1.Analysis of chondroitin sulfate content of Cervi Cornu Pantotrichum with different processing methods and different parts.
Rui-Ze GONG ; Yan-Hua WANG ; Yin-Shi SUN
China Journal of Chinese Materia Medica 2018;43(3):556-562
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.
Animals
;
Chondroitin Sulfates
;
analysis
;
Deer
;
Horns
;
chemistry
2.Adsorption of chondroitin sulfate-A to the surface of titanium.
Hong FAN ; Zhiqing CHEN ; Ping ZHANG ; Jing QIU
Journal of Biomedical Engineering 2003;20(4):650-667
To elucidate the adsorption mechanism of CS-A to the surface of titanium, 5 ml solutions of the CS-A were reacted with 2 g of native and 2 g of calcium-treated titanium powder for 48 h at 37 degrees C. Residual CS-A was detected by the carbazole elaborate method. The results showed that no CS-A attached to native titanium. Comparatively, titanium treated with calcium produced a significant adsorption of CS-A. At concentration of 60 micrograms/ml, the adsorption of CA-A to calcium-treated titanium powder attained the maximum, 83 micrograms/g. Only EDTA can liberate the bound CS-A from titanium surface. These findings suggest that calcium ion is necessary for the adsorption of CS-A to titanium.
Adsorption
;
Calcium
;
Chondroitin Sulfates
;
pharmacology
;
Static Electricity
;
Surface Properties
;
Titanium
;
chemistry
3.In vivo study of extracellular matrix coating enhancing fixation of the pedicle screw-bone's interface.
Guo-Min LIU ; Xing-Yi ZHANG ; Chuan-Jie XU ; Xiao-Min ZHU ; Jun WANG ; Yi LIU
Chinese Medical Journal 2011;124(23):3945-3952
BACKGROUNDBased on in vivo research on the effect of the coating of the extracellular matrix composition of pedicle screws on the conduction and induction of bone formation in young sheep, the aim of this study was to investigate the application of coated pedicle screws in sheep with scoliosis whose spines are under constant development.
METHODSFour groups of pedicle screws were randomly implanted into bilateral L2-L5 pedicles of 2.5- to 3-month-old sheep. A static experiment was performed on one side and a loading test was performed on the other side by implanting connecting rods at the L2-L3 and L4-L5 segments. The changes in the force on the coated screws and the combination of the surface of the coated screws with the surrounding bone in the growth process of young sheep's spines with aging were observed. After 3 months, the lumbar vertebrae with the screws were removed and examined by micro-CT, histological, and biomechanical analyses.
RESULTSUnder nonloading conditions, there is bone formation around the surfaces of coated screws. The bone forming on the surface of collagen/chondroitin sulfate/hydroxyapatite coating of pedicle screws is the most, the one of the collagen/chondroitin sulfate coating and hydroxyapatite coating is followed, and no significant difference between the two groups. In terms of the trabecular bone morphology parameters of the region of interest around the surface of the pedicle screws, such as bone mineral content, bone mineral density, tissue mineral content, tissue bone mineral density, bone volume fraction, and connection density, those associated with collagen/chondroitin sulfate/hydroxyapatite coatings are largest and those unassociated with coatings are smallest. Under nonloading conditions, the pullout strength of the collagen/chondroitin sulfate/hydroxyapatite-coated screws was largest, and that of the uncoated screws was minimal (P < 0.01). Under loading conditions, the maximum pullout strength of each group of pedicle screws was less than that of the pedicle screws in the nonloading state (P < 0.01) with no significant difference between the groups (P > 0.05).
CONCLUSIONSUnder nonloading conditions, the coatings of both organic and inorganic components of the extracellular matrix of titanium pedicle screws can conduct or induce bone formation around the surface of the screws. The ability of collagen/chondroitin sulfate/hydroxyapatite coatings to induce bone formation is stronger; under loading conditions, a large amount of connective tissue formed around the surfaces of the screws in each group. No significant differences were found between the groups.
Animals ; Biomechanical Phenomena ; Bone Screws ; Chondroitin Sulfates ; chemistry ; Collagen ; chemistry ; Durapatite ; chemistry ; Rats ; Sheep ; X-Ray Microtomography
4.Synthesis and Biocompatibility Characterizations of in Situ Chondroitin Sulfate–Gelatin Hydrogel for Tissue Engineering
Sumi BANG ; Ui Won JUNG ; Insup NOH
Tissue Engineering and Regenerative Medicine 2018;15(1):25-35
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.
Amines
;
Cartilage
;
Cell Survival
;
Chemistry
;
Chondroitin Sulfates
;
Chondroitin
;
Click Chemistry
;
Collagen
;
Extracellular Matrix
;
Gelatin
;
Hydrogel
;
Hydrogels
;
In Vitro Techniques
;
Spectrum Analysis
;
Tissue Engineering
;
Transplants
5.Analysis of type-I collagen fibrils and chondroitin sulfate distribution in human dentin by confocal laser scanning microscopy combined with dual immunofluorescent labeling technique.
Shuai LU ; Sanjun ZHAO ; Yong SUN ; Yu GAO ; Xiaojing LI ; Jihua CHEN
Chinese Journal of Stomatology 2015;50(12):746-750
OBJECTIVETo introduce the method of dual immunofluorescence labeling of human dentin matrix without demineralization of the whole dentin fragments, and to analyze the distribution of type-I collagen fibrils and chondroitin sulfate in human dentin.
METHODSForty 30 µm- thick middle coronal dentin sections were obtained from 8 freshly extracted human third molars and etched with 37% phosphoric acid(PA) gel for 15 s. After preconditioning with or without tosyl- phenylalanyl chloromethyl ketone(TPCK) treated trypsin digestion, sections were subjected to dual immunofluorescent labeling and scanned by confocal laser scanning microscopy to identify the type-I collagen fibrils and chondroitin sulfate.
RESULTSChondroitin sulfate was localized in the lumens of the dentin tubules and peritubular dentin, while the type-I collagen fibrils were localized in intertubular dentin and peritubular dentin. After preconditioning with TPCK treated trypsin digestion, the red fluorescence was decreased or disappeared.
CONCLUSIONSThe dual immunofluorescence labeling methodology can be used to study the human dentin matrix without demineralization of the whole dentin fragments. Chondroitin sulfate was localized in the lumens of the dentin tubules and peritubular dentin, while the type-I collagen fibrils were localized in intertubular dentin and peritubular dentin.
Acid Etching, Dental ; methods ; Chondroitin Sulfates ; analysis ; Collagen Type I ; analysis ; Dentin ; chemistry ; Extracellular Matrix ; Fluorescent Antibody Technique ; methods ; Humans ; Microscopy, Confocal ; Molar ; Phosphoric Acids
6.Preparation and cytocompatibility of chitosan-based carriers of corneal cells.
Xingshuang GAO ; Wanshun LIU ; Baoqin HAN ; Xiaojuan WEI
Chinese Journal of Biotechnology 2008;24(8):1381-1386
To study the possibility of using hydroxypropyl chitosan-based blend membranes as carriers of corneal cells in tissue engineering, we prepared three kinds of blend membranes labeled hydroxypropyl chitosan/chondroitin sulfate, hydroxypropyl chitosan/gelatin/chondroitin sulfate and hydroxypropyl chitosan/oxidized hyaluronic acid/chondroitin sulfate. The transparency, water content and ability of protein adsorption of the blend membranes were measured. To evaluate the cytocompatibility of the blend membranes with corneal epithelial cells, rabbit corneal epithelial cells were cultured on the surface of the carrier membranes. The morphological characteristics, cell adhesion, cell proliferation and the activity of lactate dehydrogenase (LDH) in the media were investigated. Three kinds of blend membranes had good optical transmittance, suitable water content and ability of protein adsorption. The results showed that the less injury was made to corneal epithelial cells by the hydroxypropyl chitosan/gelatin/chondroitin sulfate blend membrane than the others. This kind of membrane was favor of the growth and adhesion of corneal epithelial cells. The hydroxypropyl chitosan/gelatin/chondroitin sulfate blend membrane is a promising carrier of corneal cells and can be used in reconstruction of tissue engineered cornea.
Animals
;
Biocompatible Materials
;
chemistry
;
pharmacology
;
Cell Culture Techniques
;
methods
;
Cell Proliferation
;
drug effects
;
Cells, Cultured
;
Chitosan
;
chemistry
;
Chondroitin Sulfates
;
chemistry
;
Epithelium, Corneal
;
cytology
;
Gelatin
;
chemistry
;
Membranes, Artificial
;
Rabbits
;
Tissue Engineering
;
methods
7.Novel nano-microspheres containing chitosan, hyaluronic acid, and chondroitin sulfate deliver growth and differentiation factor-5 plasmid for osteoarthritis gene therapy.
Zhu CHEN ; Shang DENG ; De-Chao YUAN ; Kang LIU ; Xiao-Cong XIANG ; Liang CHENG ; Dong-Qin XIAO ; Li DENG ; Gang FENG
Journal of Zhejiang University. Science. B 2018;19(12):910-923
OBJECTIVE:
To construct a novel non-viral vector loaded with growth and differentiation factor-5 (GDF-5) plasmid using chitosan, hyaluronic acid, and chondroitin sulfate for osteoarthritis (OA) gene therapy.
METHODS:
Nano-microspheres (NMPs) were prepared by mixing chitosan, hyaluronic acid, and chondroitin sulfate. GDF-5 plasmid was encapsulated in the NMPs through electrostatic adsorption. The basic characteristics of the NMPs were observed, and then they were co-cultured with chondrocytes to observe their effects on extracellular matrix (ECM) protein expression. Finally, NMPs loaded with GDF-5 were injected into the articular cavities of rabbits to observe their therapeutic effects on OA in vivo.
RESULTS:
NMPs exhibited good physicochemical properties and low cytotoxicity. Their average diameter was (0.61±0.20) μm, and encapsulation efficiency was (38.19±0.36)%. According to Cell Counting Kit-8 (CCK-8) assay, relative cell viability was 75%-99% when the total weight of NMPs was less than 560 μg. Transfection efficiency was (62.0±2.1)% in a liposome group, and (60.0±1.8)% in the NMP group. There was no significant difference between the two groups (P>0.05). Immunohistochemical staining results suggested that NMPs can successfully transfect chondrocytes and stimulate ECM protein expression in vitro. Compared with the control groups, the NMP group significantly promoted the expression of chondrocyte ECM in vivo (P<0.05), as shown by analysis of the biochemical composition of chondrocyte ECM. When NMPs were injected into OA model rabbits, the expression of ECM proteins in chondrocytes was significantly promoted and the progression of OA was slowed down.
CONCLUSIONS
Based on these data, we think that these NMPs with excellent physicochemical and biological properties could be promising non-viral vectors for OA gene therapy.
Animals
;
Cell Differentiation
;
Cell Survival/drug effects*
;
Chitosan/chemistry*
;
Chondrocytes/cytology*
;
Chondroitin Sulfates/chemistry*
;
Drug Carriers
;
Extracellular Matrix/metabolism*
;
Genetic Therapy/methods*
;
Growth Differentiation Factor 5/genetics*
;
Hyaluronic Acid/chemistry*
;
Microspheres
;
Nanomedicine
;
Osteoarthritis/therapy*
;
Plasmids/metabolism*
;
Rabbits