OBJECTIVE: To investigate expression status of p-mTOR(Ser2448,an active form of mTOR)in colorectal cancer (CRC),and its correlation with clinicopathological features and prognosis. METHODS: Protein expression of p-mTOR was examined using the tissue microarrays-immunohistochemistry (TMA-IHC) technique in surgically-resected colorectal cancer tissues and adjacent normal epithelia from 441 patients. Correlations between p-mTOR expression levels,clinicopathological parameters and prognosis of patients were analyzed using the Pearson Chi-square and Fisher exact tests,and Kaplan-Meier survival and Cox regression analyses. RESULTS: Overexpression of p-mTOR was detected in 58% (254/441) of the CRC specimens but only 4% (17/441) of the normal colorectal epithelia,and the difference between them was significant (P < 0.01). In addition,elevated p-mTOR expression was positively correlated with higher pathologic T stage,lymph node metastasis,distant metastasis,and advanced clinical stage (each P < 0.05). Kaplan-Meier survival analysis showed the expression level of p-mTOR was significantly correlated with the poor prognosis of colorectal cancer patients (each P < 0.05). The disease-free survival (DFS) and overall survival (OS) of patients with high expression of p-mTOR were 52 and 60 months,respectively,which were significantly shorter than those with low expression of p-mTOR (71 and 70 months). The Cox multiple regression analyses reveal that p-mTOR overexpression was an independent prognostic factor for DFS and OS in CRC patients. CONCLUSION: p-mTOR was frequently upregulated in CRC,and its overexpression was significantly correlated with adverse outcomes;thus,it may serve as a potential prognostic marker for predicting the risk of postoperative death,recurrence and metastasis in patients with colorectal cancer.

OBJECTIVE: To screen for radiation-sensitive metabolites in rat plasma and to explore the metabolic pathways that provide scientific basis for identification of radiation-sensitive biomarkers. METHODS: Rats irradiated whole-body with ⁶⁰Co gamma rays of 0,1,3 and 5 Gy,respectively. Changes of metabolites in rat plasma were detected by non-targeted metabolomics method based on the liquid chromatography mass spectrometry platform. RESULTS: Eleven plasma metabolites were identified as the potential radiation biomarkers. Taurine and hypotaurine metabolism,nicotinic acid and nicotinamide metabolism,and primary bile acid biosynthesis were enriched after irradiation. Linoelaidylcarnitine,11Z-octadecenylcarnitine,L-palmitoylcarnitine,taurine,niacinamide,L-kynurenine and cytosine were decreased with the increased dose and had good dose-response relationships. CONCLUSION: The metabolites levels in rat plasma were significantly changed after exposure to gamma-rays. Six metabolites had the basic conditions for radiation biological dosimeters.

OBJECTIVE: To investigate whether butein protects against LPS-induced inflammation in primary bone marrow macrophages,and to analyze the role of JAK2-STAT3 in this process. METHODS: In order to obtain bone marrow-derived macrophages (BMDM),bone marrows samples from C57BL/6 mice were treated with 40 ng/mL GM-CSF for 7 d. BMDM cells were stimulated with 500 ng/mL LPS for 12 h in the presence or absence of 5,10,20 μmol/L butein. Levels of TNF-α,IL-6 and NO in the culture medium were assessed by ELISA specific kits. To detect intracellular levels of reactive oxygen species (ROS),BMDM were stained with DCFH-DA and analyzed by flow-cytometry. Moreover,Western blot assay was used to detect protein expression of iNOS,p-JAK2,JAK2,p-STAT3 and STAT3. Ratios of p-JAK2/JAK2 and p-STAT3/STAT3 were determined. RESULTS: Exposure to LPS significantly increased the levels of TNF-α,IL-6 and NO in the culture medium of BMDM cells. However,administration of 5,10 and 20 μmol/L butein inhibited the production of these proinflammatory factors in a dose-dependent manner. Flow cytometry assays indicate that butein attenuated ROS generation in LPS-activated BMDM macrophages. In addition,Western blot results suggest that LPS promoted iNOS,p-JAK2 and p-STAT3 protein expression,and butein blunted this process. CONCLUSION: Butein inhibited LPS-induced inflammation in BMDM cells by regulating expressions in the JAK2-STAT3 pathway. Therefore,butein may be useful as a drug for control of inflammatory diseases.

OBJECTIVE: To study the clinical significance of miR-429 and Bmi-1 mRNA in Diffuse large B-cell lymphoma (DLBCL) tissues. METHODS: Samples of tumor tissues were collected from 45 cases of the DLBCL patients from January 2014 to December 2015,which were compared with 28 normal lymph node tissues. Expressions of miR-429 and Bmi-1 mRNA in tumor tissues and normal lymph node tissues were detected by Reverse transcription quantitative polymerase chain reaction (RT-qPCR). Their relationships with the clinical features of the patients were analysed. RESULTS: The tumor tissues showed significantly decreased expression of miR-429 and increased expression of Bmi-1 mRNA,compared with the normal lymph node tissues. DLBCL patients with low expression of miR-429 and high expression of Bmi-1 tend to have later clinical stages,more prone to bone marrow metastasis,higher Ki-67 positive rate and higher IPI score than patients with high expression of miR-429 and low expression of Bmi-1 mRNA (P < 0.05). CONCLUSION: The results show that the expression levels of miR-429 and Bmi-1 in DLBCL tissues were closely related to the progression and prognosis of the lymphomas. Lower miR-429 expression and higher Bmi-1 mRNA expression in tumor tissues may be the predictors of poorer prognosis.

OBJECTIVE: To investigate differences in induction of cytotoxicity,inflammatory factor release level and apoptosis between two cigarettes with different tar releases. METHODS: The amounts of tar release under the suction parameters as specified by the International Organization for Standardization (ISO) were 9.4 mg per cigarette in sample 1 and 14.0 mg per cigarette in sample 2. Beas-2b cells were divided into clean air control and smoke-exposed groups. The clean air or diluted mainstream smoke generated by the VITROCELL system was used to expose the cells in the form of an air-liquid interface. After calculation,the dose for the clean air control group was 0 mainstream smoke and the doses of the smoke-exposed group were:mainstream smoke from 0.12%,0.27%,0.57%,and 1% cigarette. The neutral red uptake cytotoxicity test was used to detect the cell viability;the ELISA method was used to detect the IL-6 and IL-8 expression levels;the Annexin V-FITC/PI flow cells analytical method was used to detect the level of apoptosis. RESULTS: Both of the mainstream smoke produced by samples 1 and 2 reduced survival rates of Beas-2b cells,and significantly increased the release levels of inflammatory factors IL-6 and IL-8 and the apoptosis rates. At the 0.12% cigarettes smoke dose,there was no statistically significant difference in the cytotoxicity,inflammatory factor release level and apoptosis rate caused by the smoke of samples 1 and 2. At the 0.57% and 1% cigarette smoke doses,sample 2 had more significant cytotoxicity than sample 1. CONCLUSION: Both cigarette samples with different tar releases caused cytotoxicity,pro-inflammatory factor release,and apoptotic effects. In the range of 0.57% to 1%,cigarette smoke with high tar release was more cytotoxic than cigarette smoke with low tar release.

OBJECTIVE: To investigate the inhibitory effects of ethanol extract from Euphorbia humifusa(EEEH) on activities of BGC823,MGC803,SGC7901 and AGS in human gastric cancer (GC) cells. METHODS: The EEEH was prepared by distillation and freeze-drying technology. The GC cells were treated with different concentrations of EEEH (0,62.5,125,250 and 500 μg/mL) and the solvent control. The cell survival rates were detected by the MTS method,the apoptosis rates and cell cycle distributions were detected by flow cytometry,and the protein expression levels of cell proliferation,apoptosis and cycle were detected by Western blot method. RESULTS: After treatment with different concentrations of EEEH,the proliferation of GC cells was significantly inhibited compared with the control group and the difference was statistically significant (P < 0.05). The results of flow cytometry show that the apoptosis rates of treated AGS cells increased significantly and the cell cycle was arrested in G0/G1 phase. The differences were significant from that of the control (P < 0.05). The results of Western blot test show that the expression levels of proliferation-related protein PCNA,anti-apoptosis-related protein BCL-2 and BCL-XL in the treated cells were significantly lower than those in the control group,and apoptosis-related protein caspase-8 and caspase-3 showed obvious lytic fragments,indicating increased activities. The differences were statistically significant (P < 0.05). However,there were no significant changes in the expression levels and activities of caspase-9 protein. CONCLUSION: EEEH inhibited GC cell proliferation via the induction of apoptosis and cycle arrest.

OBJECTIVE: To analyze the expression of long non-coding RNA (lncRNA) in esophageal squamous cell carcinoma (ESCC) and adjacent tissues of Kazakh nationality in Xinjiang. METHODS: Differentially expressed lncRNAs in four groups of cancer and a control group were screened by gene chip technology,and the target genes were predicted by lncRNA-mRNA co-expression network. GO and KEGG databases were used for enrichment and Pathway analyses to clarify the function of lncRNA. RESULTS: A total of 318 lncRNAs were differentially expressed between the cancer and control groups. The expression levels of 193 lncRNAs were higher for the cancer than the control groups,and 125 lncRNAs were lower. Enrichment analyses show that there were 5 GO categories,including 3 cellular components,1 biological process and 1 molecular function,all of which were target genes regulated by trans-regulation. Pathway analyses show that these differentially expressed genes might be involved in two signal pathways,such as axonal guidance signal pathway and ECM receptor interaction pathway. CONCLUSION: Expression levels of lncRNAs in Kazakh esophageal squamous cell carcinoma were significantly different from that of the control group. The differentially expressed lncRNAs may be involved in the complex regulatory network of cancer and has a certain relationship with tumorigenesis and progression.

OBJECTIVE: To analyze relationships between NFKBIA gene polymorphism,eating habits and susceptibility to gastric cancer. METHODS: 1:1 matched case-control study was carried out with 587 newly-diagnosed primary gastric patients and 587 controls matched by sex and age in the Fujian Xianyou hospital from April 2013 to June 2017. The data of general information and eating habits were collected by a self-made questionnaire. The Sequenom MassARRAY SNP detection method was used to detect the genotype of NFKBIA gene rs696 locus. Condition logistic regression was used to analyze the effects of eating habits and genotypes on risk of gastric cancer. Forked analysis,logistic regression model and Excel table compiled by Andersson were used to evaluate the interactive effects of NFKBIA polymorphism and eating habits. RESULTS: Fast eating,irregular meals,high salt diet and often pickle intakes were the risk factors of gastric cancer. OR(95%CI) were 1.53(1.20,1.95),1.55(1.16,2.08),1.51(1.17,1.94),2.78(2.01,3.85) respectively. The NFKBIA rs696 locus AG genotype,AA genotype and dominant model (AG+AA) increased the risk of gastric cardia cancer. In addition,the interaction effects were found on rs696 AA genotype with high salt diet and pickle intakes. The interactive effects were also found on the rs696 dominant model (AG+AA) with fast eating,high salt diet and often intake pickles. CONSLUSION:Bad eating habit was a risk factor of gastric cancer. The NFKBIA rs696 AA genotype,the dominant model (AG+AA) increased the risk of gastric-cardia cancer and had an interaction effect with eating habits.

OBJECTIVE: To investigate the effects of SF1a-PRLR and its variant ΔS2 SF1a-PRLR over-expression on proliferation and microRNA expression in human breast cancer cells (MCF-7). METHODS: SF1a-PRLR and ΔS2 SF1a-PRLR cDNAs were recombined separately into lentivirus by using homologous recombination strategy,and the viruses carrying different genes (including SF1a-PRLR,or ΔS2 SF1a-PRLR cDNA) or not carrying any target gene (control) were transfected into MCF-7 cells,respectively. The stably transfected cells including MCF7-con (control group),MCF7-SF1a-PRLR (SF1a),and MCF7-ΔS2 SF1a-PRLR (ΔS2 SF1a) were obtained through multiple screening with puromycin. Total RNAs were extracted using Trizol reagent,and high-throughput sequencing of small RNAs were performed. In addition, separate sets of the cells were tested using the MTS cell proliferation assay. RESULTS: Cell proliferation assay results show that at 48 h,the D(492) values of the control,SF1a and ΔS2 SF1a groups were 1.85±0.29,2.24±0.26 and 2.57±0.23,with significant differences among them (all P < 0.05). Furthermore, sequencing data show that there were significant differences on microRNA expression among the groups. SF1a induced 176 microRNAs expression changes(32 were up-regulated and 144 were down-regulated) and ΔS2 SF1a induced 206 microRNAs expression changes (52 were up-regulated and 154 were down-regulated). For comparison,4 microRNAs (miR-4454,miR-215-5p,miR-622 and miR-797) were up-regulated and 1 (miR-210-5p) was down-regulated from ΔS2 SF1a-PRLR over-expression. The relative expressions of miR-210-5p in the control,SF1a and ΔS2 SF1a groups were 66.18、31.67 and 13.07,with significant differences among the three groups (all P < 0.05). CONCLUSION: Both SF1a-PRLR and ΔS2 SF1a-PRLR over-expression promoted proliferation of breast cancer cells (MCF-7),and significantly affected the expression of microRNAs in a similar way for most of the detected genes. Several microRNAs such as miR-4454,etc. are affected in a different mode by SF1a PRLR and ΔS2 SF1a-PRLR.

OBJECTIVE: To explore the effect of Danzhijiangtang capsule on pathological changes of heart and kidney in diabetic rats. METHODS: 70 male SD rats,randomly selected 10 as normal group,the average feed,the rest 4 weeks rats a high-fat feed,intraperitoneal injection of chain urea with streptozotocin (STZ) 34 mg/kg copy type Ⅱ diabetic rat model,building is not successful rats intraperitoneal injection of STZ injection 25 mg/kg. The model rats were divided into four groups of 10 rats per group according to blood glucose and volume:model group,high-dose Danzhijiangtang capsule group[1.08 g/(kg·d)],low-dose Danzhijiangtang capsule group[0.54 g/(kg·d)],and pionlitazone group[10 mg/(kg·d)]. The normal group and the model group were given the same volume of normal saline. Blood glucose,blood lipid,renal function (BUN,Scr,mAlb,urinary protein) were detected by automatic biochemical analyzer. Cardiac tissues were stained with HE and MASSON,and renal tissues were stained with HE,MASSON and glycogen PAS. Pathological changes of tissues were investigated. RESULTS: Compared with the normal group,the model group rats showed decreased volume,significantly increased food intake and water intake,blood sugar,blood lipid and renal function index (P < 0.01). In addition,they showed visible infiltration of heart inflammatory cells,increased myocardial fibrosis,changes in kidney baumann's capsule structure,and significantly increased cytoplasmic matrix and the degree of fibrosis. After 6 weeks of administration and compared with the model group,the volume of pioglitazone group and the high-dose Danzhijiangtang capsule group significantly increased,the food intake and water intake decreased,and the indexes of blood glucose,blood lipid and kidney function decreased (P < 0.05,P < 0.01). In addition,the pathological changes of heart and kidney slowed down. Although the indexes of blood glucose,blood lipid and renal function of rats in the low dose danzhu hypoglycemic capsule group decreased,there was no significant difference from the TC level (P > 0.05). However,there were statistical differences between the levels of HbAlc,TG,TC and mAlb and those in the pyrrolitazone group (P < 0.05,P < 0.01). CONCLUSION: Administration of either high or low dose Danzhijiangtang capsules effectively reduced blood glucose and lipid levels in rats,relieved the symptoms of polyphagia and polydipsia,and reduced the damage of hyperglycemia in myocardial and renal tissues of rats.

OBJECTIVE: To investigate correlations between toxicity in HepaRG cells and free anthraquinone contents in aqueous extracts from Polygoni Multiflori Radix (PMR),Polygoni Cuspidati Rhizoma et Radix (PCRER) and Rhei Radix et Rhizoma (RRER). METHODS: High performance liquid chromatography was used to detect the content of emodin,physcion,aloe emodin,rhein and chrysophanol from the three aqueous extracts. MTT assay was employed to measure HepaRG cell viability following treatment with theextracts in different concentrations (1.0,1.5,2.0,2.5,3.0 mg/mL) for 48 h. HepaRG cells were also treated in concentrations of 1.5,2.0,2.5 mg/mL for the detection of apoptosis by AnnexinⅤ/PI dual staining for FCM. RESULTS: The contents of free anthraquinoneins in the three extracts were 0.229%,0.545%,2.593% respectively. The survival rate of HepaRG cells was down-regulated by PMR (2.0,2.5,3.0 mg/mL),PCRER (1.5,2.0,2.5,3.0 mg/mL) and RRER (1.0,1.5,2.0,2.5,3.0 mg/mL) in dose-dependent manners. The degree of inhibition was positively correlated with the content of free anthraquinones. The same trend was observed in apoptosis cell proportion in which the concentrations of extracts were 1.5,2.0,2.5 mg/mL. CONCLUSION: The contents of free anthraquinones in the three aqueous extracts from low to high degree were PMR,PCRER and RRER. The content was positively correlated with cytotoxicity in vitro,which suggests that free anthraquinones might be the important component inducing toxicity in these three traditional Chinese medicines.

OBJECTIVE: To explore the expression and function of aurora kinase A (AURKA) in esophageal cancer (ESCA) and its prognostic value in patients. METHODS: Differences of AURKA expression in ESCA tissues and normal esophagus tissues were analyzed using GEPIA and Oncomine databases. A protein-protein interaction (PPI) network about AURKA was constructed based on STRING online database and hub genes were screened with cytoHubba application of cytoscape. Correlation expressions between AURKA and hub genes were analyzed using TCGA and GTEx datasets. GO function and KEGG pathway enrichment analyses were carried out using DAVID online database. The R2 platform was used to study the association of AURKA expression and overall survival (OS) of ESCA patients. RESULTS: Expression of AURKA in ESCA tissues was significantly higher than that in normal tissues (P < 0.05),but it was unrelated to disease stages (P > 0.05). AURKA was positively correlated with CDC20 (r=0.78),PLK1 (r=0.83) and other hub genes,and their functions were mainly enriched in cell differentiation,DNA damage detection points,cell proliferation,and negative regulation of cell apoptosis. These were involved in the regulation of cell cycle,cell senescence,P53 and FoxO signaling pathways. The AURKA high expressed group was significantly associated with poor prognosis (P < 0.05),similarly the high expressions of CCNB1,BUB1B and NEK2 were significantly associated with low survival rates (all P < 0.05). CONCLUSION: AURKA was up-regulated in ESCA significantly and associated with poor prognosis based on the Oncomine and GEPIA database analyses. It was also involved in tumor-related functions and pathways together with CCNB1,NEK2 and other hub genes. Therefore,it can be used as a biomarker for early diagnosis,treatment and prognosis of patients with ESCA.

OBJECTIVE: To find pathogenic gene(s) in a clinically diagnosed familial hypercholesterolemia (FH) family using whole exome sequencing. METHODS: Peripheral venous blood of the family members was taken to measure serum TC,TG,LDL cholesterol and HDL cholesterol. DNA of the white blood cell was extracted to perform whole exome sequencing. Four genes (LDLR,APOB,PCSK9,LDLRAP1) associated with FH were chosen to analyze single nucleotide polymorphisms (SNP) and predict the function of protein using Polyphen-2 and SIFT. RESULTS: Four SNPs (rs676210,rs679899,c.10094A > T and c.9937C > G) of APOB might be associated with the elevated lipid levels,but no co-segregating variants were found in this family. No damaging variant was found in LDLR,PCSK9 and LDLRAP1 by Polyphen-2 and SIFT. CONCLUSION: Our study found that some SNPs of APOB may be associated with elevated lipid levels in a clinically diagnosed FH family. The function of these SNPs still needs further experiments to confirm.

OBJECTIVE: Comet assays were conducted on Sprague Dawley rats treated with four chemicals with different mechanisms of action:p-chloroaniline,sodium chloride,acetaminophen and gentamicin. METHODS: Rats were orally exposed to p-chloroaniline[37.5,75 and 150 mg/(kg·d)],sodium chloride[500,1 000 and 2 000 mg/(kg·d)],acetaminophen[20,100 and 500 mg/(kg·d)],vehicle controls,and intramuscular-injected gentamicin[20,40,and 80 mg/(kg·d)] for three consecutive days (0,24 and 45 h),as well as orally treated with the positive control (EMS,200 mg/kg) for 24 and 45 h. Liver,stomach,and kidney were sampled approximately 3 h after the last dosing. After preparation of single cell suspensions,slides were prepared for lysis,unwinding,electrophoresis,neutralization,and DNA staining for comet visualization and image analysis. Histopathological examination of liver and kidney was respectively conducted for the two target-organ toxicity chemicals (acetaminophen and gentamicin). RESULTS: Among the four chemicals,p-chloroanilin produced positive results in liver,stomach,and kidney,while sodium chloride,acetaminophen,and gentamicin were all negative in liver,stomach,and kidney. Inflammation,degeneration,or necrosis of liver was noted in all the acetaminophen dosing groups. CONCLUSION: Collectively,known genotoxic compounds and non-genotoxic compounds with and without target-organ toxicity were used to verify the sensitivity and specificity of the multi-organ comet assay,which is expected to provide technical reference for the popularization of alkaline comet assay in vivo in the field of genotoxicity evaluation for new drugs.