Objective To determine the association of -429T/C and G1704T polymorphisms in the receptor for advanced glycation end products gene with proliferative diabetic retinopathy (PDR). Methods Case-control study. From the Beijing Desheng Diabetic Eye Study cohort of 1467 patients with type 2 diabetes mellitus (T2DM),atotal of 97 patients with PDR and 105 diabetic patients without retinopathy (DWR, duration of diabetes 15 years) were included for this study. Questionnaires were collected and general ophthalmologic examinations were performed. Biochemical analysis was conducted. DNA was extracted from peripheral venous blood. The -429T/C and G1704T single nucleotide polymorphisms were detected by the means of PCR-restrication fragment length polymorphisms. Results The frequency distribution of -429T/C in DWR group was 81.0% in TT, 16.1% in TC, 2.9% in CC. The frequency distribution of -429T/C in PDR group was 77.3% in TT, 20.6% in TC, 2.1% in CC. There was no significant statistical difference between the two groups (χ2=0.40, P > 0.05). Frequency of the -429T/C minor alleleCin the DWR and PDR group were 11.0% and 12.4%, respectively, with no significant statistical difference between the two groups (χ2=0.20，P > 0.05). The frequency distribution of G1704T in DWR group was 66.7% in GG, 29.5% in GT, 3.8% in TT. The frequency distribution of G1704T in PDR group was 78.4% in GG, 21.6% in GT. There was no significant statistical difference between the two groups (χ2=3.44, P > 0.05). Frequency of the G1704T minor alleleTin the DWR and PDR group were 18.6% and 10.8%, respectively, in which significant difference was found within the two groups (χ2=4.79, OR=1.88，95%CI: 1.06 - 3.33, P > 0.05). Conclusions G1704T polymorphism is associated with PDR presence and 1704G allele may increase the risk of PDR.
【摘要】 目的 觀察晚期糖基化終產物（advanced glycosylation end prodrcts，AGE）對人結腸癌細胞株SW-480增殖的影響，并探討其可能機制。 方法 不同濃度AGE干預SW-480細胞，噻唑藍（MTT）法比較各組細胞活力，流式細胞術觀察AGE對SW-480細胞周期的影響，蛋白質印跡法觀察AGE對SW-480細胞CyclinD1表達的影響，端粒重復序列擴增法（telomeric repeat amplification protocol，TRAP）銀染法觀察AGE對SW-480細胞端粒酶活性的影響。MTT測細胞活力的檢測設置空白對照組、100 μg/mL小牛血清白蛋白（bovine serum albumin，BSA）組及50、100、500 μg/mL AGE組，其余檢測只設置100 μg/mL BSA組和100 μg/mL AGE組。 結果 MTT結果示AGE促進SW-480細胞的增殖，且呈濃度依賴性。100 μg/mL BSA組與100 μg/mL AGE組72 h后的細胞G0/G1期所占百分比分別為56.02%±0.58%、51.93%±1.01%，差異有統計學意義（Plt;0.05）。蛋白質印跡法示100 μg/mL AGE組72 h后CyclinD1的表達較100 μg/mL BSA組增加，差異有統計學意義（Plt;0.05）。TRAP銀染法檢測示100 μg/mL AGE干預SW-480細胞72 h后可以增加端粒酶活性（Plt;0.05）。 結論 AGE可促進人結腸癌細胞SW-480生長，呈劑量依賴性。其作用機制可能與AGE上調CyclinD1的表達加速G1/S期轉換及增加端粒酶活性有關。【Abstract】 Objective To observe the effects of advanced glycosylation end products (AGE) on proliferation of SW-480 cells and study the possible mechanism. Methods Various concentrations of AGE were designed to have impact on SW-480 cells. Proliferation of SW-480 cells was assessed by thiazolyl blue tetrazolium bromide (MTT) assay; The impact of AGE on the cell cycle of SW-480 cells was analyzed by flow cytometry (FCM); the influence of AGE on expression of CyclinD1 was checked by Western blotting; and the impact of AGE on telomerase activity was examined by telomeric repeat amplification proctol (TRAP) sliver staining. For the MTT assay, blank control group, 100 μg/mL bovine serum albumin (BSA) group, 50, 100 and 500 μg/mL AGE groups were designed, while for other examinations, there were only 100 μg/mL BSA group and 100 μg/mL AGE group. Results MTT result showed that AGE increased the proliferation of SW-480 cells in a dose-dependent mode. The proportion of the cells at G0/G1 stage of the 100 μg/mL BSA group and the 100 μg/mL AGE experimental group were (56.02±0.58)% and (51.93±1.01)% respectively after 72 hours, with a significant difference (Plt;0.05); western blotting showed that the expression of CyclinD1 in the 100 μg/mL AGE group was significantly higher than that in the 100 μg/mL BSA group after 72 hours; TRAP silver staining demonstrated that telomerase activity increased significantly after treated with 100 μg/mL AGE for 72 hours. Conclusions AGE can promote the growth of SW-480 cells in a dose-dependent mode. Its mechanism is mainly by up-regulating the expression of CyclinD1 to shorten G0/G1 and increasing the telomerase activity significantly.
ObjectiveTo observe the effect of polypyramidine tract binding protein-associated splicing factor (PSF) towards advanced glycation end products (AGEs) induced the apoptosis of Müller cells in vitro.MethodsExperimental study. Müller cells were cultured and divided into groups according to the project design, plasmid enhanced green fluorescent protein-PSF were transfected into the cells to achieve the overexpression of PSF Müller cells in vitro, then cells were exposed to AGEs and the Morphological changes were observed by HE staining and Hoechst 33258 staining while the survival rate of cells were detected by MTT assay. The effects of PSF on AGEs-induced Müller apoptosis was measured by Cell Death Detection ELISA kit. Meanwhile, 2′,7′-dichlorofluorescin diacetate staining was performed to monitor the protective effects of PSF on AGEs-induced Müller cells ROS.ResultsThe morphology of cells in normal group was full and the cytoplasm staining was uniform. In N+AGEs group and Vec+AGEs group, cell volume decreased, cytoplasm was dense and concentrated, and eosinophilic staining was enhanced. The cell morphology of PSF+AGEs group was still full, with uniform cytoplasm staining and uniform nucleus staining. The viability of N+AGEs group, Vec+AGEs group and PSF+AGEs group were 0.42±0.11, 0.35±0.12 and 0.68±0.12. The apoptosis values were 1.08±0.16, 0.96±0.20 and 0.44±0.08. The intracellular ROS levels were 28 833.67±3 550.06, 28 356.67±4 854.81, 186 163.00±382.54. Compared with N+AGEs group and Vec+AGEs group, the cell viability of PSF+AGEs group was significantly improved (F=20.65, P=0.000), cell apoptosis value (F=43.43, P=0.000) and intracellular ROS level (F=18.86, P=0.000).ConclusionPSF overexpression play a protective role in AGEs-induced apoptosis by inhibiting the production of ROS in Müller cells.
Objective To investigate the effects of advanced glycation endproducts (AGEs) on proliferation of pericytes of bovine retinal capillary vessels and expression of transforming growth factor beta;(TGF-beta;). Methods The proliferation of pericytes detected by methyl thiazolyl tetrazolium (MTT) colorimetric assay, cellular cycle of pericytes was analyzed by flow cytometry was used to analyze cell, and TGF-beta; protein expression of pericytes was observed by immunofluorescent staining. Results AGEs inhibited the proliferation of pericytes of bovine retinal capillary vessels, stopped the cellular cycle of pericytes in synthesis phase (S phase), increased the number of apoptotic cells obviously (Plt;0.01), and promoted the expression of TGF-beta; proteinof perycytes. Conclusions AGEs may promote the apoptosis of pericytes by inhibiting the proliferation of pericytes to lead the decrease of pericytes number, and may accelerate diabetic retinopathy by promoting the expression of TGF-beta; protein of pericytes. (Chin J Ocul Fundus Dis, 2006, 22: 20-23)
Objective To investigate the effects of glycation pro ducts on growth and cytosoic free calcium (［Ca2+］i) of bovine retinal capillary pericytes. Methods The changes of growth and ［Ca2+］i of bovine retinal pericytes,which were cultured in early glycation products of bovine serum albumin (EG-BSA) and advanced glycation end products of bovine serum albumin (AGE-BSA),were studied by counting cell numbers,MTT colorimetric assay,［3H］thymidine incorporating,and fluorescent indicator fura-2 acetoxymeth1 ester (Fura-2AM). Results The number of alive pericytes in groups of EG-BSA and AGE-BSA were 17.87plusmn;2.36 and 14.77plusmn;3.72 which comparing with their control groups (20.54plusmn;0.82 and 20.31plusmn;0.93)were de creased 13．00% and 27．00% (Plt;0.01) by counting cell numbers on a counting plate after four days.The results were 0.4619plusmn;0.0946 and 0.3884plusmn;0.1031 which comparing with their control groups (0.5236plusmn;0.0539 and 0.5227plusmn;0.0519)were decreased 12．00% and 25.70% (Plt;0.01) by MTT colorimetric assay.Amount of ［3H］thymidine incorporating in groups of EG-BSA and AGE-BSA were 39450.16plusmn;887 0.68 and 33667.85plusmn;10581.70 which comparing with their control groups (56373.63plusmn;2317.97 and 56542.04plusmn;1961.23)were decreased 30．00% and 40.40% (Plt;0.01).The ［Ca2+］i concentration of pericytes in groups of EG-BSA and AGE-BSA were (129.55plusmn;30.41) nmol/L and (179.71plusmn;56.69) nmo l/L which comparing with their control groups ［(79.70plusmn;6.94) nmol/L and (83.plusmn;6.39) nmo l/L］ were increased to 163．00% and 214．00%. Conclusion Both EG-BSA and AGE-BSA can inhibit the proliferation and DNA syntheses of retinal capillary pericytes,and increased ［Ca2+］i concentration in pericytes,especially the AGE-BSA. (Chin J Ocul Fundus Dis，2000，16：139-212)
Objective To observe the value of serum soluble receptor of advanced glycation endproducts (sRAGE) combined with lung function and high resolution lung CT (HRCT) in predicting the risk of chronic obstructive pulmonary disease (COPD) developing non-small cell lung cancer (NSCLC). Methods From January 2019 to June 2021, 140 patients with COPD combined with NSCLC, 137 patients with COPD, and 133 patients with NSCLC were enrolled in the study from the People's Hospital of Ningxia Hui Autonomous Region. General data, clinical symptoms, pulmonary function indexes and HRCT emphysema indexes (EI) were collected. Serum sRAGE levels of these patients were measured by enzyme linked immunosorbent assay. Clinical characteristics of patients with COPD complicated with NSCLC were analyzed. Serum sRAGE, lung function and lung HRCT were combined to evaluate the correlation between the degree of emphysema and the occurrence of NSCLC in COPD, and receiver operator characteristic (ROC) curve analysis was performed for diagnostic efficiency. Results Compared with NSCLC group, COPD combined with NSCLC group had higher proportion of male patients, higher proportion of elderly patients, higher smoking index, and higher proportion of squamous cell carcinoma (P<0.05). FEV1 and FEV1%pred in COPD combined with NSCLC group were significantly lower than those in COPD group and NSCLC group. The Goddard score and EI values of emphysema were significantly increased (P<0.05). Serum sRAGE was significantly lower than that of COPD group and NSCLC group (P<0.05). Serum sRAGE level was positively correlated with FEV1%pred (r=0.366, P<0.001) and FEV1/FVC (r=0.419, P<0.001), and negatively correlated with Goddard score (r=–0.710, P=0.001) and EI value (r=–0.515, P<0.001). Binary multi-factor logistic regression analysis showed that age, smoking index, EI, Goddard score, RV/TLC were positively correlated with the risk of COPD developing NSCLC, while FEV1%pred, FVC, FEV1/FVC and serum sRAGE were negatively correlated with the risk of COPD developing NSCLC. ROC curve results showed that the area under the curve (AUC) of single diagnosis of sRAGE was 0.990, and the optimal cut-off value of 391.98 pg/mL with sensitivity of 93.3% and specificity of 89.7%. The AUC of sRAGE combined with age, smoking index, EI, Goddard score, FEV1%pred, FVC, FEV1/FVC, RV/TLC was 1.000 with sensitivity of 96.7%, specificity of 96.6%, and Yoden index of 0.933. Conclusion The combination of serum sRAGE, lung function and HRCT emphysema score can improve prediction of NSCLC occurrence in COPD.
Objective To investigate the effect of advanced glycation end products (AGEs) on the catalase activity and the levels of malondialdehyde in cultured bovine retinal capillary pericytes (BRPs), and to investigate the relationship between oxidative stress and diabetic retinopathy. Methods Cultured BRPs were exposed to AGEs (0, 8, 32, 125, 500, 2 000 μg/ml) for four days. Activity and the levels of catalase and malondialdehyde in cultured BRPs were examined by spectrophotometry. Results AGEs decreased the catalase activity, whereas increased the levels of malondialdehyde of cultured BRPs in a dose-dependent manner (r=-0.714, r=0.748, P＜0.01).There were significant differences between BRPs cultured in 32 μg/ml AGEs and in control group (P＜0.01), while no significant differences between BRPs cultured in non-glycated bovine serum albumin and absence of bovine serum albumin were found. Conclusion Oxidative stress may be one of the reasons why the pericyte disappears in diabetic retinopathy. (Chin J Ocul Fundus Dis, 2002, 18: 143-145)
ObjectiveTo observe and analyze the effect of HbA1c level on macular microcirculation in patients with type 2 diabetes mellitus (T2DM).MethodsA cross-sectional study. One hundred and twenty-four T2DM patients (124 eyes) without diabetic retinopathy who diagnosed by the examination of fundus color photography in Lixiang Eye Hospital Of Soochow University during September to December 2017 were enrolled in this study. There were 59 males (59 eyes) and 65 females (65 eyes), with the mean age of 65.06±7.99 years old. All patients underwent BCVA, fundus color photography, and OCT angiography (OCTA). The history of diabetes, hypertension and dyslipidemia were recorded in detail. According to the HbA1c level, patients were divided into three groups, HbA1c ideal control group (group A, HbA1c ＜7%, 67 eyes), HbA1c control group (group B, 7%≤HbA1c≤9%, 44 eyes), and HbA1c poor control group (group C, HbA1c＞9%, 13 eyes), respectively. The 3 mm×3 mm range of the macular area was scanned by OCTA instrument. The vascular density (VD) and skeleton density (SD) of nonsegmented retinal layer (NRL), superficial retinal layer (SRL) and deep retinal layer (DRL) in the macular area and foveal avascular zone (FAZ) area, non-circularity index, axial rate (AR) of SRL were measured. The correlation between HbA1c, BCVA and VD, SD of NRL, SRL, DRL was analyzed statistically with Spearman correlation test. The correlation between systemic factors and the above indicators was analyzed statistically with linear regression analysis.ResultsThe results of linear regression analysis showed that HbA1c was significantly correlated with VD (t=?3.237, ?3.156, ?2.050) and SD (t=?0.3.45, ?3.034, ?2.248) of NRL, SRL and DRL (P＜0.05); but no correlation with FAZ, non-circularity index and AR (t=1.739, 0.429, 1.155; P＞0.05). The differences of VD (F=6.349, 5.981, 3.709), SD (F=7.275, 6.085, 1.904) and AR (F=0.027) of NRL, SRL and DRL in group A, B and C were statistically significant (P＜0.05); but the differences of FAZ (F=1.904), non-circularity index (F=0.280) was not statistically significant (P＞0.05). Significant differences (P＜0.05) of VD and SD of NRL were found between group A and B (t=1.987, 2.201), group A and C (t=3.365, 3.572), group B and C (t=2.010, 2.076). Significant differences (P＜0.05) of VD and SD of SRL were found between group A and B (t=2.087, 2.168), group A and C (t=3.197, 3.194). There were significant differences (P＜0.05) in SD of DRL between group A and B (t=2.239), group A and C (t=?2.519). There was significant difference in VD of DRL between group A and C (t=2.363). The results of Spearman correlation analysis showed that HbA1c was negatively correlated with VD (r=?0.273, ?0.255, ?0.222; P=0.002, 0.004, 0.013) and SD (r=?0.275, ?0.236, ?0.254; P＜0.05) of NRL, SRL, DRL; positively correlated with FAZ and BCVA (r=0.221, 0.183; P＜0.05). BCVA was negatively correlated with VD (r=?0.210, ?0.190, ?0.245) and SD (r=?0.239, ?0.207, ?0.296) of NRL, SRL, and DRL (P＜0.05), but not correlated with FAZ (r=0.099, P＞0.05).ConclusionThe decrease of macular perfusion and the morphological change of FAZ accompanied by HbA1c increased.