The NF-κB Inhibitor, IMD-0354, Affects Immune Gene Expression, Bacterial Microbiota and Trypanosoma cruzi Infection in Rhodnius prolixus Midgut

Abstract

Rhodnius prolixus is an insect vector of Trypanosoma cruzi, the causative agent of Chagas disease in Latin America. Nuclear factor-κB (NF-κB) transcription factors (TF) are conserved components of the innate immune system in several multicellular organisms including insects. The drug IMD-0354 [N-(3,5-bis-trifluoromethyl-phenyl)-5-chloro-2-hydroxy-benzamide] is a selective inhibitor of IκB kinases. It blocks IκBα phosphorylation thus preventing nuclear translocation of the NF-κb TF. In humans, NF-κB is involved in several biological processes such as inflammation, cell proliferation and immunity. In insects, the activation of the immune system upon microbial challenge can be controlled by signaling pathways such as the immune deficiency (IMD) and Toll, to combat infection. These activated pathways signal to downstream NF-κB TF to stimulate specific immune genes, triggering the synthesis of several molecules such as the antimicrobial peptides. In Drosophila melanogaster, the activation and regulation of NF-κB TF have been elucidated, while in triatomines these mechanisms are not fully understood Therefore, the present study investigated the effects of oral administration of the drug IMD-0354 on the R. prolixus immune response to challenge with bacteria and T. cruzi, as well as the impact on the gut bacterial microbiota. R. prolixus were fed with rabbit blood containing IMD-0354 and Escherichia coli, Staphylococcus aureus, or T. cruzi. The effects of IMD-0354 on insect mortality and antimicrobial activity in insect midgut samples, as well as the relative expression of R. prolixus immune genes were recorded. The bacterial microbiota was analyzed, and viable parasites were counted in insect midgut samples. The IMD-0354 treatment modulated antibacterial activity and the gene expression patterns of defensin A, defensin B, defensin C, and prolixicin, and the genes involved in the IMD and Toll pathways. Additionally, there was an increase of bacterial microbiota in treated insects. Insects treated with IMD-0354 and concomitantly infected with bacteria or T. cruzi through the blood meal had increased mortality, while the T. cruzi population in R. prolixus midgut was reduced. The inhibitory effect of IMD-0354 indicates the importance of NF-κB TF in the innate immune responses involved in the control of bacteria and parasite infections in the R. prolixus midgut.

The Role of Bacterial Symbionts in Triatomines: An Evolutionary Perspective

“…The AMPs involved in these studies are likely controlled by the Toll and IMD pathways as silencing the NF-κB transcription factor relish (IMD pathway) reduces the intestinal bacterial population while the opposite occurs when the NF-κB transcription factor dorsal (Toll pathway) is silenced [251]. The pharmacological inhibition of these pathways also affects AMP expression and reduces the microbiota levels [250]. Despite the effects on the microbiota, T. cruzi numbers remain unaltered when either pathway is silenced, suggesting that T. cruzi can tolerate the immune responses or actively induces some of these changes to regulate the microbiota [251,267].…”

The emerging role of IMD 0354 on bone homeostasis by suppressing osteoclastogenesis and bone resorption, but without affecting bone formation

“…Therefore, we screened multiple related compounds and found that N-(3,5-bis-trifluoromethyl-phenyl)-5-chloro-2-hydroxybenzamide (IMD 0354) is a non-ATP binding competitive and selective molecular inhibitor of IKK-β. IMD 0354 has been reported in several studies as an effective agent for the treatment of acute and subacute inflammatory diseases including myocarditis, pulmonary arterial hypertension, and diabetic retinopathy via the suppression of NF-κB activation in various cells, such as pulmonary arterial smooth muscle cells or myocardial cells [18][19][20][21] . There are considerable issues in regard to the drugs for osteoporosis treatment currently.…”

Modulation of IMD, Toll, and Jak/STAT Immune Pathways Genes in the Fat Body of Rhodnius prolixus During Trypanosoma rangeli Infection

“…showed that the expression of Relish and Cactus regulates infection prevalence and intensity, highlighting the importance of the IMD and Toll pathways in anti-parasitic immune responses in vectors ( Meister et al., 2005 ; Frolet et al., 2006 ; Hu and Aksoy, 2006 ; Antonova et al., 2009 ; Louradour et al., 2019 ). In this regard, a recent report demonstrated a reduction in T. cruzi loads in the midgut of R. prolixus treated with the drug IMD-0354, which impedes the activation of NF-κB TFs ( Vieira C. S. et al., 2018 ).…”

“…Primers were designed for: α-Actin (housekeeping), Rp-Relish , Rp-Dorsal , Rp-Caspar , and Rp-STAT . Primers for Rp-Cactus were retrieved from the literature ( Vieira C. S. et al., 2018 ). RT-qPCR efficiency for each primer pair was determined using the slope of a linear regression mode.…”

“…Strikingly, the same study revealed that silencing Rp-Relish or Rp-Dorsal did not influence in T. cruzi loads detected 1 or 2 weeks after the infection ( Mesquita et al., 2015 ). However, in a recent report, a reduction in T. cruzi loads was evidenced in the midgut of R. prolixus treated with the drug IMD-0354, which blocks IκBα phosphorylation and thus prevents nuclear translocation of NF-κB TFs ( Rp-Relish and Rp-Dorsal ), indicating the importance of these pathways in the anti-parasitic innate immune responses ( Vieira C. S. et al., 2018 ).…”