ABSTRACT
Objective To investigate the role of methyl-CpG-binding protein 2 (MeCP2) in the regulation and epithelial-mesenchymal transition (EMT) of human lens epithelial cells (LECs) and its possible mechanism.Methods Human LEC lines (SRA01/04) were divided into MeCP2-mimic group,MeCP2-NC group and small interferening RNA-MeCP2 (si-MeCP2) group,and MeCP2 analog plasmid,blank plasmid and MeCP2 si-RNA plasmid was used respectively to transfect the cells.The expression of MeCP2 mRNA in the cells was detected by using real-time PCR 24 hours after transfection.At 48 hours after transfection,the migration rate of the cells was evaluated by scratching test,and the expression of Wnt3a protein in the cells was detected by immunofluorescence stainning.The relative expressions of β-catenin,E-cadherin,Vimentin,matrix metallo proteinase (MMP)-9,MMP-7 and secreted frizzled-related protein 5 (SFRP5) proteins in the cells were detected by Western blot.Results After 24 hours of transfection,the relative expression of MeCP2 mRNA in the cells was significantly different among the MeCP2-mimic group,MeCP2-NC group and si-MeCP2 group (F =4 773.00,P＜0.00 1).The migrating rate of the cells in the MeCP2-mimic group,MeCP2-NC group and si-MeCP2 group was (57.45±5.20)％,(32.71± 10.02)％ and (17.77±9.22)％,respectively,showing a significant difference among the three groups (F=124.00,P＜0.001),and the migrating rate of the cells in the si-MeCP2 group was significantly lower than that of the MeCP2-mimic group or MeCP2-NC group (both at P＜0.001).The relative expressing intensity (absorbance) of Wnt3a in the cells of the MeCP2-mimic group,MeCP2-NC group and si-MeCP2 group was 75.92 ± 6.10,52.03 ± 5.22 and 28.75 ± 3.39,respectively,with a significant difference among three the groups (F=221.30,P＜0.001),and the relative expressing intensity (absorbance) of Wnt3a in the cells was significantly lower in the si-MeCP2-mimic group than that of the MeCP2-NC group and MeCP2-mimic group (both at P＜0.001).The relative expressing level of E-cadherin protein was significantly elevated and the expressions of β-catenin,Vimentin,MMP-9 and MMP-7 were significantly reduced in the si-MeCP2 group compared with the MeCP2-mimnic group and MeCP2-NC group (all at P＜O.01).The relative expressing level of SFRP5 protein in the MeCP2-mimic group,MeCP2-NC group and si-MeCP2 group was 27.19± 0.03,47.54±0.05 and 74.93±0.05,respectively,showing a statistical difference among the three groups (F =183.49,P＜0.001),and the relative expressing level of SFRP5 in the si-MeCP2 group was significantly higher than that in the MeCP2-mimic group and MeCP2-NC group (both at P＜0.001).Conclusions MeCP2C can promote EMT of human LECs by down-regulating the expression of SFRP5 and therefore activating the Wnt3a/β-catenin signal pathway.
ABSTRACT
Objective@#To investigate the role of methyl-CpG-binding protein 2 (MeCP2) in the regulation and epithelial-mesenchymal transition (EMT) of human lens epithelial cells (LECs) and its possible mechanism.@*Methods@#Human LEC lines (SRA01/04) were divided into MeCP2-mimic group, MeCP2-NC group and small interferening RNA-MeCP2 (si-MeCP2) group, and MeCP2 analog plasmid, blank plasmid and MeCP2 si-RNA plasmid was used respectively to transfect the cells.The expression of MeCP2 mRNA in the cells was detected by using real-time PCR 24 hours after transfection.At 48 hours after transfection, the migration rate of the cells was evaluated by scratching test, and the expression of Wnt3a protein in the cells was detected by immunofluorescence stainning.The relative expressions of β-catenin, E-cadherin, Vimentin, matrix metallo proteinase (MMP)-9, MMP-7 and secreted frizzled-related protein 5 (SFRP5) proteins in the cells were detected by Western blot.@*Results@#After 24 hours of transfection, the relative expression of MeCP2 mRNA in the cells was significantly different among the MeCP2-mimic group, MeCP2-NC group and si-MeCP2 group (F=4 773.00, P<0.00 1). The migrating rate of the cells in the MeCP2-mimic group, MeCP2-NC group and si-MeCP2 group was (57.45±5.20)%, (32.71±10.02)% and (17.77±9.22)%, respectively, showing a significant difference among the three groups (F=124.00, P<0.001), and the migrating rate of the cells in the si-MeCP2 group was significantly lower than that of the MeCP2-mimic group or MeCP2-NC group (both at P<0.001). The relative expressing intensity (absorbance) of Wnt3a in the cells of the MeCP2-mimic group, MeCP2-NC group and si-MeCP2 group was 75.92±6.10, 52.03±5.22 and 28.75±3.39, respectively, with a significant difference among three the groups (F=221.30, P<0.001), and the relative expressing intensity (absorbance) of Wnt3a in the cells was significantly lower in the si-MeCP2-mimic group than that of the MeCP2-NC group and MeCP2-mimic group (both at P<0.001). The relative expressing level of E-cadherin protein was significantly elevated and the expressions of β-catenin, Vimentin, MMP-9 and MMP-7 were significantly reduced in the si-MeCP2 group compared with the MeCP2-mimnic group and MeCP2-NC group (all at P<0.01). The relative expressing level of SFRP5 protein in the MeCP2-mimic group, MeCP2-NC group and si-MeCP2 group was 27.19±0.03, 47.54±0.05 and 74.93±0.05, respectively, showing a statistical difference among the three groups (F=183.49, P<0.001), and the relative expressing level of SFRP5 in the si-MeCP2 group was significantly higher than that in the MeCP2-mimic group and MeCP2-NC group (both at P<0.001).@*Conclusions@#MeCP2C can promote EMT of human LECs by down-regulating the expression of SFRP5 and therefore activating the Wnt3a/β-catenin signal pathway.
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Objective To evaluate the relationship between anterior chamber angle and intraocular pressure (IOP) after laser peripheral iridotomy (LPI) treatment.Methods A retrospective cases control study was adopted.Fifty-eight patients (58 eyes) who were diagnosed as primary angle closure (PAC) were included in this study.Ultrasound biomicroscopy (UBM) parameters in angle opening distance (AOD),trabecular iris area (TISA) and angle recess area (ARA) examination were performed before LPI.The changes of intraocular pressure (IOP) were compared between different time-points (before and 1 hour,2 hours,8 hours,24 hours,2 weeks,6 months and 12 months after LPI).The patients were divided into IOP≤21 mmHg group (41 eyes) and IOP＞21 mmHg group (17 eyes) after LPI.Relationship between anterior chamber angle and IOP after LPI treatment was explored.This study was approved by Ethic Committee of the Henan Eye Institute and informed consent was obtained from each patient.Results The IOPs were increased in 1 hour,2 hours after LPI and lowered in 2 weeks,6 months,12 months after LPI compared with IOP before LPI,with significant differences between them (all at P＜0.01).Twelve patients suffered transient elevated IOP and recovered by self-healing or treatment.IOP of 4 patients were elevated after 6 months to 1 year follow-up.The IOPs in 2 weeks,6 months and 12 months after LPI were lowered compared with IOP before LPI,with significant differences between them (all at P＜0.01).The UBM parameters were significantly increased in 2 weeks,6 months,12 months after LPI in comparison with IOP before IPL (all at P＜0.01).IOP and UBM parameters values were significantly different between IOP＞21 mmHg group and ≤21 mmHg group after LPI.Regression analysis indicated that ARA750 (OR =0.75,P＜0.05) was correlated to the IOP after LPI rather than IOP before operation,AOD and TISA (P＞0.05).Conclusions ARA750 value is correlated with the IOP variations after LPI.UBM structured observation can improve the surgical successful rates and safty and prevent complications.
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Background Accurate measurement of corneal thickness is very important during the pre-and post-operative management of corneal surgical procedures,especially laser-assisted in-situ keratomileusis (LASIK),which is the most popular approach to the correction of refractive errors currently.This may be particularly important for the patients who have undergone previous laser refractive surgery with suboptimal outcomes and are being considered for an enhancement procedure.Objective This study was to compare the measuring outcomes of corneal thickness by slit-scanning pachymetry,non-contact specular microscope,anterior segment optical coherence tomography (AS-OCT)and ultrasound pachymetry,with a focus on central and midperipheral (from the central 3.0 mm) region of cornea in post-LASIK eyes.Methods Sixty-four right eyes of 64 patients who received LASIK were collected in Henan Eye Institute,Henan Eye Hospital from March to June 2011 with the equivalent spherical diopter of (-4.75±2.38)D and horizontal corneal diameter of (11.36±0.32)mm.Central corneal thickness was measured on each eye by using non-contact specular microscope (Topcon SP-3000P),slit-scanning pachymetry (Orbscan Ⅱ),AS-OCT and A-type ultrasound pachymetry,respectively,and the paracentral corneal thickness including 12:00,2:00,6:00 and 10:00 meridian was measured using Orbscan Ⅱ,non-contact specular microscope and AS-OCT.The measuring values and the agreement from different instruments were compared and evaluated.Results The mean central corneal thickness was (467.12±31.10)tμm for AS-OCT,(466.67±30.99)μm for ultrasound pachymetry,(441.84 ± 33.65) μm for specular microscopy and (422.51 ± 44.09) μm for Orbsan Ⅱ,respectively,showing a significant difference among the four methods (F =23.730,P =0.000).The central thickness value of the A-type ultrasound pachymetry was significantly higher than that of Orbsan Ⅱ or non-contact specular microscope (q =6.940,6.720,both at P =0.000).Compared with Orbscan Ⅱ,the measuring values of non-contact specular microscope and AS-OCT were significantly higher (q =-5.54,6.940,both at P =0.000),and the measuring value of AS-OCT was significantly higher that of non-contact specular microscope (q =6.800,P =0.000).The lowest difference value (25.3 μm)and the best agreement was found between the ultrasound pachymetry and AS-OCT.The paracentral corneal thickness values in 12:00,2:00,10:00,6:00 meridians were highest for Orbsan Ⅱ and the next for AS-OCT,and non-contact specular microscope had the lowest values,with significantly differences among them (F =5.020,22.950,67.890,18.850,all at P ＜ 0.01).Conclusions The corneal thickness values vary with the different instruments.Orbsan Ⅱ underestimates the central corneal thickness and overestimates the midperipheral corneal thickness,and non-contact specular microscope underestimates both the central and midperipheral corneal thickness.The measuring outcome from AS-OCT has a good agreement with ultrasound pachymetry and therefore they can be used interchangeably.