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General strategies for synthesizing sulfated oligosaccharides employ the protection of sulfate groups for glycosylation or post‐sulfation after the synthesis of oligosaccharides. However, the chemical behavior of free sulfate groups in glycosylation reactions has not been thoroughly studied. We examined several glycosyl donors with free sulfate groups, but neither glycosyl imidates nor thioglycosides achieved products. Conversely, activating 6‐sulfated GalNAc donors with either a 2‐NTroc group or 2‐O‐acetyl group using an ortho‐hexynylbenzoate group at the anomeric position yielded β‐glycosides in good yield. These results indicate that glycosylations with a free sulfate group can be performed and that the neighboring group participation of 2‐NHTroc and 2‐O‐acetyl groups works even with an unprotected sulfate group at the 6‐position. Ab initio calculations supported the formation of acyloxonium‐cation via 2‐O‐acyl participation. Additionally, glycosylation reactions under various counter cations of the sulfate group, such as Na+, Li+, K+, Ba2+, and pyridinium were examined. Results showed that sodium and lithium salt donors yielded the products in good yield. These results were also supported by ab initio calculations. Practical glycosylation reactions between disaccharide donor‐acceptor pairs with free sulfate groups successfully yielded the sulfated tetrasaccharides. This study also discusses how a sodium salt acts as a protecting group during glycosylation. [ABSTRACT FROM AUTHOR]

Glycosaminoglycans are often deprioritized as targets for synthetic immunotherapy due to the complexity of glyco-epitopes and limited options for obtaining specific subtype binding. Solid tumors express proteoglycans that are modified with oncofetal chondroitin sulfate (CS), a modification normally restricted to the placenta. Here, we report the design and functionality of transient chimeric antigen receptor (CAR) T cells with selectivity to oncofetal CS. Following expression in T cells, the CAR could be "armed" with recombinant VAR2CSA lectins (rVAR2) to target tumor cells expressing oncofetal CS. While unarmed CAR T cells remained inactive in the presence of target cells, VAR2-armed CAR T cells displayed robust activation and the ability to eliminate diverse tumor cell types in vitro. Cytotoxicity of the CAR T cells was proportional to the concentration of rVAR2 available to the CAR, offering a potential molecular handle to finetune CAR T cell activity. In vivo, armed CAR T cells rapidly targeted bladder tumors and increased the survival of tumor-bearing mice. Thus, our work indicates that cancer-restricted glycosaminoglycans may be exploited as potential targets for CAR T cell therapy. Synopsis: Transient VAR2-[SpyT][SpyC]-CAR T cells targeting oncofetal chondroitin sulfate (CS) show high specificity and activity against solid tumor cells, adding glycosaminoglycans to the list of potential actionable targets for CAR T cell therapy in solid tumors. Transient CAR T cells were developed to target oncofetal CS, unique to cancer cells. VAR2-[SpyT]-armed CAR T cells demonstrated robust activation, cytokine production, and cytotoxicity in vitro. The targeting mechanism offers high specificity and potential broad applicability across various solid tumors. VAR2-[SpyT][SpyC]-CAR T cells inhibited tumor growth and increased survival in vivo. The VAR2-[SpyT][SpyC]-CAR T cell system provides proof-of-concept for oncofetal CS as a potential target for CAR T cell approaches. Transient VAR2-[SpyT][SpyC]-CAR T cells targeting oncofetal chondroitin sulfate (CS) show high specificity and activity against solid tumor cells, adding glycosaminoglycans to the list of potential actionable targets for CAR T cell therapy in solid tumors. [ABSTRACT FROM AUTHOR]

Morbidities associated with natural bone have highlighted the need of advanced bioactive implants for the replacement of damaged bone. Known for their beneficial bioactivity and biocompatibility, Nigella sativa seeds (kalonji/black seed) are traditionally used in many medications. This research focuses on the positive impact of black seed extract on the chondroitin sulfate‐based hydroxyapatite nanocomposite for bone tissue regeneration applications. Novel nanocomposites with varied concentrations of black seed extract (0, 1, 2, and 3 wt%) were fabricated through a coprecipitation approach and analyzed for physicochemical and biological applications. The results of X‐ray diffraction and FTIR spectroscopy revealed that the HCSK3 nanocomposite was successfully developed with appreciably strong molecular interactions among their components. HCSK3 nanocomposite exhibits higher rough surface morphology (Ra −1.13 µm and Rz −4.8 µm), uniform particle distribution (average particle size −9.9 nm), and good thermal stability with total weight loss of 13%. It also provides better nucleation sites for apatite formation and protein adsorption stimulating cell attachment, proliferation, and differentiation. Moreover, HCSK3 nanocomposite showed excellent cell viability, higher ARS/ALP activities, better antibacterial property, and nonhemolytic property (below 3%). Thus, it can be concluded that HCSK3 nanocomposite manifested the highest biocompatibility and negligible cell toxicity. Therefore, it can be employed as an ideal bone implant in the bone tissue restoration phenomenon. [ABSTRACT FROM AUTHOR]

Accumulation of abnormal chondroitin sulfate (CS) chains in breast cancer tissue is correlated with poor prognosis. However, the biological functions of these CS chains in cancer progression remain largely unknown, impeding the development of targeted treatment focused on CS. Previous studies identified chondroitin polymerizing factor (CHPF; also known as chondroitin sulfate synthase 2) is the critical enzyme regulating CS accumulation in breast cancer tissue. We then assessed the association between CHPF‐associated proteoglycans (PGs) and signaling pathways in breast cancer datasets. The regulation between CHPF and syndecan 1 (SDC1) was examined at both the protein and RNA levels. Confocal microscopy and image flow cytometry were employed to quantify macropinocytosis. The effects of the 6‐O‐sulfated CS‐binding peptide (C6S‐p) on blocking CS functions were tested in vitro and in vivo. Results indicated that the expression of CHPF and SDC1 was tightly associated within primary breast cancer tissue, and high expression of both genes exacerbated patient prognosis. Transforming growth factor beta (TGF‐β) signaling was implicated in the regulation of CHPF and SDC1 in breast cancer cells. CHPF supported CS–SDC1 stabilization on the cell surface, modulating macropinocytotic activity in breast cancer cells under nutrient‐deprived conditions. Furthermore, C6S‐p demonstrated the ability to bind CS‐SDC1, increase SDC1 degradation, suppress macropinocytosis of breast cancer cells, and inhibit tumor growth in vivo. Although other PGs may also be involved in CHPF‐regulated breast cancer malignancy, this study provides the first evidence that a CS synthase participates in the regulation of macropinocytosis in cancer cells by supporting SDC1 expression on cancer cells. [ABSTRACT FROM AUTHOR]

Background: Cartilage is an avascular and alymphatic tissue that lacks the intrinsic ability to undergo spontaneous repair and regeneration in the event of significant injury. The efficacy of conventional therapies for invasive cartilage injuries is limited, thereby prompting the emergence of cartilage tissue engineering as a possible alternative. In this study, we fabricated three‐dimensional hydrogel films utilizing sodium alginate (SA), gelatin (Gel), and chondroitin sulfate (CS). These films were included with Wharton's jelly mesenchymal stem cells (WJ‐MSCs) and intended for cartilage tissue regeneration. Methods: The hydrogel film that were prepared underwent evaluation using various techniques including scanning electron microscope (SEM), Fourier transform infrared (FTIR) spectroscopy, assessment of the degree of swelling, degradation analysis, determination of water vapor transmission rate (WVTR), measurement of water contact angle (WCA), evaluation of mechanical strength, and assessment of biocompatibility. The rabbit ear cartilage regeneration by hydrogel films with and without of WJ‐MSCs was studied by histopathological investigations during 15, 30, and 60 days. Results: The hydrogel films containing CS exhibited superior metrics compared to other nanocomposites such as better mechanical strength (12.87 MPa in SA/Gel compared to 15.56 in SA/Gel/CS), stability, hydrophilicity, WVTR (3103.33 g/m2/day in SA/Gel compared to 2646.67 in nanocomposites containing CS), and swelling ratio (6.97 to 12.11% in SA/Gel composite compared to 5.03 to 10.90% in SA/Gel/CS). Histopathological studies showed the presence of chondrocyte cells in the lacunae on the 30th day and the complete restoration of the cartilage tissue on the 60th day following the injury in the group of SA/Gel/CS hydrogel containing WJ‐MSCs. Conclusions: We successfully fabricated a scaffold composed of alginate, gelatin, and chondroitin sulfate. This scaffold was further enhanced by the incorporation of Wharton's jelly mesenchymal stem cells. Our findings demonstrate that this composite scaffold has remarkable biocompatibility and mechanical characteristics. The present study successfully demonstrated the therapeutic potential of the SA‐Gel‐CS hydrogel containing WJ‐MSCs for cartilage regeneration in rabbits. [ABSTRACT FROM AUTHOR]

In this work, capillary electrophoresis with Gaussian distribution‐based peak fitting for improved peak resolution for simultaneous quantitative analysis of chondroitin sulfate (CS) and dermatan sulfate (DS) was developed for the first time. The effects of different pH, voltage, and peak shapes on the peak fitting were investigated. The results indicate that separation conditions that affect peak shape have a significant impact on peak fitting. Under optimized separation conditions, the peak fitting achieves high accuracy and precision, the relative standard deviation (n = 5) of peak area and migration time of CS and DS were 1.1%, 1.8% and 0.94%, 0.84%, respectively. Furthermore, the results obtained by peak fitting were close to real values. The developed method was used to analyze the purity of CS. The results had a high consistency with the labeled value; furthermore, the peak fitting could point out the number of possible impurities. The developed method has good potential for the analysis of other glycosaminoglycans with peak overlap. [ABSTRACT FROM AUTHOR]

Purpose: The study explored the enhanced skin moisturizing capabilities and moisture retention effects achieved by forming a polyion complex using sulfated glycosaminoglycan (GAG), specifically chondroitin sulfate (CS), and amino acids (AA) such as glutamine (Q) and arginine (R). The overall hydration effect of this CS‐AA complex was examined. Methods: After analyzing the CS‐AA polyion complex structure using spectroscopic methods, the ex vivo moisture retention ability was assessed under dry conditions using porcine skin samples. Additionally, the efficacy of the CS‐AA polyion complex in reducing transepidermal water loss (TEWL) and improving skin hydration was evaluated on human subjects using a digital evaporimeter and a corneometer, respectively. Results: Validating a systematic reduction in particle size, the following order was observed: CS > CS/AA simple mixture > CS‐AA complex based on dynamic light scattering (DLS) and transmission electron microscopy (TEM) analysis. Furthermore, observations revealed that the CS‐AA complex exhibits negligible surface charge. Additionally, Fourier‐transform infrared spectroscopy (FT‐IR) analysis demonstrated a distinct peak shift in the complex, confirming the successful formation of the CS‐AA complex. Subsequently, the water‐holding effect through porcine skin was assessed, revealing a notable improvement in moisture retention (weight loss) for the CS‐Q complex: 40.6% (1 h), 20.5% (2 h), and 18.7% (4 h) compared to glycerin. Similarly, the CS‐R complex demonstrated enhancements of 50.2% (1 h), 37.5% (2 h), and 33% (4 h) compared to glycerin. Furthermore, TEWL improvement efficacy on human skin demonstrated approximately 25% improvement for both the CS‐Q complex and CS‐R complex, surpassing the modest 12.5% and 18% improvements witnessed with water and glycerin applications, respectively. Finally, employing a corneometer, hydration changes in the skin were monitored over 4 weeks. Although CS alone exhibited nominal alterations, the CS‐Q complex and CS‐R complex showed a significant increase in moisture levels after 4 weeks of application. Conclusion: In this study, polyion complexes were successfully formed between CS, a sulfated GAG, and AA. Comparisons with glycerin, a well‐known moisturizing agent, confirmed that the CS‐AA complex exhibits superior moisturizing effects in various aspects. These findings suggest that the CS‐AA complex is a more effective ingredient than CS or AA alone in terms of efficacy. [ABSTRACT FROM AUTHOR]

Background: Condoliase is an enzyme used as a treatment for lumbar disc herniation (LDH). This enzyme degrades chondroitin sulfate (CS) in the nucleus pulposus of the intervertebral disc (IVD). However, there are cases in which symptoms do not improve, despite condoliase administration. This study reports histological analysis of lumbar disc tissue of LDH patients who underwent surgery because condoliase had no therapeutic effect. Methods: Between March 2019 and August 2019, 12 LDH patients who underwent full endoscopic spine surgery (FESS) discectomy at the Dezawa Akira PED Clinic were the subjects of the study. There are two study groups: six cases underwent FESS after condoliase administration, while six underwent FESS without condoliase administration. The average duration from drug administration to surgery was 152 days. Herniated disc removed at surgery was evaluated by histological staining including immunohistochemistry by anti‐CS antibodies. Results: Multiple large clusters (40–120 μm in diameter) were observed in the nucleus pulposus of those who received condoliase, but no clusters were observed in those who did not. The lumbar disc tissues, including the nucleus pulposus of recipients, were stained with anti‐CS antibodies that recognize the CS unsaturated disaccharide, but non‐administration tissue was not stained. These findings suggest that the enzyme acted on the nucleus pulposus, even in cases where symptoms were not improved by condoliase administration. Furthermore, there was no histological difference between stained images of the extracellular matrix in those who did or did not receive condoliase, suggesting that condoliase acted specifically on CS in the nucleus pulposus. Conclusions: We demonstrated that CS in the nucleus pulposus was degraded in patients in whom condoliase did not have a therapeutic effect. Moreover, condoliase acts in human IVD without causing necrosis of chondrocytes and surrounding tissues. [ABSTRACT FROM AUTHOR]

BACKGROUND: In this study, a biocompatible nano‐carrying platform using chitosan (ChI) and chondroitin sulfate (ChS) was developed for the encapsulation of cobia liver oil (CBLO) to prevent its oxidation and improve its absorption. An ionic gelation method was applied to encapsulate CBLO with different weight ratios (from 1.0 to 1.5) to obtain ChS‐ChI nano‐capsules (ChS‐ChI@CBLO NCs). RESULTS: Morphological observations of the nano‐capsules revealed a spherical shape and diameter around 267–381 nm. The maximum loading capacity (LC) and encapsulation efficiency (EE) for ChS‐ChI@CBLO NCs estimated by thermogravimetric analysis (TGA) and derivative thermogravimetric (DTG) analysis were 25.7% and 56.2%, respectively. The structural stability of ChS‐ChI@CBLO NCs was confirmed through differential scanning calorimetry (DSC) and X‐ray diffraction (XRD) analysis; moreover DSC also further confirmed the oxidative stability of ChS‐ChI@CBLO NCs. Fourier‐transform infrared (FTIR) spectra confirmed the excellent stability of ChS‐ChI@CBLO NCs against high temperature and sunlight exposure. Biocompatibility analysis also verified the non‐toxicity of ChS‐ChI@CBLO NCs, further indicating safety and potential application in complex‐nutritional supplements. CONCLUSION: Nano‐degree of ChS‐ChI@CBLO NCs has a loading capacity and encapsulation efficiency of around 16.5 ~ 25.7% and 33.4 ~ 56.2%, respectively, for encapsulation of CBLO. Characterization results also indicate that ChS‐ChI@CBLO NCs display high oxidative stability against long‐term, hyperthermal, and sunlight exposure. Bioassay results confirm that the ChS‐ChI@CBLO NCs are safe and non‐toxic. This study demonstrates that nano‐capsules are also beneficial in preventing sensitive compounds from metamorphosis, and are non‐toxic. These materials are suitable for use in the food and pharmaceutical industries. © 2023 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

Cartilage injury is a common disease in daily life. Especially in aging populations, the incidence of osteoarthritis is increasing. However, due to the poor regeneration ability of cartilage, most cartilage injuries cannot be effectively repaired. Even cartilage tissue engineering still faces many problems such as complex composition and poor integration of scaffolds and host tissues. In this study, chondroitin sulfate, one of the main components of extracellular matrix (ECM), is chosen as the main natural component of the material, which can protect cartilage in a variety of ways. Moreover, the results show that the addition of chondroitin sulfate improves the mechanical properties of gelatin methacrylate (GelMA) hydrogel, making it able to effectively bear mechanical loads in vivo. Further, chondroitin sulfate is modified to obtain the oxidized chondroitin sulfate (OCS) containing aldehyde groups via sodium periodate. This special group improves the interface integration and adhesion ability of the hydrogel to host cartilage tissue through schiff base reactions. In summary, GelMA/OCS hydrogel is a promising candidate for cartilage regeneration with good biocompatibility, mechanical properties, tissue integration ability, and excellent cartilage repair ability. [ABSTRACT FROM AUTHOR]

Anisotropy is an important cue for neural organization during morphogenesis and healing, contributing to the mechanical and functional properties of neural tissues. The ability to replicate such anisotropy in vitro holds great promise for the development of effective regeneration strategies. In this work, interfacial polyelectrolyte complexation (IPC) is applied to fabricate microfibers from charged ECM components without any chemical modification. Using flow‐focusing microfluidics, collagen (Col) and glycosaminoglycans (GAGs), such as chondroitin sulfate (CS) or heparin (Hep), form Col/CS and Col/Hep interfacial complexes that coalesce as IPC microfibers. These fibers are flexible and absorb large amounts of water but remain stable under physiological conditions. At these conditions, the tensile strength of the assembled Col/GAG microfibers is similar to the strength of the neural tissue. The fibers are biocompatible and biofunctional; PC12 neural cells adhere and orient longitudinally to the fibers. Moreover, Col/CS microfibers promote the formation of neural processes. The results demonstrate that the microfluidic‐assisted IPC complexation enables the assembly of ECM mimics by synergetic integration of anisotropic, chemical, and mechanical cues that boost the development of neural cells. [ABSTRACT FROM AUTHOR]

Necrotizing enterocolitis (NEC) continues to be a devastating disease in preterm neonates and has a paucity of medical management options. Chondroitin sulfate (CS) is a naturally occurring glycosaminoglycan (GAG) in human breast milk (HM) and has been shown to reduce inflammation. We hypothesized that supplementation with CS in an experimental NEC model would alter microbial diversity, favorably alter the cytokine profile, and (like other sulfur compounds) improve outcomes in experimental NEC via the eNOS pathway. NEC was induced in 5‐day‐old pups. Six groups were studied (n = 9–15/group): (1) WT breastfed and (2) Formula fed controls, (3) WT NEC, (4) WT NEC + CS, (5) eNOS KO (knockout) NEC, and (6) eNOS KO NEC + CS. Pups were monitored for clinical sickness score and weights. On postnatal day 9, the pups were killed. Stool was collected from rectum and microbiome analysis was done with 16 s rRNA sequencing. Intestinal segments were examined histologically using a well‐established injury scoring system and segments were homogenized and analyzed for cytokine profile. Data were analyzed using GraphPad Prism with p < 0.05 considered significant. CS supplementation in formula improved experimental NEC outcomes when compared to NEC alone. CS supplementation resulted in similar improvement in NEC in both the WT and eNOS KO mice. CS supplementation did not result in microbial changes when compared to NEC alone. Our data suggest that although CS supplementation improved outcomes in NEC, this protection is not conferred via the eNOS pathway or alteration of microbial diversity. CS therapy in NEC does improve the intestinal cytokine profile and further experiments will explore the mechanistic role of CS in altering immune pathways in this disease. [ABSTRACT FROM AUTHOR]

The brain extracellular matrix (ECM) is a highly glycosylated environment and plays important roles in many processes including cell communication, growth factor binding, and scaffolding. The formation of structures such as perineuronal nets (PNNs) is critical in neuroprotection and neural plasticity, and the formation of molecular networks is dependent in part on glycans. The ECM is also implicated in the neuropathophysiology of disorders such as Alzheimer's disease (AD), Parkinson's disease (PD), and Schizophrenia (SZ). As such, it is of interest to understand both the proteomic and glycomic makeup of healthy and diseased brain ECM. Further, there is a growing need for site‐specific glycoproteomic information. Over the past decade, sample preparation, mass spectrometry, and bioinformatic methods have been developed and refined to provide comprehensive information about the glycoproteome. Core ECM molecules including versican, hyaluronan and proteoglycan link proteins, and tenascin are dysregulated in AD, PD, and SZ. Glycomic changes such as differential sialylation, sulfation, and branching are also associated with neurodegeneration. A more thorough understanding of the ECM and its proteomic, glycomic, and glycoproteomic changes in brain diseases may provide pathways to new therapeutic options. [ABSTRACT FROM AUTHOR]

Cartilage tissue engineering strategies seek to repair damaged tissue using approaches that include scaffolds containing components of the native extracellular matrix (ECM). Articular cartilage consists of glycosaminoglycans (GAGs) which are known to sequester growth factors. In order to more closely mimic the native ECM, this study evaluated the chondrogenic differentiation of mesenchymal stem cells (MSCs), a promising cell source for cartilage regeneration, on fibrous scaffolds that contained the GAG‐mimetic cellulose sulfate. The degree of sulfation was evaluated, examining partially sulfated cellulose (pSC) and fully sulfated cellulose (NaCS). Comparisons were made with scaffolds containing native GAGs (chondroitin sulfate A, chondroitin sulfate C and heparin). Transforming growth factor‐beta3 (TGF‐β3) sequestration, as measured by rate of association, was higher for sulfated cellulose‐containing scaffolds as compared to native GAGs. In addition, TGF‐β3 sequestration and retention over time was highest for NaCS‐containing scaffolds. Sulfated cellulose‐containing scaffolds loaded with TGF‐β3 showed enhanced chondrogenesis as indicated by a higher Collagen Type II:I ratio over native GAGs. NaCS‐containing scaffolds loaded with TGF‐β3 had the highest expression of chondrogenic markers and a reduction of hypertrophic markers in dynamic loading conditions, which more closely mimic in vivo conditions. Studies also demonstrated that TGF‐β3 mediated its effect through the Smad2/3 signaling pathway where the specificity of TGF‐β receptor (TGF‐ βRI)‐phosphorylated SMAD2/3 was verified with a receptor inhibitor. Therefore, studies demonstrate that scaffolds containing cellulose sulfate enhance TGF‐β3‐induced MSC chondrogenic differentiation and show promise for promoting cartilage tissue regeneration. [ABSTRACT FROM AUTHOR]

An injectable hybrid hydrogel is synthesized, comprising silk fibroin (SF) and chondroitin sulfate (CS) through di‐tyrosine formation bond of SF chains. CS and SF are reported with excellent biocompatibility as tissue engineering scaffolds. Nonetheless, the rapid degradation rate of pure CS scaffolds presents a challenge to effectively recreate articular cartilage. As CS is one of the cartilage extracellular matrix (ECM) components, it has the potential to enhance the biological activity of SF‐based hydrogel in terms of cartilage repair. Therefore, altering the CS concentrations (i.e., 0 wt%, 0.25 wt%, 0.5 wt%, 1 wt%, and 2 wt%), which are interpenetrated between SF β‐sheets and chains, can potentially adjust the physical, chemical, and mechanical features of these hybrid hydrogels. The formation of β‐sheets by 30 days of immersion in de‐ionized (DI) water can improve the compression strength of the SF/CS hybrid hydrogels in comparison with the same SF/CS hybrid hydrogels in the dried state. Biological investigation and observation depicts proper cell attachment, proliferation and cell viability for C28/I2 cells. Gene expression of sex‐determining region YBox 9 (SOX9), Collagen II α1, and Aggrecan (AGG) exhibits positive C3H10T1/2 growth and expression of cartilage‐specific genes in the 0.25 wt% and 0.5 wt% SF/CS hydrogels. [ABSTRACT FROM AUTHOR]

Biglycan (BGN), a small leucine‐rich repeat proteoglycan, is involved in a variety of pathological processes including malignant transformation, for which the upregulation of BGN was found related to cancer cell invasiveness. Because the functions of BGN are mediated by its chondroitin/dermatan sulfate (CS/DS) chains through the sulfates, the determination of CS/DS structure and sulfation pattern is of major importance. In this study, we have implemented an advanced glycomics method based on ion mobility separation (IMS) mass spectrometry (MS) and tandem MS (MS/MS) to characterize the CS disaccharide domains in BGN. The high separation efficiency and sensitivity of this technique allowed the discrimination of five distinct CS disaccharide motifs, of which four irregulated in their sulfation pattern. For the first time, trisulfated unsaturated and bisulfated saturated disaccharides were found in BGN, the latter species documenting the non‐reducing end of the chains. The structural investigation by IMS MS/MS disclosed that in one or both of the CS/DS chains, the non‐reducing end is 3‐O‐sulfated GlcA in a rather rare bisulfated motif having the structure 3‐O‐sulfated GlcA‐4‐O‐sulfated GalNAc. Considering the role played by BGN in cancer cell spreading, the influence on this process of the newly identified sequences will be investigated in the future. [ABSTRACT FROM AUTHOR]

The development of electro‐stimulated drug release devices is an innovative approach to attain the drug delivery in accurate doses at target sites in a programmed manner. In this work, novel electroactive nanocomposite hydrogels were prepared by encapsulating green‐synthesized polypyrrole (PPy) colloids within chondroitin sulfate (CS) networks during the self‐crosslinking of CS via N‐ethyl‐N′‐(3‐dimethylaminopropyl) carbodiimide chemistry. The structural and morphological properties of CS/PPy hydrogels were studied by Fourier‐transformed infrared spectroscopy, scanning electron microscopy, and swelling kinetic measurements. The chemotherapeutic agent 5‐fluorouracil (5‐FU) was loaded into CS/PPy samples by hydrogel swelling method, or alternatively, by pre‐incubating the drug in polymer mixture before crosslinking. Different electrical stimulations can be used to switch ON and accurately tune the 5‐FU delivery from GG/PPy hydrogels. A single pulse potential of 5 V switched on the drug delivery up to 90% from nanocomposite hydrogel, in contrast to the low 5‐FU amount released in a passive form (< 20%). PPy electroactive behavior played a determining role as the main driving force in 5‐FU release activation. Cytotoxicity of hydrogels with and without 5‐FU was examined in normal and cancer cells. Considering the high cytotoxicity of 5‐FU, the ON/OFF 5‐FU release patterns evidenced the potential of CS/PPy hydrogels for electrically controlled drug delivery in implantable or transdermal drug release devices. [ABSTRACT FROM AUTHOR]

Chondroitin sulfate/dermatan sulfate (CS/DS) is a member of glycosaminoglycans (GAGs) found in animal tissues. Major CS/DS subclasses, O, A, C, D, and E units, exist based on the sulfation pattern in d‐glucuronic acid (GlcA) and N‐acetyl‐d‐galactosamine repeating units. DS is formed when GlcA is epimerized into l‐iduronic acid. Our study aimed to analyze the CS/DS profile in 3T3‐L1 cells before and after adipogenic induction. CS/DS contents, molecular weight (Mw), and sulfation pattern were analyzed by using high‐performance liquid chromatography. CS/DS synthesis‐ and sulfotransferase‐related genes were analyzed by reverse transcription real‐time PCR. CS/DS amount was significantly decreased in the differentiated (DI) group compared to the non‐differentiated (ND) group, along with a lower expression of CS biosynthesis‐related genes, chondroitin sulfate N‐acetylgalactosaminyltransferase 1 and 2, as well as chondroitin polymerizing factor. GAGs in the DI group also showed lower Mw than those of ND. Furthermore, the A unit was the major CS/DS in both groups, with a proportionally higher CS‐A in the DI group. This was consistent with the expression of carbohydrate sulfotransferase 12 that encodes chondroitin 4‐O‐sulfotransferase, for CS‐A formation. These qualitative and quantitative changes in CS/DS and CS/DS‐synthases before and after adipocyte differentiation reveal valuable insights into adipocyte development. [ABSTRACT FROM AUTHOR]

Sensory nerves are long being recognized as collecting units of various outer stimuli; recent advances indicate that the sensory nerve also plays pivotal roles in maintaining organ homeostasis. Here, this study shows that sensory nerve orchestrates intervertebral disc (IVD) homeostasis by regulating its extracellular matrix (ECM) metabolism. Specifically, genetical sensory denervation of IVD results in loss of IVD water preserve molecule chondroitin sulfate (CS), the reduction of CS bio‐synthesis gene chondroitin sulfate synthase 1 (CHSY1) expression, and dysregulated ECM homeostasis of IVD. Particularly, knockdown of sensory neuros calcitonin gene‐related peptide (CGRP) expression induces similar ECM metabolic disorder compared to sensory nerve denervation model, and this effect is abolished in CHSY1 knockout mice. Furthermore, in vitro evidence shows that CGRP regulates nucleus pulposus cell CHSY1 expression and CS synthesis via CGRP receptor component receptor activity‐modifying protein 1 (RAMP1) and cyclic AMP response element‐binding protein (CREB) signaling. Therapeutically, local injection of forskolin significantly attenuates IVD degeneration progression in mouse annulus fibrosus puncture model. Overall, these results indicate that sensory nerve maintains IVD ECM homeostasis via CGRP/CHSY1 axis and promotes IVD repair, and this expands the understanding concerning how IVD links to sensory nerve system, thus shedding light on future development of novel therapeutical strategy to IVD degeneration. [ABSTRACT FROM AUTHOR]

Background: Condoliase has been used in Japan to treat patients with lumbar disc herniation by its injection into the nucleus pulposus. The injection of condoliase together with contrast media is prohibited; because there are no data whether contrast media have any effect on condoliase activity. This study aimed to elucidate the effects of contrast media on condoliase activity. Methods: Condoliase with chondroitin sulfate (CS) and without CS were mixed with various contrast media (nonionic [iohexol or iotrolan]; ionic [amidotrizoic acid]). (i) The mixtures with CS were incubated at 37°C; (ii) the mixtures without CS were stored at 24°C for 60 min, followed by addition of CS to assess condoliase activity by measuring the amount of N‐acetylhexosamines enzymatically cleaved from CS using Morgan–Elson method. Results: (i) In the presence of CS, the ionic contrast media reduced condoliase activity within 10 min in a dose‐dependent manner, and the nonionic contrast media had no effect on condoliase activity for at least 120 min. (ii) In the absence of CS, the ionic contrast media almost completely inactivated condoliase within 15 min, and the nonionic contrast media also reduced condoliase activity; the residual activity was 65% with iotrolan and 35% with iohexol at 60 min. Conclusions: The ionic contrast media significantly reduced condoliase activity regardless of presence or absence of CS. Although the nonionic contrast media did not affect condoliase activity in the presence of CS, it reduced activity in the absence of CS. Mixing condoliase with contrast media, especially ionic type contrast media, should be avoided. [ABSTRACT FROM AUTHOR]

The dynamic properties of the polymeric components of cartilage are important for understanding the many biological functions of this biological tissue. Knowledge of the rheology of these materials is also crucial for the development of therapeutic treatments for diseases, such as arthritis, in which cartilage loses its functional physiological properties. In the present work complementary noninvasive techniques, rheology and dynamic light scattering (DLS) are used to probe the structural and dynamic properties of solutions of the principal proteoglycan components of cartilage extracellular matrix at different levels of structural hierarchy (chondroitin sulfate, aggrecan, aggrecan‐hyaluronic acid complex). It is shown that the bottlebrush shaped aggrecan molecules form large, loosely organized polyelectrolyte domains comprised of molecules organized into diffuse branched polymer network structures permeated by a diffuse cloud of counter‐ions. The rheological behaviors of these polymer solutions vary from viscoelastic solutions, consistent with the formation of dynamic polymer clusters in solution, to "weak gel" materials exhibiting approximate power law shear stress relaxation or creep over a wide range of timescales. [ABSTRACT FROM AUTHOR]

Background: Atherosclerosis is triggered by the retention of apolipoprotein B-containing lipoproteins by proteoglycans. In addition to low-density lipoprotein, remnant lipoproteins have emerged as pivotal contributors to this pathology, particularly in the context of insulin resistance and diabetes. We have previously reported antiatherogenic properties of a monoclonal antibody (chP3R99) that recognizes sulfated glycosaminoglycans on arterial proteoglycans., Methods and Results: Solid-phase assays demonstrated that chP3R99 effectively blocked >50% lipoprotein binding to chondroitin sulfate and vascular extracellular matrix in vitro. The preperfusion of chP3R99 (competitive effect) resulted in specific antibody-arterial accumulation and reduced fluorescent lipoprotein retention by ~60% in insulin resistant JCR:LA- cp rats. This competitive reduction was dose dependent (25-250 μg/mL), effectively decreasing deposition of cholesterol associated with lipoproteins. In a 5-week vaccination study in insulin resistant rats with (200 μg subcutaneously, once a week), chP3R99 reduced arterial lipoprotein retention, and was associated with the production of antichondroitin sulfate antibodies (Ab3) able to accumulate in the arteries (dot-blot). Neither the intravenous inoculation of chP3R99 (4.5 mg/kg), nor the immunization with this antibody displayed adverse effects on lipid or glucose metabolism, insulin resistance, liver function, blood cell indices, or inflammation pathways in JCR:LA -cp rats., Conclusions: Both acute (passive) and long-term administration (idiotypic cascade) of chP3R99 antibody reduced low-density lipoprotein and remnant lipoprotein interaction with proteoglycans in an insulin-resistant setting. These findings support the innovative approach of targeting proatherogenic lipoprotein retention by chP3R99 as a passive therapy or as an idiotypic vaccine for atherosclerosis.

The compound 3′‐phosphoadenosine‐5′‐phosphosulfate (PAPS) serves as a sulfate group donor in the production of valuable sulfated compounds. However, elevated costs and low conversion efficiency limit the industrial applicability of PAPS. Here, we designed and constructed an efficient and controllable catalytic system for the conversion of adenosine triphosphate (ATP) (disodium salt) into PAPS without inhibition from by‐products. In vitro and in vivo testing in Escherichia coli identified adenosine‐5′‐phosphosulfate kinase from Penicillium chrysogenum (PcAPSK) as the rate‐limiting enzyme. Based on analysis of the catalytic steps and molecular dynamics simulations, a mechanism‐guided "ADP expulsion" strategy was developed to generate an improved PcAPSK variant (L7), with a specific activity of 48.94 U·mg−1 and 73.27‐fold higher catalytic efficiency (kcat/Km) that of the wild‐type enzyme. The improvement was attained chiefly by reducing the ADP‐binding affinity of PcAPSK, as well as by changing the enzyme's flexibility and lid structure to a more open conformation. By introducing PcAPSK L7 in an in vivo catalytic system, 73.59 mM (37.32 g·L−1) PAPS was produced from 150 mM ATP in 18.5 h using a 3‐L bioreactor, and achieved titer is the highest reported to date and corresponds to a 98.13% conversion rate. Then, the PAPS catalytic system was combined with the chondroitin 4‐sulfotransferase using a one‐pot method. Finally, chondroitin sulfate was transformed from chondroitin at a conversion rate of 98.75%. This strategy has great potential for scale biosynthesis of PAPS and chondroitin sulfate. [ABSTRACT FROM AUTHOR]

Heparan sulfate (HS), a sulfated glycosaminoglycan (GAG), was reported to be a necessary host attachment factor that promotes SARS‐CoV‐2 infection. In this study, we developed GAG microarrays based on fluorescence detection for high‐sensitivity screening of the GAG‐binding specificity of proteins and applied it for the analysis of SARS‐CoV‐2 spike (S) protein. Among the 20 distinct GAGs, the S protein bound not only to heparin (HEP)/HS but also to chondroitin sulfate E (CSE) in a concentration‐dependent manner. We then analyzed the specificity of each subunit of the S protein. While the S1 subunit showed exclusive binding to HEP, the S2 subunit also bound to CSE and HEP/HS. CSE might act as an alternative attachment factor for HS in SARS‐CoV‐2 infection. [ABSTRACT FROM AUTHOR]

Midkine (MK) is a neurotrophic factor that participates in the embryonic central nervous system (CNS) development and neural stem cell regulation, interacting with sulfated glycosaminoglycans (GAGs). Chondroitin sulfate (CS) is the natural ligand in the CNS. In this work, we describe the interactions between a library of synthetic models of CS‐types and mimics. We did a structural study of this library by NMR and MD (Molecular Dynamics), concluding that the basic shape is controlled by similar geometry of the glycosidic linkages. Their 3D structures are a helix with four residues per turn, almost linear. We have studied the tetrasaccharide‐midkine complexes by ligand observed NMR techniques and concluded that the shape of the ligands does not change upon binding. The ligand orientation into the complex is very variable. It is placed inside the central cavity of MK formed by the two structured beta‐sheets domains linked by an intrinsically disordered region (IDR). Docking analysis confirmed the participation of aromatics residues from MK completed with electrostatic interactions. Finally, we test the biological activity by increasing the MK expression using CS tetrasaccharides and their capacity in enhancing the growth stimulation effect of MK in NIH3T3 cells. [ABSTRACT FROM AUTHOR]

Phospholipid Phosphatase‐Related Protein Type 1 (PLPPR1) is a member of a family of lipid phosphatase related proteins, integral membrane proteins characterized by six transmembrane domains. This family of proteins is enriched in the brain and recent data indicate potential pleiotropic functions in several different contexts. An inherent ability of this family of proteins is to induce morphological changes, and we have previously reported that members of this family interact with each other and may function co‐operatively. However, the function of PLPPR1 is not yet understood. Here we show that the expression of PLPPR1 reduces the inhibition of neurite outgrowth of cultured mouse hippocampal neurons by chondroitin sulfate proteoglycans and the retraction of neurites of Neuro‐2a cells by lysophosphatidic acid (LPA). Further, we show that PLPPR1 reduces the activation of Ras homolog family member A (RhoA) by LPA in Neuro‐2a cells, and that this is because of an association of PLPPR1with the Rho‐specific guanine nucleotide dissociation inhibitor (RhoGDI1). These results establish a novel signaling pathway for the PLPPR1 protein. [ABSTRACT FROM AUTHOR]

Promising cancer treatment requires the assistant of drug delivery systems (DDS) with the aim to increase the accumulation of drugs in tumor tissue. Herein, a hybrid DDS was successfully developed to integrate chondroitin sulfate (CS) and calcium carbonate (CC) in to one system. Anticancer drug adriamycin (Adr) was preloaded into CC nanoparticles to obtain Adr‐loaded CC nanoparticles (CC/Adr). The resulted CS‐CC/Adr nanoparticles as a biocompatible DDS was able to specifically target cancer cells to enhance the chemotherapy of lung cancer due to the surface modification of CS. Intracellular uptake as well as in vivo imaging results revealed the obtained CS‐CC/Adr nanoparticles (size of ~100 nm) showed CS mediated tumor specific accumulation into A549 and LLC cells than unmodified CC/Adr, in which the CD44 receptor might be involved, which finally resulted in stronger anticancer capability than Adr or CC/Adr. As a result, CS‐CC/Adr nanoparticles could be further extended to clinical administration in our future works. [ABSTRACT FROM AUTHOR]

Loss of chondroitin sulfate (CS) has been reported to play a key role during intervertebral disc degeneration (IDD). However, the detailed mechanism of CS and its synthases have not been elucidated. Since CS is mainly synthesized by chondroitin synthases 3 (Chsy3), here, the Chsy3 knockout mice are generated by using CRISPR‐Cas9 and semi‐cloning technology to study its mechanism during IDD. We find that CS and Chsy3 expression are decreased during IDD both in human and mice nucleus pulposus (NP) tissue, and knockout of Chsy3 shows that spontaneous IDD phenotype resembles that of human samples in the Chsy3−/− mice. Taking advantage of RNA‐Seq data, we confirm increased catabolic and decreased anabolic changes in Chsy3−/− NP cells. By using bioinformatic analysis and validation, we find that Hippo signaling pathway is significantly downregulated, and the activation of Yap1 is mainly affected in Chsy3−/− NP cells. Furthermore, functional analyses have shown that Chsy3 could regulate NP cell degeneration by Actin tension mediated activation of Yap1, which is independent of Hippo/Lats signaling. In summary, our findings reveal a novel mechanism that depletion of CS‐related Chsy3 can cause spontaneous intervertebral disc degeneration by mediating Yap activation through CS‐related actin‐tension in NP cells. [ABSTRACT FROM AUTHOR]

Tissue engineering is a technique that can be used for repairing damaged cartilage. The aim of this study was chondro‐differentiation of mesenchymal stem cells (MSCs) on polyvinyl alcohol/gelatin/chondroitin sulfate (PVA/GT/CS) blend nanofibers. Nanofibers were fabricated by electrospinning method with different concentrations of CS (10%, 15%, and 20%). MSCs were seeded on nanofibers for biocompatibility and cell viability test by MTT assay. Alcian blue staining assay was done to show that MSCs were differentiated to chondrocyte by staining the glycosaminoglycans (GAGs) that was produced by cells. Reverse transcription polymerase chain reaction (RT‐PCR) and immunocytochemistry (ICC) assay were done to confirm type II collagen expression. Scaffolds with different concentrations of CS were tested for biocompatibility, and the nanofiber with 15% of GAG showed better results (P = 0.02) compared with other nanofibers; therefore, it was chosen as the main sample for chondrogenesis tests. On the basis of scanning electron microscopy (SEM), hMSCs had a proper attachment to the nanofiber with 15% CS. Alcian blue staining showed the different rates of blue coloration, which means cells produced different types of GAGs from the GAG we used in the nanofiber. RT‐PCR and ICC showed that COL2A1 was expressed in mRNA and protein levels. This study indicates that the PVA/GT/CS nanofiber containing 15% CS can be a good candidate for cartilage tissue engineering. [ABSTRACT FROM AUTHOR]

Pain originating from an intervertebral disc (discogenic pain) is a major source of chronic low back pain. Pathological innervation of the disc by pain‐sensing nerve fibers is thought to be a key component of discogenic pain, so treatment with biomaterials that have the ability to inhibit neurite growth will greatly benefit novel disc therapeutics. Currently, disc therapeutic biomaterials are rarely screened for their ability to modulate nerve growth, mainly due to a lack of models to screen neuromodulation. To address this deficit, our lab has engineered a three dimensional in vitro disc innervation model that mimics the interface between primary sensory nerves and the intervertebral disc. Further, herein we have demonstrated the utility of this model to screen the efficacy of chondroitin sulfate biomaterials to inhibit nerve fiber invasion into the model disc. Biomaterials containing chondroitin‐4‐sulfate (CS‐A) decrease neurite growth in a uniform gel and at an interface between a growth‐permissive and a growth‐inhibitory gel, while chondroitin‐6‐sulfate (CS‐C) is less neuroinhibitory. This in vitro model holds great potential for screening inhibitors of nerve fiber growth to further improve intervertebral disc replacements and therapeutics. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 38:1016‐1026, 2020 [ABSTRACT FROM AUTHOR]

Glycosaminoglycan (GAG) chemical exchange saturation transfer (gagCEST) is a potential method for cartilage quality assessment. The aim of this study was to investigate how the gagCEST effect depends on the types and molecular organization of GAG typically found in articular cartilage. gagCEST was performed on different concentrations of GAG in various forms: free chains of chondroitin sulfate (CS) of different types (‐A and ‐C) and GAG bound to protein in aggregated and nonaggregated aggrecan extracted from calf articular cartilage. The measured magnetization transfer ratio asymmetry (MTRasym) was compared with known GAG concentrations or GAG concentrations determined through biochemical analysis. The gagCEST effect was assessed through the linear regression coefficient with 95% confidence interval of MTRasym per GAG concentration. We observed a lower gagCEST effect in phantoms containing a mixture of CS‐A and CS‐C compared with phantoms containing mainly CS‐A. The difference in response corresponds well to the difference in CS‐A concentration. GAG bound in aggrecan from calf articular cartilage, where CS‐A is assumed to be the major type of GAG, produed a similar gagCEST effect as that observed for free CS‐A. The effect was also similar for aggregated (ie, bound to hyaluronic acid) and nonaggregated aggrecan. In conclusion, our results indicate that the aggrecan structure in itself does not impact the gagCEST effect, but that the effect is strongly dependent on GAG type. In phantoms, the current implementation of gagCEST is sensitive to CS‐A while for CS‐C, the main GAG component in mature human articular cartilage, the sensitivity is limited. This difference in gagCEST sensitivity between GAG types detected in phantoms is a strong motivation to also explore the possibility of a similar effect in vivo. [ABSTRACT FROM AUTHOR]

BACKGROUND: The production and consumption of chicken has doubled in the last 17 years. Since slaughter yields range from 70 to 80%, during production various chicken waste residues are generated. One of these residues is waste created after the mechanical separation of chicken meat (W‐MSM). The goal of the circular economy is to manage this waste and transform it into a valuable product. RESULTS: W‐MSM was subjected to hydrolysis by the proteolytic enzymes pepsin and papain to obtain bioactive compounds. Papain digestion resulted in several times higher yields of chondroitin sulfate (CS), hyaluronic acid (HA), peptides and amino acids in comparison with pepsin. The hydrolysis conditions were further optimized using the software Design Expert to evaluate the impact of added enzyme, buffer and duration of the hydrolysis. In addition, the scale‐up of this process was verified with 5 kg of W‐MSM and the obtained product contained approximately 77% (w/w) of peptides, 9% (w/w) of CS, 7% (w/w) of HA and 4% (w/w) of amino acids. CONCLUSIONS: We tested and optimized the method for evaluating the direct waste from poultry processing and transformed it into a product containing bioactive compounds which can be utilized as an ingredient for improving the properties of feed. © 2019 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

Extracellular levels of soluble TIMP‐3 are low, reflecting its binding by extracellular matrix (ECM) components including sulfated glycosaminoglycans (SGAGs) and endocytosis via low density lipoprotein receptor‐related protein 1. Since TIMP‐3 inhibits ECM degradation, the ability of SGAGs to elevate extracellular TIMP‐3 is significant for osteoarthritis treatment. Previous studies of such interactions have utilized immobilized TIMP‐3 or ligands. Here, we report the thermodynamics of the interactions of the sGAG‐binding N‐domain of TIMP‐3 with chondroitin sulfate, pentosan polysulfate, and suramin in solution using isothermal titration calorimetry. All three interactions are driven by a favorable negative enthalpy change combined with an unfavorable decrease in entropy. The heat capacity changes (ΔCp) for all of the interactions are zero, indicating an insignificant contribution from hydrophobic interactions. [ABSTRACT FROM AUTHOR]

The long noncoding RNA HOTAIR plays significant roles in promoting cancer metastasis. However, how it conveys an invasive advantage in cancer cells is not clear. Here we identify the chondroitin sulfotransferase CHST15 (GalNAc4S‐6ST) as a novel HOX transcript antisense intergenic RNA (HOTAIR) target gene using RNA profiling and show that CHST15 is required for HOTAIR‐mediated invasiveness in breast cancer cells. CHST15 catalyzes sulfation of the C6 hydroxyl group of the N‐acetyl galactosamine 4‐sulfate moiety in chondroitin sulfate to form the 4,6‐disulfated chondroitin sulfate variant known as the CS‐E isoform. We show that HOTAIR is necessary and sufficient for CHST15 transcript expression. Inhibition of CHST15 by RNA interference abolished cell invasion promoted by HOTAIR but not on HOTAIR‐mediated migratory activity. Conversely, reconstitution of CHST15 expression rescued the invasive activity of HOTAIR‐depleted cells. In corroboration with this mechanism, blocking cell surface chondroitin sulfate using a pan‐CS antibody or an antibody specifically recognizes the CS‐E isoform significantly suppressed HOTAIR‐induced invasion. Inhibition of CHST15 compromised tumorigenesis and metastasis in orthotopic breast cancer xenograft models. Furthermore, the expression of HOTAIR closely correlated with the level of CHST15 protein in primary as well as metastatic tumor lesions. Our results demonstrate a novel mechanism underlying the function of HOTAIR in tumor progression through programming the context of cell surface glycosaminoglycans. Our results further establish that the invasive and migratory activities downstream of HOTAIR are distinctly regulated, whereby CHST15 preferentially controls the arm of invasiveness. Thus, the HOTAIR‐CHST15 axis may provide a new avenue toward novel therapeutic strategies and prognosis biomarkers for advanced breast cancer. What's new? The long noncoding RNA HOTAIR is known to influence cancer growth, and now researchers have identified a molecule that mediates the effect. Using both loss‐of‐function and gain‐of‐function experiments, these authors identified chondroitin sulfotransferase (CHST15) as a downstream target of HOTAIR required for invasive activity of breast cancer cells. Antibodies against cell surface chondroitin sulfate suppressed HOTAIR‐induced invasion in xenograft models. Further, HOTAIR expression correlates with CHST15 protein both in primary tumors and metastases. The identification of HOTAIR‐CHST15 regulation of cell surface moieties could lead to new therapeutic targets or diagnostic biomarkers for breast cancer. [ABSTRACT FROM AUTHOR]

Capillary electrophoresis with large‐volume sample stacking using an electroosmotic flow pump was developed for the determination of chondroitin sulfate, dermatan sulfate, and hyaluronic acid. Central composite design was used to simultaneously optimize the parameters for capillary electrophoresis separation. The optimized capillary electrophoresis conditions were 200 mM sodium dihydrogen phosphate, 200 mM butylamine, and 0.5% w/v polyethylene glycol as a background electrolyte, pH 4 and ‐16 kV. Exploiting large‐volume sample stacking using an electroosmotic flow pump, the sensitivity of the proposed capillary electrophoresis system coupled with UV detection was significantly improved with limits of detection of 3, 5, 1 mg/L for chondroitin sulfate, dermatan sulfate, and hyaluronic acid, respectively. The developed method was applied to the determination of chondroitin sulfate and hyaluronic acid in cell culture media, cerebrospinal fluid, cosmetic products, and supplementary samples with highly acceptable accuracy and precision. Therefore, the proposed capillary electrophoresis approach was found to be simple, rapid, and reliable for the determination of chondroitin sulfate, dermatan sulfate, and hyaluronic acid in cell culture media, cerebrospinal fluid, cosmetic, and supplementary samples without sample pretreatment. [ABSTRACT FROM AUTHOR]

Chondroitin sulfate E (CS‐E) is a glycosaminoglycan containing type‐E disaccharide units (sulfated at C‐4 and C‐6 of N‐acetylgalactosamine). CS‐E is covalently linked to a core protein to form chondroitin sulfate proteoglycans (PGs) that are secreted or associated with the plasma membrane of several types of cells. CS‐E‐containing PGs selectively interact with growth factors and chemokines and control various cellular and/or tissue processes. Angiogenesis is a process that is highly regulated in physiological conditions but deregulated in pathologies, leading to excess or deficient blood vessel formation. Angiogenesis regulation is orchestrated by numerous growth factors, such as vascular endothelial growth factor A, fibroblast growth factors and pleiotrophin, whose functions can be affected by CS‐containing PGs. In the present review, we focus on the emerging area of CS‐mediated angiogenesis and particularly on the critical assessment of data related to a potential role of CS‐E in controlling endothelial cell functions, focusing on angiogenesis regulation and vascular homeostasis in health and disease. [ABSTRACT FROM AUTHOR]

The four syndecans identified in mammals are membrane proteoglycans that play major roles in regulating cell behavior, cell signaling, and cell‐matrix interactions. The membrane forms of these syndecans function as receptors and co‐receptors. Their ectodomains, which are proteolytically released in the extracellular matrix by shedding, also regulate various biological processes. Apart from the cytoplasmic domain of syndecan‐4, the 3D structures of syndecans are poorly characterized, which hinders our understanding of the molecular mechanisms underlying syndecan functions that are mediated by numerous interactions. This mini‐review summarizes the structural data that are available for syndecans and provides a comprehensive syndecan interactome, which comprises three hundred and fifty‐one partners, including those identified by the high‐throughput method affinity purification‐mass spectrometry. It also gives a perspective on future studies of syndecan structures and interactions, which are required to further elucidate the molecular recognition processes that mediate the biological roles of the membrane and shed forms of syndecans. [ABSTRACT FROM AUTHOR]

The pharmacokinetic profile of a new 800‐mg tablet of nonanimal chondroitin sulfate (CS) (Mythocondro®, 800‐mg tablets, Gnosis S.p.A., Italy) was investigated vs an animal CS in healthy volunteers for a total period of 48 hours. After a single 2400‐mg dose of the test and the reference formulation, total CS, the compositional disaccharides (ΔDi6S, ΔDi4S and ΔDi0S), and the overall charge density were quantified in plasma. The safety and tolerability profile after a single dose of this new nonanimal CS tablets was excellent. After baseline‐corrected concentrations, an overall greater plasma concentration was observed after 24 hours of ∼44% and after 48 hours of ∼45% from administration of nonanimal when compared to animal‐derived CS. Moreover, nonanimal CS increases the specific sulfation in the 6‐position of N‐acetyl‐galactosamine in human plasma CS and, as a consequence, the overall charge density, reaching double values (0.91), after 48 hours compared to bovine CS and to endogenous CS. In conclusion, nonanimal CS, possessing a lower molecular weight than an animal‐derived sample, produces a greater CS concentration for a more prolonged period of time in plasma and an increase in charge density and specific 6‐sulfation of endogenous plasma CS. [ABSTRACT FROM AUTHOR]

Aims: To present a rationale for the inclusion of urothelial coating dysfunction in the etipathogenesis of bladder pain syndrome/interstitial cystitis (BPS/IC) and the preclinical and clinical evidence in support of glycosaminoglycan (GAG) replenishment therapy in the treatment of BPS/IC, supplemented by the clinical experience of medical experts in the field and patient advocates attending a symposium on GAG replenishment at ESSIC'17, the annual Meeting of the International Society for the Study of Bladder Pain Syndrome, held in Budapest, Hungary in 2017. Results: The urothelial GAG layer has a primary role in providing a permeability barrier to prevent penetration of urinary toxins and pathogens into the bladder wall. Disruption of the GAG layer contributes to the development of BPS/IC. The evidence shows that replenishment of GAGs can restore the GAG layer in BPS/IC, reducing inflammation, pain, and other symptoms. Conclusions: Although data from large randomized controlled studies are limited, long clinical observation and the experience of clinicians and patients support the beneficial effects of intravesical GAG replenishment therapy for providing symptomatic relief for patients with BPS/IC. [ABSTRACT FROM AUTHOR]

Glycosaminoglycans (GAGs) are linear, highly negatively charged carbohydrate chains present in connective tissues. Chondroitin sulfate (CS) and heparin (Hep) are also found in the numerous secretory granules of mast cells (MC), tissue immune cells involved in allergic and inflammatory reactions. CS and Hep may inhibit secretion of histamine from rat connective tissue MC, but their effect on human MC remains unknown. Human LAD2 MC were pre‐incubated with CS, Hep, or dermatan sulfate (DS) before being stimulated by either the peptide substance P (SP, 2 μM) or the cytokine IL‐33 (10 ng/mL). Preincubation with CS had no effect on MC degranulation stimulated by SP, but inhibited TNF (60%) and CXCL8 (45%) secretion from LAD2 cells stimulated by IL‐33. Fluorescein‐conjugated CS (CS‐F) was internalized by LAD2 cells only at 37 °C, but not 4 °C, indicating it occurred by endocytosis. DS and Hep inhibited IL‐33‐stimulated secretion of TNF and CXCL8 to a similar extent as CS. None of the GAGs tested inhibited IL‐33‐stimulated gene expression of either TNF or CXCL8. There was no effect of CS on ionomycin‐stimulated calcium influx. There was also no effect of CS on surface expression of the IL‐33 receptor, ST2. Neutralization of the hyaluronan receptor CD44 did not affect the internalization of CS‐F. The findings in this article show that CS inhibits secretion of TNF and CXCL8 from human cultured MC stimulated by IL‐33. CS could be formulated for systemic or topical treatment of allergic or inflammatory diseases, such as atopic dermatitis, cutaneous mastocytosis, and psoriasis. © 2018 BioFactors, 45(1):49–61, 2019 [ABSTRACT FROM AUTHOR]

Proteoglycans are diverse, complex extracellular/cell surface macromolecules composed of a central core protein with covalently linked glycosaminoglycan (GAG) chains; both of these components contribute to the growing list of important bio‐active functions attributed to proteoglycans. Increasingly, attention has been paid to the roles of proteoglycans in nervous tissue development due to their highly regulated spatio/temporal expression patterns, whereby they promote/inhibit neurite outgrowth, participate in specification and maturation of various precursor cell types, and regulate cell behaviors like migration, axonal pathfinding, synaptogenesis and plasticity. These functions emanate from both the environments proteoglycans create around cells by retaining ions and water or serving as scaffolds for cell shaping or motility, and from dynamic interactions that modulate signaling fields for cytokines, growth factors and morphogens, which may bind to either the protein or GAG portions. Also, genetic abnormalities impacting proteoglycan synthesis during critical steps of brain development and response to environmental insults and injuries, as well as changes in microenvironment interactions leading to tumors in the central nervous system, all suggest roles for proteoglycans in behavioral and intellectual disorders and malignancies. [ABSTRACT FROM AUTHOR]

Abstract: Chondroitin sulfate (Ch) is one of the main structural components of cartilage tissue, therefore, its presence in tissue engineered scaffold is expected to enhance cartilage regeneration. Previously, silk fibroin/chitosan (SF/CS) blend was proven to be a potential biomaterial for tissue development. In this study, the effect of Ch on physicochemical and biological properties of SF/CS blend was investigated and scaffolds with 0.8 wt% Ch was found to be favorable. The scaffolds possess pore size of 37–212 µm, contact angle 46.2–50.3°, showed controlled swelling and biodegradation. The biocompatibility of scaffold was confirmed by subcutaneous implantation in mouse. Human mesenchymal stem cells (hMSCs) seeded scaffolds cultured under spinner flask bioreactor promoted cell attachment, proliferation, distribution, and metabolic activity in vitro. The histology and immunofluorescence studies revealed that combined effect of Ch and dynamic condition resulted in higher glycosaminoglycan secretion and native cartilage type matrix synthesis in comparison to SF/CS scaffolds used as control. Higher expression of collagen‐II, Sox9, aggrecan and decrease in collagen‐I expression represented by quantitative polymerase chain reaction study confirmed the progression of chondrogenic differentiation. This study successfully demonstrates the potentiality of SF/CS‐Ch scaffold for hMSCs recruitment and redirecting cartilage tissue regeneration with enhanced chondrogenesis. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 2576–2587, 2018. [ABSTRACT FROM AUTHOR]

Polyelectrolyte microcapsules are modular constructs which facilitate cell handling and assembly of cell‐based tissue constructs. In this study, an electrospray (ES) encapsulation apparatus was developed for the encapsulation of mesenchymal stem cells (MSCs). Ionic complexation between glycosaminoglycans (GAGs) and chitosan formed a polyelectrolyte complex membrane at the interface. To optimize the capsules, the effect of voltage, needle size and GAG formulation on capsule size were investigated. It was observed that by increasing the voltage and decreasing the needle size, the capsule size would decrease but at voltages above 12 kV, capsule size distribution broadened significantly which yields lower circularity. Increase in GAG viscosity resulted in larger microcapsules and cell viability exhibited no significant changes during the encapsulation procedure. These results suggest that ES is a highly efficient, and scalable approach to the encapsulation of MSCs for subsequent use in bioprinting and other modular tissue engineering or regenerative medicine applications. [ABSTRACT FROM AUTHOR]

Abstract: In this study, highly neutralized, highly porous, and ultralight polymeric aerogels prepared from aqueous colloidal suspensions of chitosan (CS) and chondroitin sulfate (ChS) nanocomplexes, formulated as quasi‐equimolar amounts of both, are described. These aerogels were designed as healing agents under the inspiration of minimizing the amount of matter applied to wounds, reducing the electrostatic potential of the material and avoiding covalent cross‐linkers in order to decrease metabolic stress over wounds. Aerogels synthesized under these criteria are biocompatible and provide specific properties for the induction of wound healing. They do not affect neither the metabolic activity of cultured 3T3 fibroblasts nor the biochemical parameters of experimental animals, open wounds close significantly faster and, unlike control wounds, complete reepithelialization and scarring can be attained 14 days after surgery. Because of its hydration abilities, rapid adaptation to the wound bed and the early accelerator effect of wound closure, the CS/ChS aerogels appear to be functional inducers of the healing. Previous information show that CS/ChS aerogels improve wound bed quality, increase granulation tissue and have pain suppressive effect. CS/ChS aerogels are useful as safe, inexpensive and easy to handle materials for topical applications, such as skin chronic wounds. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 2464–2471, 2018. [ABSTRACT FROM AUTHOR]

Abstract: Chondroitin sulfate has been widely used in both medical and clinical applications. Commercial chondroitin sulfate has been mainly acquired from animal tissue extraction. Here we report a new two‐step biological strategy for producing chondroitin sulfate A and chondroitin sulfate C. First, the chondroitin biosynthesis pathway in a recombinant Bacillus subtilis strain using sucrose as carbon source was systematically optimized and the titer of chondroitin was significantly enhanced to 7.15 g/L. Then, specific sulfation transformation systems were successfully constructed and optimized by combining the purified aryl sulfotransferase IV (ASST IV), chondroitin 4‐sulfotransferase (C4ST) and chondroitin 6‐sulfotransferase (C6ST). Chondroitin sulfate A and C were enzymatically transformed from chondroitin at conversion rates of 98% and 96%, respectively. The present biological strategy has great potential to be scaled up for biosynthesis of chondroitin sulfate A and C from cheap carbon sources. [ABSTRACT FROM AUTHOR]

Abstract: Imprinting chondroitin sulfate (CS)/silica composites with Pb(II) and Cu(II) cations was explored with CS of bovine and different fish species origin. The process was based on the assumption that particular arrangements of the linear CS chains in aqueous solution, induced so as to accommodate cross complexation with the cations, would be embodied into a tridimensional matrix created through an organoalkoxysilane sol‐gel scheme. The presence of Cu(II) in the synthesis of the composites did not result in the production of significantly stronger Cu(II)‐oriented binding arrangements, and therefore, the imprinting was not successful. Inversely, for Pb(II), the materials obtained exhibited a “memory” effect for the Pb(II) ions, expressed in the observation of stronger (13%‐44%) binding as compared to the nonimprinted counterparts, and increased selectivity (1.5‐2 folds) against Cd(II). The imprinting features observed were dependent on the CS source. However, it was not possible to identify, among a set of their properties (carboxylate and sulfate abundance, percent of disulfated units, 4S/6S ratio, and molecular weight), any that correlated directly with the observed imprinting features. The augmented selectivity provided by the cation‐imprinting process may be advantageous in areas such as analytical separation, remediation, purification, sensing, and others, particularly in those cases where a certain cation is of special interest within a mixture of them. [ABSTRACT FROM AUTHOR]

Abstract: Using silver nanoparticles (AgNPs) as the nanocatalyst, l‐cysteine rapidly reduced HAuCl4 to make a stable gold nanoparticle sol (Ag/AuNP) that had a high surface‐enhanced Raman scattering (SERS) activity in the presence of Victoria blue 4R (VB4r) molecular probes. Under the selected conditions, chondroitin sulfate (Chs) reacted with the VB4r probes to form associated complexes that caused the SERS effect to decrease to 1618 cm−1. The decreased SERS intensity was linear to the Chs concentration in the range 3.1–500 ng/ml, with a detection limit of 1.0 ng/ml Chs. Accordingly, we established a simple and sensitive SERS quantitative analysis method to determine Chs in real samples, with a relative standard deviation of 1.47–3.16% and a recovery rate of 97.6–104.2%. [ABSTRACT FROM AUTHOR]

The nervous system coordinates the functions of most multicellular organisms and their response to the surrounding environment. Its development involves concerted cellular interactions, including migration, axon guidance, and synapse formation. These processes depend on the molecular constituents and structure of the extracellular matrices (ECM). An essential component of ECMs are proteoglycans, i.e., proteins containing unbranched glycan chains known as glycosaminoglycans (GAGs). A defining characteristic of GAGs is their enormous molecular diversity, created by extensive modifications of the glycans during their biosynthesis. GAGs are widely expressed, and their loss can lead to catastrophic neuronal defects. Despite their importance, we are just beginning to understand the function and mechanisms of GAGs in neuronal development. In this review, we discuss recent evidence suggesting GAGs have specific roles in neuronal patterning and synaptogenesis. We examine the function played by the complex modifications present on GAG glycans and their roles in regulating different aspects of neuronal patterning. Moreover, the review considers the function of proteoglycan core proteins in these processes, stressing their likely role as co-receptors of different signaling pathways in a redundant and context-dependent manner. We conclude by discussing challenges and future directions toward a better understanding of these fascinating molecules during neuronal development. Developmental Dynamics 247:54-74, 2018. © 2017 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

Chondroitin sulfate (CS) is a sulfated polysaccharide that plays essential physiological roles. Here, we report an enzyme-based method for the synthesis of a library of 15 different CS oligosaccharides. This library covers 4-O-sulfated and 6-O-sulfated oligosaccharides ranging from trisaccharides to nonasaccharides. We also describe the synthesis of unnatural 6-O-sulfated CS pentasaccharides containing either a 6-O-sulfo-2-azidogalactosamine or a 6-O-sulfogalactosamine residue. The availability of structurally defined CS oligosaccharides offers a novel approach to investigate the biological functions of CS. [ABSTRACT FROM AUTHOR]

In this research, fabrication of gelatin/chondroitin sulfate (GAG) nanofibrous scaffolds using electrospinning technique for skin tissue engineering was studied. The influence of GAG content on chemical, physical, mechanical and biological properties of the scaffolds were investigated. Human dermal fibroblast (HDF) cells were cultured and bioactivity of electrospun gelatin/GAG scaffolds for skin tissue engineering was assayed. Biological results illustrated that HDF cells attached and spread well on gelatin/GAG nanofibrous scaffolds displaying spindle-like shapes and stretching. MTS assay was performed to evaluate the cell proliferation on electrospun gelatin/GAG scaffolds. The results confirmed the influence of GAG content as well as the nanofibrous structure on cell proliferation and attachment of substrates. The gelatin/GAG nanofibrous scaffolds with the desired thickness for in-vivo evaluations were used on the full-thickness wounds. Pathobiological results showed that cell loaded gelatin/GAG scaffolds significantly accelerated wounds healing. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 2020-2034, 2017. [ABSTRACT FROM AUTHOR]