RESUMEN
Single nucleotide polymorphisms (SNPs) that do not change the composition of amino acids and cause synonymous mutations (sSNPs) were previously considered to lack any functional roles. However, sSNPs have recently been shown to interfere with protein expression owing to a myriad of factors related to the regulation of transcription, mRNA stability, and protein translation processes. In patients with Chagas disease, the presence of the synonymous mutation rs1129293 in phosphatidylinositol-4,5-bisphosphate 3-kinase gamma (PIK3CG) gene contributes to the development of the chronic Chagas cardiomyopathy (CCC), instead of the digestive or asymptomatic forms. In this study, we aimed to investigate whether rs1129293 is associated with the transcription of PIK3CG mRNA and its activity by quantifying AKT phosphorylation in the heart samples of 26 chagasic patients with CCC. Our results showed an association between rs1129293 and decreased PIK3CG mRNA expression levels in the cardiac tissues of patients with CCC. The phosphorylation levels of AKT, the protein target of PI3K, were also reduced in patients with this mutation, but were not correlated with PI3KCG mRNA expression levels. Moreover, bioinformatics analysis showed that rs1129293 and other SNPs in linkage disequilibrium (LD) were associated with the transcriptional regulatory elements, post-transcriptional modifications, and cell-specific splicing expression of PIK3CG mRNA. Therefore, our data demonstrates that the synonymous SNP rs1129293 is capable of affecting the PIK3CG mRNA expression and PI3Kγ activation.
Asunto(s)
Cardiomiopatía Chagásica , Cardiomiopatía Chagásica/genética , Fosfatidilinositol 3-Quinasa Clase Ib/genética , Humanos , Fosfatidilinositol 3-Quinasas , Polimorfismo de Nucleótido Simple , Proteínas Proto-Oncogénicas c-akt , ARN Mensajero/genética , ARN Mensajero/metabolismo , Mutación SilenciosaRESUMEN
Chagas disease is caused by the protozoan parasite Trypanosoma cruzi. During the chronic phase of disease, while most infected people do not present symptoms, characterizing the asymptomatic form, some patients develop the cardiac form or chronic chagasic cardiomyopathy, which is considered the most severe manifestation of this disease. Considering that the activation of the PI3Kγ signaling pathway is essential for an efficient immune response against T. cruzi infection, we evaluated the PIK3CG C > T (rs1129293) polymorphism in exon 3 of this gene, which encodes the catalytic subunit of PI3Kγ. The PIK3CG CT and TT genotypes were found to be associated with an increased risk of developing the cardiac form of the disease rather than the asymptomatic or digestive forms. In conclusion, the presence of the T allele at single or double doses may differentiate the cardiac from other clinical manifestations of Chagas disease. This finding should help in further studies to evaluate the mechanisms underlying the differential association of PIK3CG in Chagas disease.
Asunto(s)
Dominio Catalítico/genética , Cardiomiopatía Chagásica/genética , Enfermedad de Chagas/genética , Enfermedad de Chagas/parasitología , Fosfatidilinositol 3-Quinasa Clase Ib/genética , Polimorfismo de Nucleótido Simple , Trypanosoma cruzi , Cardiomiopatía Chagásica/parasitología , Fosfatidilinositol 3-Quinasa Clase Ib/metabolismo , Variación Genética , Genotipo , Corazón/parasitología , Interacciones Huésped-Parásitos , Humanos , Enfermedades Desatendidas/genética , Enfermedades Desatendidas/parasitología , Transducción de SeñalRESUMEN
This study investigates the participation of PI3Kγ in the development of joint inflammation and dysfunction in an experimental model of acute gout in mice. Acute gout was induced by injection of monosodium urate (MSU) crystals into the tibiofemoral joint of mice. The involvement of PI3Kγ was evaluated using a selective inhibitor and mice deficient for PI3Kγ (PI3Kγ-/- ) or with loss of kinase activity. Neutrophils recovered from the inflamed joint were quantified and stained for phosphorylated Akt (pAkt) and production of reactive oxygen species (ROS). The adherence of leukocytes to the joint microvasculature was assessed by intravital microscopy and cleaved caspase-1 by Western blot. Injection of MSU crystals induced massive accumulation of neutrophils expressing phosphorylated Akt. In the absence of PI3Kγ, there was reduction of pAkt expression, chemokine production, and neutrophil recruitment. Genetic or pharmacological inhibition of PI3Kγ reduced the adherence of leukocytes to the joint microvasculature, even in joints with established inflammation. Neutrophils from PI3Kγ-/- mice produced less ROS than wild-type neutrophils. There was decreased joint damage and dysfunction in the absence of PI3Kγ. In addition, in the absence of PI3Kγ activity, there was reduction of cleaved caspase-1 and IL-1ß production in synovial tissue after injection of MSU crystals and leukotriene B4 . Our studies suggest that PI3Kγ is crucial for MSU crystal-induced acute joint inflammation. It is necessary for regulating caspase-1 activation and for mediating neutrophil migration and activation. Drugs that impair PI3Kγ function may be useful to control acute gout inflammation.
Asunto(s)
Artritis Gotosa/enzimología , Artritis Gotosa/inmunología , Caspasa 1/metabolismo , Fosfatidilinositol 3-Quinasa Clase Ib/metabolismo , Infiltración Neutrófila , Enfermedad Aguda , Animales , Adhesión Celular , Movimiento Celular , Fosfatidilinositol 3-Quinasa Clase Ib/deficiencia , Citoplasma/metabolismo , Activación Enzimática , Inflamasomas/metabolismo , Inflamación/patología , Interleucina-1beta/metabolismo , Articulaciones/patología , Leucotrieno B4/metabolismo , Masculino , Ratones Endogámicos C57BL , Microvasos/patología , Neutrófilos/metabolismo , Nocicepción , Fosforilación , Proteínas Proto-Oncogénicas c-akt/metabolismo , Membrana Sinovial/irrigación sanguínea , Ácido ÚricoRESUMEN
Given the impossibility to study the lung immune response during Mycobacterium tuberculosis-latent infection, and consequently, the mechanisms that control the bacterial load, it is reasonable to determine the activation of local immunity in the early phase of the infection. The phosphatidylinositol-3-kinase gamma enzyme (PI3Kγ) is involved in the leukocyte recruitment, phagocytosis and cellular differentiation, and therefore, it is considered a promising target for the development of immunotherapies for chronic inflammatory diseases. Mice genetically deficient in PI3Kγ (PI3Kγ-/-) or WT (Wild Type) were evaluated 15 days post-infection. The enzyme deficiency improved the resistance against infection, increased the frequency of CD4+IL-17+ cells, the production of IL-17 as well as the gene and protein expression of molecules associated with Th17â¯cell differentiation and neutrophil recruitment. Our findings show, for the first time, the participation of the PI3Kγ in vivo in the M. tuberculosis-infection, and suggest an association of Th17â¯cells with protection in the early phase of tuberculosis.