Danilo Candido de Almeida

Gut Bacteria Products Prevent AKI Induced by Ischemia-Reperfusion

Short-chain fatty acids (SCFAs) are fermentation end products produced by the intestinal microbiota and have anti-inflammatory and histone deacetylase-inhibiting properties. Recently, a dual relationship between the intestine and kidneys has been unraveled. Therefore, we evaluated the role of SCFA in an AKI model in which the inflammatory process has a detrimental role. We observed that therapy with the three main SCFAs (acetate, propionate, and butyrate) improved renal dysfunction caused by injury. This protection was associated with low levels of local and systemic inflammation, oxidative cellular stress, cell infiltration/activation, and apoptosis. However, it was also associated with an increase in autophagy. Moreover, SCFAs inhibited histone deacetylase activity and modulated the expression levels of enzymes involved in chromatin modification. In vitro analyses showed that SCFAs modulated the inflammatory process, decreasing the maturation of dendritic cells and inhibiting the capacity of these cells to induce CD4(+) and CD8(+) T cell proliferation. Furthermore, SCFAs ameliorated the effects of hypoxia in kidney epithelial cells by improving mitochondrial biogenesis. Notably, mice treated with acetate-producing bacteria also had better outcomes after AKI. Thus, we demonstrate that SCFAs improve organ function and viability after an injury through modulation of the inflammatory process, most likely via epigenetic modification.

Immune Regulatory Properties of Allogeneic Adipose-Derived Mesenchymal Stem Cells in the Treatment of Experimental Autoimmune Diabetes

Adipose-derived mesenchymal stem cells (ADMSCs) display immunosuppressive properties, suggesting a promising therapeutic application in several autoimmune diseases, but their role in type 1 diabetes (T1D) remains largely unexplored. The aim of this study was to investigate the immune regulatory properties of allogeneic ADMSC therapy in T cell–mediated autoimmune diabetes in NOD mice. ADMSC treatment reversed the hyperglycemia of early-onset diabetes in 78% of diabetic NOD mice, and this effect was associated with higher serum insulin, amylin, and glucagon-like peptide 1 levels compared with untreated controls. This improved outcome was associated with downregulation of the CD4+ Th1-biased immune response and expansion of regulatory T cells (Tregs) in the pancreatic lymph nodes. Within the pancreas, inflammatory cell infiltration and interferon-γ levels were reduced, while insulin, pancreatic duodenal homeobox-1, and active transforming growth factor-β1 expression were increased. In vitro, ADMSCs induced the expansion/proliferation of Tregs in a cell contact–dependent manner mediated by programmed death ligand 1. In summary, ADMSC therapy efficiently ameliorates autoimmune diabetes pathogenesis in diabetic NOD mice by attenuating the Th1 immune response concomitant with the expansion/proliferation of Tregs, thereby contributing to the maintenance of functional β-cells. Thus, this study may provide a new perspective for the development of ADMSC-based cellular therapies for T1D.

Exploring the Role of Soluble Factors Associated with Immune Regulatory Properties of Mesenchymal Stem Cells

Mesenchymal stem cells (MSCs) are characterized as multipotent stromal cells with the capacity for both self-renewal and differentiation into mesodermal cell lineages. MSCs also have a fibroblast-like phenotype and can be isolated from several tissues. In recent years, researchers have found that MSCs secrete several soluble factors that exert immunosuppressive effects by modulating both innate (macrophages, dendritic and NK cells) and adaptive (B cells and CD4+ and CD8+ T cells) immune responses. This review summarizes the principal trophic factors that are related to immune regulation and secreted by MSCs under both autoimmune and inflammatory conditions. The understanding of mechanisms that regulate immunity in MSCs field is important for their future use as a novel cellular-based immunotherapy with clinical applications in several diseases.

Human hepatic stellate cell line (LX-2) exhibits characteristics of bone marrow-derived mesenchymal stem cells

The LX-2 cell line has characteristics of hepatic stellate cells (HSCs), which are considered pericytes of the hepatic microcirculatory system. Recent studies have suggested that HSCs might have mesenchymal origin. We have performed an extensive characterization of the LX-2 cells and have compared their features with those of mesenchymal cells. Our data show that LX-2 cells have a phenotype resembling activated HSCs as well as bone marrow-derived mesenchymal stem cells (BM-MSCs). Our immunophenotypic analysis showed that LX-2 cells are positive for activated HSC markers (αSMA, GFAP, nestin and CD271) and classical mesenchymal makers (CD105, CD44, CD29, CD13, CD90, HLA class-I, CD73, CD49e, CD166 and CD146) but negative for the endothelial marker CD31 and endothelial progenitor cell marker CD133 as well as hematopoietic markers (CD45 and CD34). LX-2 cells also express the same transcripts found in immortalized and primary BM-MSCs (vimentin, annexin 5, collagen 1A, NG2 and CD140b), although at different levels. We show that LX-2 cells are capable to differentiate into multilineage mesenchymal cells in vitro and can stimulate new blood vessel formation in vivo. LX-2 cells appear not to possess tumorigenic potential. Thus, the LX-2 cell line behaves as a multipotent cell line with similarity to BM-MSCs. This line should be useful for further studies to elucidate liver regeneration mechanisms and be the foundation for development of hepatic cell-based therapies.

Adipose tissue-derived mesenchymal stem cells increase skin allograft survival and inhibit Th-17 immune response.

Adipose tissue-derived mesenchymal stem cells (ADSC) exhibit immunosuppressive capabilities both in vitro and in vivo. Their use for therapy in the transplant field is attractive as they could render the use of immunosuppressive drugs unnecessary. The aim of this study was to investigate the effect of ADSC therapy on prolonging skin allograft survival. Animals that were treated with a single injection of donor allogeneic ADSC one day after transplantation showed an increase in donor skin graft survival by approximately one week. This improvement was associated with preserved histological morphology, an expansion of CD4+ regulatory T cells (Treg) in draining lymph nodes, as well as heightened IL-10 expression and down-regulated IL-17 expression. In vitro, ADSC inhibit naïve CD4+ T cell proliferation and constrain Th-1 and Th-17 polarization. In summary, infusion of ADSC one day post-transplantation dramatically increases skin allograft survival by inhibiting the Th-17 pathogenic immune response and enhancing the protective Treg immune response. Finally, these data suggest that ADSC therapy will open new opportunities for promoting drug-free allograft survival in clinical transplantation.

Long-Term Aerobic Exercise Protects against Cisplatin- Induced Nephrotoxicity by Modulating the Expression of IL-6 and HO-1

Nephrotoxicity is substantial side effect for 30% of patients undergoing cancer therapy with cisplatin and may force them to change or even abandon the treatment. Studies regarding aerobic exercise have shown its efficacy for the treatment of many types of diseases and its capacity to reduce tumors. However, little is known about the impact of physical exercise on cisplatin-induced acute kidney injury (AKI). In the present study, our aim was to investigate the role of physical exercise in AKI induced by cisplatin. We submitted C57Bl6 male mice to seven weeks of chronic exercise on a training treadmill and treated them with single i.p. injection of cisplatin (20 mg/kg) in the last week. Exercise efficacy was confirmed by an increased capillary-to-fiber ratio in the gastrocnemius muscle of exercised groups (EX and CIS-EX). The group submitted to exercise before cisplatin administration (CIS-EX) exhibited less weight loss and decreased serum urea levels compared to the cisplatin group (CIS). Exercise also showed a protective role against cisplatin-induced cell death in the kidney. The CIS-EX group showed a lower inflammatory response, with less TNF and IL-10 expression in the kidney and serum. In the same group, we observed an increase of IL-6 and HO-1 expression in the kidney. Taken together, our results indicate that chronic aerobic exercise is able to attenuate AKI by inducing IL-6 and HO-1 production, which results in lower inflammatory and apoptotic profiles in the kidney.

Kinin B1 receptor deficiency attenuates cisplatin-induced acute kidney injury by modulating immune cell migration

Cisplatin is a chemotherapeutic agent that causes severe renal dysfunction. The kinin B1 receptor has been associated with the migration of immune cells to injured tissue as well as with renal inflammation. To examine the role of the kinin B1 receptor in cisplatin-induced acute kidney injury, we used kinin B1 receptor knockout mice and treatment with a receptor antagonist before and after cisplatin administration. Cisplatin injection caused exacerbation of renal macrophage and neutrophil migration, higher levels of serum creatinine and blood urea, upregulation of B1 receptor mRNA and an increase in pro-inflammatory cytokines expression. B1 receptor knockout mice exhibited a reduction in serum creatinine and blood urea levels, diminished apoptosis, and decreased cisplatin-induced upregulation of inflammatory components. Moreover, treatment with the B1 receptor antagonist prior to cisplatin administration normalized serum creatinine, blood urea levels, protected from acute tubular necrosis, apoptosis-related genes, and prevented upregulation of pro-inflammatory cytokines. Thus, we propose that kinins have an important role in cisplatin-induced acute kidney injury by impairing immune cells migration to renal tissue during cisplatin nephrotoxicity.Key messageKinin B1 receptor is upregulated after cisplatin exposure.Kinin B1 receptor deficiency diminishes the nephrotoxicity caused by cisplatin.Kinin B1 receptor deficiency ameliorates the inflammatory response.Kinin B1 receptor deficiency diminishes apoptosis caused by cisplatin.Kinin B1 receptor antagonism ameliorates renal function after cisplatin injection.

Metformin exerts antitumor activity via induction of multiple death pathways in tumor cells and activation of a protective immune response

The antitumor effect of metformin has been demonstrated in several types of cancer; however, the mechanisms involved are incompletely understood. In this study, we showed that metformin acts directly on melanoma cells as well as on the tumor microenvironment, particularly in the context of the immune response. In vitro, metformin induces a complex interplay between apoptosis and autophagy in melanoma cells. The anti-metastatic activity of metformin in vivo was assessed in several mouse models challenged with B16F10 cells. Metformins activity was, in part, immune system-dependent, whereas its antitumor properties were abrogated in immunodeficient (NSG) mice. Metformin treatment increased the number of lung CD8-effector-memory T and CD4+Foxp3+IL-10+ T cells in B16F10-transplanted mice. It also decreased the levels of Gr-1+CD11b+ and RORγ+ IL17+CD4+ cells in B16F10-injected mice and the anti-metastatic effect was impaired in RAG-1−/− mice challenged with B16F10 cells, suggesting an important role for T cells in the protection induced by metformin. Finally, metformin in combination with the clinical metabolic agents rapamycin and sitagliptin showed a higher antitumor effect. The metformin/sitagliptin combination was effective in a BRAFV600E/PTEN tamoxifen-inducible murine melanoma model. Taken together, these results suggest that metformin has a pronounced effect on melanoma cells, including the induction of a strong protective immune response in the tumor microenvironment, leading to tumor growth control, and the combination with other metabolic agents may increase this effect.

Caloric Restriction Is More Efficient than Physical Exercise to Protect from Cisplatin Nephrotoxicity via PPAR-Alpha Activation

The antineoplastic drug cisplatin promotes renal injury, which limits its use. Protocols that reduce renal cisplatin toxicity will allow higher doses to be used in cisplatin treatment. Here, we compare physical exercise and caloric restriction (CR) as protocols to reduce cisplatin renal injury in mice. Male C57BL/6 were divided into four groups: Control, cisplatin, exercise + cisplatin, and 30% CR + cisplatin. Animals were injected with a single dose of cisplatin (20 mg/kg i.p.) and sacrificed 96 h after injection. Quantitative real time PCR, histological analyses, immunohistochemistry, and biochemical measurements were performed to investigate renal injury, necrosis, apoptosis, and inflammatory mechanisms. Both protocols protected against cisplatin renal injury, but CR was more effective in reducing uraemia and renal necrosis. The CR + Cisplatin group exhibited reduced serum IL-1β and TNF-α levels. No differences were noted in the renal mRNA expression of cytokines. Both interventions reduced apoptosis, but only the CR + Cisplatin group decreased TNFR2 protein expression. PPAR-α was activated in mice after CR. An antagonist of PPAR-α blocked the protective effect of CR. Both interventions attenuated the nephrotoxicity caused by cisplatin injection, but CR + Cisplatin showed a better response by modulating TNFR2. Moreover, part of the CR benefit depends on PPAR-α activation.

Role of aryl hydrocarbon receptor in mesenchymal stromal cell activation: A minireview

Mesenchymal stromal cells (MSCs) possess great therapeutic advantages due to their ability to produce a diverse array of trophic/growth factors related to cytoprotection and immunoregulation. MSC activation via specific receptors is a crucial event for these cells to exert their immunosuppressive response. The aryl-hydrocarbon receptor (AhR) is a sensitive molecule for external signals and it is expressed in MSCs and, upon positive activation, may potentially regulate the MSC-associated immunomodulatory function. Consequently, signalling pathways linked to AhR activation can elucidate some of the molecular cascades involved in MSC-mediated immunosuppression. In this minireview, we have noted some important findings concerning MSC regulation via AhR, highlighting that its activation is associated with improvement in migration and immunoregulation, as well as an increase in pro-regenerative potential. Thus, AhR-mediated MSC activation can contribute to new perspectives on MSC-based therapies, particularly those directed at immune-associated disorders.