Extracellular Vesicles from Leishmania (Leishmania) infantum Contribute in Stimulating Immune Response and Immunosuppression in Hosts with Visceral Leishmaniasis.

Visceral leishmaniasis (VL) is a chronic systemic disease. In Brazil this infection is caused by Leishmania (Leishmania) infantum. Extracellular vesicles (EVs) released by Leishmania species have different functions like the modulation of host immune systems and inflammatory responses, among others. This study evaluated the participation of EVs from L. (L.) infantum (Leish-EVs) in recognition of the humoral and cellular immune response of hosts with VL. Promastigotes were cultivated in 199 medium and, in the log phase of growth, they were centrifuged, washed, resus-pended in RPMI medium, and incubated for 2 to 24 h, at 25 °C or 37 °C to release Leish-EVs. This dynamic was evaluated using transmission (TEM) and scanning (SEM) electron microscopies, as well as nanoparticle tracking analysis (NTA). The results suggested that parasite penetration in mammal macrophages requires more Leish-EVs than those living in insect vectors, since promastigotes incubated at 37 °C released more Leish-EVs than those incubated at 25 °C. Infected THP-1 cells produced high EV concentration (THP-1 cells-EVs) when compared with those from the control group. The same results were obtained when THP-1 cells were treated with Leish-EVs or a crude Leishmania antigen. These data indicated that host-EV concentrations could be used to distinguish infected from uninfected hosts. THP-1 cells treated with Leish-EVs expressed more IL-12 than control THP-1 cells, but were unable to express IFN-γ. These same cells highly expressed IL-10, which inhibited TNF-α and IL-6. Equally, THP-1 cells treated with Leish-EVs up-expressed miR-21-5p and miR-146a-5p. In conclusion, THP-1 cells treated with Leish-EVs highly expressed miR-21-5p and miR-146a-5p and caused the dysregulation of IL-10. Indirectly, these results suggest that high expression of these miRNAs species is caused by Leish-EVs. Consequently, this molecular via can contribute to immunosuppression causing enhanced immunopathology in infected hosts.

Plasma extracellular microRNAs are related to AIDS/cerebral toxoplasmosis co-infection

This study investigated the potential of five miRNA candidates for cerebral toxoplasmosis/HIV co-infection (CT/HIV) biomarkers. miR-155-5p, miR-146a-5p, miR-21-5p, miR-125b-5p and miR-29c-3p were tested in 79 plasma divided into groups: 32 CT/HIV patients; 27 individuals with asymptomatic toxoplasmosis (AT); and 20 individuals seronegative for toxoplasmosis (NC). From each was collected peripheral blood/EDTA for laboratory diagnosis. Blood cells for DNA extractions (molecular diagnosis), plasma for RNA extractions (gene expression) and ELISA (serological diagnosis). miRNA expression was performed by qPCR, and values were expressed in Relative Quantification (RQ). Among the five miRNAs, miR-21-5p and miR-146a-5p were up-expressed in CT/HIV group when compared with AT and NC groups. RQ means for miR-21-5p and miR-146a-5p in CT/HIV group were 3.829 and 2.500, while in AT group, were 1.815 and 1.661, respectively. Differences between 3 groups were statistically significant (Kruskal-Wallis ANOVA test), as well as CT/HIV and AT groups (Mann-Whitney test). Plasma of CT/HIV and AT groups expressed similar levels of miR-29c-3p, miR-155-5p and miR-125b-5p. As NC group was different of CT/HIV and AT groups, differences between three groups were statistically significant (Kruskal-Wallis ANOVA test). No difference was shown between CT/HIV and AT groups (Mann-Whitney test). These results suggest the host miRNAs modulation by Toxoplasma gondii

Expressão gênica de miRNA na toxoplasmose cerebral / Gene expression of miRNA in cerebral toxoplasmosis

A toxoplasmose constitui um sério problema de saúde pública, principalmente em pacientes imunocomprometidos, como os portadores do vírus da imunodeficiência humana (HIV). Apesar do acesso universal e gratuito à terapia antirretroviral (TARV) e a diminuição dos casos, a toxoplasmose cerebral ainda é responsável por alta morbidade e mortalidade, além de representar um determinante de mau prognóstico nesses pacientes. Os miRNAs são moléculas que exercem um importante papel na regulação da expressão gênica em células eucariontes. Sendo assim, há muito potencial para serem utilizados para o diagnóstico, prognóstico e intervenções terapêuticas. Diante disso, este estudo avaliou os níveis de expressão dos miRNAs miR-146a, miR-21, miR-155, miR-125b e miR-29c em plasma de pacientes com toxoplasmose cerebral e aids (TC/aids). Foram analisadas 79 amostras de plasma divididas em três grupos: grupo I- 32 amostras de plasma de pacientes TC/aids; grupo II- 27 amostras de plasma de indivíduos assintomáticos e soropositivos para toxoplasmose (TA) e grupo III- 20 amostras de indivíduos soronegativos para toxoplasmose (CN). Após a extração de RNA total contendo miRNA foi realizada a síntese do DNA complementar e em seguida o perfil de expressão de cada miRNA foi determinado por PCR quantitativo em tempo real. Os resultados foram expressos em quantificação relativa e mostraram que o miR-146a e o miR-21 foram significativamente mais expressos nos pacientes com toxoplasmose cerebral e aids quando comparados com os indivíduos assintomáticos e soropositivos para toxoplasmose.

Toxoplasmosis is an important public health problem, especially in immunocompromised patients, such as those with human immunodeficiency virus (HIV). Despite free universal access to antiretroviral therapy (HAART), cerebral toxoplasmosis is still responsible for high morbidity, mortality and a determinant of worst prognosis. MicroRNAs are molecules that play an important role in gene expression in eukaryotic cells. Therefore, these molecules have the potential to be used for diagnosis, prognosis and therapeutic. This study evaluated the miRNA expression levels of miR-146a, miR-21, miR-155, miR125b and miR-29c in plasma of patients with cerebral toxoplasmosis/aids (TC/AIDS). A total of 79 plasma samples were divided into three groups. Group I: 32 plasma samples from TC/aids patients. Group II: 27 plasma samples from individuals with asymptomatic toxoplasmosis, seropositives for toxopalsmosis (TA). Group III: 20 plasma samples from seronegative individuals for toxoplasmosis (CN). After extraction of total RNA, containing miRNA, a complementary DNA synthesis was performed and, then, the expression profile of each target miRNA, which was determined by quantitative real-time PCR (qPCR). Results expressed as relative quantification (RQ) showed that miR-146a and miR-21 were significantly up-expressed in TC/AIDS patients, when compared with TA individuals. Analyzes of miR-155, miR-125b and miR-29c showed that no statistically significant differences were shown between TC/AIDS and TA groups. These results suggest the host miRNAs modulation by T. gondii