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Objective:To investigate the relationship between single nucleotide polymorphisms of transforming growth factor-β2 (TGFβ2) gene and Keshan disease (KD) in Han population of Shaanxi Province.Methods:KD region in Huangling County, Yan'an City, Shaanxi Province was selected as the investigation site in this study. Using the method of cluster random sampling, 52 families with KD in 6 administrative villages in Huangling County (Duanjiawan Village, Taoqu Village, Yaoping Village, Jianzhuang Village, Anjiao Village in Yaoping Town, and Houziping Village in Diantou Town) were selected for epidemiological investigation. According to the "Diagnosis of Keshan Disease" (WS/T 210-2011), 285 subjects were identified, including 79 patients with KD (case group) and 206 healthy controls (control group). Genomic DNA was extracted from the peripheral venous blood. The polymorphism of genetic variation of TGFβ2 gene rs6658835 was genotyped by matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF). Chi-square (χ 2) test and t-test were used to analyze the baseline data, and binary logistic regression model was used to analyze the influencing factors of KD, all samples were tested for Hardy-Weinberg equilibrium using goodness-of-fit χ 2 test, differences in genotype and allele frequencies between case and control groups were compared by χ 2 test, and logistic regression analysis was used to compare the genotype frequencies between two groups after adjusting for confounding factors. Results:Epidemiological investigation showed that there were significant differences in age and heart murmur between case group and control group ( t = 7.03, χ 2 = 9.66, P < 0.05). The analysis of binary logistic regression model showed that the influence of age on KD was statistically significant (χ 2 = 20.72, P < 0.001). The gene frequency distribution of TGFβ2 gene rs6658835 in case group and control group conformed to the Hardy-Weinberg equilibrium (χ 2 = 0.02, P = 0.900). Correlation analysis results: the difference of genotype frequency of TGFβ2 gene rs6658835 in case group (GG, GA, AA: 6.3%, 38.0%, 55.7%) and control group (GG, GA, AA: 10.7%, 43.7%, 45.6%) was not statistically significant (χ 2 = 2.78, P = 0.249). After adjustment by age, the difference of genotype frequency and dominant model of TGFβ2 gene rs6658835 in case group and control group was statistically significant (χ 2adj = 5.43, 4.86, P < 0.05), the difference of recessive model of TGFβ2 gene rs6658835 in case group and control group was not statistically significant (χ 2adj = 2.12, P = 0.145). Conclusion:TGFβ2 gene rs6658835 is associated with KD in Han population of Shaanxi Province.
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Objective:To investigate the relationship between the expression level of long non-coding RNA transforming growth factor β2-antisense RNA1 (lncRNA TGFB2-AS1) and placental spiral artery recasting in the placenta of preeclampsia.Methods:A total of 108 pregnant women with severe preeclampsia who were hospitalized in Zaozhuang Maternal and Child Health Hospital and delivered by cesarean section from Oct. 2019 to Jun. 2021 were selected as the research objects, and they were divided into the late-onset preeclampsia group (late-onset severe preeclampsia pregnant women, 56 cases) and early-onset preeclampsia group (early-onset severe preeclampsia pregnant women, 52 cases) ; at the same time, 58 normal pregnant women were selected as the normal pregnancy group. The general data of pregnant women were collected, such as age, systolic blood pressure and diastolic blood pressure. Real-time fluorescent quantitative PCR (qRT-PCR) method was used to detect the expression level of lncRNA TGFB2-AS1 in placental tissues, a scanning electron microscope was used to measure the lumen area and wall thickness of spiral arteries. Pearson correlation analysis method was used to analyze the correlation between the level of lncRNA TGFB2-AS1 in the placenta tissue and the thickness of the spiral artery wall and the area of the lumen of pregnant women with early-onset and late-onset severe preeclampsia.Results:The tube wall thickness [ (119.69±8.31) μm], systolic blood pressure [ (162.86±4.94) mmHg], diastolic blood pressure [ (103.09±2.35) mmHg], and 24-hour urine protein [ (2.17±0.31) g/24 h] in the early preeclampsia group were higher than those in the late preeclampsia group [ (101.04±5.78) μm, (146.95±6.43) mmHg, (92.13±4.74) mmHg, (1.62±0.23) g/24 h] and the normal pregnancy group [ (99.82±5.56) μm, (116.42±9.31) mmHg, (74.25±6.74) mmHg, (0.06±0.02) g/24 h], the placental tissue lncRNA TGFB2-AS1 level (0.62±0.16), lumen area [ (133.74±20.16) μm 2], gestational week of delivery [ (32.15±1.74) weeks], weight of the newborns [ (2.25±0.26) g] were lower than those in the late-onset preeclampsia group [ (0.99±0.21), (185.49±22.75) μm 2, (36.14±1.59) weeks, (3.37±0.32) g] and the normal pregnancy group [ (1.02±0.23), (186.42±23.71) μm 2, (38.19±1.56) weeks, (3.42±0.37) g] ( P<0.05). The systolic blood pressure, diastolic blood pressure, and 24-hour urine protein in the late preeclampsia group were higher than those in the normal pregnancy group, gestational week of delivery was lower than the normal pregnancy group ( P<0.05). Placental tissue lncRNA TGFB2-AS1 of pregnant women with early-onset severe preeclampsia was positively correlated with the lumen area, and negatively correlated with the thickness of the tube wall ( P<0.05). There was no correlation between lncRNA TGFB2-AS1 and the lumen area and wall thickness in the placental tissue of pregnant women with late-onset severe preeclampsia ( P>0.05) . Conclusion:The lncRNA TGFB2-AS1 expression in the placenta tissue of pregnant women with early-onset severe preeclampsia is abnormally low, which may be related to the insufficient recasting of the placental spiral artery.
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Objective To investigate the role and mechanism of transforming growth factor-β2 (TGF-β2) on regulating the phenotype transformation of vascular smooth muscle cells (SMC) during the wall remodeling after venous thrombosis. Methods Human umbilical vein smooth muscle cells (HVSMCs) were treated with different concentration (0, 1, 10, 20 ng/ml) of TGF-β2 for 24 hours, or treated with 10 ng/ml TGF-β2 at different time points (0, 2, 6, 24 h). RT-PCR was performed to detect the mRNA expressions of phenotype markers (contractile type: α-SMA and Elastin; synthetic type: Col1A1 and Col1A2). SD rat models of inferior vena cava thrombosis (IVCT) were established. Rats were randomly divided into sham surgery group (n=5), thrombus group (n=9) and TGF-β2 experimental group (n=5). The successful establishment of rat model was confirmed by HE and Masson staining of vein wall tissues on 4 and 7 days after modelling. At the 14th day after modelling, IVCT rats were locally treated with hydrogel containing TGF-β2 10 ng per rat for 24 hours. RT-PCR was used to detect the expression of SMC phenotypic marker mRNA in the vein wall of each group. Results TGF-β2 up-regulated mRNA expressions of both contractile and synthetic markers in HVSMCs in a dose- and time-dependent manner (P<0.05). Notely, the mRNA expression of contractile markers was more up-regulated than that of synthetic markers. HE and Masson staining showed the acute thrombosis phase 4 days after modelling and the chronic organized thrombus and recanalization 7 days after modelling, confirming the successful establishment of IVCT models. At the 14th day after modelling, compared with the sham surgery group, the mRNA expression in SMC venous wall contractile markers of thrombus group was down-regulated obviously, and was up-regulated markedly in synthetic markers (P<0.05). When TGF-β2 was used to locally treat the thrombus, the expressions of contractile markers mRNA were more significantly up-regulated (P<0.05). Conclusions TGF-β2 may induce the expression of contractile and synthetic markers in HVSMCs, particularly up-regulating the contractile markers. In IVCT rats model, TGF-β2 can significantly up-regulate the mRNA expressions of SMC contraction markers, which is in favour of maintenance of SMC contractile phenotype in the thrombus vein wall.
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Objective: To determine the effects of interleukin 2 (IL-2) on epithelial-mesenchymal transition (EMT) and extracellular matrix (ECM) synthesis in retinal pigment epithelial (RPE) cells. Methods: IL-2 of 10 μg/L was used to induce RPE cells. Real-time PCR and Western blot methods were used to detect the EMT marker α-smooth muscle actin (α-SMA), the extracellular matrix (ECM) markers fibronectin (Fn) and type I collagen (COL-1), transforming growth factor β2 (TGF-β2), and the activation of the JAK/STAT3 signaling pathway at corresponding time points. Furthermore, JAK/STAT3 signaling pathway was specifically blocked by WP1066, and the changes in α-SMA, COL-1, Fn and TGF-β2 mRNA and protein expressions were detected. Results: After induction by 10 μg/L of IL-2, the expressions of Fn, COL-1, TGF-β2 mRNA and protein as well as p-STAT3/STAT3 were significantly increased (P<0.05). This effect was correlated with the length of IL-2 treatment, while α-SMA mRNA and protein expression did not change significantly. JAK/STAT3 inhibitor WP1066 could effectively inhibit the expressions of Fn, COL-1 and TGF-β2 in IL-2-induced RPE cells. Conclusion: IL-2 promotes ECM synthesis and TGF-β2 expression in RPE cells via JAK/STAT3 signaling pathway, which may play an important role in proliferative vitreoretinopathy.
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Objective • To establish the transforming growth factor-β2 (TGF-β2) induced epithelial-mesenchymal transition (EMT) model of retinal pigment epithelium cells, and investigate the effect and mechanism of lutein on EMT. Methods • ARPE-19 cells were cultured and divided into 4 groups including control group, TGF-β2 group, TGF-β2+lutein group and lutein group. The mRNA levels of α-smooth muscle actin (α-SMA), fibronectin (FN) and E-cadherin were analyzed by real-time PCR. The protein expression of α-SMA, FN and occludin were assayed by Western blotting. Immunofluorescence was used to detect the change of α-SMA. Meanwhile, Western blotting was performed to detect the expression levels of pSmad3 in the TGF/Smad signaling pathway. Results • TGF-β2 induced EMT was inhibited by lutein. Lutein decreased the mRNA and protein levels of the mesenchymal markers α-SMA and FN, and increased the expression of the epithelial markers E-cadherin and occludin (all P<0.05). Immunofluorescence showed that lutein can inhibit the conversion of epithelial cells into myofibroblasts. Lutein significantly downregulated the high expression of pSmad3 in TGF-β2 treated ARPE-19 cells (P=0.001). Conclusion • Lutein inhibits TGF-β2 induced EMT by downregulating the expression of pSmad3 in TGF-β/Smad signaling pathway, indicating it may attenuate subretinal fibrosis.
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Background@#Microparticles (MPs) are small extracellular plasma membrane particles shed by activated and apoptotic cells, which are involved in the development of atherosclerosis. Our previous study found that microRNA (miR)-19b encapsulated within endothelial MPs (EMPs) may contribute to the upregulation of circulating miR-19b in unstable angina patients. Hypoxia is involved in atherosclerosis as a critical pathological stimulus. However, it still remains unclear whether the increase of miR-19b levels in EMPs is related to hypoxia and if the effect of miR-19b - wrapped within EMPs - stimulates hypoxia on vascular endothelial cells. This study aimed to explore the changes of miR-19b in EMPs induced by hypoxia as well as their effects on endothelial cells.@*Methods@#Human umbilical vein endothelial cells (HUVECs) were cultured in vitro and arranged to harvest EMPs in two parts: the first part consisted of EMP and EMP and the second part included EMP, EMP, and EMP. Cell migration was detected by scratch migration and transwell chamber migration. Angiogenesis was assessed by tube formation assays. Furthermore, we predicted the target gene of miR-19b by bioinformatics analysis, and luciferase assay was used to verify the targeted gene of miR-19b. Data were analyzed by one-way analysis of variance. Student's t-test was used when two groups were compared.@*Results@#Compared with EMP- and EMP-inhibited migration of cells by scratch migration assay (80.77 ± 1.10 vs. 28.37 ± 1.40, P < 0. 001) and transwell chamber migration assay (83.00 ± 3.46 vs. 235.00 ± 16.52, P < 0.01), the number of tube formations was markedly reduced by 70% in the EMP group (P < 0.001) in vitro analysis of HUVECs. Meanwhile, a strong inhibition of migration and tube formation of HUVECs in the presence of miR-19b-enriched EMP was observed. This effect might be due to the delivery of miR-19b in EMPs. Transforming growth factor-β2 (TGFβ2) was predicted to be one of the target genes of miR-19b, and we further confirmed that TGFβ2 was a direct target gene of miR-19b using the luciferase assay. The expression of TGFβ2 in HUVECs was inhibited by treatment with EMP and EMP.@*Conclusions@#MiR-19b in EMPs induced by hypoxia could reduce endothelial cell migration and angiogenesis by downregulating TGFβ2 expression, which may have inhibited the progression of atherosclerosis.
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Humans , Cell Hypoxia , Genetics , Physiology , Cell Movement , Genetics , Physiology , Endothelial Cells , Metabolism , Human Umbilical Vein Endothelial Cells , Metabolism , MicroRNAs , Genetics , Metabolism , Neovascularization, Physiologic , Genetics , Physiology , Transforming Growth Factor beta2 , Genetics , MetabolismABSTRACT
·AIM: To investigate the role of small interference RNA interference targeted Integrin-linked kinase (ILK SiRNA) on the proliferation and apoptosis of human Tenon fibroblasts (HTFs) induced by transforming growth factor-β2(TGF-β2). · METHODS: The HTFs were identified by immunofluorescence analysis with Vimentin and keratin. HTFs with no other addiction was as normal control;H+T group:HTFs+5μ g/L TGF-β2;H+T+NC SiRNA group:HTFs+5μ g/L TGF-β2+50nmol/L negtive SiRNA; H+T+ILK SiRNA group:HTFs+5μ g/L TGF-β2+50nmol/L ILK SiRNA. The ILK SiRNA were transfected into HTFs by lipofectamine 2000, then the cells were stimulated with 5μ g/L TGF - β2. The protein expression of ILK were analyzed by Western Blot. The proliferation levels of HTFs were analyzed by CCK-8, the apoptosis of HTFs were analyzed by Hoechst 33342/PI double staining. ·RESULTS: The protein ILK were expressed in both TGF-β2treated and control groups, and TGF- β2up-regulated the expression of ILK, ILK SiRNA inhibited the protein expression of ILK(P< 0. 05). CCK- 8 analysis showed that compared with the normal control group,the cell proliferation rate of HTFs in TGF-β2treated group increased, and in ILK SiRNA group the cell proliferation rate was suppressed after exposured to ILK SiRNA for 48h (P<0.05). Hoechst 33342/PI double staining showed that there was no change on the apoptosis of TGF - β2 stimulated group (P>0.05), compared with the normal control group, however in the ILK SiRNA group, we found lots of apoptosis cells and a few of necrotic cells (P<0.05). ·CONCLUSION: The ILK SiRNA attenuates the abnormal proliferation of HTFs induced by TGF - β2, thereby enhancing the apoptosis of HTFs.
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Posterior capsule opacification (PCO) is a common complication after cataract phacoemulsification with intraocular lens (IOL) implantation.The mechanism of lens epithelial cells (LECs) fibrosis,PCO and capsular wrinkle is mainly related with residual LECs migration,proliferation and epithelialmesenchymal transition (EMT) after cataract surgery.Transforming growth factor β2 (TGF-β2) has been proposed as the most important factor driving the EMT and pathologic fibrosis of LECs,TGF-β2 induces LECs EMT by the Smad signaling pathway.Besides,PI3K/AKT/mTOR signaling pathway was studied to participate in TGF-β2 induced EMT.As one of the gene regulation methods,RNA interference (RNAi) technology shows an important application prospect in inhibiting LECs fibrosis and proliferation by interfering signaling pathway.This review highlights RNAi effects on LECs biological behavior by interfering PI3K/AKT/mTOR and TGF-β2/Smad signaling pathways.
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Objective To study the expression and significance of CD68 and TGF-β2 in benign prostatic hyperplasia(BPH).Methods The immunohistochemistry PV two step method was used to detect the expression level of CD68 and TGF-β2 in 90 cases of benign prostatic hyperplasia and combined with clinical data were analyzed.Results Positive rate of CD68 as macrophages marker was 86%(77/90) in the epithelial cells,positive rate of TGF-β2 as transforming growth factor marker was 79%(71/90) in epithelial cells and stromal cells. Immune inlfammation mediated by macrophages,with coloring degree deepening,the degree of immune inlfammation increased,larger prostate volume(P<0.01),IPSS score higher (P<0.01),maximum urinary lfow rate lower(P<0.01),the dffierences between groups were statistically significant.Conclusions This point may indicate immune inlfammation play an important role in the development process of BPH and help to complete the pathogenisis theory.
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Background Insulin can promote the occurrence of myopia.It has been proven that insulin receptor exists in human retinal pigment epithelial (RPE) cells and can promote RPE cells to secrete transforming growth factor-β2(TGF-β2),which is one of the most important myopic signal molecules.Objective This study was to investigate if PI3K/Akt mediates the promotive effects of insulin on proliferation of human RPE cells and secretion of TGF-β2.Methods Human RPE cell line,ARPE-19 cells,were regularly cultured using DMEM containing 10％ fetal bovine serum,and 10× 103 U/ml insulin,LY294002,10× 103 U/ml insulin+LY294002,Wortmanin,10× 103 U/ml insulin+Wortmanin were added into the medium respectively for 48 hours,and the regularly cultured cells served as blank controls.The proliferation value (absorbance,A) of the cells was evaluated by MTS,and the TGF-β2 level in the cell supernatant was detected by ELISA.The relative expression of TGF-β2 mRNA in the cells was assayed using reverse transcription PCR (RT-PCT) 1 hour and 2 hours after the addition of reagents.Results MTS showed that the proliferation value of the cells in the insulin+LY294002 group was 0.75±0.03,which was significantly lower than that in the insulin group (0.98± 0.04).No significant difference was seen in the proliferative value between the insulin+Wortmanin group and the insulin group (0.97±0.07 versus 0.98± 0.04,P＞0.05).ELISA revealed that the content of TGF-β2 in the the cell supernatant was (11.59±2.85) pg/ml and (49.16± 10.94) pg/ml in the insulin + LY294002 group and the insulin + Wortmanin group,respectively,showing a significant decline in comparison with (548.50±35.18) pg/ml in the insulin group (both at P＜0.05).A significant difference was found in the TGF-β2 content between the insulin+LY294002 group and the insulin+Wortmanin group (t =8.131,P =0.000).The RT-PCR showed that 1 hour and 2 hours after addition of the reagents,the expression levels of TGF-β2 mRNA in the cells were lower in both insulin+LY294002 group and insulin+Wortmanin group than those in the insulin group (P＜0.05).The decline range of TGF-β2 mRNA expression level was more significant in the insulin+LY294002 group than that in the insulin+Wortmanin group at 1 hour (t=4.176,P=0.014) rather than at 2 hours (t=0.756,P=0.492).Conclusions Insulin can promote the proliferation of human RPE cells and secretion of TGF-β2 through PI3K/Akt pathway.This may be one of the mechanisms of insulin causes myopia.
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Background Protein kinase B (Akt) is the center of multiple cellular signaling pathways,and it participates in the regulation of cell function,such as proliferation,migration,and metabolism of cells.Phosphoinositide 3-kinase (PI3K) promotes activation of Akt and therefore triggers many signal pathways.PI3K inhibitor can silent Akt,but whether it can affect the biological behavior of lens epithelial cells (LECs) during the posterior capsular opacity (PCO) is worthy of investigation.Objective This study was to explore the effect of LY294002,a PI3K inhibitor,on Akt activation in human LECs.Methods Human LECs strain,HLEC-B3 cells,were cultured and passaged.The cells were incubated to 96-well plate for 24 hours,then LY294002 was added with the final concentration at 0,10,20,30,40,50,60,70,80 μmol/L,respectively.After incubation for 24 hours,cell counting kit-8 (CCK-8) was used to detect the inhibitory rate of cell proliferation.Transforming growth factor-β2 (TGF-β2) of 10 μg/L was added in the medium of cells as TGF-β2 group,TGF-β2 + LY294002 (20 μmol/L) was used to coculture the other group of cells,and the cells without TGF-β2 and LY294002 served as the control group.Phosphorylation of Akt (p-Akt) in the cells was detected by laser scan confocal immunofluorescence microscope.The expression level of p-Akt was evaluated using Western blot.Results CCK-8 assay showed that the A value of the cells was gradually reduced with the increase of LY294002 concentration (Fgroup =9.72,P =0.00),but the A value was significant raised along with the lapse of time (Ftime =1737.54,P=0.00).Confocal immunofluorescence revealed that a little of p-Akt was expressed in the control cells.After induced by TGF-β2,lots of red fluorescence of p-Akt was seen in cells around the cell membranes.But in the TGF-β2+LY294002 co-culture cells,the fluorescence of Akt was much weaker.Western blot showed that the expression level of p-Akt was 0.91±0.08,1.48±0.13 and 0.95±0.19 in the control group,TGF-β2 group and TGF-β2 +LY294002 group,respectively,with a significant difference among the three groups (F =15.04,P =0.00).Conclusions LY294002 can inhibit the activation of Akt induced by TGF-β2.LY294002 may have utility in the prevention and treatment of PCO.
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Background Epithelial-myofibroblast transition (EMT) of human lens epithelial cells (LECs) induced by transforming growth factor-β2 (TGF-β2) is the main mechanism in the pathogenesis of posterior capsular opacification(PCO).Seeking an effective drug capable of inhibiting this process is important for the prevention and treatment of PCO.Objective The purpose of this study was to investigate the inhibitory effect of rapamycin (RAPA)on the proliferation of human LECs and TGF-β2-induced EMT.Methods Human LEC strain(SRA01/04)was cultured in DMEM with high glucose and 10％ fetal bovine serum.The cells were consequently cultured in serumfree DMEM with 5 mg/L TGF-β2,TGF-β2+10 mg/L RAPA,TGF-β2 + 100 mg/L RAPA,TGF-β2 + 1000 mg/L RAPA or TGF-β2 +10 000 mg/L RAPA for 72 hours,and SRA01/04 cultured in serum-free DMEM were used as control.The proliferation rate(A490)of SRA01/04 in the different groups was detected using the MTT assay and the rate of inhibition of RAPA was calculated.The expressions of the α-smooth muscle actin(α-SMA) and E-cadherin(E-cad)mRNA and protein were detected by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot,respectively.The changes in the expression of α-SMA and E-cad in SRA01/04 were evaluated by Western blot 24,48 and 72 hours after TGF-β2 +400 mg/L RAPA treatment.Results The A490 value of SRA01/04 was 0.680±0.020,0.550±0.013,0.480±0.014,0.400±0.011 and 0.200±0.019 in the control group,TGF-β2 group,TGF-β2 + 10 mg/LRAPA group,TGF-β2 + 100 mg/L RAPA group,TGF-β2 + 1000 mg/L RAPA and TGF-β2 + 10 000 mg/L RAPA group,respectively,showing a gradually declining trend in SRA01/04 rate of proliferation with increasing RAPA concentrations (F =101.920,P =0.000).RT-PCR and Western blot assay showed that the relative expression levels of α-SMA mRNA (α-SMA mRNA/β-actin mRNA)and protein (α-SMA/β-actin)in the cells were significantly increased in the TGF-β2 treatment group.However,with exposure to RAPA,the relative expression levels of α-SMA mRNA and protein were significantly lowered with increasing RAPA concentrations,but the expression levels of E-cad mRNA and protein were raised (α-SMA mRNA:F =294.660,P =0.000 ; α-SMA protein:F =346.950,P =0.000 ; E-cad mRNA:F =264.250,P =0.000 ; E-cad protein:F =317.327,P =0.000).In addition,after exposure to 400 mg/L RAPA,the expression levels of α-SMA protein gradually reduced and those of E-cad protein gradually increased with increasing treatment durations,showing significant differences among the different time points (α-SMA:F =693.864,P =0.000 ;E-cad:F=369.286,P =0.000).Conclusions RAPA can inhibit the proliferation of SRA01/04 in vitro and arrest EMT of SRA01/04 induced by TGF-β2 in a dose-and time-dependent manner.
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Background Research showed that transforming growth factor-β2 (TGF-β2) promotes scar formation.But its mechanism in scarring after glaucoma filtration surgery is worthy of studying.Objective This study was to investigate the effect of TGF-β2 on myofibroblast transition of human Tenon fibroblasts (HTFs) and scarring after glaucoma filtration surgery.Methods Tenon capsular tissue was obtained from 3 patients with strabismus during the surgery and was incubated in DMEM with 10％ fetal bovine serum (FBS).The cells were collected and passaged in the free-serum medium for 24 hours,and then 1,2,5,10,20 μg/L TGF-β2 was added into the medium respectively,to induce the transformation of HTFs,and 2 μg/L or 5 μg/L TGF-β2 was used to treat the HTFs for 6,24,48 and 72 hours.The control group was not treated with TGF-β2.The expressions of α-smooth muscle actin (α-SMA) and phosphorylation of the signaling proteins (pSmad2) in HTFs were detected by Western blot assay.The expressions of α-SMA and F-actin were located by cell immunofluorescine technique under the confocal immunofluorescence microscopy.Cell contractility was determined by collagen gel contraction assays.This study was approved by Ethic Committee of Institute of Surgery Research of Daping Hospital,and informed consent was obtained from each patient or custodian initial of the study.Results The expression of α-SMA protein in the HTFs was increased significantly after the treatment of TGF-β2 in comparison with the control group and reached a peak at 24-48 hours.The α-SMA expression was gradually weakened in the 10 μg/L TGF-β2 groups.Little of α-SMA and F-actin were expressed in the control group.However,strong staining for α-SMA and F-actin were observed in the 1,2 and 5 μg/L TGF-β2 groups and then the staining weakened at the concentration of 10 μg/L.In addition,pSmad2 showed a stronger expression in the 2 μg/L TGF-β2 group than that in the PBS group and FBS group,with the strongest expression in 30 minutes through 2 hours.The untreated gel contracted (78.00±3.13)％ from its initial size,and contraction in the 1,2,5,10 μg/L TGF-β2 group were (63.88±1.78)％,(20.69±0.65)％,(19.49-±0.54)％,(16.24±0.84) ％,respectively,TGF-β2 increased HTFs contraction significantly (Fgroup =859.400,P =0.000).Conclusions TGF-β2 can induce transdifferentiation of Tenon fibroblast into myofibroblast and increase cell contractility,with a concentration-dependent and time-dependent pattern to an extent.It may be the mechanism of scar formation after glaucoma filter surgery.
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-β2 was positively correlated with OPG (r=0. 432,P<0. 01). Conclusions The reference ranges of serum TGF-β1 and TGF-β2 in healthy adult females are established. Both TGF-β1 and TGF-β2 of them are correlated with OPG and ieptin.
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AIM: To detect the gene expression of TGF-β2 in retinas of diabetic rats at different stages, to observe and analyze the effect of TGF-β2 on the retinas of diabetic rats, to explore the role of TGF-β2 in pathogenesis of diabetic retinopathy (DR), and to provide experiment data and experience for further clinic studies.METHODS: Sprague-Dawley (SD) rats were used and retinas were dissected. The total RNA was isolated from which the first strand of cDNA was prepared. Diabetes mellitus was induced by a single intraperitoneal injection of 60mg/kg streptozotocin (STZ) and the rats were held without insulin treatment until sacrifice. Besides, agematched rats treated with saline were used as controls. Tail vein blood glucose was measured after 2 days and rats were considered hyperglycemic if blood glucose reading>16.7mmol/L. Animals with blood glucose level<16.7mmol/L were excluded from the study. The rats were killed at the 4th, 8th, 12th, 16th, 20th and 24th week respectively after hyperglycemic models were established. The retinas were separated and preserved in liquid nitrogen. The expressions of TGF-β2 gene mRNA were detected by reverse transcription PCR (RT-PCR).RESULTS: The RNA of rat retina was integrative enough to be used to further carry out PCR analysis. Compared with control groups, the expression of TGF-β2 mRNA in retinas of diabetic rats was up-regulated at the 4th week, but there was no statistical difference (P>0.05); it was down-regulated at the 8th week, and there was statistical difference (P<0.05); it was also down-regulated at the 12th week, and there was statistical difference (P<0.05); at the 16th week there was no statistical difference (P>0.05); it was up-regulated at the 20th week, but there was no statistical difference (P>0.05); it continued to be up-regulated at the 24th week, and there was statistical difference (P<0.05).CONCLUSION: Since the expression of TGF-β2 mRNA in retinas of diabetic rats was down-regulated at the 8th week and 12th week statistically, up-regulated at the 24th week statistically, it has obviously shown that TGF-β2 was down-and up-regulated through the period of DR. That is, its changes are diphasic with time. It may confirm that TGF-β2, with the characteristic of diphasic regulation, played an important role in DR. It is necessary to study it furthermore.