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Objective To investigate the main chemical constituents of the low polarity extracts from pinusmassoniana Lamb. leaves and their synergetic activity with fluconazole against fluconazole-resistant Candida albicans. Methods The pinusmassoniana leaves were extracted with 80% ethanol, and then the extracts were extracted by petroleum ether to obtain the low polarity extracts. The chemical components were detected by GC-MS and elucidated by the comparison with the standard mass spectral data. The relative contents in percentage were calculated using the area normalization method. The minimal inhibitory concentrations (MIC80) of fluconazole-resistant Candida albicans strains by the low polarity extracts in combination with fluconazole were determined by checkerboard microdilution assay. Results 30 components were detected from the low polarity extracts, and 17 components were identified. The minimum inhibitory concentration (MIC80) of the 80% ethanol extracts, the low polarity extracts and the petroleum ether extracts from the pinusmassoniana leaves combined with fluconazole against fluconazole-resistant Candida albicans were 7.81 μg/ml, 31.25 μg/ml and >250 μg/ml, respectively. Conclusion The 80% ethanol extracts of pinusmassoniana leaves and its low polarity extracts have synergistic activity combined with fluconazole onfluconazole-resistant Candida albicans. The diterpenoids (53.99%) may be the effective components of the low polarity extracts.
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Objective@#To evaluate the effect and mechanism of down-regulating the expression of REV7 on the radiosensitivity of human colon cancer cell HCT116.@*Methods@#HCT116 cells were cultured and the expression of REV7 was down-regulated by RNA interference technique. HCT116 cells were divided into the blank group, negative control transfected with negative RNA oligo group and REV7 expression down-regulation transfected with REV7 RNA oligo group, respectively. The cell proliferation was determined by colony formation assay. The expression levels of the proteins of relevant genes were detected by Western blot. The level of cell apoptosis and non-homologous end joining was evaluated.@*Results@#The colony formation rate was significantly reduced in THE REV7 siRNA group after 6Gy irradiation (P<0.05). The down-regulating efficiency rate of REV7 gene was>60% in the REV7 siRNA group. The expression levels of γH2AX and Caspase9 were significantly up-regulated, whereas those of KU80 and XRCC4 were remarkably down-regulated in the REV7 siRNA group (all P<0.05).@*Conclusions@#The radiosensitivity of human colon cancer cell HCT116 can be increased by down-regulating the expression of REV7. The underlying mechanism may be related to the lower incidence rate of non-homologous end joining.
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Objective To evaluate the effect and mechanism of down-regulating the expression of REV7 on the radiosensitivity of human colon cancer cell HCT116.Methods HCT116 cells were cultured and the expression of REV7 was down-regulated by RNA interference technique.HCT116 cells were divided into the blank group,negative control transfected with negative RNA oligo group and REV7 expression down-regulation transfected with REV7 RNA oligo group,respectively.The cell proliferation was determined by colony formation assay.The expression levels of the proteins of relevant genes were detected by Western blot.The level of cell apoptosis and non-homologous end joining was evaluated.Results The colony formation rate was significantly reduced in THE REV7 siRNA group after 6Gy irradiation (P＜0.05).The down-regulating efficiency rate of REV7 gene was ＞ 60％ in the REV7 siRNA group.The expression levels of γ H2AX and Caspase9 were significantly up-regulated,whereas those of KU80 and XRCC4 were remarkably down-regulated in the REV7 siRNA group (all P＜0.05).Conclusions The radiosensitivity of human colon cancer cell HCT116 can be increased by down-regulating the expression of REV7.The underlying mechanism may be related to the lower incidence rate of non-homologous end joining.
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Objective To explore the effect of silencing Rev1 gene on the proliferation and apoptosis induction of human colon cancer cell line THC8307 after X-ray irradiation.Methods Rev1 siRNA was transferred into THC8307 cells and its transferring efficiency was tested by RT-PCR and Western blot assay.The cells were divided into blank control group,negative control group and radiation group.After 6 Gy irradiation,cell proliferation were detected by MTI assay,cell apoptosis were detected by flow cytometry (FCM),and the expressions of PCNA,γ-H2AX,P53,Bax,and Bcl-2 proteins in each group were detected by Western blot.Results Compared with the blank control group,after 6 Gy irradiation,the proliferation rate of the silenced Rev1 was reduced (t =7.53,P ＜ 0.05) and apoptosis rate was increased (t =6.23,P ＜ 0.05).The expression of PCNA and Bcl-2 decreased (t =4.39,6.13 P ＜0.05),and the expressions of γ-H2AX,P53 and Bax increased (t =5.48,5.09,3.32,P ＜ 0.05).Conclusions Silencing Rev1 gene inhibits the proliferation of colon cancer cells,promotes apoptosis,and increases cell radiosensitivity to X-rays.
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Objective To explore the effects of cyclophosphamide administered by different routes or in different doses on the embryo-fetal development in pregnant rabbits, and to determine the optimal mode of cyclophosphamide administration to induce fetal malformation.Methods Pregnant rabbits were divided into control group C (saline), group Y1 (intravenous injection of 15 mg/kg cyclophosphamide,), group Y2 (subcutaneous injection of low dose cyclophosphamide, 20 mg/kg), and group Y3 (subcutaneous injection of high dose cyclophosphamide, 30 mg/kg).Each rat was administrated according to the corresponding mode once daily on GD10~13.The day of conception was designated as GD0.The pregnant rabbits were sacrificed and dissected on GD28.Then, the number of corpora lutea and implantation, the weight of uterus with contained fetus, and fetal resorption rate were detected, the fetuses were removed and the fetal sex, body length, tail length, the number of live births and stillbirths were recorded, and the appearance of deformities, visceral deformities and skeletal malformations were detected.Results Pregnant rabbit fetuses in the cyclophosphamide intravenous injection group and subcutaneous injection of low dose cyclophosphamide group showed deformities.The appearance malformation rates in the two groups were 30.77% and 95.65%, the skeletal deformity rates were 7.69% and 73.91%, and the visceral abnormality rates were 20.51% and 47.83%, respectively.The fetal resorption rate in the high dose cyclophosphamide subcutaneous injection group was 100%.Conclusions Subcutaneous injection of 20 mg/kg cyclophosphamide to pregnant rabbits at GD10~13 can be used as a positive administrationmethod for rabbit embryo-fetal developmental toxicity test.Thismethod has the advantages of short administration period, easy operation, few fetus resorption, and high rate of fetal malformation, thus, providing the evidence for selection of appropriate model of rabbit embryo-fetal developmental toxicity.
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Objective To investigate the plasma vitamin D3 level and its association with B cell subsets of patients with primary Sj(o)gren's syndrome (pSS).The role of vitamin D3 levels in the pathogenesis of pSS was explored.Methods The expression of plasma vitamin D3 levels of 55 patients with pSS and 32 controls were analyzed by ELISA.Frequencies of peripheral blood CD19+CD27-na(i)ve B cells,CD19+CD27+ memory B cells and CD19+CD27high plasma cells were analyzed by flow cytometry in 34 pSS patients without therapy and 22 controls.The relationship between the vitamin D3 levels and B cell subsets,SSDAI,tear flow rate,saliva flow rate,rheumatoid factor,immunoglobulin was analyzed in pSS patients.Non-parametric test,t test,one-way ANOVA,x2test,Pearman's and Spearman's correlation analysis were used for statistical analysis.Results ① There was significant difference in the levels of plasma 1,25 (OH)2D3 between the pSS patients group and normal control group,1,25 (OH)2D3 was significantly lower in pSS patients than that in the normal control group [24.17(22.20,28.41) pg/ml and 41.25(23.38,62.18) pg/ml,P＜0.05],and that was also obviously lower in the active group [22.64(20.74,24.90) pg/ml] than that in the normal control group (P＜0.05),and that was also obviously lower in the active disease group than that in the inactive disease group [25.39 (23.16,33.09) pg/ml,P＜0.05],but there was no difference between the inactive group and the normal control group (P＞0.05).② The percentage of peripheral blood of CD19+CD27high plasma cells and CD19+CD27+ memory B cells in CD19+ cells was reduced in patients in the pSS group compared with the control group [(0.89±0.30)％ and (1.72±0.43)％,(24±8)％ and (34±5)％; P＜0.05],and that was also significantly lower in the active group [(1.03±0.59) ％ ; (26± 10)％] and inactive group [(1.00±0.16)％,(26± 3)％] than that in the normal controls (P＜0.05).However,there was no difference between the active group and the inactive group (P＞0.05),but the frequency of peripheral blood of CD19+ CD27-naive B cells in CD19+ B cells was increased in patients with pSS compared with normal control group [(75.4±7.5)％ and (63.9±5.2)％,P＜0.05],and that was also significantly higher in the active group [(73.4±9.7)％] and inactive group [(73.3±2.9)％] than that in the normal control (P＜0.05),there was no difference between the active group and the inactive group.③ Significant negative correlation was observed between 1,25 (OH)2D3 and the percentage of peripheral blood CD19+CD27+ memory B cells in CD19+ cells as well as immunoglobulin G(r=-0.627,P=0.039; r=-0.657,P＜0.01) level.Conclusions These results demonstrate that abnormality of vitamin D levels may play an important role in the pathogenesis of pSS.