Objective To explore the effect of HTLV-1 (human T-cell leukemia virus type 1 ) Tax protein on the DcR3 gene expression in T cells.Methods The construction of DcR3 (-1010 bp to +114 bp) luciferase reporter gene; MT2,TaxP,and Jurkat E6-1 cells were transfected with DcR3 luciferase reporter gene (pGL3-DcR3-1uc) using liposomes according to the manufacturer's instructions.For the control group,pGL3-basic replaced it respectively.At 48 h after incubation,luciferase activity was measured with a luciferase assay system; Jurkat cells were transfected with pCMV-Tax-Bam using liposomes,and total RNA was extracted from the cells at 48 h after incubation.Reverse transcription was performed using standard protocols.Then real time PCR with primers DcR3 and 3-actin was conducted;The expression of DcR3 protein was detected by flow cytometry in MT2,TaxP,and Jurkat cells.Results The construction of DcR3 luciferase reporter gene is identified correctly; The detection of luciferase activity showed that the luciferase activity of experimental group in MT2 cells was increased by (32.07±12.43)-fold,of which in TaxP cells was increased by ( 13.27±4.04)-fold,and the luciferase activity of experimental group in Jurkat cells was increased by ( 1.26±0.49 ) -fold.And there is statistic significance about the relative luciferase activity of experimental group in MT2 cells and TaxP cells compared to the relative lueiferase activity of experimental group in Jurkat cells respectively ( P＜0.01 ) ; The result of real-time PCR showed that the level of DcR3 mRNA in the experimental group was higher compared with the control group(P＜0.05) ; The result of FCM shows the expression of DcR3 protein in MT2,TaxP cells is higher than that in Jurkat cells( P＜0.05 ).Conclusion Tax protein can promote the expression of DcR3 gene in T cells.

Objective To evaluate the effects and underlying mechanisms of crocin on glutamate-induced apoptosis of retinal ganglion cells (RGCs) by affecting extracellular calcium influx.Methods Primary rat retinal ganglion cells were isolated and stimulated with glutamate at concentrations of 0.1 mmol/L and 1 mmol/L for 24 h or 48 h,respectively,to establish apoptosis model of RGCs.Afterwards,crocin of different doses (0.1,1.0 and 3.0 μmol/L) was used to treat the glutamate-induced RGCs for 12 h;then cell apoptosis was detected by Annexin V-FITC/PI staining.The intracellular calcium concentration was determined by FIuo-3/AM fluorescent labeling.Western blot was used to examine the effect of crocin on Ca2+-mediated apoptotic signal molecules calpain and CaMKII.The mitochondrial membrane potential was detected by JC-1 staining and mitochondrial apoptosis-related signaling molecules Caspase-3,Caspase-9 and Bcl-2/Bax were evaluated by Western blot,respectively.Results In comparison with the untreated controls,the cell apoptosis of RGCs exposed to 0.1 mmol/L of glutamate for 24 h did not significantly change (P＞ 0.05).However,apoptosis rate of the cells reached (43.050 ± 2.616) ％ when the stimulation time lasted for 48 h and showed a significant increase (P＜0.01).Treatment with higher-dose glutamate (1 mmol/L) significantly increased apoptosis of RGCs at 24 h (46.450±1.061)％ and 48 h (45.500±3.253)％ compared with the controls (P＜0.01).RGCs were induced by 1 mmol/L of glutamate for 12 h,followed by the treatment with crocin at concentrations of 0.1,1.0 and 3.0 μmol/L,respectively.Each dose of crocin could significantly inhibit cell apoptosis in the dose-dependent manner (P＜0.01).In addition,crocin at 1.0 μmol/L blocked glutamate-induced extracellular calcium influx,inhibited the expression of calcium-dependent proteins Calpainl and CaMK Ⅱ.Moreover,crocin at the dose of 1.0 μmol/L also increased mitochondrial membrane potential,suppressed the expressions of Caspase-3 and Caspase-9,and elevated Bcl-2/Bax ratio.Cornclusion Crocin inhibits glutamate-induced apoptosis of retinal ganglion cells through suppressing extracellular calcium influx,thereby blocking calcium-dependent and mitochondria-dependent apoptosis signaling pathways.

Objective To study the effects of the adult T cell leukemia virus type 1(HTLV-1) Tax protein on T lymphocytes self-reactive growth hormone(GH). Methods The expression of the growth hormone protein was measured by Western blot in TaxP and TaxN cells, which expresses HTLV-1 Tax or no. The location of growth hormone in the TaxP and TaxN cells were detected by LSCM(laser scanning confocal microscope). The level of growth hormone mRNA in TaxP and TaxN cells was measured by RT-PCR. The pGL2-GH-luc was transfected in TaxN and TaxP cells by Tfx-50-mediated transfection to assay transcriptional activity. The pNF-κB-luc, pAP-1-luc and pNFAT-luc was transfected into TaxP cells with pcDNA3.0-GH by Tfx50 to test bioactivity of the nuclear transcription factors. Results The mRNA and protein expression of GH could be promoted significantly by Tax. Relative to the TaxN cells, the transcriptional activity of GH was significantly increased about 6.37 times in TaxP cells(P＜0.05). Overexpression of GH can increase the activity of NF-κB about 1.7 times in TaxP cells (P＜0.05). Conclusion GH maybe involve in adult T-cell leukemia(ATL) through activating NF-κB.

Objective To standardize the selection of clinical research outcome indicators,which can objectively evaluate the clinical efficacy or effect of traditional Chinese medicine in the treatment of myasthenia gravis.This study aims to standardize the construction of the core outcome set of clinical research of traditional Chinese medicine in the treatment of myasthenia gravis.Methods We followed the core outcome set development specification(COS-STAD)to carry out research,established a research working group,which set up a Delphi-method advisory group.Two graduate students of working group conducted a document research and meetings of patients to establishe an outcome set item pool of myasthenia gravis in clinical trials of Chinese medicine under the instruction of other members.With the questionnaire based on the content of item pool,we then carried out Delphi-method expert consultations and a consensus meeting.Results The core outcome set of clinical research on myasthenia gravis treated with traditional Chinese medicine included five outcome domains:endpoint outcome,myasthenia gravis symptom evaluation,medication evaluation,quality of life evaluation and safety outcome;Nine outcome measures:recurrence rate,incidence of hormone complications,incidence of crisis,QMGS scale(MGFA quantitative myasthenia gravis score),daily activity scale of MG patients(ADL),analysis of immunosuppressant dosage,analysis of glucocorticoid dosage,analysis of cholinesterase inhibitor dosage,and incidence of adverse events.Conclusion The five outcome domains and nine outcome measures included in the core outcome set can be used as outcome options for the efficacy evaluation of myasthenia gravis clinical research.