Chondroitin Sulfate C Research Articles

Delivery of Nitric Oxide by Chondroitin Sulfate C Increases the Rate of Wound Healing through Immune Modulation.

Chronic wounds impact 2.5% of the United States population and will continue to be a major clinical challenge due to increases in population age, chronic disease diagnoses, and antibiotic-resistant infection. Nitric oxide (NO) is an endogenous signaling molecule that represents an attractive, simple therapeutic for chronic wound treatment due to its innate antibacterial and immunomodulatory function. Unfortunately, modulating inflammation for extended periods by low levels of NO is not possible with NO gas. Herein, we report the utility of a NO-releasing glycosaminoglycan biopolymer (GAG) for promoting wound healing. GAGs are naturally occurring biopolymers that are immunomodulatory and known to be involved in the native wound healing process. Thus, the combination of NO and GAG biopolymers represents an attractive wound therapeutic due to these known independent roles. The influence and contribution of chondroitin sulfate C (CSC) modified to facilitate controlled and targeted delivery of NO (CSC-HEDA/NO) was evaluated using in vitro cell proliferation and migration assays and an in vivo wound model.

Chondroitin sulfate-based microneedles for transdermal delivery of stem cell-derived extracellular vesicles to treat rheumatoid arthritis

For the non-invasive treatment of rheumatoid arthritis (RA), a chondroitin sulfate C (CSC)-based dissolving microneedles (cMN) was prepared to deliver human adipose stem cell-derived extracellular vesicles (hASC-EV) into inflamed joints. Owing to their anti-inflammatory function, the hASC-EV-bearing cMN (EV@cMN) significantly suppressed activated fibroblast-like synoviocytes (aFLS) and M1 macrophages (M1), which are responsible for the progression of RA. In addition, EV@cMN facilitated the chondrogenic differentiation of bone marrow-derived stem cells. In mice with collagen-induced arthritis, EV@cMN efficiently delivered both hASC-EV and CSC to inflamed joints. Interestingly, pro-inflammatory cytokines in the inflamed joints were remarkably downregulated by the synergistic effect of CSC and hASC-EV. Consequently, as judged from the overall clinical score and joint swelling, EV@cMN showed an outstanding therapeutic effect, even comparable to the wild-type mice, without significant adverse effects. Overall, EV@cMN might have therapeutic potential for RA by efficiently delivering CSC and hASC-EV into the inflamed joints in a non-invasive manner.

The structures and applications of microbial chondroitin AC lyase.

As an important glycosaminoglycan hydrolase, chondroitin lyases can hydrolyze chondroitin sulfate (CS) and release disaccharides and oligosaccharides. They are further divided into chondroitin AC, ABC, and B lyases according to their spatial structure and substrate specificity. Chondroitin AC lyase can hydrolyze chondroitin sulfate A (CS-A), chondroitin sulfate C (CS-C), and hyaluronic acid (HA), making it an essential biocatalyst for the preparation of low molecular weight chondroitin sulfate, analysis of the structure of the chondroitin sulfate, treatment of spinal cord injury, and purification of heparin. This paper provides an overview of reported chondroitin AC lyases, including their properties and the challenges faced in industrial applications. Up to now, although many attempts have been adopted to improve the enzyme properties, the most important factors are still the low activity and stability. The relations between the stability of the enzyme and the spatial structure were also summarized and discussed. Also perspectives for remodeling the enzymes with protein engineering are included.

Chondroitin sulfate expression around motoneurons changes after complete spinal transection of neonatal rats

Hind limb locomotor activity spontaneously recovers after complete spinal transection (CST) in neonatal rats, but the mechanisms underlying the recovery are poorly understood. The perineuronal net (PNN) surrounding the neuronal cell bodies comprises an extracellular matrix that regulates neuronal plasticity during development. Here, we examined the expression of chondroitin sulfate (CS), a major component of the PNN, on motoneurons after CST in neonatal rats, and compared it with that in juvenile rats, in which hindlimb locomotor activity does not recover spontaneously.The spinal cord was transected at the mid-thoracic level in neonatal (postnatal day 5 [P5] and P10) and juvenile (P15 and P20) rats. Two weeks after CST, the percentage of motoneurons surrounded by chondroitin sulfate C (CS-C) − positive structures was significantly lower in rats with CST at P10 than in intact rats, and tended to be higher in rats with CST at P15 than in intact rats. The percentage of motoneurons with CS-A − positive structures was significantly lower in rats with CST at P15 than in intact rats.These findings suggest that CS-A and CS-C are differentially expressed in the PNNs in rats with CST. The decrease in CS-C − positive PNNs might facilitate the formation of new synaptic contacts to motoneurons, resulting in the recovery of the hindlimb locomotor activity in rats with CST during the neonatal period.

Substrate specificity of Chondroitinase ABC I based on analyses of biochemical reactions and crystal structures in complex with disaccharides

Chondroitinase ABC I (cABC-I) is the enzyme which cleaves the β-1,4 glycosidic linkage of chondroitin sulfate (CS) by β-elimination. To elucidate more accurately the substrate specificity of cABC-I, we evaluated the kinetic parameters of cABC-I and its reactivity with CS isomers displaying less structural heterogeneity as substrates, e.g., approximately 90 percent of disaccharide units in Chondroitin sulfate A (CSA) or Chondroitin sulfate C (CSC) is D-glucuronic acid and 4-O-sulfated N-acetyl galactosamine (GalNAc) (A-unit) or D-glucuronic acid and 6-O-sulfated GalNAc (C-unit), respectively. cABC-I showed the highest reactivity to CSA and CSC among all CS isomers, and the kcat/Km of cABC-I was higher for CSA than for CSC. Next, we determined the crystal structures of cABC-I in complex with CS disaccharides, and analyzed the crystallographic data in combination with molecular docking data. Arg500 interacts with 4-O-sulfated and 6-O-sulfated GalNAc residues. The distance between Arg500 and the 4-O-sulfate group was 0.8 Å shorter than that between Arg500 and the 6-O-sulfated group. Moreover, it is likely that the 6-O-sulfated group is electrostatically repulsed by the nearby Asp490. Thus, we demonstrated that cABC-I has the highest affinity for the CSA richest in 4-O-sulfated GalNAc residues among all CS isomers. Recently, cABC-I was used to treat lumbar disc herniation. The results provide useful information to understand the mechanism of the pharmacological action of cABC-I.

Comparative Study of Electrospun Scaffolds Containing Native GAGs and a GAG Mimetic for Human Mesenchymal Stem Cell Chondrogenesis.

Articular cartilage has limited healing and self-repair capability. Damage to articular cartilage becomes irreversible leading to osteoarthritis, which can impact a person's quality of life. Approximately, 5-10% of cartilage tissue is made up of sulfated glycosaminoglycans (GAGs), which sequester growth factors as well as provide structural integrity to the native cartilage tissue. This study evaluated the chondrogenic differentiation of human mesenchymal stem cells (MSCs) on gelatin-based scaffolds containing partially sulfated cellulose (pSC), a GAG mimetic derived from cellulose, in comparison to native GAGs, chondroitin sulfate-A (CS-A) and chondroitin sulfate-C (CS-C), where pSC has similarity to CS-C in terms of degree and pattern of sulfation. Scaffolds were prepared by electrospinning gelatin with pSC or the native GAGs. All scaffolds consist of fibers having average diameters of approximately 3μm and inter-fiber spacing of approximately 30μm and were hydrolytically stable throughout the culture. MSCs cultured on pSC containing scaffolds showed early production of sulfated GAGs and higher collagen type II to type I ratio than native GAGs. Among the native GAGs, chondrogenesis was promoted to a greater extent for CS-C in comparison to CS-A containing scaffolds, which suggests the pattern of sulfation impacts chondrogenesis. Partially sulfated cellulose could be used as a potential GAG mimic for cartilage tissue engineering applications.

Size of Heparin-Imprinted Nanoparticles Reflects the Matched Interactions with the Target Molecule.

It has been shown that the faradic current at an electrode grafted with molecularly imprinted polymer (MIP) is sensitive to the specific target molecule used as the template. This phenomenon is applicable to sensors with very high selectivity, but the sensing mechanism is still a black box. We investigated the size sensitivity of nanoparticles of molecularly imprinted polymers (MIP-NPs) to a specific interaction for determination of the mechanism of the gate effect and its feasibility for new applications. Nanoparticles of poly(methacryloxy ethyl trimethylammonium chloride-co-acrylamide-co-methylenebisacrylamide) imprinted with heparin immobilized on glass beads were synthesized. The diameter of the MIP-NPs of heparin was increased by the presence of the heparin template but was insensitive to chondroitin sulfate C (CSC), the analogue of heparin. The high selectivity of the MIP-NPs was consistent with the selectivity of electrodes grafted with a heparin-imprinted polymer in our previous studies. The quartz crystal microbalance probes immobilizing heparin or CSC were sensitive to MIP-NPs, which indicates that the binding ability of MIP-NP does not discriminate between the template and other glycosaminoglycans. These results indicate that the size of the MIP-NP is sensitive to the matched binding with the template through the imprinted cavity.

Investigation of glycosaminoglycan mimetic scaffolds for neurite growth

Spinal cord injury can lead to severe dysfunction as a result of limited nerve regeneration that is due to an inhibitory environment created at the site of injury. Neural tissue engineering using materials that closely mimic the extracellular matrix (ECM) during neural development could enhance neural regeneration. Glycosaminoglycans (GAGs), which are sulfated polysaccharides, have been shown to modulate axonal outgrowth in neural tissue depending upon the position and degree of sulfation. Cellulose sulfate (CelS), which is a GAG mimetic, was evaluated for its use in promoting neurite extension. Aligned fibrous scaffolds containing gelatin blended with 0.25% partially sulfated cellulose sulfate (pCelS), having sulfate predominantly at the 6-carbon position of the glucose monomer unit, and fully sulfated cellulose sulfate (fCelS), which is sulfated at the 2-, 3-, and 6-carbon positions of the glucose monomer unit, were fabricated using the electrospinning method. Comparisons were made with scaffolds containing native GAGs, chondroitin sulfate-A (CS-A) and chondroitin sulfate-C (CS-C), which were obtained from commercial sources. CS-A and CS-C are present in neural tissue ECM. The degree of sulfation and position of sulfate groups was determined using elemental analysis, Fourier-transform infrared spectroscopy (FTIR), Raman microspectroscopy, and 13C nuclear magnetic resonance (NMR). In vitro studies examined both nerve growth factor (NGF) binding on scaffolds and neurite extension by dorsal root ganglion (DRG) neurons. NGF binding was highest on scaffolds containing pCelS and fCelS. Neurite extension was greatest for scaffolds containing fCelS followed by pCelS, with the lowest outgrowth on the CS-A containing scaffolds, suggesting that the degree and position of sulfation of CelS was permissible for neurite outgrowth. This study demonstrated that cellulose sulfate, as a GAG mimetic, could be used for future neural tissue regeneration application. Statement of signficanceScaffolds that closely mimic the native extracellular matrix (ECM) during development may be a promising approach to enhance neural regeneration. Here, we reported a glycosaminoglycan (GAG) mimetic derived from cellulose that promotes neurite extension over native GAGs, chondroitin sulfate-A (CS-A) and chondroitin sulfate-C (CS-C), which are present in neural ECM. Depending upon the degree and position of sulfation, the GAG mimetic can impact nerve growth factor binding and permissive neurite outgrowth.

Chondroitin Sulfate Expression in Perineuronal Nets After Goldfish Spinal Cord Lesion.

Perineuronal nets (PNNs) surrounding neuronal cell bodies regulate neuronal plasticity during development, but their roles in regeneration are unclear. In the PNNs, chondroitin sulfate (CS) is assumed to be involved in inhibiting contact formation. Here, we examined CS expression in PNNs in the ventral horn of a goldfish hemisected spinal cord in which descending axons regenerate beyond the lesion to connect with distal spinal neurons. In intact fish, chondroitin sulfate A (CS-A)–positive PNNs accounted for 5.0% of HuC/D-immunoreactive neurons, and 48% of choline acetyltransferase (ChAT)-immunoreactive neurons. At 2, 4 and 8 weeks after spinal hemisection, CS-A–positive PNNs accounted for 8.4%–9.9% of HuC/D-immunoreactive neurons, and 50%–60% of ChAT-immunoreactive neurons, which was not significantly different from intact fish. Chondroitin sulfate C (CS-C)–positive PNNs accounted for 6.4% of HuC/D-immunoreactive neuron, and 67% of ChAT-immunoreactive neurons in intact fish. At 2, 4 and 8 weeks after spinal hemisection, CS-C–positive PNNs accounted for 7.9%, 5.5% and 4.3%, respectively, of HuC/D-immunoreactive neurons, and 65%, 52% and 42%, respectively, of ChAT-immunoreactive neurons, demonstrating a significant decrease at 4 and 8 weeks after spinal hemisection. Among ventral horn neurons that received descending axons labeled with tetramethylrhodamine dextran amine (RDA) applied at the level of the first spinal nerve, CS-A–positive PNNs accounted for 53% of HuC/D-immunoreactive neurons. At 2 and 4 weeks after spinal hemisection, CS-A–positive PNNs accounted for 57% and 56% of HuC/D-immunoreactive neurons, which was not significantly different from intact fish. CS-C–positive PNNs, accounted for 48% of HuC/D-immunoreactive neurons that received RDA-labeled axons. At 2 and 4 weeks after spinal hemisection, CS-C–positive PNNs significantly decreased to 22% of the HuC/D-immunoreactive neurons, and by 4 weeks after spinal hemisection they had returned to 47%. These findings suggest that CS expression is maintained in the PNNs after spinal cord lesion, and that the descending axons regenerate to preferentially terminate on neurons not covered with CS-C–positive PNNs. Therefore, CS-C in the PNNs possibly inhibits new contact with descending axons, and plasticity in the spinal neurons might be endowed by downregulation of CS-C in the PNNs in the regeneration process after spinal hemisection in goldfish.

Blood heparin sensor made from a paste electrode of graphite particles grafted with molecularly imprinted polymer

A real-time heparin monitor could be used to optimize the dosage of heparin during extracorporeal circulation procedures. This report describes the development of a graphite-paste (GP) electrode with molecularly imprinted polymer (MIP) grafted onto it. Heparin-imprinted poly (methacryloxyethyltriammonium chloride -co- acrylamide -co- methylenebisacrylamide) was grafted directly onto graphite particles. The grafted particles were thoroughly mixed with oil to fabricate the MIP-GP electrode. Traditional cyclic voltammetry was performed with the electrode in physiological saline or bovine whole blood containing 5 mM ferrocyanide and 0–8 units/mL heparin. The current intensity increased with heparin concentration, due to expansion of the effective surface area resulting from heparin-promoted mobility of the oil in the MIP-GP electrode. No significant difference was found in the sensitivity of the current to unfractionated heparin among the electrodes fabricated because of the electrode homogenization resulting from thorough mixing of the MIP-grafted particles and oil. (A previous MIP-grafted indium tin oxide electrode exhibited lower sensitivity in blood than in saline.) Only 60 s were needed to stabilize the current. The current at the MIP-GP electrode was also sensitive to low-molecular-weight heparin in blood, but insensitive to chondroitin sulfate C (CSC), which is a heparin analog. The non-imprinted polymer (NIP)-grafted electrode was insensitive to heparin. Thus, the MIP-GP electrode, which operated through a new heparin-sensing mechanism, is an excellent candidate for application as a disposable sensor to monitor heparin levels in blood.

A new chemical complex can rapidly concentrate lentivirus and significantly enhance gene transduction.

In this study, we developed a new purification method using chondroitin sulfate C (CSC) and protamine sulfate (PS) to concentrate lentivirus. To evaluate the efficiency of this new method, we compared it with several previously described purification protocols, including virus concentrated by ultracentrifugation (Ultra), precipitated by polyethylene glycol (PEG), and sedimented by CSC combined with polybrene (PB). After using the different methods to purify and concentrate equivalent amounts of lentivirus supernatant, the virus pellets precipitated by the different methods were resuspended using the equivalent volumes of DMEM. Subsequently, 10μl of each lentivirus stock carrying EGFP gene was used to transduce two types of cells, human embryonic kidney 293T (HEK293T) cells and mouse mesenchymal stem cells (mMSC). It was obvious that HEK293T and mMSC appeared much intensiver green fluorescence through virus transduction from PS method than from other methods. To quantitate the transduction efficiency of the viruses, we examined virus titer in the cells after transduction using a real-time PCR-based analysis. Accordingly, we verified that PS precipitation could generate virus with a higher titer (4.39×108IU/ml) than PB (2.43×108IU/ml), Ultra (1.16×108IU/ml), and PEG (0.56×108IU/ml) in HEK293T cells. As for HEK293T cellsin mMSC, the PS method also generated virus with a higher titer (4.66×108IU/ml) than the Ultra method (2.36×108IU/ml), and a much higher titer than those of the other chemical-based precipitation methods using PB (4.82×106IU/ml) and PEG (8.98×104IU/ml). Furthermore, the HEK293T cells and mMSC transduced by PS(1X)-virus appeared to have higher cell growth ratios, respectively, than the HEK293T cells and mMSC transduced by lentivirus using the other methods. We conclude that our new method for purifying lentivirus is cost-effective, time-saving, and highly efficient, and that lentivirus purification by this means could possibly be used to transduce a variety of cells, including stem cells.

Studies of the antitumor mechanism of action of dermaseptin B2, a multifunctional cationic antimicrobial peptide, reveal a partial implication of cell surface glycosaminoglycans.

Dermaseptin-B2 (DRS-B2) is a multifunctional cationic antimicrobial peptide (CAP) isolated from frog skin secretion. We previously reported that DRS-B2 possesses anticancer and antiangiogenic activities in vitro and in vivo. In the present study, we evaluated the antiproliferative activity of DRS-B2 on numerous tumor cell lines, its cell internalization and studies of its molecular partners as well as their influences on its structure. Confocal microscopy using ([Alexa594]-(Cys0)-DRS-B2) shows that in sensitive human tumor cells (PC3), DRS-B2 seems to accumulate rapidly at the cytoplasmic membranes and enters the cytoplasm and the nucleus, while in less sensitive tumor cells (U87MG), DRS-B2 is found packed in vesicles at the cell membrane. Furthermore FACS analysis shows that PC3 cells viability decreases after DRS-B2 treatment while U87 MG seems to be unaffected. However, "pull down" experiments performed with total protein pools from PC3 or U87MG cells and the comparison between the antiproliferative effect of DRS-B2 and its synthetic analog containing all D-amino acids suggest the absence of a stereo-selective protein receptor. Pretreatment of PC3 cells with sodium chlorate, decreases the antiproliferative activity of DRS-B2. This activity is partially restored after addition of exogenous chondroitin sulfate C (CS-C). Moreover, we demonstrate that at nanomolar concentrations CS-C potentiates the antiproliferative effect of DRS-B2. These results highlight the partial implication of glycosaminoglycans in the mechanism of antiproliferative action of DRS-B2. Structural analysis of DRS-B2 by circular dichroism in the presence of increasing concentration of CS-C shows that DRS-B2 adopts an α-helical structure. Finally, structure-activity-relationship studies suggest a key role of the W residue in position 3 of the DRS-B2 sequence for its antiproliferative activity.

Chondroitin sulfates do not impede axonal regeneration in goldfish spinal cord

Chondroitin sulfate proteoglycans produced in glial scar tissue are a major inhibitory factor for axonal regeneration after central nervous system injury in mammals. The inhibition is largely due to chondroitin sulfates, whose effects differ according to the sulfation pattern. In contrast to mammals, fish nerves spontaneously regenerate beyond the scar tissue after spinal cord injury, although the mechanisms that allow for axons to pass through the scar are unclear. Here, we used immunohistochemistry to examine the expression of two chondroitin sulfates with different sulfation variants at the lesion site in goldfish spinal cord.The intact spinal cord was immunoreactive for both chondroitin sulfate-A (CS-A) and chondroitin sulfate-C (CS-C), and CS-A immunoreactivity overlapped extensively with glial processes positive for glial fibrillary acidic protein. At 1week after inducing the spinal lesion, CS-A immunoreactivity was observed in the cell bodies and extracellular matrix, as well as in glial processes surrounding the lesion center. At 2weeks after the spinal lesion, regenerating axons entering the lesion center overtook the CS-A abundant area. In contrast, at 1week after lesion induction, CS-C immunoreactivity was significantly decreased, and at 2weeks after lesion induction, CS-C immunoreactivity was observed along the regenerating axons entering the lesion center. The present findings suggest that after spinal cord injury in goldfish, chondroitin sulfate proteoglycans are deposited in the extracellular matrix at the lesion site but do not form an impenetrable barrier to the growth of regenerating axons.

* Gelatin Scaffolds Containing Partially Sulfated Cellulose Promote Mesenchymal Stem Cell Chondrogenesis.

Articular cartilage has a limited capacity to heal after damage from injury or degenerative disease. Tissue engineering constructs that more closely mimic the native cartilage microenvironment can be utilized to promote repair. Glycosaminoglycans (GAGs), a major component of the cartilage extracellular matrix, have the ability to sequester growth factors due to their level and spatial distribution of sulfate groups. This study evaluated the use of a GAG mimetic, cellulose sulfate, as a scaffolding material for cartilage tissue engineering. Cellulose sulfate can be synthesized to have a similar level and spatial distribution of sulfates as chondroitin sulfate C (CSC), the naturally occurring GAG. This partially sulfated cellulose (pSC) was incorporated into a fibrous gelatin construct by the electrospinning process. Scaffolds were characterized for fiber morphology and overall stability over time in an aqueous environment, growth factor interaction, and for supporting mesenchymal stem cell (MSC) chondrogenesis in vitro. All scaffold groups had micron-sized fibers and maintained overall stability in aqueous environments. Increasing concentrations of the transforming growth factor-beta 3 (TGF-β3) were detected on scaffolds with increasing pSC. MSC chondrogenesis was enhanced on the scaffold with the highest pSC concentration as seen with the highest collagen type II production, collagen type II immunostaining, expression of cartilage-specific genes, and ratio of collagen type II to collagen type I production. These studies demonstrated the potential of pSC sulfate as a scaffolding material for cartilage tissue engineering.

Biocompatible complex coated with glycosaminoglycan for gene delivery

The purpose of this study was to develop a ternary complex of plasmid DNA (pDNA) electrostatically assembled with dendrigraft poly-l-lysine (DGL) and biodegradable glycosaminoglycan for effective and secure gene delivery. High gene expression of pDNA/DGL complex was confirmed with slight cytotoxicity and erythrocyte agglutination. Anionic ternary complexes of 55.4–223.8 nm were formed by the addition of a glycosaminoglycan such as chondroitin sulfate A (CS-A), chondroitin sulfate B (CS-B), chondroitin sulfate C (CS-C) or hyaluronic acid (HA). Using the cell line B16-F10, most of the ternary complexes showed only weak gene expression and little cytotoxicity, although the pDNA/DGL/CS-A complexes maintained a certain level of gene expression. In particular, the pDNA/DGL/CS-A8 complexes showed significantly higher gene expression than pDNA/DGL complexes in the presence of fetal bovine serum. Gene expression from the pDNA/DGL/CS-A8 complex was inhibited by a high concentration of CS-A and endocytosis inhibitors. After intravenous administration of the pDNA/DGL/CS-A8 complex and the pDNA/DGL complex into ddY mice, high gene expression was observed in the reticuloendothelial systems, the pDNA/DGL/CS-A complex is expected to be useful for gene therapy.

Systematic protein interactome analysis of glycosaminoglycans revealed YcbS as a novel bacterial virulence factor.

Microbial pathogens have evolved several strategies for interacting with host cell components, such as glycosaminoglycans (GAGs). Some microbial proteins involved in host–GAG binding have been described; however, a systematic study on microbial proteome–mammalian GAG interactions has not been conducted. Here, we used Escherichia coli proteome chips to probe four typical mammalian GAGs, heparin, heparan sulphate (HS), chondroitin sulphate B (CSB), and chondroitin sulphate C (CSC), and identified 185 heparin-, 62 HS-, 98 CSB-, and 101 CSC-interacting proteins. Bioinformatics analyses revealed the unique functions of heparin- and HS-specific interacting proteins in glycine, serine, and threonine metabolism. Among all the GAG-interacting proteins, three were outer membrane proteins (MbhA, YcbS, and YmgH). Invasion assays confirmed that mutant E. coli lacking ycbS could not invade the epithelial cells. Introducing plasmid carrying ycbS complemented the invading defects at ycbS lacking E. coli mutant, that can be further improved by overexpressing ycbS. Preblocking epithelial cells with YcbS reduced the percentage of E. coli invasions. Moreover, we observed that whole components of the ycb operon were crucial for invasion. The displacement assay revealed that YcbS binds to the laminin-binding site of heparin and might affect the host extracellular matrix structure by displacing heparin from laminin.

Study of the enantioseparation capability of chiral dual system based on chondroitin sulfate C in CE.

It has been reported that chiral dual system is able to improve the enantioseparation of enantiomers in many cases. Currently, the dual systems involved in CE chiral separation are mostly dual CDs systems, and the polysaccharides-based chiral dual system was reported in only one paper. To the best of our knowledge, the use of chondroitin sulfate C (CSC)-based dual system for enantiomeric separation has not been reported previously. Herein, four CSC-based chiral dual systems, namely CSC/glycogen, CSC/chondroitin sulfate A (CSA), CSC/hydroxypropyl-β-CD (HP-β-CD), as well as CSC/β-CD (β-CD), were evaluated for the first time for their enantioseparation capability by CE in this paper. During the course of the work, the influences of chiral selector concentration and buffer pH values on enantioseparation in dual systems were systematically investigated. Under the optimized conditions, the dual system consisting of CSC and glycogen exhibited better separations toward nefopam, duloxetine, sulconazole, atenolol, laudanosine, and cetirizine enantiomers compared to the single CSC or glycogen system. The combination of CSC and HP-β-CD improved the separation of amlodipine and chlorphenamine enantiomers. However, no synergistic effect was observed in the CSC/CSA and CSC/β-CD systems.

Effect of chondroitin sulphate C on thein vitroandin vivochondrogenesis of mesenchymal stem cells in crosslinked type II collagen scaffolds

This study evaluates the crosslinkage effect of chondroitin sulphate C (CSC) and type II collagen (COL II) on chondrogenesis of mesenchymal stem cells (MSCs) in vitro and in vivo. In the in vitro studies, our results show that the weight ratio CSC:COL II that reaches 1.2:100 (CSC1.2/100 -COL II scaffold) can provide an optimal microenvironment for MSC chondrogenesis. When MSCs are cultured in this CSC1.2/100 -COL II scaffold, the chondrogenic gene expression of cultured cells is upregulated, while the osteogenic gene expression of these is downregulated. In addition, MSCs cultivated in the CSC1.2/100 -COL II scaffold are found to express the highest glycosaminoglycans:DNA ratio as compared to those in scaffolds of other CSC:COL II ratios. Histological and immunohistological evidence also supports the result. In the in vivo study, our results show that MSCs cultivated in the CSC1.2/100 -COL II scaffold demonstrate a better repair ability on cartilage lesions than does the COL II scaffold. After 1 month in vivo, the injected MSCs in the CSC1.2/100 -COL II scaffold show lacuna structures and stimulate the formation of type II collagen at the defective sites. Six months after transplantation, the generated cells in the CSC1.2/100 -COL II group show higher gene expressions of type II collagen and aggrecan but lower gene expression of type I collagen at the defective sites than those in the COL II group. The results strongly suggest that CSC1.2/100 -COL II scaffold can serve as a potential candidate for cartilage repair and improve the chondrogenesis of MSCs in general.

Gangliosides and chondroitin sulfate desensitize and internalize B2 bradykinin receptors

Prolonged or repeated agonist activation of G-protein-coupled receptors (GPCRs) initiates their desensitization and internalization, rendering them unresponsive to agonist activation. We analyzed how gangliosides and chondroitin sulfate affect B2 bradykinin (BK) receptors (B2Rs). Gangliosides and chondroitin sulfate did not stimulate intracellular Ca2+ release from B2R-expressing CHO-K1 cells, but repeated exposure desensitized B2Rs to BK stimulation. Microscopic observation of DsRed-fused B2Rs revealed that several gangliosides and chondroitin sulfate C (CSC) effectively internalized B2Rs. Ganglioside-CSC treatment of B2R mutant-expressing cells failed to desensitize and internalize the mutant receptors. As this mutant lacks the first extracellular domain and cannot activate GPCR kinase (GRK), gangliosides and CSC likely initiate B2R desensitization and endocytosis through GRK-mediated B2R phosphorylation.

Effects of chondroitin sulfate-C on articular cartilage destruction in murine collagen-induced arthritis.

The effects of chondroitin sulfate-C (CAS 25322-46-7, Chondroitin ZS Tab) on type II collagen (CII)-induced arthritis (CIA) in mice were evaluated. DBA/1J mice were immunized with bovine CII emulsified in Freund's complete adjuvant, followed by a booster injection 21 days later. Chondroitin sulfate-C at doses of 100, 300 and 1000 mg/kg was administered orally once daily beginning 14 days before initial immunization. An arthritis index and hind paw edema were examined from day 0 to day 49, when the mice were killed by ether anesthesia for histopathological examination. The delayed-type hypersensitivity (DTH) reaction, serum anti-CII antibody titer, and histopathologic characteristics of both synovitis and destruction of articular cartilage were analyzed. Both the arthritis index and the serum anti-CII antibody titer were reduced by treatment with chondroitin sulfate-C in a dose-dependent manner. Chondroitin sulfate-C (1000 mg/kg) significantly inhibited hind paw edema, synovitis and destruction of the articular cartilage, but not DTH reaction.