1.Effect of DCN gene on the anti-tumor and immunological functions of mice bearing S180 sarcoma
Jingjing WANG ; Wanfang YANG ; Jing LAN ; Guiqin WANG
Cancer Research and Clinic 2008;20(4):221-223
Objective To investigate the anti-tumorigenesis function of DCN gene and its effect on the immunological funotions of mice bearing S180.Methods Twenty four mice bearing S180 were divided into three groups at random(eight mice every group),and then the normal saline group(negative control),pcDNA3.1(+)vector group and pcDNA3.1(+)-DCN group were set up for the experiment.They all started to inject next day by armpit.The volume of tumors were measured at the 5th day,7th day,9th day,11th day.The transformation rate of T lymphocytes were detected by means of MTT,the mRNA level of TGF-β by RT-PCR and the level of VEGF in serum by ELISA.Results The average volume of tumor in pcDNA3.1 (+)-DCN group were deflated to(2.219±0.105)cm3 comparing with the others(P<0.001);the transformation rate of T lymphocytes were increased significantly to(67.21±6.32)%(P<0.01,P<0.05);meanwhile it can decrease the mRNA level of TGF-β in serum,and depress the expression of VEGF markedly,whose absorbance were2.6175±0.099(P<0,01,P<0.05).Conclusion DCN gene could significantly inhibit the growth of tumor,and enhance the immunological function of mice bearing S180 in vivo.
2.Effect of rhDCN on the expression of HLA molecules on HepG2 cell lines
Yonhong WANG ; Guiqin WANG ; Jingjing WANG ; Fang HAO ; Wanfang YANG ; Ying XUE
Cancer Research and Clinic 2009;21(5):298-300
Objective To observe the regulatory effect of recombinant human DCN on the expression of HLA class Ⅰ, Ⅱ molecule on hepatoma carcinoma cell HepG2 and investigate the relatively immune mechanism of human DCN on enhancing anti-tumor effect. Methods After transfected HepG2 cells with pcDNA3.1 (+)-DCN by liposome transfection, the expression of HLA class Ⅰ , Ⅱ molecule and the apoptotic indexes were analyzed by the flow cytometer. Results The expression of HLA class Ⅰ , Ⅱ molecule on the HepG2 cells transfected with pcDNA3.1 (+)-DCN was up-regulated with the apoptosis indexes being significantly higher than that of other control groups. Conclusion The human DCN can induce HepG2 apoptosis and up-regulate the expressions of HLA class Ⅰ , Ⅱ molecule which may contribute to its antitumor effect.
3.Effect of influenza virus on chemokine pathways in mice lungs and intervention of Shufeng Xuanfei Jiedu formula
Qi LIU ; Jianguo WANG ; Baozhen LI ; Lu JIA ; Yu GUO ; Wanfang YANG
Chinese Critical Care Medicine 2021;33(4):405-409
Objective:To screen the differentially expressed genes on whole expression profiles of the inflammation-related cytokines in mice infected with influenza virus by the gene chip technology, and to explore the intervention effect of Shufeng Xuanfei Jiedu formula.Methods:Male ICR mice were divided into normal group (N group), influenza virus infective model group (M group), Oseltamivir control group (C group) and Shufeng Xuanfei Jiedu formula high, medium and low dose groups (SH, SM, SL groups) according to the random number table method, with 10 rats in each group. A mouse model of influenza virus pneumonia was reproduced by nasal drip of influenza virus strain FM1 (0.05 mL). In N group, 0.05 mL normal saline was used. In SH, SM and SL groups, Shufeng Xuanfei Jiedu formula was used 2 hours after intranasal infection (2 times, equal and 1/2 of the clinical treatment dose, approximately 3.8, 1.9 and 1.0 g·mL -1·d -1) for 4 days. In C group, the dosage of Oseltamivir was 2.5 g·mL -1·d -1. In N group and M group, distilled water was given (0.2 mL once a day). On the 5th day, the whole lung of mice was taken. The lung index was calculated, and the pathological sections were observed. The total RNA of lung tissue was extracted and detected after hybridization with mice whole gene expression spectrum chip to select differentially expressed genes of chemokine pathways. The expression intensity ratio of the chip probe signal in each group vs. M group was calculated, and P < 0.05 and log 2ratio > 1 were up-regulated genes, while P < 0.05 and log 2ratio < -1 were down-regulated genes. Results:Compared with the N group, the lung index in the M group was significantly higher, and pathological changes were found in lung tissue, which suggested that the model of influenza virus infection was successfully established. Compared with the M group, the lung index of mice in C, SH, SM, SL groups was significantly lower (0.96±0.14, 1.45±0.22, 1.14±0.18, 1.22±0.21 vs. 1.72±0.15, all P < 0.05), and the extent and degree of lesions were reduced, however, there was no significant difference among the groups. Gene chip analysis showed that there were more differentially expressed genes in N group vs. M group, SH group vs. M group, SM group vs. M group, SL group vs. M group. It could be used for further signal transduction pathway screening. Compared with N group, the differential gene expression of chemokine C-C ligands (CCL-3, CCL-5) and chemokine C-X-C ligands (CXCL-9, CXCL-10) in M group were significantly up-regulated [log 2 (M group/N group) were 6.64, 3.51, 5.40, 6.64, respectively]. Compared with M group, the gene expressions of CCL-3, CCL-5, CXCL-9 and CXCL-10 were significantly down-regulated in C, SH, SM and SL groups [log 2 (C group/M group) were -3.96, -2.26, -3.12, -2.40; log 2 (SH group/M group) were -5.57, -2.37, -1.57, -1.01; log 2 (SM group/M group) were -4.35, -1.47, -1.26, -1.74; log 2 (SL group/M group) were -2.86, -1.86, -1.23, -1.39, respectively]. Conclusion:Shufeng Xuanfei Jiedu formula inhibits inflammatory damage in mice after influenza virus infection by down-regulating the expressions of CCL-3, CCL-5, CXCL-9 and CXCL-10 on chemokine pathways.
4.Effects of Shufengxuanfeijiedu formula on Janus kinase signal transducer and transcription activator pathway in mice with influenza viral pneumonia
Qi LIU ; Jianguo WANG ; Yanping MA ; Haijun YUAN ; Wanfang YANG ; Ligang GU ; Shasha LING ; Peng ZHI ; Lu XIANG
Chinese Journal of Integrated Traditional and Western Medicine in Intensive and Critical Care 2016;23(5):449-452
ObjectiveTo investigate the regulatory effects of traditional Chinese medicine (TCM) Shufengxuanfeijiedu formula on Janus kinase signal transducer and activators of transcription (JAK-STAT) of lung tissues in mice with influenza viral pneumonia.Methods According to random number table, 60 mice were randomly divided into six groups with 10 mice in each group: normal group (N), model group (M), Tamiflu control group (C) and low (SL), medium (SM), high dose (SH) Shufengxuanfeijiedu formula groups. The mouse model of influenza virus pneumonia was reproduced by dropping of 0.05 mL 4LD50 inflluenza virus FM1 strain which can be adapted to lung tissue into the nose; while the N received nose instillation of 0.05 mL normal saline. After successful modeling for 2 hours, distilled water was given orally (by lavage) to N and M; Duffy (oseltamivir) 2.5 g·mL-1·d-1 was administrated to C; the TCM SL, SM, SH were intragastrically administered with different doses of shufengxuanfeijiedu decoction into the corresponding groups respectively (the ingredients of prescription: chrysanthemum, mulberry leaf, almond, platycodon root, forsythia, bupleurum etc. forming granules), according to the suitable dose of granules used for human body surface, the dose used for mouse surface area was calculated, the high dose means the dose used in the medium dose group doubled, the low dose means 1/2 dose used in medium group, once a day, once 0.2 mL for consecutive 4 days. Afterwards, the lung tissues were collected, the mouse differential gene expressions related to JAK-STAT pathway were detected by gene chip technology, the standards for screening of differential gene expression were as follows: up-regulated gene was P < 0.05, and the log2ratio > 1; down-regulation gene wasP < 0.05, and log2ratio < -1. The levels in lung tissue kinase (JAK) andγinterferon (IFN-γ) mRNA expressions were determined by real-time fluorescence quantitative reverse transcription polymerase chain reaction (RT-qPCR).Results Compared with those in N, the differential expression gene transcription activator, STAT5 [log2 (N/M) = 2.32], interleukin 4 receptor alpha subunit [IL4RA, log2 (N/M) = 4.77], interleukin 12 receptor [IL12R, log2 (N/M) = 1.58], JAK [log2 (N/M) = 2.41] were all obviously up-regulated, and IFN was significantly down-regulated [log2 (N/M) = -1.45] in M. Compared with those in M, C group IFN [log2 (C/M) = 1.51], various TCM dose groups [log2 (SL/M) = 1.46, log2 (SM/M) = 1.72, log2 (SH/M) = 1.40] differential expression gene IFN was significantly up-regulated, STAT5 [log2 (C/M) = -2.06, log2 (SL/M) = -1.41, log2 (SM/M) = -2.10, log2 (SH/M) = -1.89], IL4RA [log2 (C/M) = -2.52, log2 (SL/M) = -1.85, log2 (SM/M) = -2.74, log2 (SH/M) = -1.39), IL12R [log2 (C/M) = -1.48, log2 (SL/M) = -0.10, log2 (SM/M) = -1.58, log2 (SH/M) = -0.53], JAK [log2 (C/M) = -1.44, log2 (SL/M) = -0.88, log2 (SM/M) = -1.74, log2 (SH/M) = -0.53] were significantly down-regulated. In M, the JAK mRNA expression was obviously elevated (2-ΔΔCt: 3.17±0.94 vs. 1.01±0.13,P < 0.05), while the IFN-γ mRNA expression was decreased (2-ΔΔCt: 0.15±0.48 vs. 1.01±0.12,P < 0.05); compared with M, the JAK mRNA expressions in C, SM and SH groups were all obviously decreased (2-ΔΔCt: 2.02±0.63, 1.19±0.30, 1.59±0.67 vs. 3.17±0.94, allP < 0.05); while the IFN-γmRNA expressions in C, SL, SM and SH groups were elevated (2-ΔΔCt: 0.61±0.12, 0.41±0.13, 0.85±0.14, 0.78±0.20 vs. 0.15±0.48, allP < 0.05).Conclusions Shufengxuanfeijiedu formula can ameliorate the mice immune pathological injury of lung tissues induced by influenza virus by regulating JAK-STAT signal pathway and balancing Th1/2 via up-regulating the expression of IFN-γ.
5.Study on a novel Rho kinase inhibitor WAR5 for treating EAE
Zhibin DING ; Hui ZHANG ; Xingwang YANG ; Haifei ZHANG ; Jiezhong YU ; Yanhua LI ; Chunyun LIU ; Wanfang YANG ; Junlian LI ; Qianjin FENG ; Yongfei ZHAO ; Baoguo XIAO ; Cungen MA
Chinese Journal of Pathophysiology 2014;(9):1610-1615
AIM:To explore the therapeutic effect of a novel Rho kinase inhibitor WAR 5 on the experimental autoimmune encephalomyelitis (EAE) and its possible mechanism.METHODS: Female C57BL/6 mice were randomly divided into EAE group and WAR5 group.EAE model was induced by the application of MOG 35-55 peptide.WAR5 was in-jected intraperitoneally every other day from post-immunization (PI) day 3 to PI day 27.The clinical score and body weight were recorded every other day .On PI day 28, the animals were sacrificed and spinal cords were obtained for HE and mye-lin staining .The splenocytes were isolated and the expression of CD 16/32 and CD206 were analyzed by flow cytometry . The protein extracts from the brains and spinal cords were collected for the measurement of inducible nitric oxide synthase ( iNOS) by Western blotting .RESULTS:The administration of WAR 5 delayed the onset of EAE and attenuated the clini-cal symptoms .The results of the pathological examination revealed that WAR 5 inhibited the infiltration of inflammatory cells and improved myelination in spinal cords , accompanied with the poralization of M 1 macrophages to M2 phenotype in the spleen.WAR5 inhibited the expression of iNOS in the central nervous system , especially in the spinal cords .CON-CLUSION:The therapeutic effect of WAR5 on EAE may be related to the shift of M1 macrophages to M2 phenotype and inhibition of inflammation in the central nerve system .
6.Effects of Shufeng Xuanfei Jiedu formula for the inflammation-related cytokines in pneumonia mice infected with influenza virus
Qi LIU ; Jianguo WANG ; Yu GUO ; Lu JIA ; Haijun YUAN ; Wanfang YANG ; Yangxuan WEN
Chinese Critical Care Medicine 2020;32(2):183-187
Objective:To analyze the differential gene expression of Shufeng Xuanfei Jiedu formula on whole expression profiles of the inflammation-related cytokines in mice infected with influenza virus by the gene chip technology.Methods:Male ICR mice were divided into normal group (N group), influenza virus pneumonia model group (M group), oseltamivir control group (C group) and Shufeng Xuanfei Jiedu formula high, medium and low dose groups (SH, SM, SL groups) according to the random number table method, with 10 mice in each group. A mouse model of influenza virus pneumonia was established by nasal drip of influenza virus strain FM1 (0.05 mL); in group N, 0.05 mL normal saline was used. In SH, SM and SL groups, Shufeng Xuanfei Jiedu formula was prescribed after 2 hours of intranasal infection (drug concentration approximately 3.8, 1.9 and 1.0 kg/L), 0.2 mL once a day for 4 days; in group C, the dosage of oseltamivir was 2.5 kg/L; in group N and group M, distilled water was given. On the 5th day, the whole lung of mice was harvested, and the total RNA of lung tissue was extracted and detected after hybridization with mice whole gene expression spectrum chip. Differential expressed genes of cytokines involved in inflammatory pathways were selected. The intensity expression ratio of the chip probe signal in each group vs. M group was calculated, and P < 0.05 and log2 ratio > 1 were defined as up-regulated genes, while P < 0.05 and log2 ratio < -1 were down-regulated genes. The mRNA expressions of interleukin (IL-1, IL-8) and intercellular adhesion molecule-1 (ICAM-1) were detected by reverse transcription-polymerase chain reaction (RT-PCR). Results:Compared with group N, the differential gene expressions of IL-1, IL-8 and ICAM-1 in group M were significantly up-regulated [log2 (N/M) were 2.62, 2.07, 1.41, respectively, all P < 0.05]. Compared with group M, the gene expressions of IL-1, IL-8, ICAM-1 were significantly down-regulated in SH, SM, SL and C groups [log2 (SH/M) were -1.91, -1.85, -0.88; log2 (SM/M) were -3.10, -1.74, -1.84; log2 (SL/M) were -1.89, -1.39, -0.53; log2 (C/M) were -2.46, -1.52, -1.44, respectively, all P < 0.05]. RT-PCR showed that the mRNA expressions of IL-1, IL-8 and ICAM-1 in group M were significantly higher than those in group N [IL-1 (2 -ΔΔCT): 4.63±0.24 vs. 1.01±0.13, IL-8 (2 -ΔΔCT): 6.28±0.13 vs. 1.02±0.09, ICAM-1 (2 -ΔΔCT): 2.90±0.18 vs. 1.02±0.12, all P < 0.05]. The mRNA expressions of IL-1, IL-8, ICAM-1 in SH, SM, SL and C groups were lower than those in group M [IL-1 (2 -ΔΔCT): 2.12±0.32, 1.71±0.07, 2.05±0.16, 1.66±0.13 vs. 4.63±0.24; IL-8 (2 -ΔΔCT): 3.89±0.13, 2.08±0.19, 2.98±0.20, 2.02±0.12 vs. 6.28±0.13; ICAM-1 (2 -ΔΔCT): 1.72±0.93, 1.34±0.14, 1.53±0.25, 1.17±0.12 vs. 2.90±0.18, all P < 0.05]. There was no significant difference among the SH, SM, SL and C groups. Conclusion:Shufeng Xuanfei Jiedu formula inhibits inflammatory damage in mice after influenza virus infection by down-regulating the expressions of IL-1, IL-8, and ICAM-1 inflammatory cytokine-related genes.
7.Effect of microRNA-181b-5p on the proliferation and invasion of cutaneous melanoma cells and its mechanisms
Li XIA ; Linhong YANG ; Li XU ; Wenguo SUN ; Liang YU ; Wanfang ZHAI ; Dongxia WANG ; Xiaowan KUANG
Chinese Journal of Dermatology 2022;55(7):588-595
Objective:To explore whether microRNA (miRNA) -181b-5p inhibits the proliferation and invasion of cutaneous melanoma cells by targeting pleckstrin (PLEK) .Methods:Bioinformatics methods were used to analyze cutaneous melanoma-associated core genes; dual-luciferase reporter assay was performed to verify the targeted interaction between miRNA-181b-5p and PLEK. Oligo RNA and small interfering RNA (siRNA) were used to regulate the expression of miRNA-181b-5p and PLEK in A375 cells respectively in this experiment, and A375 cells were divided into the following groups in detail: mimic negative control group, miRNA-181b-5p mimic group, inhibitor negative control group, miRNA-181b-5p inhibitor group, PLEK siRNA group, siRNA negative control group, miRNA-181b-5p inhibitor + control siRNA co-transfection group and miRNA-181b-5p inhibitor + PLEK siRNA3 co-transfection group. After 48-hour treatment, qPCR was performed to determine the mRNA expression of miRNA-181b-5p and PLEK in A375 cells, Western blot analysis to determine the PLEK protein expression, and Transwell assay to assess the invasive ability of A375 cells; after additional 24-96 hours of culture, cell counting kit-8 (CCK8) assay was conducted to assess the proliferative ability of A375 cells.Results:PLEK was the core gene for cutaneous melanoma. PLEK expression in the cutaneous melanoma in situ tissues was significantly higher than that in the paracancerous tissues ( P = 0.031) , but lower than that in the metastatic tissues ( P = 0.001) . Compared with human epidermal melanocytes HEMa-LP, the mRNA and protein expression of PLEK significantly increased in A375 cells (mRNA: 3.884 ± 0.156 vs. 0.997 ± 0.010, t = 18.48, P < 0.001; protein: 2.840 ± 0.301 vs. 1.029 ± 0.094, t = 5.47, P = 0.005) , but the miRNA-181b-5p expression significantly decreased in A375 cells (0.333 ± 0.042 vs. 0.967 ± 0.069, t = 7.83, P = 0.001) . Dual-luciferase reporter assay showed targeted binding of miRNA-181b-5p to PLEK. Compared with the mimic negative control group, the miRNA-181b-5p mimic group showed significantly decreased survival rate of A375 cells (48 hours: t = 7.96, P = 0.015; 72 hours: t = 7.50, P = 0.002; 96 hours: t = 7.96, P = 0.001) , and significantly decreased invasive ability of A375 cells ( t = 5.07, P = 0.007) ; on the contrary, the survival rate and invasive ability of A375 cells were significantly higher in the miRNA-181b-5p inhibitor group than in the inhibitor negative control group (survival rate: 24 hours, t =5.38, P = 0.013; 48 hours, t = 5.36, P = 0.013; 72 hours, t =7.63, P = 0.005; 96 hours, t = 5.99, P = 0.004; invasive ability: t = 7.24, P = 0.002) ; compared with the siRNA negative control group, the proliferative and invasive ability of A375 cells significantly decreased in the PLEK siRNA group (proliferative ability: 48, 72, 96 hours, P = 0.015, 0.011, 0.001, respectively; invasive ability: t = 4.93, P = 0.008) ; compared with the miRNA-181b-5p inhibitor + control siRNA co-transfection group, the miRNA-181b-5p inhibitor + PLEK siRNA co-transfection group showed significantly decreased proliferation rate and invasive ability of A375 cells (proliferation rate: 24, 48, 72, 96 hours, P = 0.042, 0.042, 0.037, 0.017, respectively; invasive ability: t = 8.52, P = 0.001) . Conclusion:miRNA-181b-5p can inhibit the proliferation and invasion of cutaneous melanoma A375 cells, likely by down-regulating the PLEK expression.
8. Effects of sustained lung inflation combined with pulmonary surfactant on neonatal respiratory distress syndrome: a prospective randomized controlled trial
Junyan ZHONG ; Haifeng ZONG ; Nan YE ; Mei HUANG ; Yurong YUAN ; Sue ZHANG ; Wanfang ZHANG ; Lin ZHU ; Shujuan ZHANG ; Zhifeng HUANG ; Yuping SHI ; Chuanzhong YANG
Chinese Journal of Perinatal Medicine 2019;22(11):781-786
Objective:
To investigate the efficacy and adverse effects of sustained lung inflation (SLI) combined with pulmonary surfactant (PS) in the treatment of neonatal respiratory distress syndrome (NRDS).
Methods:
This prospective randomized controlled trial included 124 premature infants (gestational age <34 weeks and birth weight <2 000 g) diagnosed with NRDS and in need of PS treatment in Shenzhen Maternity & Child Healthcare Hospital affiliated to Southern Medical University from July 1, 2016 to October 31, 2018. They were randomly divided into experimental or control group, with 62 cases in each. Infants in the experimental group were treated with SLI using T-piece and intratracheal PS, while those in the control group were given PS only. Blood gas analysis and measurement of fraction of inspiration O2 (FiO2) and ratio of partial pressure of oxygen (PO2) over FiO2 were performed before and 1 h after PS injection. Results of the treatments and incidence of complications were compared. Paired samples
9.Integrated mass spectrometry imaging reveals spatial-metabolic alteration in diabetic cardiomyopathy and the intervention effects of ferulic acid
Yanhua LIU ; Xin ZHANG ; Shu YANG ; Zhi ZHOU ; Lu TIAN ; Wanfang LI ; Jinfeng WEI ; Zeper ABLIZ ; Zhonghua WANG
Journal of Pharmaceutical Analysis 2023;13(12):1496-1509
Diabetic cardiomyopathy(DCM)is a metabolic disease and a leading cause of heart failure among people with diabetes.Mass spectrometry imaging(MSI)is a versatile technique capable of combining the molecular specificity of mass spectrometry(MS)with the spatial information of imaging.In this study,we used MSI to visualize metabolites in the rat heart with high spatial resolution and sensitivity.We optimized the air flow-assisted desorption electrospray ionization(AFADESI)-MSI platform to detect a wide range of metabolites,and then used matrix-assisted laser desorption ionization(MALDI)-MSI for increasing metabolic coverage and improving localization resolution.AFADESI-MSI detected 214 and 149 metabolites in positive and negative analyses of rat heart sections,respectively,while MALDI-MSI detected 61 metabolites in negative analysis.Our study revealed the heterogenous metabolic profile of the heart in a DCM model,with over 105 region-specific changes in the levels of a wide range of metabolite classes,including carbohydrates,amino acids,nucleotides,and their derivatives,fatty acids,glycerol phospholipids,carnitines,and metal ions.The repeated oral administration of ferulic acid during 20 weeks significantly improved most of the metabolic disorders in the DCM model.Our findings provide novel insights into the molecular mechanisms underlying DCM and the potential of ferulic acid as a therapeutic agent for treating this condition.