共查询到20条相似文献，搜索用时 390 毫秒
1.
AIM: To investigate the effects of obesity on mouse oocyte in vivo aging, its in vitro fertilization and embryo development. METHODS: The oocyte-cumulus granulosa cell complexes were obtained at the time points of 14 h, 18 h and 22 h after HCG injection. The in vitro fertilization and embryo culture were conducted. Hoechst 33342 and propidium iodide double staining were performed to determine the apoptotic rate of cumulus granulosa cells. JC-1 and DCFH-DA were used to detect the levels of mitochondrial membrane potential(MMP) and reactive oxygen species (ROS), respectively. The levels of ATP and glutathione (GSH) in oocytes were also detected. RESULTS: The GSH level in obese mouse oocytes just after ovulation was higher than that in control group (P<0.05). No difference of blastocyst formation rate, ATP, ROS and cumulus granulosa cell apoptotic rate between obesity group and control group was observed (P>0.05). However, 18 h after HCG injection, ATP content and blastocyst cell numbers in obese mouse oocytes were less than those in control group (P<0.05), while ROS level in oocytes and the apoptotic rate of cumulus granulosa cells were higher than those in control group (P<0.05). Compared with control group, MMP and blastocyst formation rate were significantly decreased in obesity group 22 h after HCG injection (P<0.05). CONCLUSION: Obesity promotes oocyte aging after ovulation, and oxygen free radicals may play an important role in obesity-induced decrease of maternal infertility. 相似文献
2.
AIM: To investigate the expression of microRNA-141 (miR-141) in human hepatocellular carcinoma (HCC) cell line SMMC-7721 and normal hepatocyte line HL-7702, and to analyze the effect of abnormal expression of miR-141 on the malignant biological behaviors of human hepatocarcinoma cells. METHODS: The RNA from SMMC-7721 cells and HL-7702 cells was extracted. SYBR Green real-time PCR was performed to detect the expression of miR-141. Synthetic miR-141 mimic and its negative control were transfected into the SMMC-7721 cells, and miR-141 inhibitor and its negative control were transfected into the HL-7702 cells by the method of Lipofectamine. After transfection, MTS assay and BrdU-ELISA were employed to evaluate the effect of miR-141 on the cell proliferation. Flow cytometry was used to detect cell cycle and apoptosis. The changes of migration ability were investigated by Transwell invasion assay. RESULTS: The expression of miR-141 in the SMMC-7721 cells was significantly lower than that in the HL-7702 cells (P < 0.05). Compared with blank group, Lipofectamine group and negative control group, the proliferation of the SMMC-7721 cells transfected with 25 nmol/L miR-141 mimic was significantly inhibited in a time-dependent manner (P < 0.05). The percentages of G1 phase cells and early apoptotic rate were significantly increased when miR-141 was up-regulated, but the migration ability was inhibited (P < 0.05). Compared with blank group, Lipofectamine group and negative control group, the proliferation of HL-7702 cells transfected with 50 nmol/L miR-141 inhibitor was significantly increased in a time-dependent manner (P < 0.05). When miR-141 was down-regulated, the percentages of G1 phase cells and early apoptotic rate were significantly decreased, but the migration ability was enhanced (P < 0.05). CONCLUSION: miR-141 is down-regulated in human hepatocarcinoma cell line. Up-regulation of miR-141 will not only inhibit cell proliferation and migration ability, but also affect the cell cycle and apoptosis of SMMC-7721 cells. miR-141 may function as a tumor suppressor gene during HCC development. 相似文献
3.
MA Ji GUO Cheng-shan ZHAO Xia HE Wei-dong WANG Ying-xue DONG Qiao-feng ZHAO Jing-jie 《园艺学报》2009,25(8):1570-1575
AIM: To analyze the effect of mesenchymal stem cells (MSCs) on secreting cytokines by T lymphocytes from patients with idiopathic thrombocytopenic purpura (ITP) in vitro.METHODS: Human bone marrow-derived MSCs were isolated by Ficoll Hypaque and cultured for proliferating to passage cells. Allogeneic T lymphocytes of ITP were isolated from peripheral blood by Ficoll Hypaque and nylon cotton column. Then the stromal feeder layers of different numbers (2×103, 1×104, 5×104 per well) of MSCs treated with mitomycin were co-cultured with above-mentioned T lymphocytes. The supernatant were respectively collected on day 2, 4 and 6 after co-culture, then the levels of IL-2, IFN-γ, IL-4, IL-10 secreted by T lymphocytes were measured by enzyme linked immunosorbent assay (ELISA) dynamically.RESULTS: The levels of IL-2 and IFN-γ secreted by T cells from ITP were higher than those from normal control (P<0.05, respectively). Inversely, IL-4 and IL-10 were lower than those in normal control (P<0.05, respectively). After co-cultured with T lymphocytes, MSCs significantly inhibited the cytokine levels of IL-2 and IFN-γ secreted by T lymphocytes from ITP or health adults (P<0.05, respectively) in a dose dependent manner (P<0.05, respectively), and the effect was more obvious when co-cultured for 4 days or 6 days than that for 2 days (P<0.05, respectively). However, MSCs significantly promoted the releases of IL-4 and IL-10 by T lymphocytes from ITP patients (P<0.05, respectively) in a dose dependent manner (P<0.05, respectively), and the effect on IL-10 was in a time dependent way (P<0.05), while the effect on IL-4 had no obvious difference among 2 d, 4 d and 6 d(P>0.05). As for health control group, when cell numbers exceeded above 1×104, MSCs obviously promoted IL-4 and IL-10 levels secreted by T lymphocytes (P<0.05) in a dose dependent manner (P<0.05), and both of the effects were more noticeable when co-cultured for 4 d or 6 d than that for 2 d(P<0.05, respectively).CONCLUSION: MSCs regulate the balance between Th1 and Th2 reaction and partly correct ITP Th1 polarization in vitro. 相似文献
4.
AIM: To investigate mutations of oncogene k-ras in colorectal cancer tissues and the relationship between mutations of k-ras and biological behavior of colorectal carcinoma. METHODS: The specimens of 123 patients with colorectal cancer were collected. Real-time fluorescence quantitative PCR were performed to detect k-ras mutations at codon 12 and codon 13 of exon 1, and the results were analyzed with the corresponding clinical pathological data. RESULTS: Among 123 colorectal cancer cases, point mutations were detected in 53 cases (40.8%), point mutations at codon 12 were found in 42 (34.1%) cases, and 11(8.9%) cases at codon 13. No closely relationship between mutations of k-ras and tumor size, location, invasive depth and differentiation extent was observed. The rate of k-ras mutation in the cases with more invaded lymph nodes was higher than that in the cases without invaded lymph nodes (P<0.05), and the rate of k-ras gene mutation in the cases with hepatic metastases was higher than that in no hepatic metastases (P<0.05). The rate of k-ras gene mutation was higher in TNM staging Ⅲ/Ⅳ than that in Ⅰ/Ⅱ(P<0.05). CONCLUSION: Mutation of oncogene k-ras plays an important role in the carcinogenesis and development of colorectal cancer, and it is closely associated with invaded lymph notes and hepatic metastases, suggesting that mutation of k-ras indicates a poor prognosis. 相似文献
5.
AIM: To explore the expression and activation of matrix metalloproteinase-9 (MMP-9) from monocyte-derived macrophages induced by tumor necrosis factor-α (TNF-α) and to investigate its association with progression of joint damage in patients with rheumatoid arthritis. METHODS: TNF-α and MMP-9 in serum and synovial fluid from patients with early RA and controls were tested with a double-antibody enzyme-linked immunosorbent assay. The correlation between MMP-9 and Larsen score over the first 12 months was analyzed. THP-1 cells differentiated by the treatment with TPA were stimulated with increasing concentration of TNF-α for 24 h in vitro. The protein expression of MMP-9 was determined by Western blotting. The activity of MMP-9 was measured by gelatinolytic zymography. Boyden chamber-matrigel in vitro invasion assay was used to detect the invasive capacity. RESULTS: The levels of TNF-α and MMP-9 in serum and synovial fluid of RA patients were significantly higher than those in controls (P<0.05). Serum and synovial fluid levels of MMP-9 correlated significantly with Larsen score (r=0.37 and 0.32, P<0.01). The MMP-9 activity and invasive ability of co-cultured THP-1 cells with TNF-α and TPA were higher than those of non-TNF-α treatment. CONCLUSION: TNF-α upregulates MMP-9 activation and promotes infiltration of monocyte-derived macrophages, indicating that TNF-α play an important role in the pathogenesis of RA. 相似文献
6.
AIM: To investigate the immunomodulatory effect of human umbilical cord mesenchymal stem cells with interleukin-17 receptor-like molecule overexpression (IL-17RLM-hUCMSCs) on the spleen lymphocytes from the mice with trinitrobenzene sulfonic acid (TNBS)-induced colitis for providing optimal seed cells to treat inflammatory bowel disease. METHODS: Mesenchymal stem cells were isolated from human umbilical cord. The IL-17RLM gene was transferred into mesenchymal stem cells by lentivirus vector to establish IL-17RLM-hUCMSCs. The experimental colitis mice were induced by TNBS enema, and the spleen lymphocyte suspension was isolated. The lymphocytes were co-cultured with different concentrations of IL-17RLM-hUCMSCs and hUCMSCs under the stimulation of concanavalin A (ConA) for 72 h. The proliferation of lymphocytes was detected by the methods of CCK8 assay and CFSE labeling with lymphocytes+ConA as positive control. The changes of lymphocyte subsets (Th1, Th2, Th17 and Treg) were examined by flow cytometry.RESULTS: Both hUCMSCs and IL-17RLM-hUCMSCs inhibited T-cell proliferation in vitro co-culture system (P < 0.05). When the ratios of MSCs to lymphocytes ranged from 1:1 to 1:10, the inhibitory rates were in a dose-dependent manner. IL-17RLM-hUCMSCs showed higher inhibitory rate than hUCMSCs within the effective concentration range (P < 0.05). Both hUCMSCs and IL-17RLM-hUCMSCs reduced the proportions of Th1 and Th17 cell subsets and increased Treg cell population of spleen lymphocytes from TNBS-induced colitis mice, and IL-17RLM-hUCMSCs showed a stronger inhibitory effect on Th17 cell subset (P < 0.05). CONCLUSION: IL-17RLM-hUCMSCs remarkably inhibit the proliferation of spleen lymphocytes from TNBS-induced colitis mice in a dose-dependent manner. Meanwhile, they regulate immune balance of T cells and have stronger inhibitory effect on Th17 subset. 相似文献
7.
AIM:To investigate the effects of invariant natural killer T-cells (iNKT cells) from ovalbumin (OVA)-induced asthmatic mice combined with OVA on the phenotypic and functional characteristics of bone marrow-derived dendritic cells (BMDCs) in vitro. METHODS:The BMDCs from wild-type (WT) BLAB/c mice were co-cultured with purified iNKT cells from WT mice immunized and challenged with OVA in the presence of 100 mg/L OVA (iNKT cells plus OVA group) or PBS (iNKT cells plus PBS group) for 20 h, and were also cultured with 50 mg/L LPS (LPS group), 100 mg/L OVA (OVA group) or PBS (PBS group) for 20 h. The expression of MHC-Ⅱ, CD40, CD86, and CD80 on the BMDCs was measured by flow cytometric analysis, and the levels of interleukin-12 (IL-12) p70, IL-6, tumor necrosis factor-α (TNF-α) and IL-10 in the culture supernatant were measured by ELISA. Splenic CD4+ T cells from DO11.10 transgenic mice were co-cultured for 48 h with the above mentioned BMDCs, and then the concentrations of IL-4 and interferon-γ (IFN-γ) in culture supernatants were measured by ELISA. RESULTS:The expression of MHC-Ⅱ, CD80, CD86 and CD40, and releases of proinflammatory cytokines by BMDCs in iNKT cells plus OVA group were comparable to those in the LPS group (P>0.05), but significantly higher than those in iNKT cells plus PBS group, OVA group, and PBS group (P<0.05 or P<0.01). The concentration of IL-4 in culture supernatants from BMDCs in iNKT cells plus OVA group co-cultured with DO11.10 CD4+ T cells was similar to that in LPS group (P>0.05), but markedly higher than that in iNKT cells plus PBS group, OVA group, and PBS group (P<0.01). CONCLUSION:Bone marrow-derived dendritic cells undergo immunogenic maturation upon interaction with iNKT cells in the presence of OVA. 相似文献
8.
AIM: To investigate the differential expression of annexin A2 (ANXA2) in gastric carcinoma and to analyze the relationship between ANXA2 expression and clinicopathological parameters of gastric carcinoma. METHODS: Pure gastric adenocarcinoma cells (GAC) and normal gastric epithelial cells (NGEC) in 15 patients with gastric cancer were acquired by laser capture microdissection (LCM). All peptide specimens after trypsin digestion were labeled with 18O/16O. Quantitatively identification of differential expression of the proteins betweem GAC and NGEC was performed by Nano-RPLC-MS/MS. The expression of ANXA2 in the 2 kinds of tissues was detected by Western blotting. Tissue microarray containing 75 pairs of gastric carcinoma and para-carcinoma tissues was used and the expression of ANXA2 in these specimens was detected by the method of immunohistochemistry (IHC). The relationship between ANXA2 expression and clinicopathological parameters of the pateints with gastric carcinoma was analyzed. RESULTS: A total of 78 differential proteins were identified and ANXA2 was up-expressed in GAC (2.32∶ 1), which was confirmed by Western blotting (P<0.01). The results of IHC showed that the correlations between the expression level of ANXA2 protein and invasive depth (T stage), lymph node metastasis (N stage), histological differentiation, TNM stage and the size of tumor were observed (P<0.01), but the correlations between the ANXA2 expression and sex, age and distant metastasis (M stage) were not found (P>0.05). CONCLUSION: The up-expressed ANXA2 may play an important role in the biological behavior of gastric cancer. 相似文献
9.
WU Bo YE Xin CHEN Lin YANG Jin ZHANG Han-chao LIU Jun-ying CHEN Liang DIAO Jian-jun 《园艺学报》2019,35(4):753-757
AIM:To investigate the effect of Toll-like receptor 7(TLR7) agonist on the anti-tumor activity of peripheral blood mononuclear cells (PBMCs) in the patient with renal cell carcinoma.METHODS:Primary renal cancer cells from the postoperative specimens of the patient were co-cultured with peripheral blood mononuclear cells from the same patient stimulated by TLR7 agonist. The cytokine levels in culture medium were measured by ELISA, and the cell cycle distribution of the renal cell carcinoma cells was analyzed by flow cytometry. The anti-tumor activity of PBMCs was evaluated by[51Cr] release trial. The protein levels of Skp2 and its downstream pathway molecules were determined by Western blot. RESULTS:TLR7 agonist increased the expression of interferon-γ(IFN-γ), tumor necrosis factor-α (TNF-α) and interleukin-2 (IL-2) in the co-cultured medium, which significantly inhibited the proliferation index of the renal cell carcinoma cells. The cytotoxicity of PBMCs to renal cell carcinoma cells was markedly increased (P<0.05). The protein level of Skp2 in renal cell carcinoma cells was decreased significantly after stimulation, which was consistent with the change of cell proliferation index of renal cell carcinoma cells(P<0.05). The protein level of p27 was increased significantly (P<0.05), which was opposite to the change of Skp2. However, no significant difference in the expression of p21 and p53 was observed. CONCLUSION:TLR7 agonist effectively enhances the anti-tumor activity of PBMCs and results in the growth of renal cell carcinoma cells inhibiting. The mechanism may be relate to the cell cycle arrest by inhibiting the Skp2/p27 pathway. 相似文献
10.
HAN Li-hui SUN Wen-sheng LIU Su-xia JIA Xiao-qing WANG Xiao-yan MA Chun-hong GAO Li-fen ZHANG Li-ning CAO Ying-lin 《园艺学报》2005,21(8):1486-1490
AIM: To explore the effect of TNF-related apoptosis inducing ligand (TRAIL), a new apoptotic inducing molecule on the biological activity of hepatocarcinoma cell line. METHODS: The expression of membrane binding TRAIL on HepG2 cells was detected by immuno-cytochemistry. Quantity of secretory TRAIL was assayed by ELISA method. The cytotoxicity and apoptosis induced by TRAIL was detected by MTT and TUNEL method, respectively. The telomerase activity of HepG2 cells was detected by TRAP-PCR assay kit. The expression of hTERT, the catalytic subunit of telomerase, was detected by FCM. RESULTS: TRAIL was constitutively expressed on the membrane of HepG2 cell line. Soluble TRAIL was also expressed to a certain degree. Cytotoxicity assay showed that TRAIL significantly inhibited the growth of hepatocarcinoma cells. TUNEL assay indicated that TRAIL induced apoptosis in hepatocarcinoma cells. Detection of telomerase activity showed that TRAIL inhibited telomerase activity and the expression of telomerase catalytic subunit. CONCLUSION: TRAIL is an effective molecule to inhibit the growth of hepatocarcinoma through multiple pathways, such as inducing apoptosis and inhibiting the activity of telomerase. 相似文献
11.
ATM: To investigate whether hypoxia-inducible factor 1α (HIF-1α) mediates the effect of intermittent hypoxia on A549 cell viability, apoptosis and invasive ability METHODS: A549 cells were transfected with HIF-1α-siRNA and cultured under intermittent hypoxia. The expression of HIF-1α and its downstream genes, such as Bcl-2, Bax, P53, P21 and VEGF at mRNA and protein levels was determined by real-time PCR and Western blot. The viability of the A549 cells was measured by MTT assay. The apoptosis and cell cycle distribution of the A549 cells were examined by flow cytometry. The invasive ability of the A549 cells was detected by transwell test. RESULTS: The expression levels of HIF-1α, Bcl-2 and VEGF in non-HIF-1α-siRNA transfected A549 cells cultured in intermittent hypoxia environment[blank controlgroup(IH C),empty vector control group (IH E) and negative control group (IH N)] were higher than those in the A549 cells in normoxia group (RA), but the expression levels of Bax and P21 were lower than those in RA group (P<0.05). The siRNA-mediated HIF-1α gene silencing[intermittent hypoxia silenced group (IHS)] resulted in obvious down-regulation of HIF-1α, Bcl-2 and VEGF, and significant increase in the protein expression of P21 and Bax(P<0.05). The expression level of P53 in intermittent hypoxia groups was significantly higher than that in RA group, and no significant difference of P53 expression in different intermittent hypoxia groups was observed. Compared with normoxia, intermittent hypoxia resulted in significantly enhanced cell viability, decreased apoptosis, and enhanced invasive ability of non-HIF-1α-siRNA transfected A549 cells (P<0.05). The siRNA-mediated HIF-1α gene silencing resulted in significant cell viability inhibition, elevated apoptotic rate and decreased invasive ability under hypoxic condition (P<0.05).CONCLUSION: Intermittent hypoxia promotes the viability and invasion of A549 cells by HIF-1α-mediated downstream gene expression. HIF-1α gene silencing inhibits A549 cell growth and invasion under intermittent hypoxia by inhibition of HIF-1α signal pathways in vitro. 相似文献
12.
AIM: To provide the reference for optimizing the seed cells for tissue engineering,the relationship between apoptosis and differentiation in the process of induction on embryonic stem cells(ESCs) was investigated.METHODS: Day 2-3 embryoid bodys (EBs) were derived from ESCs,and then tissue growth factor-β1(TGF-β1) was added,and co-cultured with visceral endoderm like END-2 cell conditioned medium.The induced cells were evaluated by using immunofluoresence and transmission electron micrography.Cell apoptosis was analyzed using flow cytometry.RESULTS: The total percentage of beating EBs treated with TGF-β1 was 43%.All the beating cardiomyocytes derived from ESCs expressed cardiac-specific proteins for TnT,and were observed the cardiac-specific ultrastructure.Interestingly,the total percentage of beating EBs treated with TGF-β1 combined with END-2 cell conditioned medium was 88% (P＜0.01),and the beating areas were bigger.After 3 days of induction with different conditions,the apoptotic levels were (5.58%±0.65% and 9.60%±0.75%,P＜0.05),respectively.CONCLUSION: combination of TGF-β1 with co-cultured visceral endoderm like END-2 cell conditioned medium could get a higher induction efficiency on the differentiation of ESCs into the cardiomyocytes.The partly inducible effect mechanisms may induce EDCs differentiation toward a cardiac phenotype and enhance apoptosis in cells not committed to cardiac differentiation. 相似文献
13.
AIM: To investigate the differentiation of murine embryonic stem cells (ESCs) into hematopoietic stem cells (HSCs) by the supportive effects of human aorta-gonad-mesonephros (AGM) region and fetal liver (FL) stromal cells.METHODS: E14 ESCs were induced into embryoid body (EB) first. Then the cells from EB were further co-cultured with human AGM region and FL stromal cells in non-contact system. On day 6, the cells derived from EB were collected for Sca-1+c-Kit+ cell analysis by flow cytometry, colony forming unit (CFU) assay and teratoma formation checking. BALB/c female mice conditioned with lethal dose of γ-ray irradiation were transplanted with EB cells from different culture systems. The survival rates, engraftment of donor cells, reconstitution of hematopoietic were monitored.RESULTS: Sca-1+c-Kit+ cells in EB cells co-cultured with human AGM region and FL stromal cells had the value of (21.96±2.54)%, and the total CFU was as (520±52)/105 cells, which were statistically greater than those in EB cells only cultured with human AGM region stromal cells (P＜0.05). No teratoma was found in NOD-SCID mice after subcutaneous injection of EB cells co-cultured with human AGM region and FL stromal cells. In BALB/c female mice transplanted of EB cells co-cultured with human AGM region and FL stromal cells, the survival rate was 77.8%, and the peripheral blood cell count was obviously improved on day 14. PCR results showed the recipients all had sry gene copies from donor in bone marrow. The recipient mice transplanted with EB cells only cultured with human AGM region stromal cells all died within 15 days.CONCLUSION: Stromal cells from human AGM region and FL enhance the directed differentiation of ESCs into HSCs which can reconstruct hematopoiesis in vivo. 相似文献
14.
AIM: To observe the effects of insulin-like growth factorⅡ (IGF-Ⅱ) at different expression levels on hepatocellular carcinoma Huh7 cell proliferation and migration, and to explore the role of IGF-Ⅱ in the development of HCC. METHODS: The Huh7 cells were transfected with the over-expression plasmid pcDNA3.1(+)- IGF-Ⅱ or RNA interference plasmid pLVX-shRNA-IGF-Ⅱ by Lipofectamine 2000. The quantitative real-time PCR and Western blotting were used to verify the expression of IGF-II. The biological behaviors of the Huh7 cells were analyzed by CCK-8 assay, plate clone formation assay, cell scratch test and Transwell chamber experiment.RESULTS: Over-expression of IGF-II promoted the growth and migration of hepatocellular carcinoma cells (P < 0.05), and the cell proliferation was significantly inhibited in the Huh7 cells with low IGF-II expression (P < 0.05).CONCLUSION: IGF-II is involved in the regulation of biological behavior of hepatocellular carcinoma Huh7 cells in vitro, which may play a promoting role in the development of hepatocellular carcinoma. 相似文献
15.
FU Jin-rong LIU Wen-li ZHOU Yu-feng CHEN Hui-qin ZHOU Dun-hua HUANG Shao-liang 《园艺学报》2007,23(7):1373-1377
AIM: To explore the supportive and expansive effects of aorta-gonad-mesonephros(AGM) region derived stromal cells on hematopoietic stem cells (HSC) in vitro.METHODS: The murine stromal cells were separated and cultured from AGM region of a 11 day postcoitum (dpc) mouse embryo and 6 week mouse. After identification by Wrights staining and flow cytometry, the stromal cells were co-cultured with the embryonic stem cell(ESC)-derived, cytokine-induced HSCs, and the maintenance and expansion of HSCs were evaluated by detecting CD34+，CD34+Sca-1+cells with flow cytometry.Blast colony-forming cell (BL-CFC) and high proliferative potential colony-forming cells(HPP-CFC) were determined by semi-solid medium clonal culture.RESULTS: AGM-derived and bone marrow(BM)-derived stromal cells were similar in morphology and phenotype, and had common character of stromal cells. Supported by AGM stromal cells or by BM stromal cells, more primitive progenitor cells HPP-CFC were expanded, but BL-CFC expansion was only detected in AGM-derived stromal cells. In the supporting of BM stromal cells CD34+ hematopoietic stem/progenitor cells were expanded 3-4 times, but no significant expansion in CD34+Sca-1+ cells was observed. While in the supporting of AGM stromal cells, both CD34+ hematopoietic stem/progenitor cells and CD34+Sca-1+ cells were expanded significantly from 4 to 5 times, respectively (P<0.05).CONCLUSION: AGM-derived stromal cells significantly support the expansion of HSC, and also maintain the self-renewal activity and multi-lineage differentiation of HSC in vitro. 相似文献
16.
AIM： To investigate the role of damaged mitochondria in dendritic cell (DC) apoptosis induced by Vibrio vulnificus (Vv) and its possible mechanism. METHODS：DC2.4 cells were co-cultured with Vv 1.1758 strain. Fluorescent probes DCFH-DA and Fluo-8-AM were used to detect reactive oxygen species (ROS) and intracellular Ca2+ concentration in the invaded cells, respectively. The cellular apoptotic rates and mitochondrial membrane potential (Δψm) were measured by flow cytometry. The expression of nuclear factor-kappa B p65 (NF-κB p65) and tumor necrosis factor-alpha (TNF-α) was detected by Western blotting.
RESULTS： Vv 1.1758 induced DC2.4 cell apoptosis. Vv 1.1758 bacteria invaded into the DC2.4 cells by binding with cellular membrane though the end of the body. In the invaded DC2.4 cells, the visible mitochondrial damage, elevated ROS and intracellular Ca2+ levels, and declined Δψm were presented. After 1 h of co-culture, NF-κB p65 began to rise and reached the peak at 5 h, and then slightly decreased at 6 h. The TNF-α level increased after 2 h of co-culture and reached the peak at 6 h. CONCLUSION： The damaged mitochondria play an important role in DC apoptosis induced by Vv, and its possible mechanism may associate with the elevation of ROS and intracellular Ca2+ level, and the declined Δψm. Meanwhile, NF-κB p65 and TNF-α are potential critical signaling molecules in the process of apoptosis. 相似文献
17.
PENG Cheng-hong ZHAO Zhi-ming LIU Hong HAN Bao-san WANG Yong FANG He-qing WU Yu-lian GAO Chang-you 《园艺学报》2005,21(2):289-292
AIM: To evaluate the blood compatibility of a new bioartificial reactor membranous material (propylene-acidamide grafted polypropylene membrane, PP-g-AAm) in vitro. METHODS: Contacted PP-g-AAm membrane and PP (polypropylene) membrane with platelet-rich plasma in a swing bed, 37 ℃, to simulate the conditions in vivo, and another group of PRP without any membranes was set as control group. ELISA was used to study the expression of β-thromboglobulin, and flow cytometry was used to study CD62P and CD63 expression of the activated blood platelets after contacting the two kinds of membranes with PRP. Scanning electrical microscopy was used to study the configuration and numbers of platelet cells adhered on the membranes. RESULTS: After contacting with PRP 30 min, β-TG expression showed marked difference between the two kinds of material groups and the control group (P<0.01, P<0.05), and the difference between the two kinds of membrane groups was also significant (P<0.05). There were obviously differences on the expression of CD62P and CD63 between the two kinds of membranes after contacting with PRP for 30 min (P<0.05，P<0.01). When enlarged 10 000 times, the disfiguration of the platelet cells adhered on the two kinds of membranes after one hour were found by scanning electrical microscopy, and the numbers of platelets on the PP membrane were more than the PP-g-AAm membrane markedly. CONCLUSION: The PP-g-AAm membrane has better blood compatibility than the PP membrane. 相似文献
18.
MAO Yue-ping CHEN Ming-chun QI Qing GUO Qing HUANG Yong-jing ZENG Fan-qin 《园艺学报》2008,24(12):2455-2459
AIM: To investigate the change of collagen component and the expression of TGF-β1 in the co-culture of mdMSCs and human fibroblast of scleroderma (hsFb) in vitro,and to probe the therapy possibility of mdMSCs on scleroderma.METHODS: The cultivated mdMSCs were isolated by tissue digested-adherented-subcultured in low-serum medium.The changes of hydroxyproline (Hyp) and TGF-β1 in co-cultured mdMSCs and human normal fibroblast (hnFb) were determined at day 4 and 8 by samples basic hydrolysis and ELISA respectively.RESULTS: The Hyp production and TGF-β1 expression of hsFb were significant higher than that of hnFb on day 8 (P<0.05),no difference among the various hsFb/mdMSCs co-cultured Transwell system was found (P>0.05).The TGF-β1 production in hsFb/mdMSCs 2.5×104 co-culture Transwell system was significant higher than that in hsFb culture alone on day 4 (P<0.05),but there was on difference between them on day 8 (P>0.05).No correlation between the production of Hyp and TGF-β1 in co-cultured Transwell system (r＝0.221,P>0.05) was observed.However,the production of Hyp and TGF-β1 showed significant positive correlation under the condition that hnFb or hsFb was cultured alone (P<0.05).CONCLUSION: In vitro,mdMSCs couldn't effectively reduce the production of Hyp and TGF-β1 by hsFb in Transwell system.The mdMSCs may not effectively treat scleroderma by the effect on hsFb. 相似文献
19.
HU Min HAN Yao-wu LI Lu MA Ke-long Lü Lei ZHANG Dao-qin YANG Jian-ao WANG Si-ying 《园艺学报》2019,35(12):2187-2193
AIM: To establish a three-dimensional angiogenesis model in vitro for observing the influence of tumor cells on angiogenesis, and to explore its possible molecular mechanism. METHODS: Human umbilical vein endothelial cells (HUVECs) and mouse endothelial cells SVEC4-10EE2 were coated on the surface of beads, and then mixed with fibrinogen and seeded in cell culture plates containing thrombin. The cultured endothelial cells coated on the beads grew into a vessel-like structure in a three-dimensional space containing fibrin condensate to establish an in vitro three-dimensional angiogenesis model. Tumor cells MDA-MB-231 and E0771 were co-cultured in the model to observe the effect of tumor cells on angiogenesis in vitro,respectively. The concentrations of monocyte chemoattractant protein-1 (MCP-1) and vascular endothelial growth factor (VEGF) in the tumor cells culture supernatant were measured by ELISA. An antagonist of CCR2 (receptor of MCP-1), along with SU5416 (an inhibitor of VEGF receptor), were added to the tumor cell co-culture system. The supernatant of the tumor cells was collected as a conditioned medium, which was then added to the angiogenesis system of HUVECs or SVEC4-10EE2 cultured individually. The effect of conditioned medium on angiogenesis was observed under the conditions of with or without SU5416, as well as with or without CCR2 antagonist. RESULTS: Under normal condition, HUVECs and SVEC4-10EE2 formed a multi-cellular vascular structure the three-dimensional angiogenesis model in vitro. The co-culture of MDA-MB-231 (E0771) cells significantly promoted in the formation of membrane-like structures. The ELISA results showed that the levels of MCP-1 and VEGF in the supernatant of tumor cells were significantly elevated (P<0.05). MCP-1 promoted the formation of membrane tube in three-dimensional culture system in vitro. After adding the antagonists of MCP-1 receptor CCR2 and VEGF (SU5416), the angiogenesis was significantly inhibited (P<0.05). At the same time, conditioned medium promoted the formation of extracorporeal membranes in the endothelial cells. The promoting effect was blocked by CCR2 antagonists and SU5416 (P<0.05). CONCLUSION: The in vitro three-dimensional angiogenesis model is successfully established. Tumor cells significantly promote the formation of membrane tubes. The effect of tumor cells might be related to MCP-1 and VEGF secretion. 相似文献
20.
AIM:To study the protective effect of bone marrow stromal cells (MSCs) on ischemia /reperfusion hippocampal slices.METHODS:Ischemia/reperfusion models of hippocampal slices from newborn rats were established. MSCs obtained from adult bone marrow were cultured, isolated and purified. Cell death was assessed using propidium iodide fluorescence. And brain-derived neurotrophic factor (BDNF) expression in MSCs was determined by immunocytochemistry and Western blot.RESULTS:Maximal dead cells appeared in hippocampal slices 3 to 7 days after reperfusion. When the slices were co-cultured with MSCs, only a few cells were dead. The protective effect of MSCs on the slices was diminished significantly when anti-BDNF antibody was added to the medium. The protein of BDNF was faintly expressed in MSCs under normal conditions. When MSCs were co-cultured with ischemia /reperfusion hippocampal slices, the expression of BDNF in MSCs was increased gradually especially when co-cultured for 3 to 7 days. However, MSCs co-cultured with normal hippocampal slices expressed BDNF at a lower level at any times of co-culture.CONCLUSIONS:In an in vitro model of simulated ischemia, MSCs reduce cell death. Ischemia/reperfusion hippocampal slices co-cultured with MSCs promote the expression of BDNF in MSCs, which in turn protect the ischemic neurons. 相似文献