The molecular and cellular mechanisms of bone metastasis in prostate cancer remain unclear. Current researches focus on chemotaxis of tumor cells in bone metastasis, interactions between tumor cells and bone micro-environment as well as the vicious circle among tumor cells, osteoclasts, osteoblasts and bone matrix.

To construct the eukaryotic vector that expresses the fusion protein of Axud1 and influenza virus hemagglutin HA epitope tag, the total RNA was isolated from the peripheral blood lymphocytes, and reverse transcription reaction was used to amplify the full length of human Axud1 cDNA. PCR product of Axud1 was then amplified using specific primers containing HA epitope sequence, and inserted into eukaryotic expression plasmid pcDNA3.1(+)digested with BamH Ⅰand Xba Ⅰ. The recombinant plasmid was identified by polymerase chain reaction, restriction endonuclease mapping and sequencing, and then transfected into human lung adenocarcinoma SPC-A1 cell lines.The fusion HA-Axud1 protein expression in anti-G418 clones was verified by Western blot. This study might be instrumental in further study of the function of Axud1 protein in tumor cells.

Objective To investigate the effects of exogenous prostaglandin E 1 (PGE 1) on expression of platelet derived growth factor B (PDGF-B) mRNA and its receptor ? protein in rabbit with schistosomiasis. Methods In this study, 14 rabbits were infected with cercaria of S. japonicum percutaneously. PGE 1 ( 2.5 ?g/kg?d -1 ) was given intravenously to 7 rabbits from the 60th day to day 120. The expressions of PDGF-B mRNA, PDGFR ? protein and ?-SMA were detected by RT-PCR, Western blotting and immunohistochemistry, respectively. Endogenous IFN-? was measured by in situ hybridization. Results Up-regulated expressions of PDGF-B mRNA, receptor ? protein as well as ?-SMA were observed in rabbits with Schistosome hepatic fibrosis. The increased expressions of PDGF mRNA and receptor ? were suppressed in rabbits treated with exogenous PGE 1 (29.42?5.05 vs 41.37?7.23, P

0.05 ). The rate of hypertensive gastropathy, compared with PCDV (66.74%), was significantly attenuated in patiens who underwent PCDV+ORP (22.78%, P

AIM: To determine the differences in phosphoproteome between LPS stimulated THP-1 cells with and without previous oxidative stress for screening of more potential regulators.METHODS: Differentiation of THP-1 cells into macrophages was induced by treatment with 100 ?g/L PMA for 36 h. Differentiated cells were rested for additional 36 h without PMA treatment, then treated with 100 ?mol/L H2O2 or medium for 1 h followed by LPS or medium treatment for 30 min. After desalted, phosphoproteins were enriched by phosphoprotein metal affinity column, and were run on 2-D electrophoresis, then the spots were analyzed to show the difference between LPS group (cells treated with LPS alone) and H2O2+LPS group (LPS stimulated cells also pretreated with H2O2). Finally, some of these spots were identified by MS and subsequent bioinformatic analysis was also conducted. RESULTS: Compared to LPS group, 29 reproducibly changed spots on the 2-D map in H2O2+LPS group were visualized and selected for MS analysis. Among these, 12 down-regulated spots (include those disappeared), 17 up-regulated spots (include those newly emerged) were selected. Up to now, 5 of these were identified, which were shown to be involved in various cellular processes such as proteolysis, signal transduction and protein folding. Among these, proteasome beta-4 subunit, which was dramatically down-regulated in H2O2+LPS group, was a major component of the proteasome complex and might participate in LPS signalling through various ways.CONCLUSION: With comparative phosphoprotein-affinity profiling, the interference brought by highly abundant house-keeping proteins is minimized, rendering us to detect less abundant signalling molecules. Aforementioned 5 proteins, especially proteasome beta-4 subunit, might be involved in LPS pathway reprogrammed by oxidative stress.

Objective:To observe the effect of combined operation of internal and external surgery for intraocular hemorrhage of polypoidal choroidal vascular disease (PCV).Methods:Retrospective clinical research. From January to August 2018, 14 eyes of 14 patients with PCV combined with vitreous hemorrhage (VH) with hemorrhagic retinal detachment confirmed by eye examination at the Second Affiliated Hospital of Nanchang University were enrolled in this study. The average disease course was 1.12±0.68 months. The visual acuity decreased from 2 to 14 days. The average time from hemorrhagic retinal detachment to surgery was 6.32±2.82 d. There were 9 eyes with BCVA of light sensing, 3 eyes of hand movement, and 2 eyes of counting fingers. The average macular foveal thickness (CFT) was 564.6±102.2 μm. The range of retinal detachment involves equal or greater than 2 quadrants. 23G vitrectomy surgery was used to remove VH, and subretinal hemorrhage was drained through external scleral drainage to reset the retina. One week and 1 and 2 months after the operation, the eyes were injected with 0.05 ml of ranibizumab in the vitreous cavity. The average follow-up time after surgery was 6.24±1.16 months. One week after the operation, 1, 3, and 6 patients were examined with the same equipment and methods before the operation to observe the condition of BCVA, CFT, intraocular pressure and retinal reattachment. Non-parametric tests were used to compare BCVA before and after surgery.Results:VH was completely cleared in all eyes, and no iatrogenic retinal hole appeared during the operation. Six months after the operation, there were 1 eye with BCVA of hand movement, 1 eye of counting fingers, 8 eyes of 0.1, 2 eyes of 0.2 and 2 eyes of over than 0.2; the average CFT was 336.4±54.8 μm. Compared with before surgery, the BCVA was significantly improved ( Z=-3.711, P=0.000), the CFT was significantly decreased ( t=3.212, P=0.006). OCT examination showed that the macular morphology and structure were clearly visible. Of the 14 eyes, 13 eyes were reattached, 1 eye was detached again after silicone oil removal. Intraocular pressure increased in 3 eyes within 1 week after surgery, including 1 eye with a small amount of blood in the anterior chamber. No recurrent VH was found in all eyes during follow-up. Conclusions:Combined internal and external surgery for PCV combined with VH with hemorrhagic retinal detachment is safe and effective.

OBJECTIVETo investigate AXIN1-related CSRNP1 gene expression and the mechanism of its transcriptional regulation in TGF-β1-induced tumor cells.

METHODSHuman lung carcinoma A549 cells or human prostate cancer PC3 cells were treated with TGF-β1 at different doses (0, 20, 40, and 80 ng/ml) or at 20 ng/ml for 0, 8, 12, or 24 h, and the dose and time effect of TGF-β1 on CSRNP1 mRNA expression in the tumor cells were evaluated with real-time RT-PCR. A549 cells were also treated with TGF-β1 and cycloheximide to clarify whether CSRNP1 expression induced by TGF-β1 required de novo protein synthesis. A549 cells transfected with pcDNA3.1, flag-SMAD3, or flag-SMAD3-mu, after serum starvation, were treated with or without TGF-β1 (20 ng/mL) for 24 h, and the overexpression of wild-type SMAD3 and dominant negative SMAD3-mu mutant were confirmed by Western blotting. The effect of SMAD3 or SMAD3-mu overexpression on CSRNP1 mRNA expression was also measured by real-time RT-PCR.

RESULTSIn both A549 and PC3 cells, TGF-β1 dose- and time-dependently stimulated CSRNP1 expression, which required de novo protein synthesis in A549 cells. Overexpression of wild-type SMAD3 significantly increased the expression of CSRNP1 mRNA induced by TGF-β1, while overexpression of dominant negative SMAD3 mutant remarkably reduced CSRNP1 mRNA expression in response to TGF-β1 in A549 cells.

CONCLUSIONTGF-β1 may contribute to CSRNP1 expression through SMAD3 activation and downstream signaling in tumor cells.

Apoptosis Regulatory Proteins ; genetics ; metabolism ; Axin Protein ; genetics ; metabolism ; Cell Line, Tumor ; Gene Expression Regulation, Neoplastic ; Humans ; RNA, Messenger ; genetics ; Signal Transduction ; Smad3 Protein ; genetics ; metabolism ; Transfection ; Transforming Growth Factor beta1 ; pharmacology

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.

OBJECTIVE: To study the effects of cadmium on the expression of estrogen receptor( ER) and miRAN-155,miRAN-200 c in human breast cancer MCF-7 cells. METHODS: MCF-7cells in logarithmic growth phase were randomly divided into fulvestrant( ICI182780,ICI) group and non-ICI group. The non-ICI group was treated with cadmium chloride(Cd Cl2) at the final concentrations of 0. 0,2. 5,5. 0 and 10. 0 μmol/L for 24 hours. The ICI group was pretreated at a concentration of 1. 0 μmol/L for 12 hours,and then treated with Cd Cl2 at the final concentrations 0. 0,2. 5,5. 0 and 10. 0μmol/L for 24 hours. The cell proliferation activity was measured by methyl thiazolyl tetrazolium assay. Flow cytometry was used to measured cell apoptosis. Western blot was applied to measure the relative expression of ERα and ERβ protein,and the relative expression of miRNA-155 and miRNA-200 c were detected by real-time fluorescence quantitative polymerase chain reaction. RESULTS: The proliferation rates of MCF-7 cells in 2. 5,5. 0 and 10. 0 μmol/L Cd Cl2 groups were significantly decreased than the 0. 0 μmol/L Cd Cl2 group( P < 0. 05). The proliferation rate in ICI group was lower than that of the non-ICI group( P < 0. 05). When Cd Cl2 concentration was 2. 5,5. 0 and 10. 0 μmol/L,the apoptosis rate of MCF-7 cells in non-ICI group increased compared with those cells without exposure to Cd Cl2( P < 0. 05). The relative expression of ERα,ERβ,miRNA-155 and miRNA-200 c increased( P < 0. 05). The proliferation of MCF-7 cells in ICI group decreased( P < 0. 05),and the relative apoptosis rate increased( P < 0. 05); and the relative expression of ERαand ERβ increased( P < 0. 05),the relative expression of miRNA-155 and miRNA-200 c decreased( P < 0. 05). When treated without Cd Cl2,the apoptosis rate of the ICI group increased compared with non-ICI group(P < 0. 05),the relative expression of ERα and ERβ decreased( P < 0. 05),and the relative expression of miRNA-155 and miRNA-200 c were increased( P < 0. 05). When Cd Cl2 concentration was 2. 5,5. 0 and 10. 0 μmol/L,the apoptosis rate and the relative expression of ERα,ERβ,miRNA-155 and miRNA-200 c decreased compared with the non-ICI group treated with same dose Cd Cl2(P < 0. 05). CONCLUSION: Cadmium can induce cell apoptosis and increase expression of miRNAs through the ER signaling pathway.

ObjectiveTo study the effects of Epimedii Folium polysaccharides on mice with exercise-induced fatigue and explore its possible mechanism of action. MethodICR male mice screened by swimming training were randomly divided into a control group， model group， vitamin C group， and low， medium， and high dose groups of Epimedii Folium polysaccharides， with eight mice in each group. The exercise-induced fatigue model was established by weight-bearing swimming training in each group except for the control group. After two weeks of weight-bearing swimming， the Epimedii Folium polysaccharide groups were given 100， 200， 400 mg∙kg^-1 of Epimedii Folium polysaccharides by gavage， and the vitamin C group was given 200 mg∙kg^-1 of vitamin C by gavage. The control group and the model group were given equal amounts of saline for 14 d. At the end of the experimental period， the body mass of the mice in each group and the time of last swimming due to exhaustion were recorded. Serum urea nitrogen （BUN）， lactic acid （LA）， lactate dehydrogenase （LDH）， malondialdehyde （MDA）， superoxide dismutase （SOD）， glutathione peroxidation （GSH-Px）， myoglycogen （MG） in skeletal muscle， hepatic glycogen （HG） in the liver were detected by kits. Hematoxylin-eosin （HE） staining was used to observe the pathological changes in muscle tissue. Western blot was used to detect the protein expression of p38 mitogen-activated protein kinase （p38 MAPK）， phosphorylation （p）-p38 MAPK， extracellular signal-regulated kinase1/2 （ERK1/2）， nuclear factor-κB （NF-κB）， p-NF-κB， interleukin-1β （IL-1β）， and interleukin-6 （IL-6） in muscle tissue. The immunofluorescence （IF） method was used to detect the expression of tumor necrosis factor-α （TNF-α） in skeletal muscle tissue of mice in each group. ResultCompared with the control group， the body mass of mice in the model group decreased， and the time of last swimming due to exhaustion decreased （P<0.01）. In addition， there were significantly higher serum levels of the fatigue metabolites LA， LDH， BUN， and lipid peroxidation product MDA （P<0.01） and decreased levels of MG， HG， SOD， and GSH-Px （P<0.01）. The protein expressions of p-p38 MAPK， ERK1/2， p-NF-κB， IL-1β， IL-6， and TNF-α in skeletal muscle tissue were significantly higher than those of the control group （P<0.01）. Compared with the model group， the body mass and time of last swimming due to exhaustion of the mice in the low， medium， and high dose groups of Epimedii Folium polysaccharides and the vitamin C group were increased （P<0.05， P<0.01）， and the contents of LA， LDH， BUN， and MDA were significantly decreased （P<0.05， P<0.01）. The levels of MG， HG， SOD， and GSH-Px increased （P<0.05， P<0.01）， and the protein expression levels of p-p38 MAPK， ERK， p-NF-κB， IL-1β， IL-6， and TNF-α in skeletal muscle tissue decreased （P<0.05， P<0.01）. ConclusionEpimedii Folium polysaccharides can play a role in alleviating exercise-induced fatigue by inhibiting the p38 MARK/NF-κB signaling pathway， thereby reducing the accumulation of metabolites， improving the activity of antioxidant enzymes， increasing the glycogen content of the body， and reducing inflammation in skeletal muscle.