Chloroquine (CQ), an antimalarial drug, has been shown to modulate the immune system and reduce the severity of experimental autoimmune encephalomyelitis (EAE). The mechanisms of disease suppression are dependent on regulatory T cell induction, although Tregs-independent mechanisms exist. We aimed to evaluate whether CQ is capable to modulate bone marrow-derived dendritic cells (DCs) both phenotypically and functionally as well as whether transfer of CQ-modulated DCs reduces EAE course. Our results show that CQ-treated DCs presented altered ultrastructure morphology and lower expression of molecules involved in antigen presentation. Consequently, T cell proliferation was diminished in coculture experiments. When transferred into EAE mice, DC-CQ was able to reduce the clinical manifestation of the disease through the modulation of the immune response against neuroantigens. The data presented herein indicate that chloroquine-mediated modulation of the immune system is achieved by a direct effect on DCs and that DC-CQ adoptive transfer may be a promising approach for avoiding drug toxicity.

Inflammation is a necessary process to control infection. However, exacerbated inflammation, acute or chronic, promotes deleterious effects in the organism. Violacein (viola), a quorum sensing metabolite from the Gram-negative bacterium Chromobacterium violaceum, has been shown to protect mice from malaria and to have beneficial effects on tumors. However, it is not known whether this drug possesses anti-inflammatory activity. In this study, we investigated whether viola administration is able to reduce acute and chronic autoimmune inflammation. For that purpose, C57BL/6 mice were intraperitoneally injected with 1 μg of LPS and were treated with viola (3.5mg/kg) via i.p. at the same time-point. Three hours later, the levels of inflammatory cytokines in the sera and phenotypical characterization of leukocytes were determined. Mice treated with viola presented a significant reduction in the production of inflammatory cytokines compared with untreated mice. Interestingly, although viola is a compound derived from bacteria, it did not induce inflammation upon administration to naïve mice. To test whether viola would protect mice from an autoimmune inflammation, Experimental Autoimmune Encephalomyelitis (EAE)-inflicted mice were given viola i.p. at disease onset, at the 10th day from immunization. Viola-treated mice developed mild EAE disease in contrast with placebo-treated mice. The frequencies of dendritic cells and macrophages were unaltered in EAE mice treated with viola. However, the sole administration of viola augmented the levels of splenic regulatory T cells (CD4+Foxp3+). We also found that adoptive transfer of viola-elicited regulatory T cells significantly reduced EAE. Our study shows, for the first time, that violacein is able to modulate acute and chronic inflammation. Amelioration relied in suppression of cytokine production (in acute inflammation) and stimulation of regulatory T cells (in chronic inflammation). New studies must be conducted in order to assess the possible use of viola in therapeutic approaches in human autoimmune diseases.

In previous years, several kinases, such as phosphoinositide 3-kinase (PI3K), mammalian target of rapamycin (mTOR), and extracellular-signal-regulated kinase (ERK), have been linked to important human diseases, although some kinase families remain neglected in terms of research, hiding their relevance to therapeutic approaches. Here, a review regarding the NEK family is presented, shedding light on important information related to NEKs and human diseases. NEKs are a large group of homologous kinases with related functions and structures that participate in several cellular processes such as the cell cycle, cell division, cilia formation, and the DNA damage response. The review of the literature points to the pivotal participation of NEKs in important human diseases, like different types of cancer, diabetes, ciliopathies and central nervous system related and inflammatory-related diseases. The different known regulatory molecular mechanisms specific to each NEK are also presented, relating to their involvement in different diseases. In addition, important information about NEKs remains to be elucidated and is highlighted in this review, showing the need for other studies and research regarding this kinase family. Therefore, the NEK family represents an important group of kinases with potential applications in the therapy of human diseases.

Tolerogenic dendritic cells (DCs) are widely studied for their possible use in the treatment of inflammatory disorders, such as autoimmune diseases. One of the obstacles for the use of this cell-based therapy is the characterization of drugs that are able to modulate DCs. We have previously shown that chloroquine (CQ), an antimalarial agent, has the ability to modulate DCs towards a tolerogenic phenotype. 1 These tolerogenic DCs are able to suppress the development of experimental autoimmune encephalomyelitis (EAE), a T cell-driven mouse model of human multiple sclerosis. In addition, several studies have proposed that nitric oxide (NO) plays a major role in the differentiation of regulatory T cells (Tregs) and the suppression of Th1/Th17 cells. 2,3 However, little is known about the role of DC-derived NO in the modulation of inflammatory autoimmune responses. Thus, we aimed to evaluate whether NO plays a role in the tolerogenic activity of CQ-treated DCs (CQDCs). We found that CQ induces DC production of NO and expression of indoleamine 2,3-dioxygenase (IDO), as well as inducible nitric oxide synthase (iNOS). In addition, CQ-DCs stimulated the differentiation of Tregs at the expense of Th1/ Th17 cells. On the other hand, iNOS 2/2 DCs did not acquire a tolerogenic phenotype following CQ treatment. Rather, CQDCs iNOS2/2 stimulated the differentiation of Th1/Th17 cells as well as Tregs. In a therapeutic approach, CQ-DCs iNOS2/2 were unable to suppress the development of EAE. Gene expression analyses of central nervous system (CNS) tissue from mice that received CQ-DCs iNOS2/2 showed an increased expression of inflammatory modulators compared with mice that received CQ-DCs WT . In this work, we show that NO is an important factor in the modulatory activity of tolerogenic dendritic cells.DCs are antigen-presenting cells that can dictate the course of the immune response via the modulation and activation of naive T cells. DC modulation is a possible approach to address the immunosuppression that is often caused by tumors 4 and the exacerbated immune response observed in autoimmune diseases. 5 Multiple sclerosis, one such autoimmune disease, is a debilitating condition that affects the CNS. Studies in EAE, an experimental mouse model of multiple sclerosis, have found that much of the immunological etiology of the disease development is due to the activity of Th1/Th17 cells, and these studies have found that NO plays a major role in disease progression. 2 To verify whether NO is involved in the modulatory activity of tolerogenic DCs, we generated DCs from bone marrow precursors obtained from wild-type (DCs WT ) and iNOS 2/2 (DCs iNOS2/2 ) mice and treated these DCs with CQ or vehicle (PBS-DCs WT ). All protocols involving laboratory animals were approved by the institutional committee (protocol no. 2687-1). NO measurements revealed that CQ treatment induced DCs WT to produce large amounts of NO in an iNOS-dependent manner (Figure 1a). It has been demonstrated that CQ administration results in NO production by the endot...

NIMA-related kinases, or NEKs, are a family of Ser/Thr protein kinases involved in cell cycle and mitosis, centrosome disjunction, primary cilia functions, and DNA damage responses among other biological functional contexts in vertebrate cells. In human cells, there are 11 members, termed NEK1 to 11, and the research has mainly focused on exploring the more predominant roles of NEKs in mitosis regulation and cell cycle. A possible important role of NEKs in DNA damage response (DDR) first emerged for NEK1, but recent studies for most NEKs showed participation in DDR. A detailed analysis of the protein interactions, phosphorylation events, and studies of functional aspects of NEKs from the literature led us to propose a more general role of NEKs in DDR. In this review, we express that NEK1 is an activator of ataxia telangiectasia and Rad3-related (ATR), and its activation results in cell cycle arrest, guaranteeing DNA repair while activating specific repair pathways such as homology repair (HR) and DNA double-strand break (DSB) repair. For NEK2, 6, 8, 9, and 11, we found a role downstream of ATR and ataxia telangiectasia mutated (ATM) that results in cell cycle arrest, but details of possible activated repair pathways are still being investigated. NEK4 shows a connection to the regulation of the nonhomologous end-joining (NHEJ) repair of DNA DSBs, through recruitment of DNA-PK to DNA damage foci. NEK5 interacts with topoisomerase IIβ, and its knockdown results in the accumulation of damaged DNA. NEK7 has a regulatory role in the detection of oxidative damage to telomeric DNA. Finally, NEK10 has recently been shown to phosphorylate p53 at Y327, promoting cell cycle arrest after exposure to DNA damaging agents. In summary, this review highlights important discoveries of the ever-growing involvement of NEK kinases in the DDR pathways. A better understanding of these roles may open new diagnostic possibilities or pharmaceutical interventions regarding the chemo-sensitizing inhibition of NEKs in various forms of cancer and other diseases.

SummaryDendritic cells (DCs) are professional antigen-presenting cells specifically targeted during Plasmodium infection. Upon infection, DCs show impaired antigen presentation and T-cell activation abilities. In this study, we aimed to evaluate whether cellular extracts obtained from Plasmodium berghei-infected erythrocytes (PbX) modulate DCs phenotypically and functionally and the potential therapeutic usage of PbX-modulated DCs in the control of experimental autoimmune encephalomyelitis (EAE, the mouse model for human multiple sclerosis). We found that PbX-treated DCs have impaired maturation and stimulated the generation of regulatory T cells when cultured with naive T lymphocytes in vitro. When adoptively transferred to C57BL/6 mice the EAE severity was reduced. Disease amelioration correlated with a diminished infiltration of cytokine-producing T cells in the central nervous system as well as the suppression of encephalitogenic T cells. Our study shows that extracts obtained from P. berghei-infected erythrocytes modulate DCs towards an immunosuppressive phenotype. In addition, the adoptive transfer of PbX-modulated DCs was able to ameliorate EAE development through the suppression of specific cellular immune responses towards neuro-antigens. To our knowledge, this is the first study to present evidence that DCs treated with P. berghei extracts are able to control autoimmune neuroinflammation.

The thymus plays an important role shaping the T cell repertoire in the periphery, partly, through the elimination of inflammatory auto-reactive cells. It has been shown that, during Plasmodium berghei infection, the thymus is rendered atrophic by the premature egress of CD4+CD8+ double-positive (DP) T cells to the periphery. To investigate whether autoimmune diseases are affected after Plasmodium berghei NK65 infection, we immunized C57BL/6 mice, which was previously infected with P.berghei NK65 and treated with chloroquine (CQ), with MOG35–55 peptide and the clinical course of Experimental Autoimmune Encephalomyelitis (EAE) was evaluated. Our results showed that NK65+CQ+EAE mice developed a more severe disease than control EAE mice. The same pattern of disease severity was observed in MOG35–55-immunized mice after adoptive transfer of P.berghei-elicited splenic DP-T cells. The higher frequency of IL-17+- and IFN-γ+-producing DP lymphocytes in the Central Nervous System of these mice suggests that immature lymphocytes contribute to disease worsening. To our knowledge, this is the first study to integrate the possible relationship between malaria and multiple sclerosis through the contribution of the thymus. Notwithstanding, further studies must be conducted to assert the relevance of malaria-induced thymic atrophy in the susceptibility and clinical course of other inflammatory autoimmune diseases.

Experimental autoimmune encephalomyelitis (EAE) is a T cell-dependent disease that mimics human multiple sclerosis (MS). The inflammation in the central nervous system (CNS) is a hallmark in these autoimmune inflammatory conditions [1]. Therefore, the development of new drugs that target the infiltration of inflammatory cells to the CNS is desired [2,3]. In this context, accumulating evidences place antimalarial drugs as immunomodulatory agents. For instance, administration of chloroquine, a weak base that has tropism toward acidic organelles, was shown to modulate a series of inflammatory diseases, including EAE [4,5]. On the other hand, primaquine (PQ), a drug analogous to chloroquine [6], has much less toxicity and is poorly investigated in the control of inflammation.In this study, we aimed to evaluate whether PQ administration is able to reduce EAE severity. For that purpose, we induced EAE in C57BL/6 mice through subcutaneous injection of 100 lg of MOG 35-55 peptide alongside with 200 ng of pertussis toxin, as previously described [7]. When the clinical signs of EAE started to appear, mice were treated with PQ (3 mg/kg, intraperitoneally) for five consecutive days and disease outcome was evaluated (EAE+PQ group). Mice treated with vehicle (PBS) were used as controls (EAE group). We observed that PQ treatment significantly reduced the development of EAE ( Figure 1A). Interestingly, disease amelioration correlated with lower infiltration of cells in the CNS ( Figure 1B). Notwithstanding, the frequency of Iba-1 + cells (microglia, macrophages, and dendritic cells) was found reduced in the EAE+PQ group compared with untreated EAE-bearing mice, suggesting that probably the demyelination process is reduced in the CNS of EAE+PQ mice ( Figure 1B). To evaluate whether glial cells are affected by PQ treatment, slices were stained for glial fibrillary acidic protein (GFAP) production. Our results showed that EAE+PQ mice presented lower expression of GFAP in comparison with the EAE group ( Figure 1B). Of note, PQ treatment by itself did not induce GFAP expression.Collectively, these results show that improvement in EAE+PQ mice correlated with a reduction in cellular infiltration and gliosis. To investigate whether mediators of inflammation are involved in disease amelioration, the gene expression levels of inflammationrelated cytokines and transcription factors were analyzed. Results showed that levels of FOXP3, IL-10, and TGF-b were significantly increased, while the levels of TNF-a, IL-17, IFN-c, and GATA-3 were decreased in the CNS of EAE+PQ mice compared with the control ( Figure 1C). Unexpectedly, levels of Th17 cell-related transcription factor, RORct, were found reduced in EAE-bearing mice compared with na€ ıve mice, showing that modulation of this transcription factor is not a key element in PQ-induced EAE amelioration.The increased expression in FOXP3, IL-10, and TGF-b in the CNS of EAE+PQ mice suggests that the treatment induces the expansion of regulatory T (Treg) cells. To confirm this hypothesis, na€ ıve m...