RESUMO
Activating mutations in the KEAP1/NRF2 pathway characterize a subset of non-small cell lung cancer (NSCLC) associated with chemoresistance and poor prognosis. We herein evaluated the relationship between 64 oxidative stress-related genes and overall survival data from 35 lung cancer datasets. Thioredoxin reductase-1 (TXNRD1) stood out as the most significant predictor of poor outcome. In a cohort of NSCLC patients, high TXNRD1 protein levels correlated with shorter disease-free survival and distal metastasis-free survival post-surgery, including a subset of individuals treated with platinum-based adjuvant chemotherapy. Bioinformatics analysis revealed that NSCLC tumors harboring genetic alterations in the NRF2 pathway (KEAP1, NFE2L2 and CUL3 mutations, and NFE2L2 amplification) overexpress TXNRD1, while no association with EGFR, KRAS, TP53 and PIK3CA mutations was found. In addition, nuclear accumulation of NRF2 overlapped with upregulated TXNRD1 protein in NSCLC tumors. Functional cell assays and gene dependency analysis revealed that NRF2, but not TXNRD1, has a pivotal role in KEAP1 mutant cells' survival. KEAP1 mutants overexpress TXNRD1 and are less susceptible to the cytotoxic effects of the TXNRD1 inhibitor auranofin when compared to wild-type cell lines. Inhibition of NRF2 with siRNA or ML-385, and glutathione depletion with buthionine-sulfoximine, sensitized KEAP1 mutant A549 cells to auranofin. NRF2 knockdown and GSH depletion also augmented cisplatin cytotoxicity in A549 cells, whereas auranofin had no effect. In summary, these findings suggest that TXNRD1 is not a key determinant of malignant phenotypes in KEAP1 mutant cells, although this protein can be a surrogate marker of NRF2 pathway activation, predicting tumor recurrence and possibly other aggressive phenotypes associated with NRF2 hyperactivation in NSCLC.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Tiorredoxina Redutase 1 , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Proteínas Culina , Humanos , Proteína 1 Associada a ECH Semelhante a Kelch/genética , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Recidiva Local de Neoplasia/genética , Transdução de Sinais , Tiorredoxina Redutase 1/genética , Tiorredoxina Redutase 1/metabolismoRESUMO
Obesity is a major risk factor for breast cancer, especially in post-menopausal women. In the breast tissue of obese women, cyclooxygenase-2 (COX-2)-dependent prostaglandin E2 (PGE2) production has been correlated with inflammation and local estrogen biosynthesis via aromatase. Using a mouse model of 7,12-dimethylbenz[a]anthracene/medroxyprogesterone-acetate (DMBA/MPA)-induced carcinogenesis, we demonstrated that an obesogenic diet promotes mammary tissue inflammation and local estrogen production, and accelerates mammary tumor formation in a COX-2-dependent manner. High-sugar/fat (HSF) diet augmented the levels of the pro-inflammatory mediators MCP-1, IL-6, COX-2, and PGE2 in mammary tissue, and this was accompanied by crown-like structures of breast (CLS-B) formation and aromatase/estrogen upregulation. Treatment with a COX-2 selective inhibitor, etoricoxib, decreased PGE2, IL-6, MCP-1, and CLS-B formation as well as reduced aromatase protein and estrogen levels in the mammary tissue of mice fed a HSF diet. Etoricoxib-treated mice showed increased latency and decreased incidence of mammary tumors, which resulted in prolonged animal survival when compared to HSF diet alone. Inhibition of tumor angiogenesis also seemed to account for the prolonged survival of COX-2 inhibitor-treated animals. In conclusion, obesogenic diet-induced COX-2 is sufficient to trigger inflammation, local estrogen biosynthesis, and mammary tumorigenesis.
Assuntos
Neoplasias da Mama/metabolismo , Ciclo-Oxigenase 2/metabolismo , Dieta Hiperlipídica/efeitos adversos , Dinoprostona/biossíntese , Açúcares/efeitos adversos , Regulação para Cima , 9,10-Dimetil-1,2-benzantraceno/efeitos adversos , Animais , Aromatase/metabolismo , Neoplasias da Mama/induzido quimicamente , Neoplasias da Mama/tratamento farmacológico , Linhagem Celular Tumoral , Quimiocina CCL2/metabolismo , Modelos Animais de Doenças , Etoricoxib/administração & dosagem , Etoricoxib/farmacologia , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Interleucina-6/metabolismo , Células MCF-7 , Acetato de Medroxiprogesterona/efeitos adversos , CamundongosRESUMO
Canine obesity is associated with genetic, environmental, and behavioural factors, with the latter including both the behaviour of the dog and the owner. Knowledge about owner perception of canine obesity and its treatment can inform the development of new strategies to help prevent and manage this disease. Therefore, the aim of this study was to evaluate the opinions of dog owners regarding canine obesity and weight management. Dog owners residing in the city of Sao Paulo (Brazil) completed a questionnaire, either at home or in the waiting rooms of 3 veterinary hospitals. Owners determined their dog's body condition score (BCS), and this was compared with BCS determined by a veterinarian. Questionnaire findings from dogs that were in overweight (BCS 6-7/9) or obese (BCS (8-9/9) condition were compared with those in ideal weight (4-5/9) using chi-square tests and odds ratios. A total of 926 dogs were included, of which 480 (52%), 317 (34%) and 129 (14%) were in ideal, overweight and obese condition, respectively. Many owners under-estimated their dog's weight status, with the proportion increasing as the dog's weight status increased (ideal 60/480, 13%; overweight 174/317, 55%; obese 88/129, 68%; P<0.001). Although most owners (890/926, 96%) believed that canine obesity could pose health risks, the proportion that disagreed increased as weight status increased (ideal 12/480, 2%; overweight 14/317, 4%; 10/129, 8%; P = 0.006). Finally, although most owners (880/926, 95%) stated that they would let their dog undergo weight management, only a minority (182/926; 20%) believed that a trained professional was needed, and they had various misperceptions including potential cost and what the strategies that would be effective. Based on the findings of this study, it would be advisable for veterinarians to spend time addressing these misperceptions, in the hope of both improving awareness of obesity and the outcomes of weight management.
Assuntos
Obesidade/veterinária , Inquéritos e Questionários , Animais , Atitude , Brasil , Doenças do Cão/epidemiologia , Cães , Feminino , Humanos , Masculino , Obesidade/epidemiologia , Sobrepeso/veterinária , Condicionamento Físico Animal/estatística & dados numéricos , Fatores de Risco , Inquéritos e Questionários/estatística & dados numéricos , Programas de Redução de PesoRESUMO
Canine obesity is associated with comorbidities, a shortened lifespan, and a poorer quality of life, but epidemiological studies characterizing canine obesity in Latin America are scarce. Therefore, this study aimed to determine the prevalence of canine obesity in the city of Sao Paulo, Brazil, and the possible associated causal factors. Randomly-selected households from different city regions were visited. Dogs in each household were evaluated and owners completed a questionnaire whilst their anthropometric measures were taken. Total of 285 dogs from 221 owners were included, and the combined prevalence of overweight and obesity was 40.5%. The prevalence of overweight and obesity was greater in female dogs (P = 0.003) and in dogs that were neutered (P = 0.001). There was also a positive association between BCS and frequency of visits to a veterinarian (P = 0.026), feeding frequency (P = 0.033), and higher snack intake (P = 0.011). Further, the BCS of dogs was greater when their owners reported consuming more snacks themselves (P = 0.005) and whose had a presence of elderly people in the household (P = 0.006). In conclusion, the prevalence of obesity found in a Brazilian metropolitan region was similar to that if other countries, and neutering and snack intake were associated with the development of this disease.
Assuntos
Doenças do Cão/epidemiologia , Sobrepeso/veterinária , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Ração Animal , Animais , Brasil/epidemiologia , Castração , Estudos Transversais , Dieta , Cães , Características da Família , Feminino , Hábitos , Humanos , Masculino , Pessoa de Meia-Idade , Sobrepeso/epidemiologia , Prevalência , Fatores de Risco , Estudos de Amostragem , Lanches , Fatores Socioeconômicos , Cobertura Vacinal , Adulto JovemRESUMO
BACKGROUND: The impact of exposure to multiple pathogens concurrently or consecutively on immune function is unclear. Here, immune responses induced by combinations of the bacterium Salmonella Typhimurium (STm) and the helminth Nippostrongylus brasiliensis (Nb), which causes a murine hookworm infection and an experimental porin protein vaccine against STm, were examined. METHODOLOGY/PRINCIPAL FINDINGS: Mice infected with both STm and Nb induced similar numbers of Th1 and Th2 lymphocytes compared with singly infected mice, as determined by flow cytometry, although lower levels of secreted Th2, but not Th1 cytokines were detected by ELISA after re-stimulation of splenocytes. Furthermore, the density of FoxP3+ T cells in the T zone of co-infected mice was lower compared to mice that only received Nb, but was greater than those that received STm. This reflected the intermediate levels of IL-10 detected from splenocytes. Co-infection compromised clearance of both pathogens, with worms still detectable in mice weeks after they were cleared in the control group. Despite altered control of bacterial and helminth colonization in co-infected mice, robust extrafollicular Th1 and Th2-reflecting immunoglobulin-switching profiles were detected, with IgG2a, IgG1 and IgE plasma cells all detected in parallel. Whilst extrafollicular antibody responses were maintained in the first weeks after co-infection, the GC response was less than that in mice infected with Nb only. Nb infection resulted in some abrogation of the longer-term development of anti-STm IgG responses. This suggested that prior Nb infection may modulate the induction of protective antibody responses to vaccination. To assess this we immunized mice with porins, which confer protection in an antibody-dependent manner, before challenging with STm. Mice that had resolved a Nb infection prior to immunization induced less anti-porin IgG and had compromised protection against infection. CONCLUSION: These findings demonstrate that co-infection can radically alter the development of protective immunity during natural infection and in response to immunization.
Assuntos
Nippostrongylus/imunologia , Vacinas contra Salmonella/imunologia , Salmonella typhimurium/imunologia , Infecções por Strongylida/imunologia , Animais , Anticorpos Antibacterianos/sangue , Anticorpos Anti-Helmínticos/sangue , Coinfecção/imunologia , Citocinas/biossíntese , Imunização , Switching de Imunoglobulina , Imunoglobulina G/sangue , Camundongos , Camundongos Endogâmicos BALB C , Linfócitos T/imunologiaRESUMO
The effects of Leishmania mexicana metacyclic promastigotes upon MAP kinase signalling in mouse bone marrow macrophages and subsequent expression of the disease regulatory proteins iNOS and COX-2 were studied. At a ratio of 5:1, promastigotes caused a marked increase in phosphorylation of the three major MAP kinases, ERK, p38 and JNK. MAP kinase signalling was substantially reduced in TLR-4(-/-) but not TLR-2(-/-) deficient macrophages and completely abolished in double TLR-2/4(-/-) macrophages. A similar outcome was observed using cysteine peptidase B deficient amastigotes. Furthermore, whilst promastigotes had no independent effect on iNOS or COX-2 expression, they prolonged the induction of these proteins stimulated by LPS and enhanced PGE(2) and NO production. Induction of COX-2 and iNOS was also TLR-4 dependent. Blockade of either PGE(2) or NO production with indomethacin or l-NAME reversed promastigote inhibition of LPS induced IL-12 production. Promastigotes also increased macrophage arginase-1 expression and enhanced arginase activity, both of which were substantially reduced in TLR-4 but not TLR-2 deficient macrophages. Surprisingly, arginase inhibition by Nor-NOHA also caused a reversal of promastigote mediated inhibition of macrophage IL-12 production. These data demonstrate for the first time the role of TLR-4 in mediating the effects of L. mexicana promastigotes on MAP kinase activation, up-regulation of COX-2, iNOS as well as arginase-1 expression in macrophages and further shows that PGE(2), NO and arginase activity all contribute substantially to the inhibition of host cell IL-12 production.
Assuntos
Ativação Enzimática/imunologia , Interleucina-12/biossíntese , Leishmaniose/imunologia , Macrófagos/imunologia , Receptor 4 Toll-Like/metabolismo , Animais , Arginase/biossíntese , Arginase/imunologia , Western Blotting , Ciclo-Oxigenase 2/biossíntese , Ciclo-Oxigenase 2/imunologia , MAP Quinases Reguladas por Sinal Extracelular/imunologia , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Interleucina-12/imunologia , Leishmania mexicana/imunologia , Leishmaniose/metabolismo , Macrófagos/metabolismo , Macrófagos/microbiologia , Camundongos , Óxido Nítrico Sintase Tipo II/biossíntese , Óxido Nítrico Sintase Tipo II/imunologia , Transdução de Sinais/imunologia , Receptor 4 Toll-Like/imunologiaRESUMO
Immunologically intact BALB/c mice infected with Leishmania mexicana develop non-healing progressively growing lesions associated with a biased Th2 response while similarly infected IL-4Rα-deficient mice fail to develop lesions and develop a robust Th1 response. In order to determine the functional target(s) for IL-4/IL-13 inducing non-healing disease, the course of L. mexicana infection was monitored in mice lacking IL-4Rα expression in specific cellular compartments. A deficiency of IL-4Rα expression on macrophages/neutrophils (in LysM(cre)IL-4Rα(-/lox) animals) had minimal effect on the outcome of L. mexicana infection compared with control (IL-4Rα(-/flox)) mice. In contrast, CD4(+) T cell specific (Lck(cre)IL-4Rα(-/lox)) IL-4Rα(-/-) mice infected with L. mexicana developed small lesions, which subsequently healed in female mice, but persisted in adult male mice. While a strong Th1 response was manifest in both male and female CD4(+) T cell specific IL-4Rα(-/-) mice infected with L. mexicana, induction of IL-4 was manifest in males but not females, independently of CD4(+) T cell IL-4 responsiveness. Similar results were obtained using pan-T cell specific (iLck(cre)IL-4Rα(-/lox)) IL-4Rα(-/-) mice. Collectively these data demonstrate that upon infection with L. mexicana, initial lesion growth in BALB/c mice is dependent on non-T cell population(s) responsive to IL-4/IL-13 while progressive infection is dependent on CD4(+) T cells responsive to IL-4.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Leishmania mexicana/imunologia , Leishmaniose Cutânea/imunologia , Leishmaniose Cutânea/mortalidade , Receptores de Superfície Celular/deficiência , Receptores de Superfície Celular/imunologia , Animais , Feminino , Interleucina-13/imunologia , Interleucina-4/imunologia , Leishmania mexicana/patogenicidade , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Fatores SexuaisRESUMO
In this study we generated a novel dual specific phosphatase 4 (DUSP4) deletion mouse using a targeted deletion strategy in order to examine the role of MAP kinase phosphatase-2 (MKP-2) in immune responses. Lipopolysaccharide (LPS) induced a rapid, time and concentration-dependent increase in MKP-2 protein expression in bone marrow-derived macrophages from MKP-2(+/+) but not from MKP-2(-/-) mice. LPS-induced JNK and p38 MAP kinase phosphorylation was significantly increased and prolonged in MKP-2(-/-) macrophages whilst ERK phosphorylation was unaffected. MKP-2 deletion also potentiated LPS-stimulated induction of the inflammatory cytokines, IL-6, IL-12p40, TNF-α, and also COX-2 derived PGE(2) production. However surprisingly, in MKP-2(-/-) macrophages, there was a marked reduction in LPS or IFNγ-induced iNOS and nitric oxide release and enhanced basal expression of arginase-1, suggesting that MKP-2 may have an additional regulatory function significant in pathogen-mediated immunity. Indeed, following infection with the intracellular parasite Leishmania mexicana, MKP-2(-/-) mice displayed increased lesion size and parasite burden, and a significantly modified Th1/Th2 bias compared with wild-type counterparts. However, there was no intrinsic defect in MKP-2(-/-) T cell function as measured by anti-CD3 induced IFN-γ production. Rather, MKP-2(-/-) bone marrow-derived macrophages were found to be inherently more susceptible to infection with Leishmania mexicana, an effect reversed following treatment with the arginase inhibitor nor-NOHA. These findings show for the first time a role for MKP-2 in vivo and demonstrate that MKP-2 may be essential in orchestrating protection against intracellular infection at the level of the macrophage.
Assuntos
Citocinas/metabolismo , Mediadores da Inflamação/metabolismo , Leishmania mexicana/patogenicidade , Leishmaniose Cutânea/imunologia , Leishmaniose Cutânea/prevenção & controle , Macrófagos/imunologia , Proteínas Tirosina Fosfatases/fisiologia , Animais , Arginase/metabolismo , Western Blotting , Medula Óssea/imunologia , Medula Óssea/metabolismo , Medula Óssea/patologia , Células Cultivadas , Feminino , Leishmaniose Cutânea/metabolismo , Lipopolissacarídeos/farmacologia , Macrófagos/metabolismo , Macrófagos/patologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Óxido Nítrico/metabolismo , Fosforilação , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Th1/imunologia , Células Th1/metabolismo , Células Th2/imunologia , Células Th2/metabolismo , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
Leishmania mexicana cysteine peptidases (CPs) have been identified as important parasite virulence factors. More recently, a natural inhibitor of CPs (ICP) from L. mexicana has been characterized, and ICP mutants have been created. Infection of BALB/c mice with ICP null mutants or ICP reexpressing mutants resulted in nonhealing, progressively growing lesions albeit slightly attenuated compared with the growth of lesions produced by wild-type parasites. In contrast, BALB/c mice infected with mutants overexpressing ICP were able to significantly control lesion growth or heal. While BALB/c mice infected with wild-type parasites, ICP null mutants, or ICP reexpressing mutants produced significant antibody responses, including immunoglobulin E (IgE), no Th1 response, as indicated by antigen-induced splenocyte gamma interferon (IFN-gamma) production, could be demonstrated. In contrast, BALB/c mice infected with mutants overexpressing ICP produced significantly less antibody, particularly IgE, as well as significantly reduced splenocyte interleukin-4 and enhanced IFN-gamma production. BALB/c mice were able to resolve infection following infection with one ICP overexpressing clone, which was subsequently used for vaccination studies with BALB/c mice. However, no protection was afforded these mice when they were challenged with wild-type parasites. Nevertheless, two other mouse strains susceptible to L. mexicana, C3H and C57BL/6, vaccinated with overexpressing ICP mutants were able to control challenge infection associated with an enhanced Th1 response. This study confirms that L. mexicana CPs are virulence factors and that ICPs have therapeutic potential.
Assuntos
Cisteína Endopeptidases/metabolismo , Inibidores de Cisteína Proteinase/biossíntese , Leishmania mexicana/imunologia , Leishmania mexicana/patogenicidade , Leishmaniose Cutânea/patologia , Células Th1/imunologia , Fatores de Virulência/antagonistas & inibidores , Animais , Anticorpos Antiprotozoários/sangue , Inibidores de Cisteína Proteinase/genética , Feminino , Deleção de Genes , Teste de Complementação Genética , Imunoglobulina E/sangue , Interferon gama/metabolismo , Leishmaniose Cutânea/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Índice de Gravidade de Doença , Baço/imunologia , VirulênciaRESUMO
Interleukin-18 deficient mice on a BALB/c background display increased resistance to cutaneous infection with Leishmania mexicana, with reduced lesion progression and reduced parasite burdens compared with wild-type mice. Infected IL-18-/- mice had lower antigen specific IgG1 levels and total IgE levels and conversely higher antigen specific IgG2a levels than similarly infected wild-type mice. Splenocytes isolated from infected IL-18-/- mice produced significantly lower levels of antigen induced IL-4 and higher levels of IFN-gamma than wild-type animals. Consequently IL-18 during L. mexicana infection of BALB/c mice promotes a Th2 biased response and thereby has a disease exacerbating role.