biological traits at a single cell level These advantages of on-chip biological experiments contrast to conventional methods, which require bulk samples that provide only averaged information on cell metabolism. We report on a device that integrates microfluidic technology with a magnetic tweezers array to combine the functionality of separation and encapsulation of objects such as immunomagnetically labeled cells or magnetic beads into pico-liter droplets on the same chip. The ability to control the separation throughput that is independent of the hydrodynamic droplet generation rate allows the encapsulation efficiency to be optimized. The device can potentially be integrated with on-chip labeling and/or bio-detection to become a powerful single-cell analysis device.

  4. Development of a cell culture surface conversion technique using alginate thin film for evaluating effect upon cellular differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Nakashima, Y., E-mail: yuta-n@mech.kumamoto-u.ac.jp [Graduate School of Science and Technology, Kumamoto University, 2-39-1 Kurokami, Chuo-ku, Kumamoto 096-8555 (Japan); Graduate School of Science and Engineering, Yamaguchi University, 2-16-1 Tokiwadai, Ube 755-8611 (Japan); Tsusu, K.; Minami, K. [Graduate School of Science and Engineering, Yamaguchi University, 2-16-1 Tokiwadai, Ube 755-8611 (Japan); Nakanishi, Y. [Graduate School of Science and Technology, Kumamoto University, 2-39-1 Kurokami, Chuo-ku, Kumamoto 096-8555 (Japan)

    2014-06-15

    Here, we sought to develop a cell culture surface conversion technique that would not damage living cells. An alginate thin film, formed on a glass plate by spin coating of sodium alginate solution and dipping into calcium chloride solution, was used to inhibit adhesion of cells. The film could be removed by ethylenediaminetetraacetate (EDTA) at any time during cell culture, permitting observation of cellular responses to conversion of the culture surface in real time. Additionally, we demonstrated the validity of the alginate thin film coating method and the performance of the film. The thickness of the alginate thin film was controlled by varying the rotation speed during spin coating. Moreover, the alginate thin film completely inhibited the adhesion of cultured cells to the culture surface, irrespective of the thickness of the film. When the alginate thin film was removed from the culture surface by EDTA, the cultured cells adhered to the culture surface, and their morphology changed. Finally, we achieved effective differentiation of C2C12 myoblasts into myotube cells by cell culture on the convertible culture surface, demonstrating the utility of our novel technique.

  5. Survivability of entrapped Lactobacillus rhamnosus in liquid- and gel-core alginate beads during storge and simulated gastrointestinal conditions

    OpenAIRE

    M. E. Rodríguez-Huezo; C. Lobato-Calleros; J.G. Reyes-Ocampo; O. Sandoval-Castilla; C. Pérez-Alonso; D.J. Pimentel-González

    2011-01-01

    L. rhamnosus cells were encapsulated in liquid-core (LCBR) and gel-core (GCBR) calcium alginate beads, and cell survivability under storage conditions and simulated gastrointestinal conditions were evaluated, and compared with that of non-encapsulated cells. The average external diameters of both beads (1.37 - 0.25 mm) were non-significantly dierent, and the average thickness of alginate gelled layer in LCBR was of 0.27 - 0.01 mm. The bacteria entrapped into LCBR tended to gather together for...

  6. Effect of encapsulation of selected probiotic cell on survival in simulated gastrointestinal tract condition

    Directory of Open Access Journals (Sweden)

    Hasiah Ayama

    2014-06-01

    Full Text Available The health benefits of probiotic bacteria have been led to their increasing use in foods. Encapsulation has been investigated to improve their survival. In this study, the selection, encapsulation and viability of lactic acid bacteria (LAB with probiotic properties in simulated gastrointestinal tract (GIT condition were investigated. One hundred and fifty isolates of LAB were obtained from 30 samples of raw cow and goat milk and some fermented foods. Nine isolates could survive under GIT condition and only 3 isolates exhibited an antimicrobial activity against all food-borne pathogenic bacteria. Among them, 2 isolates (CM21 and CM53 exhibited bile salt hydrolase activity on glycocholate and glycodeoxycholate agar plates and were identified as Lactobacillus plantarum. CM53 was selected for encapsulation using 1-3% alginate and 2% Hi-maize resistant starch by emulsion system. Viability and releasing ability of encapsulated CM53 in simulated GIT condition was increased in accordance to the alginate concentration and incubation time, respectively.

  7. Phenol biodegradation by immobilized Pseudomonas putida FNCC-0071 cells in alginate beads

    Science.gov (United States)

    Hakim, Lukman Nul; Rochmadi, Sutijan

    2017-06-01

    Phenol is one of industrial liquid waste which is harmful to the environment, so it must be degraded. It can be degraded by immobilized Pseudomonas putida FNCC-0071 cells. It needs the kinetics and mass transfer data to design this process which can be estimated by the proposed dynamic model in this study. This model involves simultaneous diffusion and reaction in the alginate bead and liquid bulk. The preliminary stage of phenol biodegradation process was acclimatization cells. This is the stage where cells were acclimated to phenol as carbon source (substrate). Then the acclimated cells were immobilized in alginate beads by extrusion method. The variation of the initial phenol concentration in the solution is 350 to 850 ppm where 60 g alginate bead contained by cells loaded into its solution in reactor batch, so then biodegradation occurs. In this study, the average radius of alginate bead was 0.152 cm. The occurred kinetic reaction process can be explained by Blanch kinetic model with the decreasing of parameter μmax' while the increasing values of initial phenol concentration in the same time, but the parameters KM, KM', and kt were increasing by the rising values of initial phenol concentration. The value of the parameter β is almost zero. Effective diffusivity of phenol and cells are 1.11 × 10-5±4.5% cm2 s-1 and 1.39 × 10-7± 0.04% cm2 s-1. The partition coefficient of phenol and cells are 0.39 ± 15% and 2.22 ± 18%.

  8. Alginate inhibits iron absorption from ferrous gluconate in a randomized controlled trial and reduces iron uptake into Caco-2 cells.

    Science.gov (United States)

    Wawer, Anna A; Harvey, Linda J; Dainty, Jack R; Perez-Moral, Natalia; Sharp, Paul; Fairweather-Tait, Susan J

    2014-01-01

    Previous in vitro results indicated that alginate beads might be a useful vehicle for food iron fortification. A human study was undertaken to test the hypothesis that alginate enhances iron absorption. A randomised, single blinded, cross-over trial was carried out in which iron absorption was measured from serum iron appearance after a test meal. Overnight-fasted volunteers (n = 15) were given a test meal of 200 g cola-flavoured jelly plus 21 mg iron as ferrous gluconate, either in alginate beads mixed into the jelly or in a capsule. Iron absorption was lower from the alginate beads than from ferrous gluconate (8.5% and 12.6% respectively, p = 0.003). Sub-group B (n = 9) consumed the test meals together with 600 mg calcium to determine whether alginate modified the inhibitory effect of calcium. Calcium reduced iron absorption from ferrous gluconate by 51%, from 11.5% to 5.6% (p = 0.014), and from alginate beads by 37%, from 8.3% to 5.2% (p = 0.009). In vitro studies using Caco-2 cells were designed to explore the reasons for the difference between the previous in vitro findings and the human study; confirmed the inhibitory effect of alginate. Beads similar to those used in the human study were subjected to simulated gastrointestinal digestion, with and without cola jelly, and the digestate applied to Caco-2 cells. Both alginate and cola jelly significantly reduced iron uptake into the cells, by 34% (p = 0.009) and 35% (p = 0.003) respectively. The combination of cola jelly and calcium produced a very low ferritin response, 16.5% (p ferrous gluconate alone. The results of these studies demonstrate that alginate beads are not a useful delivery system for soluble salts of iron for the purpose of food fortification. ClinicalTrials.gov NCT01528644.

  9. RGD-peptide modified alginate by a chemoenzymatic strategy for tissue engineering applications.

    Science.gov (United States)

    Sandvig, Ioanna; Karstensen, Kristin; Rokstad, Anne Mari; Aachmann, Finn Lillelund; Formo, Kjetil; Sandvig, Axel; Skjåk-Bræk, Gudmund; Strand, Berit Løkensgard

    2015-03-01

    One of the main challenges in tissue engineering and regenerative medicine is the ability to maintain optimal cell function and survival post-transplantation. Biomaterials such as alginates are commonly used for immunoisolation, while they may also provide structural support to the cell transplants by mimicking the extracellular matrix. In this study, arginine-glycine-aspartate (RGD)-peptide-coupled alginates of tailored composition were produced by adopting a unique chemoenzymatic strategy for substituting the nongelling mannuronic acid on the alginate. Alginates with and without RGD were produced with high and low content of G. Using carbodiimide chemistry 0.1-0.2% of the sugar units were substituted by peptide. Furthermore, the characterization by (1)H-nuclear magnetic resonance (NMR) revealed by-products from the coupling reaction that partly could be removed by coal filtration. Olfactory ensheathing cells (OECs) and myoblasts were grown in two-dimensional (2D) and 3D cultures of RGD-peptide modified or unmodified alginates obtained by the chemoenzymatically strategy and compared to native alginate. Both OECs and myoblasts adhered to the RGD-peptide modified alginates in 2D cultures, forming bipolar protrusions. OEC encapsulation resulted in cell survival for up to 9 days, thus demonstrating the potential for short-term 3D culture. Myoblasts showed long-term survival in 3D cultures, that is, up to 41 days post encapsulation. The RGD modifications did not result in marked changes in cell viability in 3D cultures. We demonstrate herein a unique technique for tailoring peptide substituted alginates with a precise and flexible composition, conserving the gel forming properties relevant for the use of alginate in tissue engineering.

  10. Fourier transform infrared spectroscopy studies of alginate-PLL capsules with varying compositions.

    Science.gov (United States)

    van Hoogmoed, Chris G; Busscher, Henk J; de Vos, Paul

    2003-10-01

    Microencapsulation of cells is a promising approach to prevention of rejection in the absence of immunosuppression. Clinical application, however, is hampered by insufficient insight into the factors that influence the biocompatibility of the capsules. Capsules prepared of alginates with a high guluronic (G) acid content proved to be more adequate for clinical application since they are more stable, but, unfortunately, they are less biocompatible than capsules prepared of intermediate-G alginate. In order to get some insight into the physicochemical factors that influence the biocompatibility of capsules for the encapsulation of living cells, the chemical compositions of alginate[bond]Ca beads and alginate[bond]PLL capsules were studied by Fourier transform infrared spectroscopy. We found that during the transition of the alginate[bond]Ca beads to alginate[bond]PLL capsules, Ca connecting the alginate molecules, disappeared at the surface of both high-G and intermediate-G alginate[bond]PLL capsules. At the same time, it turned out that high-G alginate[bond]PLL capsules contained more hydrogen bonding than did intermediate[bond]G alginate capsules. Thus the well-known higher stability of high-G alginate[bond]PLL compared to intermediate-G alginate[bond]PLL capsules is not caused by a higher degree of binding to Ca of the alginate molecules but rather by the presence of more hydrogen bonds. Another observation was that after the transition from bead to capsule, high-G alginate[bond]PLL capsules contained 20% more PLL than the intermediate-G alginate[bond]PLL capsules. Finally, we show that in both high-G and intermediate-G alginate[bond]PLL capsules, the PLL exists in the alpha-helix, in the antiparallel beta-sheet, and in the random coil conformation. This study shows that FT-IR allows for successful analyses of the chemical factors essential for understanding differences in the biocompatibility of alginate[bond]PLL capsules. Copyright 2003 Wiley Periodicals, Inc. J

  11. Alginic acid cell entrapment: a novel method for measuring in vivo macrophage cholesterol homeostasis

    Science.gov (United States)

    Sontag, Timothy J.; Chellan, Bijoy; Bhanvadia, Clarissa V.; Getz, Godfrey S.; Reardon, Catherine A.

    2015-01-01

    Macrophage conversion to atherosclerotic foam cells is partly due to the balance of uptake and efflux of cholesterol. Cholesterol efflux from cells by HDL and its apoproteins for subsequent hepatic elimination is known as reverse cholesterol transport. Numerous methods have been developed to measure in vivo macrophage cholesterol efflux. Most methods do not allow for macrophage recovery for analysis of changes in cellular cholesterol status. We describe a novel method for measuring cellular cholesterol balance using the in vivo entrapment of macrophages in alginate, which retains incorporated cells while being permeable to lipoproteins. Recipient mice were injected subcutaneously with CaCl2 forming a bubble into which a macrophage/alginate suspension was injected, entrapping the macrophages. Cells were recovered after 24 h. Cellular free and esterified cholesterol mass were determined enzymatically and normalized to cellular protein. Both normal and cholesterol loaded macrophages undergo measureable changes in cell cholesterol when injected into WT and apoA-I-, LDL-receptor-, or apoE-deficient mice. Cellular cholesterol balance is dependent on initial cellular cholesterol status, macrophage cholesterol transporter expression, and apolipoprotein deficiency. Alginate entrapment allows for the in vivo measurement of macrophage cholesterol homeostasis and is a novel platform for investigating the role of genetics and therapeutic interventions in atherogenesis. PMID:25465389

  12. High throughput single-cell and multiple-cell micro-encapsulation.

    Science.gov (United States)

    Lagus, Todd P; Edd, Jon F

    2012-06-15

    Microfluidic encapsulation methods have been previously utilized to capture cells in picoliter-scale aqueous, monodisperse drops, providing confinement from a bulk fluid environment with applications in high throughput screening, cytometry, and mass spectrometry. We describe a method to not only encapsulate single cells, but to repeatedly capture a set number of cells (here we demonstrate one- and two-cell encapsulation) to study both isolation and the interactions between cells in groups of controlled sizes. By combining drop generation techniques with cell and particle ordering, we demonstrate controlled encapsulation of cell-sized particles for efficient, continuous encapsulation. Using an aqueous particle suspension and immiscible fluorocarbon oil, we generate aqueous drops in oil with a flow focusing nozzle. The aqueous flow rate is sufficiently high to create ordering of particles which reach the nozzle at integer multiple frequencies of the drop generation frequency, encapsulating a controlled number of cells in each drop. For representative results, 9.9 μm polystyrene particles are used as cell surrogates. This study shows a single-particle encapsulation efficiency P(k=1) of 83.7% and a double-particle encapsulation efficiency P(k=2) of 79.5% as compared to their respective Poisson efficiencies of 39.3% and 33.3%, respectively. The effect of consistent cell and particle concentration is demonstrated to be of major importance for efficient encapsulation, and dripping to jetting transitions are also addressed. Continuous media aqueous cell suspensions share a common fluid environment which allows cells to interact in parallel and also homogenizes the effects of specific cells in measurements from the media. High-throughput encapsulation of cells into picoliter-scale drops confines the samples to protect drops from cross-contamination, enable a measure of cellular diversity within samples, prevent dilution of reagents and expressed biomarkers, and amplify

  13. Biocompatibility of microcapsules for cell immobilization elaborated with different type of alginates.

    Science.gov (United States)

    Orive, G; Ponce, S; Hernández, R M; Gascón, A R; Igartua, M; Pedraz, J L

    2002-09-01

    The biocompatibility of alginate-PLL-alginate (APA) microcapsules has been evaluated with respect to impurity levels. The impurity content of three different alginates (a raw high M-alginate, a raw high G-alginate and a purified high G-alginate) has been determined and the in vivo antigenic response of APA beads made with each alginate assessed. Results show that purification of the alginate not only reduces the total amount of impurities (63% less in polyphenols, 91.45% less in endotoxins and 68.5% less in protein in relation to raw high M-alginate), but also avoids an antibody response when microcapsules of this material are implanted in mice. In contrast, raw alginates produced a detectable antibody response though the differences in their impurity content. Consequently, this work revealed that purity of the alginate rather than their chemical composition, is probably of greater importance in determining microcapsule biocompatibility.

  14. Fuel cell electrode interconnect contact material encapsulation and method

    Science.gov (United States)

    Derose, Anthony J.; Haltiner, Jr., Karl J.; Gudyka, Russell A.; Bonadies, Joseph V.; Silvis, Thomas W.

    2016-05-31

    A fuel cell stack includes a plurality of fuel cell cassettes each including a fuel cell with an anode and a cathode. Each fuel cell cassette also includes an electrode interconnect adjacent to the anode or the cathode for providing electrical communication between an adjacent fuel cell cassette and the anode or the cathode. The interconnect includes a plurality of electrode interconnect protrusions defining a flow passage along the anode or the cathode for communicating oxidant or fuel to the anode or the cathode. An electrically conductive material is disposed between at least one of the electrode interconnect protrusions and the anode or the cathode in order to provide a stable electrical contact between the electrode interconnect and the anode or cathode. An encapsulating arrangement segregates the electrically conductive material from the flow passage thereby, preventing volatilization of the electrically conductive material in use of the fuel cell stack.

  15. The vasculature of nurse cells infected with non-encapsulated Trichinella species.

    Science.gov (United States)

    Khositharattanakool, Pathamet; Morakote, Nimit; Uparanukraw, Pichart

    2013-07-04

    The vasculature surrounding the nurse cells of encapsulated Trichinella spiralis has been described previously. It has been postulated the function of these vessels is to support the growth of the parasite. We describe here for the first time the vasculature surrounding the nurse cells of non-encapsulated T. pseudospiralis and T. papuae. Similar to the vasculature of uninfected muscle cells, the vessels surrounding non-encapsulated Trichinella nurse cells are dense and branched longitudinally along the long axis of the muscle cells; they also appear to be similar in diameter. The netting pattern of enlarged vessels found around T. spiralis (encapsulated) nurse cells is not present in non-encapsulated Trichinella infections. The vessels surrounding non-encapsulated Trichinella nurse cells seem to exist prior to parasite invasion of the muscle cell.

  16. Cytotoxicity study of novel water-soluble chitosan derivatives applied as membrane material of alginate microcapsules

    NARCIS (Netherlands)

    Sobol, Marcin; Bartkowiak, Artur; de Haan, Bart; de Vos, Paul

    2013-01-01

    The majority of cell encapsulation systems applied so far are based on polyelectrolyte complexes of alginate and polyvalent metal cations. Although widely used, these systems suffer from the risk of disintegration. This can be partially solved by applying chitosan as additional outer membrane. Howev

  17. Cytotoxicity study of novel water-soluble chitosan derivatives applied as membrane material of alginate microcapsules

    NARCIS (Netherlands)

    Sobol, Marcin; Bartkowiak, Artur; de Haan, Bart; de Vos, Paul

    The majority of cell encapsulation systems applied so far are based on polyelectrolyte complexes of alginate and polyvalent metal cations. Although widely used, these systems suffer from the risk of disintegration. This can be partially solved by applying chitosan as additional outer membrane.

  18. A Technology Platform to Test the Efficacy of Purification of Alginate

    NARCIS (Netherlands)

    Paredes-Juarez, Genaro A.; de Haan, Bart J.; Faas, Marijke M.; de Vos, Paul

    2014-01-01

    Alginates are widely used in tissue engineering technologies, e.g., in cell encapsulation, in drug delivery and various immobilization procedures. The success rates of these studies are highly variable due to different degrees of tissue response. A cause for this variation in success is, among other

  19. A Technology Platform to Test the Efficacy of Purification of Alginate

    NARCIS (Netherlands)

    Paredes-Juarez, Genaro A.; de Haan, Bart J.; Faas, Marijke M.; de Vos, Paul

    Alginates are widely used in tissue engineering technologies, e.g., in cell encapsulation, in drug delivery and various immobilization procedures. The success rates of these studies are highly variable due to different degrees of tissue response. A cause for this variation in success is, among other

  20. The control of cell orientation using biodegradable alginate fibers fabricated by near-field electrospinning.

    Science.gov (United States)

    Fuh, Yiin-Kuen; Wu, Yun-Chung; He, Zhe-Yu; Huang, Zih-Ming; Hu, Wei-Wen

    2016-05-01

    For spatially controlling cell alignment, near field electrospinning (NFES) was developed to direct-write alginate fiber patterns. Compared to randomly electrospun fibers, NFES fibers guided the extension of HEK 293T cells and the levels of cell alignment increased with decreasing fiber distances. However, these guiding fibers were unfavorable for cell adhesion and limited cell growth. To preserve cell alignment ability and improve biocompatibility, the stability of patterned alginate fibers was adjusted by regulating the level of ion crosslinking. These partially crosslinked NFES fibers demonstrated parallel line-patterns in the initial stage while gradually degraded with time. The reduction of fiber density increased the available area for cell growth and enhanced cell viability. On the other hand, aligned cells were still found on these degraded patterns, suggesting that cell morphologies were mainly guided during cell seeding. This dynamically controlled fiber pattern system fulfilled the need of controlling cell orientation and biocompatibility, thus was potential to modify scaffold surfaces for tissue engineering application.

  1. Increased Survival and Function of Mesenchymal Stem Cell Spheroids Entrapped in Instructive Alginate Hydrogels

    Science.gov (United States)

    Ho, Steve S.; Murphy, Kaitlin C.; Binder, Bernard Y.K.; Vissers, Caroline B.

    2016-01-01

    Mesenchymal stem cell (MSC)-based therapies are under broad investigation for applications in tissue repair but suffer from poor cell persistence and engraftment upon transplantation. MSC spheroids exhibit improved survival, anti-inflammatory, and angiogenic potential in vitro, while also promoting vascularization when implanted in vivo. However, these benefits are lost once cells engage the tissue extracellular matrix and migrate from the aggregate. The efficacy of cell therapy is consistently improved when using engineered materials, motivating the need to investigate the role of biomaterials to instruct spheroid function. In order to assess the contribution of adhesivity on spheroid activity in engineered materials and promote the bone-forming potential of MSCs, we compared the function of MSC spheroids when entrapped in Arg-Gly-Asp (RGD)-modified alginate hydrogels to nonfouling unmodified alginate. Regardless of material, MSC spheroids exhibited reduced caspase activity and greater vascular endothelial growth factor (VEGF) secretion compared with equal numbers of dissociated cells. MSC spheroids in RGD-modified hydrogels demonstrated significantly greater cell survival than spheroids in unmodified alginate. After 5 days in culture, spheroids in RGD-modified gels had similar levels of apoptosis, but more than a twofold increase in VEGF secretion compared with spheroids in unmodified gels. All gels contained mineralized tissue 8 weeks after subcutaneous implantation, and cells entrapped in RGD-modified alginate exhibited greater mineralization versus cells in unmodified gels. Immunohistochemistry confirmed more diffuse osteocalcin staining in gels containing spheroids compared with dissociated controls. This study demonstrates the promise of cell-instructive biomaterials to direct survival and function of MSC spheroids for bone tissue engineering applications. Significance Mesenchymal stem cell (MSC) spheroids exhibit improved therapeutic potential in vitro

  2. Programmed cell death in Laminaria japonica (Phaeophyta) tissues infected with alginic acid decomposing bacterium

    Institute of Scientific and Technical Information of China (English)

    WANG Gaoge; LIN Wei; ZHANG Lijing; YAN Xiaojun; DUAN Delin

    2004-01-01

    TdT-mediated dUTP-biotin nick end labeling (TUNEL) is a sensitive and valid method for detecting DNA cleavage in programmed cell death (PCD). Using this method, DNA cleavage was observed in Laminaria japonica sporophytic tissues, which were infected with alginic acid decomposing bacterium. It was found that DNA cleavage occurred 5 min after the infection, the fragments with 3′-OH groups of cleaved nuclear DNA increased with time of infection and spread from the infection site. Although no typical DNA ladder (200 bp/180 bp) was detected by routine agarose gel electrophoresis, the cleavage of nuclear DNA fragments of 97~48.5 kb could be detected by pulsed field gel electrophoresis (PFGE). By using CaspGLOWTM fluorescein active caspase-3 staining method, caspase-3 activity has been detected in response to the infection of alginic acid decomposing bacterium. Our results are similar to the observations in hypersensitive response (HR) of higher plant, suggesting that the rapid cell death of L. Japonica infected by alginic acid decomposing bacterium might be involved in PCD, and indicating that the occurrence of PCD is an active defense process against the pathogen's infection.

  3. A non-destructive culturing and cell sorting method for cardiomyocytes and neurons using a double alginate layer.

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    Hideyuki Terazono

    Full Text Available A non-destructive method of collecting cultured cells after identifying their in situ functional characteristics is proposed. In this method, cells are cultivated on an alginate layer in a culture dish and released by spot application of a calcium chelate buffer that locally melts the alginate layer and enables the collection of cultured cells at the single-cell level. Primary hippocampal neurons, beating human embryonic stem (hES cell-derived cardiomyocytes, and beating hES cell-derived cardiomyocyte clusters cultivated on an alginate layer were successfully released and collected with a micropipette. The collected cells were recultured while maintaining their physiological function, including beating, and elongated neurites. These results suggest that the proposed method may eventually facilitate the transplantation of ES- or iPS-derived cardiomyocytes and neurons differentiated in culture.

  4. Biocompatible Hydrogels for Microarray Cell Printing and Encapsulation

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    Akshata Datar

    2015-10-01

    Full Text Available Conventional drug screening processes are a time-consuming and expensive endeavor, but highly rewarding when they are successful. To identify promising lead compounds, millions of compounds are traditionally screened against therapeutic targets on human cells grown on the surface of 96-wells. These two-dimensional (2D cell monolayers are physiologically irrelevant, thus, often providing false-positive or false-negative results, when compared to cells grown in three-dimensional (3D structures such as hydrogel droplets. However, 3D cell culture systems are not easily amenable to high-throughput screening (HTS, thus inherently low throughput, and requiring relatively large volume for cell-based assays. In addition, it is difficult to control cellular microenvironments and hard to obtain reliable cell images due to focus position and transparency issues. To overcome these problems, miniaturized 3D cell cultures in hydrogels were developed via cell printing techniques where cell spots in hydrogels can be arrayed on the surface of glass slides or plastic chips by microarray spotters and cultured in growth media to form cells encapsulated 3D droplets for various cell-based assays. These approaches can dramatically reduce assay volume, provide accurate control over cellular microenvironments, and allow us to obtain clear 3D cell images for high-content imaging (HCI. In this review, several hydrogels that are compatible to microarray printing robots are discussed for miniaturized 3D cell cultures.

  5. Alginate Immobilization of Metabolic Enzymes (AIME) for High ...

    Science.gov (United States)

    Alginate Immobilization of Metabolic Enzymes (AIME) for High-Throughput Screening Assays DE DeGroot, RS Thomas, and SO SimmonsNational Center for Computational Toxicology, US EPA, Research Triangle Park, NC USAThe EPA’s ToxCast program utilizes a wide variety of high-throughput screening (HTS) assays to assess chemical perturbations of molecular and cellular endpoints. A key criticism of using HTS assays for toxicity assessment is the lack of xenobiotic metabolism (XM) which precludes both metabolic detoxification as well as bioactivation of chemicals tested in vitro thereby mischaracterizing the potential risk posed by these chemicals. To address this deficiency, we have developed an extracellular platform to retrofit existing HTS assays with XM activity. This platform utilizes the S9 fraction of liver homogenate encapsulated in an alginate gel network which reduces the cytotoxicity caused by direct addition of S9 to cells in culture. Alginate microspheres containing encapsulated human liver S9 were cross-linked to solid supports extending from a 96-well plate lid and were assayed using a pro-luciferin substrate specific for CYP3A4 (IPA). We demonstrate that S9 was successfully encapsulated and remained enzymatically active post-encapsulation with 5-10X the CYP3A4 activity as compared to 1 µg solubilized human liver S9. Ketoconazole, a known inhibitor of human CYP3A4, inhibited CYP3A4 activity in a concentration-dependent manner (IC50: 0.27 µM) and inhibiti

  6. Cell retention by encapsulation for the cultivation of Jurkat cells in fixed and fluidized bed reactors.

    Science.gov (United States)

    Kaiser, P; Werner, M; Jérôme, V; Hübner, H; Buchholz, R; Freitag, R

    2014-12-01

    Jurkat cells are accepted model cells for primary human T lymphocytes, for example, in medical research. Their growth to tissue-like cell densities (up to 100 × 10(6)  cells/mLcapsule ) in semi-permeable (molecular weight cut off cultivations, that is, under conditions where both encapsulated and non-encapsulated cells can be cultivated under otherwise identical conditions, showed that maximum specific growth rates were higher for the encapsulated than for the non-encapsulated cells. In the subsequent batch and repeated batch bioreactor experiments (only encapsulated cells), growth rates were similar, with the exception of the fixed bed batch reactor, where growth kinetics were significantly slower. Concomitantly, a significant fraction of the cells towards the bottom of the bed were no longer metabolically active, though apparently not dead. In the repeated batch fluidized bed reactor cellular division could be maintained for more than two weeks, albeit with a specific growth rate below the maximum one, leading to final cell densities of approximately 180 × 10(6)  cell/gcapsule . At the same time, the cell cycle distribution of the cells was shifted to the S and G2/M phases.

  7. Preparation of carboxymethyl cellulose based microgels for cell encapsulation

    Directory of Open Access Journals (Sweden)

    Y. Ke

    2014-11-01

    Full Text Available Biocompatible and biodegradable carboxymethyl cellulose (CMC has been modified with 4-hydroxybenzylamine (CMC-Ph in order to prepare CMC-based microgels through the horseradish peroxidise/hydrogen peroxide enzymatic reaction. CMC-Ph was identified as a blend, and the amount of the grafted 4-hydroxybenzylamine per 100 units of CMC was between 17 and 23 according to the molecular weight of CMC. Through a special designed co-flowing microfluidic device, CMC-Ph microgels were prepared with the radius from 100 to 500 μm via adjusting the flow rates of the disperse phase and the continuous phase, respectively. The chondrocytic cell line ATDC5 was encapsulated in the CMC-Ph microgels. The cell-laden microgels were cultured for up to 40 days, illustrating the biocompatibility of CMC-Ph and the microfluidic approach through the enzymatic crosslinking reaction primarily. CMC-Ph showed a great promise to encapsulate the cells for further fabrication of the injectable scaffolds.

  8. [Effects of alginate/collagen scaffold on cell proliferation and differentiation of human adipose-derived mesenchymal stem cells].

    Science.gov (United States)

    Cheng, W; Han, X P; Mou, S L; Yang, F; Liu, L P

    2017-04-09

    Objective: To build scaffold materials with different concentrations of alginate and collagen, and to observe the effects of alginate/collagen ratio on the proliferation of human adipose-derived mesenchymal stem cells (hAMSC) and osteogenic differentiation. The optimal concentration of alginate/collagen will be chosen for constructing hydrogel that will be used for bone tissue engineering. Methods: Soluble hydrogel scaffold materials containing alginate/collagen were prepared, and the following groups were established based on different alginate/collagen ratio: 4∶1 (group A), 2∶1 (group B), and 1∶1 (group C). Cell proliferation on the material surface was observed using the cell counting kit-8 (CCK-8) assay, while cell viability in each material group were observed using live/dead staining. Quantitative real-time PCR(qPCR) was used to measure the differential expression of osteogenesis-related genes on and in the materials. Immunofluorescence staining was used to measure the differential gene expression of osteogenesis-related proteins in each group. Results: The results from the CCK-8 assay showed increasing cell proliferation rate on the lyophilized hydrogel material surface as the collagen concentration increased, and the highest cell proliferation was observed in group C. Live/dead staining assay indicated that cells were able to proliferate in all three types of hydrogel materials, and the highest cell viability was found in material from group B ([87.50±2.65]%). qPCR showed that the expression of osteogenesis-related genes in group C was the highest, among the three groups, while the expression of osteocalcin in group B was significantly higher than those in the other two groups (Palginate/collagen scaffold materials did not show adverse effects on the cell proliferation of hAMSC and osteogenenic differentiation. Bone tissue engineering can use 10% hydrogel material, and when the sodium alginate and collagen have a ratio of 2∶1, the hydrogel can be

  9. Microencapsulation of probiotics in hydrogel particles: enhancing Lactococcus lactis subsp. cremoris LM0230 viability using calcium alginate beads.

    Science.gov (United States)

    Yeung, Timothy W; Arroyo-Maya, Izlia J; McClements, David J; Sela, David A

    2016-04-01

    Probiotics are beneficial microbes often added to food products to enhance the health and wellness of consumers. A major limitation to producing efficacious functional foods containing probiotic cells is their tendency to lose viability during storage and gastrointestinal transit. In this study, the impact of encapsulating probiotics within food-grade hydrogel particles to mitigate sensitivity to environmental stresses was examined. Confocal fluorescence microscopy confirmed that Lactococcus lactis were trapped within calcium alginate beads formed by dripping a probiotic-alginate mixture into a calcium solution. Encapsulation improved the viability of the probiotics during aerobic storage: after seven days, less than a two-log reduction was observed in encapsulated cells stored at room temperature, demonstrating that a high concentration of cells survived relative to non-encapsulated bacteria. These hydrogel beads may have applications for improving the stability and efficacy of probiotics in functional foods.

  10. Chondrogenic Differentiation of Human Umbilical Cord Blood-Derived Unrestricted Somatic Stem Cells on A 3D Beta-Tricalcium Phosphate-Alginate-Gelatin Scaffold

    Directory of Open Access Journals (Sweden)

    Masoud Soleimani

    2014-03-01

    Full Text Available Objective: Finding cell sources for cartilage tissue engineering is a critical procedure. The purpose of the present experimental study was to test the in vitro efficacy of the beta-tricalcium phosphate-alginate-gelatin (BTAG scaffold to induce chondrogenic differentiation of human umbilical cord blood-derived unrestricted somatic stem cells (USSCs. Materials and Methods: In this experimental study, USSCs were encapsulated in BTAG scaffold and cultured for 3 weeks in chondrogenic medium as chondrogenic group and in Dulbecco’s Modified Eagle’s Medium (DMEM as control group. Chondrogenic differentiation was evaluated by histology, immunofluorescence and RNA analyses for the expression of cartilage extracellular matrix components. The obtain data were analyzed using SPSS version 15. Results: Histological and immunohistochemical staining revealed that collagen II was markedly expressed in the extracellular matrix of the seeded cells on scaffold in presence of chondrogenic media after 21 days. Reverse transcription-polymerase chain reaction (RT-PCR showed a significant increase in expression levels of genes encoded the cartilage-specific markers, aggrecan, type I and II collagen, and bone morphogenetic protein (BMP-6 in chondrogenic group. Conclusion: This study demonstrates that BTAG can be considered as a suitable scaffold for encapsulation and chondrogenesis of USSCs.

  11. Biomedical-grade, high mannuronic acid content (BioMVM) alginate enhances the proteoglycan production of primary human meniscal fibrochondrocytes in a 3-D microenvironment

    Science.gov (United States)

    Rey-Rico, Ana; Klich, Angelique; Cucchiarini, Magali; Madry, Henning

    2016-01-01

    Alginates are important hydrogels for meniscus tissue engineering as they support the meniscal fibrochondrocyte phenotype and proteoglycan production, the extracellular matrix (ECM) component chiefly responsible for its viscoelastic properties. Here, we systematically evaluated four biomedical- and two nonbiomedical-grade alginates for their capacity to provide the best three-dimensional (3-D) microenvironment and to support proteoglycan synthesis of encapsulated human meniscal fibrochondrocytes in vitro. Biomedical-grade, high mannuronic acid alginate spheres (BioLVM, BioMVM) were the most uniform in size, indicating an effect of the purity of alginate on the shape of the spheres. Interestingly, the purity of alginates did not affect cell viability. Of note, only fibrochondrocytes encapsulated in BioMVM alginate produced and retained significant amounts of proteoglycans. Following transplantation in an explant culture model, the alginate spheres containing fibrochondrocytes remained in close proximity with the meniscal tissue adjacent to the defect. The results reveal a promising role of BioMVM alginate to enhance the proteoglycan production of primary human meniscal fibrochondrocytes in a 3-D hydrogel microenvironment. These findings have significant implications for cell-based translational studies aiming at restoring lost meniscal tissue in regions containing high amounts of proteoglycans. PMID:27302206

  12. Sonication-induced gelation of silk fibroin for cell encapsulation.

    Science.gov (United States)

    Wang, Xiaoqin; Kluge, Jonathan A; Leisk, Gary G; Kaplan, David L

    2008-03-01

    Purified native silk fibroin forms beta-sheet-rich, physically cross-linked, hydrogels from aqueous solution, in a process influenced by environmental parameters. Previously we reported gelation times of days to weeks for aqueous native silk protein solutions, with high ionic strength and temperature and low pH responsible for increasing gelation kinetics. Here we report a novel method to accelerate the process and control silk fibroin gelation through ultrasonication. Depending on the sonication parameters, including power output and time, along with silk fibroin concentration, gelation could be controlled from minutes to hours, allowing the post-sonication addition of cells prior to final gel setting. Mechanistically, ultrasonication initiated the formation of beta-sheets by alteration in hydrophobic hydration, thus accelerating the formation of physical cross-links responsible for gel stabilization. K(+) at physiological concentrations and low pH promoted gelation, which was not observed in the presence of Ca(2+). The hydrogels were assessed for mechanical properties and proteolytic degradation; reported values matched or exceeded other cell-encapsulating gel material systems. Human bone marrow derived mesenchymal stem cells (hMSCs) were successfully incorporated into these silk fibroin hydrogels after sonication, followed by rapid gelation and sustained cell function. Sonicated silk fibroin solutions at 4%, 8%, and 12% (w/v), followed by mixing in hMSCs, gelled within 0.5-2 h. The cells grew and proliferated in the 4% gels over 21 days, while survival was lower in the gels with higher protein content. Thus, sonication provides a useful new tool with which to initiate rapid sol-gel transitions, such as for cell encapsulation.

  13. Crystal structure of bacterial cell-surface alginate-binding protein with an M75 peptidase motif

    Energy Technology Data Exchange (ETDEWEB)

    Maruyama, Yukie; Ochiai, Akihito [Laboratory of Basic and Applied Molecular Biotechnology, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611-0011 (Japan); Mikami, Bunzo [Laboratory of Applied Structural Biology, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611-0011 (Japan); Hashimoto, Wataru [Laboratory of Basic and Applied Molecular Biotechnology, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611-0011 (Japan); Murata, Kousaku, E-mail: kmurata@kais.kyoto-u.ac.jp [Laboratory of Basic and Applied Molecular Biotechnology, Graduate School of Agriculture, Kyoto University, Uji, Kyoto 611-0011 (Japan)

    2011-02-18

    Research highlights: {yields} Bacterial alginate-binding Algp7 is similar to component EfeO of Fe{sup 2+} transporter. {yields} We determined the crystal structure of Algp7 with a metal-binding motif. {yields} Algp7 consists of two helical bundles formed through duplication of a single bundle. {yields} A deep cleft involved in alginate binding locates around the metal-binding site. {yields} Algp7 may function as a Fe{sup 2+}-chelated alginate-binding protein. -- Abstract: A gram-negative Sphingomonas sp. A1 directly incorporates alginate polysaccharide into the cytoplasm via the cell-surface pit and ABC transporter. A cell-surface alginate-binding protein, Algp7, functions as a concentrator of the polysaccharide in the pit. Based on the primary structure and genetic organization in the bacterial genome, Algp7 was found to be homologous to an M75 peptidase motif-containing EfeO, a component of a ferrous ion transporter. Despite the presence of an M75 peptidase motif with high similarity, the Algp7 protein purified from recombinant Escherichia coli cells was inert on insulin B chain and N-benzoyl-Phe-Val-Arg-p-nitroanilide, both of which are substrates for a typical M75 peptidase, imelysin, from Pseudomonas aeruginosa. The X-ray crystallographic structure of Algp7 was determined at 2.10 A resolution by single-wavelength anomalous diffraction. Although a metal-binding motif, HxxE, conserved in zinc ion-dependent M75 peptidases is also found in Algp7, the crystal structure of Algp7 contains no metal even at the motif. The protein consists of two structurally similar up-and-down helical bundles as the basic scaffold. A deep cleft between the bundles is sufficiently large to accommodate macromolecules such as alginate polysaccharide. This is the first structural report on a bacterial cell-surface alginate-binding protein with an M75 peptidase motif.

  14. An additive manufacturing-based PCL-alginate-chondrocyte bioprinted scaffold for cartilage tissue engineering.

    Science.gov (United States)

    Kundu, Joydip; Shim, Jin-Hyung; Jang, Jinah; Kim, Sung-Won; Cho, Dong-Woo

    2015-11-01

    Regenerative medicine is targeted to improve, restore or replace damaged tissues or organs using a combination of cells, materials and growth factors. Both tissue engineering and developmental biology currently deal with the process of tissue self-assembly and extracellular matrix (ECM) deposition. In this investigation, additive manufacturing (AM) with a multihead deposition system (MHDS) was used to fabricate three-dimensional (3D) cell-printed scaffolds using layer-by-layer (LBL) deposition of polycaprolactone (PCL) and chondrocyte cell-encapsulated alginate hydrogel. Appropriate cell dispensing conditions and optimum alginate concentrations for maintaining cell viability were determined. In vitro cell-based biochemical assays were performed to determine glycosaminoglycans (GAGs), DNA and total collagen contents from different PCL-alginate gel constructs. PCL-alginate gels containing transforming growth factor-β (TGFβ) showed higher ECM formation. The 3D cell-printed scaffolds of PCL-alginate gel were implanted in the dorsal subcutaneous spaces of female nude mice. Histochemical [Alcian blue and haematoxylin and eosin (H&E) staining] and immunohistochemical (type II collagen) analyses of the retrieved implants after 4 weeks revealed enhanced cartilage tissue and type II collagen fibril formation in the PCL-alginate gel (+TGFβ) hybrid scaffold. In conclusion, we present an innovative cell-printed scaffold for cartilage regeneration fabricated by an advanced bioprinting technology.

  15. Evaluation of gene expression and alginate production in response to oxygen transfer in continuous culture of Azotobacter vinelandii.

    Directory of Open Access Journals (Sweden)

    Alvaro Díaz-Barrera

    Full Text Available Alginates are polysaccharides used as food additives and encapsulation agents in biotechnology, and their functional properties depend on its molecular weight. In this study, different steady-states in continuous cultures of A. vinelandii were established to determine the effect of the dilution rate (D and the agitation rate on alginate production and expression of genes involved in alginate polymerization and depolymerization. Both, the agitation and dilution rates, determined the partitioning of the carbon utilization from sucrose into alginate and CO2 under oxygen-limiting conditions. A low D (0.07 h(-1 and 500 rpm resulted in the highest carbon utilization into alginate (25%. Quantitative real-time polymerase chain reaction was used to determine the transcription level of six genes involved in alginate polymerization and depolymerization. In chemostat cultures at 0.07 h(-1, the gene expression was affected by changes in the agitation rate. By increasing the agitation rate from 400 to 600 rpm, the algE7 gene expression decreased tenfold, whereas alyA1, algL and alyA2 gene expression increased between 1.5 and 2.8 times under similar conditions evaluated. Chemostat at 0.07 h(-1 showed a highest alginate molecular weight (580 kDa at 500 rpm whereas similar molecular weights (480 kDa were obtained at 400 and 600 rpm. The highest molecular weight was not explained by changes in the expression of alg8 and alg44 (genes involved in alginate polymerization. Nonetheless, a different expression pattern observed for lyases could explain the highest alginate molecular weight obtained. Overall, the results suggest that the control of alginate molecular weight in A. vinelandii cells growing in continuous mode is determined by a balance between the gene expression of intracellular and extracellular lyases in response to oxygen availability. These findings better our understanding of the biosynthesis of bacterial alginate and help us progress toward obtain

  16. Evaluation of Gene Expression and Alginate Production in Response to Oxygen Transfer in Continuous Culture of Azotobacter vinelandii

    Science.gov (United States)

    Díaz-Barrera, Alvaro; Martínez, Fabiola; Guevara Pezoa, Felipe; Acevedo, Fernando

    2014-01-01

    Alginates are polysaccharides used as food additives and encapsulation agents in biotechnology, and their functional properties depend on its molecular weight. In this study, different steady-states in continuous cultures of A. vinelandii were established to determine the effect of the dilution rate (D) and the agitation rate on alginate production and expression of genes involved in alginate polymerization and depolymerization. Both, the agitation and dilution rates, determined the partitioning of the carbon utilization from sucrose into alginate and CO2 under oxygen-limiting conditions. A low D (0.07 h−1) and 500 rpm resulted in the highest carbon utilization into alginate (25%). Quantitative real-time polymerase chain reaction was used to determine the transcription level of six genes involved in alginate polymerization and depolymerization. In chemostat cultures at 0.07 h−1, the gene expression was affected by changes in the agitation rate. By increasing the agitation rate from 400 to 600 rpm, the algE7 gene expression decreased tenfold, whereas alyA1, algL and alyA2 gene expression increased between 1.5 and 2.8 times under similar conditions evaluated. Chemostat at 0.07 h−1 showed a highest alginate molecular weight (580 kDa) at 500 rpm whereas similar molecular weights (480 kDa) were obtained at 400 and 600 rpm. The highest molecular weight was not explained by changes in the expression of alg8 and alg44 (genes involved in alginate polymerization). Nonetheless, a different expression pattern observed for lyases could explain the highest alginate molecular weight obtained. Overall, the results suggest that the control of alginate molecular weight in A. vinelandii cells growing in continuous mode is determined by a balance between the gene expression of intracellular and extracellular lyases in response to oxygen availability. These findings better our understanding of the biosynthesis of bacterial alginate and help us progress toward obtain tailor

  17. Influence of hydrophobic modification in alginate-based hydrogels for biomedical applications

    Science.gov (United States)

    Choudhary, Soumitra

    Alginate has been exploited commercially for decades in foods, textiles, paper, pharmaceutical industries, and also as a detoxifier for removing heavy metals. Alginate is also popular in cell encapsulation because of its relatively mild gelation protocol and simple chemistry with which biological active entities can be immobilized. Surface modification of alginate gels has been explored to induce desired cell interactions with the gel matrix. These modifications alter the bulk properties, which strongly determine on how cells feel and response to the three-dimensional microenvironment. However, there is a need to develop strategies to engineer functionalities into bulk alginate hydrogels that not only preserve their inherent qualities but are also less toxic. In this thesis, our main focus was to optimize the mechanical properties of alginate-based hydrogels, and by doing so control the performance of the biomaterials. In the first scheme, we used alginate and hydrophobically modified ethyl hydroxy ethyl cellulose as components in interpenetrating polymer network (IPN) gels. The second network was used to control gelation time and rheological properties. We believe these experiments also may provide insight into the mechanical and structural properties of more complex biopolymer gels and naturally-occurring IPNs. Next, we worked on incorporating a hydrophobic moiety directly into the alginate chain, resulting in materials for extended release of hydrophobic drugs. We successfully synthesized hydrophobically modified alginate (HMA) by attaching octylamine groups onto the alginate backbone by standard carbodiimide based amide coupling reaction. Solubility of several model hydrophobic drugs in dilute HMA solutions was found to be increased by more than an order of magnitude. HMA hydrogels, prepared by crosslinking the alginate chains with calcium ions, were found to exhibit excellent mechanical properties (modulus ˜100 kPa) with release extended upto 5 days. Ability

  18. Silver nanoparticles impregnated alginate-chitosan-blended nanocarrier induces apoptosis in human glioblastoma cells.

    Science.gov (United States)

    Sharma, Shilpa; Chockalingam, S; Sanpui, Pallab; Chattopadhyay, Arun; Ghosh, Siddhartha Sankar

    2014-01-01

    Herein, a green method for the development of a novel biodegradable silver nanoparticles (NPs) impregnated alginate-chitosan-blended nanocarrier (Ag NPs-Alg-Chi NC) is reported. The synthesis of Ag NPs-Alg-Chi NC is based on the polyelectrolyte complex formation between alginate and chitosan. The composite NC is characterized by ultraviolet-visible spectroscopy, transmission electron microscopy, scanning electron microscopy (SEM), Fourier transform infrared spectroscopy, and X-ray diffraction. The Ag NPs in the NC are found to elicit anticell proliferative effect on refractory U87MG (human glioblastoma) cells at IC50 of 2.4 μg mL(-1) for Ag NPs. The cell cycle analysis shows extensive DNA damage. Elevation in reactive oxygen species level indicates induction of oxidative stress in treated cells. Mitochondrial dysfunction in cell death is evident from the depolarization of mitochondrial membrane potential (ΔΨm ). Fluorescence and SEM images of the treated cells reveal nuclear and morphological changes characteristic of apoptosis, which is further confirmed by TUNEL assay. The induction of apoptosis at low concentration of Ag NPs present in Ag NPs-Alg-Chi NC in comparison with free Ag NPs makes it a promising tool for cancer therapy.

  19. Hybrid 3D printing and electrodeposition approach for controllable 3D alginate hydrogel formation.

    Science.gov (United States)

    Shang, Wanfeng; Liu, Yanting; Wan, Wenfeng; Hu, Chengzhi; Liu, Zeyang; Wong, Chin To; Fukuda, Toshio; Shen, Yajing

    2017-06-07

    Calcium alginate hydrogels are widely used as biocompatible materials in a substantial number of biomedical applications. This paper reports on a hybrid 3D printing and electrodeposition approach for forming 3D calcium alginate hydrogels in a controllable manner. Firstly, a specific 3D hydrogel printing system is developed by integrating a customized ejection syringe with a conventional 3D printer. Then, a mixed solution of sodium alginate and CaCO3 nanoparticles is filled into the syringe and can be continuously ejected out of the syringe nozzle onto a conductive substrate. When applying a DC voltage (∼5 V) between the substrate (anode) and the nozzle (cathode), the Ca(2+) released from the CaCO3 particles can crosslink the alginate to form calcium alginate hydrogel on the substrate. To elucidate the gel formation mechanism and better control the gel growth, we can further establish and verify a gel growth model by considering several key parameters, i.e., applied voltage and deposition time. The experimental results indicate that the alginate hydrogel of various 3D structures can be formed by controlling the movement of the 3D printer. A cell viability test is conducted and shows that the encapsulated cells in the gel can maintain a high survival rate (∼99% right after gel formation). This research establishes a reliable method for the controllable formation of 3D calcium alginate hydrogel, exhibiting great potential for use in basic biology and applied biomedical engineering.

  20. Cell proliferation on PVA/sodium alginate and PVA/poly(γ-glutamic acid) electrospun fiber

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    Yang, Jen Ming, E-mail: jmyang@mail.cgu.edu.tw [Department of Chemical and Materials Engineering, Chang Gung University, Taoyuan 333, Taiwan, ROC (China); Yang, Jhe Hao [Department of Electronic Engineering, Chang Gung University, Taoyuan, Taiwan, ROC (China); Tsou, Shu Chun; Ding, Chian Hua [Department of Chemical and Materials Engineering, Chang Gung University, Taoyuan 333, Taiwan, ROC (China); Hsu, Chih Chin [Department of Physical Medicine and Rehabilitation, Chang Gung Memorial Hospital at Keelung, Keelung, Taiwan, ROC (China); School of Traditional Chinese Medicine, Chang Gung University, Taoyuan, Taiwan, ROC (China); Yang, Kai Chiang [School of Biomedical Engineering, College of Biomedical Engineering, Taipei Medical University, Taipei 11031, Taiwan, ROC (China); Yang, Chun Chen [Department of Chemical Engineering, Ming-Chi University of Science and Technology, New Taipei City, Taiwan, ROC (China); Chen, Ko Shao [Department of Materials Engineering, Tatung University, Taipei, Taiwan, ROC (China); Chen, Szi Wen [Department of Electronic Engineering, Chang Gung University, Taoyuan, Taiwan, ROC (China); Wang, Jong Shyan [Department of Physical Therapy and the Graduate Institute of Rehabilitation Science, Chang Gung University, Taoyuan, Taiwan, ROC (China)

    2016-09-01

    To overcome the obstacles of easy dissolution of PVA nanofibers without crosslinking treatment and the poor electrospinnability of the PVA cross-linked nanofibers via electrospinning process, the PVA based electrospun hydrogel nanofibers are prepared with post-crosslinking method. To expect the electrospun hydrogel fibers might be a promising scaffold for cell culture and tissue engineering applications, the evaluation of cell proliferation on the post-crosslinking electrospun fibers is conducted in this study. At beginning, poly(vinyl alcohol) (PVA), PVA/sodium alginate (PVASA) and PVA/poly(γ-glutamic acid) (PVAPGA) electrospun fibers were prepared by electrospinning method. The electrospun PVA, PVASA and PVAPGA nanofibers were treated with post-cross-linking method with glutaraldehyde (Glu) as crosslinking agent. These electrospun fibers were characterized with thermogravimetry analysis (TGA) and their morphologies were observed with a scanning electron microscope (SEM). To support the evaluation and explanation of cell growth on the fiber, the study of 3T3 mouse fibroblast cell growth on the surface of pure PVA, SA, and PGA thin films is conducted. The proliferation of 3T3 on the electrospun fiber surface of PVA, PVASA, and PVAPGA was evaluated by seeding 3T3 fibroblast cells on these crosslinked electrospun fibers. The cell viability on electrospun fibers was conducted with water-soluble tetrazolium salt-1 assay (Cell Proliferation Reagent WST-1). The morphology of the cells on the fibers was also observed with SEM. The results of WST-1 assay revealed that 3T3 cells cultured on different electrospun fibers had similar viability, and the cell viability increased with time for all electrospun fibers. From the morphology of the cells on electrospun fibers, it is found that 3T3 cells attached on all electrospun fiber after 1 day seeded. Cell–cell communication was noticed on day 3 for all electrospun fibers. Extracellular matrix (ECM) productions were found and

  1. Calcium alginate gels as stem cell matrix-making paracrine stem cell activity available for enhanced healing after surgery.

    Directory of Open Access Journals (Sweden)

    Andreas Schmitt

    Full Text Available Regeneration after surgery can be improved by the administration of anabolic growth factors. However, to locally maintain these factors at the site of regeneration is problematic. The aim of this study was to develop a matrix system containing human mesenchymal stem cells (MSCs which can be applied to the surgical site and allows the secretion of endogenous healing factors from the cells. Calcium alginate gels were prepared by a combination of internal and external gelation. The gelling behaviour, mechanical stability, surface adhesive properties and injectability of the gels were investigated. The permeability of the gels for growth factors was analysed using bovine serum albumin and lysozyme as model proteins. Human MSCs were isolated, cultivated and seeded into the alginate gels. Cell viability was determined by AlamarBlue assay and fluorescence microscopy. The release of human VEGF and bFGF from the cells was determined using an enzyme-linked immunoassay. Gels with sufficient mechanical properties were prepared which remained injectable through a syringe and solidified in a sufficient time frame after application. Surface adhesion was improved by the addition of polyethylene glycol 300,000 and hyaluronic acid. Humans MSCs remained viable for the duration of 6 weeks within the gels. Human VEGF and bFGF was found in quantifiable concentrations in cell culture supernatants of gels loaded with MSCs and incubated for a period of 6 weeks. This work shows that calcium alginate gels can function as immobilization matrices for human MSCs.

  2. Calcium alginate gels as stem cell matrix-making paracrine stem cell activity available for enhanced healing after surgery.

    Science.gov (United States)

    Schmitt, Andreas; Rödel, Philipp; Anamur, Cihad; Seeliger, Claudine; Imhoff, Andreas B; Herbst, Elmar; Vogt, Stephan; van Griensven, Martijn; Winter, Gerhard; Engert, Julia

    2015-01-01

    Regeneration after surgery can be improved by the administration of anabolic growth factors. However, to locally maintain these factors at the site of regeneration is problematic. The aim of this study was to develop a matrix system containing human mesenchymal stem cells (MSCs) which can be applied to the surgical site and allows the secretion of endogenous healing factors from the cells. Calcium alginate gels were prepared by a combination of internal and external gelation. The gelling behaviour, mechanical stability, surface adhesive properties and injectability of the gels were investigated. The permeability of the gels for growth factors was analysed using bovine serum albumin and lysozyme as model proteins. Human MSCs were isolated, cultivated and seeded into the alginate gels. Cell viability was determined by AlamarBlue assay and fluorescence microscopy. The release of human VEGF and bFGF from the cells was determined using an enzyme-linked immunoassay. Gels with sufficient mechanical properties were prepared which remained injectable through a syringe and solidified in a sufficient time frame after application. Surface adhesion was improved by the addition of polyethylene glycol 300,000 and hyaluronic acid. Humans MSCs remained viable for the duration of 6 weeks within the gels. Human VEGF and bFGF was found in quantifiable concentrations in cell culture supernatants of gels loaded with MSCs and incubated for a period of 6 weeks. This work shows that calcium alginate gels can function as immobilization matrices for human MSCs.

  3. Calcium Alginate Gels as Stem Cell Matrix – Making Paracrine Stem Cell Activity Available for Enhanced Healing after Surgery

    Science.gov (United States)

    Schmitt, Andreas; Rödel, Philipp; Anamur, Cihad; Seeliger, Claudine; Imhoff, Andreas B.; Herbst, Elmar; Vogt, Stephan; van Griensven, Martijn; Winter, Gerhard; Engert, Julia

    2015-01-01

    Regeneration after surgery can be improved by the administration of anabolic growth factors. However, to locally maintain these factors at the site of regeneration is problematic. The aim of this study was to develop a matrix system containing human mesenchymal stem cells (MSCs) which can be applied to the surgical site and allows the secretion of endogenous healing factors from the cells. Calcium alginate gels were prepared by a combination of internal and external gelation. The gelling behaviour, mechanical stability, surface adhesive properties and injectability of the gels were investigated. The permeability of the gels for growth factors was analysed using bovine serum albumin and lysozyme as model proteins. Human MSCs were isolated, cultivated and seeded into the alginate gels. Cell viability was determined by AlamarBlue assay and fluorescence microscopy. The release of human VEGF and bFGF from the cells was determined using an enzyme-linked immunoassay. Gels with sufficient mechanical properties were prepared which remained injectable through a syringe and solidified in a sufficient time frame after application. Surface adhesion was improved by the addition of polyethylene glycol 300,000 and hyaluronic acid. Humans MSCs remained viable for the duration of 6 weeks within the gels. Human VEGF and bFGF was found in quantifiable concentrations in cell culture supernatants of gels loaded with MSCs and incubated for a period of 6 weeks. This work shows that calcium alginate gels can function as immobilization matrices for human MSCs. PMID:25793885

  4. Drying of micro-encapsulated lactic acid bacteria — Effects of trehalose and immobilization on cell survival and release properties

    Science.gov (United States)

    Li, Xiaoyan; Chen, Xiguang

    2009-03-01

    Lactic acid bacteria (LAB) were encapsulated with alginate, gelatin and trehalose additives by the extrusion method and dried at 4 °C. The microcapsules were generally spherical and had a wrinkled surface with a size of 1.7 mm ± 0.2 mm. Trehalose as a carbohydrate source in the culture medium could reduce acid production and performed no function in the positive proliferation of LAB. Using trehalose as a carbohydrate source and protective medium simultaneously had a benefit in the protection of LAB cells during the storage at 4 °C. The density of live LAB cells could be 107 CFU g-1 after 8 weeks of storage. Cells of LAB could be continuously released from the capsules from the acidic (pH 1.2) to neutral conditions (pH 6.8). The release amounts and proliferation speeds of LAB cells in neutral medium were much larger and faster than those in acidic conditions. Additionally, immobilization of LAB could improve the survival of cells when they were exposed to acidic medium (pH 1.2) with a survival rate of 76 %.

  5. Drying of Micro-Encapsulated Lactic Acid Bacteria-Effects of Trehalose and Immobilization on Cell Survival and Release Properties

    Institute of Scientific and Technical Information of China (English)

    LI Xiaoyan; CHEN Xiguang

    2009-01-01

    Lactic acid bacteria (LAB) were encapsulated with alginate, gelatin and trehalose additives by the extrusion method and dried at 4℃. The microcapsules were generally spherical and had a wrinkled surface with a size of 1.7mm±0.2mm. Trehalose as a carbohydrate source in the culture medium could reduce acid production and performed no function in the positive proliferation of LAB. Using trehalose as a carbohydrate source and protective medium simultaneously had a benefit in the protection of LAB cells during the storage at 4℃. The density of hve LAB cells could be 10- CFU g-1 after 8 weeks of storage. Cells of LAB could be con-tinuously released from the capsules from the acidic (pH 1.2) to neutral conditions (plt 6.8). The release amounts and proliferation speeds of LAB cells in neutral medium were much larger and faster than those m acidic conditions. Additionally, immobilization of LAB could improve the survival of cells when they, were exposed to acidic medium (pH 1.2) with a survival rate of 76 %.

  6. Fluid and cell behaviors along a 3D printed alginate/gelatin/fibrin channel.

    Science.gov (United States)

    Xu, Yufan; Wang, Xiaohong

    2015-08-01

    Three-dimensional (3D) cell manipulation is available with the integration of microfluidic technology and rapid prototyping techniques. High-Fidelity (Hi-Fi) constructs hold enormous therapeutic potential for organ manufacturing and regenerative medicine. In the present paper we introduced a quasi-three-dimensional (Q3D) model with parallel biocompatible alginate/gelatin/fibrin hurdles. The behaviors of fluids and cells along the microfluidic channels with various widths were studied. Cells inside the newly designed microfluidic channels attached and grew well. Morphological changes of adipose-derived stem cells (ADSCs) in both two-dimensional (2D) and 3D milieu were found on the printed constructs. Endothelialization occurred with the co-cultures of ADSCs and hepatocytes. This study provides insights into the interactions among fluids, cells and biomaterials, the behaviors of fluids and cells along the microfluidic channels, and the applications of Q3D techniques.

  7. Culture of Neural Stem Cells in Calcium-alginate Microbeads

    Institute of Scientific and Technical Information of China (English)

    Li-Song YAO; Tian-Qing LIU; Dan GE; Xue-Hu MA; Zhan-Feng CUI

    2005-01-01

    @@ 1 Introduction Recent research shows that neural stem cells may play an important role in the nerve injury reparation and nerve disease treatment. The shortage of the source and the number of NSCs, however, is the main challenge for its clinic application. In this situation, expansion of NSCs in large scale and culture in three dimensional environment are very worth of exploration. Notablely, the shear stress existed in bioreactors can cause serious cell injury especially for the shear sensitive cells like NSCs.

  8. Diffusion of Oxygen in Alginate Gels Related to the Kinetics of Methanol Oxidation by Immobilized Hansenula polymorpha Cells

    NARCIS (Netherlands)

    Hiemstra, Harry; Dijkhuizen, Lubbert; Harder, Willem

    1983-01-01

    In the yeast Hansenula polymorpha an oxygen-requiring enzyme, alcohol oxidase, catalyzes the conversion of methanol into formaldehyde. After growth on methanol cells of the organism were harvested and entrapped in barium-alginate gels. The diffusion of oxygen towards these cells is seriously hindere

  9. Chemistry and biocompatibility of alginate-PLL capsules for immunoprotection of mammalian cells

    NARCIS (Netherlands)

    de Vos, P; Hoogmoed, CG; Busscher, HJ

    Transplantation of encapsulated living cells is a promising approach for the treatment of a wide variety of diseases. Large-scale application of the technique, however, is hampered by insufficient biocompatibility of the capsules. In order to get means to study factors influencing the

  10. Chemistry and biocompatibility of alginate-PLL capsules for immunoprotection of mammalian cells

    NARCIS (Netherlands)

    de Vos, P; Hoogmoed, CG; Busscher, HJ

    2002-01-01

    Transplantation of encapsulated living cells is a promising approach for the treatment of a wide variety of diseases. Large-scale application of the technique, however, is hampered by insufficient biocompatibility of the capsules. In order to get means to study factors influencing the biocompatibili

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

    Science.gov (United States)

    Bünger, C M; Gerlach, C; Freier, T; Schmitz, K P; Pilz, M; Werner, C; Jonas, L; Schareck, W; Hopt, U T; de Vos, P

    2003-12-15

    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. Copyright 2003 Wiley Periodicals, Inc. J Biomed Mater Res 67A: 1219-1227, 2003

  12. Biochemical and structural characterization of neocartilage formed by mesenchymal stem cells in alginate hydrogels.

    Directory of Open Access Journals (Sweden)

    Magnus Ø Olderøy

    Full Text Available A popular approach to make neocartilage in vitro is to immobilize cells with chondrogenic potential in hydrogels. However, functional cartilage cannot be obtained by control of cells only, as function of cartilage is largely dictated by architecture of extracellular matrix (ECM. Therefore, characterization of the cells, coupled with structural and biochemical characterization of ECM, is essential in understanding neocartilage assembly to create functional implants in vitro. We focused on mesenchymal stem cells (MSC immobilized in alginate hydrogels, and used immunohistochemistry (IHC and gene expression analysis combined with advanced microscopy techniques to describe properties of cells and distribution and organization of the forming ECM. In particular, we used second harmonic generation (SHG microscopy and focused ion beam/scanning electron microscopy (FIB/SEM to study distribution and assembly of collagen. Samples with low cell seeding density (1e7 MSC/ml showed type II collagen molecules distributed evenly through the hydrogel. However, SHG microscopy clearly indicated only pericellular localization of assembled fibrils. Their distribution was improved in hydrogels seeded with 5e7 MSC/ml. In those samples, FIB/SEM with nm resolution was used to visualize distribution of collagen fibrils in a three dimensional network extending from the pericellular region into the ECM. In addition, distribution of enzymes involved in procollagen processing were investigated in the alginate hydrogel by IHC. It was discovered that, at high cell seeding density, procollagen processing and fibril assembly was also occurring far away from the cell surface, indicating sufficient transport of procollagen and enzymes in the intercellular space. At lower cell seeding density, the concentration of enzymes involved in procollagen processing was presumably too low. FIB/SEM and SHG microscopy combined with IHC localization of specific proteins were shown to provide

  13. Impact of carrier material on fermentative activity of encapsulated yoghurt culture in whey based substrate

    Directory of Open Access Journals (Sweden)

    Krunić Tanja Ž.

    2017-01-01

    Full Text Available The main objectives of this paper were to study the influence of the carrier material used for encapsulation and of bead size to fermentative activity and viability of the dairy starter culture ‘Lactoferm ABY 6’. Encapsulation of yoghurt culture in beads with diameter of 1mm provides better results than encapsulation in beads with larger diameter. Alginate beads and chitosan coated beads have proved to be a strong barrier for nutrients from substrate, so samples with those beads have lower viable cell count, lower titratable acidity and higher pH value after 5h of fermentation at 42°C, than samples with WPC-alginate beads. Also those beads have significantly (P < 0.05 lower cell leaking, than WPC-alginate beads and lower antioxidant capacity. Encapsulation of yoghurt culture in WPC-alginate carrier with diameter of approximately 1mm provided the best characteristics for fermented product. Samples with these beads have significantly (P < 0.05 higher increase of viable cell number after fermentation, despite of major cell leaking (19.7 %. Moreover, sample with these beads have the highest titratable acidity, the lowest pH value after fermentation (the best fermentative activity and the best antioxidant characteristics. [Projekat Ministarstva nauke Republike Srbije, br. TR 31017 i br. III 46010

  14. Incorporation of DMSO and dextran-40 into a gelatin/alginate hydrogel for controlled assembled cell cryopreservation.

    Science.gov (United States)

    Wang, Xiaohong; Xu, Huirong

    2010-12-01

    A new cell cryopreservation strategy for cell-assembling constructs was proposed. With this strategy, different concentrations of dimethysulfoxide (DMSO) and dextran-40 were directly incorporated into the cell/gelatin/alginate systems, prototyped according to a predesigned structure, cryopreserved at -80 °C for 10 days and followed a thawing process at 17 °C. The rheological properties, bonding water contents and melting points of the gelatin/alginate hydrogel systems were changed with the addition of different amounts of DMSO. The microscopy analysis, (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrasodium bromide (MTT) and hematoxylin and eosin (HE) staining indicated that the cell numbers were progressively in a selected DMSO concentration range. With DMSO 5% (v/v) alone, the metabolic rate in the construct attained (81.3±5.7)%. A synergistic effect was achieved with the combination of the DMSO/gelatin/alginate and dextran-40/gelatin/alginate hydrogel systems. These results indicated that the inclusion of DMSO and dextran-40 in the hydrogel could effectively enhance the cell preservation effects. This cryopreservation strategy holds the ability to be widely used in organ manufacturing techniques.

  15. Microencapsulation of probiotic bacteria Lactobacillus plantarum 15HN using alginate-psyllium-fenugreek polymeric blends.

    Science.gov (United States)

    Haghshenas, B; Abdullah, N; Nami, Y; Radiah, D; Rosli, R; Yari Khosroushahi, A

    2015-04-01

    Investigation on the use of herbal-based biopolymers for probiotic-Lactobacillus plantarum 15HN-encapsulation is presented. The objectives are to enhance its oral delivery, colonic release and survival rate of these probiotic cultures in gastrointestinal environment. Nine types of herbal-based polymers blend with different concentration of alginate alone or mixed with psyllium and fenugreek was used as candidate for encapsulation matrix by applying a simple extrusion method. All the blend formulations recorded high encapsulation efficiency at value >98%. The survival rate of viable probiotic cells under both low pH and high bile salt conditions was also high with value above 80% in 2% (w/v) alginate, alginate+psyllium (1·5 + 0·5%) blend and alginate+fenugreek (1·5 + 0·5%) blend as compared to other polymer formulations and nonencapsulated cells. Their release occurred after 2 h in colonic condition and sustained until the 12th hour incubation period. A value added prebiotic effect was observed in (1·5 + 0·5%) alginate-psyllium formulation. The high encapsulation efficiency, high viability of cell in low pH, high bile salt and the sustained release rates of probiotic cells in colonic condition during storage time was also observed for these herbal gel formulations. Herbal-based biopolymers offer added advantages of being prebiotic towards the enhancement of probiotic bacterial growth in the gastrointestinal environment. © 2015 The Society for Applied Microbiology.

  16. Effects of alginate hydrogel cross-linking density on mechanical and biological behaviors for tissue engineering.

    Science.gov (United States)

    Jang, Jinah; Seol, Young-Joon; Kim, Hyeon Ji; Kundu, Joydip; Kim, Sung Won; Cho, Dong-Woo

    2014-09-01

    An effective cross-linking of alginate gel was made through reaction with calcium carbonate (CaCO3). We used human chondrocytes as a model cell to study the effects of cross-linking density. Three different pore size ranges of cross-linked alginate hydrogels were fabricated. The morphological, mechanical, and rheological properties of various alginate hydrogels were characterized and responses of biosynthesis of cells encapsulated in each gel to the variation in cross-linking density were investigated. Desired outer shape of structure was maintained when the alginate solution was cross-linked with the applied method. The properties of alginate hydrogel could be tailored through applying various concentrations of CaCO3. The rate of synthesized GAGs and collagens was significantly higher in human chondrocytes encapsulated in the smaller pore structure than that in the larger pore structure. The expression of chondrogenic markers, including collagen type II and aggrecan, was enhanced in the smaller pore structure. It was found that proper structural morphology is a critical factor to enhance the performance and tissue regeneration.

  17. In Situ Transplantation of Alginate Bioencapsulated Adipose Tissues Derived Stem Cells (ADSCs via Hepatic Injection in a Mouse Model.

    Directory of Open Access Journals (Sweden)

    Mong-Jen Chen

    Full Text Available Adipose tissue derived stem cells (ADSCs transplantation has recently gained widespread enthusiasm, particularly in the perspective to use them as potential alternative cell sources for hepatocytes in cell based therapy, mainly because of their capability of hepatogenic differentiation in vitro and in vivo. But some challenges remain to be addressed, including whether ADSCs can be provided effectively to the target organ and whether subsequent proliferation of transplanted cells can be achieved. To date, intrasplenic injection is the conventional method to deliver ADSCs into the liver; however, a number of donor cells retained in the spleen has been reported. In this study, our objective is to evaluate a novel route to transplant ADSCs specifically to the liver. We aimed to test the feasibility of in situ transplantation of ADSCs by injecting bioencapsulated ADSCs into the liver in mouse model.The ADSCs isolated from human alpha 1 antitrypsin (M-hAAT transgenic mice were used to allow delivered ADSCs be readily identified in the liver of recipient mice, and alginate was selected as a cell carrier. We first evaluated whether alginate microspheres are implantable into the liver tissue by injection and whether ADSCs could migrate from alginate microspheres (study one. Once proven, we then examined the in vivo fate of ADSCs loaded microspheres in the liver. Specifically, we evaluated whether transplanted, undifferentiated ASDCs could be induced by the local microenvironment toward hepatogenic differentiation and the distribution of surviving ADSCs in major tissue organs (study two.Our results indicated ADSCs loaded alginate microspheres were implantable into the liver. Both degraded and residual alginate microspheres were observed in the liver up to three weeks. The viable ADSCs were detectable surrounding degraded and residual alginate microspheres in the liver and other major organs such as bone marrow and the lungs. Importantly, transplanted

  18. Proliferation and osteogenic differentiation of human bone marrow stromal cells on alginate-gelatine-hydroxyapatite scaffolds with anisotropic pore structure.

    Science.gov (United States)

    Bernhardt, A; Despang, F; Lode, A; Demmler, A; Hanke, T; Gelinsky, M

    2009-01-01

    Porous mineralized scaffolds are required for various applications in bone engineering. In particular, tube-like pores with controlled orientation inside the scaffold may support homogeneous cell seeding as well as sufficient nutrient supply and may facilitate blood vessel ingrowth. Scaffolds with parallely orientated tube-like pores were generated by diffusion-controlled ionotropic gelation of alginate. Incorporation of hydroxyapatite (HA) during the gelation process yielded stable scaffolds with an average pore diameter of approximately 90 microm. To evaluate the potential use of alginate-gelatine-HA scaffolds for bone tissue engineering, in vitro tests with human bone marrow stromal cells (hBMSCs) were carried out. We analysed biocompatibility and cell penetration into the capillary pores by microscopic methods. hBMSCs were also cultivated on alginate-gelatine-HA scaffolds for 3 weeks in the presence and absence of osteogenic supplements. We studied proliferation and osteogenic differentiation in terms of total lactate dehydrogenase (LDH) activity, DNA content and alkaline phosphatase (ALP) activity and found a 10-14-fold increase of cell number after 2 weeks of cultivation, as well as an increase of specific ALP activity for osteogenic-induced hBMSCs. Furthermore, the expression of bone-related genes [ALP, bone sialoprotein II (BSPII)] was analysed. We found an increase of ALP as well as BSPII expression for osteogenic-induced hBMSCs on alginate-gelatin-HA scaffolds. 2008 John Wiley & Sons, Ltd

  19. Enhancing cell migration in shape-memory alginate-collagen composite scaffolds: In vitro and ex vivo assessment for intervertebral disc repair.

    Science.gov (United States)

    Guillaume, Olivier; Naqvi, Syeda Masooma; Lennon, Kerri; Buckley, Conor Timothy

    2015-04-01

    Lower lumbar disc disorders pose a significant problem in an aging society with substantial socioeconomic consequences. Both inner tissue (nucleus pulposus) and outer tissue (annulus fibrosus) of the intervertebral disc are affected by such debilitating disorders and can lead to disc herniation and lower back pain. In this study, we developed an alginate-collagen composite porous scaffold with shape-memory properties to fill defects occurring in annulus fibrosus tissue of degenerated intervertebral discs, which has the potential to be administered using minimal invasive surgery. In the first part of this work, we assessed how collagen incorporation on preformed alginate scaffolds influences the physical properties of the final composite scaffold. We also evaluated the ability of annulus fibrosus cells to attach, migrate, and proliferate on the composite alginate-collagen scaffolds compared to control scaffolds (alginate only). In vitro experiments, performed in intervertebral disc-like microenvironmental conditions (low glucose and low oxygen concentrations), revealed that for alginate only scaffolds, annulus fibrosus cells agglomerated in clusters with limited infiltration and migration capacity. In comparison, for alginate-collagen scaffolds, annulus fibrosus cells readily attached and colonized constructs, while preserving their typical fibroblastic-like cell morphology with spreading behavior and intense cytoskeleton expression. In a second part of this study, we investigated the effects of alginate-collagen scaffold when seeded with bone marrow derived mesenchymal stem cells. In vitro, we observed that alginate-collagen porous scaffolds supported cell proliferation and extracellular matrix deposition (collagen type I), with secretion amplified by the local release of transforming growth factor-β3. In addition, when cultured in ex vivo organ defect model, alginate-collagen scaffolds maintained viability of transplanted mesenchymal stem cells for up to 5

  20. A Technology Platform to Test the Efficacy of Purification of Alginate

    Directory of Open Access Journals (Sweden)

    Genaro A. Paredes-Juarez

    2014-03-01

    Full Text Available Alginates are widely used in tissue engineering technologies, e.g., in cell encapsulation, in drug delivery and various immobilization procedures. The success rates of these studies are highly variable due to different degrees of tissue response. A cause for this variation in success is, among other factors, its content of inflammatory components. There is an urgent need for a technology to test the inflammatory capacity of alginates. Recently, it has been shown that pathogen-associated molecular patterns (PAMPs in alginate are potent immunostimulatories. In this article, we present the design and evaluation of a technology platform to assess (i the immunostimulatory capacity of alginate or its contaminants, (ii where in the purification process PAMPs are removed, and (iii which Toll-like receptors (TLRs and ligands are involved. A THP1 cell-line expressing pattern recognition receptors (PRRs and the co-signaling molecules CD14 and MD2 was used to assess immune activation of alginates during the different steps of purification of alginate. To determine if this activation was mediated by TLRs, a THP1-defMyD88 cell-line was applied. This cell-line possesses a non-functional MyD88 coupling protein, necessary for activating NF-κB via TLRs. To identify the specific TLRs being activated by the PAMPs, we use different human embryonic kidney (HEK cell-line that expresses only one specific TLR. Finally, specific enzyme-linked immunosorbent assays (ELISAs were applied to identify the specific PAMP. By applying this three-step procedure, we can screen alginate in a manner, which is both labor and cost efficient. The efficacy of the platform was evaluated with an alginate that did not pass our quality control. We demonstrate that this alginate was immunostimulatory, even after purification due to reintroduction of the TLR5 activating flagellin. In addition, we tested two commercially available purified alginates. Our experiments show that these commercial

  1. Cultivation and Differentiation of Encapsulated hMSC-TERT in a Disposable Small-Scale Syringe-Like Fixed Bed Reactor

    DEFF Research Database (Denmark)

    Weber, Christian; Pohl, Sebastian; Pörtner, Ralf

    2007-01-01

    The use of commercially available plastic syringes is introduced as disposable small-scale fixed bed bioreactors for the cultivation of implantable therapeutic cell systems on the basis of an alginate-encapsulated human mesenchymal stem cell line. The system introduced is fitted with a noninvasiv...... the fixed bed reactor an interesting option for GMP processes. The cultivation of the encapsulated cells in the fixed bed bioreactor system offered vitalities and adipogenic differentiation similar to well-mixed suspension cultures....

  2. The experimental study of polyelectrolyte coatings suitability for encapsulation of cells.

    Science.gov (United States)

    Granicka, L H; Antosiak-Iwańska, M; Godlewska, E; Hoser, G; Strawski, M; Szklarczyk, M; Dudziński, K

    2009-01-01

    Living cells encapsulated in polymeric shells are receiving increasing attention because of their possible biotechnological and biomedical applications. The aim of this work is to evaluate how different polyelectrolyte coatings, characterized by different numbers of polyelectrolyte layers and by different polyelectrolyte conformations, affect the viability of encapsulated biological material. We demonstrate the ability to individually encapsulate HL-60 cells as well as rat pancreatic islets within polymeric shells consisting of different PE layers using the layer-by-layer process. Coating of HL-60 cells allows for surviving and functioning of cells for all applied PE as well as for different numbers of layers. The islets encapsulated in applied polyelectrolytes exhibited the lower level of mitochondrial activity as compared to non-encapsulated islets. Nevertheless, encapsulated islets exhibited comparable absorbance values during the whole period of culture. Polyelectrolyte coating seems to be a promising way of allowing capsule void volume minimization in a model of encapsulated biological material for local production of biologically active substances.

  3. Encapsulated living choroid plexus cells: potential long-term treatments for central nervous system disease and trauma

    Science.gov (United States)

    Skinner, S. J. M.; Geaney, M. S.; Lin, H.; Muzina, M.; Anal, A. K.; Elliott, R. B.; Tan, P. L. J.

    2009-12-01

    In neurodegenerative disease and in acute brain injury, there is often local up-regulation of neurotrophin production close to the site of the lesion. Treatment by direct injection of neurotrophins and growth factors close to these lesion sites has repeatedly been demonstrated to improve recovery. It has therefore been proposed that transplanting viable neurotrophin-producing cells close to the trauma lesion, or site of degenerative disease, might provide a novel means for continuous delivery of these molecules directly to the site of injury or to a degenerative region. The aim of this paper is to summarize recent published information and present new experimental data that indicate that long-lasting therapeutic implants of choroid plexus (CP) neuroepithelium may be used to treat brain disease. CP produces and secretes numerous biologically active neurotrophic factors (NT). New gene microarray and proteomics data presented here indicate that many other anti-oxidant, anti-toxin and neuronal support proteins are also produced and secreted by CP cells. In the healthy brain, these circulate in the cerebrospinal fluid through the brain and spinal cord, maintaining neuronal networks and associated cells. Recent publications describe how transplanted CP cells and tissue, either free or in an immunoprotected encapsulated form, can effectively deliver therapeutic molecules when placed near the lesion or site of degenerative disease in animal models. Using simple techniques, CP neuroepithelial cell clusters in suspension culture were very durable, remaining viable for 6 months or more in vitro. The cell culture conditions had little effect on the wide range and activity of genes expressed and proteins secreted. Recently, completed experiments show that implanting CP within alginate-poly-ornithine capsules effectively protected these xenogeneic cells from the host immune system and allowed their survival for 6 months or more in the brains of rats, causing no adverse effects

  4. Effects of Accelerated Exposure Testing (AET) Conditions on Performance Degradation of Solar Cells and Encapsulants

    Energy Technology Data Exchange (ETDEWEB)

    Glick, S. H.; Pern, F. J.

    2000-01-01

    The paper briefly summarizes the results from several accelerated exposure tests (AET) studies. Causes responsible for the photothermal instability of the encapsulated Si solar cells appear to be multiple and complex.

  5. Rapid Fabrication of Cell-Laden Alginate Hydrogel 3D Structures by Micro Dip-Coating

    Science.gov (United States)

    Ghanizadeh Tabriz, Atabak; Mills, Christopher G.; Mullins, John J.; Davies, Jamie A.; Shu, Wenmiao

    2017-01-01

    Development of a simple, straightforward 3D fabrication method to culture cells in 3D, without relying on any complex fabrication methods, remains a challenge. In this paper, we describe a new technique that allows fabrication of scalable 3D cell-laden hydrogel structures easily, without complex machinery: the technique can be done using only apparatus already available in a typical cell biology laboratory. The fabrication method involves micro dip-coating of cell-laden hydrogels covering the surface of a metal bar, into the cross-linking reagents calcium chloride or barium chloride to form hollow tubular structures. This method can be used to form single layers with thickness ranging from 126 to 220 µm or multilayered tubular structures. This fabrication method uses alginate hydrogel as the primary biomaterial and a secondary biomaterial can be added depending on the desired application. We demonstrate the feasibility of this method, with survival rate over 75% immediately after fabrication and normal responsiveness of cells within these tubular structures using mouse dermal embryonic fibroblast cells and human embryonic kidney 293 cells containing a tetracycline-responsive, red fluorescent protein (tHEK cells). PMID:28286747

  6. Injectable hydrogels derived from phosphorylated alginic acid calcium complexes

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Han-Sem; Song, Minsoo, E-mail: minsoosong00@gmail.com; Lee, Eun-Jung; Shin, Ueon Sang, E-mail: usshin12@dankook.ac.kr

    2015-06-01

    Phosphorylation of sodium alginate salt (NaAlg) was carried out using H{sub 3}PO{sub 4}/P{sub 2}O{sub 5}/Et{sub 3}PO{sub 4} followed by acid–base reaction with Ca(OAc){sub 2} to give phosphorylated alginic acid calcium complexes (CaPAlg), as a water dispersible alginic acid derivative. The modified alginate derivatives including phosphorylated alginic acid (PAlg) and CaPAlg were characterized by nuclear magnetic resonance spectroscopy for {sup 1}H, and {sup 31}P nuclei, high resolution inductively coupled plasma optical emission spectroscopy, Fourier transform infrared spectroscopy, and thermogravimetric analysis. CaPAlg hydrogels were prepared simply by mixing CaPAlg solution (2 w/v%) with NaAlg solution (2 w/v%) in various ratios (2:8, 4:6, 6:4, 8:2) of volume. No additional calcium salts such as CaSO{sub 4} or CaCl{sub 2} were added externally. The gelation was completed within about 3–40 min indicating a high potential of hydrogel delivery by injection in vivo. Their mechanical properties were tested to be ≤ 6.7 kPa for compressive strength at break and about 8.4 kPa/mm for elastic modulus. SEM analysis of the CaPAlg hydrogels showed highly porous morphology with interconnected pores of width in the range of 100–800 μm. Cell culture results showed that the injectable hydrogels exhibited comparable properties to the pure alginate hydrogel in terms of cytotoxicity and 3D encapsulation of cells for a short time period. The developed injectable hydrogels showed suitable physicochemical and mechanical properties for injection in vivo, and could therefore be beneficial for the field of soft tissue engineering. - Highlights: • Preparation of water-soluble alginic acid complexes with calcium phosphate • Self-assembly of the phosphorylated alginic acid calcium complexes with sodium alginate • Preparation of injectable hydrogels with diverse gelation times within about 3–40 min.

  7. Effect of microencapsulation of Lactobacillus plantarum 25 into alginate/chitosan/alginate microcapsules on viability and cytokine induction.

    Science.gov (United States)

    Jiang, Tao; Kim, You-Kyoung; Singh, Bijay; Kang, Sang-Kee; Choi, Yun-Jaie; Cho, Chong-Su

    2013-08-01

    Lactobacillus plantarum 25 (LP25) encapsulated into alginate/chitosan/alginate (ACA) microcapsules (LP25-ACA MCs) prepared by an extrusion methods were characterized to assess their efficacy in oral delivery. The particle sizes of LP25-ACA MCs were 1.11 +/- 0.32 mm. The loading content of LP25 was 1.11 x 10(7) colony forming unit (cfu)/microcapsule and encapsulation efficiency was above 98%. The viability of LP25 in ACA MCs was more than 65% in simulated gastric fluid (SGF, pH 2.0) and 75% in simulated small intestinal fluid (SIF, pH 7.2) up to 2 h. Encapsulated LP25 were completely released from LP25-ACA MCs in SIF and simulated colon fluid (SCF, pH 6.0) within 12 h and 8 h respectively. The viability of LP25 in ACA MCs till 5 weeks was above 58%, whereas viability of free LP25 stored at room temperature up to 5 weeks was zero. Besides, LP25-ACA MCs induced the secretion of tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) from macrophages and dendritic cells showing the immunomodulatory effect of LP25. These findings demonstrate that the encapsulation of LP25 by ACA is a suitable strategy for oral delivery of probiotics.

  8. Effect of chitosan coating on a bacteria-based alginate microrobot.

    Science.gov (United States)

    Park, Sung Jun; Lee, Yu Kyung; Cho, Sunghoon; Uthaman, Saji; Park, In-Kyu; Min, Jung-Joon; Ko, Seong Young; Park, Jong-Oh; Park, Sukho

    2015-04-01

    To develop an efficient bacteria-based microrobot, first, therapeutic bacteria should be encapsulated into microbeads using biodegradable and biocompatible materials; second, the releasing rate of the encapsulated bacteria for theragnostic function should be regulated; and finally, flagellated bacteria should be attached on the microbeads to ensure the motility of the microrobot. For the therapeutic bacteria encapsulation, an alginate can be a promising candidate as a biodegradable and biocompatible material. Owing to the non-regulated releasing rate of the encapsulated bacteria in alginate microbeads and the weak attachment of flagellated bacteria on the surface of alginate microbeads, however, the alginate microbeads cannot be used as effective cargo for a bacteria-based microrobot. In this paper, to enhance the stability of the bacteria encapsulation and the adhesion of flagellated bacteria in alginate microbeads, we performed a surface modification of alginate microbeads using chitosan coating. The bacteria-encapsulated alginate microbeads with 1% chitosan coating maintained their structural integrity up to 72 h, whereas the control alginate microbead group without chitosan coating showed severe degradations after 24 h. The chitosan coating in alginate microbeads shows the enhanced attachment of flagellated bacteria on the surface of alginate microbeads. The bacteria-actuated microrobot with the enhanced flagellated bacteria attachment could show approximately 4.2 times higher average velocities than the control bacteria-actuated microrobot without chitosan coating. Consequently, the surface modification using chitosan coating enhanced the structural stability and the motility of the bacteria-based alginate microrobots.

  9. Encapsulated Lactococcus lactis with enhanced gastrointestinal survival for the development of folate enriched functional foods.

    Science.gov (United States)

    Divya, Jayakumar Beena; Nampoothiri, Kesavan Madhavan

    2015-01-01

    Two lactic acid bacteria (LAB) isolated from cow's milk were identified as Lactococcus lactis strains and designated as L. lactis CM22 and L. lactis CM28. They were immobilised by co-encapsulation using alginate and mannitol and by hybrid entrapment with skim milk, glycerol, CaCO3 and alginate. The encapsulated cells survived better in simulated gastrointestinal conditions compared to the free cells. The percentage survival of probiotics encapsulated by hybrid entrapment method was 62.74% for L. lactis CM22 and 68% for L. lactis CM28. Studies to check their efficacy in fermentative fortification of skim milk and ice cream revealed an enhancement in folate level. Copyright © 2015 Elsevier Ltd. All rights reserved.

  10. Perspectives of treatment of anemias with cells of fetal liver, immobilized in macroporous alginate-gelatin carriers

    Directory of Open Access Journals (Sweden)

    Gritsay D.V.

    2014-06-01

    Full Text Available Aim of the work was to study possibility of erythropoiesis stimulation by transplantation of fetal liver cells, seeded into macro¬porous carriers to the rats with post-hemorrhargic anemia, induced by 70% hepatectomy. Fetal liver cells (FLC were isolated from fetuses of rats with 15 days’ gestation and were cryopreserved. Decryopreserved FLC were seeded into macroporous spongy alginate-gelatin scaffolds, which were covered by alginate capsule and implanted into omentum of rats with modeled liver insufficiency. It was shown that fetal liver cells, immobilized in macroporous scaffolds after implantation have positive effect on red blood count and hemoglobin content, indicating that this approach is promising for the development of new methods of anemia treatment.

  11. Regulatory Considerations in Application of Encapsulated Cell Therapies

    NARCIS (Netherlands)

    van Zanten, J.; de Vos, Paul; Pedraz, JL; Orive, G

    2010-01-01

    The encapsulation of tissue in semi-permeable membranes is a technology with high potential and in due time several new therapies based on this technology will be tested in clinical trials. Recent, new legislation requires that these investigational medicinal products used in clinical trials Phase I

  12. Regulatory Considerations in Application of Encapsulated Cell Therapies

    NARCIS (Netherlands)

    van Zanten, J.; de Vos, Paul; Pedraz, JL; Orive, G

    2010-01-01

    The encapsulation of tissue in semi-permeable membranes is a technology with high potential and in due time several new therapies based on this technology will be tested in clinical trials. Recent, new legislation requires that these investigational medicinal products used in clinical trials Phase I

  13. Preparation of hydrogel hollow particles for cell encapsulation by a method of polyester core degradation.

    Science.gov (United States)

    Rabanel, J-M; Hildgen, P

    2004-06-01

    Implantation of encapsulated cells in particles of less than 1 mm (micro-encapsulation) has been proposed as a cell synthesized bio-molecule delivery system. Encapsulation provides immuno-isolation, protecting foreign cells from host immune system while nutrients, oxygen and therapeutic products can diffuse freely across capsule walls. A new method is described for the synthesis of a new family of hollow microparticles for cell encapsulation. Unlike other micro-encapsulation methods, encapsulation in those devices will take place after capsule synthesis, by micro-injection. The microcapsules were prepared by a three-steps original procedure: first, synthesis of a core particle, followed by coating with a layer of epichlorohydrin cross-linked amylo-pectin gel and, finally, selective degradation of the core particle to create the cavity. Initial experiments make use of amylo-pectin cross-linked with trimetaphosphate as core particle material. However, selective degradation was difficult to achieve. In further essays, polyesters were used successfully for the preparation of core particles. Optimizations were carried out and the permeability and morphology of the hollow particles were investigated. The preliminary results show that the new method has the potential to become a standard procedure to obtain hydrogel hollow particles. Moreover, the permeability study seems to be in accordance with specifications for immuno-isolation.

  14. An understanding of potential and limitations of alginate/PLL microcapsules as a cell retention system for perfusion cultures.

    Science.gov (United States)

    Demont, Aurelie; Cole, Harriet; Marison, Ian W

    2016-02-01

    Microcapsules for high cell density culture of mammalian cells have found an increasing interest, however, the poor stability of the microcapsules and the lack of characterisation methods led to few quantitative results. Alginate-poly-L-lysine (PLL) microcapsules have been studied in detail in order to form a basis for comparison of capsules made from different polymers. Since the microcapsules can be easily retained in the bioreactor without the need for a cell separation device, high cell densities were achieved with a maximum of 4 × 10(7) cell/ml(microcapsules), corresponding to a colonisation of 5% of the internal capsule volume. Measurement of microcapsule integrity and mechanical resistance showed that alginate-PLL microcapsules are not suitable for perfusion cultures since they are very sensitive to media composition, mainly the presence of non-gelling ions that have a higher affinity for alginate than PLL and Ca(2+), leading to the leakage of PLL and Ca(2+), and to microcapsule rupture.

  15. Human hepatoma cell lines on gas foaming templated alginate scaffolds for in vitro drug-drug interaction and metabolism studies.

    Science.gov (United States)

    Stampella, A; Rizzitelli, G; Donati, F; Mazzarino, M; de la Torre, X; Botrè, F; Giardi, M F; Dentini, M; Barbetta, A; Massimi, M

    2015-12-25

    Liver in vitro systems that allow reliable prediction of major human in vivo metabolic pathways have a significant impact in drug screening and drug metabolism research. In the present study, a novel porous scaffold composed of alginate was prepared by employing a gas-in-liquid foaming approach. Galactose residues were introduced on scaffold surfaces to promote cell adhesion and to enhance liver specific functions of the entrapped HepG2/C3A cells. Hepatoma cells in the gal-alginate scaffold showed higher levels of liver specific products (albumin and urea) and were more responsive to specific inducers (e.g. dexamethasone) and inhibitors (e.g. ketoconazole) of the CYP3A4 system than in conventional monolayer culture. HepG2/C3A cells were also more efficient in terms of rapid elimination of testosterone, used as a model substance, at rates comparable to those of in vivo excretion. In addition, an improvement in metabolism of testosterone, in terms of phase II metabolite formation, was also observed when the more differentiated HepaRG cells were used. Together the data suggest that hepatocyte/gas templated alginate-systems provide an innovative high throughput platform for in vitro drug metabolism and drug-drug interaction studies, with broad fields of application, and might provide a valid tool for minimizing animal use in preclinical testing of human relevance.

  16. Properties of Encapsulated CIGS Cells in 85 degrees C/85%RH

    Energy Technology Data Exchange (ETDEWEB)

    Olsen, Larry C.; Gross, Mark E.; Kundu, Sambhu N.; Shafaman, William N.

    2010-02-16

    This paper concerns studies of encapsulated cells subjected to an environment of 85ºC and 85%RH (85/85). Cells are encapsulated with PNNL multi-layer coatings (referred to as PML coatings) utilizing alternating layers of Al2O3, and an advanced polymer. The new polymer has been determined to withstand the 85/85 environment. Two types of cells were used for these studies, namely, SSI mini-modules (which are actually CIGSS devices) and CIGS cells provided by the Institute of Energy Conversion (IEC). Cells were coated and stressed at 85/85 in an environmental chamber. Current-voltage characteristics were acquired before and after coating, and periodically after being subjected to the 85/85 environment. Whereas coated SSI modules were determined to last 1000 hours when stressed at 60ºC/90%RH without degradation, the efficiency of these modules degrade to a level of 60% of the beginning-of-life value when stressed at 85/85. Encapsulated IEC cells, however, have exhibited extraordinary results. The efficiency of several encapsulated cells did not decrease for 1500 hours in an 85ºC/85%RH environment. This results establishes a benchmark for stressed, encapsulated CIGS cells.

  17. Shape-controlled fabrication of cell-laden calcium alginate-PLL hydrogel microcapsules by electrodeposition on microelectrode.

    Science.gov (United States)

    Chen, Weinan; Zhu, Bowen; Ma, Li; Hua, Xiaoqing

    2017-10-01

    In this study, we propose an electrodeposition method of fabricating shape-controlled calcium alginate-poly-L-lysine hydrogel microcapsules. The micro-patterned electrodes, which are produced by coating a patterned photoresist layer onto fluorine-doped tin oxide glass slide based on photolithography technique, are used for making different shapes of microcapsules. By the electrolysis of water in alginate gelation on micro-patterned anode electrode, the 2D alginate hydrogel structures embedded with yeast cells are formed on fluorine-doped tin oxide glass slide. Then, the 2D structures would be detached from the microelectrode surface and treated with given reagent to be transformed into 3D microcapsules while maintaining the ring and hexagon shapes. Finally, the yeast cells within the microcapsules are further promoted into compact tissues by cultivation. The experimental results indicate the method can successfully fabricate tissues which can maintain certain cells bioactivity after 24 h cultivation. The recommended method can lead to fabricating cell-laden scaffold for tissue engineering, biological studies and drug discovery with higher accuracy and efficiency.

  18. Sodium alginate and gum acacia hydrogels of ZnO nanoparticles show wound healing effect on fibroblast cells.

    Science.gov (United States)

    Raguvaran, R; Manuja, Balvinder K; Chopra, Meenu; Thakur, Rajesh; Anand, Taruna; Kalia, Anu; Manuja, Anju

    2017-03-01

    An ideal biomaterial for wound dressing applications should possess antibacterial and anti-inflammatory properties without any toxicity to the host cells while providing the maximum healing activity. Zinc oxide nanoparticles (ZnONPs) possess antimicrobial activity and enhance wound healing, but the questions regarding their safety arise before application to the biological systems. We synthesized ZnONPs-loaded-sodium alginate-gum acacia hydrogels (SAGA-ZnONPs) by cross linking hydroxyl groups of the polymers sodium alginate and gum acacia with the aldehyde group of gluteradehyde. Here, we report the wound healing properties of sodium alginate/gum acacia/ZnONPs, circumventing the toxicity of ZnONPs simultaneously. We demonstrated the concentration-dependent zones of inhibition in treated cultures of Pseudomonas aerigunosa and Bacillus cereus and biocompatability on peripheral blood mononuclear/fibroblast cells. SAGA-ZnONPs hydrogels showed a healing effect at a low concentration of ZnONPs using sheep fibroblast cells. Our findings suggest that high concentrations of ZnONPs were toxic to cells but SAGA-ZnONPs hydrogels significantly reduced the toxicity and preserved the beneficial antibacterial and healing effect. Copyright © 2016 Elsevier B.V. All rights reserved.

  19. Improvement of Biodesulfurization Rate of Alginate Immobilized Rhodococcus erythropolis R1

    OpenAIRE

    Derikvand, Peyman; Etemadifar, Zahra

    2014-01-01

    Background: Sulfur oxides released from the burning of oil causes severe environmental pollution. The sulfur can be removed via the 4S pathway in biodesulfurization (BDS). Immobilization approaches have been developed to prevent cell contamination of oil during the BDS process. Objectives: The encapsulation of Rhodococcus erythropolis R1 in calcium alginate beads was studied in order to enhance conversion of dibenzothiophene (DBT) to 2-hydroxy biphenyl (2-HBP) as the final product. Also the e...

  20. Interpenetrating Fibrin-Alginate Matrices for in vitro Ovarian Follicle Development

    OpenAIRE

    Shikanov, Ariella; Xu, Min; Woodruff, Teresa K.; Shea, Lonnie D.

    2009-01-01

    In this report, we investigate the fibrin-alginate inter penetrating network (FA-IPN) to provide dynamic cell-responsive mechanical properties, which we apply to the in vitro growth of ovarian follicles. The mechanical properties and polymerization rate of the gels were investigated by rheology, and the fiber structure was imaged by electron microscopy. Using a mouse model, two-layered secondary follicles were encapsulated in FA-IPNs, and growth, morphology, hormone production, fibrin degrada...

  1. Microencapsulation of a probiotic and prebiotic in alginate-chitosan capsules improves survival in simulated gastro-intestinal conditions.

    Science.gov (United States)

    Chávarri, María; Marañón, Izaskun; Ares, Raquel; Ibáñez, Francisco C; Marzo, Florencio; Villarán, María del Carmen

    2010-08-15

    Chitosan was used as a coating material to improve encapsulation of a probiotic and prebiotic in calcium alginate beads. Chitosan-coated alginate microspheres were produced to encapsulate Lactobacillus gasseri (L) and Bifidobacterium bifidum (B) as probiotics and the prebiotic quercetin (Q) with the objective of enhancing survival of the probiotic bacteria and keeping intact the prebiotic during exposure to the adverse conditions of the gastro-intestinal tract. The encapsulation yield for viable cells for chitosan-coated alginate microspheres with quercetin (L+Q and B+Q) was very low. These results, together with the study about the survival of microspheres with quercetin during storage at 4 degrees C, demonstrated that probiotic bacteria microencapsulated with quercetin did not survive. Owing to this, quercetin and L. gasseri or B. bifidum were microencapsulated separately. Microencapsulated L. gasseri and microencapsulated B. bifidum were resistant to simulated gastric conditions (pH 2.0, 2h) and bile solution (3%, 2h), resulting in significantly (p<0.05) improved survival when compared with free bacteria. This work showed that the microencapsulation of L. gasseri and B. bifidum with alginate and a chitosan coating offers an effective means of delivery of viable bacterial cells to the colon and maintaining their survival during simulated gastric and intestinal juice. Copyright 2010 Elsevier B.V. All rights reserved.

  2. Synthesis of collagen by bovine chondrocytes cultured in alginate; posttranslational modifications and cell-matrix interaction

    NARCIS (Netherlands)

    Beekman, B.; Verzijl, N.; Bank, R.A.; Von Der Mark, K.; TeKoppele, J.M.

    1997-01-01

    The extracellular matrix synthesized by articular chondrocytes cultured in alginate beads was investigated. Collagen levels increased sigmoidally with time and remained constant after 2 weeks of culture. The presence of cartilage-specific type II collagen was confirmed immunohistochemically.

  3. Cell proliferation on PVA/sodium alginate and PVA/poly(γ-glutamic acid) electrospun fiber.

    Science.gov (United States)

    Yang, Jen Ming; Yang, Jhe Hao; Tsou, Shu Chun; Ding, Chian Hua; Hsu, Chih Chin; Yang, Kai Chiang; Yang, Chun Chen; Chen, Ko Shao; Chen, Szi Wen; Wang, Jong Shyan

    2016-09-01

    To overcome the obstacles of easy dissolution of PVA nanofibers without crosslinking treatment and the poor electrospinnability of the PVA cross-linked nanofibers via electrospinning process, the PVA based electrospun hydrogel nanofibers are prepared with post-crosslinking method. To expect the electrospun hydrogel fibers might be a promising scaffold for cell culture and tissue engineering applications, the evaluation of cell proliferation on the post-crosslinking electrospun fibers is conducted in this study. At beginning, poly(vinyl alcohol) (PVA), PVA/sodium alginate (PVASA) and PVA/poly(γ-glutamic acid) (PVAPGA) electrospun fibers were prepared by electrospinning method. The electrospun PVA, PVASA and PVAPGA nanofibers were treated with post-cross-linking method with glutaraldehyde (Glu) as crosslinking agent. These electrospun fibers were characterized with thermogravimetry analysis (TGA) and their morphologies were observed with a scanning electron microscope (SEM). To support the evaluation and explanation of cell growth on the fiber, the study of 3T3 mouse fibroblast cell growth on the surface of pure PVA, SA, and PGA thin films is conducted. The proliferation of 3T3 on the electrospun fiber surface of PVA, PVASA, and PVAPGA was evaluated by seeding 3T3 fibroblast cells on these crosslinked electrospun fibers. The cell viability on electrospun fibers was conducted with water-soluble tetrazolium salt-1 assay (Cell Proliferation Reagent WST-1). The morphology of the cells on the fibers was also observed with SEM. The results of WST-1 assay revealed that 3T3 cells cultured on different electrospun fibers had similar viability, and the cell viability increased with time for all electrospun fibers. From the morphology of the cells on electrospun fibers, it is found that 3T3 cells attached on all electrospun fiber after 1day seeded. Cell-cell communication was noticed on day 3 for all electrospun fibers. Extracellular matrix (ECM) productions were found and

  4. Influence of epoxy resin as encapsulation material of silicon photovoltaic cells on maximum current

    Directory of Open Access Journals (Sweden)

    Acevedo-Gómez David

    2017-01-01

    Full Text Available This work presents an analysis about how the performance of silicon photovoltaic cells is influenced by the use of epoxy resin as encapsulation material with flat roughness. The effect of encapsulation on current at maximum power of mono-crystalline cell was tested indoor in a solar simulator bench at 1000 w/m² and AM1.5G. The results show that implementation of flat roughness layer onto cell surface reduces the maximum current inducing on average 2.7% less power with respect to a cell before any encapsulation. The losses of power and, in consequence, the less production of energy are explained by resin light absorption, reflection and partially neutralization of non-reflective coating.

  5. Effect of sodium-alginate and laminaran on Salmonella Typhimurium infection in human enterocyte-like HT-29-Luc cells and BALB/c mice.

    Science.gov (United States)

    Kuda, Takashi; Kosaka, Misa; Hirano, Shino; Kawahara, Miho; Sato, Masahiro; Kaneshima, Tai; Nishizawa, Makoto; Takahashi, Hajime; Kimura, Bon

    2015-07-10

    Brown algal polysaccharides such as alginate, polymers of uronic acids, and laminaran, beta-1,3 and 1,6-glucan, can be fermented by human intestinal microbiota. To evaluate the effects of these polysaccharides on infections caused by food poisoning pathogens, we investigated the adhesion and invasion of pathogens (Salmonella Typhimurium, Listeria monocytogenes and Vibrio parahaemolyticus) in human enterocyte-like HT-29-Luc cells and in infections caused in BALB/c mice. Both sodium Na-alginate and laminaran (0.1% each) inhibited the adhesion of the pathogens to HT-29-Luc cells by approximately 70-90%. The invasion of S. Typhimurium was also inhibited by approximately 70 and 80% by Na-alginate and laminaran, respectively. We observed that incubation with Na-alginate for 18 h increased the transepithelial electrical resistance of HT-29-Luc monolayer cells. Four days after inoculation with 7 log CFU/mouse of S. Typhimurium, the faecal pathogen count in mice that were not fed polysaccharides (control mice) was about 6.5 log CFU/g while the count in mice that were fed Na-alginate had decreased to 5.0 log CFU/g. The liver pathogen count, which was 4.1 log CFU/g in the control mice, was also decreased in mice that were fed Na-alginate. In contrast, the mice that were fed laminaran exhibited a more severe infection than that exhibited by control mice.

  6. Bio-inspired encapsulation and functionalization of living cells with artificial shells.

    Science.gov (United States)

    Wang, Sha; Guo, Zhiguang

    2014-01-01

    In nature, most single cells do not have structured shells to provide extensive protection apart from diatoms and radiolarians. Fabrication of biomimetic structures based on living cells encapsulated with artificial shells has a great impact on the area of cell-based sensors and devices as well as fundamental studies in cell biology. The past decade has witnessed a rapid increase of research concerning the new fabrication strategies, functionalization and applications of this kind of encapsulated cells. In this review, the latest fabrication strategies on how to encapsulate living cells with functional shells based on the diversity of artificial shells are discussed: hydrogel matrix shells, sol-gel shells, polymeric shells, and induced mineral shells. Classical different types of artificial shells are introduced and their advantages and disadvantages are compared and explained. The biomedical applications of encapsulated cells with particular emphasis on cell implant protection, cell separation, biosensors, cell therapy and tissue engineering are also described and a recap of this review and the future perspectives on these active areas is given finally.

  7. Cell-mediated remodeling of biomimetic encapsulating hydrogels triggered by adipogenic differentiation of adipose stem cells

    Science.gov (United States)

    Clevenger, Tracy N; Luna, Gabriel; Boctor, Daniel; Fisher, Steven K; Clegg, Dennis O

    2016-01-01

    One of the most common regenerative therapies is autologous fat grafting, which frequently suffers from unexpected volume loss. One approach is to deliver adipose stem cells encapsulated in the engineered hydrogels supportive of cell survival, differentiation, and integration after transplant. We describe an encapsulating, biomimetic poly(ethylene)-glycol hydrogel, with embedded peptides for attachment and biodegradation. Poly(ethylene)-glycol hydrogels containing an Arg–Gly–Asp attachment sequence and a matrix metalloprotease 3/10 cleavage site supported adipose stem cell survival and showed remodeling initiated by adipogenic differentiation. Arg–Gly–Asp–matrix metalloprotease 3/10 cleavage site hydrogels showed an increased number and area of lacunae or holes after adipose stem cell differentiation. Image analysis of adipose stem cells in Arg–Gly–Asp–matrix metalloprotease 3/10 cleavage site hydrogels showed larger Voronoi domains, while cell density remained unchanged. The differentiated adipocytes residing within these newly remodeled spaces express proteins and messenger RNAs indicative of adipocytic differentiation. These engineered scaffolds may provide niches for stem cell differentiation and could prove useful in soft tissue regeneration. PMID:27733898

  8. Biomimetic alginate/polyacrylamide porous scaffold supports human mesenchymal stem cell proliferation and chondrogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Peng [Department of ENT-Head and Neck Surgery, EENT Hospital, Shanghai 200031 (China); Shanghai Medical School, Fudan University, 210029 (China); Yuan, Yasheng, E-mail: yuanyasheng@163.com [Department of ENT-Head and Neck Surgery, EENT Hospital, Shanghai 200031 (China); Shanghai Medical School, Fudan University, 210029 (China); Eaton-Peabody Laboratory, Massachusetts Eye and Ear Infirmary, Harvard Medical School, Boston, MA 02114 (United States); Chi, Fanglu [Department of ENT-Head and Neck Surgery, EENT Hospital, Shanghai 200031 (China); Shanghai Medical School, Fudan University, 210029 (China)

    2014-09-01

    We describe the development of alginate/polyacrylamide (ALG/PAAm) porous hydrogels based on interpenetrating polymer network structure for human mesenchymal stem cell proliferation and chondrogenesis. Three ALG/PAAm hydrogels at molar ratios of 10/90, 20/80, and 30/70 were prepared and characterized with enhanced elastic and rubbery mechanical properties, which are similar to native human cartilage tissues. Their elasticity and swelling properties were also studied under different physiological pH conditions. Finally, in vitro tests demonstrated that human mesenchymal stem cells could proliferate on the as-synthesized hydrogels with improved alkaline phosphatase activities. These results suggest that ALG/PAAm hydrogels may be a promising biomaterial for cartilage tissue engineering. - Highlights: • ALG/PAAm hydrogels were prepared at different molar ratios for cartilage tissue engineering. • ALG/PAAm hydrogels feature an interpenetrating polymer network structure. • ALG/PAAm hydrogels demonstrate strengthened elastic and rubbery mechanical properties. • hMSCs could be cultured on the ALG/PAAm hydrogels for proliferation and chondrogenesis.

  9. Performance of polymer nano composite membrane electrode assembly using Alginate as a dopant in polymer electrolyte membrane fuel cell

    Science.gov (United States)

    Mulijani, S.

    2017-01-01

    Polymer membrane and composite polymer for membrane electrode assembly (MEAs) are synthesized and studied for usage in direct methanol fuel cell (DMFC). In this study, we prepared 3 type of MEAs, polystyrene (PS), sulfonated polystyrene (SPS) and composite polymer SPS-alginat membrane via catalyst hot pressed method. The performance and properties of prepared MEAs were evaluated and analyzed by impedance spectrometry and scanning electron microscopy (SEM). The result showed that, water up take of MEA composite polymer SPS-alginate was obtained higher than that in SPS and PS. The proton conductivity of MEA-SPS-alginate was also higher than that PS and PSS. SEM characterization revealed that the intimate contact between the carbon catalyst layers (CL) and the membranes, and the uniformly porous structure correlate positively with the MEAs prepared by hot pressed method, exhibiting high performances for DMFC.

  10. Performance of polymer nano composite membrane electrode assembly using Alginate as a dopant in polymer electrolyte membrane fuel cell

    Science.gov (United States)

    Mulijani, S.

    2016-11-01

    Polymer membrane and composite polymer for membrane electrode assembly (MEAs) are synthesized and studied for usage in direct methanol fuel cell (DMFC). In this study, we prepared 3 type of MEAs, polystyrene (PS), sulfonated polystyrene (SPS) and composite polymer SPS-alginat membrane via catalyst hot pressed method. The performance and properties of prepared MEAs were evaluated and analyzed by impedance spectrometry and scanning electron microscopy (SEM). The result showed that, water up take of MEA composite polymer SPS-alginate was obtained higher than that in SPS and PS. The proton conductivity of MEA-SPS-alginate was also higher than that PS and PSS. SEM characterization revealed that the intimate contact between the carbon catalyst layers (CL) and the membranes, and the uniformly porous structure correlate positively with the MEAs prepared by hot pressed method, exhibiting high performances for DMFC.

  11. Impact of culture conditions on β-carotene encapsulation using Yarrowia lipolytica cells

    Science.gov (United States)

    Dang, Tran Hai; Minh, Ho Thi Thu; Van Nhi, Tran Nguyen; Ngoc, Ta Thi Minh

    2017-09-01

    Yeast cell was reported as an effective natural preformed material for use in encapsulation of hydrophobic compounds. The encapsulation process was normally considered as passive transfer through cellular wall and cellular membrane. Beside solubility of hydrophobic compound in phospholipid membrane or plasmolysis, membrane characteristics of yeast cell which are differed between strains and influenced by culture conditions are main factors involving the accumulation of hydrophobic compound into yeast cell. In this study, the oleaginous yeast Yarrowia lipolytica was used as micro-container shell to encapsulate a high hydrophobic compound - β-carotene. Yeast cell was cultured under different conditions and wet yeast biomass was incubated with β-carotene which was dissolved in soybean oil overnight. β-carotene accumulation was then extracted and evaluated by UV-VIS spectrometry. Optimization of culture condition was investigated using the Box-Behnken model. β-carotene encapsulation efficiency in Y. lipolytica was showed to be affected by both pH of medium and agitation conditions. The highest β-carotene encapsulation efficiency was optimized at 42.8 μg/g with Y. lipolytica cultured at pH 4.5, medium volume equal to 115 ml and agitation speed at 211 rpm.

  12. Investigation of test methods, material properties, and processes for solar cell encapsulants. Annual report

    Energy Technology Data Exchange (ETDEWEB)

    Willis, P. B.; Baum, B.; White, R. A.

    1978-06-01

    Springborn Laboratories is engaged in a study of evaluating potentially useful encapsulating materials for the encapsulation task of the Low-Cost Solar Array project (LSA) funded by the Department of Energy. The goal of this program is to identify, evaluate, and recommend encapsulant materials (other than glass) and processes for the production of cost-effective, long-life photovoltaic solar modules. The results of an investigation of solar module encapsulation systems applicable to the Low-Cost Solar Array project 1986 cost and performance goals are presented. The 1986 cost goal for a 20 year life solar cell module is $0.50 per watt or $5 per square foot (in 1975 dollars). Out of this cost goal, $0.25 per square foot is currently allocated for the encapsulation in terms of raw materials, exclusive of labor. Assuming the flat-plate collector to be the most efficient module design, six basic construction elements were identified and their specific uses in module construction defined. In order to generate a comparative analysis, a uniform costing basis was established for each element. Extensive surveys into commercially available materials were then conducted in order to identify either general classes or specific products suitable for use for each construction element. The survey results were also useful in revealing price ranges for classes of materials and estimating the cost allocation for each element within the encapsulation cost goal.

  13. Alginate microencapsulated hepatocytes optimised for transplantation in acute liver failure.

    Directory of Open Access Journals (Sweden)

    Suttiruk Jitraruch

    Full Text Available BACKGROUND AND AIM: Intraperitoneal transplantation of alginate-microencapsulated human hepatocytes is an attractive option for the management of acute liver failure (ALF providing short-term support to allow native liver regeneration. The main aim of this study was to establish an optimised protocol for production of alginate-encapsulated human hepatocytes and evaluate their suitability for clinical use. METHODS: Human hepatocyte microbeads (HMBs were prepared using sterile GMP grade materials. We determined physical stability, cell viability, and hepatocyte metabolic function of HMBs using different polymerisation times and cell densities. The immune activation of peripheral blood mononuclear cells (PBMCs after co-culture with HMBs was studied. Rats with ALF induced by galactosamine were transplanted intraperitoneally with rat hepatocyte microbeads (RMBs produced using a similar optimised protocol. Survival rate and biochemical profiles were determined. Retrieved microbeads were evaluated for morphology and functionality. RESULTS: The optimised HMBs were of uniform size (583.5±3.3 µm and mechanically stable using 15 min polymerisation time compared to 10 min and 20 min (p<0.001. 3D confocal microscopy images demonstrated that hepatocytes with similar cell viability were evenly distributed within HMBs. Cell density of 3.5×10(6 cells/ml provided the highest viability. HMBs incubated in human ascitic fluid showed better cell viability and function than controls. There was no significant activation of PBMCs co-cultured with empty or hepatocyte microbeads, compared to PBMCs alone. Intraperitoneal transplantation of RMBs was safe and significantly improved the severity of liver damage compared to control groups (empty microbeads and medium alone; p<0.01. Retrieved RMBs were intact and free of immune cell adherence and contained viable hepatocytes with preserved function. CONCLUSION: An optimised protocol to produce GMP grade alginate-encapsulated

  14. Non-Ionic Highly Permeable Polymer Shells for Encapsulation of Living Cells

    Science.gov (United States)

    2011-05-01

    thermal gelation. Artif Cells Blood Substit Immobil Biotechnol 2004, 32 (2), 275-91. 2. Wilson, J. T.; Chaikof, E. L., Challenges and emerging...R.; Korsgren, O.; Nilsson, B., Islet surface heparinization prevents the instant blood -mediated inflammatory reaction in islet transplantation...R. J.; Poncelet, D., Encapsulation of brewers yeast in chitosan coated carrageenan microspheres by emulsification/thermal gelation. Artif Cells

  15. Cell-penetrating peptides meditated encapsulation of protein therapeutics into intact red blood cells and its application.

    Science.gov (United States)

    He, Huining; Ye, Junxiao; Wang, Yinsong; Liu, Quan; Chung, Hee Sun; Kwon, Young Min; Shin, Meong Cheol; Lee, Kyuri; Yang, Victor C

    2014-02-28

    Red blood cells (RBCs) based drug carrier appears to be the most appealing for protein drugs due to their unmatched biocompatability, biodegradability, and long lifespan in the circulation. Numerous methods for encapsulating protein drugs into RBCs were developed, however, most of them induce partial disruption of the cell membrane, resulting in irreversible alterations in both physical and chemical properties of RBCs. Herein, we introduce a novel method for encapsulating proteins into intact RBCs, which was meditated by a cell penetrating peptide (CPP) developed in our lab-low molecular weight protamine (LMWP). l-asparaginase, one of the primary drugs used in treatment of acute lymphoblastic leukemia (ALL), was chosen as a model protein to illustrate the encapsulation into erythrocytes mediated by CPPs. In addition current treatment of ALL using different l-asparaginase delivery and encapsulation methods as well as their associated problems were also reviewed.

  16. Biodegradable polymer based encapsulation of neem oil nanoemulsion for controlled release of Aza-A.

    Science.gov (United States)

    Jerobin, Jayakumar; Sureshkumar, R S; Anjali, C H; Mukherjee, Amitava; Chandrasekaran, Natarajan

    2012-11-06

    Azadirachtin a biological compound found in neem have medicinal and pesticidal properties. The present work reports on the encapsulation of neem oil nanoemulsion using sodium alginate (Na-Alg) by cross linking with glutaraldehyde. Starch and polyethylene glycol (PEG) were used as coating agents for smooth surface of beads. The SEM images showed beads exhibited nearly spherical shape. Swelling of the polymeric beads reduced with coating which in turn decreased the rate of release of Aza-A. Starch coated encapsulation of neem oil nanoemulsion was found to be effective when compared to PEG coated encapsulation of neem oil nanoemulsion. The release rate of neem Aza-A from the beads into an aqueous environment was analyzed by UV-visible spectrophotometer (214 nm). The encapsulated neem oil nanoemulsion have the potential for controlled release of Aza-A. Neem oil nanoemulsion encapsulated beads coated with PEG was found to be toxic in lymphocyte cells. Copyright © 2012 Elsevier Ltd. All rights reserved.

  17. 3D bioprinting of heterogeneous aortic valve conduits with alginate/gelatin hydrogels.

    Science.gov (United States)

    Duan, Bin; Hockaday, Laura A; Kang, Kevin H; Butcher, Jonathan T

    2013-05-01

    Heart valve disease is a serious and growing public health problem for which prosthetic replacement is most commonly indicated. Current prosthetic devices are inadequate for younger adults and growing children. Tissue engineered living aortic valve conduits have potential for remodeling, regeneration, and growth, but fabricating natural anatomical complexity with cellular heterogeneity remain challenging. In the current study, we implement 3D bioprinting to fabricate living alginate/gelatin hydrogel valve conduits with anatomical architecture and direct incorporation of dual cell types in a regionally constrained manner. Encapsulated aortic root sinus smooth muscle cells (SMC) and aortic valve leaflet interstitial cells (VIC) were viable within alginate/gelatin hydrogel discs over 7 days in culture. Acellular 3D printed hydrogels exhibited reduced modulus, ultimate strength, and peak strain reducing slightly over 7-day culture, while the tensile biomechanics of cell-laden hydrogels were maintained. Aortic valve conduits were successfully bioprinted with direct encapsulation of SMC in the valve root and VIC in the leaflets. Both cell types were viable (81.4 ± 3.4% for SMC and 83.2 ± 4.0% for VIC) within 3D printed tissues. Encapsulated SMC expressed elevated alpha-smooth muscle actin, while VIC expressed elevated vimentin. These results demonstrate that anatomically complex, heterogeneously encapsulated aortic valve hydrogel conduits can be fabricated with 3D bioprinting.

  18. Influence of viscosity and uronic acid composition of alginates on the properties of alginate films and microspheres produced by emulsification.

    Science.gov (United States)

    Lee, Huey Ying; Chan, Lai Wah; Dolzhenko, Anton V; Heng, Paul Wan Sia

    2006-12-01

    This study investigated the influence of viscosity and uronic acid composition of alginates on the properties of alginate films and microspheres produced by emulsification. Tensile properties of films were determined while the yield, size, drug contents and release characteristics of the microspheres were examined. Tensile properties of calcium alginate matrix were significantly affected by the orientation and arrangement of the polymer chains. High viscosity alginates gave rise to higher yields and bigger microspheres. Generally, microspheres with high drug content and slower rate of drug release had high Ca2+ contents and were produced from alginates of higher viscosity. Within an alginate microsphere batch, small sized microsphere fractions had higher drug contents but showed faster drug release rates. Microspheres having a defined size range revealed great dependence of encapsulation efficiency and drug release rates on viscosity and extent of Ca2+-alginate interaction. Viscosity appeared to exert a predominant influence on the microsphere properties.

  19. Improving the controlled delivery formulations of caffeine in alginate hydrogel beads combined with pectin, carrageenan, chitosan and psyllium.

    Science.gov (United States)

    Belščak-Cvitanović, Ana; Komes, Draženka; Karlović, Sven; Djaković, Senka; Spoljarić, Igor; Mršić, Gordan; Ježek, Damir

    2015-01-15

    Alginate-based blends consisting of carrageenan, pectin, chitosan or psyllium husk powder were prepared for assessment of the best formulation aimed at encapsulation of caffeine. Alginate-pectin blend exhibited the lowest viscosity and provided the smallest beads. Alginate-psyllium husk blend was characterised with higher viscosity, yielding the largest bead size and the highest caffeine encapsulation efficiency (83.6%). The release kinetics of caffeine indicated that the porosity of alginate hydrogel was not reduced sufficiently to retard the diffusion of caffeine from the beads. Chitosan coated alginate beads provided the most retarded release of caffeine in water. Morphological characteristics of beads encapsulating caffeine were adversely affected by freeze drying. Bitterness intensity of caffeine-containing beads in water was the lowest for alginate-psyllium beads and chitosan coated alginate beads. Higher sodium alginate concentration (3%) for production of hydrogel beads in combination with psyllium or chitosan coating would present the most favourable carrier systems for immobilization of caffeine.

  20. Encapsulated Whole Bone Marrow Cells Improve Survival in Wistar Rats after 90% Partial Hepatectomy

    Directory of Open Access Journals (Sweden)

    Carolina Uribe-Cruz

    2016-01-01

    Full Text Available Background and Aims. The use of bone marrow cells has been suggested as an alternative treatment for acute liver failure. In this study, we investigate the effect of encapsulated whole bone marrow cells in a liver failure model. Methods. Encapsulated cells or empty capsules were implanted in rats submitted to 90% partial hepatectomy. The survival rate was assessed. Another group was euthanized at 6, 12, 24, 48, and 72 hours after hepatectomy to study expression of cytokines and growth factors. Results. Whole bone marrow group showed a higher than 10 days survival rate compared to empty capsules group. Gene expression related to early phase of liver regeneration at 6 hours after hepatectomy was decreased in encapsulated cells group, whereas genes related to regeneration were increased at 12, 24, and 48 hours. Whole bone marrow group showed lower regeneration rate at 72 hours and higher expression and activity of caspase 3. In contrast, lysosomal-β-glucuronidase activity was elevated in empty capsules group. Conclusions. The results show that encapsulated whole bone marrow cells reduce the expression of genes involved in liver regeneration and increase those responsible for ending hepatocyte division. In addition, these cells favor apoptotic cell death and decrease necrosis, thus increasing survival.

  1. Different Patterns of Mast Cells Distinguish Diffuse from Encapsulated Neurofibromas in Patients with Neurofibromatosis 1

    Science.gov (United States)

    Tucker, Tracy; Riccardi, Vincent M.; Sutcliffe, Margaret; Vielkind, Juergen; Wechsler, Janine; Wolkenstein, Pierre; Friedman, Jan M.

    2011-01-01

    Multiple neurofibromas are cardinal features of neurofibromatosis 1 (NF1). Several different types of NF1-associated neurofibromas occur, each distinct in terms of pathological details, clinical presentation, and natural history. Mast cells are present in most neurofibromas and have been shown to be critical to the origin and progression of neurofibromas in both human NF1 and relevant mouse models. In this investigation, the authors determined whether mast cell involvement is the same for all types of NF1-associated neurofibromas. They examined the density and distribution of mast cells within 49 NF1-associated neurofibromas classified histopathologically as diffuse or encapsulated on the basis of the presence or absence of the perineurium or its constituent cells. They made two observations: (1) Diffuse neurofibromas had significantly higher densities of mast cells than did encapsulated neurofibromas, and (2) mast cells were evenly distributed throughout diffuse neurofibromas but were primarily restricted to the periphery of encapsulated neurofibromas. The differences in mast cell density and distribution differentiate the two basic types of NF1-associated neurofibromas, suggesting that the pathogenesis of diffuse and encapsulated neurofibromas may be significantly different. PMID:21525187

  2. Immobilization of Nicotiana tabacum plant cell suspensions within calcium alginate gel beads for the production of enhanced amounts of scopolin.

    Science.gov (United States)

    Gilleta; Roisin; Fliniaux; Jacquin-Dubreuil; Barbotin; Nava-Saucedo

    2000-02-01

    Scopolin-producing cells of Nicotiana tabacum were immobilized within Ca-alginate gel beads. Free cell suspensions accumulated scopolin within cytoplasmic compartments and cell disruption was necessary to recover scopolin. On the contrary, immobilized plant cells excreted considerable amounts of scopolin. Scopolin diffused throughout the gel matrix and reached the culture media. A large fraction of produced scopolin could then be recovered from the culture medium without disrupting cells. Immobilized N. tabacum cells produced more scopolin than free cell suspensions did (3.8 mg/g fresh weight biomass [into the culture media] versus 0.2 mg/g fresh weight biomass [intracellular]). Variation of the immobilization conditions revealed a marked influence on the behavior of N. tabacum plant cells: production of scopolin and enhanced excretion, cell growth, and morphological aspect of plant cell colonies. This excretion phenomenon could be used advantageously at an industrial production level.

  3. Exopolysaccharide Production and Prevention of Syneresis in Starch Using Encapsulated Probiotic Lactobacillus plantarum

    Directory of Open Access Journals (Sweden)

    Bindhumol Ismail

    2010-01-01

    Full Text Available Encapsulation of probiotic bacteria with a matrix can increase their survival rate by protecting them from adverse conditions and at the same time without affecting the production of metabolites. An effort has been made to encapsulate the probiotic Lactobacillus plantarum using calcium alginate. Box-Behnken model of response surface methodology (RSM was employed in the optimization of major encapsulation conditions such as concentration of sodium alginate, calcium chloride and curing time. The second-order quadratic model with the optimum conditions (sodium alginate 2 % (by mass per volume, calcium chloride 0.5 M and curing time 3 h resulted in a maximum titre of (0.9±0.1 g/L of exopolysaccharides (EPS at 72 h. The nearness of the coefficient of determination (R²=0.97 to 1 ensures the satisfactory adjustment of the quadratic model to the experimental data. The efficiency of EPS production by encapsulated cells was compared with free cells. The efficacy of secreted EPS in the prevention of syneresis in starch was investigated.

  4. Alginate Microencapsulation of Human Islets Does Not Increase Susceptibility to Acute Hypoxia

    Directory of Open Access Journals (Sweden)

    I. K. Hals

    2013-01-01

    Full Text Available Islet transplantation in diabetes is hampered by the need of life-long immunosuppression. Encapsulation provides partial immunoprotection but could possibly limit oxygen supply, a factor that may enhance hypoxia-induced beta cell death in the early posttransplantation period. Here we tested susceptibility of alginate microencapsulated human islets to experimental hypoxia (0.1–0.3% O2 for 8 h, followed by reoxygenation on viability and functional parameters. Hypoxia reduced viability as measured by MTT by 33.8±3.5% in encapsulated and 42.9±5.2% in nonencapsulated islets (P<0.2. Nonencapsulated islets released 37.7% (median more HMGB1 compared to encapsulated islets after hypoxic culture conditions (P<0.001. Glucose-induced insulin release was marginally affected by hypoxia. Basal oxygen consumption was equally reduced in encapsulated and nonencapsulated islets, by 22.0±6.1% versus 24.8±5.7%. Among 27 tested cytokines/chemokines, hypoxia increased the secretion of IL-6 and IL-8/CXCL8 in both groups of islets, whereas an increase of MCP-1/CCL2 was seen only with nonencapsulated islets. Conclusion. Alginate microencapsulation of human islets does not increase susceptibility to acute hypoxia. This is a positive finding in relation to potential use of encapsulation for islet transplantation.

  5. Pluronic F127 as a cell encapsulation material: utilization of membrane-stabilizing agents.

    Science.gov (United States)

    Khattak, Sarwat F; Bhatia, Surita R; Roberts, Susan C

    2005-01-01

    Thermoreversible gelation of the copolymer Pluronic F127 (generic name, poloxamer 407) in water makes it a unique candidate for cell encapsulation applications, either alone or to promote cell seeding and attachment in tissue scaffolds. At concentrations of 15-20% (w/w), aqueous Pluronic F127 (F127) solutions gel at physiological temperatures. The effect of F127 on viability and proliferation of human liver carcinoma cells (HepG2) was determined for both liquid and gel formulations. Cell concentration and viability over a 5-day period were measured by the trypan blue assay via hemocytometry and results were confirmed in both the MTT and LDH assays. With 0.1-5% (w/w) F127 (liquid), cells proliferated and maintained high viability over 5 days. However, at 10% (w/w) F127 (liquid), there was a significant decrease in cell viability and no cell proliferation was evident. HepG2 cell encapsulation in F127 concentrations ranging from 15 to 20% (w/w) (gel) resulted in complete cell death by 5 days. This was also true for the HMEC-1 (endothelial) and L6 (muscle) cell lines evaluated. Cell-seeding density did not affect cell survival or proliferation. Membrane-stabilizing agents (hydrocortisone, glucose, and glycerol) were added to the F127 gel formulations to improve cell viability. The steroid hydrocortisone demonstrated the most significant improvement in viability, from 70% (with 60 nM hydrocortisone added). These results suggest that F127 formulations supplemented with membrane-stabilizing agents can serve as viable cell encapsulation materials. In addition, hydrocortisone may be generally useful in the promotion of cell viability for a wide range of encapsulation materials.

  6. The promotion of in vitro vessel-like organization of endothelial cells in magnetically responsive alginate scaffolds.

    Science.gov (United States)

    Sapir, Yulia; Cohen, Smadar; Friedman, Gary; Polyak, Boris

    2012-06-01

    One of the major challenges in engineering thick, complex tissues such as cardiac muscle, is the need to pre-vascularize the engineered tissue in vitro to enable its efficient integration with host tissue upon implantation. Herein, we explored new magnetic alginate composite scaffolds to provide means of physical stimulation to cells. Magnetite-impregnated alginate scaffolds seeded with aortic endothelial cells stimulated during the first 7 days out of a total 14 day experimental course showed significantly elevated metabolic activity during the stimulation period. Expression of proliferating cell nuclear antigen (PCNA) indicated that magnetically stimulated cells had a lower proliferation index as compared to the non-stimulated cells. This suggests that the elevated metabolic activity could instead be related to cell migration and re-organization. Immunostaining and confocal microscopy analyses supported this observation showing that on day 14 in magnetically stimulated scaffolds without supplementation of any growth factors, cellular vessel-like (loop) structures, known as indicators of vasculogenesis and angiogenesis were formed as compared to cell sheets or aggregates observed in the non-stimulated (control) scaffolds. This work is the first step in our understanding of how to accurately control cellular organization to form tissue engineered constructs, which together with additional molecular signals could lead to a creation of an efficient pre-vascularized tissue construct with potential applicability for transplantation.

  7. Alginate-modifying enzymes: Biological roles and biotechnological uses

    Directory of Open Access Journals (Sweden)

    Helga eErtesvåg

    2015-05-01

    Full Text Available Alginate denotes a group of industrially important 1-4-linked biopolymers composed of the C-5-epimers β-D-mannuronic acid (M and α-L-guluronic acid (G. The polysaccharide is manufactured from brown algae where it constitutes the main structural cell wall polymer. The physical properties of a given alginate molecule, e.g. gel-strength, water-binding capacity, viscosity and biocompatibility, are determined by polymer length, the relative amount and distribution of G residues and the acetyl content, all of which are controlled by alginate modifying enzymes. Alginate has also been isolated from some bacteria belonging to the genera Pseudomonas and Azotobacter, and bacterially synthesized alginate may be O-acetylated at O-2 and/or O-3. Initially, alginate is synthesized as polymannuronic acid, and some M residues are subsequently epimerized to G residues. In bacteria a mannuronan C-5-epimerase (AlgG and an alginate acetylase (AlgX are integral parts of the protein complex necessary for alginate polymerisation and export. All alginate-producing bacteria use periplasmic alginate lyases to remove alginate molecules aberrantly released to the periplasm. Alginate lyases are also produced by organisms that utilize alginate as carbon source. Most alginate-producing organisms encode more than one mannuronan C-5 epimerase, each introducing its specific pattern of G residues. Acetylation protects against further epimerization and from most alginate lyases. One enzyme with alginate deacetylase activity from Pseudomonas syringae has been reported. Functional and structural studies reveal that alginate lyases and epimerases have related enzyme mechanisms and catalytic sites. Alginate lyases are now utilized as tools for alginate characterization. Secreted epimerases have been shown to function well in vitro, and have been engineered further in order to obtain enzymes that can provide alginates with new and desired properties for use in medical and

  8. Alginate-modifying enzymes: biological roles and biotechnological uses.

    Science.gov (United States)

    Ertesvåg, Helga

    2015-01-01

    Alginate denotes a group of industrially important 1-4-linked biopolymers composed of the C-5-epimers β-D-mannuronic acid (M) and α-L-guluronic acid (G). The polysaccharide is manufactured from brown algae where it constitutes the main structural cell wall polymer. The physical properties of a given alginate molecule, e.g., gel-strength, water-binding capacity, viscosity and biocompatibility, are determined by polymer length, the relative amount and distribution of G residues and the acetyl content, all of which are controlled by alginate modifying enzymes. Alginate has also been isolated from some bacteria belonging to the genera Pseudomonas and Azotobacter, and bacterially synthesized alginate may be O-acetylated at O-2 and/or O-3. Initially, alginate is synthesized as polymannuronic acid, and some M residues are subsequently epimerized to G residues. In bacteria a mannuronan C-5-epimerase (AlgG) and an alginate acetylase (AlgX) are integral parts of the protein complex necessary for alginate polymerization and export. All alginate-producing bacteria use periplasmic alginate lyases to remove alginate molecules aberrantly released to the periplasm. Alginate lyases are also produced by organisms that utilize alginate as carbon source. Most alginate-producing organisms encode more than one mannuronan C-5 epimerase, each introducing its specific pattern of G residues. Acetylation protects against further epimerization and from most alginate lyases. An enzyme from Pseudomonas syringae with alginate deacetylase activity has been reported. Functional and structural studies reveal that alginate lyases and epimerases have related enzyme mechanisms and catalytic sites. Alginate lyases are now utilized as tools for alginate characterization. Secreted epimerases have been shown to function well in vitro, and have been engineered further in order to obtain enzymes that can provide alginates with new and desired properties for use in medical and pharmaceutical applications.

  9. Cell Membrane Capsules for Encapsulation of Chemotherapeutic and Cancer Cell Targeting in Vivo.

    Science.gov (United States)

    Peng, Li-Hua; Zhang, Yuan-Hong; Han, Li-Jie; Zhang, Chen-Zhen; Wu, Jia-He; Wang, Xia-Rong; Gao, Jian-Qing; Mao, Zheng-Wei

    2015-08-26

    Systemic administration of chemotherapeutic agents can cause indiscriminate drug distribution and severe toxicity. Until now, encapsulation and targeting of drugs have typically relied on synthetic vehicles, which cannot minimize the clearance by the renal system and may also increase the risk of chemical side effects. Cell membrane capsules (CMCs) provide a generic and far more natural approach to the challenges of drug encapsulation and delivery in vivo. Here aptamer AS1411, which can recognize and bind overexpressed nucleolin on a cancer cell membrane, was chemically conjugated onto CMCs. As a result, AS1411 modified CMCs showed enhanced ingestion in certain cancer cells in vitro and accumulation in mouse cancer xenografts in vivo. Chemotherapeutics and contrast agents with therapeutically significant concentrations can be packaged into CMCs by reversible permeating their plasma membranes. The systematic administration of cancer targeting CMCs loaded with doxorubicin hydrochloride can significantly inhibit tumor growth in mouse xenografts, with significantly reduced toxicity compared to free drug. These findings suggest that cancer targeting CMCs may have considerable benefits in drug delivery and cancer treatment.

  10. Long-term survival of encapsulated GDNF secreting cells implanted within the striatum of parkinsonized rats.

    Science.gov (United States)

    Grandoso, Laura; Ponce, Sara; Manuel, Ivan; Arrúe, Aurora; Ruiz-Ortega, Jose A; Ulibarri, Isabel; Orive, Gorka; Hernández, Rosa M; Rodríguez, Alicia; Rodríguez-Puertas, Rafael; Zumárraga, Mercedes; Linazasoro, Gurutz; Pedraz, Jose Luis; Ugedo, Luisa

    2007-10-01

    Several findings suggest that glial cell line-derived neurotrophic factor (GDNF) may be a useful tool to treat parkinsonism by acting as a neuroprotective and neurotrophic factor for dopaminergic neurotransmission systems. In the present study, we implanted alginate-poly-L-lysine-alginate microcapsules containing immobilized Fischer rat 3T3 fibroblasts transfected to produce GDNF in vitro into the striatum of 6-hydroxydopamine (6-OHDA) lesioned rats. Microencapsulated GDNF secreting cells were stable for at least 3 weeks in vitro. Intrastriatal implantation of microencapsulated GDNF secreting cells into 6-OHDA lesioned rats resulted in a decrease in apomorphine-induced rotations by 84%, 64%, 84%, 60% and 52% (2, 5, 8, 16 and 24 weeks, respectively) with respect to the value before implantation and with respect to the value obtained from the empty microcapsule implanted-group at each time point. Six months after transplantation, immunohistochemical detection of GDNF revealed strong immunoreactivity in the striatal tissue surrounding the microcapsules in the absence of tissue damage due to microcapsule implantation. No changes in the levels of dopamine and its metabolites or of tyrosine hydroxylase immunoreactivity were detected in the striatum. In summary, the implantation of microencapsulated GDNF secreting cells allows the delivery of this molecule into the rat striatum for at least 6 months and results in substantial behavioral improvement.

  11. Patterned three-dimensional encapsulation of embryonic stem cells using dielectrophoresis and stereolithography.

    Science.gov (United States)

    Bajaj, Piyush; Marchwiany, Daniel; Duarte, Carlos; Bashir, Rashid

    2013-03-01

    Controlling the assembly of cells in three dimensions is very important for engineering functional tissues, drug screening, probing cell-cell/cell-matrix interactions, and studying the emergent behavior of cellular systems. Although the current methods of cell encapsulation in hydrogels can distribute them in three dimensions, these methods typically lack spatial control of multi-cellular organization and do not allow for the possibility of cell-cell contacts as seen for the native tissue. Here, we report the integration of dielectrophoresis (DEP) with stereolithography (SL) apparatus for the spatial patterning of cells on custom made gold micro-electrodes. Afterwards, they are encapsulated in poly (ethylene glycol) diacrylate (PEGDA) hydrogels of different stiffnesses. This technique can mimic the in vivo microscale tissue architecture, where the cells have a high degree of three dimensional (3D) spatial control. As a proof of concept, we show the patterning and encapsulation of mouse embryonic stem cells (mESCs) and C2C12 skeletal muscle myoblasts. mESCs show high viability in both the DEP (91.79% ± 1.4%) and the no DEP (94.27% ± 0.5%) hydrogel samples. Furthermore, we also show the patterning of mouse embryoid bodies (mEBs) and C2C12 spheroids in the hydrogels, and verify their viability. This robust and flexible in vitro platform can enable various applications in stem cell differentiation and tissue engineering by mimicking elements of the native 3D in vivo cellular micro-environment.

  12. Comparison of Different Encapsulating Adhesives to Enhance the Efficiencies and Lifetimes of Polymeric Solar Cells

    Science.gov (United States)

    Chung, Ming-Hua; Chen, Chen-Ming; Hsieh, Tsung-Eong; Tang, Rong-Ming; Tsai, Yu Sheng; Chu, Wei-Ping; Liu, Mark O.; Juang, Fuh-Shyang

    2009-04-01

    Polymeric solar cells (PSCs) with a derivative of C60 [[6,6]-phenyl C61-butyric acid methyl ester (PCBM)], and 3-hexylthiophene (P3HT) as active layers have been fabricated. The PSC devices were also packaged with glass and novel UV glues to improve their lifetimes and power conversion efficiencies (PCEs). After encapsulation with UV glue I, II, and III, the PCEs of PSCs reached 4, 4.82, and 6%, respectively, and their half-lifetimes increased to 16-18, 26-28, and 90 h, respectively, while the PCEs and half-lifetimes of PSCs without encapsulation were 3.76% and 2.5 h, respectively.

  13. Investigation of test methods, material properties, and processes for solar cell encapsulants. Annual report

    Energy Technology Data Exchange (ETDEWEB)

    Willis, P. B.; Baum, B.; Schnitzer, H. S.

    1980-07-01

    The goal of this program is to identify, evaluate, and recommend encapsulant materials and processes for the production of cost-effective, long-life solar cell modules. Technical activities during the past year have covered a number of topics and have emphasized the development of solar module encapsulation technology that employs ethylene/vinyl acetate, copolymer (EVA) as the pottant. These activities have included: (1) continued production of encapsulation grade EVA in sheet form to meet the needs of the photovoltaic industry; (2) investigations of three non-blocking techniques for EVA sheet; (3) performed an economic analysis of the high volume production of each pottant in order to estimate the large volume selling price (EVA, EPDM, aliphatic urethane, PVC plastisol, and butyl acrylate); (4) initiated an experimental corrosion protection program to determine if metal components could be successfully protected by encapsulation; (5) began an investigation to determine the maximum temperature which can be tolerated by the candidate pottant material in the event of hot spot heating or other temperature override; (6) continuation of surveys of potentially useful outer cover materials; and (7) continued with the accelerated artificial weathering of candidate encapsulation materials. Study results are presented. (WHK)

  14. Generation of continuous packed bed reactor with PVA-alginate blend immobilized Ochrobactrum sp. DGVK1 cells for effective removal of N,N-dimethylformamide from industrial effluents

    Energy Technology Data Exchange (ETDEWEB)

    Sanjeev Kumar, S.; Kumar, M. Santosh [Department of Biochemistry, Gulbarga University, Gulbarga 585106, Karnataka (India); Siddavattam, D. [Department of Animal Sciences, University of Hyderabad, Hyderabad 500046 (India); Karegoudar, T.B., E-mail: goudartbk@gmail.com [Department of Biochemistry, Gulbarga University, Gulbarga 585106, Karnataka (India)

    2012-01-15

    Highlights: Black-Right-Pointing-Pointer Removal of DMF was compared by free and immobilized cells of Ochrobactrum sp. DGVK1. Black-Right-Pointing-Pointer Ochrobactrum sp. DGVK1 cells entrapped in PVA-alginate have shown more tolerance. Black-Right-Pointing-Pointer PVA-alginate beads removed DMF even in the presence of other organic solvents. Black-Right-Pointing-Pointer Removal of DMF from industrial effluents by PVA-alginate blended batch operations. Black-Right-Pointing-Pointer Development of industrially feasible remediation strategy for DMF removal. - Abstract: Effective removal of dimethylformamide (DMF), the organic solvent found in industrial effluents of textile and pharma industries, was demonstrated by using free and immobilized cells of Ochrobactrum sp. DGVK1, a soil isolate capable of utilizing DMF as a sole source of carbon, nitrogen. The free cells have efficiently removed DMF from culture media and effluents, only when DMF concentration was less than 1% (v/v). Entrapment of cells either in alginate or in polyvinyl alcohol (PVA) failed to increase tolerance limits. However, the cells of Ochrobactrum sp. DGVK1 entrapped in PVA-alginate mixed matrix tolerated higher concentration of DMF (2.5%, v/v) and effectively removed DMF from industrial effluents. As determined through batch fermentation, these immobilized cells have retained viability and degradability for more than 20 cycles. A continuous packed bed reactor, generated by using PVA-alginate beads, efficiently removed DMF from industrial effluents, even in the presence of certain organic solvents frequently found in effluents along with DMF.

  15. Sodium alginate-cross-linked polymyxin B sulphate-loaded solid lipid nanoparticles: Antibiotic resistance tests and HaCat and NIH/3T3 cell viability studies.

    Science.gov (United States)

    Severino, Patrícia; Chaud, Marco V; Shimojo, Andrea; Antonini, Danilo; Lancelloti, Marcelo; Santana, Maria Helena A; Souto, Eliana B

    2015-05-01

    Polymyxins are a group of antibiotics with a common structure of a cyclic peptide with a long hydrophobic tail. Polymyxin B sulphate (PLX) has cationic charge, which is an obstacle for the efficient loading into Solid Lipid Nanoparticles (SLN). In the present paper, we describe an innovative method to load PLX into SLN to achieve the sustained release of the drug. PLX was firstly cross-linked with sodium alginate (SA) at different ratios (1:1, 1:2 and 1:3 SA/PLX), and loaded into SLN produced by high pressure homogenization (HPH). Optimized SLN were produced applying 500bar pressure and 5 homogenization cycles. The best results were obtained with SA/PLX (1:1), recording 99.08±1.2% for the association efficiency of the drug with SA, 0.99±10g for the loading capacity and 212.07±5.84% degree of swelling. The rheological profile of aqueous SA solution followed the typical behaviour of concentrated polymeric solutions, whereas aqueous SA/PLX solution exhibited a gel-like dynamic behaviour. Micrographs show that SA/PLX depicted a porous and discontinuous amorphous phase in different ratios. The encapsulation efficiency of SA/PLX (1:1) in SLN, the mean particle diameter, polydispersity index and zeta potential were, respectively, 82.7±5.5%; 439.5±20.42nm, 0.241±0.050 and -34.8±0.55mV. The effect of SLN on cell viability was checked in HaCat and NIH/3T3 cell lines, and the minimal inhibitory concentrations (MIC) were determined in Pseudomonas aeruginosa strains. SA/PLX-loaded SLN were shown to be less toxic than free PLX. Minimal inhibitory concentrations (MIC) showed the presence of the cross-linker polymer-drug complex, and SLN were shown to enhance MIC in the evaluated strains.

  16. TATVHL peptide-grafted alginate/poly(γ-glutamic acid) scaffolds with inverted colloidal crystal topology for neuronal differentiation of iPS cells.

    Science.gov (United States)

    Kuo, Yung-Chih; Chung, Chiu-Yen

    2012-12-01

    The neuronal differentiation of induced pluripotent stem (iPS) cells in scaffolding biomaterials is an emerging issue in nervous regeneration and repair. This study presents the production of neuron-lineage cells from iPS cells in inverted colloidal crystal (ICC) scaffolds comprising alginate, poly(γ-glutamic acid) (γ-PGA), and TATVHL peptide. The ability of iPS cells to differentiate toward neurons in the constructs was demonstrated by flow-cytometeric sorting and immunochemical staining. The results revealed that hexagonally arrayed microspheres molded alginate/γ-PGA hydrogel into ICC topology with adequate interconnected pores. An increase in the quantity of surface TATVHL peptide enhanced the atomic ratio of nitrogen and the adhesion efficiency of iPS cells in constructs. However, the effect of TATVHL peptide on the viability of iPS cells was insignificant. The adhesion and viability of iPS cells in ICC constructs was higher than those in freeform ones. TATVHL peptide raised the percentage of β III tubulin-identified cells differentiating from iPS cells, indicating that TATVHL peptide stimulated the neuronal development in alginate/γ-PGA ICC constructs. TATVHL peptide-grafted alginate/γ-PGA ICC scaffolds can be promising for establishing nerve tissue from iPS cells.

  17. Cell-laden poly(ɛ-caprolactone)/alginate hybrid scaffolds fabricated by an aerosol cross-linking process for obtaining homogeneous cell distribution: fabrication, seeding efficiency, and cell proliferation and distribution.

    Science.gov (United States)

    Lee, HyeongJin; Ahn, SeungHyun; Bonassar, Lawrence J; Chun, Wook; Kim, GeunHyung

    2013-10-01

    Generally, solid-freeform fabricated scaffolds show a controllable pore structure (pore size, porosity, pore connectivity, and permeability) and mechanical properties by using computer-aided techniques. Although the scaffolds can provide repeated and appropriate pore structures for tissue regeneration, they have a low biological activity, such as low cell-seeding efficiency and nonuniform cell density in the scaffold interior after a long culture period, due to a large pore size and completely open pores. Here we fabricated three different poly(ɛ-caprolactone) (PCL)/alginate scaffolds: (1) a rapid prototyped porous PCL scaffold coated with an alginate, (2) the same PCL scaffold coated with a mixture of alginate and cells, and (3) a multidispensed hybrid PCL/alginate scaffold embedded with cell-laden alginate struts. The three scaffolds had similar micropore structures (pore size = 430-580 μm, porosity = 62%-68%, square pore shape). Preosteoblast cells (MC3T3-E1) were used at the same cell density in each scaffold. By measuring cell-seeding efficiency, cell viability, and cell distribution after various periods of culturing, we sought to determine which scaffold was more appropriate for homogeneously regenerated tissues.

  18. Three-dimensional photopatterning of hydrogels using stereolithography for long-term cell encapsulation.

    Science.gov (United States)

    Chan, Vincent; Zorlutuna, Pinar; Jeong, Jae Hyun; Kong, Hyunjoon; Bashir, Rashid

    2010-08-21

    Cell-encapsulated hydrogels with complex three-dimensional (3D) structures were fabricated from photopolymerizable poly(ethylene glycol) diacrylate (PEGDA) using modified 'top-down' and 'bottoms-up' versions of a commercially available stereolithography apparatus (SLA). Swelling and mechanical properties were measured for PEGDA hydrogels with molecular weights (M(w)) ranging from 700 to 10 000 Daltons (Da). Long-term viability of encapsulated NIH/3T3 cells was quantitatively evaluated using an MTS assay and shown to improve over 14 days by increasing the M(w) of the hydrogels. Addition of adhesive RGDS peptide sequences resulted in increased cell viability, proliferation, and spreading compared to pristine PEG hydrogels of the same M(w). Spatial 3D layer-by-layer cell patterning was successfully demonstrated, and the feasibility of depositing multiple cell types and material compositions into distinct layers was established.

  19. Identification of enzymes responsible for extracellular alginate depolymerization and alginate metabolism in Vibrio algivorus.

    Science.gov (United States)

    Doi, Hidetaka; Tokura, Yuriko; Mori, Yukiko; Mori, Kenichi; Asakura, Yoko; Usuda, Yoshihiro; Fukuda, Hiroo; Chinen, Akito

    2017-02-01

    Alginate is a marine non-food-competing polysaccharide that has potential applications in biorefinery. Owing to its large size (molecular weight >300,000 Da), alginate cannot pass through the bacterial cell membrane. Therefore, bacteria that utilize alginate are presumed to have an enzyme that degrades extracellular alginate. Recently, Vibrio algivorus sp. SA2(T) was identified as a novel alginate-decomposing and alginate-utilizing species. However, little is known about the mechanism of alginate degradation and metabolism in this species. To address this issue, we screened the V. algivorus genomic DNA library for genes encoding polysaccharide-decomposing enzymes using a novel double-layer plate screening method and identified alyB as a candidate. Most identified alginate-decomposing enzymes (i.e., alginate lyases) must be concentrated and purified before extracellular alginate depolymerization. AlyB of V. algivorus heterologously expressed in Escherichia coli depolymerized extracellular alginate without requiring concentration or purification. We found seven homologues in the V. algivorus genome (alyB, alyD, oalA, oalB, oalC, dehR, and toaA) that are thought to encode enzymes responsible for alginate transport and metabolism. Introducing these genes into E. coli enabled the cells to assimilate soluble alginate depolymerized by V. algivorus AlyB as the sole carbon source. The alginate was bioconverted into L-lysine (43.3 mg/l) in E. coli strain AJIK01. These findings demonstrate a simple and novel screening method for identifying polysaccharide-degrading enzymes in bacteria and provide a simple alginate biocatalyst and fermentation system with potential applications in industrial biorefinery.

  20. Pseudomonas aeruginosa Aggregate Formation in an Alginate Bead Model System Exhibits In Vivo-Like Characteristics.

    Science.gov (United States)

    Sønderholm, Majken; Kragh, Kasper Nørskov; Koren, Klaus; Jakobsen, Tim Holm; Darch, Sophie E; Alhede, Maria; Jensen, Peter Østrup; Whiteley, Marvin; Kühl, Michael; Bjarnsholt, Thomas

    2017-05-01

    Alginate beads represent a simple and highly reproducible in vitro model system for diffusion-limited bacterial growth. In this study, alginate beads were inoculated with Pseudomonas aeruginosa and followed for up to 72 h. Confocal microscopy revealed that P. aeruginosa formed dense clusters similar in size to in vivo aggregates observed ex vivo in cystic fibrosis lungs and chronic wounds. Bacterial aggregates primarily grew in the bead periphery and decreased in size and abundance toward the center of the bead. Microsensor measurements showed that the O2 concentration decreased rapidly and reached anoxia ∼100 μm below the alginate bead surface. This gradient was relieved in beads supplemented with NO3(-) as an alternative electron acceptor allowing for deeper growth into the beads. A comparison of gene expression profiles between planktonic and alginate-encapsulated P. aeruginosa confirmed that the bacteria experienced hypoxic and anoxic growth conditions. Furthermore, alginate-encapsulated P. aeruginosa exhibited a lower respiration rate than the planktonic counterpart and showed a high tolerance toward antibiotics. The inoculation and growth of P. aeruginosa in alginate beads represent a simple and flexible in vivo-like biofilm model system, wherein bacterial growth exhibits central features of in vivo biofilms. This was observed by the formation of small cell aggregates in a secondary matrix with O2-limited growth, which was alleviated by the addition of NO3(-) as an alternative electron acceptor, and by reduced respiration rates, as well as an enhanced tolerance to antibiotic treatment.IMPORTANCEPseudomonas aeruginosa has been studied intensively for decades due to its involvement in chronic infections, such as cystic fibrosis and chronic wounds, where it forms biofilms. Much research has been dedicated to biofilm formation on surfaces; however, in chronic infections, most biofilms form small aggregates of cells not attached to a surface, but embedded in

  1. Potential Biomedical Application of Enzymatically Treated Alginate/Chitosan Hydrosols in Sponges—Biocompatible Scaffolds Inducing Chondrogenic Differentiation of Human Adipose Derived Multipotent Stromal Cells

    Directory of Open Access Journals (Sweden)

    Anna Zimoch-Korzycka

    2016-08-01

    Full Text Available Current regenerative strategies used for cartilage repair rely on biomaterial functionality as a scaffold for cells that may have potential in chondrogenic differentiation. The purpose of the research was to investigate the biocompatibility of enzymatically treated alginate/chitosan hydrosol sponges and their suitability to support chondrogenic differentiation of human adipose derived multipotent stromal cells (hASCs. The alginate/chitosan and enzyme/alginate/chitosan sponges were formed from hydrosols with various proportions and were used as a biomaterial in this study. Sponges were tested for porosity and wettability. The porosity of each sponge was higher than 80%. An equal dose of alginate and chitosan in the composition of sponges improved their swelling ability. It was found that equal concentrations of alginate and chitosan in hydrosols sponges assure high biocompatibility properties that may be further improved by enzymatic treatment. Importantly, the high biocompatibility of these biomaterials turned out to be crucial in the context of hydrosols’ pro-chondrogenic function. After exposure to the chondrogenic conditions, the hASCs in N/A/C and L/A/C sponges formed well developed nodules and revealed increased expression of collagen type II, aggrecan and decreased expression of collagen type I. Moreover, in these cultures, the reactive oxygen species level was lowered while superoxide dismutase activity increased. Based on the obtained results, we conclude that N/A/C and L/A/C sponges may have prospective application as hASCs carriers for cartilage repair.

  2. Enhanced bone tissue formation by alginate gel-assisted cell seeding in porous ceramic scaffolds and sustained release of growth factor.

    Science.gov (United States)

    Florczyk, Stephen J; Leung, Matthew; Jana, Soumen; Li, Zhensheng; Bhattarai, Narayan; Huang, Jerry I; Hopper, Richard A; Zhang, Miqin

    2012-12-01

    Increasing cell seeding efficiency in a tissue engineering construct can enhance cellular activity and tissue formation in vivo. Here, we demonstrate the use of alginate gel as a secondary phase material in 3D porous β-tricalcium phosphate scaffolds to improve cell seeding and provide controlled release of growth factors for bone tissue engineering. Cells were seeded in scaffolds in three ways: conventional seeding (CS), alginate gel-assisted seeding (GS), and alginate GS with bone morphogenetic protein-2 (BMP-2, GSB). In vitro study with MG-63 cells showed that cell seeding efficiency and cell population 1 week after seeding were significantly elevated in GS and GSB samples compared to CS samples. The GSB system demonstrated a sustained, steady release of BMP-2 over 2 weeks. In vivo, scaffolds seeded with rat mesenchymal stem cells were implanted ectopically into Sprague-Dawley rats for 8 weeks. GS and GSB samples exhibited improved osteogenic activity, with the GSB samples inducing the greatest osteocalcin and osteoid deposition. This study suggests that the alginate gel-assisted cell seeding increases seeding efficiency and allows for sustained release of growth factors. The use of the secondary phase polymer bolsters bone formation in vivo and has the potential for improving outcome in other tissue engineering applications.

  3. Controlled malolactic fermentation in cider using Oenococcus oeni immobilized in alginate beads and comparison with free cell fermentation.

    Science.gov (United States)

    Herrero; Laca; García; Díaz

    2001-01-02

    Cells of Oenococcus oeni (formerly Leuconostoc oenos) immobilized in alginate beads were used as starter culture to conduct malolactic fermentation in cider production. Concentrations of major organic acids and volatile compounds were monitored during the process, and results were compared to those obtained when using free cells in the same conditions. The rates of malic acid consumption were similar but lower ethanoic acid content and higher concentration of alcohols were detected with immobilized cells. These features have beneficial effects on the organoleptic properties of cider. A comparison between the kinetic behavior in immobilized and free cells, based on the data obtained for the malic acid consumption, has been developed solving the homogeneous diffusion model when it is applied to the system with immobilized cells.

  4. Efficient, air-stable colloidal quantum dot solar cells encapsulated using atomic layer deposition of a nanolaminate barrier

    KAUST Repository

    Ip, Alexander H.

    2013-12-23

    Atomic layer deposition was used to encapsulate colloidal quantum dot solar cells. A nanolaminate layer consisting of alternating alumina and zirconia films provided a robust gas permeation barrier which prevented device performance degradation over a period of multiple weeks. Unencapsulated cells stored in ambient and nitrogen environments demonstrated significant performance losses over the same period. The encapsulated cell also exhibited stable performance under constant simulated solar illumination without filtration of harsh ultraviolet photons. This monolithically integrated thin film encapsulation method is promising for roll-to-roll processed high efficiency nanocrystal solar cells. © 2013 AIP Publishing LLC.

  5. Production of isomaltulose obtained by Erwinia sp. cells submitted to different treatments and immobilized in calcium alginate

    Directory of Open Access Journals (Sweden)

    Haroldo Yukio Kawaguti

    2011-03-01

    Full Text Available In recent decades, there has been an increase in the studies of isomaltulose obtainment, due to its physicochemical properties and physiological health benefits. These properties, which include low cariogenicity, low glycemic index and greater stability, allow the use of this sweetener as a substitute for sucrose in foods; besides the fact that it can be converted to isomalt, a dietary non-cariogenic sugar alcohol used in pharmaceuticals as well as in the food industry. Isomaltulose (6-O-α-D-glucopyronosyl-1-6-D-fructofuranose is a disaccharide reducer obtained by the enzymatic conversion of sucrose - the α-glucosyltransferase enzyme. Different treatments were performed for the preparation of whole cells; lysed cells; and crude enzyme extract of Erwinia sp. D12 strain immobilized in calcium alginate. The packed bed column of granules, containing Erwinia sp. cells sonicated and immobilized in calcium alginate (CSI, reached a maximum conversion of 53-59% sucrose into isomaltulose and it presented activity for 480 hours. The converted syrup was purified and the isomaltulose crystallization was performed through the lowering of temperature. The isomaltulose crystals presented purity of 96.5%.

  6. Magnetic separation of encapsulated islet cells labeled with superparamagnetic iron oxide nano particles.

    Science.gov (United States)

    Mettler, Esther; Trenkler, Anja; Feilen, Peter J; Wiegand, Frederik; Fottner, Christian; Ehrhart, Friederike; Zimmermann, Heiko; Hwang, Yong Hwa; Lee, Dong Yun; Fischer, Stefan; Schreiber, Laura M; Weber, Matthias M

    2013-01-01

    Islet cell transplantation is a promising option for the restoration of normal glucose homeostasis in patients with type 1 diabetes. Because graft volume is a crucial issue in islet transplantations for patients with diabetes, we evaluated a new method for increasing functional tissue yield in xenogeneic grafts of encapsulated islets. Islets were labeled with three different superparamagnetic iron oxide nano particles (SPIONs; dextran-coated SPION, siloxane-coated SPION, and heparin-coated SPION). Magnetic separation was performed to separate encapsulated islets from the empty capsules, and cell viability and function were tested. Islets labeled with 1000 μg Fe/ml dextran-coated SPIONs experienced a 69.9% reduction in graft volume, with a 33.2% loss of islet-containing capsules. Islets labeled with 100 μg Fe/ml heparin-coated SPIONs showed a 46.4% reduction in graft volume, with a 4.5% loss of capsules containing islets. No purification could be achieved using siloxane-coated SPIONs due to its toxicity to the primary islets. SPION labeling of islets is useful for transplant purification during islet separation as well as in vivo imaging after transplantation. Furthermore, purification of encapsulated islets can also reduce the volume of the encapsulated islets without impairing their function by removing empty capsules.

  7. Electroporation of micro-droplet encapsulated HeLa cells in oil phase

    KAUST Repository

    Xiao, Kang

    2010-08-27

    Electroporation (EP) is a method widely used to introduce foreign genes, drugs or dyes into cells by permeabilizing the plasma membrane with an external electric field. A variety of microfluidic EP devices have been reported so far. However, further integration of prior and posterior EP processes turns out to be very complicated, mainly due to the difficulty of developing an efficient method for precise manipulation of cells in microfluidics. In this study, by means of a T-junction structure within a delicate microfluidic device, we encapsulated HeLa cells in micro-droplet of poration medium in oil phase before EP, which has two advantages: (i) precise control of cell-encapsulating droplets in oil phase is much easier than the control of cell populations or individuals in aqueous buffers; (ii) this can minimize the electrochemical reactions on the electrodes. Finally, we successfully introduced fluorescent dyes into the micro-droplet encapsulated HeLa cells in oil phase. Our results reflected a novel way to realize the integrated biomicrofluidic system for EP. © 2010 Wiley-VCH Verlag GmbH & Co. KGaA.

  8. Alginate Biosynthesis Factories in Pseudomonas fluorescens: Localization and Correlation with Alginate Production Level.

    Science.gov (United States)

    Maleki, Susan; Almaas, Eivind; Zotchev, Sergey; Valla, Svein; Ertesvåg, Helga

    2015-12-11

    Pseudomonas fluorescens is able to produce the medically and industrially important exopolysaccharide alginate. The proteins involved in alginate biosynthesis and secretion form a multiprotein complex spanning the inner and outer membranes. In the present study, we developed a method by which the porin AlgE was detected by immunogold labeling and transmission electron microscopy. Localization of the AlgE protein was found to depend on the presence of other proteins in the multiprotein complex. No correlation was found between the number of alginate factories and the alginate production level, nor were the numbers of these factories affected in an algC mutant that is unable to produce the precursor needed for alginate biosynthesis. Precursor availability and growth phase thus seem to be the main determinants for the alginate production rate in our strain. Clustering analysis demonstrated that the alginate multiprotein complexes were not distributed randomly over the entire outer cell membrane surface.

  9. Production Process for Stem Cell Based Therapeutic Implants: Expansion of the Production Cell Line and Cultivation of Encapsulated Cells

    Science.gov (United States)

    Weber, C.; Pohl, S.; Poertner, R.; Pino-Grace, Pablo; Freimark, D.; Wallrapp, C.; Geigle, P.; Czermak, P.

    Cell based therapy promises the treatment of many diseases like diabetes mellitus, Parkinson disease or stroke. Microencapsulation of the cells protects them against host-vs-graft reactions and thus enables the usage of allogenic cell lines for the manufacturing of cell therapeutic implants. The production process of such implants consists mainly of the three steps expansion of the cells, encapsulation of the cells, and cultivation of the encapsulated cells in order to increase their vitality and thus quality. This chapter deals with the development of fixed-bed bioreactor-based cultivation procedures used in the first and third step of production. The bioreactor system for the expansion of the stem cell line (hMSC-TERT) is based on non-porous glass spheres, which support cell growth and harvesting with high yield and vitality. The cultivation process for the spherical cell based implants leads to an increase of vitality and additionally enables the application of a medium-based differentiation protocol.

  10. Comparative study of bio-ethanol production from mahula (Madhuca latifolia L.) flowers by Saccharomyces cerevisiae cells immobilized in agar agar and Ca-alginate matrices

    Energy Technology Data Exchange (ETDEWEB)

    Behera, Shuvashish; Mohanty, Rama Chandra [Department of Botany, Utkal University, Vani Vihar, Bhubaneswar 751004, Orissa (India); Kar, Shaktimay; Ray, Ramesh Chandra [Microbiology Laboratory, Central Tuber Crops Research Institute (Regional Centre), Bhubaneswar 751019, Orissa (India)

    2010-01-15

    Batch fermentation of mahula (Madhuca latifolia L., a tree commonly found in tropical rain forest) flowers was carried out using immobilized cells (in agar agar and calcium alginate) and free cells of Saccharomyces cerevisiae. The ethanol yields were 151.2, 154.5 and 149.1 g kg{sup -1} flowers using immobilized (in agar agar and calcium alginate) and free cells, respectively. Cell entrapment in calcium alginate was found to be marginally superior to those in agar agar (2.2% more) as well as over free cell (3.5% more) as regard to ethanol yield from mahula flowers is concerned. Further, the immobilized cells were physiologically active at least for three cycles [150.6, 148.5 and 146.5 g kg{sup -1} (agar agar) and 152.8, 151.5 and 149.5 g kg{sup -1} flowers (calcium alginate) for first, second and third cycle, respectively] of ethanol fermentation without apparently lowering the productivity. Mahula flowers, a renewable, non-food-grade cheap carbohydrate substrate from non-agricultural environment such as forest can serve as an alternative to food grade sugar/starchy crops such as maize, sugarcane for bio-ethanol production. (author)

  11. Alginate/PEG based microcarriers with cleavable crosslinkage for expansion and non-invasive harvest of human umbilical cord blood mesenchymal stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Li, Chunge [Tianjin Key Laboratory of Composite and Functional Materials, School of Materials Science and Engineering, Tianjin University, No. 92, Weijin Road, Tianjin 300072 (China); Qian, Yufeng [Department of Chemistry and Biochemistry, University of Texas at Austin, 2500 Speedway, Austin, TX 78712 (United States); Zhao, Shuang [Tianjin Key Laboratory of Composite and Functional Materials, School of Materials Science and Engineering, Tianjin University, No. 92, Weijin Road, Tianjin 300072 (China); Yin, Yuji, E-mail: yinyuji@tju.edu.cn [Tianjin Key Laboratory of Composite and Functional Materials, School of Materials Science and Engineering, Tianjin University, No. 92, Weijin Road, Tianjin 300072 (China); Li, Junjie, E-mail: li41308@tju.edu.cn [Tianjin Key Laboratory of Composite and Functional Materials, School of Materials Science and Engineering, Tianjin University, No. 92, Weijin Road, Tianjin 300072 (China); Department of Advanced Interdisciplinary Studies, Institute of Basic Medical Sciences and Tissue Engineering Research Center, Academy of Military Medical Sciences, No. 27, Taiping Road, Beijing 100850 (China)

    2016-07-01

    Porous microcarriers are increasingly used to expand and harvest stem cells. Generally, the cells are harvested via proteolytic enzyme treatment, which always leads to damages to stem cells. To address this disadvantage, a series of alginate/PEG (AL/PEG) semi-interpenetrating network microcarriers are prepared in this study. In this AL/PEG system, the chemically cross-linked alginate networks are formed via the reaction between carboxylic acid group of alginate and di-terminated amine groups of cystamine. PEG is introduced to modulate the degradation of microcarriers, which does not participate in this cross-linked reaction, while it interpenetrates in alginate network via physical interactions. In addition, chitosan are coated on the surface of AL/PEG to improve the mechanical strength via the electrostatic interactions. Biocompatible fibronectin are also coated on these microcarriers to modulate the biological behaviors of cells seeded in microcarriers. Results suggest that the size of AL/PEG microcarriers can be modulated via adjusting the contents and molecular weight of PEG. Moreover, the microcarriers are designed to be degraded with cleavage of disulfide crosslinkage. By changing the type and concentration of reductant, the ratio of AL to PEG, and the magnitude of chitosan coating, the degradation ability of AL/PEG microcarriers can be well controlled. In addition, AL/PEG microcarriers can support the attachment and proliferation of human umbilical cord blood mesenchymal stem cells (hUCB-MSCs). More importantly, the expanded hUCB-MSCs can be detached from microcarriers after addition of reductant, which indeed reduce the cell damage caused by proteolytic enzyme treatment. Therefore, it is convinced that AL/PEG based microcarriers will be a promising candidate for large-scale expansion of hUCB-MSCs. - Graphical abstract: Alginate/PEG IPN microcarriers can support the attachment and expansion of hUCB-MSCs. More importantly, the expanded cells can be harvested

  12. Enhanced cellular activities of polycaprolactone/alginate-based cell-laden hierarchical scaffolds for hard tissue engineering applications.

    Science.gov (United States)

    Lee, HyeongJin; Kim, GeunHyung

    2014-09-15

    Biomedical scaffolds have been widely investigated because they are essential for support and promotion of cell adhesion, proliferation and differentiation in three-dimensional (3D) structures. An ideal scaffold should be highly porous to enable efficient nutrient and oxygen transfer and have a 3D structure that provides optimal micro-environmental conditions for the seeded cells to obtain homogeneous growth after a long culture period. In this study, new hierarchical osteoblast-like cell (MG-63)-laden scaffolds consisting of micro-sized struts/inter-layered micro-nanofibres and cell-laden hydrogel struts with mechanically stable and biologically superior properties were introduced. Poly(ethylene oxide) (PEO) was used as a sacrificial component to generate pores within the cell-laden hydrogel struts to attain a homogeneous cell distribution and rapid cell growth in the scaffold interior. The alginate-based cell-laden struts with PEO induced fast/homogeneous cell release, in contrast to nonporous cell-laden struts. Various weight fractions (0.5, 1, 2, 3 and 3.5 wt%) of PEO were used, of which 2 wt% PEO in the cell-laden strut resulted in the most appropriate cell release and enhanced biological activities (cell proliferation and calcium deposition), compared to nonporous cell-laden struts.

  13. Engineering alginate as bioink for bioprinting.

    Science.gov (United States)

    Jia, Jia; Richards, Dylan J; Pollard, Samuel; Tan, Yu; Rodriguez, Joshua; Visconti, Richard P; Trusk, Thomas C; Yost, Michael J; Yao, Hai; Markwald, Roger R; Mei, Ying

    2014-10-01

    Recent advances in three-dimensional (3-D) printing offer an excellent opportunity to address critical challenges faced by current tissue engineering approaches. Alginate hydrogels have been used extensively as bioinks for 3-D bioprinting. However, most previous research has focused on native alginates with limited degradation. The application of oxidized alginates with controlled degradation in bioprinting has not been explored. Here, a collection of 30 different alginate hydrogels with varied oxidation percentages and concentrations was prepared to develop a bioink platform that can be applied to a multitude of tissue engineering applications. The authors systematically investigated the effects of two key material properties (i.e. viscosity and density) of alginate solutions on their printabilities to identify a suitable range of material properties of alginates to be applied to bioprinting. Further, four alginate solutions with varied biodegradability were printed with human adipose-derived stem cells (hADSCs) into lattice-structured, cell-laden hydrogels with high accuracy. Notably, these alginate-based bioinks were shown to be capable of modulating proliferation and spreading of hADSCs without affecting the structure integrity of the lattice structures (except the highly degradable one) after 8days in culture. This research lays a foundation for the development of alginate-based bioink for tissue-specific tissue engineering applications.

  14. In vitro culture and oxygen consumption of NSCs in size-controlled neurospheres of Ca-alginate/gelatin microbead

    Energy Technology Data Exchange (ETDEWEB)

    Song, Kedong, E-mail: Kedongsong@dlut.edu.cn [State Key Laboratory of Fine Chemicals, Dalian R and D Center for Stem Cell and Tissue Engineering, Dalian University of Technology, Dalian 116024 (China); Yang, Yanfei; Li, Shixiao; Wu, Meiling; Wu, Yixing [State Key Laboratory of Fine Chemicals, Dalian R and D Center for Stem Cell and Tissue Engineering, Dalian University of Technology, Dalian 116024 (China); Lim, Mayasari [Division of Bioengineering, School of Chemical and Biomedical Engineering, Nanyang Technological University, Singapore (Singapore); Liu, Tianqing, E-mail: liutq@dlut.edu.cn [State Key Laboratory of Fine Chemicals, Dalian R and D Center for Stem Cell and Tissue Engineering, Dalian University of Technology, Dalian 116024 (China)

    2014-07-01

    Neural stem cells (NSCs) forming neurospheres in a conventional culture tend to develop necrotic/apoptotic centers due to mass transport limitations. In this study, the internal pore structure of calcium-alginate/gelatin (CAG) microbeads was tuned and controlled to provide a suitable three-dimensional environment supporting NSC proliferation. Direct impact of three-dimensional space availability was quantified by oxygen consumption rates of NSCs and cells were cultured in three different methods: neurospheres, single cell suspension of NSCs, and encapsulated NSCs in microbeads. Our results showed that encapsulated NSCs in CAG microbeads maintained higher cell viability than in conventional culture. In addition, NSCs encapsulated in CAG microbeads preserved their original stemness and continued to express nestin, CNPase, GFAP and β-tubulin-III post-encapsulation. Oxygen consumption rates of encapsulated NSCs in CAG microbeads were the lowest as compared to the other two culture methods. The optimal cell density supporting high cell proliferation in CAG microbeads was found to be 1.5 × 10{sup 5} cells/mL. The glucose consumption curve suggests that encapsulated NSCs in microbeads had a slower growth profile. This study presents an alternative method in hybrid microbead preparation to generate a highly favorable three-dimensional cell carrier for NSCs and was successfully applied for its effective in vitro expansion. - Highlights: • CAG microbeads effectively restricted the size of neurospheres.

  15. [Prediction of Encapsulation Temperatures of Copolymer Films in Photovoltaic Cells Using Hyperspectral Imaging Techniques and Chemometrics].

    Science.gov (United States)

    Lin, Ping; Chen, Yong-ming; Yao, Zhi-lei

    2015-11-01

    A novel method of combination of the chemometrics and the hyperspectral imaging techniques was presented to detect the temperatures of Ethylene-Vinyl Acetate copolymer (EVA) films in photovoltaic cells during the thermal encapsulation process. Four varieties of the EVA films which had been heated at the temperatures of 128, 132, 142 and 148 °C during the photovoltaic cells production process were used for investigation in this paper. These copolymer encapsulation films were firstly scanned by the hyperspectral imaging equipment (Spectral Imaging Ltd. Oulu, Finland). The scanning band range of hyperspectral equipemnt was set between 904.58 and 1700.01 nm. The hyperspectral dataset of copolymer films was randomly divided into two parts for the training and test purpose. Each type of the training set and test set contained 90 and 10 instances, respectively. The obtained hyperspectral images of EVA films were dealt with by using the ENVI (Exelis Visual Information Solutions, USA) software. The size of region of interest (ROI) of each obtained hyperspectral image of EVA film was set as 150 x 150 pixels. The average of reflectance hyper spectra of all the pixels in the ROI was used as the characteristic curve to represent the instance. There kinds of chemometrics methods including partial least squares regression (PLSR), multi-class support vector machine (SVM) and large margin nearest neighbor (LMNN) were used to correlate the characteristic hyper spectra with the encapsulation temperatures of of copolymer films. The plot of weighted regression coefficients illustrated that both bands of short- and long-wave near infrared hyperspectral data contributed to enhancing the prediction accuracy of the forecast model. Because the attained reflectance hyperspectral data of EVA materials displayed the strong nonlinearity, the prediction performance of linear modeling method of PLSR declined and the prediction precision only reached to 95%. The kernel-based forecast models were

  16. Engineering three-dimensional macroporous hydroxyethyl methacrylate-alginate-gelatin cryogel for growth and proliferation of lung epithelial cells.

    Science.gov (United States)

    Singh, Deepti; Zo, Sun Mi; Kumar, Ashok; Han, Sung Soo

    2013-01-01

    Three-dimensional (3D) growth of cell is of particular interest in the field of tissue engineering and regenerative medicine. Scaffolds used for this purpose are often tailor-made to mimic the microenvironment and the extracellular matrix of the tissue with defined role such as to provide appropriate structural, chemical, and mechanical support. The aim of the study was to design the macroporous matrix with potential in the field of tissue engineering especially for lung muscle regeneration. Blend of hydroxyethyl methacrylate-alginate-gelatin (HAG) cryogel scaffold was synthesized using cryogelation technique and this polymer material combination is being reported first time. The rheology study showed the elastic property of the material in wet state with no variation in storage modulus (G'), loss modulus (G″), and phase angle upon temperature variation. The microcomputer tomography (micro-CT) analysis confirmed the homogenous polymer structure with average pore diameter of 84 μm. Scaffold synthesized using polymer combinations which is mixture of polysaccharide (alginate) and protein (gelatin) provides supportive environment for human lung epithelial cell proliferation confirmed by cytoskeletal stain phalloidin and nuclei staining 4',6-diamidino-2-phenylindole checked for over three weeks. The in vivo biocompatibility was further performed which showed integration of scaffold to the surrounding tissue with ability to recruit cells. However, at first week, small amount of infiltrating mast cells were found which subsequently diminished in following weeks. Immunohistochemistry for dendritic cells confirmed in vivo biocompatible nature of the HAG scaffold. The mechanical strength, stiffness, elastic measurements, in vivo compatibility, and in vitro lung cell proliferation show the potentiality of HAG materials for lung tissue engineering.

  17. Preparation and in vitro evaluation of amoxicillin encapsulated in ...

    African Journals Online (AJOL)

    encapsulated in alginate-coated chitosan microparticles. Ebele Onuigbo* ... antibacterial property against Salmonella typhi and Staphylococcus aureus. Keywords: Amoxicillin .... detached was blotted dry with a filter paper and weighed.

  18. Fabrication of engineered heart tissue grafts from alginate/collagen barium composite microbeads

    Energy Technology Data Exchange (ETDEWEB)

    Bai, X P; Zheng, H X; Fang, R; Wang, T R; Li, Y; Tian, W M [Department of Life Science and Engineering, Harbin Institute of Technology, Harbin, 150080 (China); Hou, X L [The Fourth Affiliated Hospital of Harbin Medical University, Harbin, 150001 (China); Chen, X B, E-mail: tianweiming@gmail.com [Department of Mechanical Engineering, University of Saskatchewan, Saskatoon (Canada)

    2011-08-15

    Cardiac tissue engineering holds great promise for the treatment of myocardial infarction. However, insufficient cell migration into the scaffolds used and inflammatory reactions due to scaffold biodegradation remain as issues to be addressed. Engineered heart tissue (EHT) grafts fabricated by means of a cell encapsulation technique provide cells with a tissue-like environment, thereby potentially enhancing cellular processes such as migration, proliferation, and differentiation, and tissue regeneration. This paper presents a study on the fabrication and characterization of EHT grafts from novel alginate/collagen composite microbeads by means of cell encapsulation. Specifically, the microbeads were fabricated from alginate and collagen by barium ion cross-linking, with neonatal rat cardiomyocytes encapsulated in the composite microbeads during the fabrication of the EHT grafts. To evaluate the suitablity of these EHT grafts for heart muscle repair, the growth of cardiac cells in the microbeads was examined by means of confocal microscopy and staining with DAPI and F-actin. The EHT grafts were analyzed by scanning electron microscopy and transmission electron microscopy, and the contractile function of the EHT grafts monitored using a digital video camera at different time points. The results show the proliferation of cardiac cells in the microbeads and formation of interconnected multilayer heart-like tissues, the presence of well-organized and dense cell structures, the presence of intercalated discs and spaced Z lines, and the spontaneous synchronized contractility of EHT grafts (at a rate of 20-30 beats min{sup -1} after two weeks in culture). Taken together, these observations demonstrate that the novel alginate/collagen composite microbeads can provide a tissue-like microenvironment for cardiomyocytes that is suitable for fabricating native heart-like tissues.

  19. Investigation of test methods, material properties, and processes for solar cell encapsulants. Seventh annual report

    Energy Technology Data Exchange (ETDEWEB)

    Willis, P.B.

    1983-01-01

    The goal of the program is to identify and evaluate encapsulation materials and processes for the protection of silicon solar cells for service in a terrestrial environment. Aging and degradation studies were performed including: thermal aging, sunlamp exposures, aging in controlled environment reactors and outdoor photothermal aging devices, and metal catalyzed degradation. Other tests addressed water absorption, primers and adhesives, soiling experiments, and corrosion protection. (LEW)

  20. Gel-based optical waveguides with live cell encapsulation and integrated microfluidics.

    Science.gov (United States)

    Jain, Aadhar; Yang, Allen H J; Erickson, David

    2012-05-01

    In this Letter, we demonstrate a biocompatible microscale optical device fabricated from agarose hydrogel that allows for encapsulation of cells inside an optical waveguide. This allows for better interaction between the light in the waveguide and biology, since it can interact with the direct optical mode rather than the evanescent field. We characterize the optical properties of the waveguide and further incorporate a microfluidic channel over the optical structure, thus developing an integrated optofluidic system fabricated entirely from agarose gel.

  1. Accelerated Lifetime Testing of Organic-Inorganic Perovskite Solar Cells Encapsulated by Polyisobutylene.

    Science.gov (United States)

    Shi, Lei; Young, Trevor L; Kim, Jincheol; Sheng, Yun; Wang, Lei; Chen, Yifeng; Feng, Zhiqiang; Keevers, Mark J; Hao, Xiaojing; Verlinden, Pierre J; Green, Martin A; Ho-Baillie, Anita W Y

    2017-08-02

    Metal halide perovskite solar cells (PSCs) have undergone rapid progress. However, unstable performance caused by sensitivity to environmental moisture and high temperature is a major impediment to commercialization of PSCs. In the present work, a low-temperature, glass-glass encapsulation technique using high performance polyisobutylene (PIB) as the moisture barrier is investigated on planar glass/FTO/TiO2/FAPbI3/PTAA/gold perovskite solar cells. PIB was applied as either an edge seal or blanket layer. Electrical connections to the encapsulated PSCs were provided by either the FTO or Au layers. Results of a "calcium test" demonstrated that a PIB edge-seal effectively prevents moisture ingress. A shelf life test was performed and the PIB-sealed PSC was stable for at least 200 days. Damp heat and thermal cycling tests, in compliance with IEC61215:2016, were used to evaluate different encapsulation methods. Current-voltage measurements were performed regularly under simulated AM1.5G sunlight to monitor changes in PCE. The best results we have achieved to date maintained the initial efficiency after 540 h of damp heat testing and 200 thermal cycles. To the best of the authors' knowledge, these are among the best damp heat and thermal cycle test results for perovskite solar cells published to date. Given the modest performance of the cells (8% averaged from forward and reverse scans) especially with the more challenging FAPbI3 perovskite material tested in this work, it is envisaged that better stability results can be further achieved when higher performance perovskite solar cells are encapsulated using the PIB packaging techniques developed in this work. We propose that heat rather than moisture was the main cause of our PSC degradation. Furthermore, we propose that preventing the escape of volatile decomposition products from the perovskite solar cell materials is the key for stability. PIB encapsulation is a very promising packaging solution for perovskite solar

  2. Viral infection of human progenitor and liver-derived cells encapsulated in three-dimensional PEG-based hydrogel

    Energy Technology Data Exchange (ETDEWEB)

    Cho, Nam-Joon; Elazar, Menashe; Xiong, Anming; Glenn, Jeffrey S [Department of Medicine, Division of Gastroenterology and Hepatology, Stanford University School of Medicine, CCSR Building Room 3115A, 269 Campus Drive, Stanford, CA 94305 (United States); Lee, Wonjae [Mechanical Engineering, Stanford University, Stanford, CA 94305 (United States); Chiao, Eric; Baker, Julie [Department of Genetics, Stanford University School of Medicine, Stanford, CA 94305 (United States); Frank, Curtis W, E-mail: jeffrey.glenn@stanford.ed, E-mail: curt.frank@stanford.ed [Department of Chemical Engineering, Stanford University, Stanford, CA 94305 (United States)

    2009-02-15

    We have studied the encapsulation of human progenitor cells into 3D PEG hydrogels. Replication-incompetent lentivirus promoter reporter vectors were found to efficiently detect the in vivo expression of human hepatic genes in hydrogel-encapsulated liver progenitor cells. Similarly, hydrogel-encapsulated cells could be efficiently infected with hepatitis C virus, and progeny infectious virus could be recovered from the media supernatants of the hydrogels. Provocatively, the diameters of these virus particles range from {approx}50 to 100 nm, while the calculated mesh size of the 8 k hydrogel is 44.6 +- 1.7 A. To reconcile how viral particles can penetrate the hydrogels to infect the encapsulated cells, we propose that microfractures/defects of the hydrogel result in a functional pore size of up to 20 fold greater than predicted by theoretical mesh calculations. These results suggest a new model of hydrogel structure, and have exciting implications for tissue engineering and hepatitis virus studies. (communication)

  3. Microencapsulation of a probiotic bacteria with alginate-gelatin and its properties.

    Science.gov (United States)

    Li, Xiao Yan; Chen, Xi Guang; Cha, Dong Su; Park, Hyun Jin; Liu, Cheng Sheng

    2009-06-01

    Lactobacillus casei ATCC 393-loaded microcapsules based on alginate and gelatin had been prepared by extrusion method and the product could increase the cell numbers of L. casei ATCC 393 to be 10(7) CFU g(-1) in the dry state of microcapsules. The microparticles homogeneously distributed with size of 1.1 ± 0.2 mm. Four kinds of microcapsules (S(1), S(2), S(3) and S(4)) exhibited swelling in simulated gastric fluid (SGF) while the beads eroded and disintegrated rapidly in simulated intestinal fluid (SIF). Cells of L. casei ATCC 393 could be continuously released from the microcapsules during simulated gastrointestinal tract (GIT) and the release amounts and speeds in SIF were much higher and faster than that in SGF. Encapsulation in alginate-gelatin microcapsules successfully improved the survival of L. casei ATCC 393 and this approach might be useful in delivery of probiotic cultures as a functional food.

  4. Efficient functionalization of alginate biomaterials.

    Science.gov (United States)

    Dalheim, Marianne Ø; Vanacker, Julie; Najmi, Maryam A; Aachmann, Finn L; Strand, Berit L; Christensen, Bjørn E

    2016-02-01

    Peptide coupled alginates obtained by chemical functionalization of alginates are commonly used as scaffold materials for cells in regenerative medicine and tissue engineering. We here present an alternative to the commonly used carbodiimide chemistry, using partial periodate oxidation followed by reductive amination. High and precise degrees of substitution were obtained with high reproducibility, and without formation of by-products. A protocol was established using l-Tyrosine methyl ester as a model compound and the non-toxic pic-BH3 as the reducing agent. DOSY was used to indirectly verify covalent binding and the structure of the product was further elucidated using NMR spectroscopy. The coupling efficiency was to some extent dependent on alginate composition, being most efficient on mannuronan. Three different bioactive peptide sequences (GRGDYP, GRGDSP and KHIFSDDSSE) were coupled to 8% periodate oxidized alginate resulting in degrees of substitution between 3.9 and 6.9%. Cell adhesion studies of mouse myoblasts (C2C12) and human dental stem cells (RP89) to gels containing various amounts of GRGDSP coupled alginate demonstrated the bioactivity of the material where RP89 cells needed higher peptide concentrations to adhere.

  5. Effect of microcavitary alginate hydrogel with different pore sizes on chondrocyte culture for cartilage tissue engineering

    Energy Technology Data Exchange (ETDEWEB)

    Zeng, Lei; Yao, Yongchang [School of Materials Science and Engineering, South China University of Technology, Guangzhou 510641 (China); National Engineering Research Center for Tissue Restoration and Reconstruction, Guangzhou 510006 (China); Wang, Dong-an, E-mail: DAWang@ntu.edu.sg [National Engineering Research Center for Tissue Restoration and Reconstruction, Guangzhou 510006 (China); Division of Bioengineering, School of Chemical and Biomedical Engineering, Nanyang Technological University, Singapore 637457 (Singapore); Chen, Xiaofeng, E-mail: chenxf@scut.edu.cn [School of Materials Science and Engineering, South China University of Technology, Guangzhou 510641 (China); National Engineering Research Center for Tissue Restoration and Reconstruction, Guangzhou 510006 (China)

    2014-01-01

    In our previous work, a novel microcavitary hydrogel was proven to be effective for proliferation of chondrocytes and maintenance of chondrocytic phenotype. In present work, we further investigated whether the size of microcavity would affect the growth and the function of chondrocytes. By changing the stirring rate, gelatin microspheres in different sizes including small size (80–120 μm), middle size (150–200 μm) and large size (250–300 μm) were prepared. And then porcine chondrocytes were encapsulated into alginate hydrogel with various sizes of gelatin microspheres. Cell Counting Kit-8 (CCK-8), Live/dead staining and real-time PCR were used to analyze the effect of the pore size on cell proliferation and expression of specific chondrocytic genes. According to all the data, cells cultivated in microcavitary hydrogel, especially in small size, had preferable abilities of proliferation and higher expression of cartilaginous markers including type II collagen, aggrecan and cartilage oligomeric matrix protein (COMP). Furthermore, it was shown by western blot assay that the culture of chondrocytes in microcavitary hydrogel could improve the proliferation of cells potentially by inducing the Erk1/2-MAPK pathway. Taken together, this study demonstrated that chondrocytes favored microcavitary alginate hydrogel with pore size within the range of 80–120 μm for better growth and ECM synthesis, in which Erk1/2 pathway was involved. This culture system would be promising for cartilage tissue engineering. - Highlights: • A novel model with microcavitary structure was set up to study the interaction between cells and materials. • Microcavitary alginate hydrogel could enhance the proliferation of chondrocytes and promote the expression of cartilaginous genes as compared with plain alginate hydrogel. • Cells in microcavitary alginate hydrogel with pore size within the range of 80–120 μm were capable of better growth and ECM synthesis.

  6. Lactic acid fermentation by cells immobilised on various porous cellulosic materials and their alginate/poly-lactic acid composites.

    Science.gov (United States)

    Kumar, Mrinal Nishant; Gialleli, Angelika-Ioanna; Masson, Jean Bernard; Kandylis, Panagiotis; Bekatorou, Argyro; Koutinas, Athanasios A; Kanellaki, Maria

    2014-08-01

    Porous delignified cellulose (or tubular cellulose, abbr. TC) from Indian Mango (Mangifera indica) and Sal (Shorea robusta) wood and Rice husk, and TC/Ca-alginate/polylactic acid composites, were used as Lactobacillus bulgaricus immobilisation carriers leading to improvements in lactic acid fermentation of cheese whey and synthetic lactose media, compared to free cells. Specifically, shorter fermentation rates, higher lactic acid yields (g/g sugar utilised) and productivities (g/Ld), and higher amounts of volatile by-products were achieved, while no significant differences were observed on the performance of the different immobilised biocatalysts. The proposed biocatalysts are of food grade purity, cheap and easy to prepare, and they are attractive for bioprocess development based on immobilised cells. Such composite biocatalysts may be used for the co-immobilisation of different microorganisms or enzymes (in separate layers of the biocatalyst), to efficiently conduct different types of fermentations in the same bioreactor, avoiding inhibition problems of chemical or biological (competition) nature.

  7. Conformal nano-thin modified polyelectrolyte coatings for encapsulation of cells.

    Science.gov (United States)

    Granicka, L H; Antosiak-Iwańska, M; Godlewska, E; Strawski, M; Szklarczyk, M; Maranowski, B; Kowalewski, C; Wiśniewsk, J

    2011-10-01

    Encapsulation of cells in polymeric shells allows for separation of biological material from produced factors, which may find biotechnological and biomedical applications. Human T-lymphocyte cell line Jurkat as well as rat pancreatic islets were encapsulated using LbL technique within shells of polyelectrolyte modified by incorporation of biotin complexed with avidin to improve cell coating and to create the potential ability to elicit specific biochemical responses. The coating with nano-thin modified shells allowed for maintenance of the evaluated cells' integrity and viability during the 8-day culture. The different PE impact may be observed on different biological materials. The islets exhibited lower mitochondrial activity than the Jurkat cells. Nevertheless, coating of cells with polyelectrolyte modified membrane allowed for functioning of both model cell types: 10 μm leukemia cells or 150 μm islets during the culture. Applied membranes maintained the molecular structure during the culture period. The conclusion is that applied modified membrane conformation may be recommended for coating shells for biomedical purposes.

  8. [Gelatin/alginate hydrogel scaffolds prepared by 3D bioprinting promotes cell adhesion and proliferation of human dental pulp cells in vitro].

    Science.gov (United States)

    Yu, Hai-Yue; Ma, Dan-Dan; Wu, Bu-Ling

    2017-05-20

    To evaluate the cytotoxicity of gelatin/alginate hydrogel scaffolds prepared by 3D bioprinting in human dental pulp cells (HDPCs) and compare the cell adhesion and proliferation of the cells seeded in the biomaterial using two different methods. HDPCs isolated by tissue block culture and enzyme digestion were cultured and passaged. Gelatin/alginate hydrogel scaffolds were printed using a bioplotter, and the cytotoxicity of the aqueous extracts of the scaffold material was tested in the third passage of HDPCs using cell counting kit-8. Scanning electron microscopy and trypan blue were used to assess the adhesion and proliferation of the cells seeded in the scaffold material at a low or high concentration. The aqueous extract of the scaffolds at different concentrations showed no obvious cytotoxicity and promoted the proliferation of HDPCs. The scaffolds had a good biocompatibility and HDPCs seeded in the scaffold showed good cell growth. Cell seeding at a high concentration in the scaffold better promoted the adhesion of HDPCs and resulted in a greater cell number on the scaffold surface compared with low-concentration cell seeding after a 5-day culture (Padhesion to the scaffold material.

  9. Injectable, Biomolecule-Responsive Polypeptide Hydrogels for Cell Encapsulation and Facile Cell Recovery through Triggered Degradation.

    Science.gov (United States)

    Xu, Qinghua; He, Chaoliang; Zhang, Zhen; Ren, Kaixuan; Chen, Xuesi

    2016-11-16

    Injectable hydrogels have been widely investigated in biomedical applications, and increasing demand has been proposed to achieve dynamic regulation of physiological properties of hydrogels. Herein, a new type of injectable and biomolecule-responsive hydrogel based on poly(l-glutamic acid) (PLG) grafted with disulfide bond-modified phloretic acid (denoted as PLG-g-CPA) was developed. The hydrogels formed in situ via enzymatic cross-linking under physiological conditions in the presence of horseradish peroxidase and hydrogen peroxide. The physiochemical properties of the hydrogels, including gelation time and the rheological property, were measured. Particularly, the triggered degradation of the hydrogel in response to a reductive biomolecule, glutathione (GSH), was investigated in detail. The mechanical strength and inner porous structure of the hydrogel were influenced by the addition of GSH. The polypeptide hydrogel was used as a three-dimensional (3D) platform for cell encapsulation, which could release the cells through triggered disruption of the hydrogel in response to the addition of GSH. The cells released from the hydrogel were found to maintain high viability. Moreover, after subcutaneous injection into rats, the PLG-g-CPA hydrogels with disulfide-containing cross-links exhibited a markedly faster degradation behavior in vivo compared to that of the PLG hydrogels without disulfide cross-links, implying an interesting accelerated degradation process of the disulfide-containing polypeptide hydrogels in the physiological environment in vivo. Overall, the injectable and biomolecule-responsive polypeptide hydrogels may serve as a potential platform for 3D cell culture and easy cell collection.

  10. Control of Microbial Growth in Alginate/Polydopamine Core/Shell Microbeads.

    Science.gov (United States)

    Kim, Beom Jin; Park, Taegyun; Park, So-Young; Han, Sang Woo; Lee, Hee-Seung; Kim, Yang-Gyun; Choi, Insung S

    2015-10-01

    Microbial microencapsulation not only protects microorganisms from harmful environments by physically isolating them from the outside media but also has the potential to tailor the release profile of the encapsulated cells. However, the microbial release has not yet been controlled tightly, leading to undesired detrimental exposure of microorganisms to the outside. In this work, we suggest a simple method for controlling the cell release by suppressing the microbial growth in the microbeads. Alginate microbeads, encapsulating yeast cells, were coated with ultrathin but robust polydopamine shells, and the resulting core/shell structures effectively reduced the growth rate, while maintaining the cell viability. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  11. Bioengineering of injectable encapsulated aggregates of pluripotent stem cells for therapy of myocardial infarction

    Science.gov (United States)

    Zhao, Shuting; Xu, Zhaobin; Wang, Hai; Reese, Benjamin E.; Gushchina, Liubov V.; Jiang, Meng; Agarwal, Pranay; Xu, Jiangsheng; Zhang, Mingjun; Shen, Rulong; Liu, Zhenguo; Weisleder, Noah; He, Xiaoming

    2016-10-01

    It is difficult to achieve minimally invasive injectable cell delivery while maintaining high cell retention and animal survival for in vivo stem cell therapy of myocardial infarction. Here we show that pluripotent stem cell aggregates pre-differentiated into the early cardiac lineage and encapsulated in a biocompatible and biodegradable micromatrix, are suitable for injectable delivery. This method significantly improves the survival of the injected cells by more than six-fold compared with the conventional practice of injecting single cells, and effectively prevents teratoma formation. Moreover, this method significantly enhances cardiac function and survival of animals after myocardial infarction, as a result of a localized immunosuppression effect of the micromatrix and the in situ cardiac regeneration by the injected cells.

  12. Production of endothelial cell-enclosing alginate-based hydrogel fibers with a cell adhesive surface through simultaneous cross-linking by horseradish peroxidase-catalyzed reaction in a hydrodynamic spinning process.

    Science.gov (United States)

    Liu, Yang; Sakai, Shinji; Taya, Masahito

    2012-09-01

    We developed an alginate-based hydrogel fiber enabling to enclose endothelial cells, degradable on-demand by alginate lyase, and having a cell adhesive surface. The hydrogel fiber was obtained by extruding an aqueous solution of 4% (w/v) alginate derivative possessing phenolic hydroxyl moieties (Alg-Ph) and horseradish peroxidase (HRP) into a flow of aqueous solution containing 0.3 mM H(2)O(2) and gelatin derivative possessing Ph moieties (Gelatin-Ph). In the process, cross-linking of Alg-Ph resulting in a hydrogel fiber and immobilization of Gelatin-Ph on the surface of the hydrogel fiber were simultaneously accomplished by an HRP-catalyzed cross-linking reaction between Ph moieties. The diameter of the hydrogel fiber and the quantity of immobilized Gelatin-Ph on the fiber were controllable by changing the flow rates of the solutions and the concentration of HRP in the Alg-Ph-containing solution, respectively. The viability of the human endothelial cells enclosed in the hydrogel fibers obtained by 10 s of flowing in the H(2)O(2)-containing solution was 87.1%. In addition, the cells harvested from the hydrogel fibers through degradation using alginate lyase grew on tissue culture dishes in the same fashion as the cells seeded by a conventional subculture protocol. Human smooth muscle cells adhered, grew and achieved confluence on the surface of the hydrogel fibers. By degrading the hydrogel fibers using alginate lyase, a tubular cell construct was successfully obtained.

  13. Protein aggregation with poly(vinyl) alcohol surfactant reduces double emulsion-encapsulated mammalian cell-free expression

    Science.gov (United States)

    Ho, Kenneth K. Y.; Lee, Jin Woo; Durand, Grégory; Majumder, Sagardip

    2017-01-01

    Development of artificial cell models requires encapsulation of biomolecules within membrane-bound compartments. There have been limited studies of using mammalian cell-free expression (CFE) system as the ‘cytosol’ of artificial cells. We exploit glass capillary droplet microfluidics for the encapsulation of mammalian CFE within double emulsion templated vesicles. The complexity of the physicochemical properties of HeLa cell-free lysate poses a challenge compared with encapsulating simple buffer solutions. In particular, we discovered the formation of aggregates in double emulsion templated vesicles encapsulating mammalian HeLa CFE, but not with bacterial CFE. The aggregates did not arise from insolubility of the proteins made from CFE nor due to the interaction of mammalian CFE with the organic solvents in the middle phase of the double emulsions. We found that aggregation is dependent on the concentration of poly(vinyl) alcohol (PVA) surfactant, a critical double emulsion-stabilizing surfactant, and the lysate concentration in mammalian CFE. Despite vesicle instability and reduced protein expression, we demonstrate protein expression by encapsulating mammalian CFE system. Using mass spectrometry and Western blot, we identified and verified that actin is one of the proteins inside the mammalian CFE that aggregated with PVA surfactant. Our work establishes a baseline description of mammalian CFE system encapsulated in double emulsion templated vesicles as a platform for building artificial cells. PMID:28358875

  14. Protein aggregation with poly(vinyl) alcohol surfactant reduces double emulsion-encapsulated mammalian cell-free expression.

    Science.gov (United States)

    Ho, Kenneth K Y; Lee, Jin Woo; Durand, Grégory; Majumder, Sagardip; Liu, Allen P

    2017-01-01

    Development of artificial cell models requires encapsulation of biomolecules within membrane-bound compartments. There have been limited studies of using mammalian cell-free expression (CFE) system as the 'cytosol' of artificial cells. We exploit glass capillary droplet microfluidics for the encapsulation of mammalian CFE within double emulsion templated vesicles. The complexity of the physicochemical properties of HeLa cell-free lysate poses a challenge compared with encapsulating simple buffer solutions. In particular, we discovered the formation of aggregates in double emulsion templated vesicles encapsulating mammalian HeLa CFE, but not with bacterial CFE. The aggregates did not arise from insolubility of the proteins made from CFE nor due to the interaction of mammalian CFE with the organic solvents in the middle phase of the double emulsions. We found that aggregation is dependent on the concentration of poly(vinyl) alcohol (PVA) surfactant, a critical double emulsion-stabilizing surfactant, and the lysate concentration in mammalian CFE. Despite vesicle instability and reduced protein expression, we demonstrate protein expression by encapsulating mammalian CFE system. Using mass spectrometry and Western blot, we identified and verified that actin is one of the proteins inside the mammalian CFE that aggregated with PVA surfactant. Our work establishes a baseline description of mammalian CFE system encapsulated in double emulsion templated vesicles as a platform for building artificial cells.

  15. Asymmetric bioreduction of acetophenones by Baker's yeast and its cell-free extract encapsulated in sol-gel silica materials

    Science.gov (United States)

    Kato, Katsuya; Nakamura, Hitomi; Nakanishi, Kazuma

    2014-02-01

    Baker's yeast (BY) encapsulated in silica materials was synthesized using a yeast cell suspension and its cell-free extract during a sol-gel reaction of tetramethoxysilane with nitric acid as a catalyst. The synthesized samples were fully characterized using various methods, such as scanning electron microscopy, nitrogen adsorption-desorption, Fourier transform infrared spectroscopy, thermogravimetry, and differential thermal analysis. The BY cells were easily encapsulated inside silica-gel networks, and the ratio of the cells in the silica gel was approximately 75 wt%, which indicated that a large volume of BY was trapped with a small amount of silica. The enzyme activity (asymmetric reduction of prochiral ketones) of BY and its cell-free extract encapsulated in silica gel was investigated in detail. The activities and enantioselectivities of free and encapsulated BY were similar to those of acetophenone and its fluorine derivatives, which indicated that the conformation structure of BY enzymes inside silica-gel networks did not change. In addition, the encapsulated BY exhibited considerably better solvent (methanol) stability and recyclability compared to free BY solution. We expect that the development of BY encapsulated in sol-gel silica materials will significantly impact the industrial-scale advancement of high-efficiency and low-cost biocatalysts for the synthesis of valuable chiral alcohols.

  16. Laser-assisted fabrication of highly viscous alginate microsphere

    Science.gov (United States)

    Lin, Yafu; Huang, Yong

    2011-04-01

    Encapsulated microspheres have been widely used in various biomedical applications. However, fabrication of encapsulated microspheres from highly viscous materials has always been a manufacturing challenge. The objective of this study is to explore a novel metallic foil-assisted laser-induced forward transfer (LIFT), a laser-assisted fabrication technique, to make encapsulated microspheres using high sodium alginate concentration solutions. The proposed four-layer approach includes a quartz disk, a sacrificial and adhesive layer, a metallic foil, and a transferred suspension layer. It is found that the proposed four-layer modified LIFT approach provides a promising fabrication technology for making of bead-encapsulated microspheres from highly viscous solutions. During the process, the microsphere only can be formed if the direct-writing height is larger than the critical direct-writing height; otherwise, tail structured droplets are formed; and the encapsulated microsphere diameter linearly increases with the laser fluence and decreases with the sodium alginate concentration.

  17. Multi-compartment encapsulation of communicating droplets and droplet networks in hydrogel as a model for artificial cells

    Science.gov (United States)

    Bayoumi, Mariam; Bayley, Hagan; Maglia, Giovanni; Sapra, K. Tanuj

    2017-01-01

    Constructing a cell mimic is a major challenge posed by synthetic biologists. Efforts to this end have been primarily focused on lipid- and polymer-encapsulated containers, liposomes and polymersomes, respectively. Here, we introduce a multi-compartment, nested system comprising aqueous droplets stabilized in an oil/lipid mixture, all encapsulated in hydrogel. Functional capabilities (electrical and chemical communication) were imparted by protein nanopores spanning the lipid bilayer formed at the interface of the encapsulated aqueous droplets and the encasing hydrogel. Crucially, the compartmentalization enabled the formation of two adjoining lipid bilayers in a controlled manner, a requirement for the realization of a functional protocell or prototissue. PMID:28367984

  18. The incorporation of extracellular matrix proteins in protein polymer hydrogels to improve encapsulated beta-cell function.

    Science.gov (United States)

    Beenken-Rothkopf, Liese N; Karfeld-Sulzer, Lindsay S; Davis, Nicolynn E; Forster, Ryan; Barron, Annelise E; Fontaine, Magali J

    2013-01-01

    Biomaterial encapsulation of islets has been proposed to improve the long-term success of islet transplantation by recreating a suitable microenvironment and enhancing cell-matrix interactions that affect cellular function. Protein polymer hydrogels previously showed promise as a biocompatible scaffold by maintaining high cell viability. Here, enzymatically-crosslinked protein polymers were used to investigate the effects of varying scaffold properties and of introducing ECM proteins on the viability and function of encapsulated MIN6 β-cells. Chemical and mechanical properties of the hydrogel were modified by altering the protein concentrations while collagen IV, fibronectin, and laminin were incorporated to reestablish cell-matrix interactions lost during cell isolation. Rheology indicated all hydrogels formed quickly, resulting in robust, elastic hydrogels with Young's moduli similar to soft tissue. All hydrogels tested supported both high MIN6 β-cell viability and function and have the potential to serve as an encapsulation platform for islet cell delivery in vivo.

  19. Cell encapsulation in sub-mm sized gel modules using replica molding.

    Directory of Open Access Journals (Sweden)

    Alison P McGuigan

    Full Text Available For many types of cells, behavior in two-dimensional (2D culture differs from that in three-dimensional (3D culture. Among biologists, 2D culture on treated plastic surfaces is currently the most popular method for cell culture. In 3D, no analogous standard method--one that is similarly convenient, flexible, and reproducible--exists. This paper describes a soft-lithographic method to encapsulate cells in 3D gel objects (modules in a variety of simple shapes (cylinders, crosses, rectangular prisms with lateral dimensions between 40 and 1000 microm, cell densities of 10(5-10(8 cells/cm(3, and total volumes between 1x10(-7 and 8x10(-4 cm(3. By varying (i the initial density of cells at seeding, and (ii the dimensions of the modules, the number of cells per module ranged from 1 to 2500 cells. Modules were formed from a range of standard biopolymers, including collagen, Matrigel, and agarose, without the complex equipment often used in encapsulation. The small dimensions of the modules allowed rapid transport of nutrients by diffusion to cells at any location in the module, and therefore allowed generation of modules with cell densities near to those of dense tissues (10(8-10(9 cells/cm(3. This modular method is based on soft lithography and requires little special equipment; the method is therefore accessible, flexible, and well suited to (i understanding the behavior of cells in 3D environments at high densities of cells, as in dense tissues, and (ii developing applications in tissue engineering.

  20. The effect on lactic fermentation of concentrating inert material with immobilised cells in a calcium alginate biocatalyser

    Directory of Open Access Journals (Sweden)

    Juan Carlos Serrato

    2010-04-01

    Full Text Available Colombia is one of the world’s main sugarcane cultivating countries but it has not diversified its fermentation industry; a few fermentation industries produce alcohol and yeasts. Lactic acid and its derivatives then become alternatives providing added value to the sugar produced, thus benefiting the regions producing the sugar.This work evaluated the kinetics of lactic acid production using immobilised cells in calcium alginate at different concentrations of inert material. Lactobacillus delbrueckiI was the microorganism used and fermentation broth mainly consisted of sucrose and yeast exact. CSTR reactors were used without pH control. The results suggested that 2% to 3% inert material in the biocatalyst increased cellular retention and diffusiveness, leading to improved conversion and reaction rate.

  1. Fabrication, characterization and in vitro profile based interaction with eukaryotic and prokaryotic cells of alginate-chitosan-silica biocomposite.

    Science.gov (United States)

    Balaure, Paul Catalin; Andronescu, Ecaterina; Grumezescu, Alexandru Mihai; Ficai, Anton; Huang, Keng-Shiang; Yang, Chih-Hui; Chifiriuc, Carmen Mariana; Lin, Yung-Sheng

    2013-01-30

    This work is focused on the fabrication of a new drug delivery system based on polyanionic matrix (e.g. sodium alginate), polycationic matrix (e.g. chitosan) and silica network. The FT-IR, SEM, DTA-TG, eukaryotic cell cycle and viability, and in vitro assay of the influence of the biocomposite on the efficacy of antibiotic drugs were investigated. The obtained results demonstrated the biocompatibility and the ability of the fabricated biocomposite to maintain or improve the efficacy of the following antibiotics: piperacillin-tazobactam, cefepime, piperacillin, imipenem, gentamicin, ceftazidime against Pseudomonas aeruginosa ATCC 27853 and cefazolin, cefaclor, cefuroxime, ceftriaxone, cefoxitin, trimethoprim/sulfamethoxazole against Escherichia coli ATCC 25922 reference strains.

  2. Calcium deposition in photocrosslinked poly(Pro-Hyp-Gly) hydrogels encapsulated rat bone marrow stromal cells.

    Science.gov (United States)

    Nurlidar, Farah; Yamane, Keisuke; Kobayashi, Mime; Terada, Kayo; Ando, Tsuyoshi; Tanihara, Masao

    2017-07-17

    Reproducing the features of the extracellular matrix is important for fabricating three-dimensional (3D) scaffolds for tissue regeneration. A collagen-like polypeptide, poly(Pro-Hyp-Gly), is a promising material for 3D scaffolds because of its excellent physical properties, biocompatibility, and biodegradability. In this paper, we present a novel photocrosslinked poly(Pro-Hyp-Gly) hydrogel as a 3D scaffold for simultaneous rat bone marrow stromal cell (rBMSC) encapsulation. The hydrogels were fabricated using visible-light photocrosslinking at various concentrations of methacrylated poly(Pro-Hyp-Gly) (20-50 mg/mL) and irradiation times (3 or 5 min). The results show that the rBMSCs encapsulated in the hydrogels survived seven days of incubation. Calcium deposition on the encapsulated rBMSCs was assessed with SEM observation, Alizarin Red S and von Kossa staining. The most strongly stained area was observed in the hydrogel formed with 30 mg/mL of methacrylated poly(Pro-Hyp-Gly) with 5 min irradiation. These findings demonstrate that poly(Pro-Hyp-Gly) hydrogels support rBMSC viability and differentiation, as well as demonstrating the feasibility of using poly(Pro-Hyp-Gly) hydrogels as a cytocompatible, biodegradable 3D scaffold for tissue regeneration. This article is protected by copyright. All rights reserved.

  3. Study of Carbon Nano-Tubes Effects on the Chondrogenesis of Human Adipose Derived Stem Cells in Alginate Scaffold

    Directory of Open Access Journals (Sweden)

    Ali Valiani

    2014-01-01

    Full Text Available Background: Osteoarthritis is one of the most common diseases in middle-aged populations in the World and could become the fourth principal cause of disability by the year 2020. One of the critical properties for cartilage tissue engineering (TE is the ability of scaffolds to closely mimic the extracellular matrix and bond to the host tissue. Therefore, TE has been presented as a technique to introduce the best combination of cells and biomaterial scaffold and to stimulate growth factors to produce a cartilage tissue resembling natural articular cartilage. The aim of study is to improve differentiation of adipose derived stem cells (ADSCs into chondrocytes in order to provide a safe and modern treatment for patients suffering from cartilage damages. Methods: After functionalization, dispersions and sterilizing carbon nano-tubes (CNTs, a new type of nanocomposite gel was prepared from water-soluble CNTs and alginate. ADSCs seeded in 1.5% alginate scaffold and cultured in chondrogenic media with and without transforming growth factor-β1 (TGF-β1 for 7 and 14 days. The genes expression of sex determining region Y-box 9 (SOX9, types II and X collagens was assessed by real-time polymerase chain reaction and the amount of aggrecan (AGC and type I collagen was measured by ELISA. Results: Our findings showed that the expression of essential cartilage markers, SOX9, type II collagen and AGC, in differentiated ADSCs at the concentration of 1 μg/ml CNTs in the presence of TGF-β1 were significantly increased in comparison with the control group (P < 0.001. Meanwhile, type X collagen expression and also type I collagen production were significantly decreased (P < 0.001. Conclusions: The results showed that utilized three-dimensional scaffold had a brilliant effect in promoting gene expression of chondrogenesis.

  4. Thin tantalum-silicon-oxygen/tantalum-silicon-nitrogen films as high-efficiency humidity diffusion barriers for solar cell encapsulation

    Energy Technology Data Exchange (ETDEWEB)

    Heuer, H. [Institut fuer Halbleiter-und Mikrosystemtechnik (IHM) Technische Universitaet Dresden, Helmholtzstrasse 10, 01062 Dresden (Germany)]. E-mail: Henning.Heuer@izfp-d.fraunhofer.de; Wenzel, C. [Institut fuer Halbleiter-und Mikrosystemtechnik (IHM) Technische Universitaet Dresden, Helmholtzstrasse 10, 01062 Dresden (Germany); Herrmann, D. [Institut fuer Halbleiter-und Mikrosystemtechnik (IHM) Technische Universitaet Dresden, Helmholtzstrasse 10, 01062 Dresden (Germany); Zentrum fuer Sonnenenergie-und Wasserstoff-Forschung (ZSW) Industriestrasse 6, 70565 Stuttgart (Germany); Huebner, R. [Institut fuer Halbleiter-und Mikrosystemtechnik (IHM) Technische Universitaet Dresden, Helmholtzstrasse 10, 01062 Dresden (Germany); Leibniz Institut fuer Festkoerper-und Werkstoffforschung Dresden (IFW) Helmholtzstrasse 20, 01069, Dresden (Germany); Zhang, Z.L. [Institut fuer Halbleiter-und Mikrosystemtechnik (IHM) Technische Universitaet Dresden, Helmholtzstrasse 10, 01062 Dresden (Germany); Max-Planck-Gesellschaft fuer Metallforschung (MPI) Heisenbergstrasse 3, 70569 Stuttgart (Germany); Bartha, J.W. [Institut fuer Halbleiter-und Mikrosystemtechnik (IHM) Technische Universitaet Dresden, Helmholtzstrasse 10, 01062 Dresden (Germany)

    2006-12-05

    Flexible thin-film solar cells require flexible encapsulation to protect the copper-indium-2 selenide (CIS) absorber layer from humidity and aggressive environmental influences. Tantalum-silicon-based diffusion barriers are currently a favorite material to prevent future semiconductor devices from copper diffusion. In this work tantalum-silicon-nitrogen (Ta-Si-N) and tantalum-silicon-oxygen (Ta-Si-O) films were investigated and optimized for thin-film solar cell encapsulation of next-generation flexible solar modules. CIS solar modules were coated with tantalum-based barrier layers. The performance of the thin-film barrier encapsulation was determined by measuring the remaining module efficiency after a 1000 h accelerated aging test. A significantly enhanced stability against humidity diffusion in comparison to non-encapsulated modules was reached with a reactively sputtered thin-film system consisting of 250 nm Ta-Si-O and 15 nm Ta-Si-N.

  5. Effect of concentration and substrate flow rate on isomaltulose production from sucrose by Erwinia sp. cells immobilized in calcium-alginate using packed bed reactor.

    Science.gov (United States)

    Kawaguti, Haroldo Yukio; Harumi Sato, Hélia

    2010-09-01

    Isomaltulose was obtained from sucrose solution by immobilized cells of Erwinia sp. D12 using a batch and a continuous process. Parameters for sucrose conversion into isomaltulose were evaluated using both experimental design and response surface methodology. Erwinia sp. D12 cells were immobilized in different alginates, and the influence of substrate flow rate and concentration parameters to produce isomaltulose from sucrose were observed. Response surface methodology demonstrated that packed bed columns containing cells immobilized in low-viscosity sodium alginate (250 cP) presented a mean isomaltulose conversion rate of 47%. In a continuous process, both sucrose substrate concentration and substrate flow rate parameters had a significant effect (p < 0.05) and influenced the conversion of sucrose into isomaltulose. Higher conversion rates of sucrose into isomaltulose, from 53-75% were obtained using 75 g of immobilized cells at a substrate flow rate of 0.6 mL/min.

  6. Enhanced production of alkaline thermostable keratinolytic protease from calcium alginate immobilized cells of thermoalkalophilic Bacillus halodurans JB 99 exhibiting dehairing activity.

    Science.gov (United States)

    Shrinivas, Dengeti; Kumar, Raghwendra; Naik, G R

    2012-01-01

    The thermoalkalophilic Bacillus halodurans JB 99 cells known for production of novel thermostable alkaline keratinolytic protease were immobilized in calcium alginate matrix. Batch and repeated batch cultivation using calcium alginate immobilized cells were studied for alkaline protease production in submerged fermentation. Immobilized cells with 2.5% alginate and 350 beads/flask of initial cell loading showed enhanced production of alkaline protease by 23.2% (5,275 ± 39.4 U/ml) as compared to free cells (4,280 ± 35.4 U/ml) after 24 h. In the semicontinuous mode of cultivation, immobilized cells under optimized conditions produced an appreciable level of alkaline protease in up to nine cycles and reached a maximal value of 5,975 U/ml after the seventh cycle. The enzyme produced from immobilized cells efficiently degraded chicken feathers in the presence of a reducing agent which can help the poultry industry in the management of keratin-rich waste and obtaining value-added products.

  7. Investigation of test methods, material properties, and processes for solar cell encapsulants. Annual report

    Energy Technology Data Exchange (ETDEWEB)

    Willis, P. B.; Baum, B.

    1979-06-01

    The goal of this program is to identify, evaluate, and recommend encapsulant materials and processes for the production of cost-effective, long-life solar cell modules. During the past year, the technical activities emphasized the reformulation of a commercial grade of ethylene/vinyl acetate copolymer for use as a pottant in solar cell module manufacture. After experimenting with a variety of techniques, a vacuum-bag process was developed and found to be an excellent encapsulation method. Adhesive strengths and primers for the bonding of ethylene/vinyl acetate to superstrate and substrate materials was assessed with encouraging results. The weathering effects on ten other polymers exposed to twelve months of weathering in Arizona, Florida, and under EMMAQUA were evaluated by determination of tensile strengths, elongations, optical transmission, etc. As may be expected, the best overall retention of mechanical properties is found for the fluorocarbon polymers, especially FEP. Hard coatings containing ultraviolet absorbers were investigated for the purpose of providing a soil resistant surface and additional weathering stability to the soft EVA pottant. Corrosion studies using a standard salt spray test were used to determine the degree of protection offered to a variety of metals by encapsulation in EVA pottant. A survey of scrim materials was also conducted. These open hole weaves are intended for use as spacers between the cell and substrate to provide a mechanical barrier, improve insulation resistance and prevent migration of the pigmented pottant over the cell surface. A mechanical engineering analysis of composite structural materials for use as substrates was performed. Results are presented in detail. (WHK)

  8. Macrophage depletion improves survival of porcine neonatal pancreatic cell clusters contained in alginate macrocapsules transplanted into rats.

    NARCIS (Netherlands)

    Omer, A; Keegan, M; Czismadia, E; Vos, P De; Rooijen, van N.; Bonner-Weir, S; Weir, GC

    2003-01-01

    BACKGROUND: Macrophages can accumulate on the surface of empty and islet-containing alginate capsules, leading to loss of functional tissue. In this study, the effect of peritoneal macrophage depletion on the biocompatibility of alginate macrocapsules and function of macroencapsulated porcine neonat

  9. Macrophage depletion improves survival of porcine neonatal pancreatic cell clusters contained in alginate macrocapsules transplanted into rats

    NARCIS (Netherlands)

    Omer, A; Keegan, M; Czismadia, E; De Vos, P; Van Rooijen, N; Bonner-Weir, S; Weir, GC

    2003-01-01

    Background: Macrophages can accumulate on the surface of empty and islet-containing alginate capsules, leading to loss of functional tissue. In this study, the effect of peritoneal macrophage depletion on the biocompatibility of alginate macrocapsules and function of macroencapsulated porcine neonat

  10. Graphene oxide increases the viability of C2C12 myoblasts microencapsulated in alginate.

    Science.gov (United States)

    Ciriza, J; Saenz del Burgo, L; Virumbrales-Muñoz, M; Ochoa, I; Fernandez, L J; Orive, G; Hernandez, R M; Pedraz, J L

    2015-09-30

    Cell microencapsulation represents a great promise for long-term drug delivery, but still several challenges need to be overcome before its translation into the clinic, such as the long term cell survival inside the capsules. On this regard, graphene oxide has shown to promote proliferation of different cell types either in two or three dimensions. Therefore, we planned to combine graphene oxide with the cell microencapsulation technology. We first studied the effect of this material on the stability of the capsules and next we analyzed the biocompatibility of this chemical compound with erythropoietin secreting C2C12 myoblasts within the microcapsule matrix. We produced 160 μm-diameter alginate microcapsules with increasing concentrations of graphene oxide and did not find modifications on the physicochemical parameters of traditional alginate microcapsules. Moreover, we observed that the viability of encapsulated cells within alginate microcapsules containing specific graphene oxide concentrations was enhanced. These results provide a relevant step for the future clinical application of graphene oxide on cell microencapsulation.

  11. A cytoprotective and degradable metal-polyphenol nanoshell for single-cell encapsulation.

    Science.gov (United States)

    Park, Ji Hun; Kim, Kyunghwan; Lee, Juno; Choi, Ji Yu; Hong, Daewha; Yang, Sung Ho; Caruso, Frank; Lee, Younghoon; Choi, Insung S

    2014-11-10

    Single-cell encapsulation promises the cytoprotection of the encased cells against lethal stressors, reminiscent of the sporulation process in nature. However, the development of a cytocompatible method for chemically mimicking the germination process (i.e., shell degradation on-demand) has been elusive, despite the shell degradation being pivotal for the practical use of functional cells as well as for single cell-based biology. We report that an artificial shell, composed of tannic acid (TA) and Fe(III) , on individual Saccharomyces cerevisiae controllably degrades on-demand, while protecting the yeast from multiple external aggressors, including UV-C irradiation, lytic enzymes, and silver nanoparticles. Cell division is suppressed by the TA-Fe(III) shell, but restored fully upon shell degradation. The formation of a TA-Fe(III) shell would provide a versatile tool for achieving the chemical version of "sporulation and germination".

  12. Increasing light capture in silicon solar cells with encapsulants incorporating air prisms to reduce metallic contact losses.

    Science.gov (United States)

    Chen, Fu-Hao; Pathreeker, Shreyas; Kaur, Jaspreet; Hosein, Ian D

    2016-10-31

    Silicon solar cells are the most widely deployed modules owing to their low-cost manufacture, large market, and suitable efficiencies for residential and commercial use. Methods to increase their solar energy collection must be easily integrated into module fabrication. We perform a theoretical and experimental study on the light collection properties of an encapsulant that incorporates a periodic array of air prisms, which overlay the metallic front contacts of silicon solar cells. We show that the light collection efficiency induced by the encapsulant depends on both the shape of the prisms and angle of incidence of incoming light. We elucidate the changes in collection efficiency in terms of the ray paths and reflection mechanisms in the encapsulant. We fabricated the encapsulant from a commercial silicone and studied the change in the external quantum efficiency (EQE) on an encapsulated, standard silicon solar cell. We observe efficiency enhancements, as compared to a uniform encapsulant, over the visible to near infrared region for a range of incident angles. This work demonstrates exactly how a periodic air prism architecture increases light collection, and how it may be designed to maximize light collection over the widest range of incident angles.

  13. Encapsulation of plasmid DNA in calcium phosphate nanoparticles: stem cell uptake and gene transfer efficiency

    Directory of Open Access Journals (Sweden)

    Cao X

    2011-12-01

    Full Text Available Xia Cao*, Wenwen Deng*, Yuan Wei*, Weiyan Su, Yan Yang, Yawei Wei, Jiangnan Yu, Ximing XuDepartment of Pharmaceutics, School of Pharmacy, and Center for Nano Drug/Gene Delivery and Tissue Engineering, Jiangsu University, Jingkou District, Zhenjiang, People's Republic of China*These authors contributed equally to this workBackground: The purpose of this study was to develop calcium phosphate nanocomposite particles encapsulating plasmid DNA (CP-pDNA nanoparticles as a nonviral vector for gene delivery.Methods: CP-pDNA nanoparticles employing plasmid transforming growth factor beta 1 (TGF-β1 were prepared and characterized. The transfection efficiency and cell viability of the CP-pDNA nanoparticles were evaluated in mesenchymal stem cells, which were identified by immunofluorescence staining. Cytotoxicity of plasmid TGF-β1 and calcium phosphate to mesenchymal stem cells were evaluated by MTT assay.Results: The integrity of TGF-β1 encapsulated in the CP-pDNA nanoparticles was maintained. The well dispersed CP-pDNA nanoparticles exhibited an ultralow particle size (20–50 nm and significantly lower cytotoxicity than Lipofectamine™ 2000. Immunofluorescence staining revealed that the cultured cells in this study were probably mesenchymal stem cells. The cellular uptake and transfection efficiency of the CP-pDNA nanoparticles into the mesenchymal stem cells were higher than that of needle-like calcium phosphate nanoparticles and a standard calcium phosphate transfection kit. Furthermore, live cell imaging and confocal laser microscopy vividly showed the transportation process of the CP-pDNA nanoparticles in mesenchymal stem cells. The results of a cytotoxicity assay found that both plasmid TGF-β1 and calcium phosphate were not toxic to mesenchymal stem cells.Conclusion: CP-pDNA nanoparticles can be developed into an effective alternative as a nonviral gene delivery system that is highly efficient and has low cytotoxicity.Keywords: calcium

  14. Dextran vesicular carriers for dual encapsulation of hydrophilic and hydrophobic molecules and delivery into cells.

    Science.gov (United States)

    Pramod, P S; Takamura, Kathryn; Chaphekar, Sonali; Balasubramanian, Nagaraj; Jayakannan, M

    2012-11-12

    Dextran vesicular nanoscaffolds were developed based on polysaccharide and renewable resource alkyl tail for dual encapsulation of hydrophilic and hydrophobic molecules (or drugs) and delivery into cells. The roles of the hydrophobic segments on the molecular self-organization of dextran backbone into vesicles or nanoparticles were investigated in detail. Dextran vesicles were found to be a unique dual carrier in which water-soluble molecules (like Rhodamine-B, Rh-B) and polyaromatic anticancer drug (camptothecin, CPT) were selectively encapsulated in the hydrophilic core and hydrophobic layer, respectively. The dextran vesicles were capable of protecting the plasma-sensitive CPT lactone pharmacophore against the hydrolysis by 10× better than the CPT alone in PBS. The aliphatic ester linkage connecting the hydrophobic tail with dextran was found to be cleaved by esterase under physiological conditions for fast releasing of CPT or Rh-B. Cytotoxicity of the dextran vesicle and its drug conjugate were tested on mouse embryonic fibroblast cells (MEFs) using MTT assay. The dextran vesicular scaffold was found to be nontoxic to living cells. CPT loaded vesicles were found to be 2.5-fold more effective in killing fibroblasts compared to that of CPT alone in PBS. Confocal microscopic images confirmed that both Rh-B and CPT loaded vesicles to be taken up by fibroblasts compared to CPT alone, showing a distinctly perinuclear localization in cells. The custom designed dextran vesicular provides new research opportunities for dual loading and delivering of hydrophilic and hydrophobic drug molecules.

  15. Chondrogenesis of human bone marrow mesenchymal stromal cells in highly porous alginate-foams supplemented with chondroitin sulfate

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Zhao [Department of Orthopaedic, Trauma and Reconstructive Surgery, Charité-Universitätsmedizin-Berlin Campus Benjamin Franklin, Berlin (Germany); Nooeaid, Patcharakamon [Institute of Biomaterials, Department of Materials Science and Engineering, University of Erlangen-Nuremberg (Germany); Kohl, Benjamin [Department of Orthopaedic, Trauma and Reconstructive Surgery, Charité-Universitätsmedizin-Berlin Campus Benjamin Franklin, Berlin (Germany); Roether, Judith A.; Schubert, Dirk W. [Institute of Polymer Materials, Department of Materials Science and Engineering, University of Erlangen-Nuremberg (Germany); Meier, Carola [Department of Orthopaedic, Trauma and Reconstructive Surgery, Charité-Universitätsmedizin-Berlin Campus Benjamin Franklin, Berlin (Germany); Boccaccini, Aldo R. [Institute of Biomaterials, Department of Materials Science and Engineering, University of Erlangen-Nuremberg (Germany); Godkin, Owen; Ertel, Wolfgang; Arens, Stephan [Department of Orthopaedic, Trauma and Reconstructive Surgery, Charité-Universitätsmedizin-Berlin Campus Benjamin Franklin, Berlin (Germany); Schulze-Tanzil, Gundula, E-mail: gundula.schulze@pmu.ac.at [Department of Orthopaedic, Trauma and Reconstructive Surgery, Charité-Universitätsmedizin-Berlin Campus Benjamin Franklin, Berlin (Germany); Institute of Anatomy, Paracelsus Medical University, Nuremberg (Germany)

    2015-05-01

    To overcome the limited intrinsic cartilage repair, autologous chondrocyte or bone-marrow-derived mesenchymal stromal cell (BM-MSC) was implanted into cartilage defects. For this purpose suitable biocompatible scaffolds are needed to provide cell retention, chondrogenesis and initial mechanical stability. The present study should indicate whether a recently developed highly porous alginate (Alg) foam scaffold supplemented with chondroitin sulfate (CS) allows the attachment, survival and chondrogenesis of BM-MSCs and articular chondrocytes. The foams were prepared using a freeze-drying method; some of them were supplemented with CS and subsequently characterized for porosity, biodegradation and mechanical profile. BM-MSCs were cultured for 1–2 weeks on the scaffold either under chondrogenic or maintenance conditions. Cell vitality assays, histology, glycosaminoglycan (sGAG) assay, and type II and I collagen immunolabelings were performed to monitor cell growth and extracellular matrix (ECM) synthesis in the scaffolds. Scaffolds had a high porosity ~ 93–95% with a mean pore sizes of 237 ± 48 μm (Alg) and 197 ± 61 μm (Alg/CS). Incorporation of CS increased mechanical strength of the foams providing gradually CS release over 7 days. Most of the cells survived in the scaffolds. BM-MSCs and articular chondrocytes formed rounded clusters within the scaffold pores. The BM-MSCs, irrespective of whether cultured under non/chondrogenic conditions and chondrocytes produced an ECM containing sGAGs, and types II and I collagen. Total collagen and sGAG contents were higher in differentiated BM-MSC cultures supplemented with CS than in CS-free foams after 14 days. The cell cluster formation induced by the scaffolds might stimulate chondrogenesis via initial intense cell–cell contacts. - Highlights: • Alginate foam scaffolds revealed a high porosity and mean pore size of 197–237 μm. • Chondroitin sulfate was released over 14 days by the scaffolds. • Chondrocytes

  16. Electrostatic extrusion as a dispersion technique for encapsulation of cells and bioactive compounds

    Directory of Open Access Journals (Sweden)

    Kostić Ivana T.

    2012-01-01

    Full Text Available Significant development of cells and bioactive compound encapsulation technologies is taking place due to an exceptional possibility of their application in various scientific disciplines, including biomedicine, pharmacy, cosmetology, food and agricultural sciences, beverage production, industrial waste treatment. Despite the broad application of microencapsulation, the literature reviews on dispersion techniques for microcapsule/microbead production, their advantages, restrictions and drawbacks are scarce. The purpose of this paper is to assess the possibilities of electrostatic extrusion for encapsulation of biological material, including living cells in hydrogel microbeads. The paper presents an overview of the mechanisms of droplet formation and controlling experimental parameters for producing microbeads by means of electrostatic extrusion. Electrostatic droplet formation utilizes a special type of physical process taking advantage of electrostatic effects occurring in flowing conductive liquids after introduction of an electric field.When an electrostatic field is applied to the metal needle and an electric charge is induced in the liquid flowing out of the needle, the size of droplet detaching from the needle tip decreases as a funcion of applied electrostatic field. It has been shown that few parameters affect microbead size: applied voltage, electrode geometry, needle size, polarity arrangement and polymer concentration. The electrostatic droplet formation is one of the most precise methods, which enables one to produce spherical and uniform particles ranging from 100 μm up to 1000 μm. Most of the authors report that the encapsulated compounds (drugs, enzymes and living cells remain unaltered after electrostatic extrusion. This technique seems to be particularly promising in biotechnology, pharmaceutical and cosmetics industries, where a low-temperature process, preserving heat-sensitive material is a prerequisite. Future efforts in

  17. In vitro stimulation of murine peritoneal monocytes induced by alginates.

    Science.gov (United States)

    Pasquali, Paolo; Zalcman, Amy; Murtas, Susanna; Adone, Rosanna; Brambilla, Gianfranco; Marianelli, Cinzia; Cagiola, Monica; Ciuchini, Franco

    2005-08-01

    In this trial we assessed the effect of soluble alginates on murine cells. Mouse peritoneal monocytes were stimulated in vitro with a solution of alginate. The production of TNF-alpha and nitric oxide (NO), the expression of surface molecules CD80 and CD86, and the ability of monocytes to phagocyte bacteria were assessed, in order to evaluate the effect of alginate on cell functionality. We showed that mouse peritoneal monocytes stimulated with alginate produce NO and TNF-alpha. In addition, alginate is able also to increase their phagocytic activity and to a lesser extent also to increase the expression of CD80. Even with different degrees, it implies that alginates per se act directly on immune response, being able to effectively stimulate proinflammatory activity. These findings corroborate the idea that alginates can represent interesting adjuvants to use to increase the efficacy of antigenic stimulation.

  18. Microencapsulation of probiotics using sodium alginate

    Directory of Open Access Journals (Sweden)

    Mariana de Araújo Etchepare

    2015-07-01

    Full Text Available The consumption of probiotics is constantly growing due to the numerous benefits conferred on the health of consumers. In this context, Microencapsulation is a technology that favors the viability of probiotic cultures in food products, mainly by the properties of protection against adverse environmental conditions and controlled release. Currently there are different procedures for microencapsulation using polymers of various types of natural and synthetic origin. The use of sodium alginate polymers is one of the largest potential application in the encapsulation of probiotics because of their versatility, biocompatibility and toxicity exemption. The aim of this review is to present viable encapsulation techniques of probiotics with alginate, emphasizing the internal ionic gelation and external ionic gelation, with the possibility of applying, as well as promising for improving these techniques.

  19. Alginate-chitosan coacervation in production of artificial seeds.

    Science.gov (United States)

    Tay, L F; Khoh, L K; Loh, C S; Khor, E

    1993-08-05

    Survival of secondary embryoids of winter oilseed rape (Brassica napus ssp. oleifera cv. Primor) has been used as an assay for the development of artificial seeds involving complex coacervation of alginate (polyanion) with chitosan (polycation). Germination frequency of 100% was achieved for encapsulated embryoids when alginate formed the inner matrix and chitosan the outer layer. When the matrix makeup was reversed, there was no germination of embryoids. The artificial seeds produced were hardened in dilute alkaline solutions of NaOH and Ca(OH)(2). An optimum setting time could be selected based on a quantitative measurement of resistance of hardened capsules to compression and the germination frequency of the encapsulated embryoids.

  20. Fabrication of supramolecular hydrogels for drug delivery and stem cell encapsulation.

    Science.gov (United States)

    Wu, De-Qun; Wang, Tao; Lu, Bo; Xu, Xiao-Ding; Cheng, Si-Xue; Jiang, Xue-Jun; Zhang, Xian-Zheng; Zhuo, Ren-Xi

    2008-09-16

    Supramolecular hydrogels self-assembled by alpha-cyclodextrin and methoxypolyethylene glycol-poly(caprolactone)-(dodecanedioic acid)-poly(caprolactone)-methoxypolyethylene glycol (MPEG-PCL-MPEG) triblock polymers were prepared and characterized in vitro and in vivo. The sustained release of dextran-fluorescein isothiocyanate (FITC) from the hydrogels lasted for more than 1 month, which indicated that the hydrogels were promising for controlled drug delivery. ECV304 cells and marrow mesenchymal stem cells (MSC) were encapsulated and cultured in the hydrogels, during which the morphologies of the cells could be kept. The in vitro cell viability studies and the in vivo histological studies demonstrated that the hydrogels were non-cytotoxic and biocompatible, which indicated that the hydrogels prepared were promising candidates as injectable scaffolds for tissue engineering applications.

  1. Microencapsulation of a synbiotic into PLGA/alginate multiparticulate gels.

    Science.gov (United States)

    Cook, Michael T; Tzortzis, George; Charalampopoulos, Dimitris; Khutoryanskiy, Vitaliy V

    2014-05-15

    Probiotic bacteria have gained popularity as a defence against disorders of the bowel. However, the acid sensitivity of these cells results in a loss of viability during gastric passage and, consequently, a loss of efficacy. Probiotic treatment can be supplemented using 'prebiotics', which are carbohydrates fermented specifically by probiotic cells in the body. This combination of probiotic and prebiotic is termed a 'synbiotic'. Within this article a multiparticulate dosage form has been developed, consisting of poly(d,l-lactic-co-glycolic acid) (PLGA) microcapsules containing prebiotic Bimuno™ incorporated into an alginate-chitosan matrix containing probiotic Bifidobacterium breve. The aim of this multiparticulate was that, in vivo, the probiotic would be protected against gastric acid and the release of the prebiotic would occur in the distal colon. After microscopic investigation, this synbiotic multiparticulate was shown to control the release of the prebiotic during in vitro gastrointestinal transit, with the release of galacto-oligosaccharides (GOS) initially occurred over 6h, but with a triphasic release pattern giving further release over 288 h. Encapsulation of B. breve in multiparticulates resulted in a survival of 8.0 ± 0.3 logCFU/mL cells in acid, an improvement over alginate-chitosan microencapsulation of 1.4 logCFU/mL. This was attributed to increased hydrophobicity by the incorporation of PLGA particles. Copyright © 2014. Published by Elsevier B.V.

  2. Evaluation of Encapsulant Adhesion to Surface Metallization of Photovoltaic Cells: Preprint

    Energy Technology Data Exchange (ETDEWEB)

    Tracey, Jared; Dauskardt, Reinhold; Bosco, Nick

    2017-06-14

    Delamination of encapsulant materials from PV cell surfaces often appears to originate at regions with metallization. Using a fracture mechanics based metrology, the adhesion of EVA encapsulant to screen printed silver metallization was evaluated. At room temperature, the fracture energy, Gc [J/m2], of the EVA/silver interface (952 J/m2) was ~70% lower than that of the EVA/AR coating (>2900 J/m2) and ~60% lower than that of the EVA to the surface of cell (2265 J/m2). After only 300 hours of damp heat aging, the adhesion energy of the silver interface dropped to and plateaued at ~50-60 J/m2, while that of the EVA/AR coating and EVA/cell remained mostly unchanged. Elemental surface analysis showed that the EVA separates from the silver in a purely adhesive manner, indicating that bonds at the interface were likely displaced in the presence of humidity and elevated temperature, and may explain the propensity for delamination to occur at metallized surfaces in the field.

  3. New insights into Pseudomonas fluorescens alginate biosynthesis relevant for the establishment of an efficient production process for microbial alginates.

    Science.gov (United States)

    Maleki, Susan; Mærk, Mali; Hrudikova, Radka; Valla, Svein; Ertesvåg, Helga

    2017-07-25

    Alginate denotes a family of linear polysaccharides with a wide range of industrial and pharmaceutical applications. Presently, all commercially available alginates are manufactured from brown algae. However, bacterial alginates have advantages with regard to compositional homogeneity and reproducibility. In order to be able to design bacterial strains that are better suited for industrial alginate production, defining limiting factors for alginate biosynthesis is of vital importance. Our group has been studying alginate biosynthesis in Pseudomonas fluorescens using several complementary approaches. Alginate is synthesised and transported out of the cell by a multiprotein complex spanning from the inner to the outer membrane. We have developed an immunogold labelling procedure in which the porin AlgE, as a part of this alginate factory, could be detected by transmission electron microscopy. No time-dependent correlation between the number of such factories on the cell surface and alginate production level was found in alginate-producing strains. Alginate biosynthesis competes with the central carbon metabolism for the key metabolite fructose 6-phosphate. In P. fluorescens, glucose, fructose and glycerol, are metabolised via the Entner-Doudoroff and pentose phosphate pathways. Mutational analysis revealed that disruption of the glucose 6-phosphate dehydrogenase gene zwf-1 resulted in increased alginate production when glycerol was used as carbon source. Furthermore, alginate-producing P. fluorescens strains cultivated on glucose experience acid stress due to the simultaneous production of alginate and gluconate. The combined results from our studies strongly indicate that the availability of fructose 6-phosphate and energy requires more attention in further research aimed at the development of an optimised alginate production process.

  4. Simultaneous Alcoholic and Malolactic Fermentations by Saccharomyces cerevisiae and Oenococcus oeni Cells Co-immobilized in Alginate Beads

    Science.gov (United States)

    Bleve, Gianluca; Tufariello, Maria; Vetrano, Cosimo; Mita, Giovanni; Grieco, Francesco

    2016-01-01

    Malolactic fermentation (MLF) usually takes place after the end of alcoholic fermentation (AF). However, the inoculation of lactic acid bacteria together with yeast starter cultures is a promising system to enhance the quality and safety of wine. In recent years, the use of immobilized cell systems has been investigated, with interesting results, for the production of different fermented foods and beverages. In this study we have carried out the simultaneous immobilization of Saccharomyces cerevisiae and Oenococcus oeni in alginate beads and used them in microvinifications tests to produce Negroamaro wine. The process was monitored by chemical and sensorial analyses and dominance of starters and cell leaking from beads were also checked. Co-immobilization of S. cerevisiae and O. oeni allowed to perform an efficient fermentation process, producing low volatile acidity levels and ethanol and glycerol concentrations comparable with those obtained by cell sequential inoculum and co-inoculum of yeast and bacteria cells in free form. More importantly, co-immobilization strategy produced a significant decrease of the time requested to complete AF and MLF. The immobilized cells could be efficiently reused for the wine fermentation at least three times without any apparent loss of cell metabolic activities. This integrated biocatalytic system is able to perform simultaneously AF and MLF, producing wines similar in organoleptic traits in comparison with wines fermented following traditional sequential AF and MLF with free cell starters. The immobilized-cell system, that we here describe for the first time in our knowledge, offers many advantages over conventional free cell fermentations, including: (i) elimination of non-productive cell growth phases; (ii) feasibility of continuous processing; (iii) re-use of the biocatalyst. PMID:27379072

  5. Glucose concentration and medium volume influence cell viability and glycosaminoglycan synthesis in chondrocyte-seeded alginate constructs.

    Science.gov (United States)

    Heywood, Hannah K; Bader, Dan L; Lee, David A

    2006-12-01

    Increasing the thickness of tissue-engineered cartilage is associated with loss of chondrocyte viability and biosynthetic activity at the tissue center. Exceptionally high volumes of culture medium, however, can maintain cellularity and glycosaminoglycan synthesis throughout 4-mm-thick constructs. We hypothesized that glucose supplementation could replicate the augmentation of tissue formation achieved by medium volume. Chondrocyte-alginate constructs (40x10(6) cells/mL) were cultured for 14 days in 0.4-6.4 mL/10(-6) cells of either low- (5.1 mM) or high- (20.4 mM) glucose medium. Glucose was critical to chondrocyte viability, and glucose uptake increased significantly (P cells of low-glucose medium had a mass of 172 +/- 6.1 mg and glycosaminoglycan (GAG) content of 0.32 +/- 0.03 mg (mean +/- standard deviation). A 4-fold increase in medium volume increased the final construct mass by 44% and GAG content by 207%. An equivalent increase in glucose supply in the absence of volume change increased these parameters by just 10% and 73%, respectively. A similar trend was observed from 0.8 to 3.2 mL/10(-6) cells, when maximal values of construct GAG content and mass were obtained. Therefore, medium volume remains an important consideration for the optimal culture of tissue-engineered cartilage.

  6. Alginate/polyoxyethylene and alginate/gelatin hydrogels: preparation, characterization, and application in tissue engineering.

    Science.gov (United States)

    Aroguz, Ayse Z; Baysal, Kemal; Adiguzel, Zelal; Baysal, Bahattin M

    2014-05-01

    Hydrogels are attractive biomaterials for three-dimensional cell culture and tissue engineering applications. The preparation of hydrogels using alginate and gelatin provides cross-linked hydrophilic polymers that can swell but do not dissolve in water. In this work, we first reinforced pure alginate by using polyoxyethylene as a supporting material. In an alginate/PEO sample that contains 20 % polyoxyethylene, we obtained a stable hydrogel for cell culture experiments. We also prepared a stable alginate/gelatin hydrogel by cross-linking a periodate-oxidized alginate with another functional component such as gelatin. The hydrogels were found to have a high fluid uptake. In this work, preparation, characterization, swelling, and surface properties of these scaffold materials were described. Lyophilized scaffolds obtained from hydrogels were used for cell viability experiments, and the results were presented in detail.

  7. Characterization of novel photocrosslinked carboxymethylcellulose hydrogels for encapsulation of nucleus pulposus cells.

    Science.gov (United States)

    Reza, Anna T; Nicoll, Steven B

    2010-01-01

    Back pain is a significant clinical concern often associated with degeneration of the intervertebral disc (IVD). Tissue engineering strategies may provide a viable IVD replacement therapy; however, an ideal biomaterial scaffold has yet to be identified. One candidate material is carboxymethylcellulose (CMC), a water-soluble derivative of cellulose. In this study, 90 and 250 kDa CMC polymers were modified with functional methacrylate groups and photocrosslinked to produce hydrogels at different macromer concentrations. At 7 days, bovine nucleus pulposus (NP) cells encapsulated in these hydrogels were viable, with values for the elastic modulus ranging from 1.07 + or - 0.06 to 4.29 + or - 1.25 kPa. Three specific formulations were chosen for further study based on cell viability and mechanical integrity assessments: 4% 90 kDa, 2% 250 kDa and 3% 250 kDa CMC. The equilibrium weight swelling ratio of these formulations remained steady throughout the 2 week study (46.45 + or - 3.14, 48.55 + or - 2.91 and 42.41 + or - 3.06, respectively). The equilibrium Young's modulus of all cell-laden and cell-free control samples decreased over time, with the exception of cell-laden 3% 250 kDa CMC constructs, indicating an interplay between limited hydrolysis of interchain crosslinks and the elaboration of a functional matrix. Histological analyses of 3% 250 kDa CMC hydrogels confirmed the presence of rounded cells in lacunae and the pericellular deposition of chondroitin sulfate proteoglycan, a phenotypic NP marker. Taken together, these studies support the use of photocrosslinked CMC hydrogels as tunable biomaterials for NP cell encapsulation.

  8. Mass transfer characterization of gamma-aminobutyric acid production by Enterococcus faecium CFR 3003: encapsulation improves its survival under simulated gastro-intestinal conditions.

    Science.gov (United States)

    Divyashri, Gangaraju; Prapulla, Siddalingaiya Gurudatt

    2015-03-01

    Gamma-aminobutyric acid (GABA) production by free and Ca-alginate encapsulated cells of Enterococcus faecium CFR 3003 was investigated. Mass transfer rates characterizing the GABA production process using encapsulated cells were investigated. Experiments were performed to investigate external film and internal pore diffusion mass transfer rates. The Damkohler and Thiele analysis provides a good description of external film and internal pore diffusion resistances, respectively. The experiments revealed that the external film effects could be neglected but the process is affected to the greater extent by internal mass transfer effects and was found to be the principal rate-controlling step. Protective effect of encapsulation on cell survivability was tested under digestive environment, when challenged to salivary α-amylase, simulated gastric fluid and intestinal fluid. Viability of encapsulated cells was significantly higher under simulated gastro-intestinal conditions and could produce higher GABA than those observed with free cells. The results indicate that the Ca-alginate encapsulated probiotics could effectively be delivered to the colonic site for effective inhibitory action.

  9. Activation of cell signaling via optical manipulation of gold-coated liposomes encapsulating signaling molecules

    Science.gov (United States)

    Orsinger, Gabriel V.; Leung, Sarah J.; Romanowski, Marek

    2013-02-01

    Many diseases involve changes in cell signaling cascades, as seen commonly in drug resistant cancers. To better understand these intricate signaling events in diseased cells and tissues, experimental methods of probing cellular communication at a single to multi-cell level are required. We recently introduced a general platform for activation of selected signaling pathways by optically controlled delivery and release of water soluble factors using gold-coated liposomes. In the example presented here, we encapsulated inositol trisphosphate (IP3), a ubiquitous intracellular secondary messenger involved in GPCR and Akt signaling cascades, within 100 nm gold-coated liposomes. The high polarizability of the liposome's unique gold pseudo-shell allows stable optical trapping for subcellular manipulation in the presence of cells. We take this optical manipulation further by optically injecting IP3-containing liposomes into the cytosol of a single cell to initiate localized cell signaling. Upon optical injection of liposomal IP3 into a single ovarian carcinoma cell, we observed localized activation as reported by changes in Indo-1 fluorescence intensity. With established gap junctions between the injected cell and neighboring cells, we monitored propagation of this signaling to and through nearby cells.

  10. Technological process for cell disruption, extraction and encapsulation of astaxanthin from Haematococcus pluvialis.

    Science.gov (United States)

    Machado, Francisco R S; Trevisol, Thalles C; Boschetto, Daiane L; Burkert, Janaína F M; Ferreira, Sandra R S; Oliveira, J Vladimir; Burkert, Carlos André V

    2016-01-20

    In this work, the effectiveness of different enzymatic techniques for cell wall disruption of Haematococcus pluvialis for the extraction of carotenoids and subsequent encapsulation of extracts in the co-polymer poly(hydroxybutyrate-co-hydroxyvalerate) (PHBV) using the Solution Enhanced Dispersion by Supercritical fluids (SEDS) technique was investigated. Glucanex(®) performed best compared with Lyticase(®) and Driselase(®). The conditions for enzymatic lysis using this enzyme preparation were established as a pH of 4.5, a temperature of 55 °C, an initial activity of β-1,3-glucanase of 0.6 U mL(-1) and a reaction time of 30 min. Enzymatic lysis assisted by ultrasound without biomass freezing was shown to be a promising and simple one-step technique for cell wall disruption, reaching 83.90% extractability. In the co-precipitation experiments, the highest encapsulation efficiency (51.21%) was obtained when using a higher biomass to dichloromethane ratio (10 mg mL(-1)) at the carotenoid extraction step and a lower pressure of precipitation (80 bar). In these conditions, spherical particles in the micrometer range (0.228 μm) were obtained. Copyright © 2015 Elsevier B.V. All rights reserved.

  11. Chemical treatment and chitosan coating of yeast cells to improve the encapsulation and controlled release of bovine serum albumin.

    Science.gov (United States)

    Shi, Guorong; Liu, Yating; He, Zijun; Zhou, Jihen

    2016-08-10

    We investigate the encapsulation of bovine serum albumin (BSA) in chemical-treated and chitosan-coated yeast cells, Saccharomyces cerevisiae (S. cerevisiae), for the controlled release of BSA. The chemical treatment can sufficiently enlarge the small-sized cell-wall cavities and/or break the integrity for the entrance of BSA to the interior of yeast cells, and the additional chitosan coating can well prevent the rapid release of encapsulated BSA from the yeast-derived microcapsules. The sodium hydroxide pretreated S. cerevisiae gives a maximum encapsulation yield of (10.1 ± 0.2)% for BSA. An additional coating of S. cerevisiae with chitosan can reduce the initial burst release of BSA and extend the release period from 24 h in the chitosan-free case to 48 h in phosphate buffer at pH 7.4. The prepared microcapsules can well keep the shapes and sizes of yeast cells and thus show uniform sizes of 3.85 ± 0.81 μm. The encapsulated BSA well retains its pristine ultraviolet spectroscopic and chromatographic behaviors. The present microencapsulation protocol has the advantages of convenient and mild operation, high encapsulation efficiency, and organic solvent-free nature, which is of reference value for establishing high-performance controllable biomacromolecule-delivery systems.

  12. Development of a morphogenetically active scaffold for three-dimensional growth of bone cells: biosilica-alginate hydrogel for SaOS-2 cell cultivation.

    Science.gov (United States)

    Müller, Werner E G; Schröder, Heinz C; Feng, Qingling; Schlossmacher, Ute; Link, Thorben; Wang, Xiaohong

    2015-11-01

    Polymeric silica is formed from ortho-silicate during a sol-gel formation process, while biosilica is the product of an enzymatically driven bio-polycondensation reaction. Both polymers have recently been described as a template that induces an increased expression of the genes encoding bone morphogenetic protein 2 (BMP-2) and osteoprotegerin in osteoblast-related SaOS-2 cells; simultaneously or subsequently the cells respond with enhanced hydroxyapatite formation. In order to assess whether the biocompatible polymeric silica/biosilica can serve as a morphogenetically active matrix suitable for three-dimensional (3D) cell growth, or even for 3D cell bioprinting, SaOS-2 cells were embedded into a Na-alginate-based hydrogel. Four different gelatinous hydrogel matrices were used for suspending SaOS-2 cells: (a) the hydrogel alone; (b) the hydrogel with 400 μM ortho-silicate; (c) the hydrogel supplemented with 400 μM ortho-silicate and recombinant silicatein to allow biosilica synthesis to occur; and (d) the hydrogel with ortho-silicate and BSA. The SaOS-2 cells showed an increased growth if silica/biosilica components were present in the hydrogel. Likewise intensified was the formation of hydroxyapatite nodules in the silica-containing hydrogels. After an incubation period of 2 weeks, cells present in silica-containing hydrogels showed a significantly higher expression of the genes encoding the cytokine BMP-2, the major fibrillar structural protein collagen 1 and likewise of carbonic anhydrase. It is concluded that silica, and to a larger extent biosilica, retains its morphogenetic/osteogenic potential after addition to Na-alginate-based hydrogels. This property might qualify silica hydrogels to be also used as a matrix for 3D cell printing.

  13. Increased intestinal delivery of viable Saccharomyces boulardii by encapsulation in microspheres.

    Science.gov (United States)

    Graff, Sandrine; Hussain, Sajjad; Chaumeil, Jean-Claude; Charrueau, Christine

    2008-06-01

    Although probiotics are of a major potential therapeutic interest, their efficacy is usually limited by poor bioavailability of viable microorganisms on site. The aim of this study was to protect the probiotic Saccharomyces boulardii from degradation in order to ensure a greater number of viable yeast in the colon. Alginate microspheres coated with or not with chitosan were used to encapsulate the yeast by an extrusion method. The efficiency of encapsulation was assessed both in vitro and in vivo. In vitro, less than 1% of the non-encapsulated probiotic survived after 120 min at pH 1.1, whereas the majority of encapsulated yeast cells remained entrapped within both types of microspheres. Further exposure to a pH 6.8 allowed the release of about 35% of viable yeasts. In vivo, the percentage of viable yeast excreted over 96 h after a single oral dose of 2 x 10(8) cfu/100 g in rats was 2.5% for nonencapsulated yeast and reached 13.3 and 9.0% of the dose administered for the uncoated and chitosan-coated microspheres, respectively. Given the dose-dependent efficacy of S. boulardii and the efficiency of microencapsulation in protecting the yeast from degradation, alginate microspheres could be of great interest in therapeutic applications of the yeast.

  14. Pluronic-based micelle encapsulation potentiates myricetin-induced cytotoxicity in human glioblastoma cells.

    Science.gov (United States)

    Tang, Xiang-Jun; Huang, Kuan-Ming; Gui, Hui; Wang, Jun-Jie; Lu, Jun-Ti; Dai, Long-Jun; Zhang, Li; Wang, Gang

    As one of the natural herbal flavonoids, myricetin has attracted much research interest, mainly owing to its remarkable anticancer properties and negligible side effects. It holds great potential to be developed as an ideal anticancer drug through improving its bioavailability. This study was performed to investigate the effects of Pluronic-based micelle encapsulation on myricetin-induced cytotoxicity and the mechanisms underlying its anticancer properties in human glioblastoma cells. Cell viability was assessed using a methylthiazol tetrazolium assay and a real-time cell analyzer. Immunoblotting and quantitative reverse transcriptase polymerase chain reaction techniques were used for determining the expression levels of related molecules in protein and mRNA. The results indicated that myricetin-induced cytotoxicity was highly potentiated by the encapsulation of myricetin. Mitochondrial apoptotic pathway was demonstrated to be involved in myricetin-induced glioblastoma cell death. The epidermal growth factor receptor (EGFR)/PI3K/Akt pathway located in the plasma membrane and cytosol and the RAS-ERK pathway located in mitochondria served as upstream and downstream targets, respectively, in myricetin-induced apoptosis. MiR-21 inhibitors interrupted the expression of EGFR, p-Akt, and K-Ras in the same fashion as myricetin-loaded mixed micelles (MYR-MCs) and miR-21 expression were dose-dependently inhibited by MYR-MCs, indicating the interaction of miR-21 with MYR-MCs. This study provided evidence supportive of further development of MYR-MC formulation for preferentially targeting mitochondria of glioblastoma cells.

  15. Enhanced photodynamic efficacy towards melanoma cells by encapsulation of Pc4 in silica nanoparticles.

    Science.gov (United States)

    Zhao, Baozhong; Yin, Jun-Jie; Bilski, Piotr J; Chignell, Colin F; Roberts, Joan E; He, Yu-Ying

    2009-12-01

    Nanoparticles have been explored recently as an efficient means of delivering photosensitizers for cancer diagnosis and photodynamic therapy (PDT). Silicon phthalocyanine 4 (Pc4) is currently being clinically tested as a photosensitizer for PDT. Unfortunately, Pc4 aggregates in aqueous solutions, which dramatically reduces its PDT efficacy and therefore limits its clinical application. We have encapsulated Pc4 using silica nanoparticles (Pc4SNP), which not only improved the aqueous solubility, stability, and delivery of the photodynamic drug but also increased its photodynamic efficacy compared to free Pc4 molecules. Pc4SNP generated photo-induced singlet oxygen more efficiently than free Pc4 as measured by chemical probe and EPR trapping techniques. Transmission electron microscopy and dynamic light scattering measurements showed that the size of the particles is in the range of 25-30 nm. Cell viability measurements demonstrated that Pc4SNP was more phototoxic to A375 or B16-F10 melanoma cells than free Pc4. Pc4SNP photodamaged melanoma cells primarily through apoptosis. Irradiation of A375 cells in the presence of Pc4SNP resulted in a significant increase in intracellular protein-derived peroxides, suggesting a Type II (singlet oxygen) mechanism for phototoxicity. More Pc4SNP than free Pc4 was localized in the mitochondria and lysosomes. Our results show that these stable, monodispersed silica nanoparticles may be an effective new formulation for Pc4 in its preclinical and clinical studies. We expect that modifying the surface of silicon nanoparticles encapsulating the photosensitizers with antibodies specific to melanoma cells will lead to even better early diagnosis and targeted treatment of melanoma in the future.

  16. Immobilization of Electroporated Cells for Fabrication of Cellular Biosensors: Physiological Effects of the Shape of Calcium Alginate Matrices and Foetal Calf Serum

    Directory of Open Access Journals (Sweden)

    Nikos Katsanakis

    2009-01-01

    Full Text Available In order to investigate the physiological effect of transfected cell immobilization in calcium alginate gels, we immobilized electroporated Vero cells in gels shaped either as spherical beads or as thin membrane layers. In addition, we investigated whether serum addition had a positive effect on cell proliferation and viability in either gel configuration. The gels were stored for four weeks in a medium supplemented or not with 20% (v/v foetal calf serum. Throughout a culture period of four weeks, cell proliferation and cell viability were assayed by optical microscopy after provision of Trypan Blue. Non-elaborate culture conditions (room temperature, non-CO2 enriched culture atmosphere were applied throughout the experimental period in order to evaluate cell viability under less than optimal storage conditions. Immobilization of electroporated cells was associated with an initially reduced cell viability, which was gradually increased. Immobilization was associated with maintenance of cell growth for the duration of the experimental period, whereas electroporated cells essentially died after a week in suspension culture. Considerable proliferation of immobilized cells was observed in spherical alginate beads. In both gel configurations, addition of serum was associated with increased cell proliferation. The results of the present study could contribute to an improvement of the storability of biosensors based on electroporated, genetically or membrane-engineered cells.

  17. Antiproliferative effect of Antrodia camphorata polysaccharides encapsulated in chitosan-silica nanoparticles strongly depends on the metabolic activity type of the cell line

    Science.gov (United States)

    Kong, Zwe-Ling; Chang, Jenq-Sheng; Chang, Ke Liang B.

    2013-09-01

    Chitosan molecules interact with silica and encapsulate the Antrodia camphorata extract (ACE) polysaccharides to form composite nanoparticles. The nanoparticle suspensions of ACE polysaccharides encapsulated in silica-chitosan and silica nanoparticles approach an average particle size of 210 and 294 nm in solution, respectively. The encapsulation efficiencies of ACE polysaccharides are 66 and 63.5 %, respectively. Scanning electron micrographs confirm the formation of near-spherical nanoparticles. ACE polysaccharides solution had better antioxidative capability than ACE polysaccharides encapsulated in silica or silica-chitosan nanoparticles suspensions. The antioxidant capacity of nanoparticles increases with increasing dissolution time. The antitumor effects of ACE polysaccharides, ACE polysaccharides encapsulated in silica, or silica-chitosan nanoparticles increased with increasing concentration of nanoparticles. This is the first report demonstrating the potential of ACE polysaccharides encapsulated in chitosan-silica nanoparticles for cancer chemoprevention. Furthermore, this study suggests that antiproliferative effect of nanoparticle-encapsulated bioactive could significantly depend on the metabolic activity type of the cell line.

  18. Antiproliferative effect of Antrodia camphorata polysaccharides encapsulated in chitosan-silica nanoparticles strongly depends on the metabolic activity type of the cell line

    Energy Technology Data Exchange (ETDEWEB)

    Kong, Zwe-Ling, E-mail: kongzl@mail.ntou.edu.tw; Chang, Jenq-Sheng; Chang, Ke Liang B. [National Taiwan Ocean University, Department of Food Science (China)

    2013-09-15

    Chitosan molecules interact with silica and encapsulate the Antrodia camphorata extract (ACE) polysaccharides to form composite nanoparticles. The nanoparticle suspensions of ACE polysaccharides encapsulated in silica-chitosan and silica nanoparticles approach an average particle size of 210 and 294 nm in solution, respectively. The encapsulation efficiencies of ACE polysaccharides are 66 and 63.5 %, respectively. Scanning electron micrographs confirm the formation of near-spherical nanoparticles. ACE polysaccharides solution had better antioxidative capability than ACE polysaccharides encapsulated in silica or silica-chitosan nanoparticles suspensions. The antioxidant capacity of nanoparticles increases with increasing dissolution time. The antitumor effects of ACE polysaccharides, ACE polysaccharides encapsulated in silica, or silica-chitosan nanoparticles increased with increasing concentration of nanoparticles. This is the first report demonstrating the potential of ACE polysaccharides encapsulated in chitosan-silica nanoparticles for cancer chemoprevention. Furthermore, this study suggests that antiproliferative effect of nanoparticle-encapsulated bioactive could significantly depend on the metabolic activity type of the cell line.

  19. Poly(lactic-co-glycolic acid) encapsulated gadolinium oxide nanoparticles for MRI-based cell tracking.

    Science.gov (United States)

    Bennewitz, Margaret F; Williams, Simone S; Nkansah, Michael K; Shapiro, Erik M

    2013-06-01

    Superparamagnetic iron oxide particles have proven useful for cell tracking applications by monitoring cell transplantation and migration in living organisms. However, one perceived drawback is that these particles cause dark contrast in MRI, sometimes yielding confusion with other biological phenomena, which also yield dark contrast. To that end, researchers have investigated the use of gadolinium oxide (Gd2O3) based contrast agents for MRI-based cell tracking, as Gd2O3 has favorable r1 molar relaxivity. We synthesized Gd2O3 nanocrystals and encapsulated them within PLGA matrices to form approximatley to 150 nm nanoparticles. r1 was 1.9 mM(-1) sec(-1) and r2 was 8.4 mM(-1) sec(-1). Cell labeling with particles was well tolerated by cells except at very high doses. MRI of labeled cells showed that labeled cells could achieve both R1 and R2 enhancements due to the internalized particles. R2 enhancements were approximately to twice that of R1 enhancements suggesting the use of very short echo times when using Gd2O3 based contrast agents for MRI-based cell tracking.

  20. Ambient-processed colloidal quantum dot solar cells via individual pre-encapsulation of nanoparticles.

    Science.gov (United States)

    Debnath, Ratan; Tang, Jiang; Barkhouse, D Aaron; Wang, Xihua; Pattantyus-Abraham, Andras G; Brzozowski, Lukasz; Levina, Larissa; Sargent, Edward H

    2010-05-05

    We report colloidal quantum dot solar cells fabricated under ambient atmosphere with an active area of 2.9 mm(2) that exhibit 3.6% solar power conversion efficiency. The devices are based on PbS tuned via the quantum size effect to have a first excitonic peak at 950 nm. Because the formation of native oxides and sulfates on PbS leads to p-type doping and deep trap formation and because such dopants and traps dramatically influence device performance, prior reports of colloidal quantum dot solar cells have insisted on processing under an inert atmosphere. Here we report a novel ligand strategy in which we first encapsulate the quantum dots in the solution phase with the aid of a strongly bound N-2,4,6-trimethylphenyl-N-methyldithiocarbamate ligand. This allows us to carry out film formation and all subsequent device fabrication under an air atmosphere.