Objective To detect the effect of osthole on proliferation and apoptosis of HL-60 cells and its molecular mechanism.Methods HL-60 cells proliferation was measured through the CCK8 assay method.The cell morphological changes were observed by Hoechst33342 staining after 8 h of drug effect.Induction of apoptosis was determined by flow cytometry and fluorescent microscopy.Expressions of Bcl-2 and Bax mRNA were evaluated by RT-PCR,and the expressions of cleaved caspase-3,caspase-8,caspase-9,Fas and FasL were evaluated by using western bolt assay.Results Osthole could inhibit the proliferation of HL-60 cells,the maximum inhibiting rate was (90.7 ±4.5)％,F =138.46,P =0.000; the apoptosis rate was 33.6％,F =27.75,P =0.006.The changes of apoptosis of cells and nucleus were shown in cell morphological observation.Osthole affected the decrease of the mRNA levels of Bcl-2 and the increase of the Bax mRNA levels via a dosedependent manner(F =210.12,P =0.000).Western blotting demonstrated that osthole could lead to the increase of the expression levels of cleaved caspase-3,caspase-8,caspase-9,Fas and FasL in the HL-60 cell line via a time-dependent manner.Conclusion Data suggests that osthole inhibits proliferation and induces apoptosis of HL-60 cells through mitochondria-dependent pathway and death-receptor pathway.

Objective To study the molecular regulation mechanism of VEGF in the model of ATRA induced differentiation in HL-60 cells,and to provide new targets for leukemia anti-angiogenic therapy.Methods The morphology was observed by Wright-Gimesa staining; HL-60 cells differentiation was detected by NBT reduction experiment.VEGF,STAT3,c-myc mRNAs were measured by reverse transcription-PCR;VEGF,STAT3 and c-myc proteins were determined by Western blot.Results The proliferation of HL-60 cells was inhibited obviously by ATRA(1 μmol/L) with the induction of differentiation,NBT positive rate was 82.59％ (t =-24.157,P ＜ 0.01) ; VEGF mRNA (t =7.339,P ＜ 0.05),STAT3 mRNA (t =3.667,P ＜0.05) and c-myc mRNA (t =6.858,P ＜ 0.05) were all down-regulated.VEGF protein (t =3.386,P ＜0.05),STAT3 protein(t =4.074,P ＜ 0.05) and c-myc protein (t =3.333,P ＜ 0.05) were all down-regulated.Conclusion VEGF expression level is reduced with the procession of differentiation of HL-60 cells,which may be largely correlated with the down regulation of STAT3 and c-myc.

Objective To explore the effect of vascular endothelial growth factor antisense oligonucleotides(AS-VEGF)on HL60 cell and HL60/VCR multidrug resistance cell and analyze the function of P-gp and the expression of related multidrug resistance genes including Bcl-2,Mcl-1,MDR1,MRP,GST?,TopoⅡ? and TopoⅡ?.Methods A vector AS-VEGF which expressed in eukaryotic cell was established,then transfected the vector into HL60 and HL60/VCR by limposome transfection technology,observed and drew the growth curve by Tapanlan taining,RT-PCR was used to detect the expression of Bcl-2,Mcl-1,MDR1,MRP,GST?,TopoⅡ? and TopoⅡ? in mRNA level after transfected 24 h and 48 h.Western Blot was used to detect the expression of P-gp in proteinum level after transfected 24 h and 48 h.Results The growth of HL60 and HL60/VCR was inhibited by AS-VEGF(1.25 mmol/L).Between HL60 and HL60/VCR,AS-VEGF decreased the expression of MDR1,MRP,GST? and TopoⅡ? but could not influence the expression of Bcl-2,Mcl-1 and TopoⅡ?,and the expression of P-gp was also obviously decreased in 48 h compared with that in 24 h.Conclusions AS-VEGF can inhibite the growth of HL60 and HL60/VCR and reverse multidrug resistance by changing cell microenvironment and the cell membrane correlated protein transportating channel,reduce the cell disintoxicating and the self-repair ability.

Objective To study the molecular mechanism of different sensitivities to apoptosis induced by low concentration of As2O3 in PML-RARα negative HL-60 cells and PML-RARα positive NB4 cells. Meth-ods NB4 and HL-60 cells were cultured with As2O3 for 1 to 4 days; cell proliferation were detected by MTT method; the apoptosis was detected by flow cytometry,Bcl-2,Bax and Fas mRNA were determined by RT-PCR. Results The proliferation of NB4 cells was inhibited obviously by As2O3(1.0 μmol/L)with the induction of apoptosis( P <0.05) ,which was accompanied by the down-regulation of Bcl-2 mRNA expression( P <0.05)and the ratio of Bcl-2/Bax(P <0.05), but there was no obvious variation of Bax and Fas expression( P >0.05). Inhibition of proliferation and apoptosis were not obvious in PML-RARα negative HL-60 cells induced by low concentration As2O3 ( P >0.05), and there was no obvious variation of Bcl-2, Bax, Fas mRNA expres-sion or Bcl/Bax ratio( P >0.05). Conclusion The ratio of Bcl-2/Bax is contributed to the different sensitiv-ities of PML-RARα negative HL-60 cells and positive NB4 cells induced by low concentration of As2O3.

AIMTo investigate the changes in the expression of four kinds of calcium regulatory proteins mRNA on the isolated ischemia/ reperfusion (IR) hearts.

METHODSThe rat hearts were divided into two groups: control group and IR group which received 45 min exposure to Krebs-Henseleit solution after 15 min zero-flow global ischemia. The indexes of left ventricular function, such as LVDP, +dp/dt(max), -dp/dt(max), and an arrhythmia scoring system were compared between the two groups. The messenger ribonucleic acid (mRNA) amount of sarcoplasmic reticulum (SR) Ca2+ ATPase (SERCA), phospholamban (PLB), inositol 1,4,5-trisphosphate receptor2 (IP3R2) and ryanodine receptor2 (RyR2) was measured by reverse transcription-polymerase chain reaction (RT-PCR) and normalized to the mRNA levels of beta-actin.

RESULTSIn the IR group, LVDP, +dp/dt(max) and -dp/dt(min) of the isolated hearts were depressed and the high rate of arrhythmias occurred during reperfusion. The levels of SERCA, IP3R2, RyR2 mRNA were lower in the IR isolated hearts group than those in the control group, while there was no difference in the level of phospholamban.

CONCLUSIONThese data suggest that myocardial ischemia/reperfusion can induce the depression of cardiac performance and an increased risk of arrhythmias, concomitant with the decrease in SERCA, IP3R2, RyR2 mRNA steady state levels.

Animals ; Myocardial Reperfusion Injury ; metabolism ; Myocardium ; metabolism ; RNA, Messenger ; genetics ; Rats ; Rats, Sprague-Dawley ; Sarcoplasmic Reticulum Calcium-Transporting ATPases ; genetics ; metabolism

OBJECTIVETo investigate the effect of Astragalus membranaceus(AM) on vascular circles and the underlying mechanisms.

METHODSThe study was performed with the model of isolate rat thoracic aorta rings in organ bath. When the endothelium of rat thoracic aorta was removed,the effect of accumulated AM on aorta rings in resting tension, or pre-constricted with KCl, or pre-constricted with phenylephrine (PE) was observed. And to explove the mechanism, the aorta rings were incubated with Ca(2+)-free medium alone, or Ca(2+)-free medium plus heparin, or propranolol alone before pre-contraction with PE.

RESULTSAM had no significant effects on aorta rings in resting tension or pre-constricted with KCl. When the concentration of AM was cumulated to 10(-1), 3 x 10(-1),10(0), 3 x 10(0) g/L, it caused concentration-dependent relaxation while aorta rings were pre-constricted with PE(3 x 10(-7)mol/L), compared with the control [(90.4 +/-4.2)% compared with (94.7 +/-2.4)%,(86.1 +/-5.0)% compared with (92.6 +/-3.2)%, (82.3 +/-5.9)% compared with (90.4 +/-3.6) %, (78.3 +/-6.0)% compared with (88.1 +/-4.0)%]. This effect was not inhibited by Ca(2+)-free medium or propranolol alone. However, the effect was attenuated by the co-incubation with heparin and Ca(2+)-free medium [without heparin:(76.2+/-4.3)% compared with (92.3 +/-5.9)%, with heparin: (95.3+/-0.5)% compared with (95.1+/-0.6)%].

CONCLUSIONThe results indicate that AM can relax the rat thoracic aorta rings without endothelium. The mechanism may include the inhibition of intracellular calcium ions release by the 1,4,5-triphosphate inositol-receptor-dependent pathway in vascular smooth muscle cells.

Animals ; Aorta, Thoracic ; cytology ; Astragalus membranaceus ; Calcium ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; In Vitro Techniques ; Male ; Muscle, Smooth, Vascular ; cytology ; Phosphatidylinositols ; metabolism ; Rats ; Rats, Sprague-Dawley ; Vasodilator Agents ; pharmacology

Objective To investigate the clinical efficacy of herbal cake moxibustion plus acupuncture in treating post-stroke urinary incontinence.Methods Fifty patients with post-stroke urinary incontinence were randomized to a treatment group (26 cases) and a control groups (24 cases). The treatment group received herbal cake moxibustion plus acupuncture and the control group, acupuncture alone. Twenty-four-hour urine void number, incontinence severity and the clinical symptom score were recorded in the two groups before and after treatment.Results There were statistically significant pre-/post-treatment differences in twenty-four-hour urine void number and the clinical symptom score in the two groups (P<0.05). There were statistically significant differences in pre-/post-treatment twenty-four- hour urine void number difference value and clinical symptom score difference value between the treatment and control groups (P<0.05). There was a statistically significant pre-/post-treatment difference in incontinence severity in the two groups (P<0.05). There was a statistically significant post-treatment difference in incontinence severity between the treatment and control groups (P<0.05).Conclusion Herbal cake moxibustion plus acupuncture is an effective way to treat post-stroke urinary incontinence.

Severe fever with thrombocytopenia syndrome bunyavirus is a newly emerging virus in China, enveloped with a tripartite, single-stranded RNA genome of negative polarity. The regulatory elements for viral transcription and replication, as well as encapsidation and packaging signals, are thought to be located within these noncoding regions (NCRs). The terminal nucleotides are genus specific and highly conserved. The function of the remaining nucleotides of the NCRs is still not well understood. In this study, we developed the plasmid-driven RNA polymerase I minireplicon system for SFTSV firstly, using reporter genes GFP and luciferase. The function of the noncoding regions of the three Bunyaviridae RNA segments (L, M, S) in transcription was analyzed. Reporter genes are successfully expressed in SFTSV minireplicon system. Our results suggest that the NCRs of SFTSV from all three segments contain the necessary signals to initiate transcription. Quantitative detection of the luciferase expression level shows that promoter activity in the three segments is different.
Bunyaviridae Infections ; virology ; Cloning, Molecular ; Genome, Viral ; Humans ; Phlebovirus ; genetics ; physiology ; Replicon ; Viral Proteins ; genetics ; metabolism ; Virus Replication

OBJECTIVETo investigate the effects of Weile Powder (WLP) on bicarbonate transporters in rats with gastric ulcers, and to probe its functional mechanisms.

METHODSThe 48 SD rats were randomly divided into the normal control group, the model group, the low dose WLP group (at the daily dose of 0.075 g/mL), the middle dose WLP group (at the daily dose of 0.150 g/mL), the high dose WLP group (at the daily dose of 0.030 g/mL), and the ranitidine group (at the daily dose of 0.030 g/mL), 8 in each group. The gastric ulcer rat model was prepared by the glacial acetic acid cauterization method. Rats in each medication group were administered from the 2nd day of modeling. Rats were sacrificed after 14-day successive medication. The protein was extracted from the ulcer tissue. The protein expressions of solute carrier26A3 (SLC26A3)and solute carrier26A6 (SLC26A6) were detected using Western blot. The gastric ulcer and its peripheral tissue were sectioned. The changes of cystic fibrosis transmembrane conductance regulator (CFTR) were measured by immunofluorescence.

RESULTSCompared with the model control group, the expression levels of SLC26A3 increased in the high dose WLP group and the ranitidine group with statistical difference (P < 0.05). The expression levels of SLC26A6 increased in the high and middle dose WLP groups and the ranitidine group with statistical difference (P < 0.05). The expression level of CFTR also obviously increased in the high and middle dose WLP groups (P < 0.01).

CONCLUSIONWLP could elevate the expression levels of SLC26A6, SLC26A3, and CFTR, increase the secretion of bicarbonate, thus protecting the gastric mucosa.

Animals ; Antiporters ; metabolism ; Bicarbonates ; metabolism ; Cystic Fibrosis Transmembrane Conductance Regulator ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Female ; Gastric Mucosa ; drug effects ; metabolism ; Male ; Rats ; Rats, Sprague-Dawley ; Stomach Ulcer ; metabolism

OBJECTIVETo discuss the central modulating mechanism of Sanyinjiao (SP 6) and the amplitude of low-frequency fluctuation (ALFF) differences of the functional brain regions between SP 6 and sham acupoint.

METHODSSixteen volunteers accepted right Sanyinjiao (SP 6) (SP 6 group) or right sham (sham group) acupuncture for two times after 24 h sleep deprivation (SD), there was two weeks between two acupunctures. The needle was retained for 4 minutes and the brain fMRI scanning was performed by 3.0 TMR every time. The ALFF consequent data was processed by REST. One-sample t-test and two-pair t-test were performed by SPM5.

RESULTSSP 6 group mainly caused ALFF change in inferior parietal lobule, posterior cingulated while sham group in precuneus, posterior cingulated. SP 6 group increased ALFF in left fusiform temporal gyrus, medial frontal gyrus with no decreased regions compared with sham group.

CONCLUSIONSRetaining needle dose influence the activity of the brain region during resting-state in both group, especially the mood-related regions. And its mechanism is probably that the relevant effect is carried out by regulating function to target organs of complex brain network comprised of relevant functional center and related brain region. There are differences between SP 6 and Sham acupoint.

Acupuncture Points ; Acupuncture Therapy ; Brain ; diagnostic imaging ; physiopathology ; Female ; Humans ; Magnetic Resonance Imaging ; Male ; Radiography ; Sleep Deprivation ; diagnostic imaging ; physiopathology ; therapy ; Young Adult