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OBJECTIVE@#To explore the potential mechanism of resistance to axitinib in clear cell renal cell carcinoma (ccRCC), with a view to expanding the understanding of axitinib resistance, facilitating the design of more specific treatment options, and improving the treatment effectiveness and survival prognosis of patients.@*METHODS@#By exploring the half maximum inhibitory concentration (IC50) of axitinib on ccRCC cell lines 786-O and Caki-1, cell lines resistant to axitinib were constructed by repeatedly stimulated with axitinib at this concentration for 30 cycles in vitro. Cell lines that were not treated by axitinib were sensitive cell lines. The phenotypic differences of cell proliferation and apoptosis levels between drug resistant and sensitive lines were tested. Genes that might be involved in the drug resistance process were screened from the differentially expressed genes that were co-upregulated in the two drug resistant lines by transcriptome sequencing. The expression level of the target gene in the drug resistant lines was verified by real-time quantitative polymerase chain reaction (RT-qPCR) and Western blot (WB). The expression differences of the target gene in ccRCC tumor tissues and adjacent tissues were analyzed in the Gene Expression Profiling Interactive Analysis (GEPIA) public database, and the impact of the target gene on the prognosis of ccRCC patients was analyzed in the Kaplan-Meier Plotter (K-M Plotter) database. After knocking down the target gene in the drug resistant lines using RNA interference by lentivirus vector, the phenotypic differences of the cell lines were tested again. WB was used to detect the levels of apoptosis-related proteins in the different treated cell lines to find molecular pathways that might lead to drug resistance.@*RESULTS@#Cell lines 786-O-R and Caki-1-R resistant to axitinib were successfully constructed in vitro, and their IC50 were significantly higher than those of the sensitive cell lines (10.99 μmol/L, P < 0.01; 11.96 μmol/L, P < 0.01, respectively). Cell counting kit-8 (CCK-8) assay, colony formation, and 5-ethynyl-2 '-deoxyuridine (EdU) assay showed that compared with the sensitive lines, the proliferative ability of the resistant lines decreased, but apoptosis staining showed a significant decrease in the level of cell apoptosis of the resistant lines (P < 0.01). Although resistant to axitinib, the resistant lines had no obvious new replicated cells in the environment of 20 μmol/L axitinib. Nuclear protein 1 (NUPR1) gene was screened by transcriptome sequencing, and its RNA (P < 0.0001) and protein expression levels significantly increased in the resistant lines. Database analysis showed that NUPR1 was significantly overexpressed in ccRCC tumor tissue (P < 0.05); the ccRCC patients with higher expression ofNUPR1had a worse survival prognosis (P < 0.001). Apoptosis staining results showed that knockdown ofNUPR1inhibited the anti-apoptotic ability of the resistant lines to axitinib (786-O, P < 0.01; Caki-1, P < 0.05). WB results showed that knocking downNUPR1decreased the protein level of B-cell lymphoma-2 (BCL2), increased the protein level of BCL2-associated X protein (BAX), decreased the protein level of pro-caspase3, and increased the level of cleaved-caspase3 in the resistant lines after being treated with axitinib.@*CONCLUSION@#ccRCC cell lines reduce apoptosis through theNUPR1 -BAX/ BCL2 -caspase3 pathway, which is involved in the process of resistance to axitinib.
Subject(s)
Humans , Carcinoma, Renal Cell/metabolism , Axitinib/pharmacology , Kidney Neoplasms/metabolism , bcl-2-Associated X Protein , Nuclear Proteins , Cell Line, Tumor , Apoptosis , Cell ProliferationABSTRACT
Objective To investigate whether Danhong injection can enhance the therapeutic effect of neural stem cell (NSC) transplantation in repairing cerebral ischemia injury by regulating the nuclear factor E2-related factor 2 (Nrf2) signaling pathway. Methods Forty male SD rats were randomly divided into the NSC transplantation group (NSC group), Danhong injection group (DH group), NSC+ Danhong injection group (N+D group), NSC+ Danhong injection group +ML385 group(N+D+M group) and PBS control group (PBS group), 8 rats in each group. All rat models of cerebral ischemia were established by embolization of the middle cerebral artery. Reperfusion was performed at 1.5 h after embolization. All rats in each group received corresponding interventions at 3 d after reperfusion. The neurological function score was evaluated before and 1, 2, 4 weeks after NSC transplantation. All rats were sacrificed at 4 weeks after NSC transplantation. The parameters related to oxidative stress were detected. The expression levels of neuron-specific nuclear protein (NeuN) and von Willebrand factor (vWF) were determined by immunofluorescence staining. Results Before NSC transplantation, the neurological function scores did not significantly differ among different groups (all P > 0.05). At postoperative 1, 2 and 4 weeks, the neurological function scores in the NSC, DH and N+D groups were significantly lower than those in the PBS and N+D+M groups (all P < 0.05). Compared with the PBS and N+D+M groups, the malondialdehyde (MDA) levels were significantly decreased, whereas the superoxide dismutase (SOD) and glutathione peroxidase (GPX) levels were considerably increased in the NSC, DH and N+D groups (all P < 0.05). The GPX level in the N+D+M group was significantly lower than that in the PBS group (P < 0.05). Immunofluorescence staining showed that the transplant NSC in the rat brain migrated to the surrounding area of cerebral infarction and survived, and expressed neuronal marker NeuN and neovascularization marker vWF. However, the number of living NSC in the N+D+M group was significantly lower compared with those in the remaining groups. Conclusions Danhong injection may improve the microenvironment of stem cell transplantation, enhance the survival rate of transplant NSC and improve the therapeutic effect of NSC transplantation for cerebral ischemia injury probably by regulating the Nrf2 signaling pathway.
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The protein proteolysis-targeting chimeras (PROTAC) are a kind of bifunctional compound that can recruit target proteins and degrade the enzyme of target proteins. The mechanism of PROTAC is using the ubiquitin-proteasome pathway to degrade target protein specifically. Because of its potential to target non-proprietary proteins and to play roles in drug resistance, PROTAC has attracted wide attention. This review summarizes the application of small molecule PROTAC in previous studies of different targets, such as nuclear proteins, membrane proteins and cytoplasmic proteins.
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Nuclear protein of the testis midline carcinoma (NMC) is a rare malignant tumor that is mostly located in the upper trachea,mediastinal midline,and paravertebral midline,and few literature has described the imaging features of NMC in the nasal cavity and paranasal sinuses. In this article we summarize the clinical,radiologic,and pathologic data of one case of pathologically confirmed NMC in the nasal cavity and paranasal sinus by focusing on its CT and magnetic resonance imaging features.
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Humans , Magnetic Resonance Imaging , Nasal Cavity , Pathology , Nose Neoplasms , Diagnostic Imaging , Nuclear Proteins , Paranasal Sinus Neoplasms , Diagnostic Imaging , Paranasal Sinuses , Pathology , Tomography, X-Ray ComputedABSTRACT
Objective To investigate the relationship between two single nucleotide polymorphisms (rs356219,rs356165 sites) and cognitive dysfunction in Parkinson disease.Methods 236 patients with Parkinson's disease were randomly selected from November 2014 to November 2017.According to the results of MoCA cognitive function evaluation,the patients were divided into group A (cognitive dysfunction group)and group B (normal cognition group).At the same time,65 patients were randomly selected as group C (Health control group).The allele frequency and genotype distribution of rs356219 and rs356165 were compared,and the differences among the three group were compared.Results In the rs356165 allele frequency,group A (G:57.14％,A:42.86％),group B (G:56.45％,A:43.55％) and group C (G:52.31％,A:47.69％) had no statistical significance (P＞ 0.05).In the rs356165 genotype,G/G (21.43％) and A/A (14.29％) in group A were higher than group C (G/G:4.62％,A/A:1.54％),G/G (22.58％) in group B and A/A (14.52％) were higher than group C G/G (4.62％) and A/A (1.54％) (P＜ 0.05).In the rs356219 allele frequency,group A (G:64.29％,A:64.29％) and group B (G:64.52％,A:35.486％) and group C (G:46.15％,A:53.85％) was statistically significant (P＜0.05),but no statistical significance between group A and group B (P＞0.05);In the rs356219 genotype,group A (G/G:35.71％,A/A:21.43％,A/G:42.86％),group B (G/G:35.48％,A/A:22.58％,A/G:41.94％) and group C (G/G:30.77％,A/A:26.15％,A/G:43.08％) had no statistical significance (P＞ 0.05),and there was no statistical significance between group A and group B (P＞0.05).Conclusions The polymorphism of rs356219 and rs356165 sites in rho-synaptic nucleoprotein plays an important role in the pathogenesis of Parkinson disease.However,there was no correlation with cognitive dysfunction in patients with Parkinson disease.
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Colon cancer is one of the common malignant tumors of digestive system.In our previous study, it has been demonstrated that colon cancer specific antigen thioredoxin like-2b (Txl-2b) can interact with Ras-related nuclear protein (Ran).However, there are few reports about the role and mechanism of Ran in tumorigenesis of colon cancer.Aims: To investigate the effect of Ran-targeting RNA interference on apoptosis and expressions of caspase-3 and poly(ADP-ribose) polymerase (PARP) in colon cancer cell lines.Methods: 20, 40 and 60 nmol/L siRNA-1 (si-1 group), siRNA-2 (si-2 group), siRNA-3 (si-3 group) targeting to Ran gene and normal control siRNA (NC group) were transfected into colon cancer cell line HCT116 and DLD-1, respectively.The interference efficiency of siRNAs and expressions of caspase-3 and PARP were detected by Western blotting.Cell apoptosis was determined by flow cytometry.Results: 20 nmol/L siRNA-1 and siRNA-2 had the best effect on inhibiting expression of Ran.Early apoptosis rates of HCT116 and DLD-1 cells in si-1 group were significantly higher than those in NC group (19.37%±7.57% vs.4.83%±1.72%;16.53%±3.38% vs.6.27%±3.13%;P all <0.05).Late apoptosis rates of HCT116 and DLD-1 cells in si-1 and si-2 groups were significantly higher than those in NC group (15.97%±3.31%, 16.33%±5.40% vs.6.40%±1.05%;22.93%±1.57%, 11.50%±0.70% vs.6.20%±0.98%;P all <0.05).Compared with NC group, expressions of cleaved caspase-3 (active form) and cleaved PARP (inactive form) were significantly increased in DLD-1 cells of si-1 and si-2 groups.Conclusions: Silencing Ran gene can significantly promote the apoptosis of colon cancer cells, and its mechanism is related to the regulation of caspase-3 and PARP expression.
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Objective To detect the expression of DNA repair enzyme O6-methylguanine DNA methyl-transferase(MGMT),cell proliferation-related nuclear protein(Ki-67)and P53 protein in gliomas,and inves-tigate the relationship and clinical significance among them and gloma grade. Methods 61 cases of brain glio-ma specimes and 16 cases of internal decompression of brain trauma were used to detect the expression of MGMT,Ki-67 and P53 by using immunohistochemical SP method. Results MGMT protein expression (19. 67%∶0,χ2 =3. 729,P=0. 062),Ki-67 protein expression(39. 34% ∶0,χ2 =5. 722,P=0. 016)and P53 protein expression(27. 87% ∶0,χ2 =9. 146,P=0. 002)showed significant differences in gliomas com-pared to normal brain tissues,the expression of Ki-67 was significantly higher in high-grade gliomas(Ⅲ-Ⅳ) than that in low-grade gliomas(Ⅰ-Ⅱ)(14 ∶10,χ2 =11. 718,P =0. 001). No significant difference was found between the MGMT and P53. Conclusion MGMT protein can be used as a biomarker in gliomas detec-tion;Ki-67 has a positive correlation with tumor grade,and can be used as a reference indicator of pathological grade;P53 protein expression may be used as a potential target for the treatment of gliomas.
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Objective To study the expression of connective tissue growth factor(CTGF) and PEST containing nuclear pro‐tein(PCNP) and their significance in pancreatic cancer .Methods The expressions of CTGF and PCNP proteins were tested by im‐munohistochemistry and Immunofluorescence in 39 cases of pancreatic carcinomas and adjacent paracancerous tissues .Results The positive rate of CTGF and PCNP in pancreatic carcinomas was significantly higher than adjacent paracancerous tissues(χ2 =60 .41 , 51 .46 ,all P<0 .01) .The differences of the expression of CTGF and PCNP in pancreatic carcinoma of tumor differentiation ,TNM stage and lymph node metastasis was significant (P< 0 .05) .Conclusion The expression of CTGF and PCNP in the pancreatic cancer was obviously increased ,and the level of CTGF ,PCNP had remarkable connection with the stages of tumor and the condition of lymph node metastasis .
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As a seasonal, pandemic and worldwide communicable disease, the threat of influenza against human health is increasingly rigorous with the accelerated mutation of influenza viruses together with the increased rate of recombinant between different types of viruses. Novel anti-influenza drugs based on new targets are always the first choice of defenses against influenza viruses for safeguarding humans health because of the relatively retarded R&D of virus specific vaccines. The research and development of modern anti-influenza drugs are effectively promoted by the continuously progressive understandings on the mechanisms of infection and multiplication of influenza viruses. The present paper briefly reviews the current advances in the research on major targets of anti-influenza drugs for the reference in further R&D of new drugs.
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Hypoxia inducible factor 1 (HIF-1), a nuclear protein with transcription activity, can make the body produce adaptive response to hypoxia/ischemia by binding to target gene, transcription and post-transcriptional control. Ischemic tolerance refers to the adaptive response to transient ischemia and reperfusion, which can improve tissue tolerance during the following damage caused by more severe ischemic events. The recent studies have found that the expression of HIF-1 has an important significance in ischemic tolerance. HIF-1 may be a key factor of the oxygen signal transduction pathway in the development of cerebral ischemic tolerance.