| Background:Temporomandibular joint disorders(TMD)is a complex degenerative change in the structural and functional abnormalities of the temporomandibular joint.The pathogenesis of this disease is still unclear,many current treatment methods have certain deficiencies,and some patients may even need to remove the damaged joint disc in the later stage of the disease.In recent years,the development of tissue engineering has provided a new choice for the treatment of temporomandibular joint disorders.The temporomandibular joint disc tissue engineering is mainly for the regenerative treatment of temporomandibular joint diseases that require surgery.The goal is to create new tissue with similar tissue structure and function as well as good biological properties to repair damaged tissue.Bioscaffolds play a key role in supporting cells and/or bioactive molecules to promote damaged tissue repair and/or tissue regeneration,and finding suitable materials to replace and repair damaged tissue has become a current problem to be solved.Our group explored the mechanical properties and biological properties of poly(glycerol-sebacate)(PGS)/ poly-L lactic acid(PLLA)composite nano scaffold.The experimental results showed that PGS/PLLA scaffold has certain application prospects in temporomandibular joint tissue engineering.However,the hydrophilicity of the PGS/PLLA scaffold surface is poor,which is not conducive to cell adhesion.To solve this problem,the subject believes that it is necessary to improve the hydrophilicity of the scaffold surface to facilitate cell adhesion.Objective:The PGS/PLLA scaffold surface was plasma treated to improve its hydrophilicity,and the mechanism of plasma treatment on the surface performance of the scaffold was explored to provide a new method for promoting the application of PGS/PLLA scaffold in temporomandibular joint disc tissue engineering.Method:The PGS/PLLA composite nano scaffolds were prepared by hydrothermal phase separation in a specific mold,and the scaffolds sources were divided into two groups: the pre-treatment and post-treatment ones.Cell origin: In vitro isolation,and culture of 2-3-month-old goat temporomandibular joint disc cells,subcultured to P1 generation.The surface treatment of the stent was carried out by a plasma treatment apparatus under the following conditions: 100 W and 970 S.Performance characterization and biological performance evaluation of PGS/PLLA scaffolds before and after treatment were performed.Scanning electron microscopy(SEM)was used to observe the changes of the surface morphology of the stent before and after plasma treatment;atomic force microscopy(AFM)was used to detect the roughness of the surface of the stent;Fourier transform infrared spectroscopy(FTIR)was used to detect the chemical bonds of the stent before and after treatment;X-ray photoelectron spectroscopy(XPS)is adopted to detect changes in the content of related elements;contact angle analyzer is used to measure the hydrophilicity of the surface of the stent;the disc cells are inoculated on the scaffold,and the adhesion is observed by scanning electron microscopy.The live/dead cell test kit detects the viability of cells on the scaffold;CCK-8 detects the proliferation of articular disc cells on both kinds of scaffolds;RT-PCR measures the expression levels of type I collagen and type II collagen.Results:1.Scanning electron microscope and atomic force microscope showed that the surface of the scaffold before the oxygen plasma treatment was smooth,while the surface of the treated scaffold became uneven,which had pit-like structures,and the surface roughness of the stent significantly increased.The pore size of the stent before treatment was mainly 20 ?m and 30 ?m,but the pore diameter of the stent after treatment was mainly 10 ?m and 20 ?m.And the pore diameter tended to be more uniform.2.Fourier transform infrared spectroscopy and X-ray photoelectron spectroscopy showed no difference in the infrared spectra of PGS/PLLA scaffolds before and after oxygen plasma treatment.The content of oxygen groups in the treated scaffold is higher than that of the untreated scaffold,and the proportion of oxygen element is higher than that of the untreated scaffold.3.The results of the contact angle analyzer: the water contact angle of the stent surface before treatment was 103.7± 10.1°,and the water contact angle of the treated stent surface was 64.7± 12.6°.4.The results of the cell adhesion: under the scanning electron microscope,the adhesion of the articular disc cells with spherical structure on the surface of the untreated scaffold was poor;while the cells(in a pseudopod extended)on the surface of the treated stent adhered well.5.The results of live/dead cell staining: goat articular disc cells were observed for cell viability after 2 and 4 days of culture on PGS/PLLA on both treated scaffolds and untreated scaffolds.Most of the cells showed an active state on both scaffolds,indicating that the two scaffolds have good biocompatibility,no bio toxicity or low toxicity.At the same time,the cell number and cell density of the treated PGS/PLLA scaffolds were superior to the untreated PGS/PLLA scaffolds after 2 and 4 days of culture.6.The results of cell proliferation: Articular disc cells were cultured on PGS/PLLA scaffolds for 1 day,3 days,and 5 days.As the culture time prolonged,the number of cells on the scaffold increased significantly,and the treated scaffold surface had more cells than the untreated one.7.RT-PCR results: After 14 days of culture on both treated scaffolds and untreated scaffolds,gene expression of type I collagen was significantly increased compared with 7 days of culture.And the gene expression level of cells cultured on PGS/PLLA-OH scaffold was higher than that of cells on untreated scaffold.Similarly,the expression level of collagen type II collagen in oxygen plasma-treated scaffold cells was higher than that in untreated scaffolds,while the expression levels of type II collagen in cells cultured on PGS/PLLA and PGS/PLLA-OH scaffolds for 7d and 14 d were not significant differences.Conclusion: The PGS/PLLA composite nano scaffolds before and after oxygen plasma treatment are suitable for the adhesion and growth of goat articular disc cells.The hydrophobicity of the treated scaffolds is improved,which is more conducive to cell adhesion and proliferation. |
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