Hydrophilicity and adsorptivity of a novel scaffold twice embedded by lecithin and poly-L-lysine to human nasal septum chondrocytes / 中国组织工程研究

ABSTRACT

BACKGROUND:Chitosan is a kind of natural biomaterial and is characterized by great biocompatibility, progressive degeneration and absorption and excellent mechanical property; however, whether it may become an ideal cytoskeleton in the engineering of cartilage tissue or not should be researched further.OBJECTIVE: To observe the hydrophilicity and adsorptivity to human nasal septum chondrocytes and the effect of its function of a novel scaffold made by [poly (dl-lactide-co-glycolide)] (PLGA)/chitosan nonwoven cloth embedded with lecithin (LEC) and poly-L-lysine (PLYS).DESIGN: Blank control study.SETTING: Department of Otolaryngology-Head and Neck Surgery, Nanjing General Hospital of Nanjing Military Area Command of Chinese PLA.MATERIALS The experiment was carried out in the Department of Otolaryngology-Head and Neck Surgery, Nanjing General Hospital of Nanjing Military Area Command of Chinese PLA from October 2005 to June 2006. Chitosan nonwoven cloth was provided by Hainan Xinlong Company. The mainly technical parameters were detailed as the follows degree of deacetylation ≥ 90% and relative molecular weight 2-5 × 105. PLGA/chitosan nonwoven cloth scaffold was made in Department of Otolaryngology-Hean and Neck Surgery, Nanjing General Hospital of Nanjing Military Area Command of Chinese PLA and High Polymer Institute of Sun Yat-sen University. The mainly technical parameters were detailed as the follows mole ratio between monome lactide and glycolide 7525, porosity 82%-86%, pore diamater 100-300 μm, shear strength 48 MPa, depth 1.5 mm and completely degenerated duration 14-18 weeks. Human nasal septum chondrocytes were the septal cartilage of nose which was derived from operated patients with deflection of nasal septum under sterile condition.METHODS: PLGA/chitosan nonwoven cloth was sheared into pieces with the size of 1.5 cm × 1.5 cm, dipped in 0.01 volume fraction of LEC anhydrous alcohol and 1 g/L PLYS for 6 hours, dealt with ultraviolet radiation after dehydration for 1hour, dipped in 0.75 volume fraction of ethanol for 24 hours, washed with Hanks solution, and incubated for 24 hours. After operations mentioned above, two kinds of novel scaffolds containing various components were obtained, and they were simple scaffold and scaffold embedded with LEC and PLYS. Cells derived from the third generation of human nasal septum chondrocytes were used to make suspension. In addition, cell suspension was grown on those two scaffolds to determine the degree of hydrophilicity through observing diffused degree of cell suspension. Whether cell suspension of human nasal septum chondrocytes was wafted on scaffolds or not were observed under phase contrast microscope so as to determine adsorptivity between cells and scaffolds; meanwhile, growth of cells and production of matrix were also observed.MAIN OUTCOME MEASURES: ① Hydrophilicity of those two kinds of scaffolds (diffused degree of cell suspension of human nasal septum chondrocytes on scaffold) and adsorptivity to cells (whether cell suspension of human nasal septum chondrocytes was wafted on scaffolds or not); ② growth of cells and production of matrix.RESULTS: ① When simple scaffold was put in cell suspension of human nasal septum chondrocytes, cell suspension showing like balls attached to the surface of scaffold, and then, scattered into space of scaffold gradually. During the period of culture, phase contrast microscope indicated that masses of cells attached to the surface of fiber of scaffold.When the petri dish was shaken, cell groups drifted irregularly. The adherent rate was (21±3.7)%. With the cultured time passing by, matrix was not produced. ② When scaffold embedded with LEC and PLYS was put in cell suspension of human nasal septum chondrocytes, cell suspension scattered into space of scaffold rapidly. The adherent rate was (89±5.6)%, which was higher than that of single scaffold group. This suggested that scaffold showed a strong hydrophilicity.Phase contrast microscope indicated that chondrocytes as the form of monome or community were distributed between scaffold and fiber averagely and attached to the surface of fiber of scaffold. When the petri dish was shaken, cell groups did not drift irregularly, and only a few of cells were scarred at the bottom of petri dish. This suggested that scaffold had a strong adsorptivity to cells. One week after culture, matrix showing like cobweb was produced among fibers of scaffold.With the cultured time passing by, matrix was produced abundantly.CONCLUSION: The novel scaffold of PLGA/chitosan nonwoven cloth embedded with LEC and PLYS is characterized by an excellent hydrophilicity and adsorptivity to human nasal septum chondrocytes; meanwhile, it also can secrete matrix.