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M2-polarized tumor-associated macrophages (M2-TAMs) infiltrating the tumor microenvironment contribute to hepatocellular carcinoma (HCC) progression. It was reported that cancer cells undergoing EMT will acquire stemness characteristics. Here, the HCC SMMC-7721 cell line was co-cultured with M2-TAMs polarized from THP-1 cells in vitro. In in vivo studies, we used nude mice subcutaneous tumor model to test whether the growth of the tumor was affected by M2-TAMs. Subsequently, EMT, stemness and Wnt/ß-catenin pathway related markers were detected in cells and subcutaneous tumor tissues. TNF-α was also assessed in both the co-culture system supernatants and in nude mice serum. We found that SMMC-7721 underwent EMT and acquired stemness after co-culture with M2-TAMs, and resulted in larger tumor size following subcutaneous injection of SMMC-7721 suspended in M2-TAMs supernatants compared with SMMC-7721 alone. Enzyme linked immunosorbent assay showed that TNF-α expression was elevated in supernatants of M2-TAMs and positively correlated with tumor size in the serum of nude mice. Furthermore, we found that the Wnt/ß-catenin pathway was a downstream target of TNF-α and that the Wnt/ß-catenin inhibitor ICG-001 partially reversed EMT and attenuated cancer stemness. Our results indicate that TNF-α derived from M2-TAMs promote EMT and cancer stemness cells via the Wnt/ß-catenin pathway.
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Carcinoma Hepatocelular/metabolismo , Transição Epitelial-Mesenquimal , Neoplasias Hepáticas/metabolismo , Macrófagos/metabolismo , Células-Tronco Neoplásicas/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Via de Sinalização Wnt , Animais , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Meios de Cultivo Condicionados/farmacologia , Humanos , Neoplasias Hepáticas/patologia , Camundongos , Camundongos Nus , Células-Tronco Neoplásicas/efeitos dos fármacos , Células THP-1 , Microambiente Tumoral , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
BACKGROUND: Ovarian cancer (OC) typically develops an immunosuppressive microenvironment by funtional changes of host immune cells. Dysregulated m6A level is associated with cancer progression via the intrinsic oncogenic pathways. However, the role of m6A in regulating host immune cell function during anti-tumor immunity needs comprehensive analysis. This study aimed to investigate the role of METTL3, a catalytic subunit of the methyltransferase complex, in regulating host immune cell response against OC. METHODS: In this study, myeloid-specific Mettl3 gene knockout (Mettl3-cKO) mice were bred using the Cre-LoxP system. Intraperitoneally injection of ID8 cells was used as a syngeneic OC model. Furthermore, the compositions of immune cell populations were analyzed by flow cytometry and single-cell sequencing. Moreover, chemokines and cytokines secretion were assessed using ELISA. Lastly, the role of METTL3 in regulating IL-1ß secretion and inflammasome activation in bone marrow-derived macrophages cocultured with ID8 cells was specified by ELISA and immunoblotting. RESULTS: It was revealed that OC cell growth was enhanced in Mettl3-cKO mice. Furthermore, a shift of decreased M1 to increased M2 macrophage polarization was observed during OC progression. Moreover, Mettl3 depletion in myeloid lineage cells increased secretion of CCL2 and CXCL2 in peritoneal lavage fluild. Interestingly, Mettl3 deficiency enhanced IL-1ß secretion induced by viable ID8 cells independent of inflammasome activation and cell death. Therefore, OC cells in tumor-bearing mice trigger a slight inflammatory response with a low-to-moderate secretion of pro-inflammatory cytokines and chemokines. CONCLUSION: This study provides new insights into METTL3-mediated m6A methylation, which regulates host immune response against OC.
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PURPOSE: At present, there are few studies on the mechanisms underlying postoperative recurrence of liver cancer, and the mechanism of action of miR-602 in postoperative recurrence of liver tumors is not clear. Our goals were to investigate the effects of miR-602 on the expression of the Ras-associated domain family 1A (RASSF1A) gene and the regulation of primary and recurrent hepatic tumors to clarify the molecular mechanisms of miR-602 in postoperative hepatocellular carcinoma. METHODS: We constructed a mouse liver orthotopic tumor model and a mouse liver recurrent tumor model. We measured the expression levels of the RASSF1A gene and then analyzed the effects of miR-602 on the regulation of RASSF1A. We transiently transfected the miR-602 gene into cells that stably overexpressed RASSF1A and examined relevant indicators to elucidate the mechanisms by which miR-602 regulates the RASSF1A/c-Jun N-terminal kinase (JNK) pathway in recurrence and dormancy in liver cancer. RESULTS: RASSF1A expression was inversely related to that of JNK, activating transcription factor 2 (ATF-2), and c-Jun in SMMC7721 cells stably transfected with the RASSF1A gene and in recurrent mouse tumor tissues. After transient transfection of cells with miR-602 mimic or miR-602 inhibitor, the expression of miR-602 was inversely related to that of RASSF1A. CONCLUSION: MiR-602 might inhibit the JNK signaling pathway by inhibiting the expression of RASSF1A, thereby promoting recurrence of liver cancer after surgery. The low expression levels of miR-602 in liver cancer tissues were closely related to postoperative recurrence; they could be used as a marker to judge the prognosis of patients with liver cancer.
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The traditional Chinese medicine formula Jianpi-Huayu (JPHY) has been reported to be effective in the treatment of hepatocellular carcinoma (HCC). However, its underlying mechanism remains unclear. In this article, we employed an orthotopic transplantation model in nude mice to explore whether JPHY could inhibit the development of HCC by regulating miR-602, which targets the Ras association domain-containing protein 1A (RASSF1A) pathway. HCC SMMC-7721 cells were treated with JPHY to test whether the RASSF1A gene as mediated by miR-602 affected the proliferation and apoptosis of tumor cells. We subsequently detected miR-602, RASSF1A, and tumor cell apoptosis-related markers in cells and liver tumor tissues. We observed that mice treated with JPHY had smaller tumors and higher survival rates than untreated ones. Similarly, JPHY-treated SMMC-7721 cells exhibited alterations in morphology and higher cytotoxicity compared with the control group. Furthermore, we found that JPHY decreased overexpression of miR-602 and increased protein expression levels of the RASS1A gene, which in turn altered protein expression levels of tumor cell apoptosis-related genes in the cells and liver tumor tissues of drug-treated mice. These results indicated that JPHY could potentially be used to treat HCC by targeting miR-602, which targets the RASSF1A gene, which in turn plays a major role in HCC pathogenesis.
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Carcinoma Hepatocelular , Medicamentos de Ervas Chinesas/farmacologia , MicroRNAs/genética , Transdução de Sinais/efeitos dos fármacos , Proteínas Supressoras de Tumor/genética , Animais , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/metabolismo , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/metabolismo , Camundongos , Camundongos NusRESUMO
Tumor-induced immunosuppressive microenvironment in which myeloid-derived suppressor cells (MDSCs) plays an important role, remains an obstacle for effective oncotherapy currently. Inducing MDSCs into maturation was confirmed as an effective method to reduce the tumor-bearing host's immunosuppression. Traditional Chinese medicines (TCM) possess characteristics of alleviating immunosuppression of cancer patients and low toxicity. Jianpi Huayu Decoction (JHD) was an experienced formula of TCM for oncotherapy based on TCM theory and clinical practice. We previously observed that JHD attenuated the expression of interleukin-10 (IL-10) and transforming growth factor beta (TGF-ß) in tumor. IL-10 and TGF-ß were found to be cytokines positively related to immunosuppression induced by MDSCs. Here, our study was designed to further investigate the regulation of JHD on the immune system in the H22 liver-cancer mouse model. Mainly, flow cytometry was used to detect the proportion of immune cells, to analyze the apoptosis, differentiation and reactive oxygen species of MDSCs. We found that JHD significantly reduced the destruction of spleen structure, reduced the proportion of regulatory T cells (Treg) and T helper 17 cells (Th17), and increased the proportion of cytotoxic T lymphotes (CTL), Dendritic cells (DC) and CD11b+Gr-1+cells in spleen, but with no significant change of T helper 1 cells (Th1), T helper 2 cells (Th2) and macrophages. In vitro experiments showed that apoptosis of MDSCs was decreased as the time of JHD stimulation increased, which partly explained the increase of CD11b+Gr-1+cells in the spleen. Meanwhile, JHD could promote the differentiation of MDSCs into macrophages and dendritic cells, attenuate expression of ROS in MDSCs and reduce its inhibition on the proliferation of CD4+ T cells, in vitro. Therefore, that the proportion of CD11b+Gr-1+ cells increased in the spleen of tumor-bearing hosts may not be villainy after treatment, when these drugs suppress the immunosuppressive ability of CD11b+Gr-1+ cells and promote it mature to replenish dendritic cell, at the same time. Generally, JHD may be a complementary and alternative drug for attenuating the immunosuppressive status induced by hepatocellular carcinoma, possibly by promoting differentiation and inhibiting the immunosuppressive activity of MDSCs.
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OBJECTIVE: To study the changes of pulmonary function in children with right lung middle lobe syndrome before and after treatment. METHODS: Thirty children with right lung middle lobe syndrome were classified into two age groups: < or =4 years old and >4 years old. Pulmonary function was tested by the 2600-type and the MIR-type pulmonary function spirometry in the < or =4 years and the >4 years age groups, respectively before and after treatment. Terminal flows/peak expiratory flow (25/PF) and the percentage of tidal volume to peak tidal expiratory flow (% V-PF) were measured in the <4 years age group. Forced vital capacity (FVC), forced expiratory volume in one second (FEV1) and peak expiratory flow (PEF) were measured in the >4 years age group. RESULTS: The values of 25/PF and %V-PF in the < or =4 years age group were 0.42+/-0.08 and 0.28+/-0.03, respectively before treatment. The values were improved after treatment (0.58+/-0.12 and 0.39+/-0.06 respectively) (P<0.05). The values of FVC, FEV1 and PEF were 1.75+/-0.32, 1.36+/-0.52 and 2.56+/-0.78, respectively in the >4 years age group before treatment. The values were also improved after treatment (2.37+/-0.78, 2.08+/-0.65 and 3.68+/-0.80 respectively) (P<0.05). CONCLUSIONS: There are significant differences in the pulmonary function before and after treatment in children with right lung middle lobe syndrome. The pulmonary function can return to normal after treatment.
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Pulmão/fisiopatologia , Síndrome do Lobo Médio/fisiopatologia , Criança , Pré-Escolar , Feminino , Volume Expiratório Forçado , Humanos , Lactente , Masculino , Capacidade VitalRESUMO
Oxidative stress and neuroinflammation are the key and early events during the pathological process of Parkinson's disease (PD). Thus, therapeutic intervention to regulate oxidative stress and neuroinflammation would be an effective strategy to alleviate the progression of PD. Astragaloside IV, the main active component isolated from Astragalus membranaceus, has been shown to possess anti-inflammatory and anti-oxidant properties in neurodegeneration diseases, however, the molecular mechanisms of Astragaloside IV in the pathology of PD are still unclear. In this study, we explored the mechanisms of Astragaloside IV of PD on 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced mice model and lipopolysaccharide (LPS)-induced BV2 microglia cells. Our results showed Astragaloside IV significantly alleviated behavioral impairments and dopaminergic neuron degeneration induced by MPTP. Also, Astragaloside IV inhibited microglia activation and reduced the oxidative stress of MPTP mouse model. In addition, Astragaloside IV significantly inhibited NFκB mediated NLRP3 inflammasome activation and activated Nrf2 both in vivo and in vitro. Furthermore, Astragaloside IV lessened reactive oxygen species (ROS) generation in LPS-induced BV2 microglia cells remarkably. These findings demonstrate that Astragaloside IV protects dopaminergic neuron from neuroinflammation and oxidative stress which are largely dependent upon activation of the Nrf2 pathways and suppression of NFκB/NLRP3 inflammasome signaling pathway. Therefore, Astragaloside IV is a promising neuroprotective agent that should be further developed for neurodegeneration diseases.
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Anti-Inflamatórios/uso terapêutico , Intoxicação por MPTP/tratamento farmacológico , Fármacos Neuroprotetores/uso terapêutico , Saponinas/uso terapêutico , Triterpenos/uso terapêutico , Animais , Anti-Inflamatórios/farmacologia , Linhagem Celular , Modelos Animais de Doenças , Neurônios Dopaminérgicos/efeitos dos fármacos , Inflamassomos/imunologia , Lipopolissacarídeos/farmacologia , Intoxicação por MPTP/imunologia , Intoxicação por MPTP/fisiopatologia , Masculino , Camundongos Endogâmicos C57BL , Microglia/efeitos dos fármacos , Microglia/imunologia , Atividade Motora/efeitos dos fármacos , Fator 2 Relacionado a NF-E2 , Proteína 3 que Contém Domínio de Pirina da Família NLR/antagonistas & inibidores , Proteína 3 que Contém Domínio de Pirina da Família NLR/imunologia , Fármacos Neuroprotetores/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Ratos , Saponinas/farmacologia , Triterpenos/farmacologiaRESUMO
OBJECTIVE: The efficacy of bronchodilator in asthmatoid bronchitis remains controversial. This study was designed to investigate the effects of bronchodilators, salbutamo and ipraopium bromide, on the pulmonary function in young children with this disease. METHODS: Pulmonary function tests were performed in 20 children with asthmatoid bronchitis (2 months-2.5 years of age) before and 30, 60, and 120 minutes after salbutamo and ipratropium bromide inhalation. The indexes of pulmonary function measured included tidal breathing flow volume (TBFV) loop, percent of tidal volume to peak tidal expiratory flow (%V-PF), terminal flows per peak expiratory flow (25/PF), peak tidal expiratory flow (PTEF), rate of mid-expiratory to mid-inspiratory flow (ME/MI), respiratory rate (RR) and tidal volume per kilogram (TV/kg). RESULTS: Before drug inhalation, the descending branch of the TBFV loop was depressed. The PTEF shifted forward and %V-PF (0.19 +/- 0.04) and 25/PF (0.42 +/- 0.11) decreased. These changes did not improve and the remaining indexes, RR, ME/MI and TV/kg, 30, 60, and 120 minutes after drug inhalation also remained similar to before inhalation. CONCLUSIONS: Salbutamo and ipratropium bromide inhalation did not improve the airway resistance and ventilation function in children with asthmatoid bronchitis. This suggests that the efficacy of bronchodilator in the treatment of this disease is doubtful.