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Destruction of erythropoiesis process leads to various diseases, including thrombocytopenia, anaemia, and leukaemia. miR-429-CT10 regulation of kinase-like (CRKL) axis involved in development, progression and metastasis of cancers. However, the exact role of miR-429-CRKL axis in leukaemic cell differentiation are still unknown. The current work aimed to uncover the effect of miR-429-CRKL axis on erythropoiesis. In the present study, CRKL upregulation was negatively correlated with miR-429 downregulation in both chronic myeloid leukaemia (CML) patient and CR patient samples. Moreover, CRKL expression level was significantly decreased while miR-429 expression level was increased during the erythroid differentiation of K562 cells following hemin treatment. Functional investigations revealed that overexpression and knockdown of CRKL was remarkably effective in suppressing and promoting hemin-induced erythroid differentiation of K562 cells, whereas, miR-429 exhibited opposite effects to CRKL. Mechanistically, miR-429 regulates erythroid differentiation of K562 cells by downregulating CRKL via selectively targeting CRKL-3'-untranslated region (UTR) through Raf/MEK/ERK pathway. Conversely, CRKII had no effect on erythroid differentiation of K562 cells. Taken together, our data demonstrated that CRKL (but not CRKII) and miR-429 contribute to development, progression and erythropoiesis of CML, miR-429-CRKL axis regulates erythropoiesis of K562 cells via Raf/MEK/ERK pathway, providing novel insights into effective diagnosis and therapy for CML patients.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Diferenciação Celular , Células Eritroides , Hemina , Leucemia Mielogênica Crônica BCR-ABL Positiva , MicroRNAs , Proteínas Proto-Oncogênicas c-crk , Humanos , Regiões 3' não Traduzidas , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Diferenciação Celular/efeitos dos fármacos , Células Eritroides/metabolismo , Células Eritroides/efeitos dos fármacos , Células Eritroides/patologia , Células Eritroides/citologia , Eritropoese/genética , Eritropoese/efeitos dos fármacos , Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , Hemina/farmacologia , Células K562 , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , MicroRNAs/genética , MicroRNAs/metabolismo , Proteínas Proto-Oncogênicas c-crk/metabolismo , Proteínas Proto-Oncogênicas c-crk/genéticaRESUMO
As a member of transcription factor E-Twenty Six (ETS) family, ETS variant 6 (ETV6) plays significant role in hematopoiesis and embryonic development. ETV6 dysexpression also involved in the occurrence, development and progression of cancers and leukemia. In current work, we hypothesized that ETV6 plays a role in erythroid differentiation of chronic myeloid leukemia (CML). We found the protein expression level of ETV6 was significantly upregulated during hemin-induced erythroid differentiation of K562 cells. Moreover, overexpression of ETV6 inhibited erythroid differentiation in hemin-induced K562 cells with decreased numbers of benzidine-positive cells and decreased expression levels of erythroid differentiation specific markers glycophorin (GPA), CD71, hemoglobin A (HBA), α-globin, γ-globin and ε-globin. Conversely, ETV6 knockdown promoted erythroid differentiation in hemin-induced K562 cells. Furthermore, ETV6 expression level slightly positively with the proliferation capacity of K562 cells treated with hemin. Mechanistically, ETV6 overexpression inhibited fibrosarcoma/mitogen activated extracellular signal-regulated kinase/extracellular regulated protein kinase (Raf/MEK/ERK) pathway, ETV6 knockdown activated the Raf/MEK/ERK pathway. Collectively, the current work demonstrates that ETV6 plays an inhibitory role in the regulation of K562 cell erythroid differentiation via Raf/MEK/ERK pathway, it would be a potentially therapeutic target for dyserythropoiesis.
Assuntos
Hemina , Leucemia Mielogênica Crônica BCR-ABL Positiva , Sistema de Sinalização das MAP Quinases , Proteínas Proto-Oncogênicas c-ets , Proteínas Repressoras , Quinases raf , Diferenciação Celular , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Hemina/farmacologia , Humanos , Células K562 , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Proteínas Proto-Oncogênicas c-ets/metabolismo , Proteínas Repressoras/metabolismo , Quinases raf/metabolismo , Variante 6 da Proteína do Fator de Translocação ETSRESUMO
Abnormal glucose metabolism may contribute to cancer progression. As a member of the CRK (v-crk sarcoma virus CT10 oncogene homologue) adapter protein family, CRKL (CRK-like) associated with the development and progression of various tumours. However, the exact role and underlying mechanism of CRKL on energy metabolism remain unknown. In this study, we investigated the effect of CRKL on glucose metabolism of hepatocarcinoma cells. CRKL and PI3K were found to be overexpressed in both hepatocarcinoma cells and tissues; meanwhile, CRKL up-regulation was positively correlated with PI3K up-regulation. Functional investigations revealed that CRKL overexpression promoted glucose uptake, lactate production and glycogen synthesis of hepatocarcinoma cells by up-regulating glucose transporters 1 (GLUT1), hexokinase II (HKII) expression and down-regulating glycogen synthase kinase 3ß (GSK3ß) expression. Mechanistically, CRKL promoted glucose metabolism of hepatocarcinoma cells via enhancing the CRKL-PI3K/Akt-GLUT1/HKII-glucose uptake, CRKL-PI3K/Akt-HKII-glucose-lactate production and CRKL-PI3K/Akt-Gsk3ß-glycogen synthesis. We demonstrate CRKL facilitates HCC malignancy via enhancing glucose uptake, lactate production and glycogen synthesis through PI3K/Akt pathway. It provides interesting fundamental clues to CRKL-related carcinogenesis through glucose metabolism and offers novel therapeutic strategies for hepatocarcinoma.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Carcinoma Hepatocelular/etiologia , Carcinoma Hepatocelular/metabolismo , Glucose/metabolismo , Neoplasias Hepáticas/etiologia , Neoplasias Hepáticas/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Suscetibilidade a Doenças , Regulação Neoplásica da Expressão Gênica , Glicogênio/biossíntese , Humanos , Neoplasias Hepáticas/patologia , Proteômica/métodos , Transdução de SinaisRESUMO
MicroRNAs (miRNAs) are a class of single-stranded noncoding and endogenous RNA molecules with a length of 18-25 nucleotides. Previous work has shown that miR-124-3p leads to malignant progression of cancer including cell apoptosis, migration, invasion, drug resistance, and also recovers neural function, affects adipogenic differentiation, facilitates wound healing through control of various target genes. miR-124-3p has been mainly previously characterized as a tumor suppressor regulating tumorigenesis and progression in several cancers, such as hepatocellular carcinoma (HCC), gastric cancer (GC), bladder cancer, ovarian cancer (OC), and leukemia, as a tumor promotor in breast cancer (BC), and it has been also widely studied in a variety of neurological diseases, like Parkinson's disease (PD), dementia and Alzheimer's disease (AD), and cardiovascular diseases, ulcerative colitis (UC), acute respiratory distress syndrome (ARDS). To lay the groundwork for future therapeutic strategies, in this review we mainly focus on the most recent years of literature on the functions of miR-124-3p in related major cancers, as well as its downstream target genes. Although current work as yet provides an incomplete picture, miR-124-3p is still worthy of more attention as a practical and effective clinical biomarker.
Assuntos
Carcinogênese/metabolismo , MicroRNAs/metabolismo , Animais , Carcinogênese/genética , Doenças Cardiovasculares/genética , Doenças Cardiovasculares/metabolismo , Humanos , MicroRNAs/genética , Neoplasias/genética , Neoplasias/metabolismo , Doenças Neurodegenerativas/genética , Doenças Neurodegenerativas/metabolismoRESUMO
AIM: To investigate ANXA5 overexpression on in vitro and in vivo malignancies of murine Hca-P cells. MATERIALS & METHODS: Hca-P with low lymph node metastasis (LNM) potential was used as cell model. TEM, CCK-8 and Boyden transwell assays were performed for in vitro Hca-P behaviors. Hca-P-transplanted mouse model was established for in vivo experiment. RESULTS: ANXA5-overexpressing monoclonal Anxa5-Hca-P-1, Anxa5-Hca-P-2 and Anxa5-Hca-P-3 cells were obtained. ANXA5 upregulation alters the proliferation, morphology and rough endoplasmic reticulum of Hca-P cells, enhances in vitro migration and invasions of Hca-P, promotes in vivo malignant degree and LNM rate of Anxa5-Hca-P-3-transplanted mice. CONCLUSION: As a potential indicator for malignancy and lymphatic metastasis, ANXA5 overexpression increases in vitro migration and invasion of Hca-P cell, promotes in vivo malignancy, LNM rate and level of Hca-P-transplanted mice.
Assuntos
Anexina A5/biossíntese , Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/genética , Metástase Linfática/genética , Animais , Anexina A5/genética , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células/genética , Modelos Animais de Doenças , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/patologia , Metástase Linfática/patologia , Camundongos , Invasividade Neoplásica/genética , Invasividade Neoplásica/patologiaRESUMO
V-crk avian sarcoma virus CT10 oncogene homolog-like (CRKL) is a member of CRK family. It acts as an adaptor protein in intracellular signal transduction. CRKL has been reported overexpressed in a variety of cancers affecting the aggressive and malignant behaviors of cancer cells. CRKL seems to show a tumor-promotion role in gastric cancer, glioblastoma multiforme, hepatocellular carcinoma, bladder cancer, lung cancer, colon cancer, ovarian cancer, leukemia, breast cancer, head and neck cancer, rhabdomyosarcoma and neuroblastoma. The association of CRKL with malignant tumors and its potential action mechanisms were summarized. CRKL has the potential to be used as a biomarker for the diagnosis, treatment and prognosis of certain tumors.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Transformação Celular Neoplásica/genética , Neoplasias/genética , Proteínas Nucleares/genética , Prognóstico , Biomarcadores Tumorais/genética , Humanos , Neoplasias/classificação , Neoplasias/diagnóstico , Neoplasias/patologia , Transdução de Sinais/genéticaRESUMO
The ETV6-MECOM fusion gene, produced by the rare and recurrent chromosomal translocation t(3; 12) (q26; p13), is associated with high mortality and short survival in myeloid leukemia. However, its function and underlying mechanisms in leukemia progression remain unknown. In this study, leukemia-stable K562 cells expressing the ETV6-MECOM fusion protein were used to investigate the effects of the ETV6-MECOM oncoprotein. K562-ETV6-MECOM cells were undifferentiated and had reduced colony formation, increased cell migration and invasion, and increased sphere number and diameter in a spheroid formation assay, presenting epithelial-to-mesenchymal transition (EMT) traits. The expression of E-cadherin, a hallmark of EMT, was significantly downregulated at the transcriptional and translational level in K562-ETV6-MECOM cells to explore the mechanistic basis of EMT. Stepwise truncation, DNA sequence deletion, mutation analysis for E-cadherin promoter transactivation, and a dual luciferase assay indicated that the regulatory region of ETV6-MECOM is located in the DNA motif -1116 TTAAAA-1111 of E-cadherin promoter. Moreover, a chromatin immunoprecipitation assay showed that this oncoprotein binds to the DNA motif -1116 TTAAAA-1111 with the anti-EVI1 antibody. Although ETV6-MECOM upregulated the expressions of EMT master regulators, including SNAIL, SLUG, ZEB2, and TWIST2, their knockdown had no effect on EMT-related properties. However, overexpression of E-cadherin eliminated EMT traits in the presence of the ETV6-MECOM oncoprotein. These data confirmed that the ETV6-MECOM oncoprotein, not SNAIL, SLUG, ZEB2, or TWIST2, plays a critical role in inducing EMT traits in leukemia K562 cells. ETV6-MECOM induces EMT-related properties by downregulating the transcriptional expression of E-cadherin and repressing its transactivation activity by binding to its core motif -1116TTAAAA-1111 in leukemia K562 cells. These findings could contribute to the development of a therapeutic target for patients with myeloid leukemia characterized by ETV6-MECOM.
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The annexins are a well-known, closely related, multigene superfamily of Ca(2+)-regulated, phospholipid-dependent, membrane-binding proteins. As a member of the annexins, Anxa1 participates in a variety of important biological processes, such as cellular transduction, membrane aggregation, inflammation, phagocytosis, proliferation, differentiation and apoptosis. Accumulated evidence has indicated that Anxa1 deregulations are associated with the development, invasion, metastasis, occurrence and drug resistance of cancers. The research evidence in recent years indicates that Anxa1 might specifically function either as a tumor suppressor or a tumor promoter candidate for certain cancers depending on the particular type of tumor cells/tissues. This article summarizes the associations between Anxa1 and malignant tumors, as well as potential action mechanisms. Anxa1 has the potential to be used in the future as a biomarker for the diagnosis, treatment and prognosis of certain tumors.
Assuntos
Anexina A1/fisiologia , Neoplasias/metabolismo , Animais , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Biomarcadores Tumorais/metabolismo , Resistencia a Medicamentos Antineoplásicos , Humanos , Invasividade Neoplásica , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Regulação para CimaRESUMO
Lymphatic metastasis is the most common form in breast cancer (BC) progression. Previously, we observed that lnc045874, a most conservative homology of Homo Sapiens NONHSAT021545 (lnc021545), miR-330-3p, and EREG may have some effects in mouse hepatocarcinoma cell lines with different lymphatic metastasis potentials. Through data from TCGA and GEO database analysis, we speculated that miR-330-3p might be a tumor promoter, while EREG could be a tumor suppressor in BC. MiR-330-3p was upregulated, while lnc021545 and EREG were downregulated in 50 BC tissues. MiR-330-3p advanced the metastatic behaviors of BC cells, whereas lnc021545 and EREG resulted in the opposite effects. The three molecules' expressions were correlated respectively and showed that miR-330-3p targeted lnc021545 and EREG to affect their expressions. Lnc021545/miR-330-3p axis affected BC metastasis by regulating EREG in epithelial-to-mesenchymal transition. In 50 BC patients, these three molecules and their cooperation are associated with aggressive tumor phenotypes, patient outcomes, and trastuzumab therapy. We finally discovered that lnc021545, miR-330-3p, and EREG formed a multi-gene co-regulation system that affected the metastasis of BC and the cooperation reflects the synergistic effects of the three molecules, recommending that their cooperation may provide a more accurate index for anti-metastasis therapeutic and prognostic evaluation of BC.
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Cellular senescence serves as a powerful tumor suppressing mechanism that inhibits the proliferation of cancer cells bearing oncogenic mutations at the initial stage of cancer development. RNA-binding proteins (RBPs) play important roles in cancer progression and treatment through distinct functions. However, functions and mechanisms of RNA binding proteins in regulating senescence remain elusive. Here we reported that the RNA binding protein RBM4 contributed to cellular senescence. Depletion of RBM4 induced senescence in different types of cells, including multiple cancer cells. Meanwhile, RBM4 ablation inhibited cancer cell progression both in vitro and in vivo. Specifically, knockdown of RBM4 significantly increased the level of SERPINE1, a known promoter of senescence, thereby inducing the senescence of lung cancer cells. Mechanistically, miR-1244 bound to the 3'-UTR of SERPINE1 to suppress its expression, whereas depletion of RBM4 reduced the level of miR-1244 by promoting the degradation of primary miR-1244 transcripts (pri-miR1244), thus increasing the expression of SERPINE1 and inducing subsequent senescence. Moreover, either SERPINE1 inhibitor or miR-1244 mimics attenuated the RBM4 depletion-induced senescence. Altogether, our study revealed a novel mechanism of RBM4 in the regulation of cancer progression through controlling senescence, providing a new avenue for targeting RBM4 in cancer therapeutics.
Assuntos
Neoplasias Pulmonares , MicroRNAs , Humanos , Processamento Alternativo , Senescência Celular/genética , MicroRNAs/genética , MicroRNAs/metabolismo , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Inibidor 1 de Ativador de Plasminogênio/genética , Inibidor 1 de Ativador de Plasminogênio/metabolismoRESUMO
To examine the differential expression pattern of Ech1 protein in mouse Hca-F and Hca-P hepatocarcinoma cell lines with high and low rates of lymphatic metastasis, respectively, and to investigate the relationships between Ech1 expression and adhesion of Hca-F cells. Fluorescence two-dimensional difference in-gel electrophoresis (2D DIGE) and mass spectrometry were used to detect Ech1 expression. Ech1 gene silencing was achieved by stable transfection of Hca-F cells with a plasmid vector harboring short hairpin RNA (shRNA) targeting Ech1, pGPU6/GFP/Neo-shRNA-Ech1. Ech1 mRNA and protein expressions were detected by real-time quantitative polymerase chain reaction (qRT-PCR) and Western blotting analysis, respectively. Adhesive properties of cells were assessed by hematoxylin-eosin staining and fluorimetric detection of extracellular matrix (ECM) proteins. Endogenous Ech1 protein level was remarkably higher in the highly metastatic Hca-F cell line than in the Hca-P cell line (2.7-fold by 2D DIGE; 1.5-fold by Western blotting). shRNA-induced silencing of Ech1 significantly reduced the adhesion ability of Hca-F cells, as evidenced by decreased absorbance values of fibronectin and collagen I (Hca-F cells vs. pGPU6/GFP/Neo-shRNA-Ech1 cells: 1.42+/-0.26 vs. 1.01+/-0.27 and 1.14+/-0.07 vs. 0.90+/-0.09, respectively; P less than 0.05). Down-regulation of Ech1 can inhibit the adhesive capacity of metastatic Hca-F cells.
Assuntos
Isomerases de Ligação Dupla Carbono-Carbono/metabolismo , Carcinoma Hepatocelular/patologia , Adesão Celular , Neoplasias Hepáticas/patologia , RNA Interferente Pequeno/genética , Animais , Isomerases de Ligação Dupla Carbono-Carbono/genética , Carcinoma Hepatocelular/enzimologia , Carcinoma Hepatocelular/genética , Linhagem Celular Tumoral , Regulação para Baixo , Eletroforese em Gel Bidimensional , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas/enzimologia , Neoplasias Hepáticas/genética , Linfonodos/patologia , Metástase Linfática , Camundongos , Plasmídeos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA Interferente Pequeno/farmacologia , TransfecçãoRESUMO
A better understanding of tumor metastasis is urgently required for the treatment and prognosis of hepatocarcinoma patients. Current work contributes a novel ceRNA feedback regulation pathway composed of epiregulin (EREG), microRNA-330-3p (miR-330-3p) and long non-coding RNA 021545 (lncRNA021545) in regulating hepatocarcinoma malignancy via epithelial-mesenchymal transition (EMT) process. Closely correlated, the deficiencies of EREG and lncRNA021545 and the overexpression of miR-330-3p were involved in the clinical progression of hepatocarcinoma. In vitro results showed that 1) lncRNA021545 downregulation promoted, 2) miR-330-3p dysexpression positively correlated, and 3) EREG dysexpression reversely correlated with the migratory and invasive properties of hepatocarcinoma HCCLM3 and Huh7 cell lines. By directly binding to EREG and lncRNA021545, miR-330-3p expression change reversely correlated with their expressions in HCCLM3 and Huh7 cells, which was also confirmed in primary tumors from HCCLM3-xenograft mice in responding to miR-330-3p change. LncRNA021545 and EREG positively regulated each other, and lncRNA021545 negatively regulated miR-330-3p, while, EREG dysregulation unchanged miR-330-3p expression in hepatocarcinoma cells. Furthermore, systemic in vitro cellular characterizations showed that the malfunctions of the three molecules mediated the invasiveness of hepatocarcinoma cells via EMT process through affecting the expressions of E-cadherin, N-cadherin, vimentin, snail and slug, which was further confirmed by in vivo miR-330-3p promotion on the tumorigenicity and metastasis of HCCLM3 bearing nude mice and by in vitro miR-330-3p promotion on the migration and invasion of hepatocarcinoma cells to be antagonized by EREG overexpression through acting on EMT process. Our work indicates, that by forming a circuit signaling feedback pathway, the homeostatic expressions of lncRNA021545, miR-330-3p and EREG are important in liver health. Its collapse resulted from the downregulations of lncRNA021545 and EREG together with miR-330-3p overexpression promote hepatocarcinoma progression by enhancing the invasiveness of tumor cells through EMT activation. These discoveries suggest that miR-330-3p/lncRNA021545/EREG axis plays a critical role in hepatocarcinoma progression and as a candidate for its treatment.
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Introduction: Globally, hepatocellular carcinoma (HCC) is the sixth most common malignancy and it has the fourth highest mortality. MicroRNAs play a significant part in biological processes in cell formation and advancement by targeting genes in many cancers including HCC. Objective: In the present study we examine the involvement of miR-4521 and FAM129A correlations in HCC occurrence and progression. Methods: Expression levels of miR-4521 and FAM129A in HCC tissues and cells were detected. Immunohistochemistry was carried out to detect expression of FAM129A, MMP9 and TIMP-1 in HCC tissues. Western blot assays were used to examine expression levels of different genes involve in signaling pathways. Transwell chamber, MTT and wound healing assays were performed to check cell migration, invasion and proliferation rates. Results: Overexpression of FAM129A positively correlated with upregulation of MMP9 and negatively correlated with TIMP-1 in HCC patient samples, which encouraged progression and metastasis of HCC. An antagonistic relation between miR-4521 and FAM129A was detected in current study, down-regulation of miR-4521 and up-regulation of FAM129A was demonstrated in HCC tissues and cell lines as compare to normal tissue samples and the normal cell line LO2. Overexpressing miR-4521 and silencing FAM129A impaired HCC cell migratory and invasive properties and suppressed cell proliferation. Mutually, miR-4521-FAM129A axial regulation inhibited in vitro proliferation of cells by promoting apoptosis through the p-FAK/p-AKT/MDM2/P53 and p-FAK/p-AKT/BCL-2/BAX/Cytochrome-C/Caspase-3/Caspase-9 pathways, respectively, and suppressed the migration and invasion capabilities of HCCLM3 and HepG2 cells via the TIMP-1/MMP9/MMP2 and p-FAK/p-AKT pathway. Conclusion: Our work found the axial regulation mechanism of miR-4521-FAM129A in HCC. Deficiency of miR-4521 and abundance of FAM129A synergistically enhanced cancer progression by increasing cell proliferation and malignant invasion and by inhibiting apoptosis. These discoveries suggest that miR-4521/FAM129A might play a vital role in hepatic cancer progression and could be a candidate for its therapy.
Assuntos
Biomarcadores Tumorais/antagonistas & inibidores , Carcinoma Hepatocelular , Neoplasias Hepáticas , MicroRNAs , Proteínas de Neoplasias/antagonistas & inibidores , Biomarcadores Tumorais/metabolismo , Carcinoma Hepatocelular/metabolismo , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Hepáticas/metabolismo , MicroRNAs/genética , MicroRNAs/metabolismo , Invasividade Neoplásica/genética , Proteínas de Neoplasias/metabolismo , Fosforilação , Proteínas Proto-Oncogênicas c-akt/genéticaRESUMO
To investigate the mechanism and biologic effects of 37 nm magnetic nano FeOx powders (MNPs) on human hepatoma-bearing nude mice. 37 nm MNPs were prepared by coprecipitation methods and then injected into human hepatoma (Bel-7402) bearing-nude mice through the tail vein. After injection of MNPs, the mice were first exposed under static magnetic field and then treated under extremely low frequency altering-electric magnetic field directing to the tumor area. The migration and trafficking of MNPs were determined by MMR. Tumor growth was monitored with calipers every 5 days and tumor volume was calculated on the basis of three-dimensioned measurements. The apoptosis of tumor cells was analyzed by flow cytometry analysis. The expressions of apoptosis-associated proteins Bcl-2, Bax and HSP27 were determined using western-blot analysis. Static magnetic field could direct the migration and trafficking of MNPs to the tumor site with a higher ratio of 98.9%. Extremely Low Frequency Electric-Magnetic Field (EMF) treatment could inhibit the proliferation of tumor cells and prolong the survive time of tumor-bearing mice injected with MNPs. In addition, the survival time of tumor-bearing mice and percentages of prohibition on tumor cell growth were 27.4+/-0.7 days and 37.5+/-0.8% (F = 0.005, P is less than to 0.05), respectively. The results of flow cytometry analyses showed that about 18.1+/-0.6% (F = 0.030, P is less than to 0.05) of tumor cells were induced into early apoptosis. Furthermore, expressions of apoptosis-associated proteins Bcl-2 and Bax were significantly induced by MNPs under EMF treatment. The ratio of Bcl/Bax in both MNPs and EMF treatment group was 0.07+/-0.01, which was much lower than that of control group (0.23+/-0.02) (F = 0.016, P is less than to 0.05). Heat shock protein-27 (Hsp-27) was not significantly induced in different treatment groups. Injection of MNPs with EMF exposure on human hepatoma-bearing nude mice could significantly prolong the survival time, inhibit the tumor proliferation and growth, and induce tumor cells into apoptosis.
Assuntos
Proteínas Proto-Oncogênicas c-bcl-2 , Proteína X Associada a bcl-2 , Animais , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Produtos Biológicos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Citometria de Fluxo , Humanos , Campos Magnéticos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Transplante de Neoplasias , PósRESUMO
Clinical and experimental findings support the view that activation of hippocampus microglia through NADPH oxidase contributes to cognitive impairment in Parkinson's disease (PD). Taurine, an antioxidant, displays an exclusive physical property on brain function, such as learning and memory. To date, the role of taurine in improving cognitive impairment in PD is not fully uncovered. Hence, we evaluated the protective effect of taurine on cognitive ability and explored the related mechanism in the model built by paraquat and maneb (P + M)-induced PD mice. Then the ability of learning and memory was observed by Morris water maze, neuron loss was evaluated by immunohistochemistry in hippocampus, the level of postsynaptic density 95 (PSD95) and microglia activation was assessed by immunostaining, the molecules (gp91phox, p47phox, mac1, p-Src/Src and p-Erk/Erk) were examined by western blot. The results showed that taurine could alleviate the impairments in learning and memory induced by P + M injection in mice (decreased escape latency on day 4, P < 0.01; decreased swimming distance on day 4, P < 0.05; increased percent time in target quadrant, P < 0.05), corresponding with activation of microglia (decreased IBa-1 density, P < 0.001; decreased the protein expression of p47phox, P < 0.05; decreased protein expression of gp91phox, P < 0.01; decreased p-Src/Src, P < 0.01; decreased p-Erk/Erk, P < 0.01; decreased mac 1, P < 0.01), decreased neuron loss (increased number of NeurN+ neuron, P < 0.001; increased protein expression of NeruN, P < 0.01; decreased protein expression of caspase 3, P < 0.01) and increased PSD95 level in hippocampus (P < 0.01). The results indicated that mac1 and Src-Erk signaling was involved in increased NADPH oxidase expression in hippocampus microglia of P + M mice, and taurine could improve injuries in learning and memory through mac1 reduction. The new findings in mac1 triggering hippocampal microglia NADPH oxidase through Src/Erk pathway of the present study might provide a therapy target for PD.
Assuntos
Antiparkinsonianos/farmacologia , Comportamento Animal/efeitos dos fármacos , Cognição/efeitos dos fármacos , Disfunção Cognitiva/prevenção & controle , Hipocampo/efeitos dos fármacos , Microglia/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Transtornos Parkinsonianos/tratamento farmacológico , Taurina/farmacologia , Animais , Disfunção Cognitiva/metabolismo , Disfunção Cognitiva/fisiopatologia , Disfunção Cognitiva/psicologia , Modelos Animais de Doenças , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Hipocampo/metabolismo , Hipocampo/patologia , Hipocampo/fisiopatologia , Antígeno de Macrófago 1/metabolismo , Masculino , Maneb , Aprendizagem em Labirinto/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Microglia/metabolismo , Microglia/patologia , NADPH Oxidase 2/metabolismo , NADPH Oxidases/metabolismo , Degeneração Neural , Neurônios/metabolismo , Neurônios/patologia , Paraquat , Transtornos Parkinsonianos/metabolismo , Transtornos Parkinsonianos/patologia , Transtornos Parkinsonianos/fisiopatologia , Fosforilação , Transdução de Sinais , Quinases da Família src/metabolismoRESUMO
In mouse models, the recovery of liver volume is mainly mediated by the proliferation of hepatocytes after partial hepatectomy that is commonly accompanied with ischemia-reperfusion. The identification of differently expressed genes in liver following partial hepatectomy benefits the better understanding of the molecular mechanisms during liver regeneration (LR) with appliable clinical significance. Briefly, studying different gene expression patterns in liver tissues collected from the mice group that survived through extensive hepatectomy will be of huge critical importance in LR than those collected from the mice group that survived through appropriate hepatectomy. In this study, we performed the weighted gene coexpression network analysis (WGCNA) to address the central candidate genes and to construct the free-scale gene coexpression networks using the identified dynamic different expressive genes in liver specimens from the mice with 85% hepatectomy (20% for seven-day survial rate) and 50% hepatectomy (100% for seven-day survial rate under ischemia-reperfusion condition compared with the sham group control mice). The WGCNA combined with Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) enrichment analyses pinpointed out the apparent distinguished importance of three gene expression modules: the blue module for apoptotic process, the turquoise module for lipid metabolism, and the green module for fatty acid metabolic process in LR following extensive hepatectomy. WGCNA analysis and protein-protein interaction (PPI) network construction highlighted FAM175B, OGT, and PDE3B were the potential three hub genes in the previously mentioned three modules. This work may help to provide new clues to the future fundamental study and treatment strategy for LR following liver injury and hepatectomy.
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Introduction: As a member of annexin family proteins, annexin A3 (ANXA3) has 36-kDa and 33-kDa isoforms. ANXA3 plays crucial roles in the tumorigenesis, aggressiveness and drug-resistance of cancers. However, previous studies mainly focused on the role of total ANXA3 in cancers without distinguishing the distinction between the two isoforms, the role of 33-kDa ANXA3 in cancer remains unclear. Objectives: Current work aimed to investigate the function and regulation mechanism of 33-kDa ANXA3 in hepatocarcinoma. Methods: The expressions of ANXA3, CRKL, Rac1, c-Myc and pAkt were analyzed in hepatocarcinoma specimens by Western blotting. The biological function of 33-kDa ANXA3 in the growth, metastasis, apoptosis, angiogenesis, chemoresistance of hepatocarcinoma cells with the underlying molecular mechanism were investigated using gain-of-function strategy in vitro or in vivo. Results: 33-kDa ANXA3 was remarkably upregulated in tumor tissues compared with corresponding normal liver tissues of hepatocarcinoma patients. Its stable knockdown decreased the in vivo tumor growing velocity and malignancy of hepatocarcinoma HepG2 cells transplanted in nude mice. The in vitro experimental results indicated 33-kDa ANXA3 knockdown suppressed the proliferation, colony forming, migration and invasion abilities of HepG2 cells through downregulating CRKL, Rap1b, Rac1, pMEK, pERK2 and c-Myc in ERK pathway; inhibited angiogenesisability of HepG2 cells through inactivating PI3K/Akt-HIF pathway; induced apoptosis and enhanced chemoresistance of HepG2 cells through increasing Bax/decreasing Bcl-2 expressions and inactivating caspase 9/caspase 3 in intrinsic apoptosis pathway. Accordingly, CRKL, Rac1, c-Myc and pAkt were also upregulated in hepatocarcinoma patients ' tumor tissues compared with corresponding normal liver tissues. Conclusions: The overexpression of 33-kDa ANXA3 is involved in the clinical progression of hepatocarcinoma and in the malignancy, angiogenesis and apoptosis of hepatocarcinoma cells. It is of potential use in hepatocarcinoma diagnosis and treatment.
Assuntos
Anexina A3/metabolismo , Apoptose , Carcinoma Hepatocelular/metabolismo , Neoplasias Hepáticas/metabolismo , Animais , Carcinogênese/metabolismo , Carcinoma Hepatocelular/patologia , Movimento Celular , Proliferação de Células , Resistencia a Medicamentos Antineoplásicos , Feminino , Células Hep G2 , Humanos , Neoplasias Hepáticas/patologia , Sistema de Sinalização das MAP Quinases , Masculino , Camundongos , Camundongos Nus , Pessoa de Meia-Idade , Fosfatidilinositol 3-Quinases/metabolismo , Isoformas de Proteínas/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismoRESUMO
OBJECTIVE: To study the localization and expression of CLIC1 in mouse hepatocarcinoma ascites cell lines with different metastatic potentials. METHODS: Mouse hepatocarcinoma ascites models (a high potential of lymphatic metastasis cell line-Hca-F, and a low potential of lymphatic metastasis cell line-Hca-P) were investigated using fluorescent two-dimensional difference-gel electrophoresis (2-D DIGE) and mass spectrometry for detecting the localization and expression of CLIC1. Immunofluorescence, immunocytochemistry and Western blot were used to assess CLIC1 protein status in the two cell lines. RESULTS: CLIC1 expression was obtained in the cytoplasm and plasma membrane of cells in both cell lines. 2-D DIGE showed that CLIC1 was overexpressed in Hca-F cells, 1.6 folds higher than that of the Hca-P cells. Hca-F cells also had a higher integral membrane CLIC1 in the Hca-P cells. CONCLUSIONS: Although CLIC1 expression is detected in both Hca-F and Hca-P cell lines, a higher protein expression level is present in Hca-F cells. CLIC1 may play an important role in tumor metastasis.
Assuntos
Ascite/metabolismo , Canais de Cloreto/metabolismo , Neoplasias Hepáticas Experimentais/metabolismo , Animais , Ascite/patologia , Western Blotting , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Citoplasma/metabolismo , Imuno-Histoquímica , Neoplasias Hepáticas Experimentais/patologia , Metástase Linfática , Camundongos , Camundongos Endogâmicos , Eletroforese em Gel Diferencial BidimensionalRESUMO
Clear cell renal cell carcinoma (ccRCC) exhibits the highest mortality among all urological malignancies. The investigation of the potential disease progression markers can improve ccRCC diagnosis and treatment. Gene associated with retinoid-interferon-induced mortality-19 (GRIM-19) is involved in carcinogenesis and cancer progression in a variety of cancer types including RCC. While, its role in ccRCC remains unclear, this cancer type is considered the most aggressive RCC subtype. In the present study, RT-qPCR, western blotting and immunohistochemical (IHC) assays demonstrated that GRIM-19 protein and mRNA levels were downregulated in ccRCC tumor tissues compared with the corresponding levels noted in paracancerous non-tumor tissues. The deficiency of this protein contributed in relaxed and/or collapsed structures of the kidney tubules and collecting duct noted in tumor tissues. Moreover, the reduction in GRIM-19 expression was associated with high tumor, lymph nodes and metastasis (TNM) stage and Fuhrman grade of ccRCC tumors. The data suggested that GRIM-19 acted as a tumor suppressor and that its deficiency promoted ccRCC development and progression. GRIM-19 can be considered a potential tumor marker for ccRCC.
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Actin filament-associated protein 1 antisense RNA 1 (AFAP1-AS1), a long non-coding RNA transcribed from the antisense strand of protein coding gene AFAP1, has attracted attention in cancer research. Despite, its biological function and regulatory mechanism in hepatocellular carcinoma still unknown. The present study revealed AFAP1-AS1 mediated hepatocarcinoma progression through targeting CRKL. The bidirectional interaction of AFAP1-AS1 and oncogenic protein CRKL, and the deregulation of AFAP1-AS1 effects on Ras, MEK and c-Jun activities were investigated in depth. AFAP1-AS1 was upregulated in surgical tumorous tissues from hepatocarcinoma patients compared with the paired paracancerous non-tumor liver tissues, and in hepatocarcinoma Huh7, HCCLM3 and HepG2 cell lines compared with LO2, a normal liver cell line. AFAP1-AS1 knockdown noticeably suppressed the proliferative, migratory and invasive properties, and the epithelial-mesenchymal transition (EMT) process of HepG2 and HCCLM3 through upregulating E-cadherin and downregulating N-cadherin and vimentin. CRKL knockdown reduced AFAP1-AS1 expression levels in HepG2 and HCCLM3 cells. AFAP1-AS1 suppression impaired CRKL expression in HepG2 and HCCLM3. AFAP1-AS1 level change was positively correlated with the expression level changes of Ras, MEK and c-Jun in mediating the invasiveness of hepatocarcinoma cells. Current work demonstrated AFAP1-AS1 to be an applicable progression indicator of hepatocarcinoma. AFAP1-AS1 probably promotes the proliferation, EMT progression and metastasis of hepatocarcinoma cells via CRKL mediated Ras/MEK/c-Jun and cadherin/vimentin signaling pathways. AFAP1-AS1-CRKL bidirectional feedback signaling is worthy of further study on the monitoring, diagnosis and treatment of cancers.