RESUMO
Objective:To explore whether the lipopolysaccharide (LPS)-induced modification of O-linked N-acetylglucosamine (O-GlcNAc) is involved in the inflammatory signaling pathway of endothelial cells.Methods:Human umbilical vein endothelial cells (HUVEC) were cultured in vitro, and cells in logarithmic growth phase were used for experiments. Cells were divided into blank control group, LPS group (2 000 mg/L LPS), O-GlcNAc transferase (OGT) overexpression (OGT-OE)+LPS group (plasmid transfection OGT+2 000 mg/L LPS), protein kinase C (PKC) inhibitor+LPS group (10 μmol/L Go 6983+2 000 mg/L LPS), RhoA inhibitor+LPS group (40 μmol/L Rhoin hydrochloride+2 000 mg/L LPS), phosphatidylinositol-3-kinase (PI3K) inhibitor+LPS group (1 μmol/L SL-2052+2 000 mg/L LPS), serine/threonine kinase (Akt) inhibitor+LPS group (10 μmol/L PP2+2 000 mg/L LPS) and small interfering RNA (siRNA) treated Akt (si-AKT)+LPS group (si-Akt+2 000 mg/L LPS). After 24 hours of LPS treatment, real-time fluorescence quantitative reverse transcription-polymerase chain reaction (RT-qPCR) was used to detect the transcription levels of inflammatory cytokines [interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1)]. The protein expression or phosphorylation of OGT, O-GlcNAc, Akt, extracellular signal-regulated kinase (ERK), p38 mitogen-activated protein kinase (p38MAPK), nuclear factor-κB p65 (NF-κB p65), and signal transducer and activator of transcription 3 (STAT3) were determined by Western blotting. Results:Compared with the blank control group, the expression of OGT and the modification of O-GlcNAc in the LPS group were decreased, while the expressions of phosphorylated ERK, p38MAPK, and STAT3 were increased, and the transcript levels of inflammatory cytokines were also significantly increased [IL-6 mRNA (2 -ΔΔCt): 4.71±0.60 vs. 1.03±0.29, TNF-α mRNA (2 -ΔΔCt): 1.89±0.11 vs. 1.04±0.35, ICAM-1 mRNA (2 -ΔΔCt): 2.06±0.18 vs. 1.02±0.21, VCAM-1 mRNA (2 -ΔΔCt): 2.94±0.57 vs. 1.01±0.17, all P < 0.05], indicating that LPS could decrease O-GlcNAc modification, activate inflammatory signaling pathways and increase inflammatory cytokines expression. Compared with the LPS group, the expressions of phosphorylated ERK, p38MAPK, NF-κB p65, and STAT3 in the endothelial cells of the OGT-OE+LPS group were decreased, and the expression of inflammatory factors were significantly decreased [IL-6 mRNA (2 -ΔΔCt): 0.12±0.01 vs. 0.90±0.17, TNF-α mRNA (2 -ΔΔCt): 0.31±0.01 vs. 0.91±0.14, ICAM-1 mRNA (2 -ΔΔCt): 0.64±0.02 vs. 1.13±0.16, VCAM-1 mRNA (2 -ΔΔCt): 0.11±0.01 vs. 0.93±0.11, all P < 0.05], indicating that the increase of OGT level could inhibit the partial activation of the endothelial inflammatory signal pathway under the LPS stimulation. Compared with the blank control group, the phosphorylation level of Akt in the LPS group was increased. Compared with the LPS group, both OGT expression and O-GlcNAc modification were down-regulated after pretreatment of PKC inhibitor, RhoA inhibitor, PI3K inhibitor, or Akt inhibitor. Compared with the LPS group, the transcript levels of IL-6, TNF-α and ICAM-1 in the PP2+LPS group were significantly decreased [IL-6 mRNA (2 -ΔΔCt): 1.46±0.16 vs. 3.55±0.87, TNF-α mRNA (2 -ΔΔCt): 0.98±0.14 vs. 1.76±0.10, ICAM-1 mRNA (2 -ΔΔCt): 1.39±0.24 vs. 2.04±0.13, all P < 0.05], but there was no significant change in VCAM-1. Compared with the LPS group, the expression of OGT and O-GlcNAc modification in the si-Akt+LPS group were decreased, while the transcript levels of inflammatory cytokines were also significantly decreased [IL-6 mRNA (2 -ΔΔCt): 0.75±0.03 vs. 0.99±0.09, TNF-α mRNA (2 -ΔΔCt): 0.69±0.01 vs. 1.10±0.08, ICAM-1 mRNA (2 -ΔΔCt): 0.76±0.01 vs. 0.99±0.02, VCAM-1 mRNA (2 -ΔΔCt): 0.93±0.08 vs. 1.20±0.21, all P < 0.05], indicating that Akt participated in the action process of LPS on OGT and affected the inflammatory factor expression. Conclusions:The decreased level of O-GlcNAc modification in endothelial cells stimulated with LPS promotes partial activation of inflammatory signaling pathways, mainly involving ERK, p38MAPK, and STAT3, and affects the expression of inflammatory factors. AKT may be involved in the effect of LPS on the inhibition of O-GlcNAc modification.
RESUMO
Objective To investigate infection status and distribution characteristics of human immunodeficiency virus (HIV)infection among voluntary blood donors in Xiangyang area in 2009-2014,and provide evidence for re-cruiting blood donors from low-risk population. Methods Blood specimens of 326 856 voluntary blood donors in Xiangyang area between January 2009 and December 2014 were detected anti-HIV with enzyme linked immunosor-bent assay(ELISA),positive specimens in HIV screening were performed confirmatory testing,epidemiological analysis was conducted on the positive detection results.Results 41 (0.13‰ )voluntary blood donors were con-firmed positive for HIV in Xiangyang area,positive rate among each year was significantly different (χ2 =27 801.87,P<0.01). Except marital status and infection routes,difference in positive rates among donors of dif-ferent gender,age,area,occupation,educational attainments,and number of blood donation were all significantly different(all P<0.01). Among HIV-infected voluntary blood donors,87.80% were male,41.46% and 31.71%were aged between 18-25 and 36-45 respectively,56.10% were through heterosexual transmission,and 43.90%were through male homosexual transmission. The percentage of workman and public service personnel infected with HIV were high,accounting for 21 .95% and 17.07% respectively,60.97% of confirmed positive HIV were from city area,56.10% only received junior middle school/technical secondary school education,65.85% of HIV-infec-ted persons were infected at the initial donation.Conclusion In order to ensure blood safety,blood center should in-crease publicity efforts,carry out health consultation before blood donation,and recruit blood donors from low-risk populations,nucleic acid detection should be carried out to shorten HIV detection window period.
RESUMO
OBJECTIVE： To study the anti－ metastatic action of equal/unequal－ fork－ peptide of YIGSR.METHODS： To observe the influence of drugs on metastasis of melanoma B16 to lung in mice.RESULTS： The metastatic rate of lung in 46 mice of control and experimental groups was 100％ ,21 days after inoculating melanoma B16 cells via caudal vein.Injection of YIGSR in combination with tumor cells could reduce the number of metastatic nodules in a dose－ dependent manner.The numbers of metastatic nodules in 100μ g～ 200μ g equal fork peptide group and same dosage unequal fork peptide group were significantly different from that in control group(P<0.05;P<0.01 respectively).50μ g equal fork peptide group was different from control group.The synthetic peptide could not decrease the weight of the lung with metastatic lesions.CONCLUSION： YIGSR derivants have effective antimetastatic actions.Its mechanism may related to the inhibiting effect on formation of cancer cell emboli,which retain in microvessels of remote target organs,and multiplicate and penetrate blood vessels to form micrometastatic lesions.