2.Effects of emodin on proliferation cycle and apoptosis of human bladder cancer cell line BIU87
Zejun YAN ; Yue CHENG ; Junhui JIANG ; Guohai XIE ; Jiasheng HU
Journal of Chinese Physician 2011;13(8):1037-1040
ObjectiveTo study the suppressive role of emodin on the growth and its effect on the proliferation cycle and apoptosis of human bladder cancer cell line BIU87.MethodsThe effect of different concentration of emodin at different time point on cell proliferation of BIU87 were measured with methylthiazole (MTT) chromatometry, the cell proliferation cycle were detected with flow cytometry, expressions of bc1-2 and caspase-3 were detected by SP of immunohistochemistry.ResultsWithin a certain range, the higher concentration of emodin (10 ~ 80 μg/ml) and the longer action time are positive related the more significant inhibition of tumor cell growth and the higher apoptosis rate [(9.84 ± 1.13)%, (18.32 ±2.14)% ,(29.73+1.42)% ,(42.13 +2.36)% ,respectively].Compared with control group [(2.01 ±0.92)%], the differences were statistically significant(F =531.85, P <0.01).Emodin could inhibit the proliferation of human bladder cancer cell BIU87 by blocking BIU87 cell in G0/G1 stage, thus cut down cell proportion in stage of S [(33.27 +1.26)% ,(29.17 ±1.39)%, (16.94 ±0.86)% ,(10.85 ± 1.47)%,respectively], compared with the control group [(35.45 ± 0.38) %], the differences were statistically significant(F =524.64, P <0.01).After 48 h of emodin treatment, the bc1-2 expression(Grayscale values:122.65 + 2.12,131.37 ± 1.62,134.81 ± 1.36,145.55 ± 2.01, respectively) was decreased and the caspase-3 expression(Grayscale values : 135.26 + 1.41,130.22 ± 1.74,126.11 ± 1.77,118.36 + 1.53, respectively) was increased in a dose dependent manner.Compared with control group (Grayseale values:108.42 + 3.73,149.35 ± 1.82, respectively), the differences were statistically significant (F = 216.23,224.83, P <0.01).ConclusionsEmodin could significantly inhibit the growth and induce apoptosis of BIU87 cells in vitro, which may be through down regulation of bc1-2, and up regulation of caspase-3, and blocking BIU87 cell in G0/G1 stage.
3.Experimental study of the anti-melanoma effect of dentritic cells pulsed with RPL8 peptide
Yan LI ; Lingsi YIN ; Huan YUE ; Junqiong HUANG ; Yonglin HU
China Oncology 2014;(4):279-283
Background and purpose:Studies have shown that ribosomal protein L8 (RPL8) is shared by melanomas, gliomas and ovarian carcinomas. A peptide of RPL8 signiifcantly stimulated proliferation and cytokine expression of the hepler T cell clone and lymphocytes in melanoma patients. RPL8 may stimulate anti-tumor immunity, making RPL8 an attractive candidate for therapeutic intervention. In this study, we prepared DC pulsed by RPL8 (RPL8-DC) and investigate the anti-tumor effect of RPL8-DC on melanoma in mice.Methods: The recombinant protein was achieved through IPTG induction in E. coli and identiifed with Western blot. Bone marrow-derived DC was loaded with RPL8 protein. RPL8 and CD11c, CD80, MHC-Ⅰ, MHC-Ⅱmolecules on dentritic cells were monitored by lfuorescence microscope and FACS analysis, respectively. The anti-tumor effect of T cells in vitro was detected by MTT assay. Subcutaneous tumors were induced in C57BL/6 mice using B16 cells. The tumor volumes were measured after injection with RPL8-DC. Results:The puriifed protein was combined with speciifc antibodies. DCs pulsed by RPL8 were visualized under lfuorescent microscopy. CD11c, CD80, MHC-Ⅰ, MHC-Ⅱmolecules on DCs were up-regulated after stimulation with RPL8 and LPS. B16 cells were inhibited by T cells stimulated with RPL8-DC. The inhibition rate of tumor cells was 70%in RPL8-DC group when effector-to-target ratio was 30∶1, which was higher than PBS and DC groups. Inhibition of growth could be observed more signiifcantly in mice after the treatment with RPL8-DC. The mice receiving the therapy of RPL8-DC were able to survive much longer than the mice receiving control therapy. Conclusion:The DC pulsed by RPL8 protein can inhibit the growth of melanoma.
4.Protective effects of tempol on ultraviolet-B exposed human foreskin fibroblasts
Shuxian YAN ; Yu XU ; Yue HU ; Kanghuang LIAO
Chinese Journal of Geriatrics 2003;0(10):-
Objective To determine the effects of tempol(a nitroxide), in the exposure of ultraviolet-B (UVB), on cell proliferation, superoxide enzyme (SOD) activity, lipid peroxidation, and expression of matrix metalloproteinases (MMP)-1,MMP-3 in human foreskin fibroblasts in vitro. Methods Fibroblasts were irradiated by a single exposure of 36 seconds to 40 mJ/cm 2 UVB and at the same time incubated with, or without, tempol and detected twenty-four hours later. SOD activity and lipid peroxidation,as shown by accumulation malondialdehyde (MDA) were detected by biochemical assay. Expression of MMP-1 and MMP-3 (mRNA level) were examined by Semi-quantitative RT-PCR. Results 40 mJ/cm 2 UVB significantly inhibited cell proliferation rate to (84?8)% (P
5.Appraisal of Murine Local Lymph Node Assay Combined with Measurement of Ear Swelling for Identification of Photosensitive Agents
Li MA ; Chunlin YAN ; Kanghuang LIAO ; Yue HU
Chinese Journal of Dermatology 2003;0(10):-
Objective To establish a method of murine local lymph node assay (LLNA) combined with measurement of ear swelling for the identification of photosensitive agents. Methods The female BALB/c mice received test chemicals or vehicle alone on the back skin of both ears and then were exposed to UVA irradiation on three consecutive days. The post-auricular lymph nodes draining the ear tissue were excised 24 h following the last irradiation. Lymph node cell counts and lymphocyte proliferative rates were determined, which indicated lymph node hyperplasia. While ear swelling was also measured, to reveal skin irritating reactions. Some cytokines (IFN-?, IL-2 and IL-4) released by the draining lymph node were also analyzed by ELISA. Results Tetrachlorosalicylanilide, a strong photoallergen, caused lymph node hyperplasia without ear swelling. The mild photoallergen (6-methylcoumarin) showed neither lymph node proliferation nor ear swelling reactions. But 8-methoxypsoralen, a phototoxic substance, induced both lymph node hyperplasia and ear swelling. The level of IL-4 was significantly increased when the mice were previously sensitized with tetrachlorosalicylanilide (P
6.Determination of furbenicillin sodium and its related substances by HPLC.
Yan CHANG ; Ye TIAN ; Yue MA ; Chang-qin HU
Acta Pharmaceutica Sinica 2015;50(12):1632-1636
Furbenicillin is a broad-spectrum semisynthetic penicillin with strong antibacterial activity against Gram-negative bacteria. Furbenicillin sodium is determined by volumetric method in current criteria. However, the criteria does not contain an assay of related substances of furbenicillin sodium. In this study, we established a method for detection and analysis of furbenicillin sodium and its related substances by HPLC. The analysis was performed with a C18 column under a gradient elution, the detection wavelength was 225 nm, and the column temperature was 35 degrees C. The reliability and accuracy of established method was validated in this study. Pure samples of furbenicillin sodium and its related substances were prepared. The structures, biological activities, and chromatographic retention behaviors of furbenicillin sodium and its related substances were identified using NMR, CLSI agar dilution method, and HPLC. All results in the current study provide ample evidence that this method is able to determine the reasonable limits in the quality-control protocol for furbenicillin sodium.
Anti-Bacterial Agents
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chemistry
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Chromatography, High Pressure Liquid
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Magnetic Resonance Spectroscopy
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Penicillins
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Quality Control
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Reproducibility of Results
7.Narrow-band ultraviolet B phototherapy regimens for the treatment of chronic actinic dermatitis and analysis of factors influencing treatment compliance
Li MA ; Yue HU ; Yu XU ; Shuxian YAN ; Kanghuang LIAO
Chinese Journal of Dermatology 2017;50(8):571-574
Objective To investigate the optimal regimen of narrow-band ultraviolet B (NB-UVB) phototherapy in the treatment of chronic actinic dermatitis (CAD),and to analyze factors influencing treatment compliance.Methods Demographic data,results of photobiological tests,treatment parameters and clinical responses were collected from CAD patients who received NB-UVB phototherapy in Huashan Hospital affiliated to Fudan University from January 2008 to June 2015,and were reviewed retrospectively.Statistical analysis was done by using two independent samples t-test and chi-square test with SAS9.3 software to compare the clinical data between patients who completed and did not complete the NB-UVB phototherapy.Results A total of 79 CAD patients with Fitzpatrick skin type Ⅳ received NB-UVB phototherapy.Of these patients,61 (77%) completed the whole treatment,while 18 (23%) dropped out because of intolerance to the NB-UVB radiation.Among the 61 patients who completed the treatment,the average initial,final and cumulative radiation doses of NB-UVB were (0.08 ± 0.01) J/cm2,(0.32 ± 0.08) J/cm2and (5.9 ± 2.5) J respectively,and patients received (28 ± 8) times of treatment in average.When the radiation dose went up to 0.30 J/cm2,most skin lesions were cleared in 52 (85%) patients.A total of 19patients received phototesting again after the end of phototherapy.Among 16 patients sensitive to ultraviolet A (UVA) before the treatment,6 had normal minimal erythema dose to UVA (UVA-MED),and another 6 had improved UVA-MED after the treatment.Among 16 patients sensitive to UVB before the treatment,11 got normal UVB-MED and another 3 had improved UVB-MED after the treatment.Univariate analysis showed no significant differences in gender,age,duration of the disease,sensitivity to UVA and UVB radiation,results of photopatch test and patch test between the patients who completed and did not complete the treatment (all P > 0.05).Conclusions The appropriate NB-UVB phototherapy for CAD patients should start at an initial radiation dose of 0.08 J/cm2 in spring and end at a final radiation dose of 0.30 J/cm2 for about 28 sessions,which can effectively reduce the photosensitivity to both UVA and UVB in CAD patients.Additionally,NB-UVB phototherapy can be applied in CAD patients of different gender,age,disease duration and photosensitive condition.
8.Inhibitorv effect of emodin on heterotransplanted human bladder cancer in nude mice and its mechanism
Zejun YAN ; Yue CHENG ; Junhui JIANG ; Guohai XIE ; Jiasheng HU
Journal of Chinese Physician 2011;13(10):1330-1334
Objective To investigate the inhibitory effect of emodin on hetertransplanted human bladder cancer in nude mice and explore its mechanism.Methods Heterotransplanted models of human bladder cancer cell line BIU87 cells in nude mice were established.The mice were randomly divided into 4 groups during the experiment:blank control group,Z-VAD-FMK group,emodin group and emodin combined with Z-VAD-FMK group.The growth of tumors was observed and the growth curve was mapped.The nude mice were sacrificed 4 weeks later,the tumors were isolated and weighed.The pathological changes of tumor were observed after HE staining,the cells apoptosis were detected with flow cytometry,and the expression of AIF and Endo G were examined by reverse transcription PCR (RT-PCR) and Western blot.Results The tumor growth rate in emodin group was lower than that in the other three groups.The tumor quality in emodin group [(0.41 ± 0.05 ) g] and emodin combined with Z-VAD-FMK group [( 0.69 ±0.07)g]were lighter than that in the other two groups[(1.08 ±0.13,1.04 ±0.09)g,],and the differences were statistically significant( F =90.56,27.49,P <0.01 ).The quality difference in emodin group and emodin combined with Z-VAD-FMK group was statistically significant ( t =10.01,P < 0.01 ).The apoptosis rate in emodin group [(42.71 ±2.69)%]was significantly higher than that in emodin combined with Z-VAD-FMK group[(34.38 ± 1.73)%] ( t =6.38,P <0.01 ).The expression of AIF and Endo G in emodin combined with Z-VAD-FMK group was significantly increased than other groups [( 1.65 ±0.12)vs(1.24±0.08),(0.51 ±0.07),(0.48 ±0.04);(2.12 ±0.16)vs(1.75 ±0.13),(0.57 ±0.06),(0.59±0.07);(2.42±0.13)vs(1.73 ±0.11),(0.78 ±0.07),(0.75 ±0.08);(3.13 ±0.25)vs(2.15± 0.18 ),(0.85 ± 0.09 ),(0.81 ± 0.14 )],and the differences were significant ( F =303.22,319.32,409.38,258.53,P < 0.01 ).Conclusions Emodin could significantly inhibit the growth of hetertransplanted human bladder cancer in nude mice.The mechanism might be partly due to the expression increase of AIF and Endo G in bladder cancer cells,which might induce apoptosis through Caspase-independent pathway.
9.Mechanism of apoptosis of bladder cancer cells BIU87 induced by emodin through Caspase-independent pathway
Zejun YAN ; Yue CHENG ; Junhui JIANG ; Guohai XIE ; Jiasheng HU
Chinese Journal of Urology 2012;33(4):259-263
Objective To study the effects of emodin on apoptosis and mRNA and protein of apoptosis inducing factor (AIF) and Endonuclease G (Endo G) in human bladder cancer cells BIU87,and to investigate the anticancer mechanism of emodin. Methods The BIU87 cells were divided into 4 groups,control group,Z-VAD-FMK group,emodin group,emodin combined with Z-VAD-FMK group.The effects of different concentrations of emodin at different action time on cells proliferation of BIU87 in vitro culture were measured by methylthiazole (MTT) chromatometry,the cells apoptosis were detected by flow cytometry,and expression of AIF and Endo G were examined by reverse transcription PCR (RT-PCR) and Western blot.Results MTT assay demonstrated that the higher concentration of emodin and the longer action time,the more significant inhibition of tumor cell growth.Based on the IC50 value,80 μmol/L and 72 h of emodin intervention were selected as an intervention condition.The apoptosis rate in emodin group (44.57 ± 1.52 ) %was significantly higher than that in emodin combined with Z-VAD-FMK group (35.58 ± 1.61 ) % ( P <0.01).RT-PCR and Western blot showed that the mRNA and protein of AIF in emodin combined with Z-VAD-FMK group,emodin group,control group,Z-VAD-FMK group were ( 1.74 ± 0.11 ) and (2.59 ±0.13),(1.36±0.08) and (1.89±0.14),(0.37 ±0.02) and (0.53±0.11),(0.42 ±0.06) and (0.44 ± 0.07),respectively.There were significant differences between emodin group and the other groups (P <0.01 ).The mRNA and protein of Endo G in emodin combined with Z-VAD-FMK group,emodin group,control group,Z-VAD-FMK group were (2.28±0.15) and (3.31 ±0.36),(1.85 ±0.13) and (2.15 ±0.27),(0.53 ±0.07) and (0.71 ±0.16),(0.61 ±0.04) and (0.67 ±0.22),respectively.The differences were significant between emodin group and the other groups ( P < 0.01 ). Coneltusion Emodin can upgrade the expression of AIF and Endo G in bladder cancer cells BIU87,which can induce apoptosis through Caspase-independent pathway.
10.Expression and Clinical Significance of Minichromosome Maintenance Protein 7 in Sinonasal Squamous Cell Carcinoma
Yue HU ; Feifei JIANG ; Jiali HAN ; Xiaoxu DING ; Aihui YAN
Journal of China Medical University 2017;46(4):330-334
Objective To investigate the expression of minichromosome maintenance protein 7 (MCM7) in sinonasal squamous cell carcinoma (SNSCC),and evaluate its relationship with tumor differentiation and prognosis of patients.Methods Using immunohistochemistry,MCM7 expression in SNSCC and nasal polyps with chronic sinusitis (NPCRS) were studied,and relationships between markers and clinicapathological features were analyzed.Results In NPCRS,MCM7 positive cells were mainly distributed in the epithelial basal layer and the expression rate was low,whereas in SNSCC,MCM7 positive cells were diffuse and the expression rate was high.MCM7 expression was significantly higher in SNSCC than in NPCRS (P < 0.001) and related to tumor differentiation (P =0.001),increasing gradually with decreasing degree of differentiation.The overall 3-and 5-year survival rates of patients with SNSCC were 61.3% and 46%,respectively.The 3-year survival rates for patients with stage Ⅰ and Ⅳ were 90% and 25.6%,respectively,and the 5-year survival rates were 70% and 17.1%,respectively;the difference was statistically significant (P < 0.001).The 3-and 5-year survival rates of MCM7-negative patients were 36.0% and 18.0%,respectively,and those of MCM7-positive patients were 59.9% and 34.2%,respectively;the difference was not statistically significant (P =0.297).Conclusion In SNSCC,MCM7 expression is significantly increased,inversely associated with the degree of tumor differeutiation,and unrelated to the survival rates of patients.