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Aim To establish subcutaneous and orthotopic transplantation models of human lung cancer in nude mice, and compare the anti-cancer effects of digoxin between the two models. Methods After subcutaneous inoculation of H460 tissues in nude mice, the tumor volume was measured; HE staining and immunohistochemistry were performed; H460-Luc cell suspension was injected into the lung of nude mice toestablish orthotopic tumor model, the in vivo imaging and fluorescence values were recorded, and the tumor lesions in other organs were observed after dissection. Results Compared with control group, the gemcitabine group had a significant anti-tumor effect (P 0.05). HE staining showed that the cell density in each treatment group decreased, and necrosis and/or fibrous hyperplasia were obvious. Immunohistochemistry indicated that the protein expression of p-p38, p-ERK and Nur77 in each treatment group significantly increased in the subcutaneous transplantation model; in the orthotopic transplantation model, the gemcitabine, the middle (P < 0.05) and low dose of digoxin group could inhibit the tumor growth, while the high dose of digoxin group accelerated the development of tumor (P < 0.05). Conclusion Digoxin is more sensitive to orthotopic transplanted tumor than subcutaneous transplanted tumor, anddigoxin may inhibit the tumor growth by up-regulating the expression of p-p38, pERK and Nur77.
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ObjectiveTo investigate the role of TGF-β1/Nur 77 in HBV-induced liver fibrosis.MethodsHE, Masson's and immunohistochemistry (IHC) staining were used to detect Nur 77 expression in liver tissue. NTCP-Huh7.5.1 was constructed by transfecting pCMV6-NTCP into Huh 7.5.1 cells to explore effects of HBV replication and TGF-β1 expression. LX-2 cells was incubated with TGF-β1 in the presence of Nur 77 gRNA. QPCR was used to observe the effect of TGF-β1/Nur 77 on LX-2 activation and liver fibrosis. Our study engaged a co-culture system in the presence of Nur 77 gRNA to observe the relationship between HBV replication andα-SMA, TIMP-1 and CoL1A1 expression.ResultsIHC showed a higher Nur 77 level in severe fibrosis liver (≥S2) than in mild fibrosis liver (≤S1). Western blot showed that pCMV6-NTCP could express in Huh 7.5.1 cells. At 3 days after infection, HBV DNA level in supernatant of NTCP-Huh 7.5.1 cells was 5.39±0.96×103 copies/ml (P<0.01), HBV infection enhanced TGF-β1 mRNA expression in NTCP-Huh 7.5.1 cells (1.94 vs 1.00，P<0.01). At 72 h after TGF-β1 incubation, mRNA expression of Nur 77, α-SMA, TIMP-1 and CoL1A1 expression was up-regulated significantly compared to control group (P<0.05), which can be inhibited by Nur 77 gRNA (P<0.05). Co-cultured with HBV infected NTCP-Huh 7.5.1 cells for 72 h, mRNA expression of Nur 77, α-SMA, TIMP-1 and CoL1A1 expression in LX-2 cells was up-regulated significantly compared to control group (P<0.05), which can be inhibited by Nur 77 gRNA (P <0.05).ConclusionHBV contributes to liver fibrosis through up-regulating TGF-β1/Nur 77 signaling pathway. Our study provides a new target for the treatment of HBV induced liver fibrosis.
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Objective: To investigate the effect of memantine on Parkinson's disease cell models. Methods: Parkinson's disease cell models were established using PC12 cells incubated with 6-hydroxydopamine (6-OHDA). Flow cytometry and microscopy were used to investigate the apoptotic process and the percentage of different apoptotic stages. PC12 cells were infected with lentiviral vectors to knockdown Nur77. Western blotting was used to detect the expression of Nur77 and caspase -3, -8, -9, -12 in PC12 cells withdifferent concentrations of 6-OHDA or 6-OHDA+memantine. Results: 6-OHDA led to apoptosis PC12 cells, and increased the expression of Nur77 and caspases. Memantine significantly inhibited 6-OHDA-induced apoptosis of PC12 cells. Meanwhile, memantine could mitigate apoptosis of PC12 cells by regulating the Nur77 and caspase pathway. Conclusions: Memantine has a protective effect on the PC12 cell model via regulating the Nur77 and caspases pathway.
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Objective: To investigate the effect of memantine on Parkinson's disease cell models. Methods: Parkinson's disease cell models were established using PC12 cells incubated with 6-hydroxydopamine (6-OHDA). Flow cytometry and microscopy were used to investigate the apoptotic process and the percentage of different apoptotic stages. PC12 cells were infected with lentiviral vectors to knockdown Nur77. Western blotting was used to detect the expression of Nur77 and caspase -3, -8, -9, -12 in PC12 cells withdifferent concentrations of 6-OHDA or 6-OHDA+memantine. Results: 6-OHDA led to apoptosis PC12 cells, and increased the expression of Nur77 and caspases. Memantine significantly inhibited 6-OHDA-induced apoptosis of PC12 cells. Meanwhile, memantine could mitigate apoptosis of PC12 cells by regulating the Nur77 and caspase pathway. Conclusions: Memantine has a protective effect on the PC12 cell model via regulating the Nur77 and caspases pathway.
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Objective: To investigate the clinical significance of Nur77 expression in prostate cancer tissues, and to explore the mechanism of its effect on the metastasis of prostate cancer. Methods: The expression of Nur77 in 72 cases of prostate cancer and 24 cases of benign prostatic hyperplasia was detected by immunohistochemistry. The relationship between Nur77 expression and the clinicopathological features of prostate cancer was analyzed. The co-localization of Nur77 and P-tubulin in prostate cancer PC-3 cells was detected by immunofluorescence. The interaction between Nur77 and β-tubulin was verified by co-immunoprecipitation assay. PC-3 cells were transfected with the different plasmids including pcDNA3.1-Nur77, Nur77ΔDBD, Nur77ΔTUBD and Nur77-shRNA by liposome, respectively. The soluble and polymerized tubulin fractions were differentially extracted and detected by Western blotting. The migration ability of PC-3 cells was detected by Transwell chamber assay. Results: The expression of Nur77 protein in prostate cancer tissues was higher than that in the benign prostatic hyperplasia tissues (P < 0.01). The expression level of Nur77 was downregulated with the ascending of N staging, M staging and Gleason score in prostate cancer (all P < 0.05). The Nur77 and β-tubulin were co-located and interacted with each other in the cytoplasm of PC-3 cells. Compared with the blank control group, the ratio of polymerized/soluble (P/S) tubulin was significantly increased in pcDNA3.1-Nur77 and Nur77ΔDBD groups, while the count of migratory cells was significantly decreased in pcDNA3.1-Nur77 and Nur77 ΔDBD group (all P < 0.05). In contrast, P/S ratio was significantly decreased, and the count of migratory cells was significantly increased in Nur77-shRNA group (both P < 0.05). Conclusion: Nur77 can inhibit the migration of PC-3 cells, which may be related to promoting tubulin polymerization into microtubule.
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This study examined the relationship between PDGF-induced proliferation of vascular smooth muscle cells (VSMCs) and Nur77 expression and the effect of atorvastatin on VSMC proliferation and Nur77 in PDGF-treated VSMCs.Rat VSMCs were isolated and cultured.After incubation with atorvastatin or Nur77 siRNA,the cells were stimulated with PDGF and detected for BrdU incorporation to measure the proliferation of the VSMCs.Quantitative PCR and Western blotting were used to determine the Nur77 protein and the CREB phosphorylation level,to observe their relations with PDGF-induced VSMC proliferation.Our results showed that PDGF increased the BrdU incorporation in VSMCs,suggesting that it induced the proliferation of the cells.The VSMC proliferation was associated with increased Nur77 expression and elevated CREB phosphorylation.Atorvastatin inhibited the PDGF-induced VSMC proliferation,suppressed Nur77 expression.After silencing of Nur77 gene,the PDGF-induced VSMC proliferation was decreased.It was concluded that PDGF-induced VSMC proliferation was related to the Nur77 expression and CREB phosphorylation.Atorvastatin reduced the Nur77 expression and,at the same time,inhibited the VSMC proliferation.