miR-142-5p inhibits the invasion and migration of lung adenocarcinoma H1650 cells by affecting epithelial mesenchymal transformation / 中国肿瘤生物治疗杂志

@# Objective: To study the expression of miR-142-5p in lung adenocarcinoma tissues, and to explore its effect on proliferation, invasion, migration and epithelieal-mesenchymal transition (EMT) of H1650 cells and the potential mechanisms. Methods:Atotal of 107 pairs of lung adenocarcinoma tissues and corresponding para-cancerous tissues from patients, who underwent tumor resection and were pathologically confirmed at the Department of Thoracic Surgery, the Fourth Hospital of Hebei Medical University between Jan. 2014 and Jan. 2015, were collected for this study; in addition, human lung adenocarcinoma cell lines (H1650, HCC827, A549, H1975, PC9) and human bronchial epithelial BEAS-2B cells were also used in this study. qPCR was used to detect the expression of miR-142-5p in lung adenocarcinoma tissues and cell lines. The correlation between expression of miR-142-5p and clinical features was analyzed.After transfection with miR-142-5p mimics or miR-negative control (miR-NC) plasmid, the proliferation, invasion and migration of H1650 cells were detected with CCK-8, Transwell invasion assay and Wound healing assay, respectively. The bioinforamtics tool was used to predict the target genes of miR-142-5p, and Luciferase reporter gene assay was performed to validate the regulation of miR-142-5p on target gene. Western blotting (WB) was used to detect the expressions of cyclin-dependent kinase 5 (CDK5) and EMTrelated protein. Results: Compared to Para-cancerous tissues and BEAS-2B cells, the expression of miR-142-5p was lower in lung adenocarcinoma tissues and cell lines (all P<0.01). Of the 107 cases of lung adenocarcinoma tissues, 61 cases (57.01%) showed decreased miR-142-5 expression, which was correlated with the TNM stage and lymph node metastasis (both P<0.01). Transfection of miR-142-5p mimics significantly up-regulated the expression of miR-142-5p and decreased the proliferation, invasion and migration of H1650 cells (all P<0.05 or P<0.01). Bioinformatics showed that CDK5 was a target gene of miR-142-5p. Luciferase reporter gene assay and WB validated that miR-142-5p could significantly down-regulate CDK5 expression in H1650 cells, up-regulate the expression of E-cadherin and down-regulate the expressions of N-cadherin, Twist and Snail in H1650 cells (all P<0.01). Conclusion: miR-142-5p is low expressed in lung adenocarcinoma tissues and cell lines; it suppresses the EMT process to inhibit, invasion and migration of H1650 cells via down-regulating the expression of CDK5.

knocking out mediated by CRISPR-Cas9 genome editing for PD-L1 attenuation and enhanced antitumor immunity

Blocking the programmed death-ligand 1 (PD-L1) on tumor cells with monoclonal antibody therapy has emerged as powerful weapon in cancer immunotherapy. However, only a minority of patients presented immune responses in clinical trials. To develop an alternative treatment method based on immune checkpoint blockade, we designed a novel and efficient CRISPR-Cas9 genome editing system delivered by cationic copolymer aPBAE to downregulate PD-L1 expression on tumor cells specifically knocking out Cyclin-dependent kinase 5 () gene . The expression of PD-L1 on tumor cells was significantly attenuated by knocking out , leading to effective tumor growth inhibition in murine melanoma and lung metastasis suppression in triple-negative breast cancer. Importantly, we demonstrated that aPBAE/Cas9-Cdk5 treatment elicited strong T cell-mediated immune responses in tumor microenvironment that the population of CD8 T cells was significantly increased while regulatory T cells (Tregs) was decreased. It may be the first case to exhibit direct PD-L1 downregulation CRISPR-Cas9 genome editing technology for cancer therapy. It will provide promising strategy for preclinical antitumor treatment through the combination of nanotechnology and genome engineering.

Effect of Wuzang Wenyang Huayu Decoction on Neurofibrillary Tangles by Inhibiting Cyclin-dependent Kinase-5 Over-expression in SAMP8 Mice / 中国实验方剂学杂志

Objective: To explore the mechanism of Wuzang Wenyang Huayu decoction in improving the cognitive competence and the pharmacological mechanism for neurofibrillary tangles related to cyclin-dependent kinase-5(CDK-5).Method: The 10 SAMR1 mice were used as normal group,40 SAMP8 mice were randomly divided into model group,donepezil group (0.4 mg·kg-1·d-1),high and low dose Wuzang Wenyang Huayu decoction groups (5,1.25 g·kg-1·d-1).Drugs were administered by gastric lavage for 4 continuous weeks.Directional navigation and space exploration ability were evaluated with Morris amaze.Real-time PCR was used to measure the mRNA expression of CDK-5 in brain nerve tissues.Western blot was used to detect the protein expression of CDK-5 and phosphorylation of Tau protein.Meanwhile,neurofibrillary tangles in brain tissue were detected with silver staining method.Result: As compared with normal group,both CDK-5 expression and Tau protein phosphorylation in brain nerve tissues were remarkably increased in model group (PPPPPPConclusion: Wuzang Wenyang Huayu decoction can markedly improve the cognitive competence of SAMP8 mice,and the mechanism may be related to its inhibition on CDK-5 over-expression,and down-regulation of Tau protein phosphorylation and neurofibrillary tangles in brain tissue.

Regulation of thin recipe of Buyang Huanwudecoction on Cdk5 expression of rats after cerebral ischemia / 中国药理学通报

Aim To evaluate the regulation of thin recipe of Buyang Huanwu decoction on cyclin-dependent kinase 5(Cdk5)expressions in hippocampus tissue of rats after cerebral ischemia.Methods Male SD rats were divided into sham-operation group,MCAO group,Buyang Huanwu decoction group(ig.3.15 g·kg-1)and its thin recipe composition group(ig.2.41 g·kg-1).Each group was then divided into five subgroups based on the time after administration for 1,3,7,14,28 d respectively.Cdk5 protein and mRNA levels in each group were examined by using immunohistochemistry,Western blot and real-time PCR respectively.Results The up-regulation of Cdk5 was observed in model rat hippocampus after cerebral ischemia 1 day,and kept increasing with the aggravation of ischemia injury,the peaked expression was observed after 7～14 d,while the downtrend was observed after 28 days compared with the corresponding sham-operation groups(P0.05).Conclusion The thin recipe of Buyang Huanwu decoction could exert the protective effect by regulating Cdk5 after cerebral ischemia.

Research on the change of cyclin-dependent kinase 5 in hippocampus CA1 after conditioned fear extinction / 中华行为医学与脑科学杂志

Objective To investigate the change of cyclin-dependent kinase 5(CDK5)during 1 week after conditioned fear extinction.Methods Forty-two male adult SD rats were randomly divided into native group,extinction control group and 24h extinction group.Extinction retention test and immunohistochemistry of CDK5 of hippocampus were done at 1st,3rd,7th day after fear extinction.The numbers of CDK5 positive cell in hippocam-pus CAI was measured by computer analytic system.Results (1)Extinction retention scores were increasedgradually in extinction group compared with native group(75.60±2.51)%,the scores of extinction control group was decreased at 1st day(15.62±10.28)% and 3th day(20.58±7.79)% after extinction(P＜0.01);24h extinction group(71.04±11.65)% were better than extinction control group(35.48±12.37)at 7th day after extinction(P＜0.01).(2)The number of CDK5 positive cell was increased significantly in 24h extinction group (24.94±5.20;32.25±6.14;33.28±6.56)compared with native group(75.60±2.51;P＜0.01).In 24h extinction group the number was increased significantly at 3rd day(P＜0.01)and 7th day(P＜0.05)compared with extinction control group(25.09±4.83;26.70±4.57),and decreased at 1st day compared to 3rd day(P＜0.05)and 7th day(P＜0.01).Conclusion From 1st to 7th day after extinction,CDK5 may contribute to the retention of fear extinction in hippocampal CA1.