17β-estradiol inhibits TGF-β-induced collagen gel contraction mediated by human Tenon fibroblasts via Smads and MAPK signaling pathways

AIM:To investigate the impact of 17β-estradiol on the collagen gels contraction(CGC)and inflammation induced by transforming growth factor(TGF)-βin human Tenon fibroblasts(HTFs).METHODS:HTFs were three-dimensionally cultivated in type I collagen-generated gels with or without TGF-β(5 ng/mL),17β-estradiol(12.5 to 100μmol/L),or progesterone(12.5 to 100μmol/L).Then,the collagen gel diameter was determined to assess the contraction,and the development of stress fibers was analyzed using immunofluorescence staining.Immunoblot and gelatin zymography assays were used to analyze matrix metalloproteinases(MMPs)and tissue inhibitors of metalloproteinases(TIMPs)being released into culture supernatants.Enzyme-linked immunosorbent assay(ELISA)and reverse transcription-quantitative polymerase chain reaction(RT-PCR)were used to detect interleukin(IL)-6,monocyte chemoattractant proteins(MCP)-1,and vascular endothelial growth factor(VEGF)in HTFs at the translational and transcriptional levels.The phosphorylation levels of Sma-and Mad-related proteins(Smads),mitogen-activated protein kinases(MAPKs),and protein kinase B(AKT)were measured by immunoblotting.Statistical analysis was performed using either the Tukey-Kramer test or Student’s unpaired t-test to compare the various treatments.RESULTS:The CGC caused by TGF-βin HTFs was significantly inhibited by 17β-estradiol(25 to 100μmol/L),and a statistically significant difference was observed when comparing the normal control group with 17β-estradiol concentrations exceeding 25μmol/L(P<0.05).The suppressive impact of 17β-estradiol became evident 24h after administration and peaked at 72h(P<0.05),whereas progesterone had no impact.Moreover,17β-estradiol attenuated the formation of stress fibers,and the production of MMP-3 and MMP-1 in HTFs stimulated by TGF-β.The expression of MCP-1,IL-6,and VEGF mRNA and protein in HTFs were suppressed by 100μmol/L 17β-estradiol(P<0.01).Additionally,the phosphorylation of Smad2 Smad3,p38,and extracellular signal-regulated kinase(ERK)were downregulated(P<0.01).CONCLUSION:17β-estradiol significantly inhibits the CGC and inflammation caused by TGF-βin HTFs.This inhibition is likely related to the suppression of stress fibers,inhibition of MMPs,and attenuation of Smads and MAPK(ERK and p38)signaling.17β-estradiol may have potential clinical benefits in preventing scar development and inflammation in the conjunctiva.

Overexpression of fibroblast growth factor 13 ameliorates amyloid-β-induced neuronal damage

Previous studies have shown that fibroblast growth factor 13 is downregulated in the brain of both Alzheimer’s disease mouse models and patients,and that it plays a vital role in the learning and memory.However,the underlying mechanisms of fibroblast growth factor 13 in Alzheimer’s disease remain unclear.In this study,we established rat models of Alzheimer’s disease by stereotaxic injection of amyloid-β(Aβ_(1-42))-induced into bilateral hippocampus.We also injected lentivirus containing fibroblast growth factor 13 into bilateral hippocampus to overexpress fibroblast growth factor 13.The expression of fibroblast growth factor 13 was downregulated in the brain of the Alzheimer’s disease model rats.After overexpression of fibroblast growth factor 13,learning and memory abilities of the Alzheimer’s disease model rats were remarkably improved.Fibroblast growth factor 13 overexpression increased brain expression levels of oxidative stress-related markers glutathione,superoxide dismutase,phosphatidylinositol-3-kinase,AKT and glycogen synthase kinase 3β,and anti-apoptotic factor BCL.Furthermore,fibroblast growth factor 13 overexpression decreased the number of apoptotic cells,expression of pro-apoptotic factor BAX,cleaved-caspase 3 and amyloid-βexpression,and levels of tau phosphorylation,malondialdehyde,reactive oxygen species and acetylcholinesterase in the brain of Alzheimer’s disease model rats.The changes were reversed by the phosphatidylinositol-3-kinase inhibitor LY294002.These findings suggest that overexpression of fibroblast growth factor 13 improved neuronal damage in a rat model of Alzheimer’s disease through activation of the phosphatidylinositol-3-kinase/AKT/glycogen synthase kinase 3βsignaling pathway.

RGD多肽水凝胶对Tenon囊成纤维细胞活化和YAP信号的影响

目的 探究不同浓度的精氨酸-甘氨酸-天冬氨酸(RGD)多肽水凝胶,对Tenon囊成纤维细胞(HTF)活化和Yes相关蛋白(YAP)表达的影响。方法 制备0.5%、1.0%、2.0%三种浓度RGD多肽水凝胶,透射电子显微镜观察其内部微观结构,流变学检测弹性模量(E)。构建SD雄性大鼠结膜损伤模型后,分为空白组(不作处理)、低浓度RGD组(结膜下注射0.5%RGD多肽水凝胶)、中浓度RGD组(结膜下注射1.0%RGD多肽水凝胶)、高浓度RGD组(结膜下注射2.0%RGD多肽水凝胶),观察1周后取材,Masson染色观察大鼠结膜处胶原纤维增生情况,免疫组织化学检测大鼠结膜YAP、平滑肌肌动蛋白-α(α-SMA)表达情况。利用转化生长因子-β2(TGF-β2)刺激HTF活化构建瘢痕化细胞模型,按照实验要求分为对照组、 0.5%RGD水凝胶组、1.0%RGD水凝胶组、2.0%RGD水凝胶组、正常组,培养24 h后,Western blot检测HTF中YAP、结缔组织生长因子(CTGF)、α-SMA、纤连蛋白(FN)和I型胶原蛋白(Col I)的蛋白相对表达量;激光共聚焦显微镜观察细胞骨架蛋白(F-actin)和YAP蛋白表达定位。结果 电镜下观察可见,RGD多肽水凝胶内部纳米纤维交联密度随多肽浓度增加而升高,流变学测得0.5%、1.0%及2.0%RGD多肽水凝胶弹性模量(E)依次约为0.067 kPa、0.150 kPa、2.170 kPa。Masson染色结果显示,除中浓度RGD组大鼠结膜胶原纤维增生面积比明显低于空白组外(P<0.05),低浓度组、高浓度组与空白组差异均无统计学意义(均为P>0.05)。免疫组织化学结果显示,与空白组相比,低浓度RGD组、中浓度RGD组、高浓度RGD组大鼠YAP及α-SMA蛋白相对表达量均下降,中浓度RGD组下降最明显,差异均有统计学意义(均为P<0.05)。细胞免疫荧光染色结果显示,与对照组相比,0.5%RGD水凝胶组、1.0%RGD水凝胶组、2.0%RGD水凝胶组HTF的F-actin绿色荧光减弱,YAP红色荧光从细胞核转位于细胞质表达。Western blot检测结果显示,与对照组相比,0.5%RGD水凝胶组、1.0%RGD水凝胶组、2.0%RGD水凝胶组HTF中YAP、CTGF、α-SMA、FN及Col I蛋白相对表达量均下降,1.0%RGD水凝胶组下降最明显,差异均有统计学意义(均为P<0.05)。结论 RGD多肽水凝胶的最佳作用浓度为1.0%,其通过抑制YAP蛋白的表达水平及其核转位,减少HTF的活化及其纤维化蛋白的表达,从而产生抗瘢痕效果。

Interference of Y-27632 on the signal transduction of transforming growth factor beta type 1 in ocular Tenon capsule fibroblasts

AIM: To investigate the interfering effect of Y-27632, a ROCK-I selective inhibitor, on the signal transduction pathway of transforming growth factor-beta 1 (TGF-beta 1) in ocular Tenon capsule fibroblasts (OTFS) in vitro. METHODS: After OTFS from passages 4 to 6 47 vitro were induced by TGF-beta 1 and then treated by Y-27632, the changes of the OTFS cell cycles were analyzed via flow cytometry, and the proteins expression of the alpha -smooth muscular actin (alpha -SMA), connective tissue growth factor (CTGF), collagen I were calculated by Western blot. After OTFS treated by the different concentrations of Y-27632, the expression levels of the alpha -SMA, CTGF and collagen I mRNA were assayed by RT-PCR. RESULTS: Y-27632 had no markedly effect on the OTFS cell cycles. After treated by TGF-beta 1, OTFS in G1 period significantly increased. The cell cycles distribution by both TGF-beta 1 and Y-27632 had no remarkable difference from that in control group. Y-27632 significantly inhibited the proteins expressions of both alpha -SMA and CTGF, while to some extent inhibited that of collagen I. TGF-beta 1 significantly promoted the proteins expressions of alpha -SMA, CTGF and collagen I. After OTFS treated by both TGF-beta 1 and Y-27632, of alpha -SMA, the protein expression was similar with that in control group (P=0.066>0.05), but the protein expression of CTGF or collagen I, respectively, was significantly different from that in control group (P=0.000<0.01). The differences of expressions of the alpha -SMA, CTGF and collagen I mRNA in 30, 150, 750 mu mol/L Y-27632 group were statistically significant, compared with those in control group, respectively (alpha -SMA, P=0.002, 0.000, 0.000; CTGF, P=0.014, 0.002, 0.001; collagen I,P=0.003, 0.002, 0.000). CONCLUSION: Blocking the Rho/ROCK signaling pathway by using of Y-27632 could inhibit the cellular proliferation and the expression of both CTGF and alpha -SMA whatever OTFS induced by TGF-beta 1 or not. Y-27632 suppressed the expression of collagen I mRNA without induction.

Effect of Apigenin on Gap Junctional Intercellular Communication in Human Tenon's Capsule Fibroblasts

Purpose:To investigate the effect of apigenin on gap junctional intercellular communication (GJIC) in human Tenon's capsule fibroblasts (HTFs) and its underlying mechanism. Methods:After a 48 h treatment of cultured HTFs with apigenin.(80 μmol/L),the GJIC was detected by a scrape-loading/dye transfer technique with Lucifer yellow dye and rhodamine (Rh) dextran. The coupling index represents a quantification of GJIC where a high coupling index is associated with a greater number of cells demonstrating cell-cell communication through gap junction channels.The changes in connexin 43 (Cx43) distribution and the expression of Cx43 at the protein and mRNA levels were statistically compared between the two groups by means of immunocytochemistry, western blotting,and real-time polymerase chain reaction (PCR). Results:The functioning of GJIC in the HTFs was significantly enhanced after 48 hours by apigenin treatment when compared with the control cells. In the apigenin group, the intercellular dye transfer grade was above 9, while this value was only grade 3-4 in the control group. The coupling index was significantly increased up to 9.205±0.3621 in the apigenin group,compared with 5.1775 ±0.3177 in the control group (F=279.581, P=0.000). The expression of Cx43 at the protein and mRNA levels was significantly up-regulated in the apigenin group compared with the control group. Conclusion:Apigenin can significantly enhance the function of GJIC in HTFs by up-regulating the expression of Cx43 at both the protein and mRNA levels,suggesting that the enhancement of GJIC in HTFs by apigenin probably acts as an important mechanism underlying the inhibitory effect of apigenin on HTF proliferation.

Triptolide inhibits TGF-β-induced matrix contraction and fibronectin production mediated by human Tenon fibroblasts

AIM： To determine if triptolide influences the contractility and fibronectin production in human Tenon fibroblasts（HTFs）. METHODS： HTFs were cultured in type I collagen gels with or without transforming growth factor beta（TGF-β） and/or triptolide. The diameter of the collagen gel was used to measure contraction. Immunoblot analysis was used to quantify myosin light chain（MLC） phosphorylation and integrin expression. Laser confocal fluorescence microscopy was used to monitor the formation of actin stress fibers. Fibronectin production was measured with an enzyme immunoassay. RESULTS： Triptolide inhibition of contraction in TGF-β-induced collagen gel mediated by HTFs was dosedependent and statistically significant at 3 nmol/L（P〈0.05） and maximal at 30 nmol/L and significantly time dependent at 2 d（P〈0.05）. Triptolide reduced TGF-β-induced expression of integrins α5 and β1, phosphorylation of MLC, and formation of stress fibers in HTFs. Furthermore, the inhibition of triptolide on the attenuated TGF-β-induced production of fibronectin by HTFs was concentration-dependent and significant at 1 nmol/L（P〈0.05） and maximal at 30 nmol/L. CONCLUSION： Triptolide suppress the contractility of HTFs induced by TGF-β and the production of fibronectin by these cells. It is promising that triptolide treatment may possibly inhibit scar formation after glaucoma filtration surgery.

Anti-scarring effects of butaprost on human subconjunctival Tenon's fibroblasts

AIM： To investigate the toxicity of the E-prostanoid 2（EP2） receptor agonist, butaprost against human subconjunctival（Tenon＇s capsule） fibroblasts, and to determine the underlying mechanism. METHODS： We isolated Tenon＇s fibroblasts from the subconjunctival area of healthy subjects and evaluated the types of EP receptors expressed using quantitative realtime reverse transcription polymerase chain reaction（RTPCR）. The toxicity of butaprost against the fibroblasts was evaluated using methyl thiazolyl tetrazolium and lactic dehydrogenase assays. The inhibition of conjunctival fibroblast proliferation by butaprost was assessed by measuring α-actin levels. The underlying mechanism was assessed by measuring intracellular cyclic adenosine monophosphate（c AMP） levels. Intergroup differences were statistically analyzed using an independent t-test. Densitometry of the Western blot bands was performed using the Image J software. RESULTS： Quantitative real-time RT-PCR revealed that the fibroblast EP2 receptor levels were higher than those of the other EP receptors. Butaprost did not show toxicity against Tenon＇s tissue, but inhibited conjunctival fibroblast proliferation by reducing collagen synthesis. EP2 receptor activation enhanced the c AMP cascade, which might be an important mechanism underlying this effect.CONCLUSION： Butaprost effectively reduces the subconjunctival scarring response. Given the significanceof wound healing modulation in blebs, butaprost＇s inhibitory effect on subconjunctival Tenon＇s fibroblasts may be beneficial in managing postoperative scarring in glaucoma surgery.

Inhibitive effect of TAK-242 on Tenon’s capsule fibroblasts proliferation in rat eyes

AIM: To study the inhibition effect of TAK-242 on the proliferation of rat eye Tenon's capsule fibroblasts via the toll-like receptor 4(TLR4) signaling pathway.METHODS: SD rat Tenon's capsule fibroblasts were extracted and cultured, then the cells were divided into normal control group, lipopolysaccharide(LPS) group(10 g/m L LPS) and TAK-242 group(1 μmol/L TAK-242, and 10 μg/m L LPS after 30 min). The expressions of TLR4, transforming growth factor-β1(TGF-β1) and interleukin-6(IL-6) in each group were detected by Western blot and reverse transcriptase-polymerase chain reaction(RT-PCR). Cell proliferation was detected by cell counting kit-8(CCK-8).RESULTS: Double immunofluorescent labeling in the extracted cells showed negative keratin staining and positive vimentin staining. Western blot showed that the LPS group had the highest expression of TLR4 and TGF-β1(P<0.01). Enzyme linked immunosorbent assay(ELISA) also showed that the secretion of IL-6 was the highest in LPS group(P<0.01). But there was no significant difference in TLR4 and TGF-1, as well as IL-6 expressions between the TAK-242 group and the normal control group(P>0.05). RT-PCR showed that the IL-6 m RNA expression in LPS group was the highest in the three groups(P<0.01). CONCLUSION: TAK-242 inhibits the proliferation of LPSinduced Tenon's capsule fibroblasts and the release of inflammatory factors by regulating the TLR4 signalingpathway, providing a new idea for reducing the scarring of the filter passage after glaucoma filtration surgery.

Bifunctional Effect of Human IFN-γ on Cultured Human Fibroblasts from Tenon's Capsule

Purpose:To study the effect of human IFN-γon in vitro cultured human fibroblasts form Tenon's capsule.Materials and methods:The effect of different concentrations of human INF-γand mitomycin-C(MMC),5-fluorouracil(5-Fu）on cultured human Tenon's capsule fibroblasts(HTCF)was measured using a MTT[3-(4,5-dimethylthiazo-2-yl)]-2,5-diphenyltetrazolium bromide;Thiazolyl blue)colorimetric assay,The results were analyzed using ANOVA of the statistical package for social sciences(SPSS)9.0 version.The difference was considered to be significant if P<0.05.Results:The effects of MMC and 5-Fu on the growth of HTCF were negative,while the effects of IFN-γon the growth of HTCF were both negative(10^2-10^4units/ml in two experiments)and positive(10^6,10^5,10units/ml in two experiments).The inhibition rate of MMC ranged from5.73%to46.9%,which was similar to the inhibition rate of 5-Fu ranged from12.49%to38.92%(P=0.351),The inhibition rate of IFN-γin two experiments was smaller than MMC and 5-Fu(P<0.05).Conclusion:IFN-γhas bifunctional effect(both enhancement and inhibition)on proliferation of cultured HTCF,The antiproliferative effect of IFN-γwas weatker than MMCand 5-FU,Further study has to bd carride out to document the inhibition of scar formation of filtration bleb by IFN-γand the molecular mechanisms of its bifunctional effect on HTCF proliferation.Eye Science2000;16:43-47.

TGF-β_1对Tenon’s囊成纤维细胞增殖和结缔组织生长因子表达的影响

目的研究转化生长因子β1(TGFβ1)对Tenon’s囊成纤维细胞增殖和诱导表达结缔组织生长因子(CTGF)的影响。方法用MTT法检测不同质量浓度TGFβ1(0、0.1、1、10ng/mL)对Tenon’s囊成纤维细胞增殖的影响;免疫细胞化学染色法检测不同质量浓度TGFβ1(0、0.1、1、10ng/mL)对Tenon’s囊成纤维细胞诱导表达CTGF的影响。结果GFβ1能够剂量依赖性地促进Tenon’s囊成纤维细胞增殖(P<0.05);TGFβ1能够促进CTGF的表达,有随质量浓度的增加而增强的趋势(P<0.05),但1ng/mL和10ng/mLTGFβ1组之间无统计学差异(P>0.05)。结论GFβ1能够促进Tenon’s囊成纤维细胞增殖,并促进其下游介质CTGF的表达,可能是青光眼滤过术后滤过泡瘢痕形成的重要机制。

HSV-tk/GCV系统对人Tenon囊成纤维细胞的旁观者效应及其机制研究

目的观察HSV-tk/GCV系统对人Tenon囊成纤维细胞(HTFs)的旁观者效应并研究其作用机制。方法将转HSV-tk基因HTFs(HTFs-tk)与正常HTFs于两种接种密度下以不同比例混合,加入5μg/m l更昔洛韦作用5 d,MTT法检测对HTFs的杀伤作用;染料传输法观察HTFs细胞间隙连接细胞间通讯(G JIC)的能力;免疫细胞化学检测Cx43分布;RT-PCR及W estern b lot检测Cx43表达。结果旁观者效应随HTFs-tk/HTFs比例增加而增强,高细胞接种密度组明显强于低密度组;染料通过G JIC传输6级以上;Cx43主要分布于HTFs细胞膜并检测到其mRNA与蛋白表达。结论旁观者效应需要细胞间密切接触,Cx43介导G JIC可能是HSV-TK/GCV系统对HTFs产生旁观者效应的主要机制,HTFs表达Cx43可能在结膜伤口愈合反应中发挥重要作用。

人Tenon's囊成纤维细胞的离体培养及生长特性观察

目的离体培养人Tenon′s囊成纤维细胞,观察其生长特性。方法将人Tenon′s囊成纤维细胞进行离体培养及生长抑制实验,采用含10%胎牛血清的DMEM培养基,细胞计数法及MTT法描述细胞生长曲线。并通过免疫组织化学法对细胞进行鉴定。结果人Tenon′s囊成纤维细胞离体培养48 h至7 d处于对数生长期。细胞角蛋白染色阴性,波蛋白染色阳性。结论人Tenon′s囊成纤维细胞在体外易于生长,是细胞离体实验研究的良好材料。

环孢素A对人Tenon’s囊成纤维细胞增殖及TGF-β_1分泌的影响

目的 观察环孢素A(CsA)对体外培养的人Tenon’s囊成纤维细胞 (HTFs)增殖及其转化生长因子 β1 (TGF β1 )分泌的影响。方法 以体外培养的人HTFs为实验对象 ,分别加入 0 0 0 1～ 10 μg/mLCsA作用 48h ,细胞计数观察CsA对细胞增殖的影响。收集 0 0 0 1～ 5 μg/mLCsA处理后细胞培养上清液 ,采用ELISA方法检测细胞TGF β1 的分泌量。光镜及透射电镜观察细胞的结构变化。结果 CsA在 0 1～ 10 μg/mL范围内以质量浓度依赖方式抑制人HTFs的增殖 ,IC50 为0 5 5 μg/mL ,并在 0 0 1～ 5 μg/mL范围内刺激细胞TGF β1 的分泌。CsA作用后细胞形态学改变的主要特征为胞浆内大量空泡形成。结论 一定质量浓度的CsA可能通过直接的细胞损伤作用在促进TGF β1 分泌的同时仍显著抑制人HTFs的增殖。

雷帕霉素对培养的人Tenon′s囊成纤维细胞的作用

目的:观察雷帕霉素(rapamycin)对培养的人Tenon′s囊成纤维细胞的增殖及细胞周期的作用。方法:将培养的人Tenon′s囊成纤维细胞用含不同雷帕霉素浓度的培养液培养24 h后,用MTT法测定细胞的增殖情况,用流式细胞仪测定细胞周期的变化情况。结果:用含10、20、40、80和160 ng/ml雷帕霉素的培养液培养细胞时,细胞增殖抑制率分别为9.32%,22.55%,41.42%,42.25%,43.57%。细胞周期分析显示,雷帕霉素(40 ng/ml)作用后,G0/G1期细胞较对照组增多(41.76%vs 44.19%,P<0.05),S期细胞较对照组减少(45.42%vs 40.18%,P<0.05)。结论:雷帕霉素具有抑制人Tenon′s囊成纤维细胞增殖的效果,且随浓度增加抑制作用增强,并将细胞抑制在G1期的限制点内。

重组γ-干扰素作用人Tenons囊成纤维细胞体外研究

目的 体外研究人重组γ－干扰素对人Ｔｅｎｏｎｓ囊成纤维细胞作用。方法 以人Ｔｅｎｏｎｓ囊成纤维细胞作为实验对象，分别加入０．１～１０＾６ Ｕ／ｍｌ人重组γ－干扰素（ＩＦＮ－γ）、０．００１～１０ｍｇ／Ｌ丝裂霉素（ＭＭＣ）、０．１～１０３ｍｇ／Ｌ５－氟尿嘧啶（５－Ｆｕ）孵育４８ｈ，然后以ＭＴＴ法进行检测。结果 高浓度和低浓度ＩＦＮ－γ实验组ＯＤ值较对照组升高（Ｐ〈０．０５）。

羊膜、结膜及Tenon's膜移植在兔结膜重建中的实验研究

目的比较羊膜移植(amniotic membrane transplantation,AMT)、自体结膜移植(autologous conjunctivaltransplantation,ACF)、自体Tenon’s膜移植(autologous Tenon’s fasia transplantation,ATFT)在结膜重建中的作用。方法新西兰大白兔24只随机分组:对照8眼不作任何处理,滴bFGF眼药水8眼;AMT组16眼,ACT组8眼,ATFT组8眼。于第2,3,4,6周时杀死兔各2只,HE及透射电镜观察。结果 AMT组3w时结膜上皮移行至羊膜表面,6w时再生上皮的超微结构接近正常,但杯状细胞较正常少。ACT组从一开始就保持了上皮的完整性且杯状细胞丰富。ATFT组上皮再生与AMT组接近但有轻度结膜下纤维化。结论 AMT可有效地修复结膜表面,抑制炎症和瘢痕化。ATFT可改善眼表的基质和血供。ACT是修复结膜缺损的最有效方法。

特异性转化生长因子-β_2 shRNA对人眼Tenon囊成纤维细胞的抑制作用

目的观察转化生长因子-β2(TGF-β2)特异性短发夹RNA(shRNA)对人眼Tenon囊成纤维细胞(TCFs)TGF-β2表达的影响及对人TCFs增生的抑制作用。方法根据小干扰RNA(siRNA)的设计原则,针对TGF-β2基因序列特征构建特异性shRNA载体,转染培养的人TCFs。MTT比色法检测细胞的增生情况;ELISA法检测shRNA对TGF-β2蛋白表达的抑制作用。结果 TGF-β2shRNA转染组的细胞增生及TGF-β2蛋白的表达均受到抑制。转染24、48、72h后MTT比色法检测显示,TGF-β2shRNA转染组培养的Tenon囊的成纤维细胞的增生值分别为0.5426±0.05、0.5210±0.03、0.4055±0.04,明显低于阴性对照组的0.5655±0.03、0.5378±0.03、0.5268±0.04,3个组和各时间点的总体差异均有统计学意义(F组=54.691,P=0.000;Ftime=66.888,P=0.000)。ELISA法检测显示,TGF-β2shRNA转染组TGF-β2蛋白的表达水平下降,24、48、72hTGF-β2蛋白的表达量分别为(543.58±25.32)、(400.26±30.74)、(202.72±23.24)ng/L,明显低于阴性对照组的(659.78±20.95)、(547.45±24.65)、(442.87±17.43)ng/L及空白组的(721.75±30.19)、(756.61±30.19)、(787.60±18.37)ng/L,3个组和各时间点的总体差异均有统计学意义(F组=163.082,P=0.000;F时间=31.498,P=0.000)。结论 TGF-β2特异性shRNA能显著抑制人TCFsTGF-β2的表达,载体介导的TGF-β2特异性shRNA对眼前节术后的抗瘢痕化治疗具有潜在的可能性。

TGF-β1对Tenon囊成纤维细胞增生和HSP47表达的影响

目的探讨转化生长因子β1(TGF-β1)对人Tenon囊成纤维细胞(HTF)增生和热休克蛋白47(HSP47)表达的影响。方法青光眼患者Tenon囊组织体外培养,用不同质量浓度0.01、0.1、1、5、10ng/mLTGF-β1处理细胞,继续培养24h,用MTT比色法检测吸光度值(A),用免疫细胞化学检测HSP47和Ⅰ型胶原纤维蛋白含量,用实时荧光定量PCR检测HSP47和Ⅰ型胶原mRNA表达。结果TGF-β1能促进HTF的增生(P<0.05),诱导HTF表达HSP47和Ⅰ型胶原纤维蛋白(P<0.05),促进HTF表达HSP47和Ⅰ型胶原纤维mRNA(P<0.05),其作用随TGF-β1质量浓度的增加而增强。结论TGF-β1通过促进HTF纤维增生,诱导下游介质HSP47和Ⅰ型胶原蛋白表达而促进HTF纤维化,可能是青光眼滤过手术后瘢痕形成的重要机制。

保留Tenon’s囊的翼状胬肉切除联合羊膜移植术的临床初步研究

目的：探讨一种治疗初发性翼状胬肉的新的手术方法，防止初发性翼状胬肉术后复发的最佳手术方法。方法：24例（24眼）初发性翼状胬肉，在手术显微镜下进行手术，保留翼状胬肉表面的球结膜及巩膜表面的Tenon’s囊，在自然解剖状态下彻底分离切除球结膜下的翼状胬肉组织，同时行羊膜移植。术后随访14-35个月，平均（17.21±3.45）月。结果：24例（24眼）均治愈，均无复发。术中术后未见并发症。结论：手术显微镜下行保留Tenon’s囊的翼状胬肉切除联合球结膜下羊膜移植具有手术损伤小，最大限度保留了正常眼表面的解剖结构，术后复发率低，不影响白内障、青光眼的手术的优势，是治疗初发性翼状胬肉的一种新的有效方法。