ABSTRACT
@#[摘 要] 目的：分析miR-19对PTEN及PI3K-Akt通路的调节作用，揭示miR-19和PTEN对子宫内膜癌KLE细胞侵袭、迁移的影响及其机制。方法：收集2017年5月至2020年8月河北医科大学第一医院收治的74例子宫内膜癌患者经手术切除（术前未接受放化疗等治疗）且病理确诊为子宫内膜癌的肿瘤组织及对应癌旁组织，采用qPCR和WB法检测PTEN在子宫内膜癌组织中的表达水平以及转染miR-19模拟物或抑制物对KLE细胞中PTEN表达的影响。通过TargetScan及双荧光素酶报告基因实验预测并验证PTEN是否为miR-19的靶基因，将KLE细胞随机分为对照（NC）组、miR-19模拟物组和miR-19+PTEN组，分别转染阴性对照片段（miR-NC）、miR-19模拟物或共转染miR-19模拟物+PTEN过表达载体，采用Transwell和细胞划痕实验检测miR-19和PTEN对KLE细胞迁移和侵袭能力的影响，WB法检测对PI3K-Akt通路相关蛋白表达的影响。结果：子宫内膜癌组织中PTEN的mRNA和蛋白表达水平均明显低于癌旁组织（均P<0.01）。miR-19能与PTEN mRNA的3’-UTR特异性结合，上调miR-19的表达能抑制PTEN的mRNA和蛋白的表达，下调miR-19的表达则出现相反的结果（均P<0.01）。与miR-NC组相比，miR-19 模拟物组的迁移、侵袭细胞数以及划痕愈合率均显著升高（均P<0.01），而miR-19+PTEN组的细胞迁移和侵袭能力以及划痕愈合率较miR-19摸拟物组则明显降低（均P<0.01）；与miR-NC组相比，miR-19模拟物组中PTEN蛋白的表达明显降低，而p-Akt 308和p-Akt 473蛋白的表达明显升高，Akt蛋白的表达无明显变化（均P<0.01）；而在miR-19+PTEN组中，p-Akt 308和p-Akt 473蛋白的表达均明显低于miR-19模拟物组（均P<0.01）。结论：miR-19可能通过抑制PTEN的表达从而活化PI3K-Akt通路，进而促进子宫内膜癌KLE细胞的迁移和侵袭。
ABSTRACT
MicroRNAs (miRNAs) have been identified as potential biomarkers for endometrial carcinoma (EC) diagnosis, prognosisand therapy. The purpose of the present study was to investigate the detailed role and molecular mechanism of miR-195 inEC metastasis. qRT-PCR assay was performed to assess the expression of miR-195 and SRY-related high-mobility groupbox 4 (SOX4) mRNA in EC tissues and cells. The levels of N-cadherin, Vimentin, E-cadherin and SOX4 protein weredetermined by western blot. SOX4 protein expression in EC tissues was also determined by Immunohistochemistry (IHC)experiment. Transwell assay was used to analyze cell migration and invasion abilities. Dual-luciferase reporter assay andRNA Immunoprecipitation (RIP) assay were performed to confirm the targeted interaction between miR-195 and SOX4.Our data supported that miR-195 was downregulated and SOX4 was upregulated in EC tissues and cell lines. Upregulationof miR-195 inhibited migration, invasion and epithelial-mesenchymal transition (EMT) of EC cells. Moreover, SOX4 was adirect target of miR-195. MiR-195 overexpression-mediated anti-migration, anti-invasion and anti-EMT effects wereantagonized by SOX4 restoration in EC cells. In conclusion, our study suggested that miR-195 inhibited the migration,invasion and epithelial mesenchymal transition (EMT) of EC cells at least partly by targeting SOX4. Our study provided anovel underlying mechanism for EC metastasis and a promising therapeutic target for EC management.
ABSTRACT
Objective: To study the chemical constituents of Bletillae Formosanae Rhizoma and the distribution characteristics of active components in the fingerprint by establishing its high performance liquid chromatography (HPLC) fingerprints. Method: HPLC was used to establish the fingerprint of Bletillae Formosanae Rhizoma. Four reference substances,i.e. militarine,coelonin,4-methoxy-9,10-dihydrophenanthrene-1,2,7-triol and batatasin Ⅲ were used to identify chromatographic peaks. The fingerprints of 17 batches of Bletillae Formosanae Rhizoma fingerprints were analyzed and compared by "Computer-aided-similarity evaluation soft" and stoichiometry,and then compared with the fingerprint of Bletillae Rhizoma. Result: The established HPLC fingerprint method of Bletillae Formosanae Rhizoma showed good repeatability and stability. 20 common peaks were marked,four of which were identified by reference substances; militarine was No.8 common peak,and others corresponded to No. 10, No. 14 and No. 18 common peaks. Results showed that the similarities of samples except S4 were higher than 0.85, but the relative peak area of common peaks was quite different. Within the cluster distance 10,the samples are clustered into 5 categories, reflecting certain origin correlation. Principal component analysis (PCA) showed that the difference in samples was mainly caused by the common peaks located after No. 9 peak,where chemical constituents such as bibenzyl and dihydrophenanthrene were distributed. Bletillae Formosanae Rhizoma and Bletillae Rhizoma showed similar chemical constituents. Conclusion: The method provided a theoretical basis for the further clinical application and quality control of Bletillae Formosanae Rhizoma,as a substitute for Bletillae Rhizoma.
ABSTRACT
Objective To investigate the expression levels of retinoblastoma protein-interacting zinc finger gene 1 (RIZ1) in gliomas of different grades and its association with patient prognosis. Methods The expression of RIZ1 and Ki-67 in 51 glioma tissues of different grades were evaluated immunohistochemically. The correlation of RIZ1 immunoreactivity with the clinicopathological features and the prognostic value of RIZ1 in patients were analyzed. Results Immunohistochemistry results showed that RIZ1 was mainly expressed in the cytoplasm and Ki-67 was mainly located in the nuclei. We found that low grade glioma tissue had a significantly higher expression of RIZ1 compared with the high grade one ([7.55±4.39]% vs [48.83±4.34]%,P<0.000 1). There was a negative correlation between RIZ1 and Ki-67 immunoreactivity (Spearman: r=-0.813 0, P<0.000 1). It was also suggested that RIZ1 expression was negatively associated with tumor grade and patient age (P<0.05). Kaplan-Meier survival analysis revealed a positive correlation between expression of RIZ1 and progress-free survival (PFS) and overall survival (OS) of patients(P<0.000 1). Univariate analysis revealed that high RIZ1 expression and low Ki-67 expression were associated with better patient prognosis. Multivariate analysis using the Cox proportional hazards model revealed that high RIZ1 expression was significantly correlated with prognosis of patients with gliomas(P=0.000,HR=0.164,95%CI:0.071-0.378). Conclusion It is concluded that RIZ1 expression is decreased with the increase of pathological grade of gliomas, and it is associated with patients prognosis; RIZ1 may serve as a marker for treatment and prognosis prediction of gliomas.