【摘要】 目的 观察晚期糖基化终产物（advanced glycosylation end prodrcts，AGE）对人结肠癌细胞株SW-480增殖的影响，并探讨其可能机制。 方法 不同浓度AGE干预SW-480细胞，噻唑蓝（MTT）法比较各组细胞活力，流式细胞术观察AGE对SW-480细胞周期的影响，蛋白质印迹法观察AGE对SW-480细胞CyclinD1表达的影响，端粒重复序列扩增法（telomeric repeat amplification protocol，TRAP）银染法观察AGE对SW-480细胞端粒酶活性的影响。MTT测细胞活力的检测设置空白对照组、100 μg/mL小牛血清白蛋白（bovine serum albumin，BSA）组及50、100、500 μg/mL AGE组，其余检测只设置100 μg/mL BSA组和100 μg/mL AGE组。 结果 MTT结果示AGE促进SW-480细胞的增殖，且呈浓度依赖性。100 μg/mL BSA组与100 μg/mL AGE组72 h后的细胞G0/G1期所占百分比分别为56.02%±0.58%、51.93%±1.01%，差异有统计学意义（Plt;0.05）。蛋白质印迹法示100 μg/mL AGE组72 h后CyclinD1的表达较100 μg/mL BSA组增加，差异有统计学意义（Plt;0.05）。TRAP银染法检测示100 μg/mL AGE干预SW-480细胞72 h后可以增加端粒酶活性（Plt;0.05）。 结论 AGE可促进人结肠癌细胞SW-480生长，呈剂量依赖性。其作用机制可能与AGE上调CyclinD1的表达加速G1/S期转换及增加端粒酶活性有关。【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.
Objective To study the expression of receptor of advanced glycation end products (RAGE) in autogenous vein graft of streptozotocin induced diabetic rats and the inhibitory effects of aminoguanidine on intimal hyperplasia. Methods Sixty male Sprague-Dawley rats were randomly divided into three groups: aminoguanidine group, distilled water group and control group. Autogenous vein graft models were established in all groups. Streptozotocin was injected into abdominal cavity to induce diabetes in both aminoguanidine group and distilled water group, and they were intragastric administrated with aminoguanidine or distilled water, respectively before and after transplantation. Specimens were collected from autogenous vein graft 7 days and 14 days after surgery to undergo histological examination. At the same time, the level of serum advanced glycation end products (AGE) was tested. Western blotting and immunohistochemistry were used to detect the protein expression of RAGE and NF-κB p65. RAGE and NF-κB p65 mRNA were measured by reverse transcription-PCR. Results The mRNA and protein expressions of RAGE, NF-κB p65, the level of serum AGE and the intimal thickness of vein graft in distilled water group increased in comparison with those in control group 7 days and 14 days after surgery （P＜0.05). The level of serum AGE, mRNA and protein expressions of NF-κB p65 and the intimal thickness of vein graft in aminoguanidine group were lower than those in distilled water group （P＜0.05), and showed no significant difference compared with control group （P＞0.05). Conclusion The over-expression of RAGE in vein graft activats NF-κB in streptozotocin-induced diabetic rat, which has a close relation with intimal hyperplasia. Aminoguanidine can block the binding of AGE and RAGE by inhibiting the production of AGE, which will prevent intimal hyperplasia of vein graft.
ObjectiveTo observe the protective effect of tanshinone Ⅱ A on the mouse liver ischemia-reperfusion injury (IRI) model and preliminarily explore its mechanism of alleviating liver injury.MethodsThe IRI mouse model was established after the pre-treating with tanshinone Ⅱ A. Then, the serum and liver tissue of mice were collected to detect the changes of liver function, histopathology, liver cell apoptosis, and inflammatory factors. In addition, the protein expression levels of high mobility group box 1 (HMGB1), advanced glycosylation end-product specific receptor (RAGE), and Toll like receptor 4 (TLR4) in the liver tissues were detected by the Western blot method.ResultsAll data were analyzed by the homogeneity of variance test. The results of factorial design showed that the levels of ALT and AST in the serum, the pathological score and apoptosis index, the inflammatory response, as well as the expressions of HMGB1, TLR4 and RAGE proteins in the liver tissues were decreased significantly (P<0.05) in the sham operatation plus tanshinone Ⅱ A mice, which were increased significantly (P<0.05) in the IRI mice, which were antagonized synergistically by the tanshinone ⅡA and IRI (P<0.05).ConclusionsTanshinone ⅡA could reduce the liver IRI and inflammatory response in mouse. These effects might be related to the down-regulations of TLR4, HMGB1, and RAGE expressions.
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.