ABSTRACT
Objeetive To explore the influence of extracellular high glucose on the proliferation,migration and biomarkers of corneal limbal stem cells.Methods Establishment of a model of high glucose in cultured human limbal stem cells to observe and investigate the effects of extracellular high glucose on the proliferation and migration of corneal limbal stem cells by immunoflurescence,CCK-8 and Transwell assay,respectively.Totally 16 SPF rats were collected and induced diabetic model by streptozotocin as the high-glucose group,and the normal rats of the same age served as the control group.Corneal epidermises of rats in both groups were scraped to observe the repair of corneal epithelium.And the corneas were treated with HE staining and immunohistochemical staining to detect the expression of biomarkers of corneal limbal stem cells and the modality changes of cells.Results The proliferation rate of human limbal epithelial cells was significantly decreased when exposed to high glucose,and the rate at 24 h,48 h and 72 h was 0.728,0.345 and 0.395,respectively,which was markedly lower than that in the control group,with a significant difference (P ＜ 0.05);meanwhile the cell migration rate of the high-glucose group was 17.6％ at 48 h,which was significantly slower than that of the control group (100％).And the inhibition was accompanied by the decreased expression of β-catenin and vimentin.Furthermore,the expression levels of β-catenin and vimentin mRNA and protein were down-regulated,with abnormal location,in the high-glucose group.And diabetic rats had poor corneal epithelial healing.The epithelial layer became thinner and the structures were disorganized in diabetic rats through HE staining.The immunohistochemical assay revealed the expression of β-catenin and vimentin of cornea limbal stem cells was down-regulated in high-glocose group when compared with the control group.Conclusion High glucose can significantly inhibit the proliferation and migration of cornea limbal stem cells,and its main damage mechanism is correlated with the abnormalities of β-catenin and vimentin.
ABSTRACT
Objective To screen a kind of polyvinyl alcohol (PVA) hydrogel with similar physical properties to human vitreous body as to provide basis for exploring new vitreous substitutes.Methods Different concentrations (1％,3％,7％) PVA solution were crosslinked by γ-irradiation (Co60),and 1％,3％ and 7％ PVA hydrogels were harvested.Then PVA hydrogels with similar properties to natural vitreous were screened by detecting the physical and optical properties (density,water content,light transmittance,refractive index,pH value,swelling property) and rheological properties and cytotoxicity of PVA hydrogels.Results As for the physical and optical properties of the hydrogels,the water content,refractive index,light transmittance,pH value and density of 1％,3％ and 7％ PVA hydrogel was 98.90％ vs.98.10％ vs.93.50％,1.335 5 vs.1.336 1 vs.1.342 5,94.80％ vs.93.20％ vs.88.20％,7.22 vs.7.25 vs.7.41 and 1003.9 g · L-1d vs.1014.4 g · L-1 vs.1114.7 g · L-1,respectively.Finally,1％ and 3％PVA hydrogels had the similar physical and optical properties to the human vitreous.As for rheological properties,the storage modulus (G') of the three PVA hydrogels was larger than the loss modulus (G"),indicating that these three hydrogels were viscoelastic gel,and the elastic properties was greater than the viscosity.1％ PVA:G'was (3.2 ±1.1)Pa,G" was (0.8 ±0.5)Pa;3％ PVA:G'was (6.1 ± 1.3)Pa,G" was (1.3 ±0.9)Pa;7％ PVA:G'was (106.5 ± 18.6) Pa and G" was (18.3 ± 12.8) Pa.According to resilience,the hydrogels can be ranked as follows:7％ PVA ＞ 3％ PVA ＞ 1％ PVA;in terms of creep,3％ PVA ＞ 7％ PVA ＞ 1％ PVA.Additionally,MTT colorimetric assay showed that there was no significant difference in A values among the three groups by cells culture in 1％,3％ and 7％ PVA for 72 hours (all P ＞0.05),suggesting that all the three hydrogels showed no obvious cytotoxicity and had good biocompatibility.Conelusion 3％ PVA hydrogel has similar optical,physical and rheological properties to the human vitreous,and shows good biocompatibility,which can be one of vitreous substitutes.