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Objective:To investigate the expression of p21-activated kinase 2 (PAK2) in laryngeal squamous cell carcinoma and its relationship with the clinicopathological characteristics and chemosensitivity of patients.Methods:Transcriptome sequencing (RNA-seq) data for laryngeal squamous cell carcinoma were downloaded from the Cancer Genome Atlas (TCGA) database, and 123 patients were included in the study (12 cases had cancer tissues and normal tissues data, and the remaining 111 only had cancer tissues data). Differential expression of PAK2 in cancer and para-cancer tissues was analyzed by using R software, and the potential function of PAK2 in laryngeal squamous cell carcinoma was investigated by using the Kyoto Encyclopedia of Genes and Genomes (KEGG) database signaling pathway enrichment. A total of 34 patients with primary laryngeal squamous cell carcinoma tissues and corresponding para-carcinoma 34 tissue specimens who underwent surgical resection were retrospectively selected from Chaoyang Central Hospital between April 2016 and June 2021, and 20 cases of normal laryngeal mucosa tissues were selected as the controls. Immunohistochemistry was used to detect the expression of PAK2 in various tissues, and its correlation with clinicopathological factors was analyzed. A total of 35 supraglottic primary laryngeal squamous cell carcinoma patients were retrospectively collected before induction chemotherapy during the same period, including 20 patients sensitive to chemotherapy and 15 patients resistant to chemotherapy. Real-time fluorescence quantitative polymerase chain reaction (qRT-PCR) was used to detect the relative expression level of PAK2 mRNA in cancer tissues.Results:Analysis of TCGA database data showed that PAK2 expression was increased in cancer tissues compared with that in para-cancer tissues ( P = 0.012); KEGG database signaling pathways showed that the high expression of PAK2 in laryngeal squamous cell carcinoma was related to signal transduction pathways, cell cycle, and cancer. Immunohistochemistry showed that the proportion of PAK2 positive in 34 cases of laryngeal squamous cell carcinoma tissues was higher than that in adjacent tissues and normal tissues [58.82% (20/34) vs. 0.03% (1/34), 0 (0/20), all P < 0.001]. There were statistically significant differences in the proportion of PAK2 positive patients stratified with different degrees of differentiation [high differentiation vs. low or middle differentiation: 33.33% (6/18)vs. 87.50% (14/16)], lymph node metastasis [presence vs. absence: 90.91% (10/11) vs. 43.48% (10/23)], TNM staging [stage Ⅲ-Ⅳ vs. stage Ⅰ-Ⅱ: 82.35% (14/17) vs. 35.29% (6/17)] (all P < 0.05), and PAK2 positive patients were not associated with clinical type, tumor size, smoking history, drinking history, and age (all P > 0.05). qRT-PCR showed that the relative expression level of PAK2 mRNA in the chemotherapy-resistant group was higher than that in the chemotherapy-sensitive group (3.89±0.12 vs. 0.78±0.23, P < 0.001). Conclusions:The expression level of PAK2 in laryngeal squamous cell carcinoma tissues is increased, and the high expression of PAK2 is closely related to the malignant clinical characteristics of patients with laryngeal squamous cell carcinoma. The high expression of PAK2 may indicate the insensitivity to traditional chemotherapy regimens, and PAK2 may be a potential gene that targets and regulates the chemosensitivity of laryngeal squamous cell carcinoma.
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The serine/threonine p21-activated kinases (PAKs), as main effectors of the Rho GTPases Cdc42 and Rac, represent a group of important molecular switches linking the complex cytoskeletal networks to broad neural activity. PAKs show wide expression in the brain, but they differ in specific cell types, brain regions, and developmental stages. PAKs play an essential and differential role in controlling neural cytoskeletal remodeling and are related to the development and fate of neurons as well as the structural and functional plasticity of dendritic spines. PAK-mediated actin signaling and interacting functional networks represent a common pathway frequently affected in multiple neurodevelopmental and neurodegenerative disorders. Considering specific small-molecule agonists and inhibitors for PAKs have been developed in cancer treatment, comprehensive knowledge about the role of PAKs in neural cytoskeletal remodeling will promote our understanding of the complex mechanisms underlying neurological diseases, which may also represent potential therapeutic targets of these diseases.
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Animais , Humanos , Citoesqueleto/genética , Doenças do Sistema Nervoso/genética , Neurônios/enzimologia , Transdução de Sinais , Quinases Ativadas por p21/metabolismoRESUMO
Objective: To investigate the effects of microRNA-129-5p (miR-129-5p) on the proliferation and apoptosis of osteosarcoma cells, and to explore the possible molecular mechanism. Methods: The expression of miR-129-5p in osteosarcoma MG63 cells and osteoblasts hFOB1.19 cells was determined by real-time fluorescent quantitative PCR (RFQ-PCR). After miR-129-5p mimics were transfected into osteosarcoma MG63 cells, the expression of miR-129-5p was detected by RFQ-PCR to verify the transfection efficiency. CCK-8 assay and plate clony formation experiment were used to detect the cell proliferative ability, while FCM was performed to analyze the cell apoptosis. Double luciferase reporter gene system was used to verify the interaction between miR-129-5p and the target gene p21-activated kinase 5 (PAK 5). The expressions of PAK5 mRNA and protein in MG63 cells with miR-129-5p overexpression were tested by RFQ-PCR and Western blotting, while the expression levels of phospho-glycogen synthase kinase-3β (p-GSK3β) and β-catenin were detected by Western blotting. After siRNA-PAK5, PAK5 overexpressed plasmids and miR-129-5p mimics＋ PAK5 overexpressed plasmids were separately transfected into osteosarcoma MG63 cells, the above methods were used to further verify the molecular mechanism of miR-129-5p regulating the proliferation and apoptosis of osteosarcoma cells. Results: The level of miR-129-5p in MG63 cells was lower than that in hFOB1.19 cells (P < 0.05). After the transfection with miR-129-5p mimics, the proliferation and clony formation abilities of MG63 cells were decreased (P < 0.001, P < 0.01), while the apoptosis rate was increased (P < 0.05). miR-129-5p regulated negatively the expression of PAK 5 gene (P < 0.05). After the transfection with miR-129-5p mimics or siRNA-PCK5, the expression levels of PAK5 mRNA and protein were decreased (all P < 0.01), while the expressions of p-GSK-3β and β-catenin proteins were also decreased (both P < 0.05, both P < 0.01). The overexpression of PAK5 could restore the effects of miR-129-5p on proliferation and apoptosis of osteosarcoma cells (both P < 0.05). Conclusion: miR-129-5p can inhibit the proliferation ability of osteosarcoma cells, and promote apoptosis by down-regulating the expression of PAK 5 gene. The mechanism may be associated with the expression regulation of p-GSK-3β and β-catenin proteins.
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Objective@#To study effects of cyclin D1 overexpression on the proliferation and differentiation of cervical squamous cell carcinoma SiHa cells and to investigate related signaling molecules.@*Methods@#Primers were designed to amplify the full length of cyclin D1 gene and cyclin D1 gene was amplified by PCR for constructing pcDNA3.1 plasmid vector. The construct was then transfected into SiHa cells, and the cells with stable overexpression of cyclin D1 were established, cyclin D1 gene and protein expression were detected by RT-PCR and Western blot, respectively. Cell growth curve was documented by MTT assay. CK7, E-cadherin, vimentin, Snail gene and protein expression in transfected cells were detected by RT-PCR and Western blot. RT-PCR was used to detect the mRNA expression of proliferation and differentiation-related genes like CDK4, CDK2, p21, p27, cyclin E, Rb, E2F, E6/E7 and Ki-67. After synchronization of cells, RT-PCR was used to detect of cyclin D1 and p21 mRNA expression at different time points of the cell cycle.@*Results@#The G-3 cells with cyclin D1 overexpression were successfully established. The growth curve and Ki-67 mRNA expression accelerated in G-3 cells.Vimentin and Snail expression significantly increased at both gene and protein levels, while E-cadherin, CK7 gene and protein expression significantly decreased, indicating epithelial mesenchymal transitionoccurred in G-3 cells.Meanwhile, mRNA expression of cyclin D1, CDK4, CDK2, p21, p27, cyclin E, E2F and Rb increased, while E6/E7 and p16 showed no significant change. The expression trends of p21 and cyclin D1 were almost identical with fluctuation at different time points in the cell cycle.@*Conclusions@#Overexpression of cyclin D1 induced by gene transfection promotes proliferation and epithelial mesenchymal transition in SiHa cells.The process is accompanied by up-regulation of CDK4, CDK2, p21, p27 and cyclin E genes.p21 expression increases synchronously with cyclin D1, suggesting a regulatory role in epithelial mesenchymal transition by affecting expression of vimentin in G-3 cells.
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Objective To investigate the expression characteristics of PAK5,CyclinD1 in HCC and its relationship with clinicopathological factors and prognosis.Methods The immunohistochemical method was used to detect the expression of PAK5 and CyclinDl in 76 cases of HCC tissue and 31 cases of adjacent liver tissue.The correlation between the abnormal expression of PAK5 and CyclinD1 and clinicopathological factors and prognosis was also analyzed.Results The PAK5 expression in tumor lesions and adjacent tissue was 86.8％ (66/76) and 54.8％ (17/31) (x2 =12.962,P ＜ 0.05).Meanwhile,the CyclinD1 was positive in 73.7％ (56/76) of the HCC tissue and in 41.9％ (13/31) of the adjacent tissue (x2 =9.691,P ＜0.05).The expression of PAK5 was significantly correlated with sex (x2 =5.063,P =0.024),tumor diameter(x2 =9.159,P =0.002) and portal vein-emboli (x2 =4.469,P =0.035).CyclinD1 expression was correlated with portal vein-emboli (x2 =4.842,P =0.028),TNM (x2 =7.930,P =0.005).PAK5 expression was positively correlated with CylinD1 (γ =0.284,P =0.033).The mean survival time of PAK5positive patients after operation was shorter than the PAK5 negative patients(x2 =7.104,P =0.008).By Cox multiple factors analysis,the PAK5 expression was independent impact factors affecting the prognosis of HCC patients (RR =0.186,P =0.012).Conclusion In HCC patients positive PAK5 expression predicts poor postoperative survival.
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Objective To explore the mechanism of protective effect of Rhodioloside in cerebral ischemia-reperfusion rats and its relevance to phosphatidylinositol 3-kinases ( PI3-K)/protein serine-threonine kinases ( AKT) signaling pathway .Methods Forty eight Sprague-Dawley rats were randomly divided into four groups: sham-operation group , ischemia-reperfusion group , and Rhodiolo-side treatment groups (5 and 10 mg/kg).The model of right middle cerebral artery occlusion was established with thread ligation meth -od.The score of the neurological deficit was estimated 2 h followed by 24 h reperfusion.Histopathological changes were observed by hematoxylin-eosin(HE) staining.The infarct volume was measured with triphenyltetrazolium chloride (TTC) staining.Apoptotic cells were assessed with terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) method.The expressions of PI3-K and p-AKT were evaluated with immunohistochemistry .Results The score of the neurological deficit was decreased more ob-viously, the number of apoptotic were decreased more significantly , the expressions of PI3-K and p-AKT were increased more signifi-cantly in the Rhodioloside treatment groups (5 and 10 mg/kg) than in the ischemia-reperfusion group ( P <0.05).The score of the neurological deficit was decreased , the number of apoptotic was decreased , and the expressions of PI 3-K and p-AKT were increased in the Rhodioloside treatment group (10 mg/kg) than the Rhodioloside treatment group (5 mg/kg) ( P <0.05).Conclusions The protective mechanism of Rhodioloside therapy against cerebral ischemia r-eperfusion injury might be associated with activating the PI 3-K/AKT signaling pathway and then inhibiting neuronal apoptosis .
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The Paks (p21-activated kinases) are members of a family of serine/threonine kinases which are highly conserved throughout evolution in a variety of organisms. Paks can be activated by a variety of upstream signaling molecules, especially by Rac and Cdc42 from Rho family small G proteins. The activated Paks play an important role in regulating many signaling pathways and cell function. The aberrant expression of Paks has been observed in a variety of cancers. Paks promote tumorigenesis through regulating cytoskeleton remodeling, cell motility, cell proliferation, cell differentiation, apoptosis, mitosis and angiogenesis. It appears promising to consider Paks as potential therapeutic targets in cancer therapy. Copyright © 2012 by TUMOR.
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p21-activated kinases (PAKs) are a family of serine/threonine protein kinases comprised of six isoform (PAK1-6), all of which are direct targets of the small GTPases Rac and Cdc42. PAKs have recently been shown to regulate various cellular activities, including cell motility, survival and proliferation, the organization and function of cytoskeleton and extracellular matrix, transcription and translation. PAKs are overexpressed or hyperactivated in several human tumor, such as breast cancer, gastric cancer, ovarian cancer etc., which makes them an attractive new therapeutic targets. Thus, there has been considerable interest in the development of inhibitors to the PAKs, as biological markers and leads for the development of therapeutics.