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Lysophosphatidic acid receptor 5 (LPAR5) is involved in mediating thyroid cancer progression, but the underlying mechanism needs to be further revealed. In this study, we confirmed that LPAR5 is upregulated in papillary thyroid carcinoma (PTC), especially in BRAF-like PTC, by analyzing The Cancer Genome Atlas (TCGA) database and performing immunohistochemistry assay in human thyroid cancer tissues. LPAR5-specific antagonist TC LPA5 4 treatment inhibited CGTH-W3, TPC-1, B-CPAP, and BHT-101 cell proliferation, CGTH-W3 and TPC-1 cell migration significantly. In vivo, TC LPA5 4 treatment could delay CGTH-W3 xenograft growth in nude mice. We also found that LPAR5-specific antagonist TC LPA5 4, PI3K inhibitor wortmannin, or mTOR inhibitor rapamycin pretreatment abrogated phosphorylation of Akt and p70S6K1 stimulated by LPA in CGTH-W3 and TPC-1 cells. Stimulating CGTH-W3 cells transfected with pEGFPC1-Grp1-PH fusion protein with LPA resulted in the generation of phosphatidylinositol (3,4,5)-triphosphate, which indicates that PI3K was activated by LPA directly. The p110ß-siRNA instead of p110α-siRNA transfection abrogated the increase of levels of phosphorylated Akt and S6K1 stimulated by LPA. Furthermore, immunoprecipitation assay confirmed an interaction between LPAR5 and p110ß. Overall, we provide new insights that the downregulation of LPAR5 decreased the proliferation and migration phenotype via the PI3K/Akt pathway. Inhibition of LPAR5 or the PI3K/Akt signal may be a novel therapeutic strategy for treating thyroid cancer.
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Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Classe Ia de Fosfatidilinositol 3-Quinase/metabolismo , Receptores de Ácidos Lisofosfatídicos/metabolismo , Neoplasias da Glândula Tireoide/metabolismo , Neoplasias da Glândula Tireoide/patologia , Animais , Domínio Catalítico/fisiologia , Linhagem Celular Tumoral , Regulação para Baixo/fisiologia , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Quinases S6 Ribossômicas 70-kDa/metabolismo , Transdução de Sinais/fisiologia , Câncer Papilífero da Tireoide/metabolismo , Câncer Papilífero da Tireoide/patologiaRESUMO
Everolimus inhibits mammalian target of rapamycin complex 1 (mTORC1) and is known to cause induction of autophagy and G1 cell cycle arrest. However, it remains unknown whether everolimus-induced autophagy plays a critical role in its regulation of the cell cycle. We, for the first time, suggested that everolimus could stimulate autophagy-mediated cyclin D1 degradation in breast cancer cells. Everolimus-induced cyclin D1 degradation through the autophagy pathway was investigated in MCF-10DCIS.COM and MCF-7 cell lines upon autophagy inhibitor treatment using Western blot assay. Everolimus-stimulated autophagy and decrease in cyclin D1 were also tested in explant human breast tissue. Inhibiting mTORC1 with everolimus rapidly increased cyclin D1 degradation, whereas 3-methyladenine, chloroquine, and bafilomycin A1, the classic autophagy inhibitors, could attenuate everolimus-induced cyclin D1 degradation. Similarly, knockdown of autophagy-related 7 (Atg-7) also repressed everolimus-triggered cyclin D1 degradation. In addition, everolimus-induced autophagy occurred earlier than everolimus-induced G1 arrest, and blockade of autophagy attenuated everolimus-induced G1 arrest. We also found that everolimus stimulated autophagy and decreased cyclin D1 levels in explant human breast tissue. These data support the conclusion that the autophagy induced by everolimus in human mammary epithelial cells appears to cause cyclin D1 degradation resulting in G1 cell cycle arrest. Our findings contribute to our knowledge of the interplay between autophagy and cell cycle regulation mediated by mTORC1 signaling and cyclin D1 regulation.
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Antineoplásicos/farmacologia , Autofagia/efeitos dos fármacos , Neoplasias da Mama/tratamento farmacológico , Proliferação de Células/efeitos dos fármacos , Ciclina D1/metabolismo , Everolimo/farmacologia , Pontos de Checagem da Fase G1 do Ciclo Celular/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Proteína 7 Relacionada à Autofagia/genética , Proteína 7 Relacionada à Autofagia/metabolismo , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Ciclina D1/genética , Feminino , Humanos , Células MCF-7 , Alvo Mecanístico do Complexo 1 de Rapamicina/antagonistas & inibidores , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Proteólise , Transdução de Sinais , Técnicas de Cultura de TecidosRESUMO
Both Von Hippel-Lindau tumor suppressor (VHL) and Never-in-mitosis A (NIMA)-related kinase 1 (NEK1) are involved in primary cilium formation, but whether VHL could regulate NEK1 is unknown. Here, we demonstrated that renal cancer cells Caki-1 and ACHN with wild-type VHL expressed lower level of NEK1 than that of VHL-defective cells including 786-O, 769-P and A498â¯cells. VHL-overexpression down-regulated NEK1 in 769-P cells, while VHL-knockdown up-regulated NEK1 in Caki-1â¯cells. In addition, we found the hypoxia response element (HRE) in the promoter sequence of NEK1 and hypoxia induced NEK1 expression both in vitro and in vivo. HIF-2α knockdown blocked hypoxia induced NEK1 upregulation instead of HIF-1α, which indicates that NEK1 may be a new target of HIF-2α. Moreover, we confirmed the association between VHL and NEK1 in Caki-1â¯cell, then showed VHL promoted NEK1 protein degradation and ubiquitination. In conclusion, our findings showed VHL regulates NEK1 via both HIF-2α pathway and ubiquitin-proteasome pathway in renal cancer cells.
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Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Neoplasias Renais/metabolismo , Quinase 1 Relacionada a NIMA/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Transdução de Sinais , Ubiquitina/metabolismo , Proteína Supressora de Tumor Von Hippel-Lindau/metabolismo , Animais , Linhagem Celular Tumoral , Humanos , Neoplasias Renais/patologia , Ratos Sprague-DawleyRESUMO
NEK8 (never in mitosis gene A (NIMA)-related kinase 8) is involved in cytoskeleton, cilia, and DNA damage response/repair. Abnormal expression and/or dysfunction of NEK8 are related to cancer development and progression. However, the mechanisms that regulate NEK8 are not well declared. We demonstrated here that pVHL may be involved in regulating NEK8. We found that CAK-I cells with wild-type vhl expressed a lower level of NEK8 than the cells loss of vhl, such as 786-O, 769-P, and A-498 cells. Moreover, pVHL overexpression down-regulated the NEK8 protein in 786-O cells, whereas pVHL knockdown up-regulated NEK8 in CAK-I cells. In addition, we found that the positive hypoxia response elements (HREs) are located in the promoter of the nek8 sequence and hypoxia could induce nek8 expression in different cell types. Consistent with this, down-regulation of hypoxia-inducible factors α (HIF-1α or HIF-2α) by isoform-specific siRNA reduced the ability of hypoxia inducing nek8 expression. In vivo, NEK8 and HIF-1α expression were increased in kidneys of rats subjected to an experimental hypoxia model of ischemia and reperfusion. Furthermore, NEK8 siRNA transfection significantly blocked pVHL-knockdown-induced cilia disassembling, through impairing the pVHL-knockdown-up-regulated NEK8 expression. These results support that nek8 may be a novel hypoxia-inducible gene. In conclusion, our findings show that nek8 may be a new HIF target gene and pVHL can down-regulate NEK8 via HIFs to maintain the primary cilia structure in human renal cancer cells.
Assuntos
Regulação Enzimológica da Expressão Gênica , Proteínas Quinases/fisiologia , Proteínas Serina-Treonina Quinases/fisiologia , Proteína Supressora de Tumor Von Hippel-Lindau/metabolismo , Animais , Hipóxia Celular , Linhagem Celular Tumoral , Cílios/metabolismo , Perfilação da Expressão Gênica , Humanos , Rim/enzimologia , Neoplasias Renais/metabolismo , Masculino , Quinases Relacionadas a NIMA , RNA Interferente Pequeno/metabolismo , Ratos , Ratos Sprague-Dawley , Traumatismo por Reperfusão , TransfecçãoRESUMO
Blockade of mammalian target of rapamycin (mTOR) is a promising area in breast cancer therapy. However, in clinical trials, objective response rate with mTOR inhibitor monotherapy in breast cancer was modest. Biomarker studies designed to identify the responders of rapalogs are of increasing interest. We validated p27KIP1 expression levels as a candidate predictive biomarker of response to rapalogs. We also analyzed the correlation between rapamycin activity and p27KIP1 expression in the primary breast cancer cells and the patient-derived breast tumor xenograft models. The cells isolated from the breast tumor tissues expressing high levels of p27KIP1 were sensitive to rapamycin, whereas the cells from the tissues expressing low levels of p27KIP1 exhibited resistance to rapamycin. The correlation between p27KIP1 expression and rapamycin antitumor activity was also observed in the patient-derived breast tumor xenograft models. Moreover, we also found rapamycin significantly decreased phosphorylated p70S6K1 and phosphorylated 4EBP1 in both samples. It seemed that the different sensitivity of tumor cells to rapamycin did not owe to its different potency against mTOR activity. We further propose p27KIP1 expression level may be also a candidate predictive biomarker of rapalogs for breast cancer therapy, which requires additional clinical validation.
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Biomarcadores Tumorais/biossíntese , Neoplasias da Mama/metabolismo , Inibidor de Quinase Dependente de Ciclina p27/biossíntese , Adulto , Idoso , Animais , Antineoplásicos/farmacologia , Biomarcadores Tumorais/genética , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Inibidor de Quinase Dependente de Ciclina p27/genética , Feminino , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Fosforilação , Sirolimo/farmacologia , Serina-Treonina Quinases TOR/biossíntese , Serina-Treonina Quinases TOR/genética , Ensaios Antitumorais Modelo de Xenoenxerto/métodosRESUMO
The aim of this study was to investigate and compare the microbial community structures of eubacteria and archaea in the pit mud of Chinese Luzhou-flavor liquor from the wall (C(w)) and bottom (C(b)) of cellar through nested PCR-denaturing gradient gel electrophoresis (DGGE). The Shannon-Wiener index (H) calculated from the DGGE profiles showed that the community diversities of eubacteria and archaea in samples from C(b) were almost higher than that from C(w). In addition, cluster analysis of the DGGE profiles revealed that some differences were found in the microbial community structure in samples from different locations. The closely relative microorganisms of all eubacterial 16S rRNA gene sequences fell into four phyla (Firmicutes, Proteobacteria, Bacteroidetes and Actinobacteria), including 12 genera and 2 uncultured eubacteria. Moreover, 37.1% eubacteria were affiliated with Clostridium. Particularly, genus Acinetobacter was absent in all samples from C(b) but present in all samples from C(w). The closely relative microorganisms of all archaeal 16S rRNA gene sequences fell into four genera, which included Methanobrevibacter, Methanoculleus, Methanobacterium and Methanosaeta, while the dominant archaea in samples from C(w) and C(b) were similar. Results presented in this study provide further understanding of the spatial differences in microbial community structure in the pit mud, and is of great importance for the production and quality improvement of Luzhou-flavor liquor.
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Bebidas Alcoólicas/microbiologia , Archaea/classificação , Archaea/isolamento & purificação , Bactérias/classificação , Bactérias/isolamento & purificação , Biota , Archaea/genética , Bactérias/genética , China , Análise por Conglomerados , DNA Arqueal/química , DNA Arqueal/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Eletroforese em Gel de Gradiente Desnaturante , Genes de RNAr , Dados de Sequência Molecular , Filogenia , Reação em Cadeia da Polimerase , RNA Arqueal/genética , RNA Ribossômico 16S/genética , Análise de Sequência de DNA , Homologia de Sequência do Ácido NucleicoRESUMO
Heparanase (HPSE), an endo-ß-D-glucuronidase, cleaves heparan sulfate and serves an important role in the tumor microenvironment and thus in tumorigenesis. HPSE is known to promote tumor cell evasion of apoptosis. However, the underlying mechanism of this requires further study. In the present study, the results demonstrated that myeloid cell leukemia-1 (MCL-1), an antiapoptotic protein, and HPSE were upregulated in prostate cancer tissues compared with adjacent normal tissues. In addition, the HPSE inhibitor, OGT 2115, inhibited PC-3 and DU-145 prostate cancer cell viability in a dose-dependent manner, with IC50 values of 20.2 and 97.2 µM, respectively. Furthermore, annexin V/PI double-staining assays demonstrated that OGT 2115 induced apoptosis in prostate cancer cells. OGT 2115 treatment markedly decreased MCL-1 protein expression levels, whereas RNA interference-mediated downregulation of MCL-1 and OGT 2115 drug treatment synergistically induced apoptosis in PC-3 and DU-145 cells. In vivo, OGT 2115 40 mg/kg (ig) significantly inhibited PC-3 cell xenograft growth in nude mice and increased the positive TUNEL staining rate of xenograft tissues. It was therefore hypothesized that MCL-1 was an important signaling molecule in OGT 2115-induced apoptosis. The results of the present study also demonstrated that the proteasome inhibitor, MG-132, markedly inhibited the downregulation of MCL-1 protein expression levels induced by OGT 2115. However, the protein synthesis inhibitor, cycloheximide, did not affect the role of OGT 2115 in regulating MCL-1. In summary, the results of the present study demonstrated that the proapoptotic activity of OGT 2115 was achieved by downregulating MCL-1.
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CXCL12/CXCR4 plays an important role in metastasis of gastric carcinoma. Rapamycin has been reported to inhibit migration of gastric cancer cells. However, the role of mTOR pathway in CXCL12/CXCR4-mediated cell migration and the potential of drugs targeting PI3K/mTOR pathway remains unelucidated. We found that CXCL12 activated PI3K/Akt/mTOR pathway in MKN-45 cells. Stimulating CHO-K1 cells expressing pEGFP-C1-Grp1-PH fusion protein with CXCL12 resulted in generation of phosphatidylinositol (3,4,5)-triphosphate, which provided direct evidence of activating PI3K by CXCL12. Down-regulation of p110ß by siRNA but not p110α blocked phosphorylation of Akt and S6K1 induced by CXCL12. Consistently, p110ß-specific inhibitor blocked the CXCL12-activated PI3K/Akt/mTOR pathway. Moreover, CXCR4 immunoprecipitated by anti-p110ß antibody increased after CXCL12 stimulation and G(i) protein inhibitor pertussis toxin abrogated CXCL12-induced activation of PI3K. Further studies demonstrated that inhibitors targeting the PI3K/mTOR pathway significantly blocked the chemotactic responses of MKN-45 cells triggered by CXCL12, which might be attributed primarily to inhibition of mTORC1 and related to prevention of F-actin reorganization as well as down-regulation of active RhoA, Rac1, and Cdc42. Furthermore, rapamycin inhibited the secretion of CXCL12 and the expression of CXCR4, which might form a positive feedback loop to further abolish upstream signaling leading to cell migration. Finally, we found cells expressing high levels of cxcl12 were sensitive to rapamycin in its activity inhibiting migration as well as proliferation. In summary, we found that the mTOR pathway played an important role in CXCL12/CXCR4-mediated cell migration and proposed that drugs targeting the mTOR pathway may be used for the therapy of metastatic gastric cancer expressing high levels of cxcl12.
Assuntos
Movimento Celular/efeitos dos fármacos , Quimiocina CXCL12/fisiologia , Receptores CXCR4/metabolismo , Sirolimo/farmacologia , Serina-Treonina Quinases TOR/antagonistas & inibidores , Actinas/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Quimiocina CXCL12/genética , Quimiocina CXCL12/metabolismo , Classe Ia de Fosfatidilinositol 3-Quinase/metabolismo , Regulação para Baixo , Retroalimentação Fisiológica , Furanos/farmacologia , Expressão Gênica , Humanos , Fosfatidilinositol 3-Quinases/metabolismo , Fosforilação , Ligação Proteica , Proteínas Proto-Oncogênicas c-akt/metabolismo , Piridinas/farmacologia , Pirimidinas/farmacologia , Receptores CXCR4/genética , Proteínas Quinases S6 Ribossômicas 70-kDa/metabolismo , Transdução de Sinais , Neoplasias Gástricas , Serina-Treonina Quinases TOR/metabolismoRESUMO
Gut microbiota affects the gut-brain axis; hence, the modulation of the microbiota has been proposed as a potential therapeutic strategy for cerebral ischemia/reperfusion injury (CIRI). However, the role and mechanism of the gut microbiota in regulating microglial polarization during CIRI remain poorly understood. Herein, using a middle cerebral artery occlusion and reperfusion (MCAO/R) rat model, we evaluated changes in the gut microbiota after CIRI and the potential effects of fecal microbiota transplant (FMT) on the brain. Rats underwent either MCAO/R or sham surgery, and then they received FMT (started 3 days later; continued for 10 days). 2,3,5-Triphenyltetrazolium chloride staining, neurological outcome scale, and Fluoro-Jade C staining showed that MCAO/R induced cerebral infarction, neurological deficits, and neuronal degeneration. In addition, immunohistochemistry or real-time PCR assay showed increased expression levels of M1-macrophage markers-TNF-α, IL-1ß, IL-6, and iNOS-in the rats following MCAO/R. Our finding suggests that microglial M1 polarization is involved in CIRI. 16 S ribosomal RNA gene sequencing data revealed an imbalance in the gut microbiota of MCAO/R animals. In contrast, FMT reversed this MCAO/R-induced imbalance in the gut microbiota and ameliorated nerve injury. In addition, FMT prevented the upregulation in the ERK and NF-κB pathways, which reversed the M2-to-M1 microglial shift 10 days after MCAO/R injury in rats. Our primary data showed that the modulation of the gut microbiota can attenuate CIRI in rats by inhibiting microglial M1 polarization through the ERK and NF-κB pathways. However, an understanding of the underlying mechanism requires further study.
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BACKGROUND: Total hip arthroplasty or hemiarthroplasty are used to treat displaced femoral neck fractures. However, the optimal treatment of these fractures remained controversial. OBJECTIVE: To assess the effects that compare total hip arthroplasty with hemiarthroplasty for the treatment of femoral neck fractures in the elderly. METHODS: We searched MEDLINE (January 1980 to 2010), EMBASE (January 1980 to 2010), and the Cochrane Library 2010; issue 1. Only prospective randomized controlled trials (RCTs) that compare total hip arthroplasty with hemiarthroplasty for the treatment of femoral neck fracture in the elderly were included. The analysis was performed with software RevMan5.0 from the Cochrane Collaboration. RESULTS: We identified seven relevant randomized controlled trials with a total of 828 participants. The meta-analysis showed relative risk of re-operation was 0.40 (95% CI = 0.24-0.67, P = 0.0004), the dislocation was 2.02 (95% CI = 1.26-3.25, P = 0.002), the mobility as functional outcome was 1.70 (95% CI = 1.21-2.38, P = 0.002). It was reported that the average operating room times and blood loss volumes in total hip arthroplasty were more than in hemiarthroplasty (P < 0.001). Other results were not significantly different. CONCLUSIONS: Total hip arthroplasty is associated with better functional outcome and lower reoperation rate than hemiarthroplasty in treatment of displaced femoral neck fractures in the elderly patients.
Assuntos
Artroplastia de Quadril/métodos , Fraturas do Colo Femoral/cirurgia , Idoso , Idoso de 80 Anos ou mais , Humanos , Recuperação de Função Fisiológica , Reoperação , Resultado do TratamentoRESUMO
BACKGROUND: Killer cell immunoglobulin-like receptor 2DL4 (KIR2DL4) is a transmembrane glycoprotein that is expressed by natural killer (NK) cells and certain subsets of T cells. However, its expression profiles and functions in solid tumor progression remain poorly defined. METHODS: In the present study, using bioinformatics analysis, immunohistochemistry, immunoblotting, MTT cell viability assay, soft agar colony formation assay and a human renal cell carcinoma (RCC) cell xenograft model in nude mice, we examined whether KIR2DL4 is expressed by RCC and its possible roles in RCC progression. RESULTS: We confirmed that KIR2DL4 is overexpressed by RCC cells. MTT and soft agar cloning assays showed that KIR2DL4 knockdown delayed cell proliferation and viability in RCC cell lines, Caki-1 and 769-P, in vitro. By contrast, KIR2DL4 overexpression promoted Caki-1 cell proliferation both in vitro and in vivo, which was observed in a BALB/c-nu/nu xenograft mouse model. Moreover, RNA sequencing data demonstrated that the differentially expressed genes found between parallel-controlled and Caki-1 cells overexpressing KIR2DL4 were highly associated with cancer development, of which those related to the phosphatidylinositol-3-kinase (PI3K)/protein kinase B (AKT) signaling pathway were particularly enriched, immunoblotting data showed that the level of AKT phosphorylation was higher or lower in KIR2DL4 overexpressing or KIR2DL4 knocking-down Caki-1 cells compared with that in the parallel-controlled cells. In addition, PI3K inhibitor wortmannin treatment and KIR2DL4-shRNA transfection further deregulated the levels of phosphorylated AKT and Caki-1 cell proliferation. CONCLUSIONS: Our results indicate that KIR2DL4 is also expressed by RCC cells, which promotes RCC progression associated with PI3K/AKT activation.
Assuntos
Carcinoma de Células Renais/metabolismo , Proliferação de Células/fisiologia , Neoplasias Renais/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores KIR2DL4/biossíntese , Animais , Biomarcadores Tumorais/metabolismo , Carcinoma de Células Renais/patologia , Linhagem Celular Tumoral , Feminino , Células HEK293 , Humanos , Neoplasias Renais/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Ensaios Antitumorais Modelo de Xenoenxerto/métodosRESUMO
Background: The prognosis of breast cancer varies according to the molecular subtype. Transmembrane 4 L six family 1 (TM4SF1) exhibits different expression patterns among the molecular subtypes of breast cancer. However, the expression profile of TM4SF1 in hormone receptor HR+HER2- breast cancer remains unclear. Methods: TM4SF1 mRNA levels were examined in major subclasses of breast cancer by analyzing The Cancer Genome Atlas (TCGA) datasets. In addition, TM4SF1 protein and mRNA levels in HR+HER2- breast cancer tissue samples were determined by immunohistochemistry and Western blot assay. The effect of TM4SF1 on cell proliferation was evaluated using MTT, colony formation, 3D organoid, and xenograft models, following the TM4SF1 overexpression or knockdown. Results: TCGA database analysis demonstrated that TM4SF1 was downregulated in breast cancer compared with the healthy adjacent breast tissue. In addition, the expression of TM4SF1 in basal-like one and the mesenchymal TNBC tissue was higher than that of the healthy adjacent breast tissue. Other types, including the luminal androgen receptor-positive TNBC tissue, expressed lower levels of TM4SF1. Immunohistochemistry and real-time quantitative PCR assays demonstrated that the TM4SF1 protein and mRNA levels were downregulated in the HR+HER2- breast cancer tissue compared with the healthy adjacent tissue. Moreover, the TM4SF1 overexpression reduced the viability of MCF-7 and ZR-75-1 breast cancer cells, whilst reducing the number of colonies and 3D-organoids formed by these cell lines. By contrast, TM4SF1 knockdown led to an increased MCF-7 cell proliferation. However, in the TNBC cell line, MDA-MB-231, TM4SF1 silencing reduced cell proliferation. In vivo, the TM4SF1 overexpression inhibited MCF-7 xenograft growth in a nude mouse model, which was associated with the downregulation of the Ki-67 expression, apoptosis induction, and inhibition of the mTOR pathway. Conclusion: TM4SF1 is downregulated in HR + HER2-breast cancer, and the overexpression of TM4SF1 suppresses cell proliferation in this cancer subtype.
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The papillary thyroid carcinoma (PTC) metastasizes through lymphatic spread, but the follicular thyroid cancer (FTC) metastasis occurs by following hematogenous spread. To date, the molecular mechanism underlying different metastatic routes between PTC and FTC is still unclear. Here, we showed that specifically androgen-regulated gene (SARG) was significantly up-regulated in PTC, while obviously down-regulated in FTC through analyzing the Gene Expression Omnibus (GEO) database. Immunohistochemistry assay verified that the PTC lymph node metastasis was associated with higher levels of SARG protein in clinical PTC patient samples. SARG-knockdown decreased TPC-1 and CGTH-W3 cells viability and migration significantly. On the contrary, SARG-overexpressed PTC cells possessed more aggressive migratory ability and viability. In vivo, SARG overexpression dramatically promoted popliteal lymph node metastasis of xenografts from TPC-1 cells mouse footpad transplanting. Mechanistically, SARG overexpression and knockdown significantly increased and decreased the expression of vascular endothelial growth factor C (VEGF-C) and VEGF receptor 3 (VEGFR-3), respectively, thereby facilitating or inhibiting the tube formation in HUVECs. The tube formation experiment showed that SARG overexpression and knockdown promoted or inhibited the number of tube formations in HUVEC cells, respectively. Taken together, we showed for the first time the differential expression profile of SARG between PTC and FTC, and SARG promotes PTC lymphatic metastasis via VEGF-C/VEGFR-3 signal. It indicates that SARG may represent a target for clinical intervention in lymphatic metastasis of PTC.
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OBJECTIVE: To investigate whether 25-hydroxyvitamin D [25(OH)D] can mediate effects without being converted to 1α,25-dihydroxyvitamin D [1,25(OH)2D]. METHODS: Vitamin D3 (VD3) was injected intramuscularly to 25-hydroxyvitamin D-1α-hydroxylase [1α(OH)ase] gene knockout (KO) male mice with a dose of 10,000 IU per week for 4 weeks. Skeleton Parameters and Serum biochemistry in mice were assayed. RESULTS: Serum 25(OH)D3 levels increased from 41 to 212 ng/mL in KO mice injected with VD3. Our results show that VD3 injections significantly increased the body weight of KO mice and there were no significant differences in body weight at 7 weeks of age between VD3-treated KO mice and wildtype (WT) mice. After 1 month injection, serum calcium and phosphorus levels of the KO mice were found indistinguishable from those of their WT littermates. Serum parathyroid hormone level declined significantly, but remained higher in treated KO mice. The dry weight, percentage ash weight, and calcium content of femur were returned to normal levels in VD3-treated KO mice whereas the femoral length, although increased significantly, remained significantly smaller than that of WT mice. VD3 injections also normalized the growth plate of KO mice within normal width. CONCLUSIONS: Our results demonstrate that high-dose VD3 injections can partially rescue the phenotype in 1α-hydroxylase gene KO mice. 25-Hydroxyvitamin D can mediate effects in the absence of conversion to 1α,25-dihydroxyvitamin D was confirmed in this study.
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25-Hidroxivitamina D3 1-alfa-Hidroxilase/deficiência , Colecalciferol/farmacologia , 25-Hidroxivitamina D3 1-alfa-Hidroxilase/genética , Animais , Cálcio/sangue , Colecalciferol/sangue , Fêmur/crescimento & desenvolvimento , Técnicas de Inativação de Genes , Masculino , Camundongos , Camundongos Knockout , Fenótipo , Fosfatos/sangueRESUMO
Doublecortin-like kinase 1 (DCLK1) is a cancer stem cell marker that is highly expressed in various types of human cancer, and a protein kinase target for cancer therapy that is attracting increasing interest. However, no drug candidates targeting DCLK1 kinase have been developed in clinical trials to date. XMD-17-51 was found herein to possess DCLK1 kinase inhibitory activities by cell-free enzymatic assay. In non-small cell lung carcinoma (NSCLC) cells, XMD-17-51 inhibited DCLK1 and cell proliferation, while DCLK1 overexpression impaired the anti-proliferative activity of XMD-17-51 in A549 cell lines. Consequently, XMD-17-51 decreased Snail-1 and zinc-finger-enhancer binding protein 1 protein levels, but increased those of E-cadherin, indicating that XMD-17-51 reduces epithelial-mesenchymal transition (EMT). Furthermore, sphere formation efficiency was significantly decreased upon XMD-17-51 treatment, and XMD-17-51 reduced the expression of stemness markers such as ß-catenin, and pluripotency factors such as SOX2, NANOG and OCT4. However, the percentage of ALDH+ cells was increased significantly following treatment with XMD-17-51 in A549 cells, possibly due to EMT inhibition. In combination, the present data indicated that XMD-17-51 inhibited DCLK1 kinase activity in a cell-free assay with an IC50 of 14.64 nM, and decreased DCLK1 protein levels, cell proliferation, EMT and stemness in NSCLC cell lines. XMD-17-51 has the potential to be a candidate drug for lung cancer therapy.
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OBJECTIVE: The sodium-glucose transporter 2 (SGLT2) inhibitors Canagliflozin and Dapagliflozin are recently approved medications for type 2 diabetes. Recent studies indicate the potential ability of SGLT2 inhibitors to attenuate cancer growth of SGLT2-expressing cancer cells, but there is little known about the effects of SGLT2 inhibitors on breast cancer. The goal in this research was to assess the anticancer activity of SGLT2 inhibitors in breast cancerin vitro and in vivo. METHODS: We test the SGLT2 expression in breast cancer using immunohistochemistry and immunoblot assay. MTT cytotoxicity assay, colony formation assay and human breast cancer cells nude mice xenograft model were performed to detect the effects of SGLT2 inhibitors on cancer cell proliferation and growth. Flow Cytometry assay was performed to determine if the SGLT2 inhibitors induced cell cycle arrest and apoptosis. RESULTS: We proved that SGLT2 expresses in breast cancer cell lines and human breast tumor tissue samples. SGLT2 inhibitors Dapagliflozin and Canagliflozin exhibited a potent anti-proliferative effect in breast cancer cells as demonstrated by MTT, clonogenic survival assay in vitro and xenograft growth model in vivo. Furthermore, we found that SGLT2 inhibitors arrested cell cycle in G1/G0 phase and induced cell apoptosis. Western blot analysis demonstrated that treatment with SGLT2 inhibitors increased the phosphorylation of Amp-activated protein kinase (AMPK) and decreased the phosphorylation of 70 kDa ribosomal protein S6 kinase 1 (p70S6K1) in breast cancer cells. CONCLUSIONS: These findings indicate that SGLT2 inhibitor-therapy induced AMPK-mediated cell cycle arrest and apoptosis, which is a potential novel strategy for the treatment of breast cancer.
Assuntos
Compostos Benzidrílicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Canagliflozina/farmacologia , Glucosídeos/farmacologia , Inibidores do Transportador 2 de Sódio-Glicose/farmacologia , Proteínas Quinases Ativadas por AMP/metabolismo , Adulto , Idoso , Animais , Apoptose/efeitos dos fármacos , Neoplasias da Mama/patologia , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Glucose/metabolismo , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Pessoa de Meia-Idade , Serina-Treonina Quinases TOR/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Chemotherapy and radiotherapy are not as successful in the case of renal cell carcinoma (RCC) although some targeted drugs were approved for RCC therapy recently. Analysis of whole genomic data will lead to improvements in understanding RCC and identifying novel anticancer targets. Here, we found the differential mRNA expression and copy number variation (CNV) of Carbonic anhydrase-related protein VIII (CA8) gene in RCC through integrated bioinformatics analysis of TCGA database, which was confirmed in 5 cases of samples collected from RCC patients who underwent radical nephrectomy by analysis of CA8 mRNA and protein levels using RT-PCR immunohistochemical assay. However, we got a completely opposite result that CA8 promoted RCC progression, those are CA8 overexpression promoted the proliferative and migratory ability of Caki-1 and 769-P cells in vitro as determined with MTT and transwell assay, and CA8 overexpression could also promote Caki-1 xenograft growth in BALB/Cnu/nu mice. On the contrary, CA8-knockdown reduced Caki-1 and 769-P cell proliferation and migration. Moreover, knockdown of CA8 decreased pAKT and MMP2 protein levels in Caki-1 cells while overexpressing CA8 increased pAKT and MMP2. In conclusion, we showed that CA8 promoted RCC cell proliferation and migration, but it was down-regulated in RCC, which requires an additional mechanism study.
Assuntos
Biomarcadores Tumorais/genética , Carcinoma de Células Renais/patologia , Proliferação de Células/genética , Neoplasias Renais/patologia , Animais , Carcinoma de Células Renais/genética , Linhagem Celular Tumoral , Variações do Número de Cópias de DNA , Técnicas de Silenciamento de Genes , Humanos , Neoplasias Renais/genética , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , RNA Mensageiro/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
PURPOSE: We evaluated the role of everolimus in the prevention of ductal carcinoma in situ (DCIS) to invasive ductal carcinoma (IDC) progression. EXPERIMENTAL DESIGN: The effects of everolimus on breast cancer cell invasion, DCIS formation, and DCIS progression to IDC were investigated in a 3D cell culturing model, intraductal DCIS xenograft model, and spontaneous MMTV-Her2/neu mouse model. The effect of everolimus on matrix metalloproteinase 9 (MMP9) expression was determined with Western blotting and IHC in these models and in patients with DCIS before and after a window trial with rapamycin. Whether MMP9 mediates the inhibition of DCIS progression to IDC by everolimus was investigated with knockdown or overexpression of MMP9 in breast cancer cells. RESULTS: Everolimus significantly inhibited the invasion of human breast cancer cells in vitro. Daily intragastric treatment with everolimus for 7 days significantly reduced the number of invasive lesions from intraductal DCIS foci and inhibited DCIS progression to IDC in the MMTV-Her2/neu mouse mammary tumor model. Mechanistically, everolimus treatment decreased the expression of MMP9 in the in vitro and in vivo models, and in breast tissues from patients with DCIS treated with rapamycin for 1 week. Moreover, overexpression of MMP9 stimulated the invasion, whereas knockdown of MMP9 inhibited the invasion of breast cancer cell-formed spheroids in vitro and DCIS in vivo. Knockdown of MMP9 also nullified the invasion inhibition by everolimus in vitro and in vivo. CONCLUSIONS: Targeting mTORC1 can inhibit DCIS progression to IDC via MMP9 and may be a potential strategy for DCIS or early-stage IDC therapy.
Assuntos
Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Carcinoma Intraductal não Infiltrante/tratamento farmacológico , Everolimo/farmacologia , Metaloproteinase 9 da Matriz/metabolismo , Animais , Biomarcadores Tumorais/metabolismo , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Carcinoma Intraductal não Infiltrante/metabolismo , Carcinoma Intraductal não Infiltrante/patologia , Linhagem Celular Tumoral , Movimento Celular , Progressão da Doença , Regulação para Baixo , Feminino , Humanos , Metaloproteinase 9 da Matriz/química , Camundongos , Camundongos Nus , Camundongos Transgênicos , Receptor ErbB-2/genética , Receptor ErbB-2/metabolismo , Esferoides Celulares/efeitos dos fármacos , Esferoides Celulares/metabolismo , Esferoides Celulares/patologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
OBJECTIVE: To evaluate and compare the postoperative implant-related complications after treatment of intertrochanteric fracture by gamma nail or by sliding hip screw. METHODS: Literatures on randomized controlled trials (RCTs) of the postoperative implant-related complications after treatment of intertrochanteric fracture by gamma nail or by sliding hip screw published before the end of June 2007 were retrieved from Cochrane library, Medline, Embase databank, and Chinese Biomedical Database. Relevant articles were handsearched in several Chinese journals. Methodology quality of the trials was critically assessed and data on postoperative implant-related complications were extracted. The software RevMan4.2.10 provided by the Cochrane Collaboration was used for meta-analysis. RESULTS: 10 RCTs and 3 quasi-RCTs involving 1857 cases were included. The results of meta-analysis showed that gamma nailing significantly increased the rate of postoperative femoral shaft fractures [RR = 3.75, 95%CI (1.47, 9.55), P = 0.006]. However, there were no statistic differences in the rates of lag screw cut-out and total mechanical complications between gamma nail and sliding hip screw treatment. CONCLUSION: Compared with sliding hip screw for intertrochanteric fractures, gamma nailing can lead to higher incidence of postoperative femoral shaft fractures. Fracture stability may influence the rate of postoperative implant-related complications. Further large sample and high quality RCTs based on fracture stability are required in order to provide more sufficient and reliable evidence.
Assuntos
Pinos Ortopédicos , Parafusos Ósseos , Fixação Interna de Fraturas/instrumentação , Fraturas do Quadril/cirurgia , Complicações Pós-Operatórias , Adulto , Fixação Interna de Fraturas/efeitos adversos , Fixação Interna de Fraturas/métodos , Humanos , Complicações Pós-Operatórias/cirurgia , Ensaios Clínicos Controlados Aleatórios como AssuntoRESUMO
Previous research has revealed that the von-Hippel-Lindau tumor suppressor protein (pVHL) may downregulate never-in-mitosis A-related kinase 8 (NEK8) via hypoxia-inducible factor-α (HIF-α). The HIF-independent functions of pVHL also serve an important role in its tumor-suppressor action. In the present study, the association between pVHL and NEK8 was demonstrated in the human gastric cancer cell line, SGC-7901, indicating a direct interaction of pVHL with NEK8. Subsequently, it was reported that MG-132, a specific proteasome inhibitor, may attenuate pVHL overexpression-induced reductions in NEK8 protein expression levels. In addition, the present study revealed that pVHL may stimulate the rapid degradation of NEK8 protein and promote its ubiquitination. The association between the expression profile of NEK8 and the survival status of patients with gastric cancer was analyzed from an online database. Kaplan-Meier survival plots indicated that higher expression levels of NEK8 may lead to poor survival, as suggested by the transcriptomic data of 1,065 patients with gastric cancer. It was found that NEK8-knockdown mediated by RNA interference inhibited SGC-7901 and SNU-1 proliferation, colony formation and migration in vitro, and tumor growth in vivo. Collectively, the present study proposed that NEK8 may be a novel target of pVHL as a ubiquitin E3 ligase, and may serve a role as a potential oncoprotein in human gastric cancer.