Progress of research on application of chondroitin sulfate in osteogenic repair materials / 口腔医学

@#Chondroitin sulfate is an important component of extracellular matrix (ECM) in animal and human body. In recent years, chondroitin sulfate has been proven to have potential efficacy in biomedical application and has been widely used in bone regeneration and osteogenesis, especially in craniofacial reconstruction and dental medicine. Research shows that chondroitin sulfate derivatives and chondroitin sulfate composite scaffolds have great potential in promoting osteogenesis and biomineralization. However, due to the variety of chondroitin sulfate and various application forms, study on its mechanism of osteogenic repair is still insufficient. In this paper, biological characteristics, bone regeneration and osteogenesis of chondroitin sulfate, its application in different biomaterial design and future prospect are discussed.

Development and validation of a HILIC-HPLC-ELSD method for simultaneous determination of glucosamine hydrochloride and chondroitin sulfate in dietary supplements

Abstract The objective of the present study is to develop and validate a simple, selective and accurate hydrophilic interaction liquid chromatography - a high performance liquid chromatography incorporating an evaporative light scattering detector (HILIC-HPLC-ELSD) method for simultaneously determining glucosamine hydrochloride and chondroitin sulfate in dietary supplements. The chromatographic separation was carried out on a ZIC-HILIC column (150 mm x 4.6 mm x 5µm) in isocratic system mode with a mobile phase of acetonitrile, 30 mM ammonium formate and water (77:20:3, v/v/v) at pH 4.5, a column temperature of 35°C, a flow rate of 1 mL.min-1, and an injection volume of 5 µL. An evaporative light scattering (ELS) detector was used. Effective separation was achieved by means of analyte resolution of more than 1.5 with an analysis run time of approximately 20 minutes. The linearity of glucosamine hydrochloride and chondroitin sulfate ranged from 0.4 to 2.5 mg.mL-1. The limits of the detection and quantification of glucosamine hydrochloride were 20 and 80 mg.mL-1 respectively, while for chondroitin sulfate they were 80 and 400 mg.mL-1. All validation parameters satisfied the acceptance criteria in accordance with International Conference on Harmonisation (ICH) guidelines. The method was successfully applied to the assay of commercial dietary supplement samples

Chondroitin sulfate from fish waste exhibits strong intracellular antioxidant potential

Chondroitin sulfate (CS) is a type of glycosaminoglycan described as an antioxidant molecule that has been found in animal species such as fish. Tilapia (Oreochromis niloticus) represents an eco-friendly source of this compound, since its economical processing generates usable waste, reducing the negative environmental impact. This waste was used for CS extraction, purification, characterization by enzymatic degradation, and evaluation of its antioxidant effect. CS obtained from tilapia presented sulfation mainly at carbon 4 of galactosamine, and it was not cytotoxic at concentrations up to 200 µg/mL. Furthermore, 100 µg/mL of CS from tilapia reduced the levels of reactive oxygen species to 47% of the total intracellular reactive oxygen species level. The ability of CS to chelate metal ions in vitro also suggested an ability to react with other pathways that generate oxidative radicals, such as the Haber-Weiss reaction, acting intracellularly in more than one way. Although the role of CS from tilapia remains unclear, the pharmacological effects described herein indicate that CS is a potential molecule for further study of the relationship between the structures and functions of chondroitin sulfates as antioxidants.

Research progress of sulfation modification of chondroitin sulfate and chondroitin sulfate preparation in Kashin-Beck disease and osteoarthritis / 中华地方病学杂志

Chondroitin sulfate (CS) is a sulfurated glycosaminoglycan, a major component of the extracellular matrix, widely distributed in skin, cartilage and vascular tissue. CS plays an important role in the physiological state regulation of articular cartilage, which affects tensile strength and elasticity of tissues by influencing aggrecan. Previous studies have shown that CS sulfate modification may be related to the growth and development disorders of cartilage tissue and the occurrence of osteoarticular diseases. At the same time, CS is also a common joint supplement, often used in the treatment of osteoarthritis and Kashin-Beck disease. In this paper, the research progress of CS sulfate modification characteristics in Kashin-Beck disease and osteoarthritis and the application of the preparation in the treatment of Kashin-Beck disease and osteoarthritis are reviewed, aiming to provide help for the investigation of the etiology of Kashin-Beck disease and the treatment of osteoarthritis and Kashin-Beck disease.

Expression of GFAP, β-tubulin Ⅲ and synaptophsin and intervention effects of chondroitin sulfate in brain of rats with chronic fluorosis / 中华地方病学杂志

Objective:To observe the expression levels of glial fibrillary acidic protein (GFAP), β-tubulin Ⅲ and synaptophsin, and explore the role of tripartite synapse in the mechanism of central nervous system (CNS) injury and the neuroprotective effect of chondroitin sulfate (CS).Methods:One month old clean grade, 48 female Sparague-Dawley rats and 48 male Sparague-Dawley rats, were randomly divided into 8 groups according to body weight (90 - 120 g) by random number table method, with 12 rats in each group, half male and half female. These rats were fed with water containing different concentrations of sodium fluoride (NaF) [ < 0.5 mg/L (control, CN), 10.0 mg/L (low dose fluoride, LF) and 50.0 mg/L (high dose fluoride, HF)]. Some rats were fed directly for 185 days (CN, LF and HF groups). In addition, rats of CN + normal saline (NS), LF + NS, HF + NS groups and LF + CS, HF + CS groups, were intraperitoneally injected with NS or 0.66 mg/kg CS for 5 consecutive days after 180 days of feeding. After the experiment, the pathological changes of hippocampal CA4 of brain tissue in each group were observed by hematoxylin eosin staining under light microscope, and the expression and distribution of GFAP, β-tubulin Ⅲ and synaptophsin in hippocampal CA4 of rats were detected by immunohistochemistry, the expression of GFAP, β-tubulin Ⅲ and synaptophsin at protein level in hippocampus of rats were detected by Western blotting.Results:Under light microscope, eosinophilic change, loss and irregular arrangement of neuron in the hippocampal CA4 were observed in LF, HF, LF + NS and HF + NS groups. The morphology of LF + CS and HF + CS groups was not significantly changed compared with CN group, but was significant changed compared with LF, HF, LF + NS and HF + NS groups. Immunohistochemical results showed that the rates of positive area of GFAP, β-tubulin Ⅲ and synaptophsin in female and male rats in LF and HF groups were significantly decreased than those in CN group ( P < 0.05); the positive area rates of female and male rats in LF + CS and HF + CS groups were higher than those in LF and HF groups, respectively ( P < 0.05). Western blotting results showed that the proten expression levels of GFAP, β-tubulin Ⅲ and synaptophsin of female and male rats in LF and HF groups (LF group: 0.90 ± 0.09, 0.82 ± 0.08, 1.43 ± 0.14, 0.92 ± 0.02, 1.21 ± 0.15, 0.87 ± 0.02, HF group: 0.58 ± 0.14, 0.73 ± 0.03, 0.63 ± 0.06, 0.67 ± 0.03, 0.87 ± 0.04, 0.70 ± 0.05) were lower than those in CN group (1.24 ± 0.08, 1.09 ± 0.10, 2.64 ± 0.30, 1.54 ± 0.09, 1.72 ± 0.10, 1.13 ± 0.06, P < 0.05). Conclusions:The tripartite synapse and extracellular matrix may take part in pathogenesis of the damages of CNS results from chronic fluorosis; CS may reduce the injury to a certain extent.

Comparative study of glucosamine sulfate and chondroitin sulfate in the treatment of adult Kashin-Beck disease / 中华地方病学杂志

Objective:To observe and compare the efficacy and safety of glucosamine sulfate (GS) and chondroitin sulfate (CS) in the treatment of adult Kashin-Beck disease (KBD), so as to provide effective medical evidence for the standardized treatment of adult KBD.Methods:A clinical randomized controlled trial was conducted in Fuyu County and Shangzhi City, KBD historical seriously ill areas in Heilongjiang Province. A total of 247 patients were selected according to the standard of "Diagnosis of Kashin-Beck Disease" (WS/T 207-2010). According to gender, age and KBD condition, they were randomly divided into GS and CS groups, 124 and 123 respectively. Follow up once a month to investigate the medication and clinical symptoms of patients, and distribute drugs for the next stage. Fasting blood and urine samples were collected before, during and at the end of treatment (0, 90 and 180 d). Serum interleukin (IL)-1β content and urine pyridine (PYD) level were measured by enzyme-linked immunosorbent assay (ELISA). The visual analogue scale (VAS) score, affected joints, self-evaluation of curative effect and side effects were evaluated through the questionnaire, joint dysfunction and drug efficacy were evaluated according to the criteria of "Evaluation of Therapeutic Effect of Kashin-Beck Disease" (WS/T 79-2011).Results:Expression of cytokines related to cartilage metabolism: at 180 d of treatment, serum IL-1β contents and urinary PYD levels in GS and CS groups were lower than those at 0 d of treatment ( Z = - 2.461, - 2.160, - 5.075, - 5.471, P < 0.05). VAS score: at 90 and 180 d of treatment, the scores of knee pain, stiffness and function in GS and CS groups were lower than those at 0 d of treatment ( P < 0.05); and at 180 d of treatment, the scores of knee stiffness and function in GS group were lower than those in CS group ( P < 0.05). Evaluation of affected joints: at 90 and 180 d of treatment, the scores of joint pain, swelling and stiffness in GS and CS groups were lower than those at 0 d of treatment ( P < 0.05). Self-evaluation of curative effect: at 180 d of treatment, the self-evaluation of curative of CS group were better than that at 90 d of treatment (χ 2 = 9.376, P < 0.05). Evaluation of side effects: at 90 and 180 d of treatment, the side effects in GS and CS groups were mainly gastrointestinal symptoms. Joint dysfunction score: at 90 d of treatment, the sum of effective rate and markedly effective rate in GS group was higher than that in CS group (χ 2 = 4.042, P < 0.05), but there was no significant difference between the two groups at 180 d of treatment (χ 2 = 0.869, P > 0.05). Conclusion:GS and CS have certain therapeutic effects on adult KBD, which can improve symptoms and reduce serum IL-1β content and urinary PYD level, but GS takes effects quickly, and its effect on improving joint stiffness and function are better than CS.

Comparative study on chemical constituents of Cervi Cornu Pantotrichum in different forms of Cervus nipport / 中草药

Objective: In order to study the differences in chemical constituents of Cervi Cornu Pantotrichum in different forms of Cervus nipport. Methods: The content of polysaccharides, crude protein, amino acids, collagen, fatty acids, mineral elements, biogenic amines, nucleosides, and chondroitin sulfate composition of Cervi Cornu Pantotrichum in different forms of C. nipport was determined and compared by using UV spectrophotometer, Dumas azotometer, automatic amino acid analyzer, UPLC, HPLC, gas chromatograph, inductively coupled plasma mass spectrometer and atomic fluorescence photometer. Principal component analysis was used to evaluate the quality of Cervi Cornu Pantotrichum. Results: The content of polysaccharide, crude protein, amino acid, collagen, fatty acid, mineral element, biogenic amine, nucleoside and chondroitin sulfate in one-branched velvet antler was 8.40 mg/g, 44.82%, 42.24%, 23.23%, 6 234.69 mg/kg, 145.69 mg/g, 55.12 mg/kg, 2 271.87 mg/kg, and 0.74 mg/g, respectively; The content in two-branched velvet antler was 8.14 mg/g, 52.12%, 48.57%, 21.50%, 8 684.27 mg/kg, 126.40 mg/g, 76.14 mg/kg, 3 438.37 mg/kg, and 1.94 mg/g, respectively; The content in three-branched velvet antler was 8.64 mg/g, 51.86%, 45.49%, 22.31%, 9 100.78 mg/kg, 138.36 mg/g, 70.75 mg/kg, 2 507.82 mg/kg, and 1.84 mg/g, respectively. Conclusion: The chemical composition of Cervi Cornu Pantotrichum in different forms of C. nipport was different. The results of principal component analysis showed that the quality of the two-branched velvet antler was the best, the three-branched velvet antler was the second, the one-branched velvet antler was the worst. This study provides a reference for the quality evaluation and grading standards of different forms of Cervi Cornu Pantotrichum.

Oral glucosamine and chondroitin sulfate on synovial fluid biomarkers from osteoarthritic equine joints / Efeitos da glucosamina e condroitim sulfato oral sobre biomarcadores do líquido sinovial de articulações osteoartríticas de equinos

ABSTRACT: Several studies, mainly in vitro, have shown that chondroitin sulfate (CS) and glucosamine (GlcN) do have chondro protective and anti-inflammatory actions. The aim of the present study was to investigate whether oral CS/GlcN supplementation has effects on the CS, hyaluronic acid (HA) and prostaglandin E2 (PGE2) concentrations on synovial fluid of equine osteoarthritic joints. Horses with mild osteoarthritis (OA) in tibiotarsal joint received daily PO doses of CS and GlcN (2.8/3.1 g) for 25 days. Synovial fluid (SF) and urine samples were collected before treatment (day 0), and every 7 days, until day 55 (30 days after the end of treatment). Urinary CS increased upon oral treatment, indicating that this compound was systemically distributed. Concerning the SF, CS concentration increased after the end of the treatment and returning to baseline afterwards, while HA and PGE2 concentrations did not change. Despite the systemic distribution, oral supplementation of CS/GlcNfor 25 days was insufficient as an anti-inflammatory support. However, it is possible to infer that there was an anabolic effect upon cartilage matrix.

RESUMO: Vários estudos, principalmente in vitro, têm mostrado que o condroitim sulfato (CS) e a glucosamina (GLcN) possuem ação condroprotetora e anti-inflamatória. O objetivo deste trabalho foi investigar se a administração oral de CS/GLcN possui efeito sobre as concentrações de CS, ácido hialurônico (AH) e prostaglandina E2 (PGE2) do liquido sinovial (LS) de articulações equinas com osteoartrite (OA). Cavalos diagnosticados com OA tibiotársica grau leve receberam por via oral doses diárias de CS e GlcN (2,8g/3,1g) por 25 dias. Amostras de LS e urina foram coletadas antes do tratamento (dia 0) e a cada 7 dias até o dia 55 (30 dias após o fim do tratamento). Houve aumento do CS urinário, indicando a distribuição sistêmica desse composto. No LS, a concentração de CS aumentou após o final do tratamento e retornou aos valores basais em seguida, enquanto as concentrações de HA e PGE2 não apresentaram alterações. Apesar da distribuição sistêmica, a suplementação oral de CS/GlcNpor 25 dias foi insuficiente como medida anti-inflamatória. Contudo, pode-se inferir que houve efeito anabólico sobre a matriz cartilagínea.

Neuroprotective Effects of Low-molecular-weight Chondroitin Sulfate on Dopaminergic Neurons in MPTP-induced Parkinson ’s Disease Model Mice / 中国药房

OBJECTIVE： To observe neuroprotective effects of low-molecular-weight chondroitin sulfate （CS） on dopaminergic neurons in Parkinson’s disease （PD） mice model induced by 1-methyl-4-phenyl-1，2，3，6-tetrahydropyridine （MPTP）. METHODS： C57BL/6 mice were randomly divided into control group， MPTP injury group， low-molecular-weight CS low-dose and high-dose groups （100， 400 mg/kg）. Control group and MPTP injury group were given constant volume of normal saline intragstrically， administration groups were given relevant medicine intragastrically， once a day， for consecutive 17 d. Since 11th day after medication， except for control group， other groups were given MPTP solution （20 mg/kg） intraperitoneally to induce PD model， once a day， consecutive 5 d. After last medication， behavioral changes of mice （10 mice in each group） were evaluated by rotary rod fatigue tester. The damage of dopamine neurons （the percentage of TH positive cell and the percentage of fluorescence intensity） in substantia nigra of mice （3 mice in each group） was detected by immunohistochemistry and immunofluorescence. The content of dopamine in striatum was determined by HPLC （6 mice in each group）. The changes of oxidant stress indexes （SOD， GSH-Px， MDA） in substantia nigra of mice were determined by chemical colorimetry （6 mice in each group）. RESULTS： Compared with control group， retention time of mice on rotating rods was shortened significantly in MPTP injury group； TH positive cells of substantia nigra were decreased significantly， fluorescence intensity was obviously weakened； the percentage of positive cells and fluorescence intensity， the content of dopamine in striatum， the activities of SOD and GSH-Px in substantia nigra were decreased significantly， while the content of MDA was increased significantly （P＜0.01）. Compared with MPTP injury group， retention time of mice on the rotating rods was prolonged significantly in low-molecular-weight CS groups， the number of TH positive cells was increased significantly in substantia nigra and fluorescence intensity was increased significantly； the percentage of positive cells， the percentage of fluorescence intensity and the content of dopamine in striatum were increased significantly， while above indexes of high-dose group were significantly longer or higher than those of low-dose group （P＜0.05 or P＜0.01）. The activities of SOD and GSH-Px in substantia nigra were increased significantly in low-molecular-weight CS groups， while the content of MDA in substantia nigra was decreased significantly in low-molecular-weight CS high-dose group （P＜0.05 or P＜0.01）. CONCLUSIONS： Prophylactic administration of low-molecular-weight CS can relieve the damage of dopaminergic neurons in substantia nigra of PD model mice induced by MPTP in a dose-dependent manner， and increase the secretion of dopamine in striatum. The effect may be related to the inhibition of lipid peroxidation and the enhancement of antioxidant capacity of tissues.

Expression of Erk kinase, AMPA receptor subunits GluR1 and GluR2, and protection of chondroitin sulfate in brain of rats with chronic fluorosis / 中华地方病学杂志

Objective To study the mechanism of central nervous system (CNS) injury in chronic fluorosis and the neuroprotective effect of chondroitin sulfate (CS).Methods Forty-eight female Sprague-Dawley rats weighting 90-120 g were divided into 8 groups according to body weight by random number table,6 rats in each group:control group,drinking tap water freely;low dose and high dose fluoride groups,freely drinking tap water with fluoride content of 10 and 50 mg/L,respectively;control + normal saline (NS),low dose fluoride + NS,and high dose fluoride + NS groups,each group was fed for 180 d,and treated with intraperitoneal injection of 0.66 mg/kg NS for 5 d (once a day);low dose fluoride + CS and high dose fluoride + CS groups,each group was fed for 180 d,0.66 mg/kg CS was injected intraperitoneally for 5 d (once a day).All groups were fed standard nutritive animal feed for 185 d and dissected for brain tissue.The pathologic change was observed after hematoxylin-eosin (HE)staining;the expression levels of phosphorylated extracellular signal-regulated protein kinase 1/2 (phospho-Erk1/2)and glutamate receptors 1,2 (GluR1,GluR2) in the brain cortex were detected by immunohistochemistry;the protein levels of Erk1/2,phospho-Erk1/2,GluR1,and GluR2 in the brain cortex were detected by Western blotting.Results Brain cortex of all rats in the fluoride groups showed eosinophilic degeneration,loss and disordered arrangement of neurons,and the brain morphological changes in each fluoride + CS groups were significantly improved compared with those in the fluoride groups.Immunohistochemistry results showed that compared with the control group [(0.44 ± 0.09)％,(1.49 ± 0.05)％,(2.51 ± 0.54)％],the expression levels of phospho-Erk1/2 [(1.47 ±0.09)％,(1.03 ± 0.05)％],and GluR2 [(2.37 ± 0.06)％,(3.38 ± 0.12)％] in the low dose and high dose fluoride groups were increased,and the expression levels of GluR1 [(1.49 ± 0.02)％,(0.99 ± 0.19)％] were decreased (P ＜ 0.05).Western blotting results showed that compared with the control group (1.00 ± 0.12,1.76 ± 0.33),the protein levels of Erk1/2 (3.10 ± 0.76,1.99 ± 0.01) and phospho-Erk1/2 (3.27 ± 0.25,2.67 ± 0.05) in low dose and high dose fluoride groups were significantly increased (P ＜ 0.05);compared with low dose fluoride group,the protein levels of Erk1/2,and phospho-Erk1/2 (1.30 ± 0.31,2.20 ± 0.34) in low dose fluoride + CS group decreased significantly (P ＜0.05).Compared with control group (1.86 ± 0.47,1.17 ± 0.27),the protein levels of GluR1 (1.09 ± 0.26,0.61 ± 0.14) in low dose and high dose fluoride groups decreased significantly,while the protein level of GluR2 (1.99 ± 0.42,3.38 ±0.27) increased significantly (P ＜ 0.05);compared with low dose and high dose fluoride groups,the protein levels of GluR2 in low dose fluoride + CS and high dose fluoride + CS groups (1.53 ± 0.41,2.65 ± 0.32) decreased significantly (P ＜ 0.05).The protein level of phospho-Erk1/2 was negatively correlated with GluR1 protein level (r =-0.975,-0.991,P ＜ 0.05) in low dose and high dose fluoride groups,and it was positively correlated with the protein level of GluR2 (r =0.986,0.993,P ＜ 0.05).Conclusion The CNS injury caused by chronic fluorosis may be related to GluR1 and GluR2 activated Erk1/2 signaling pathway,and CS has certain protection to the injury.

Glucosamine combined with chondroitin sulfate for knee osteoarthritis:a systematic review / 中华风湿病学杂志

Objective To evaluate the efficacy and safety of glucosamine (GS) combined with chondroitin sulfate (CS) for knee osteoarthritis (OA).Methods We searched the CENTRAL (Issue 1,2016),PUBMED (1946 to 2016.1.20),EMBASE (1947 to 2016.1.20),CBM (1978 to 2016.1.20),CNKI (1994 to 2016.1.20),WanFang Data (1980 to 2016.1.20) and VIP (1989 to 2016.1.20) and we searched randomized controlled trials (RCT) of GS combined with CS for knee OA.Two reviewers independently identified the included trials,evaluated the quality of methodology and extracted data.The Review manager 5.3 software was used for data analysis.Results Four RCTs were included in this systematic review.The combination group compared with GS group,a total number of 1 145 patients,and the combination group compared with CS group,a total number of 1 067 patients.The outcomes showed:① Among those 4 RCTs that comparing the combination with GS,three RCTs reported western ontario and mcmaster universities osteoarthritis index (WOMAC) score.Summarized results of these 3 RCTs showed no significant difference between combination group and GS group (all P values ＞0.05).MDs of WOMAC score in pain,stiffnessand function were-6.60[95％CI(-18.79,5.59),-15.90(-43.09,11.29) and-6.44 (-16.46,3.59)].② Two long-term (≥6 months) study (n=937) compared the combination with CS group and showed no significant difference (all P values ＞0.05).Pooled MD of WOMAC score in pain and function were-0.80 (-4.96,3.36) and-1.76 (-4.46,0.94) respectively.③ Subgroup analysis in 1 RCT showed that in moderate-to-severe knee pain group (n=142) combination obviously improve the WOMAC score in pain,stiffnessand function compared with CS groupand the differences werestatistically significant (all P values ＜0.05).MDs were-10.46(-17.98,-2.94),-9.70(-18.48,0.92) and 9.39 (-17.33,-1.45).Conclusion There is no evidence to support that the combination of GS and CS for knee OA cansignificantlyimprovethe WOMAC score compared with either GS or CS.For patients with-moderate-to-severe knee pain,combination might be superior toeither GS or CS.

Production and application of 3-phosphoadenosine-5- phosphosulfate / 生物工程学报

Sulfated compounds are widely present in cytoplasm, on cell surface, and in extracellular matrix. These compounds play important roles in cell development, differentiation, immune response, detoxication, and cell signal transduction. 3-Phosphoadenosine-5-phosphosulfate (PAPS) is the universal sulfate group donor for the biosynthesis of sulfated compounds. Up to now, the synthesis of PAPS is still too expensive for industrial applications. This review focuses on the recent progress of PAPS production and summaries the application of PAPS, particularly in the production of glucosinolate, heparin, condroitin sulfate, and oxamniquine production.

Correlation be tween serum expression levels of decorin, chondroitin sulfate and C-terminal crosslinking telopeptide of type Ⅱ collagen and clinical features in arthritis patients / 中华风湿病学杂志

Objective To investigate changes in the serum expression levels of decorin (DCN),chondroitin sulfate (CS) and C-terminal crosslinking telopeptide of type Ⅱ collagen (CTX-Ⅱ) and analyze the relationship between the expression levels and clinical characteristics,aiming to provide evidence for clinical diagnosis and treatment of arthritis.Methods Ninety subjects were divided into the osteoarthritis (OA),rheumatoid arthritis (RA) and control groups (n=30 for each group).The OA and RA patients were diagnosed in Department of Rheumatism Immunity of Shanxi Dayi Hospital,and the healthy volunteers who were doing physical check-up in the Physical Check-up Center of Shanxi Dayi Hospital were recruited as the control subjects.Visual analogue scale (VAS) was utilized to evaluate the degree of pain.X-ray was performed to assess the severity of joint lesions.Enzyme linked immunosorbent assay (ELISA) was adopted to quantitatively measure the serum expression levels of DCN,CS and CTX-Ⅱ.The correlation among these parameters was statistically analyzed.Measurement data among different groups were statistically compared by one-way analysis of variance (ANOVA)and comparison between two groups was conducted by Tambane's T2.The correlation between two factors was analyzed by using Spearman correlation analysis.Results The difference of the serum expression levels of DCN in the OA,RA and control groups [(3.19±1.38) ng/ml,(1.90±0.62) ng/ml,(1.33 ±0.33) ng/ml] was statistically significant,which of the OA group was higher than the control group (P＜0.01),which of the RA group was higher than control group (P＜0.01),which of the OA group was higher than the RA group (P＜0.001).The difference of the serum expression levels of CS in the OA,RA and control groups [(0.57±0.12) ng/ml,(0.95±0.47) ng/ml,(0.36±0.09) ng/ml] was statistically significant,in which,the OA group was higher than the control group (P＜0.01),RA group was higher than control group (P＜0.01),OA group was lower than RA group (P＜0.01).The serum expression levels of CTX-Ⅱ in the OA and RA groups [(3.08±0.86) ng/ml,(3.03±1.14) ng/ml] were significantly up-regulated compared with those in the control group [(2.17±0.82) ng/ml](P＜0.001,P=0.005).The expression level of CTX-Ⅱ did not significantly different between the OA and RA groups (P=0.996).In the OA and RA groups,the serum levels of DCN,CS,CTX-Ⅱ were po-sitively related to X-ray and VAS,the correlation of DCN was the highest,the correlation was high in both OA (r=0.777,P＜0.01;r=0.622,P＜0.01) and RA (r=0.640,P＜0.01;r=0.493,P=0.006).Conclusion The serum levels of DCN,CS and CTX-Ⅱ in arthritis patients are significantly up-regulated compared with those in healthy controls,which are positively correlated with the degree of pain,and the severity of cartilage and bone defects.DCN is probably valuablein the differential diagnosis of early OA and RA.However,the clinical application remains to be further validated by evidence-based studies.

Effectiveness of glucosamine chondroitin sulfate calcium tablets for knee osteoarthritis / 中华创伤骨科杂志

Objective To study the effectiveness,side effects and safety of glucosamine chondroitin sulfate calcium (Keylid(R)) tablets for osteoarthritis of the knee.Methods To review the clinical data of 103 cases of primary knee osteoarthritis who had been treated between March 2013 and February 2014.The intervention group included 53 cases (23 males and 30 females with an age of 71.0 ± 9.1 years) who received Keylid(R) tablets (2 tablets twice per day) accompanied with exercise therapy for 24 weeks;the control group included 50 cases (22 males and 28 females with an age of 69.8 ± 8.8 years) who received placebo (2 tablets twice per day) accompanied with exercise therapy for 24 weeks.No NSAIDs,calcium or other articular dietary supplements were taken by the 2 groups during the study period.T-test was used to analyse the results.Results After treatment,the range of knee motion for the intervention group (123.27° ± 10.22°) was significantly larger than that for the control group (116.34° ± 8.76°),and the average pain visual analogue score after 40 m walking for the former (2.92 ± 0.36) significantly lower than that for the latter (3.30 ± 0.22) (P ＜ 0.05).There were no significant differences between the 2 groups in Western Ontario and McMaster Universities Arthritis Index,time for 40 m walking or circumference of the knee (P ＞ 0.05).No side effects were observed in either group during the study period.Conclusion Keylid(R) tablets complemented with exercise therapy may reduce pain and improve knee function with no adverse events in the patients with knee osteoarthritis.

Analysis of chondroitin sulfate content of Cervi Cornu Pantotrichum with different processing methods and different parts / 中国中药杂志

The differences and the variations of chondroitin sulfate content in different parts of Cervi Cornu Pantotrichum(CCP) with different processing methods were investigated. The chondroitin sulfate from velvet was extracted by dilute alkali-concentrated salt method. Next， the chondroitin sulfate was digested by chondroitinase ABC.The contents of total chondroitin sulfate and chondroitin sulfate A， B and C in the samples were determined by high performance liquid chromatography(HPLC).The content of chondroitin sulfate in wax，powder，gauze，bone slices of CCP with freeze-drying processing is 14.13，11.99，1.74，0.32 g·kg⁻¹， respectively. The content of chondroitin sulfate in wax，powder，gauze，bone slices of CCP with boiling processing is 10.71，8.97，2.21，1.40 g·kg⁻¹， respectively. The content of chondroitin sulfate in wax，powder，gauze，bone slices of CCP without blood is 12.47，9.47，2.64，0.07 g·kg⁻¹， respectively. And the content of chondroitin sulfate in wax，powder，gauze，bone slices of CCP with blood is 8.22，4.39，0.87，0.28 g·kg⁻¹ respectively. The results indicated that the chondroitin sulfate content in different processing methods was significantly different.The content of chondroitin sulfate in CCP with freeze-drying is higher than that in CCP with boiling processing.The content of chondroitin sulfate in CCP without blood is higher than that in CCP with blood. The chondroitin sulfate content in differerent paris of the velvet with the same processing methods was arranged from high to low as： wax slices， powder， gauze slices， bone slices.

Synthesis and Biocompatibility Characterizations of in Situ Chondroitin Sulfate–Gelatin Hydrogel for Tissue Engineering

Novel hydrogel composed of both chondroitin sulfate (CS) and gelatin was developed for better cellular interaction through two step double crosslinking of N-(3-diethylpropyl)-N-ethylcarbodiimide hydrochloride (EDC) chemistries and then click chemistry. EDC chemistry was proceeded during grafting of amino acid dihydrazide (ADH) to carboxylic groups in CS and gelatin network in separate reactions, thus obtaining CS–ADH and gelatin–ADH, respectively. CS–acrylate and gelatin–TCEP was obtained through a second EDC chemistry of the unreacted free amines of CS–ADH and gelatin–ADH with acrylic acid and tri(carboxyethyl)phosphine (TCEP), respectively. In situ CS–gelatin hydrogel was obtained via click chemistry by simple mixing of aqueous solutions of both CS–acrylate and gelatin–TCEP. ATR-FTIR spectroscopy showed formation of the new chemical bonds between CS and gelatin in CS–gelatin hydrogel network. SEM demonstrated microporous structure of the hydrogel. Within serial precursor concentrations of the CS–gelatin hydrogels studied, they showed trends of the reaction rates of gelation, where the higher concentration, the quicker the gelation occurred. In vitro studies, including assessment of cell viability (live and dead assay), cytotoxicity, biocompatibility via direct contacts of the hydrogels with cells, as well as measurement of inflammatory responses, showed their excellent biocompatibility. Eventually, the test results verified a promising potency for further application of CS–gelatin hydrogel in many biomedical fields, including drug delivery and tissue engineering by mimicking extracellular matrix components of tissues such as collagen and CS in cartilage.

Protective effect and mechanism of chondroitin sulfate on brain injury induced by experimental chronic fluorosis in rats / 中华地方病学杂志

Objective To investigate the changes of extracellular regulated protein kinases (Erk)1/2,phospho-Erk1/2,matrix metalloproteinases (MMP)-2 and MMP-9 in rats with experimental chronic fluorosis and the role of chondroitin sulfate in treatment of rat with experimental chronic fluorosis.Methods Using a group design and cell culture methods,the SH-SY5Y cells were divided into control group (culture medium with 0.0 mmol/L fluoride ion),fluoride group (fluoride ion:4.0 mmol/L) and chondroitin sulfate group (fluoride ion:4.0 mmol/L,chondroitin sulfate:0.4 g/L).The ultrastructural changes of the SH-SY5Y cells were observed through electron microscope after 24 h treatment.The SH-SY5Y cells were cultured for 72 h,the number of cells survived in three groups.were detected after stained by trypan blue.Fifteen clean grade SD rats with body weight of 100-120 g were divided into control group (tap water:fluorine content less than 0.5 mg/L),fluoride group (fluoride ion:10.0 mg/L) and chondroitin sulfate group (fluoride ion:10.0 mg/L,the rats were performed intraperitoneal injection with 0.66 mg/kg chondroitin sulfate for 5 days after intaking fluoride for 90 days) on the basis of random number table.Five rats were in each group,and the experiment was carried out for 95 days.The capability of learning and memory of rats were tested by Morris water maze test;the expression of phospho-Erk1/2,Erk1/2,MMP-2 and MMP-9 protein in brain tissue was detected by Western blotting;the expression of phospho-Erk1/2,MMP-2 and MMP-9 in hippocampus CA2 area of brain was detected by immunohistochemistry.Results More vesicles and swelling of mitochondria or endoplasmic reticulum were observed in SH-SY5Y cell treated with fluoride through electron microscope,but relatively less in chondroitin sulfate group.Survival rate and amount of SH-SY5Y treated with chondroitin sulfate [(92 ± 23)％ and (7.83 ± 1.38) × 106/ml] were significantly higher than that of fluoride group [(55 ± 2)％,(2.19 ± 1.26) × 106/ml,P ＜ 0.05].Animal experiment results showed that most rats in control group and chondroitin sulfate group used spatial direct search strategy,and the amount of this search strategy (2.20 ± 1.09,3.40 ± 1.34) was more than that in fluoride group (0.40 ± 0.54,P ＜ 0.05).The expression of phospho-Erk1/2 in brain tissue of rats in fluoride group (3.26 ± 0.88) was significantly higher than that in control group (1.53 ± 0.28) and chondroitin sulfate group (2.36 ± 0.87,P ＜ 0.05).Immunohistochemistry results showed that average gray value of phospho-Erk1/2 in chondroitin sulfate group (220.20 ± 3.09) was significantly higher than that of the control group and the fluoride group (100.00 ± 0.00,130.98 ± 1.27,P ＜ 0.05).The average gray value of MMP-2 in the fluoride group (294.52 ± 5.18) was significantly higher than that in control group and chondroitin sulfate group (100.00 ± 0.00,117.95 ± 1.55,P ＜ 0.05).The average gray value of MMP-9 protein of the fluoride group (993.64 ± 3.66) and the chondroitin sulfate group (1 167.30 ± 239) was significantly higher than that of control group (100.00 ± 0.00,P ＜ 0.05).Conclusions Erk1/2 pathway possibly maintains the stability of cell survival by regulating the expression of MMP-2 and MMP-9.Chondroitin sulfate can protective nerve cells and reduce the nervous damage caused by fluorosis to some certain extent.

Molecular characterization of CHST11 and its potential role in nacre formation in pearl oyster Pinctada fucata martensii

Background: C4ST-1 catalyzes the transfer of sulfate groups in the sulfonation of chondroitin during chondroitin sulfate synthesis. Chondroitin sulfate consists of numerous copies of negatively charged sulfonic acid groups that participate in the nucleation process of biomineralization. In the present study, we obtained two CHST11 genes (PmCHST11a and PmCHST11b) which encoded the C4ST-1 and explored the functions of these genes in the synthesis of chondroitin sulfate and in the formation of the nacreous layer of shells. Results: Both PmCHST11a and PmCHST11b had a sulfotransferase-2 domain, a signal peptide and a transmembrane domain. These properties indicated that these genes localize in the Golgi apparatus. Real-time PCR revealed that both PmCHST11a and PmCHST11b were highly expressed in the central zone of the mantle tissue. Inhibiting PmCHST11a and PmCHST11b via RNA interference significantly decreased the expression levels of these genes in the central zone of the mantle tissue and the concentration of chondroitin sulfate in extrapallial fluid. Moreover, shell nacre crystallized irregularly with a rough surface after RNA interference. Conclusions: This study indicated that PmCHST11a and PmCHST11b are involved in the nacre formation of Pinctada fucata martensii through participating in the synthesis of chondroitin sulfate.

Fabrication and characterization of three-dimensional cartilage nano-scaffolds based on extracellular matrix / 军事医学

Objective To produce three-dimensional cartilage nanoscaffolds based on extracellular matrix.Methods Nanoscaffolds of collagen type Ⅱ(Col-Ⅱ), hyaluronic acid(HA)and chondroitin sulfate(CS)were prepared by mixing water,trifluoroethanol and hexafluoroisopropanol as a solvent.The structure, morphology, thermal property, mechanical performance and hydrophobicity of the scaffolds were characterized.Results There were interactions between Col-Ⅱ,HA and CS.The scaffolds were hydrophobic.The Col-Ⅱ triple-helix structure wasn't completely damaged.In the study, scaffold fibers were smooth,slender and dimensionally stable.The scaffolds had good thermal stability and optimal tensile properties could be obtained at the mass ratio of 7:1:1.Conclusion In this study, scaffolds have good thermal, mechanical and structural properties and are expected to be used in cartilage repair.

Determination of Thimerosal of Chondroitin Sulfate Eye Drops in Different Packing Materials by Volt-am-pere Polarography Method / 中国药师

Objective:To establish a volt-ampere polarography method for the determination of thimerosal in chondroitin sulfate eye drops, and study the adsorption of thimerosal in different packing materials. Methods:The hanging mercury drop electrode ( HM-DE) in multi-functional electrode was used and the sample solution was added to potassium chloride-hydrochloric acid buffer solution. Nitrogen deoxygenation time was 200 s, the enrichment time was 60 s, and the equilibrium time was 5 s. The potentiometric scan was performed using a two standard addition method. Results:Thimerosal had a good linear relationship within the range of 0. 01-1. 0 mg· ml-1(r=0. 9990). The detection limit was 1. 78 μg·ml-1 and the average recovery was 99. 1% (RSD=2. 7%, n=9). Conclu-sion:The method is simple and accurate, and suitable for the determination of thimerosal in chondroitin sulfate eye drops. Low-density polyethylene and polypropylene both show obvious adsorption of thimerosal,so chondroitin sulfate eye drops should not be packed in the above packing materials.