Objective To investigate the prevalence rate of insomnia and its influence factor in adults from Bozhou region,and determine the correlation of sleep hygiene with insomnia in adult population.Methods Eight hundred and three subjects were recruited in the study by multi-stage stratified random sampling from people aged 18 years or more than 18 years.Athens Insomnia Scale (AIS)and sleep hygiene practices scale were used to evaluate the insomnia and its related factors.Results One hundred and ninety-two adults(23.9％)were identified as insomnia with dissatisfaction of sleep quality,shortage of sleep time(＜56.5 h/night)and earlier wake-up than expectation as its main manifestations; and the prevalence rate ofinsomnia in females(98/354,27.7 ％)was significantly higher as compared with that in males(94/449,20.9％,P＜0.05); prepared military personnel enjoyed the highest insomnia rate(37/99,37.3％),followed by freelance/temporaries(55/181,30.4％)with significant difference between these 2 groups(P＜0.05).The correlation analysis between the total scores of AIS and problems of sleep hygiene indicated that such factors as prescription of sleep medication,hunger at bed,drinking in purpose of helping sleeping,and worry about insomnia beofore sleep or at daytime were positively related to the total scores ofAIS(P＜0.05); nap or snooze was negatively related to the total scores of AIS(P＜0.05).Conclusion There is a high proportion of insomnia in prepared military personnel and freelance/temporaries from Bozhou region; poor sleep hygiene habits as prescription of sleep medication,hunger at bed,drinking in purpose of helping sleeping,and worry about insomnia beofore sleep or at daytime can induce insomnia.

BACKGROUND AND OBJECTIVECytokine-induced killer (CIK) cells and autologous dendritic cells-CIK (DC-CIK) cells co-cultured with autologous dendritic cells (DCs) and CIK cells are commonly used for immunotherapy recently. We compared the anti-tumor immune response of CIK cells, autologous DC-CIK cells, and semi-allogeneic DC-CIK cells to explore a more effective anti-tumor adoptive immunotherapy approach.

METHODSPeripheral monocytes were isolated from patients with renal carcinoma, lung cancer, or maxillary squamous cell carcinoma and their healthy adult children. Isolated cells were cultured and induced as DCs and CIK cells in vitro. CIK cells from patients were co-cultured with autologous DCs and DCs from their children respectively, generating DC-CIK cells and semi-allogeneic DC-CIK cells. The anti-tumor activities of autologous CIK cells, autologous DC-CIK cells, and semi-allogeneic DC-CIK cells were measured by LDH assay. Intracellular staining was used to test the secretion of cytokines. Flow cytometry was applied for detecting the phonotype changes of these three types of cells. Cell proliferation and cell apoptosis were detected by 5,6-carboxyfluorescein diacetate succinimidyl ester (CFSE) and Annexin V/PI respectively.

RESULTSCompared with autologous CIK cells and DC-CIK cells, semi-allogeneic DC-CIK cells significantly enhanced the anti-tumor activity and IFN-gamma secretion, reduced IL-4 secretion, increased the ratio of CD3(+)CD56(+) cells and CD3(+)CD8(+) cells, decreased the number of CD4(+)CD25(+) cells, promoted cell proliferation, and lessened cell apoptosis.

CONCLUSIONSSemi-allogeneic DC-CIK cells had a stronger anti-tumor effect than did autologous CIK cells and DC-CIK cells. Our results provided experimental evidence for clinical application of DC-CIK cells.

Apoptosis ; Carcinoma, Squamous Cell ; metabolism ; pathology ; Cell Line, Tumor ; Cell Proliferation ; Cells, Cultured ; Coculture Techniques ; Cytokine-Induced Killer Cells ; cytology ; immunology ; metabolism ; Cytokines ; metabolism ; Cytotoxicity, Immunologic ; Dendritic Cells ; cytology ; immunology ; metabolism ; Hep G2 Cells ; Humans ; Immunotherapy, Adoptive ; Interferon-gamma ; secretion ; Interleukin-4 ; secretion ; K562 Cells ; Kidney Neoplasms ; metabolism ; pathology ; L-Lactate Dehydrogenase ; metabolism ; Lung Neoplasms ; metabolism ; pathology ; Maxillary Neoplasms ; metabolism ; pathology

In this study, UPLC-Q-Exactive-MS/MS was used to rapidly analyze the chemical constituents of Meconopsis quintupli-nervia, and the anti-liver fibrosis mechanism of M. quintuplinervia was preliminarily analyzed by network pharmacology, molecular docking, and cell experiments. The chemical constituents of M. quintuplinervia were identified according to the information of MS~1 and MS~2, as well as the data in the literature and databases. SwissTargetPrediction and TargetNet were used to predict the potential targets. The targets related to liver fibrosis were collected from GeneCards and OMIM. The protein-protein interaction(PPI) network was constructed by STRING. Cytoscape 3.6.1 was used to construct and analyze the "constituent-target-disease" network to obtain key targets and their corresponding constituents in the network. DAVID 6.8 was used for GO analysis and KEGG signaling pathway enrichment analysis. Finally, the preliminary verification was carried out by molecular docking and cell experiments. As a result, 106 chemical constituents were identified from M. quintuplinervia, including 66 flavonoids, 16 alkaloids, 18 phenolic acids, 1 anthocyanin, and 5 other constituents. Among them, 3 constituents were identified as potential new compounds, and 59 constituents were reported in M. quintuplinervia for the first time. Network pharmacology analysis showed that M. quintuplinervia presumably acted on AKT1, SRC, JUN, EGFR, STAT3, HSP90 AA1, MAPK3, and other core targets through luteolin, isorhamnetin, quercetin, apigenin, kaempferide, amurine, 2-methylflavinantine, allocryptopine, the multi and other active compounds, thereby regulating the PI3 K/AKT signaling pathway, pathways in cancer, proteoglycans in cancer, FoxO signaling pathway, and other pathways to exert anti-liver fibrosis effects. M. quintuplinervia extract(MQE) could significantly down-regulate PI3 K and AKT protein levels in the HSC-T6 cell model induced by TGF-β1, suggesting that MQE may have the ability to regulate the PI3 K/AKT signaling pathway. The findings of this study indicated that the anti-liver fibrosis effect of M. quintuplinervia had multi-constituent, multi-target, and multi-pathway characteristics, which may provide a scientific basis for the research on the pharmacodynamic materials, action mechanism, and quality markers of M. quintupli-nervia.
Tandem Mass Spectrometry ; Molecular Docking Simulation ; Network Pharmacology ; Proto-Oncogene Proteins c-akt ; Papaveraceae ; Liver Cirrhosis ; Drugs, Chinese Herbal/pharmacology*

OBJECTIVE: To establish the technique that take the advantages of flow cytometry combined fluorescence in situ hybridization (Flow-FISH) to identify the Epstein-Barr virus(EBV) infected lymphocyte subtypies in patients' peripheral blood sample. METHODS: Peripheral Blood monocyte from 9 patients with EBV infection enrolled at Children's Hospital in Chongqing Medical University were isolated by Ficoll-paque centrifugal separation. The expressions of EBER1, EBER2 in cell were detected by qRT-PCR. The surface markers of cell were detected by Flow cytometry after staining with their antibodies. The cell was treated Fix-Permeabilization Buffer before hybridization with fluorescent labeled probe at 37 ℃ overnight. The cell status, surface markers and targeted mRNA are detected by flow cytometry and fluorescence microscope. RESULTS: It was optimized that the Fix-Permeabilization Buffer and recipe with 0.2% Tween-20 were picked out as providing a good cell integrity and high resolution of surface markers. Hybridization with 20% formamide and 7% dextran sulfate at 37 ℃ overnight is the optimal hybridization condition as a good hybridization effect, a detectable cell integrity and a high resolution of cell markers under flow cytometry detection. Finally, upon the established Flow-FISH method, lymphocyte subpopulations of the EBV⁺ cells from cell lines and blood samples of patients were identified successfully. CONCLUSION A Flow-FISH technology is established, which can be applied in the identification of EBV infected cell subtypes. This research provides a foundmental for its application in clinical test in EBV⁺ related proliferative diseases.
Epstein-Barr Virus Infections ; Flow Cytometry/methods* ; Herpesvirus 4, Human ; Humans ; In Situ Hybridization, Fluorescence/methods* ; Lymphocyte Subsets

OBJECTIVE: To investigate the function of RAS protein on the progression of the T-ALL cell lines in vitro. METHODS: The DNA of the T-ALL cells was purified then amplified the coding regions of three RAS genes (KRAS, NRAS, HRAS) by PCR reaction. After T-A cloning, the coding regions of KRAS, NRAS and HRAS were sequenced by Sanger Sequencing. The siRNA oligonucleotides were cloned into the pSEH-361 vector, which were then packaged into retroviral together with pAMPHO and pVSVG in the HEK-293T cells. The T-ALL cells were infected with the retrovirus. The gene expressions were detected by qRT-PCR and Western blot. The T-ALL cells were stained with Annexin V-PE/7-AAD and the apoptotic cells were detected by flow cytometry. The T-ALL cells were stained with Hoechst 33258, and the cell cycle distribution was determined by flow cytometry. The expression of cleaved-Caspase 3 was stained with antibody and observed with fluorescence microscope. RESULTS: For RAS genes, beside the Loucy and the P12-ICH cells harbored KRAS c.6187G>A (p.KRAS^G12D) homozygous mutant, no missense mutation of RAS was found in other T-ALL cells genome. The pan RAS inhibitor compound 3144 showed toxicity to all tested T-ALL cells, except PEER (IC₅₀=47.916 μmol/L). Similarly, Tipifarnib induced apoptosis of multiple T-ALL cell lines except for the PEER cells (IC₅₀=94.2265 μmol/L). After KRAS knock-down, the T-ALL cells showed significant apoptosis and an arrested cell cycle. CONCLUSION The KRAS protein is vital for the progression of the T-ALL cells in vitro, it is a potential therapeutic target for T-ALL patients.
Apoptosis ; Cell Line ; Cell Proliferation ; Humans ; Mutation ; Precursor T-Cell Lymphoblastic Leukemia-Lymphoma ; Proto-Oncogene Proteins p21(ras)/genetics*