Microbiological-Chemical Sourced Chondroitin Sulfates Protect Neuroblastoma SH-SY5Y Cells against Oxidative Stress and Are Suitable for Hydrogel-Based Controlled Release
TL;DR: In this paper, the neuroprotective role of released chondroitin sulfates (CS) against H2O2-induced oxidative stress was evaluated using a 1.9-dimethylmethylene blue (DMMB) assay.
Abstract: Chondroitin sulfates (CS) are a class of sulfated glycosaminoglycans involved in many biological processes. Several studies reported their protective effect against neurodegenerative conditions like Alzheimer’s disease. CS are commonly derived from animal sources, but ethical concerns, the risk of contamination with animal proteins, and the difficulty in controlling the sulfation pattern have prompted research towards non-animal sources. Here we exploited two microbiological-chemical sourced CS (i.e., CS-A,C and CS-A,C,K,L) and Carbopol 974P NF/agarose semi-interpenetrating polymer networks (i.e., P.NaOH.0 and P.Ethanol.0) to set up a release system, and tested the neuroprotective role of released CS against H2O2-induced oxidative stress. After assessing that our CS (1–100 µM) require a 3 h pre-treatment for neuroprotection with SH-SY5Y cells, we evaluated whether the autoclave type (i.e., N- or B-type) affects hydrogel viscoelastic properties. We selected B-type autoclaves and repeated the study after loading CS (1 or 0.1 mg CS/0.5 mL gel). After loading 1 mg CS/0.5 mL gel, we evaluated CS release up to 7 days by 1,9-dimethylmethylene blue (DMMB) assay and verified the neuroprotective role of CS-A,C (1 µM) in the supernatants. We observed that CS-A,C exhibits a broader neuroprotective effect than CS-A,C,K,L. Moreover, sulfation pattern affects not only neuroprotection, but also drug release.
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TL;DR: In this paper , the authors proposed a highly controllable method to optimize the printability of internally crosslinked polysaccharides, without the need for additives or post-printing treatments.
Abstract: Introduction: In the view of 3D-bioprinting with cell models representative of neural cells, we produced inks to mimic the basic viscoelastic properties of brain tissue. Moving from the concept that rheology provides useful information to predict ink printability, this study improves and expands the potential of the previously published 3D-reactive printing approach by introducing pH as a key parameter to be controlled, together with printing time. Methods: The viscoelastic properties, printability, and microstructure of pectin gels crosslinked with CaCO3 were investigated and their composition was optimized (i.e., by including cell culture medium, HEPES buffer, and collagen). Different cell models representative of the major brain cell populations (i.e., neurons, astrocytes, microglial cells, and oligodendrocytes) were considered. Results and Discussion: The outcomes of this study propose a highly controllable method to optimize the printability of internally crosslinked polysaccharides, without the need for additives or post-printing treatments. By introducing pH as a further parameter to be controlled, it is possible to have multiple (pH-dependent) crosslinking kinetics, without varying hydrogel composition. In addition, the results indicate that not only cells survive and proliferate following 3D-bioprinting, but they can also interact and reorganize hydrogel microstructure. Taken together, the results suggest that pectin-based hydrogels could be successfully applied for neural cell culture.
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TL;DR: A review of the advancements in metabolic engineering strategies and the biotechnological processes towards chondroitin manufacturing can be found in this paper , where the authors present an overview of the chemical approaches applied to obtain specific structural features and targeted decoration.
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TL;DR: In this paper, an exponential relation M t /M ∞ = kt n may be used to describe the Fickian and non-Fickian release behavior of release systems which are prepared by incorporation of a drug in a hydrophilic, initially glassy polymer.
3,522 citations
TL;DR: A modified form of the dim methylmethylene blue assay is described that has improved specificity for sulphated glycosaminoglycans, and it is shown that in conjunction with specific polysaccharidases, the dimethylmethyleneblue assay can be used to quantitate individual sulphated sugarcans.
3,345 citations
TL;DR: A rapid spectrophotometric procedure is described for the estimation of sulfated glycosaminoglycans in cartilage cultures that is substantially free from interference, is sensitive to less than 1 microgram (4 micrograms/ml) of chondroitin sulfate, and provides a simple alternative to the traditional methods for gly cosaminoglycan determinations.
Abstract: A rapid spectrophotometric procedure is described for the estimation of sulfated glycosaminoglycans in cartilage cultures. Papain digestion of tissue or culture medium provides glycosaminoglycans in solution for assay; an aliquot of the digest is mixed with the dye 1,9-dimethylmethylene blue. The assay is based on the metachromatic shift in absorption maximum which occurs when the dye is complexed with sulfated glycosaminoglycans. The reagent is stable, and the method is substantially free from interference, is sensitive to less than 1 microgram (4 micrograms/ml) of chondroitin sulfate, and provides a simple alternative to the traditional methods for glycosaminoglycan determinations.
1,307 citations
TL;DR: The inability of the adult glial scar tissue to support neurite outgrowth was best correlated with the expression of CS-PG and CT, suggesting that these molecules may be involved in limiting the growth of regenerating axons in the CNS after injury.
Abstract: The extracellular matrix (ECM) molecules chondroitin-6-sulfate proteoglycan (CS-PG) and cytotactin/tenascin (CT), present on subpopulations of astroglia or their precursors during development, can inhibit neurite outgrowth in vitro. However, it is not known whether these molecules are expressed within the mature CNS following injury, where they could contribute to regenerative failure. Thus, the expression of various ECM molecules that affect axon growth was examined in areas of reactive gliosis caused by implanting a piece of nitrocellulose into the cortex of neonatal and adult animals. The expression of these molecules was compared to the amount of neurite outgrowth that occurred in vitro when the damaged CNS tissue from animals of various ages was removed intact and used as a substrate in explant culture. The results demonstrate that the growth-promoting molecules laminin, collagen type IV, and fibronectin were present around the implant in all experimental groups. In comparison, CS-PG and CT were present within and around the area of the lesion only in adult animals. In vivo, these molecules were colocalized with intensely glial fibrillary acidic protein (GFAP)-positive astrocytes in and immediately adjacent to the scar, but not with other equally intensely GFAP-positive astrocytes in the cortex away from the site of injury. CT and CS-PG were present in gray matter areas of the cortex that had been directly damaged during the implant procedure and in the corpus callosum when lesioned during implantation. In vitro, the glial tissue removed from the lesion site of neonatal animals supported neurite outgrowth, while scars removed from adult animals did not. The inability of the adult glial scar tissue to support neurite outgrowth was best correlated with the expression of CS-PG and CT, suggesting that these molecules may be involved in limiting the growth of regenerating axons in the CNS after injury.
1,208 citations
TL;DR: In this paper, the authors give an overview of the recent design concepts of IPN hydrogels and their applications in controlled drug delivery, and separation processes, and a special concern is given to the macroporous IPN composite cryogels, which are very attractive materials for separation processes being endowed with a high reusability.
709 citations