RESUMO
Impaired functionality and loss of islet ß-cells are the primary abnormalities underlying the pathogenesis of both type 1 and 2 diabetes (T1DM and T2DM). However, specific therapeutic and preventive mechanisms underlying these conditions remain unclear. Mitogen-activated protein kinase phosphatase-5 (MKP-5) has been implicated in carcinogenesis, lipid metabolism regulation, and immune cell activation. In a previous study, we demonstrated the involvement of exogenous MKP-5 in the regulation of obesity-induced T2DM. However, the role of endogenous MKP-5 in the T1DM and T2DM processes is unclear. Thus, mice with MKP-5 knockout (KO) were generated and used to establish mouse models of both T1DM and T2DM. Our results showed that MKP-5 KO exacerbated diabetes-related symptoms in mice with both T1DM and T2DM. Given that most phenotypic studies on islet dysfunction have focused on mice with T2DM rather than T1DM, we specifically aimed to investigate the role of endoplasmic reticulum stress (ERS) and autophagy in T2DM KO islets. To accomplish this, we performed RNA sequence analysis to gain comprehensive insight into the molecular mechanisms associated with ERS and autophagy in T2DM KO islets. The results showed that the islets from mice with MKP-5 KO triggered 5' adenosine monophosphate-activated protein kinase (AMPK)-mediated autophagy inhibition and glucose-regulated protein 78 (GRP-78)-dominated ERS. Hence, we concluded that the autophagy impairment, resulting in islet dysfunction in mice with MKP-5 KO, is mediated through GRP-78 involvement. These findings provide valuable insights into the molecular pathogenesis of diabetes and highlight the significant role of MKP-5. Moreover, this knowledge holds promise for novel therapeutic strategies targeting MKP-5 for diabetes management.
Assuntos
Diabetes Mellitus Tipo 1 , Diabetes Mellitus Tipo 2 , Ilhotas Pancreáticas , Camundongos , Animais , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Diabetes Mellitus Tipo 1/metabolismo , Fosfatos/metabolismo , Ilhotas Pancreáticas/metabolismoRESUMO
PURPOSE: To compare the clinical characteristics, virus serotype, and outcome in cases of mild and severe enteroviral infection at a tertiary neonatal intensive care unit in China. METHODS: A retrospective analysis of cases hospitalized between June and August 2019. Samples (stool or throat swabs) were examined using reverse transcription polymerase chain reaction. Positive cases were divided into two groups: mild infection and severe infection. RESULTS: A total of 149 cases were assigned to one of two groups: mild infection (n = 104) and severe infection (n = 45). There were no significant differences between the groups in terms of sex, gestational age, birth weight, mode of delivery, and onset within 7 days. Clinical symptoms in both groups mostly resembled sepsis (fever, rash, poor feeding, and lethargy); however, there were significant variations in concomitant symptoms such as hepatitis, thrombocytopenia, encephalitis, coagulopathy, and myocarditis. Severe cases were more likely to have abnormal complete blood counts, biochemical parameters, and cerebrospinal fluid markers. The predominant serotypes implicated in neonatal enterovirus infections were echoviruses and Coxsackievirus B. Invasive ventilation, intravenous immunoglobulin, vasoactive medications, and blood product transfusions were often required, with high mortality rates among severe cases. CONCLUSION: We found significant differences between mild and severe cases of neonatal enterovirus infection with respect to complications, laboratory findings, and enterovirus serotypes. It is crucial to exercise caution when newborns exhibit symptoms of sepsis, during an enterovirus outbreak. Anemia, thrombocytopenia, abnormal liver function, and coagulation dysfunction should be monitored closely as they could indicate the presence of a severe enteroviral infection.
Assuntos
Infecções por Enterovirus , Centros de Atenção Terciária , Humanos , Infecções por Enterovirus/epidemiologia , Infecções por Enterovirus/virologia , Estudos Retrospectivos , Recém-Nascido , Masculino , Feminino , China/epidemiologia , Centros de Atenção Terciária/estatística & dados numéricos , Estudos de Casos e Controles , Enterovirus/isolamento & purificação , Enterovirus/classificação , Enterovirus/genética , Unidades de Terapia Intensiva Neonatal , Índice de Gravidade de Doença , Sorogrupo , População do Leste AsiáticoRESUMO
Dendritic cell (DC)-based immunotherapy has been a promising strategy for colon cancer therapy, but the efficacy of dendritic cell vaccines is in part limited by immunogenicity of loaded antigens. In this study, we aimed to identify a putative tumor antigen that can generate or enhance anti-tumor immune responses against colon cancer. CD44+ colon cancer stem cells (CCSCs) were isolated from mouse colorectal carcinoma CT-26 cell cultures and induced to form defective ribosomal products-containing autophagosome-rich blebs (DRibbles) by treatment with rapamycin, bortezomib, and ammonium chloride. DRibbles were characterized by western blot and transmission electron microscopy. DCs generated from the mice bone marrow monocytes were cocultured with DRibbles, then surface markers of DCs were analyzed by flow cytometry. Meanwhile, the efficacy of DRibble-DCs was examined in vivo. Our results showed that CCSC-derived DRibbles upregulated CD80, CD86, major histocompatibility complex (MHC)-I, and MHC-II on DCs and induced proliferation of mouse splenic lymphocytes and CD8+ T cells. In a model of colorectal carcinoma using BALB/c mice with robust tumor growth and mortality, DC vaccine pulsed with CCSC-derived DRibbles suppressed tumor growth and extended survival. A lactate dehydrogenase test indicated a strong cytolytic activity of cytotoxic T-cells derived from mice vaccinated with CCSC-derived DRibbles against CT-26 cells. Furthermore, flow cytometry analyses showed that the percentages of IFN-γ-producing CD8+ T-cells were increased in SD-DC group compare with the other groups. These findings provide a rationale for novel immunotherapeutic anti-tumor approaches based on DRibbles derived from colon cancer stem cells.
Assuntos
Vacinas Anticâncer/administração & dosagem , Carcinoma/terapia , Neoplasias Colorretais/terapia , Células-Tronco Neoplásicas/imunologia , Cloreto de Amônio/farmacologia , Animais , Autofagossomos/efeitos dos fármacos , Autofagossomos/imunologia , Bortezomib/farmacologia , Vacinas Anticâncer/imunologia , Carcinoma/imunologia , Carcinoma/patologia , Linhagem Celular Tumoral , Neoplasias Colorretais/imunologia , Neoplasias Colorretais/patologia , Células Dendríticas/imunologia , Modelos Animais de Doenças , Feminino , Humanos , Imunogenicidade da Vacina , Camundongos , Células-Tronco Neoplásicas/efeitos dos fármacos , Cultura Primária de Células , Sirolimo/farmacologia , Linfócitos T Citotóxicos/imunologiaRESUMO
Hyperglycemia and hyperlipidemia (glycolipotoxicity)-triggered islet ß-cell dysfunction is known to drive the progression of obesity-related type 2 diabetes, however the underlying mechanisms have not been clearly elucidated. The current study aimed to investigate the role of mitogen-activated protein kinase phosphatase 5 (MKP-5) in islet cells under glucolipotoxic conditions. Using gene overexpression and knockdown approaches, we demonstrated that MKP-5 could alleviate glucolipotoxicity-induced apoptosis via the endoplasmic reticulum (ER) stress and mitochondrial apoptosis pathways owing to the altered regulation of caspase family members and ER stress-related molecules in MIN6 and primary islet cells. Overexpression of MKP-5 reversed the glucose and palmitic acid (GP)-induced impairment of insulin secretion as well as the abnormal decreases in the expression of islet functional genes, thereby maintaining the normal insulin secretory functionality, whereas the absence of MKP-5 aggravated islet cell dysfunction. In parallel, the production of ROS and increased inflammation-associated genes in response to GP were also reduced upon MKP-5 overexpression. Further, inhibition of JNK or P38 MAPK pathways resisted to glucolipotoxicity observed in MKP-5 knockdown MIN6 cells. These findings indicate that MKP-5 is an important mediator for glucolipotoxicity-induced islet cell dysfunction and apoptosis, with JNK and P38 as the critical downstream pathways.
Assuntos
Apoptose/fisiologia , Fosfatases de Especificidade Dupla/fisiologia , Estresse do Retículo Endoplasmático/fisiologia , Glucose/toxicidade , Ilhotas Pancreáticas/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/fisiologia , Fosfatases da Proteína Quinase Ativada por Mitógeno/fisiologia , Palmitatos/toxicidade , Animais , Linhagem Celular Tumoral , Dieta Hiperlipídica/efeitos adversos , Fosfatases de Especificidade Dupla/genética , Técnicas de Silenciamento de Genes , Humanos , Insulina/metabolismo , Insulinoma/patologia , Ilhotas Pancreáticas/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mitocôndrias/metabolismo , Fosfatases da Proteína Quinase Ativada por Mitógeno/genética , Neoplasias Pancreáticas/patologia , Proteínas Recombinantes/metabolismo , Regulação para CimaRESUMO
Mitogen-activated protein kinase phosphatase-5 (MKP-5) is a regulator of extracellular signaling that is known to regulate lipid metabolism. In this study, we found that obesity caused by a high-fat diet (HFD) decreased the expression of MKP-5 in the pancreas and primary islet cells derived from mice. Then, we further investigated the role of MKP-5 in the protection of islet cells from lipotoxicity by modulating MKP-5 expression. As a critical inducer of lipotoxicity, palmitic acid (PA) was used to treat islet ß-cells. We found that MKP-5 overexpression restored PA-mediated autophagy inhibition in Rin-m5f cells and protected these cells from PA-induced apoptosis and dysfunction. Consistently, a lack of MKP-5 aggravated the adverse effects of lipotoxicity. Islet cells from HFD-fed mice were infected using recombinant adenovirus expressing MKP-5 (Ad-MKP-5), and we found that Ad-MKP-5 was able to alleviate HFD-induced apoptotic protein activation and relieve the HFD-mediated inhibition of functional proteins. Notably, HFD-mediated impairments in autophagic flux were restored by Ad-MKP-5 transduction. Furthermore, the autophagy inhibitor 3-methyladenine (3-MA) was used to treat Rin-m5f cells, confirming that the MKP-5 overexpression suppressed apoptosis, dysfunction, inflammatory response, and oxidative stress induced by PA via improving autophagic signaling. Lastly, employing c-Jun amino-terminal kinas (JNK), P38, or extracellular-regulated kinase (ERK) inhibitors, we established that the JNK and P38 MAPK pathways were involved in the MKP-5-mediated apoptosis, dysfunction, and autophagic inhibition observed in islet ß cells in response to lipotoxicity.
Assuntos
Autofagia/genética , Fosfatases de Especificidade Dupla/genética , Ilhotas Pancreáticas/enzimologia , Metabolismo dos Lipídeos/genética , Obesidade/genética , Adenina/análogos & derivados , Adenina/farmacologia , Adenoviridae/genética , Adenoviridae/metabolismo , Animais , Apoptose/efeitos dos fármacos , Apoptose/genética , Autofagia/efeitos dos fármacos , Dieta Hiperlipídica/efeitos adversos , Fosfatases de Especificidade Dupla/metabolismo , Regulação da Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Ilhotas Pancreáticas/citologia , Ilhotas Pancreáticas/efeitos dos fármacos , Proteínas Quinases JNK Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases JNK Ativadas por Mitógeno/genética , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Metabolismo dos Lipídeos/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteína Quinase 1 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 1 Ativada por Mitógeno/genética , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 3 Ativada por Mitógeno/genética , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Obesidade/enzimologia , Obesidade/etiologia , Obesidade/patologia , Ácido Palmítico/antagonistas & inibidores , Ácido Palmítico/toxicidade , Cultura Primária de Células , Inibidores de Proteínas Quinases/farmacologia , Transdução de Sinais , Transdução Genética , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
Researchers have shown that long noncoding RNAs (lncRNAs) are closely associated with the pathogenesis of colorectal cancer (CRC). In here, we aimed to explore the function of lncRNA MAFG-AS1 in tumorigenesis of CRC. Firstly, we found that the expression of MAFG-AS1 was upregulated in CRC tissues and positively correlated with the advanced tumor stage. A reciprocal repression was found between MAFG-AS1 and miR-147b. The expression of miR-147b was downregulated in CRC tissues and inversely correlated with MAFG-AS1. Both the low-expression of miR-147b expression and the advanced tumor stage were independent factor for poor survival probability. Furthermore, overexpression of MAFG-AS1 promoted cell proliferation, cell cycle progression, and invasion, and inhibited apoptosis, while transduction of miR-147b partially reversed the effect of MAFG-AS1 on cellular processes. Consistently, stable over-expression of MAFG-AS1 contributed to the growth of colon cancer cell xenografts in vivo. NDUFA4 was identified as a direct target of miR-147b and knockdown of NDUFA4 abolished the oncogenic role of miR-147b inhibitor. Besides, MAFG-AS1 contributed to cell glycolysis by sponging miR-147b and activation of NDUFA4, causing an upregulation of PDK1, PFK1 and PKM2. Taken together, our study suggested that MAFG-AS1 functions as a novel oncogenic lncRNA in the development of CRC by regulating miR-147b/NDUFA4.
Assuntos
Neoplasias Colorretais/patologia , Progressão da Doença , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Fator de Transcrição MafG/genética , MicroRNAs/antagonistas & inibidores , RNA Longo não Codificante/fisiologia , Proteínas Repressoras/genética , Animais , Apoptose , Ciclo Celular , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Complexo IV da Cadeia de Transporte de Elétrons/antagonistas & inibidores , Glicólise , Xenoenxertos , Humanos , Camundongos , MicroRNAs/fisiologiaRESUMO
NK cells hold promise for protecting hosts from cancer and pathogen infection through direct killing and expressing immune-regulatory cytokines. In our study, a genetically modified K562 cell line with surface expression of 4-1BBL and MICA was constructed to expand functional NK cells in vitro for further adoptive immunotherapy against cancer. After a long-term up to 21 day co-culture with newly isolated peripheral blood mononuclear cells (PBMCs) in the presence of soluble IL-21 (sIL-21), notable increase in proportion of expanded NK cells was observed, especially the CD56(bright)CD16(+) subset. Apparent up-regulation of activating receptors CD38, CD69 and NKG2D was detected on expanded NK cells, so did inhibitory receptor CD94; the cytotoxicity of expanded NK cells against target tumor cells exceeded that of NK cells within fresh PBMCs. The intracellular staining showed expanded NK cells produced immune-regulatory IFN-γ. Taken together, we expanded NK cells with significant up-regulation of activating NKG2D and moderate enhancement of cytotoxicity, with IFN-γ producing ability and a more heterogeneous population of NK cells. These findings provide a novel perspective on expanding NK cells in vitro for further biology study and adoptive immunotherapy of NK cells against cancer.
Assuntos
Ligante 4-1BB/biossíntese , Antígenos de Histocompatibilidade Classe I/biossíntese , Interleucinas/biossíntese , Células Matadoras Naturais/imunologia , Neoplasias/imunologia , Ligante 4-1BB/genética , ADP-Ribosil Ciclase 1/biossíntese , Antígenos CD/biossíntese , Antígenos de Diferenciação de Linfócitos T/biossíntese , Antígeno CD56/biossíntese , Linhagem Celular Tumoral , Técnicas de Cocultura , Proteínas Ligadas por GPI/biossíntese , Células HeLa , Células Hep G2 , Antígenos de Histocompatibilidade Classe I/genética , Humanos , Imunoterapia Adotiva , Interferon gama/biossíntese , Interferon gama/imunologia , Interleucinas/genética , Interleucinas/farmacologia , Lectinas Tipo C/biossíntese , Ativação Linfocitária/imunologia , Glicoproteínas de Membrana/biossíntese , Subfamília D de Receptores Semelhantes a Lectina de Células NK/biossíntese , Subfamília K de Receptores Semelhantes a Lectina de Células NK/biossíntese , Neoplasias/terapia , Receptores de IgG/biossínteseRESUMO
OBJECTIVE: To explore the role of PD-L1/PD-1 blockage in the cytotoxicity of natural killer cell in NSCLC. METHODS: Two NSCLC cell lines, Calu-1 and H460, were tested for susceptibility to the cytolytic activity of freshly isolated healthy donor NK cells by a non-radioactive cellular cytotoxicity assay kit. Western blot analysis, FACS, ELISA and antibody blockage experiments were conducted to determine the mechanisms. NK cells isolated from NSCLC patients were also collected for functional assays. RESULTS: Calu-1 and H460 cells were lysed by NK cells in a dose-dependent manner. H460 cells showed less susceptibility to NK cell-mediated lysis than Calu-1 cells at all ratios. The expression of PD-L1 on H460 cells was higher than that on Calu-1 cells, as determined by FACS and western blot analysis. The specific lysis of H460 cells by NK cells was enhanced when the PD-L1/PD-1 interaction was blocked by anti-PD-L1 antibody. This finding was also demonstrated in NK cells isolated from NSCLC patients. CONCLUSIONS: The present study revealed that PD-L1/PD-1 blockage enhanced the cytotoxicity of natural killer cells in NSCLC via granzyme B secretion. This study will greatly facilitate the precise treatment of lung cancer through determination of PD-L1 expression in tumors.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Humanos , Carcinoma Pulmonar de Células não Pequenas/patologia , Neoplasias Pulmonares/patologia , Receptor de Morte Celular Programada 1/metabolismo , Granzimas/metabolismo , Linhagem Celular Tumoral , Células Matadoras NaturaisRESUMO
Because the demand for rabies post exposure prophylaxis (PEP) treatment has increased exponentially in recent years, the limited supply of human and equine rabies immunoglobulin (HRIG and ERIG) has failed to provide an adequate amount of the required passive immune component in PEP in countries where canine rabies is endemic. The replacement of HRIG and ERIG with a potentially cheaper and efficacious alternative biological for the treatment of rabies in humans, therefore, remains a high priority. In this study, we set out to assess a human single-chain Fv antibody fragment fused with the Fc of an IgG1 targeting a rabies antigen to develop a product that can be used as a component of the PEP cocktail. We cloned the ScFv fragment from a human ScFv library that was established previously and inserted this fragment into the expression vector pPICZαC/Fc. An active recombinant ScFv-Fc fusion protein was successfully expressed in Pichia pastoris. The production of ScFv-Fc was optimized and scaled up in an 80L fermentor with yields exceeding 60mg/L. The ScFv-Fc protein was purified to more than 95% purity using a two-step scheme: ammonium sulfate fractionation and Protein A Sepharose CL-4B. The ScFv-Fc fusion protein neutralized rabies virus in a standard in vivo neutralization assay in which the virus was incubated with the ScFv-Fc molecules before intracranial inoculation in mice. Our results suggest that functional antibodies can be produced in P. pastoris and that ScFv-Fc fusion proteins have the potential to serve as therapeutic candidates.
Assuntos
Anticorpos Antivirais/imunologia , Antígenos Virais/imunologia , Fragmentos Fc das Imunoglobulinas/imunologia , Imunoglobulina G/imunologia , Vírus da Raiva/imunologia , Anticorpos de Cadeia Única/isolamento & purificação , Animais , Anticorpos Neutralizantes/imunologia , Anticorpos Neutralizantes/metabolismo , Western Blotting , Eletroforese em Gel de Poliacrilamida , Vetores Genéticos/genética , Humanos , Fragmentos Fc das Imunoglobulinas/genética , Dose Letal Mediana , Camundongos , Testes de Neutralização , Pichia/genética , Pichia/metabolismo , Plasmídeos/genética , Vírus da Raiva/genética , Vírus da Raiva/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/imunologia , Proteínas Recombinantes de Fusão/isolamento & purificação , Anticorpos de Cadeia Única/genética , Anticorpos de Cadeia Única/imunologiaRESUMO
P53 is an attractive target in molecular cancer therapeutics because of its critical role in regulating cell cycle arrest and apoptosis. The limitations in the development of p53-based cancer therapeutic strategy include its inefficient transmission through cell membrane of tumor cells and low protein yields in the expression system. In the present study, p53 was fused with HIV TAT protein, which can cross cell membranes, and expressed by Pichia pastoris. Stable production of Tat-p53 was achieved. After being transduced with Tat-p53 protein, the growth of cancer cell line, HepG2, was inhibited by increased apoptosis in culture. This expression system could thus be utilized to produce human Tat-p53 fusion protein.
Assuntos
Apoptose , Hepatócitos/efeitos dos fármacos , Hepatócitos/fisiologia , Proteína Supressora de Tumor p53/isolamento & purificação , Produtos do Gene tat do Vírus da Imunodeficiência Humana/isolamento & purificação , Expressão Gênica , Células Hep G2 , Humanos , Pichia/genética , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/isolamento & purificação , Transdução Genética , Proteína Supressora de Tumor p53/biossíntese , Proteína Supressora de Tumor p53/genética , Produtos do Gene tat do Vírus da Imunodeficiência Humana/biossíntese , Produtos do Gene tat do Vírus da Imunodeficiência Humana/genéticaRESUMO
OBJECTIVE: To explore the cytotoxic responses of spleen T lymphocytes (CTL) in BALB/c mice induced by recombinant HSP110-HER2/neu ICD complex. METHODS: Tumor-bearing mouse model was immunized by HSP110-HER2/neu ICD complex. The IFN-γ level secreted by activated spleen T lymphocytes was detected by enzyme linked immunospot assay (ELISPOT). The corresponding CTL activity was measured by granzyme release assay. RESULTS: The BALB/c mouse model of human mammary tumor highly expressing HER2/neu was established. HSP110-HER2/neu ICD complex immunization led to a significantly higher level of INF-γ than that in HSP110-P(789-797) immunized and HER2/neu ICD immunized mice. HSP110-HER2/neu ICD complex immunized animals also show significant CTL activity. The results of immunohistochemical staining showed that the number of blue spots in the PBS group was 4.57 ± 1.33, HSP110 group 6.83 ± 2.08, HER2/neu ICD group 16.17 ± 2.86, HSP110-P(789-797) group 43.67 ± 4.78, and SP110-HER2/neu ICD group 76.51 ± 8.17. The number of IFN-γ-secreting spleen lymphocytes in the HSP110-HER2/neu ICD group was significantly higher than that in the HSP110-P(789-797) group, and that of HSP110-P(789-797) group was significantly higher than that of HER2/neu ICD group (P < 0.01). The target cell-killing rate of the PBS group was (8.15 ± 1.27)%, HSP110 group (9.51 ± 1.51)%, HER2/neu ICD group (14.03 ± 2.45)%, HSP110-P(789-797) group (25.99 ± 3.04)% and HSP110-HER2/neu ICD group (38.15 ± 3.95)% (all P < 0.01). CONCLUSIONS: HSP110-HER2/neu ICD complex can promote the proliferation and maturation of T lymphocytes into CTLs, and might be used as anti-tumor vaccine to induce potent cytotoxic T lymophocyte immunoresponse against specific tumor cells.
Assuntos
Neoplasias da Mama/patologia , Proteínas de Choque Térmico HSP110/imunologia , Receptor ErbB-2/imunologia , Linfócitos T Citotóxicos/imunologia , Linfócitos T/citologia , Animais , Neoplasias da Mama/metabolismo , Vacinas Anticâncer/imunologia , Linhagem Celular Tumoral , Proliferação de Células , Feminino , Humanos , Interferon gama/metabolismo , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos BALB C , Transplante de Neoplasias , Distribuição Aleatória , Receptor ErbB-2/metabolismo , Proteínas Recombinantes/imunologia , Baço/citologia , Baço/imunologia , Linfócitos T/imunologia , Linfócitos T/metabolismo , Vacinas Sintéticas/imunologiaRESUMO
The mononuclear phagocyte system (MPS) consists of monocytes, dendritic cells and macrophages, which play vital roles in innate immune defense against cancer. Hepatocellular carcinoma (HCC) is a complex disease that is affected or initiated by many factors, including chronic hepatitis B virus infection, hepatitis C virus infection, metabolic disorders or alcohol consumption. Liver function, tumor stage and the performance status of patients affect HCC clinical outcomes. Studies have shown that targeted treatment of tumor microenvironment disorders may improve the efficacy of HCC treatments. Cytokines derived from the innate immune response can regulate T-cell differentiation, thereby shaping adaptive immunity, which is associated with the prognosis of HCC. Therefore, it is important to elucidate the function of the MPS in the progression of HCC. In this review, we outline the impact of HCC on the MPS. We illustrate how HCC reshapes MPS cell phenotype remodeling and the production of associated cytokines and characterize the function and impairment of the MPS in HCC.
Assuntos
Carcinoma Hepatocelular , Hepatite B Crônica , Neoplasias Hepáticas , Humanos , Carcinoma Hepatocelular/patologia , Neoplasias Hepáticas/patologia , Hepatite B Crônica/complicações , Sistema Fagocitário Mononuclear , Citocinas , Microambiente TumoralRESUMO
Multifunctionality has becoming essential for bone tissue engineering materials, such as drug release. In this study, icariin (ICA)-incorporated poly(glycolide-co-caprolactone) (PGCL) porous microcarriers were fabricated and then coated with decellularized extracellular matrix (dECM) which was derived from bone marrow mesenchymal stem cells (BMSC). The porous structure was generated due to the soluble gelatin within the microcarriers. The initial released ICA in microcarriers regulated osteogenic ECM production by BMSCs during ECM formation. The dECM could further synergistically enhance the migration and osteogenic differentiation of BMSCs together with ICA as indicated by the transwell migration assay, ALP and ARS staining, as well as gene and protein expression. Furthermore, in vivo results also showed that dECM and ICA exhibited excellent synergistic effects in repairing rat calvarial defects. These findings suggest that the porous microcarriers loaded with ICA and dECM coatings have great potential in the field of bone tissue engineering.
RESUMO
With significant high incidence and death rates, liver cancer has become one of the most common cancers all over the world. Hence, novel strategies are needed for the management of this malignancy. Apoptotic related proteins Noxa and Puma are the members of BH3-only family. In this study, human Noxa or Puma coding sequences have been inserted into plasmid pcDNA 3.1 regulated by human TERT promoter. The transfection of HepG2 cells with pcTERT-Noxa or pcTET-Puma resulted in the significant suppression of cell proliferation as well as finally led to apoptosis via mitochondrial and death receptor pathways, and also exhibited significantly reduced the ability of invasion and metastasis. Moreover, an in vivo study revealed that intratumoral injections of pcTERT-Noxa or pcTERT-Puma plasmids effectively suppressed the tumor growth and can exhibit anti-neoplastic effects by recruiting CD3, CD8, CD45 positive T lymphocytes in the tumor tissues. Overall, our findings illustrated that pcTERT-Noxa and pcTERT-Puma may exhibit significant anti-tumor effects both in vivo and in vivo.
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Apolipoprotein C-I (ApoC-I) is a small, basic apolipoprotein which is mainly secreted by the liver as a component of triglyceride-rich lipoproteins and high density lipoproteins whose importance in plasma lipoprotein metabolism is increasingly evident. At present, the only way to obtain native ApoC-I is separating it from human plasma. The methods have some restrictions on source, the complicated technology, the potential infections and a high cost which limits the research and application of native ApoC-I. Because of its small size, ApoC-I has previously been prepared by peptide synthesis which is also limited by a high cost. Therefore, in this study, a Pichia pastoris expression system was first used to obtain a high level expression of secreted, recombinant human ApoC-I (rhApoC-I).
Assuntos
Apolipoproteína C-I/biossíntese , Apolipoproteína C-I/isolamento & purificação , Pichia/genética , Proteínas Recombinantes/isolamento & purificação , Animais , Apolipoproteína C-I/genética , Apolipoproteína C-I/farmacologia , Biotecnologia/métodos , Western Blotting , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Eletroforese em Gel de Poliacrilamida , Ensaio de Imunoadsorção Enzimática , Humanos , Concentração de Íons de Hidrogênio , Miócitos de Músculo Liso , Pichia/química , Pichia/metabolismo , Ratos , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/genética , Proteínas Recombinantes/farmacologiaRESUMO
Ginseng, a perennial plant belonging to genus Panax, has been widely used in traditional herbal medicine in East Asia and North America. Ginsenosides are the most important pharmacological component of ginseng. Variabilities in attached positions, inner and outer residues and types of sugar moieties may be associated with the specific pharmacological activities of each ginsenoside. Ginsenoside Rg5 (Rg5) is a minor ginsenoside synthesized during ginseng steaming treatment that exhibits superior pharmaceutical activity compared with major ginsenosides. With high safety and various biological functions, Rg5 may act as a potential therapeutic candidate for diverse diseases. To date, there have been no systematic studies on the activity of Rg5. Therefore, in this review, all available literature was reviewed and discussed to facilitate further research on Rg5.
RESUMO
Following the publication of this paper, it was drawn to the Editors' attention by a concerned reader that certain of the flow cytometric and western blotting data shown in Fig. 3A and C respectively, and the tumor images shown in Fig. 7A, bore unexpected similarities to data appearing in different form in other articles by different authors. Owing to the fact that some of the contentious data in the above article had already been published elsewhere, or were already under consideration for publication, prior to its submission to Oncology Reports, the Editor has decided that this paper should be retracted from the Journal. After having been in contact with the authors, they agreed with the decision to retract the paper. The Editor apologizes to the readership for any inconvenience caused. [the original article was published in Oncology Reports 33: 448-456, 2015; DOI: 10.3892/or.2014.3591].
RESUMO
Esophageal squamous cell carcinoma accounts for a large proportion of cancer-associated mortalities in both men and women. Melittin is the major active component of bee venom, which has been reported to possess anti-inflammatory, antibacterial and anti-cancer properties. The aim of the present study was to construct a tumor targeted recombinant plasmid [pc-telomerase reverse transcriptase (TERT)-melittin] containing a human TERT promoter followed by a melittin coding sequence and to explore the effects of this plasmid in esophageal cell carcinoma and investigate preliminarily the underlying mechanisms of this effect. TE1 cells were transfected with pcTERT-melittin and the resulting apoptosis was subsequently examined. The viability of TE1 cells transfected with pcTERT-melittin was measured using a Cell Counting Kit-8 assay, which indicated inhibited proliferation. The disruption of mitochondrial membranes and the concomitant production of reactive oxygen species demonstrated an inducible apoptotic effect of melittin in TE1 cells. Apoptotic cells were also counted using an Annexin V-FITC and PI double-staining assay. The upregulation of cleaved caspase-9, cleaved caspase-3, Bax and poly(ADP-ribose) polymerase 1 in pcTERT-melittin transfected TE1 cells, suggested that pcTERT-melittin-induced apoptosis was associated with the mitochondrial pathway. TE1 cells were also arrested in the G0/G1 phase when transfected with pcTERT-melittin, followed by the decline of CDK4, CDK6 and cyclin D1 expression levels. As cell invasion and metastasis are common in patients with esophageal cancer, a cell migration assay was conducted and it was found that pcTERT-melittin transfection reduced the migratory and invasive abilities of TE1 cells. The findings of the present study demonstrated that pcTERT-melittin may induce apoptosis of esophageal carcinoma cells and inhibit tumor metastasis.
RESUMO
OBJECTIVE: In obese individuals, chronic low-grade inflammation resulting from adipocyte-macrophage interactions is a major cause of adipose tissue dysfunction and metabolic disease. This study investigated the role of MAP kinase phosphatase-5 (MKP-5) in obesity-induced inflammation during macrophage and adipocyte interactions. METHODS: High-fat diet-induced obese mice were used to explore the role of MKP-5 in obesity-induced adipose tissue inflammation. Macrophage polarization was determined by inflammatory cytokine expression in MKP-5-overexpressed or -silenced Raw264.7 cells exposed to palmitate (PA) or M1/M2 macrophage inducers. To uncover the role of MKP-5 during macrophage-adipocyte interactions, a coculture system composed of differentiated 3T3-L1 and Raw264.7 cells was employed. MAPK inhibitors were used to investigate the involvement of MAPK signaling. RESULTS: Increased MKP-5 expression was observed in adipose stromal vascular cells (SVCs) of obese mice. In Raw264.7 cells, MKP-5 promoted the switching of M1 macrophages to an M2 phenotype. Notably, MKP-5 reduced inflammation during the interaction of macrophages and adipocytes. MKP-5 overexpression in primary SVCs attenuated the expression of inflammatory mediators and increased the number of obesity-induced adipose tissue macrophages. MKP-5 suppressed PA-induced inflammation through the inactivation of P38, JNK, and ERK MAPKs. CONCLUSIONS: MKP-5 promotes macrophages to switch from the M1 to the M2 phenotype and is an inflammatory inhibitor involved in obesity-induced adipose tissue inflammation and PA-triggered macrophage inflammation via the P38, JNK, and ERK MAPK pathways. MKP-5 may be developed into a potential therapeutic target for obesity-related diseases, including type 2 diabetes mellitus and insulin resistance.
Assuntos
Adipócitos/fisiologia , Comunicação Celular/genética , Fosfatases de Especificidade Dupla/fisiologia , Macrófagos/fisiologia , Obesidade/patologia , Células 3T3-L1 , Tecido Adiposo/patologia , Animais , Técnicas de Cocultura , Dieta Hiperlipídica , Fosfatases de Especificidade Dupla/genética , Células HEK293 , Humanos , Resistência à Insulina/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Obesos , Obesidade/etiologia , Obesidade/genética , Células RAW 264.7RESUMO
Oncostatin M (OSM) is a multifunctional cellular regulator that belongs to the IL-6 subfamily and can act on a wide variety of cells, which has potential roles in the regulation of gene activation, cell survival, proliferation and differentiation. In order to achieve the higher level yield of recombinant human Oncostatin M (rhOSM), we determined the optimal pH condition of rhOSM expressed in the methylotrophic yeast Pichia pastoris X-33 and carried out the fermentation culture of rhOSM in 80 L fermentor in a fed-batch mode. SDS-PAGE and Western blotting assays demonstrated that rhOSM was successfully expressed and secreted into the culture medium with an apparent molecular weight of 28 kDa. N-terminals were correctly processed through amino-terminal sequencing. The maximum yield of rhOSM was 280 mg/L. rhOSM was purified by phenyl Sepharose hydrophobic interaction chromatography and SP Sepharose Fast Flow cation exchange chromatography, which resulted in a final yield of purified rhOSM of 6.94 g with a recovery of 62% and a purity of 95%. The purified rhOSM had a specific growth inhibition activity of 6.26 x 10(4)RU/microg, which was commensurate with typical values (6.2 x 10(4)RU/microg) obtained with standard hOSM.