2.Preparation of fusiform aneurysms model in rabbits
Lianfu ZHANG ; Shanshui XU ; Xinggen FANG ; Zifu LI ; Guoquan JIANG
Journal of Interventional Radiology 2014;(8):711-715
Objective To establish the carotid fusiform aneurysm model in rabbits carrying similar characteristics of human intracranial aneurysms by using induction method with porcine pancreatic elastase. Methods Twenty-five New Zealand white rabbits were randomly divided into normal control group (n=5), saline control group (n = 5) and study group (n = 15). The rabbits of the study group were randomly and equally subdivided into 7-day subgroup, 14-day subgroup and 21-day subgroup. By using induction method with porcine pancreatic elastase to digest right common carotid the fusiform aneurysm model was established in all the rabbits of the study group. DSA examination , HE staining and elastic fiber staining pathologic examination were carried out at 7, 14 and 21 days after the procedure to observe the imaging and pathologic changes of the fusiform aneurysm models. Results DSA angiography showed that the mean vascular diameters of the normal control group and the saline control group were (1.64 ± 0.17) mm and (1.66 ± 0.24) mm respectively. The mean length and width of the fusiform aneurysm of the 7-day subgroup, 14-day subgroup and 21-day subgroup were (19.33 ± 1.65) mm and (2.86 ± 0.21) mm, (19.66 ± 1.18) mm and (3.95 ± 0.54) mm, and (19.84 ± 0.82) mm and (4.03 ± 0.95) mm, respectively. Pathologically, rupture of internal elastic membrane, disordered structure of tunica media smooth muscle and distortion of cell shape were observed in the rabbits of 7-day subgroup. Gradually stabilized aneurysmal lumen intimal hyperplasia was seen in the rabbits of 14-day subgroup. Remarkable structure changes at the aneurysmal neck-cavity junction were found in the rabbits of 21-day subgroup. Elastic fiber staining demonstrated that strikingly thinned elastic layer was observed in the rabbits of 7-day subgroup, gradually thinning elastic layer at the aneurysmal neck-cavity junction was seen in the rabbits of 14-day subgroup, and the thinned elastic layer became stable in the rabbits of 21-day subgroup. Conclusion Using simple surgical method combined with porcine pancreatic elastase to digest vascular wall, carotid fusiform aneurysm models can be reliably established in New Zealand white rabbits which carry similar morphologic and pathologic characteristics of human intracranial aneurysms.
3.Vascular endothelial growth factor promotes the proliferation and osteogenic differentiation of mouse bone marrow mesenchymal stem cells
Lianfang ZHANG ; Hui KANG ; Lianfu DENG ; Huilin YANG
Chinese Journal of Tissue Engineering Research 2014;(41):6591-6596
BACKGROUND:Vascular endothelial growth factor (VEGF) is a crucial factor for regulating bone marrow mesenchymal stem cells in microenvironment, which can promote endothelial differentiation of bone marrow mesenchymal stem cells. But there is no report about VEGF effect on regulating the proliferation and osteogenic differentiation of bone marrow mesenchymal stem cells. OBJECTIVE:To investigate the regulatory role of VEGF in the proliferation and osteogenic differentiation of bone marrow mesenchymal stem cells. METHODS:The bone marrow mesenchymal stem cells were isolated and cultured from mouse long bones. cellCounting Kit-8 was used to test the proliferation of bone marrow mesenchymal stem cells cultured with different concentrations of recombinant VEGF proteins. The appropriate concentration of VEGF recombinant protein was selected to test their effects on the osteogenic differentiation of bone marrow mesenchymal stem cells. The mRNA and protein levels of Osterix, Runx2, alkaline phosphatase, osteocalcin and heme oxygenase-1 in bone marrow mesenchymal stem cells under induction of VEGF were detected by molecular biology methods. RESULTS AND CONCLUSION:(1) VEGF promoted the proliferation of bone marrow mesenchymal stem cells dose-dependently, and 100μg/L was the optimum concentration. (2) VEGF promoted the expression of Osterix, Runx2, alkaline phosphatase and osteocalcin in bone marrow mesenchymal stem cells under osteogenic induction. Similar results were obtained by alizarin red staining and quantification of numbers of calcium nodules. (3) VEGF induced the expression of heme oxygenase-1 in bone marrow mesenchymal stem cells at mRNA and protein levels. These findings indicate that VEGF can induce the expression of heme oxygenase-1 i in bone marrow mesenchymal stem cells, and promote the proliferation and osteogenic differentiation of bone marrow mesenchymal stem cells.
4.Induction and mechanism of valdecoxib on the apoptosis of human esophageal cancer cells
Yujun ZHANG ; Shuxia LIU ; Fengying QI ; Lianfu ZUO ; Jianwen GUO
Chinese Pharmacological Bulletin 1986;0(05):-
Aim To investigate the effect and mechanism of valdecoxib on the apoptosis of human esophageal cancer cells.Methods Flow cytometry was used to observe the effect of valdecoxib on apoptosis and the cell cycle distribution of Eca109 cells.Transmission electron microscope was further used to detect the cell apoptosis.The content of LDH was examined using LDH kit.The expressions of p-p38MAPK,Fas and FasL protein were detected using flow cytometry.Results Valdecoxib of 25~400 ?mol?L~(-1) significantly induced the apoptosis of Eca109 cell line,and the rate of apoptosis was increased from(2.95?0.83)% to(48.46?0.73)%,50~400 ?mol?L~(-1) valdecoxib also decreased the proliferation index and the proportion of cells in the S phase,increased the proportion of cells in the G_0/G_1 phase,but had no effect on the proportion of cells in the G_2/M phase.Compared with those in Eca109 cells cultured in the medium with solvent,the expression of p-p38MAPK,Fas and FasL was higher in the Eca109 cells exposed to valdecoxib in a dose-dependent manner in 72 h.Conclusion Valdecoxib can induce apoptosis of Eca109 cell line partly by up-regulating the expression of p-p38MAPK/Fas/FasL.
5.The expression of STATI/SOCS1 on the rat synoviocytes induced by high mobility group box
Huifang GUO ; Shuxia LIU ; Yujun ZHANG ; Jingkun ZHANG ; Lianfu ZUO ; Jianwen GUO ; Xin ZHANG ; Huichao ZHANG
Chinese Journal of Rheumatology 2008;12(7):-
Objective To investigate the effect and possible mechanisms of high mobility group box (HMGB) 1 on the proliferation of RSC-364 synoviocytes. Methods ① RSC-364 cells stimulated by 10 μg/L TNF-α and cells of the normal control groups were collected at 6, 12, 24 h respectively in vitro. HMGB1mRNA and protein was detected by reverse transcription polymerase chain reaction (RT-PCR) and immunocytochemistry (ICC); ②RSC-364 cells induced by 10 μg/L HMGB1 were collected in 6, 12, 24 h respectively, so did normal control group cells in vitro. The expression of signal transducer and activator of transcription (STAT) mRNA 1 was detected by RT-PCR. The expression of STATland SOCSI proteins were detected by ICC and flow cytometry analysis (FCM). The expression of PCNA was detected by ICC. Results ① Compared with the control group, TNF-α markedly up-regulated HMGBI mRNA at 6, 12, 24 h respectively [0.86, 0.92, 1.06 vs 0.70, P<0.01 ], as well as protein expression level. Positive signal of HMGB1 proteins was not only expressed in nuclear but also in cytoplasm after stimulation. ② Compared with normal group, HMGBI increased the expression of P-STAT1 mRNA and protein at 6, 12, 24 h respectively [0.30, 0.69, 1.05 vs 0.24, P<0.01 ] and [1.34±0.09,1.55±0.16,1.74±0.13 vs 1.00±0.15,P<0.01]. The expression of SOCSI protein increased significantly in HMGB1 group at 6 and 12 hours ( 1.43±0.10 vs 1.58±0.05), but it decreased at 24 hours (1.24±0.15). ③The expression of p-STATI protein was negatively correlated with that of SOCS1 protein. Conclusion HMGB1 appears to be an important mediator in the proliferation of RSC-364 cells, partly by up-regulating the expression and aetivity of p-STAT1.
6.Analysis of the Long-term Curative Effects of Radical Nephrectomy
Jinsheng XU ; Junxia ZHANG ; Tonghui GENG ; Yuefen WANG ; Aili ZHANG ; Lianfu ZUO
Chinese Journal of Clinical Oncology 2009;36(14):784-787
Objective:To analyze the factors that affect patient prognosis after radical nephrectomy.Meth-ods:A total of 389 cases of renal cell carcinoma treated with radical nephrectomy between January 1 993 and December 2006 were reviewed.All the data were encoded.inserted into an Excel database and then ana-lyzed by SPSS 1 3.0 software.The cumulative survival rates were calculated by life-table method.We as-sessed the impact of multiple covariates on survival time with the Cox Regression model.Results:The patho-logical results showed that 307 cases were clear call carcinoma,51 cases were papillary renal cell carcinoma,21 cases were chromophobic renal cell carcinoma,2 cases were collecting duct carcinoma.and 8 cases were unclassified.One hundred and ninety-eight cases were of T1N0M0, 113 cases were of T2N0M0, 3 cases were of T1N1M0,10 cases were of T2N1M0, 51 cases were of T3N0M0, and 14 cases were of T3N1M0, Two hundred and sixty-eight cases were followed up.The 1-year survival rate was 96.5%,the 3-year survival rate was 90.7%.the 5-year survival rate was 75.7%.and the 10-year survival rate was 65.8%.Multivariable analysis revealed that significant prognostic factors included TNM stage,Robson stage.vena cava and supplementary treat-ment(X2=22.50.P=0.001).The most important prognostic factor was pathological stage(TNM and Robson).The regression coefficients were 0.533 and 0.674,and the relative risk was 1.941 and 2.01 1(P=0.004 and p=0.002).Conclusion:Radical nephrectomy is safe and effective.TNM stage.Robson stage and vena cava are prognostic factors.Supplementary treatment is a protective factor.
7.Arsenic trioxide inhibits proliferation of rat synoviocyte line stimulated with TNF-? and its mechanism
Huifang GUO ; Shuxia LIU ; Yujun ZHANG ; Lianfu ZUO ; Jianwen GUO ; Xin ZHANG
Basic & Clinical Medicine 2006;0(04):-
Objective To investigate the effect and possible mechanism of arsenic trioxid(As2O3)on proliferation of RSC-364 synoviocyte lines stimulated with TNF-?.Methods RSC-364 synoviocytes were cultured with standard medium as control group or medium supplemented with 10 ?g/LTNF-? and different concentrations of As2O3 respectively. MTT assay were carried out to study cell proliferation. Proliferation index (PI) and cell cycle were detected by flow cytometry (FCM). RT-PCR was used to detect the mRNA expression of High mobility group box chromosomal protein (HMGB)1. HMGB-1 and proliferation cell nuclear antigen (PCNA) proteins were detected by immunocytochemistry and FCM. Results (1)As2O3 inhibited proliferation of cell lines stimulated by TNF-? time-dependently and dose-dependently. (2)Compared with normal group, TNF-? up-regulated HMGB-1 protein and mRNA as well as PCNA protein. HMGB-1 protein was not only in nuclear but also in cytoplasm by immunocy-tochemistry. As2O3 down-regulated mRNA and protein of HMGB-1 in a dose-dependent manner; so did PCNA proteins (P
8.Expression of core-binding factor a1 by human skin fibroblasts induced in vitro.
Lianfu DENG ; Wei FENG ; Yue ZHANG ; Yaping ZHU
Chinese Journal of Surgery 2002;40(8):592-595
OBJECTIVETo investigate the probabilities of core-biding factor a1 (Cbfa1) expression by human skin fibroblasts induced in vitro.
METHODSThe fibroblasts were isolated, purified from human skin, and were grown in incubation in the media of TNF-alpha, BMP-2, and combined TNF-alpha and BMP-2 at certain concentrations, respectively. The changes in biological features of these fibroblasts correlated with osteogenesis were detected by immunohistochemistry and RT-PCR assay.
RESULTSTNF-alpha could switch phenotype of collagen in fibroblasts from Type I and III to Type I and induce fibroblasts to express Ras and BMP type I receptor (BMPR-IA). TNF-alpha in combination with BMP-2 could induce fibroblasts to express Cbfa1 and osteocalcin mRNA.
CONCLUSIONHuman skin fibroblast could be induced into pro-osteoblast expressing Cbfa1, an osteoblast-specific transcription factor and a regulation of osteoblast differentiation, and combined use of TNF-alpha and BMP-2 was one of the regulating factors.
Bone Morphogenetic Protein 2 ; Bone Morphogenetic Protein Receptors, Type I ; Bone Morphogenetic Proteins ; pharmacology ; Cells, Cultured ; Collagen ; biosynthesis ; Core Binding Factor Alpha 1 Subunit ; Core Binding Factors ; Fibroblasts ; metabolism ; Humans ; Neoplasm Proteins ; Osteocalcin ; biosynthesis ; Protein-Serine-Threonine Kinases ; biosynthesis ; RNA, Messenger ; analysis ; Receptors, Growth Factor ; biosynthesis ; Skin ; cytology ; Transcription Factors ; biosynthesis ; genetics ; Transforming Growth Factor beta ; Tumor Necrosis Factor-alpha ; pharmacology
9.Inducing common carotid fusiform aneurysms in rabbit with the porcine pancreatic elastase extravascular digestion method
Zifu LI ; Guoquan JIANG ; Xinggen FANG ; Lianfu ZHANG ; Yibin FANG ; Qinghai HUANG ; Jianmin LIU
Chinese Journal of Cerebrovascular Diseases 2014;(8):420-423
Objective To investigate the feasibility and effectiveness of inducing rabbit common carotid fusiform aneurysms via the common carotid extravascular digestion method. Methods Sixteen New Zealand white rabbits were randomly assigned into either an experiment group ( n=12 ) or a control group (n=4). Porcine pancreatic elastase 80-400 U were used to incubate and digest 2 to 4 cm segment of artery distal to the origin of right common carotid artery. One week after modeling,intravenous angiography was performed and the length and width of fusiform dilatation of common carotid artery were measured. The fusiform dilated artery was examined with hematoxylin and eosin staining and the vascular morphological changes were observed with scanning electron microscope. Isotonic saline solution was used to incubate common carotid arteries of the 4 New Zealand white rabbits in the control group. After one week,the same method was used to observe the lumen of common carotid artery and intimal changes. Results After the digestion of common carotid artery adventitia,the angiography of 12 New Zealand white rabbits of the experimental group revealed fusiform dilatation of common carotid artery of the 10 model rabbits. The widest diameter of the fusiform artery was 3. 70 ± 0. 32 mm;two rabbits had common carotid artery occlusion. Compared with the control group,the right common carotid artery diameter enlarged significantly in the experimental group (1. 80 ± 0. 16 mm,P<0. 01). The HE staining showed that the lumen widened, adventitia and media reduced. Scanning electron microscope showed intimal inflammatory injury and thrombus attachment. Conclusion Using porcine pancreatic elastase to digest the adventitia of common carotid artery can make fusiform dilatation of common carotid artery in rabbits. Using this method may effectively induce a model of fusiform aneurysm,and it has certain feasibility.
10.The value of HMGB1 and Toll-like receptor 4 in peripheral blood mononuclear cells and serum of rheumatoid arthritis
Huifang GUO ; Shuxia LIU ; Xiaolei LIU ; Lianfu ZUO ; Lijuan TANG ; Jianghui LIU ; Mingfeng ZHANG ; Liyan MA
Chinese Journal of Rheumatology 2009;13(5):333-336
Objective To investigate the effect and possible mechanism of high mobility group box (HMGB) 1 in the development and progress of rheumatoid arthritis.Methods PBMC and serum samples were obtained from 74 RA patients (38 in active stage and 36 in stable stage) and 26 healthy controls.The expression of HMGB1 mRNA and protein was detected by RT-PCR and ELISA.Flow cytometry analysis ( FCM ) was used to detect the expression of Toll-like receptor 4 on PBMC.Results ①The expression of HMGBI mRNA and protein in active RA patients was significantly higher than that in healthy controls and inactive RA patients [2.63 vs 0.71,0.93 and (10.2±1.2) vs (7.5±1.8),(8.3±1.8) ng/ml,respectively](P<0.01 ).② The relative expression of TLR4 protein on CD14+ monocytes and CD3+ lymphocytes in active RA patients was increased than that in inactive RA and healthy controls (P<0.05 or P<0.01 ).It was also higher in inactive RA than in healthy controls (P<0.05 or P<0.01 ).③ Level of HMGB1 protein in serum of RA patients was positively correlated with ESR,CRP,RF,the numbers of tender joints and swollen joints as well as radiographic changes.Conclusion HMGB1 can be synthesized and released by PBMC of active RA patients,and then bind to TLR4 of PBMC to promote inflammatory responses and bone erosion.