In vitro evaluation of human intravenous immunoglobulin in combination with antimicrobials and human serum against multidrug-resistant isolates of Acinetobacter baumannii.

The high incidence of multidrug-resistant (MDR) Acinetobacter baumannii has been a challenge for health worldwide, due to the reduction of therapeutic options, making the use of antimicrobial combinations necessary for the treatment, such as meropenem, amikacin, and colistin. Antibodies against bacterial species, mainly immunoglobulins G (IgG), are produced for acting as effector mechanisms (neutralization, opsonization, phagocytosis, and complement system activation). Some studies have demonstrated promising results of IgG in combination with antimicrobial preparations against bacterial infections, in which the direct action of IgG has restored the immune system balance. Serious problem caused by the increase of MDR A. baumannii isolates results in a constant search for therapeutic alternatives to defeat these infections. However, this study aims to verify in vitro the phagocytosis rate of the A. baumannii-infected human monocytes, as well as to analyze possible morphological changes induced by intravenous immunoglobulin G (IVIG) with human serum in association with antimicrobials. The phagocytosis rate and bacterial cell binding capacity of IVIG were determined for two A. baumannii isolates submitted to 4 mg/mL of human IVIG alone and in combination with different sub-minimum inhibitory concentrations (sub-MICs) of meropenem, amikacin, and colistin and processed for indirect immunofluorescence. Subsequently, these isolates were resubmitted and coupled with human serum and processed for scanning electron microscopy. There was no statistical difference for phagocytosis rates in the isolates tested. Bacterial isolates showed alterations in cell morphology when exposed to IVIG/human serum alone and in combination with antimicrobials such as alteration in shape, wrinkling, membrane depression, and especially cell rupture with extravasation of cytoplasmic material. The isolates visually differed in the IVIG binding to the bacterial cell, with higher fluorescence intensity, which corresponds to the highest IVIG binding, in the isolate more sensitive to meropenem, amikacin, and colistin. No differences between treatments were observed in the IVIG binding to the bacterial cell. The combined action of IVIG with meropenem, amikacin, and colistin against A. baumannii MDR isolates induced several bacterial cell damages. And when associated with human serum, a massive destruction of cells can be observed. These results may suggest the analysis of the use of IgG preparations for the treatment of A. baumannii MDR infections.

Benznidazole treatment decreases IL-6 levels in Trypanosoma cruzi-infected human adipocytes differentiated from adipose tissue-derived stem cells.

BACKGROUND: Trypanosoma cruzi, which causes Chagas disease (CD), is a versatile haemoparasite that uses several strategies to evade the host's immune response, including adipose tissue (AT), used as a reservoir of infection. As it is an effective barrier to parasite evasion, the effectiveness of the drug recommended for treating CD, Benznidazole (BZ), may be questionable. OBJECTIVE: To this end, we evaluated the parasite load and immunomodulation caused by BZ treatment in the culture of adipocytes differentiated from human adipose tissue-derived stem cells (ADSC) infected with T. cruzi. METHODS: The ADSC were subjected to adipogenic differentiation. We then carried out four cultures in which we infected the differentiated AT with trypomastigote forms of the Y strain of T. cruzi and treated them with BZ. After the incubation, the infected AT was subjected to quantitative polymerase chain reaction (qPCR) to quantify the parasite load and transmission electron microscopy (TEM) to verify the infection. The supernatant was collected to measure cytokines, chemokines, and adipokines. FINDINGS: We found elevated secretion of IL-6, CXCL-10/IP-10, CCL2/MCP-1, CCL5/RANTES, and leptin in infected fat cells. However, treatment with BZ promoted a decrease in IL-6. MAIN CONCLUSION: Therefore, we believe that BZ has a beneficial role as it reduces inflammation in infected fat cells.

Benzonidazole treatment has a beneficial effect on cells infected with the Colombian strain of Trypanosoma cruzi.

Benznidazole (Bz) is the recommended drug for the treatment of Chagas disease; however, its efficacy may vary according to the sensitivity of Trypanosoma cruzi strains to the drug and host immune background. The study evaluated the immune response of peripheral blood mononuclear cells (PBMC) that were infected in vitro with the Colombian strain (Col) and treated with Bz. The co-cultures were incubated for 24 h, 5 and 10 days, where cytokine dosage was performed in the supernatant and evaluation of the cells for CD28+ and CTLA-4+ molecules in CD4+ and CD8+ lymphocytes, and CD80+ , CD86+ and HLA-DR+ in CD14+ cells. The results showed that Col induced a strong inflammatory response, with an increase in IFN-γ and TNF early in the infection (24 h), however, from 5 days of infection on, TNF production declined, and IL-10 production increased, which may be associated with a control mechanism of the exacerbated inflammatory response. The Bz treatment did not significantly alter the frequencies of the phenotypes evaluated both T cell subsets and CD14+ cells. Therefore, this study reinforces the need for typing the patient's strain to guide therapy and promote individualized treatment protocols due to the heterogeneous genetic background among T. cruzi strains.

Benznidazole treatment decreases IL-6 levels in Trypanosoma cruzi-infected human adipocytes differentiated from adipose tissue-derived stem cells

BACKGROUND Trypanosoma cruzi, which causes Chagas disease (CD), is a versatile haemoparasite that uses several strategies to evade the host's immune response, including adipose tissue (AT), used as a reservoir of infection. As it is an effective barrier to parasite evasion, the effectiveness of the drug recommended for treating CD, Benznidazole (BZ), may be questionable. OBJECTIVE To this end, we evaluated the parasite load and immunomodulation caused by BZ treatment in the culture of adipocytes differentiated from human adipose tissue-derived stem cells (ADSC) infected with T. cruzi. METHODS The ADSC were subjected to adipogenic differentiation. We then carried out four cultures in which we infected the differentiated AT with trypomastigote forms of the Y strain of T. cruzi and treated them with BZ. After the incubation, the infected AT was subjected to quantitative polymerase chain reaction (qPCR) to quantify the parasite load and transmission electron microscopy (TEM) to verify the infection. The supernatant was collected to measure cytokines, chemokines, and adipokines. FINDINGS We found elevated secretion of IL-6, CXCL-10/IP-10, CCL2/MCP-1, CCL5/RANTES, and leptin in infected fat cells. However, treatment with BZ promoted a decrease in IL-6. MAIN CONCLUSION Therefore, we believe that BZ has a beneficial role as it reduces inflammation in infected fat cells.

Synthesis, in vitro schistosomicidal activity and ultrastructural alterations caused by thiosemicarbazones and thiazolidinones against juvenile and adult Schistosoma mansoni worms (Sambon, 1907).

Schistosomiasis is a neglected disease that affects about 258 million people worldwide. Caused by Schistosoma mansoni, helminth which, in Brazil, it is present on 19 states and capital. Praziquantel (PZQ) treatment presents low efficacy and adverse effects in parasites juvenile stages. Thiosemicarbazones and thiazolidinones are rising as potent chemical groups that have biological activity wide spectrum, and with radical modifications, they may become more effective and selective. Aiming to evaluate the action of these molecules against S. mansoni, JF series thiosemicarbazones and thiazolidinones (LqIT/UFPE) were synthesized: JF30, JF31, JF33, JF34, JF35, JF36, JF38, JF39, JF42 and JF43. Several parameters were evaluated, such as: their cytotoxicity in VERO cells, in vitro schistosomicidal activity for juvenile and adult worms and their action on worms through ultrastructural changes. Cytotoxicity indices ranged from 272 µM to 725 µM. When evaluating mortality rate, adult and juvenile worms showed 100 % mortality rate within 24 h and 48 h, respectively, when exposed to the compounds JF31 and JF43 at a dose of 200 µM. Also, motility, mortality and oviposition parameters were evaluated: JF31 and JF43 presented a score of 0 in 24 h, meaning total absence of movement, whereas no eggs and soft tissue damage were observed under optical microscopy. Through scanning electron microscopy, integumentary alterations caused by the compounds JF31 and JF43 were observed, such as: exposure of the musculature, formation of integumentary bubbles, integuments with abnormal morphology and destruction of tubercles and spikes. The results shoerd that the compound JF31 was 2.39 times more selective for adult worms and JF43 was 3.74 times more selective for juvenile worms. Thus, the compounds JF43 and JF31 are the most promising for presenting schistosomicidal activity of S. mansoni.

Effects of Migonemyia migonei salivary gland homogenates on Leishmania (Viannia) braziliensis infection in BALB/c mice.

Cutaneous leishmaniasis caused by Leishmania (Viannia) braziliensis is the most widespread clinical form of leishmaniasis in the Americas. Migonemyia migonei is a widely distributed phlebotomine sand fly species in Brazil and has been implicated as a vector for L. (V.) braziliensis. In the present study, we investigated the effects of salivary gland homogenates (SGH) of Mg. migonei on the course of L. (V.) braziliensis infection in BALB/c mice. Mice were separated into four groups (six mice per group): CTRL (uninfected mice); SGH (mice inoculated with Mg. migonei SGH); SGH+LEISH (mice inoculated with Mg. migonei SGH plus L. (V.) braziliensis promastigotes); LEISH (mice inoculated with L. (V.) braziliensis promastigotes). Mice were followed up for 8 weeks and the cellular immune response was evaluated by flow cytometry at the end of the experiment. Analysis of cytokine production by splenic cells stimulated with 0.5 SGH, 0.25 SGH of Mg. migonei or L. (V.) braziliensis soluble antigen stimulation (LSA) demonstrated that upon stimulation with SGH 0.25, the production of IL-17A and TNF was not sustained in the SGH group, with decreasing levels of these cytokines after 5 days compared to 3 days of incubation. Analyzing the production of cytokines after LSA stimulation, we observed lower levels of IL-17A in the SGH group after 5 days compared to 3 days. The same was observed for IFN-γ in the SGH group. Yet, the levels of TNF were significantly higher in the LEISH group after 5 days compared to 3 days. Among SGH+LEISH and LEISH mice, three animals in each group developed skin lesions on the tail, the mean lesion size was significantly higher in the LEISH group. Our study suggests that Mg. migonei SGH may modulate BALB/c immune response, as reflected by the low production or early decrease of pro-inflammatory cytokines in splenic cell cultures following stimulation with L. (V.) braziliensis antigen. Our data also suggest that Mg. migonei saliva may reduce the lesion size in BALB/c mice, but further research with a larger sample size is needed to confirm this hypothesis.

Avaliação das quimiocinas e da expressão de seus receptores em células mononucleares de pacientes portadores da doença de Chagas submetidas à infecção in vitro com Trypanosoma cruzi e tratamento com benzonidazol / Evaluation of chemokines and their receptors expression in patient mononuclear patients of Chagas disease undergoing in vitro infection with Trypanosoma cruzi and treatment with benznidazole

Atualmente o Brasil apresenta 3 milhões de indivíduos portadores da cardiomiopatia chagásica. Porém, tratamento etiológico com o fármaco Benzonidazol (BZ) na fase crônica da doença ainda não está elucidado. Acredita-se que a recomendação do BZ nessa fase, pode prevenir ou retardar a evolução clínica da cardiomiopatia na Doença Chagas (DC). Assim o objetivo do estudo é avaliar a produção de quimiocinas e expressão de seus receptores em Células mononucleares do sangue periférico - PBMC (de portadores crônicos da doença de Chagas) submetidas in vitro ao tratamento com BZ, após a infecção com T.cruzi. Foram selecionados 11 pacientes na fase crônica da doença. Amostras de sangue desses pacientes foram coletadas para obtenção de PBMC, em que foram cultivadas em placas de cultivo na concentração de 106 células/ml por poço. Após a adesão das células aderentes (principalmente macrófagos), as células não aderentes (principalmente linfócitos) foram removidas e as formas tripomastigotas foram adicionadas ao cultivo para infecção das células aderentes. Subsequente a incubação, as células não aderentes foram adicionadas novamente ao cultivo juntamente com o fármaco Bz (1µg/mL), ficando um co-cultivo de células aderentes infectadas com T.cruzi, células não aderentes e o BZ (C+T+BZ). As placas de cultura foram incubadas por períodos de 24h e 5 dias. Para uma análise fidedigna da ação do BZ nas células aderentes e não aderentes foi necessário a criação dos controles: células (C), células e tripomastigotas (C+T) e células e o BZ (C+BZ). Após o cultivo, foram coletados os sobrenadantes das culturas, para avaliação da produção de quimiocinas (CCL2, CXL9, CXL10, CCL5 e CXCL8) por CBA (Cytometric Bead Array). Posteriormente foi realizada a imunofenotipagem...