Plasma Sphingomyelin Disturbances: Unveiling Its Dual Role as a Crucial Immunopathological Factor and a Severity Prognostic Biomarker in COVID-19.

SARS-CoV-2 infection triggers distinct patterns of disease development characterized by significant alterations in host regulatory responses. Severe cases exhibit profound lung inflammation and systemic repercussions. Remarkably, critically ill patients display a "lipid storm", influencing the inflammatory process and tissue damage. Sphingolipids (SLs) play pivotal roles in various cellular and tissue processes, including inflammation, metabolic disorders, and cancer. In this study, we employed high-resolution mass spectrometry to investigate SL metabolism in plasma samples obtained from control subjects (n = 55), COVID-19 patients (n = 204), and convalescent individuals (n = 77). These data were correlated with inflammatory parameters associated with the clinical severity of COVID-19. Additionally, we utilized RNAseq analysis to examine the gene expression of enzymes involved in the SL pathway. Our analysis revealed the presence of thirty-eight SL species from seven families in the plasma of study participants. The most profound alterations in the SL species profile were observed in patients with severe disease. Notably, a predominant sphingomyelin (SM d18:1) species emerged as a potential biomarker for COVID-19 severity, showing decreased levels in the plasma of convalescent individuals. Elevated SM levels were positively correlated with age, hospitalization duration, clinical score, and neutrophil count, as well as the production of IL-6 and IL-8. Intriguingly, we identified a putative protective effect against disease severity mediated by SM (d18:1/24:0), while ceramide (Cer) species (d18:1/24:1) and (d18:1/24:0)were associated with increased risk. Moreover, we observed the enhanced expression of key enzymes involved in the SL pathway in blood cells from severe COVID-19 patients, suggesting a primary flow towards Cer generation in tandem with SM synthesis. These findings underscore the potential of SM as a prognostic biomarker for COVID-19 and highlight promising pharmacological targets. By targeting sphingolipid pathways, novel therapeutic strategies may emerge to mitigate the severity of COVID-19 and improve patient outcomes.

sTREM-1 Predicts Disease Severity and Mortality in COVID-19 Patients: Involvement of Peripheral Blood Leukocytes and MMP-8 Activity.

Uncontrolled inflammatory responses play a critical role in coronavirus disease (COVID-19). In this context, because the triggering-receptor expressed on myeloid cells-1 (TREM-1) is considered an intrinsic amplifier of inflammatory signals, this study investigated the role of soluble TREM-1 (sTREM-1) as a biomarker of the severity and mortality of COVID-19. Based on their clinical scores, we enrolled COVID-19 positive patients (n = 237) classified into mild, moderate, severe, and critical groups. Clinical data and patient characteristics were obtained from medical records, and their plasma inflammatory mediator profiles were evaluated with immunoassays. Plasma levels of sTREM-1 were significantly higher among patients with severe disease compared to all other groups. Additionally, levels of sTREM-1 showed a significant positive correlation with other inflammatory parameters, such as IL-6, IL-10, IL-8, and neutrophil counts, and a significant negative correlation was observed with lymphocyte counts. Most interestingly, sTREM-1 was found to be a strong predictive biomarker of the severity of COVID-19 and was related to the worst outcome and death. Systemic levels of sTREM-1 were significantly correlated with the expression of matrix metalloproteinases (MMP)-8, which can release TREM-1 from the surface of peripheral blood cells. Our findings indicated that quantification of sTREM-1 could be used as a predictive tool for disease outcome, thus improving the timing of clinical and pharmacological interventions in patients with COVID-19.

AeMOPE-1, a Novel Salivary Peptide From Aedes aegypti, Selectively Modulates Activation of Murine Macrophages and Ameliorates Experimental Colitis.

The sialotranscriptomes of Aedes aegypti revealed a transcript overexpressed in female salivary glands that codes a mature 7.8 kDa peptide. The peptide, specific to the Aedes genus, has a unique sequence, presents a putative secretory nature and its function is unknown. Here, we confirmed that the peptide is highly expressed in the salivary glands of female mosquitoes when compared to the salivary glands of males, and its secretion in mosquito saliva is able to sensitize the vertebrate host by inducing the production of specific antibodies. The synthetic version of the peptide downmodulated nitric oxide production by activated peritoneal murine macrophages. The fractionation of a Ae. aegypti salivary preparation revealed that the fractions containing the naturally secreted peptide reproduced the nitric oxide downmodulation. The synthetic peptide also selectively interfered with cytokine production by murine macrophages, inhibiting the production of IL-6, IL-12p40 and CCL2 without affecting TNF-α or IL-10 production. Likewise, intracellular proteins associated with macrophage activation were also distinctively modulated: while iNOS and NF-κB p65 expression were diminished, IκBα and p38 MAPK expression did not change in the presence of the peptide. The anti-inflammatory properties of the synthetic peptide were tested in vivo on a dextran sulfate sodium-induced colitis model. The therapeutic administration of the Ae. aegypti peptide reduced the leukocytosis, macrophage activity and nitric oxide levels in the gut, as well as the expression of cytokines associated with the disease, resulting in amelioration of its clinical signs. Given its biological properties in vitro and in vivo, the molecule was termed Aedes-specific MOdulatory PEptide (AeMOPE-1). Thus, AeMOPE-1 is a novel mosquito-derived immunobiologic with potential to treat immune-mediated disorders.

Endothelinergic Contractile Hyperreactivity in Rat Contralateral Carotid to Balloon Injury: Integrated Role for ETB Receptors and Superoxide Anion.

Temporal consequences of neurocompensation to balloon injury on endothelinergic functionality in rat contralateral carotid were evaluated. Rats underwent balloon injury in left carotid and were treated with CP-96345 (NK1 antagonist). Concentration-response curves for endothelin-1 were obtained in contralateral (right) carotid at 2, 8, 16, 30, or 45 days after surgery in the absence or presence of BQ-123 (ETA antagonist), BQ-788 (ETB antagonist), or Tempol (superoxide-dismutase mimic). Endothelin-1-induced calcium mobilization was evaluated in functional assays carried out with BQ-123, BQ-788, or Tempol. Endothelin-1-induced NADPH oxidase-driven superoxide generation was measured by lucigenin chemiluminescence assays performed with BQ-123 or BQ-788. Endothelin-1-induced contraction was increased in contralateral carotid from the sixteenth day after surgery. This response was restored in CP-96345-treated rats. Endothelium removal or BQ-123 did not change endothelin-1-induced contraction in contralateral carotid. This response was restored by BQ-788 or Tempol. Contralateral carotid exhibited an increased endothelin-1-induced calcium mobilization, which was restored by BQ-788 or Tempol. Contralateral carotid exhibited an increased endothelin-1-induced lucigenin chemiluminescence, which was restored by BQ-788. We conclude that the NK1-mediated neurocompensatory response to balloon injury elicits a contractile hyperreactivity to endothelin-1 in rat contralateral carotid by enhancing the muscular ETB-mediated NADPH oxidase-driven generation of superoxide, which activates calcium channels.

Amelioration of experimental colitis after short-term therapy with glucocorticoid and its relationship to the induction of different regulatory markers.

The clinical benefits of short-term therapy with glucocorticoids (GC) in patients with inflammatory bowel disease (IBD) are widely known. However, the effects of this treatment towards the re-establishment of the regulatory network in IBD are not fully explored. We have evaluated the immunological effects of the abbreviated GC therapy in experimental colitis induced by 3% dextran sulphate sodium in C57BL/6 mice. Treatment with GC improved disease outcome, constrained circulating leucocytes and ameliorated intestinal inflammation. The control of the local inflammatory responses involved a reduction in the expression of interferon-γ and interleukin-1ß, associated with augmented mRNA levels of peroxisome proliferator-activated receptors (α and γ) in intestine. Furthermore, there was a reduction of CD4+ T cells producing interferon-γ, together with an increased frequency of the putative regulatory population of T cells producing interleukin-10, in spleen. These systemic alterations were accompanied by a decrease in the proliferative potential of splenocytes of mice treated in vivo with GC. Notably, treatment with GC also led to an increase in the frequency of the regulatory markers GITR, CTLA-4, PD-1, CD73 and FoxP3, more prominently in spleen. Taken together, our results pointed to a role of GC in the control of leucocyte responsiveness and re-establishment of a regulatory system, which probably contributed to disease control and the restoration of immune balance. Finally, this is the first time that GC treatment was associated with the modulation of a broad number of regulatory markers in an experimental model of colitis.

Toxoplasma gondii soluble tachyzoite antigen triggers protective mechanisms against fatal intestinal pathology in oral infection of C57BL/6 mice.

Toxoplasma gondii induces a potent IL-12 response early in infection that results in IFN-γ-dependent control of parasite growth. It was previously shown that T. gondii soluble tachyzoite antigen (STAg) injected 48 hr before intraperitoneal infection reduces lipoxin A4 and 5-lipoxygenase (5-LO)-dependent systemic IL-12 and IFN-γ production as well as hepatic immunopathology. This study investigated the ability of STAg-pretreatment to control the fatal intestinal pathology that develops in C57BL/6 mice orally infected with 100 T. gondii cysts. STAg-pretreatment prolonged the animals' survival by decreasing tissue parasitism and pathology, mainly in the ilea. Protection was associated with decreases in the systemic IFN-γ levels and IFN-γ and TNF message levels in the ilea and with increased TGF-ß production in this tissue, but protection was independent of 5-LO and IL-4. STAg-pretreatment decreased CD4(+) T cell, NK cell, CD11b(+) monocyte and CD11b(+)CD11c(+) dendritic cell numbers in the lamina propria and increased CD8(+) T cells in the intestinal epithelial compartment. In parallel, decreases were observed in iNOS and IL-17 expression in this organ. These results demonstrate that pretreatment with STAg can induce the recruitment of protective CD8(+) T cells to the intraepithelial compartment and decrease proinflammatory immune mechanisms that promote intestinal pathology in T. gondii infection.

Enrichment of regulatory T cells in invasive breast tumor correlates with the upregulation of IL-17A expression and invasiveness of the tumor.

Breast cancer is a leading cause of neoplasia-associated death in women worldwide. Regulatory T (Treg) and Th17 cells are enriched within some tumors, but the role these cells play in invasive ductal carcinoma (IDC) of the breast is unknown. We show that CD25(+) CD4(+) T cells from PBMCs and tumor express high levels of Foxp3, GITR, CTLA-4, and CD103, indicating that tumor-infiltrating Treg cells are functional and possibly recruited by CCL22. Additionally, we observed upregulation of Th17-related molecules (IL-17A, RORC, and CCR6) and IL-17A produced by tumor-infiltrating CD4(+) and CD8(+) T lymphocytes. The angiogenic factors CXCL8, MMP-2, MMP-9, and vascular endothelial growth factor detected within the tumor are possibly induced by IL-17 and indicative of poor disease prognosis. Treg and Th17 cells were synchronically increased in IDC patients, with positive correlation between Foxp3, IL-17A, and RORC expression, and associated with tumor aggressiveness. Therefore, Treg and Th17 cells can affect disease progression by Treg-cell-mediated suppression of the effector T-cell response, as indicated by a decrease in the proliferation of T cells isolated from PBMCs of IDC patients and induction of angiogenic factors by IL-17-producing Th17. The understanding of regulation of the Treg/Th17 axis may result in novel perspectives for the control of invasive tumors.

Different parasite inocula determine the modulation of the immune response and outcome of experimental Trypanosoma cruzi infection.

During infection, the host response develops effector mechanisms to combat the parasite. However, this response can become uncontrolled or regulated by mechanisms that modulate the inflammatory reaction. The number of parasites that infects the host, such as trypomastigotes in Chagas disease, may also influence immune activation and disease pathology. We evaluated the inflammation and immune regulation that follows Trypanosoma cruzi infection with low (300), intermediate (3000) or high (30000) parasite loads. Our results showed that the load of parasite inoculum influenced disease outcome: the higher the number of parasites in the inoculum, the lower were the survival rates. There was a strong association between parasitism and inflammatory infiltrate in the heart and the parasite inoculum determined cytokine interplay in this tissue, as shown by increased interferon-γ, tumour necrosis factor-α, interleukin-17 (IL-17) and IL-23 in the 300 and 30000 inoculum groups, higher IL-4 and IL-10 in the intermediate-inoculum mice, and elevated IL-6 production in the heart of mice in the 3000 and 30000 groups. The number of T cells and antigen-presenting cells was augmented in the infected groups, especially for the splenic CD4(+) CD25(+) regulatory T cells expressing CD45RB(low) , GITR, PD-1 and FoxP3 in the group with the highest inoculum. Interestingly, these mice also presented an apparent decrease in CD4(+) CD25(+) FoxP3(+) cells in the cardiac infiltrate, in contrast to the intermediate inoculum group, which showed elevated numbers of these regulatory leucocytes in the heart. Finally, our results demonstrated that parasite load during T. cruzi infection is linked to the response pattern that will result in parasite/inflammation control or tissue damage.

Human mucosal leishmaniasis: neutrophils infiltrate areas of tissue damage that express high levels of Th17-related cytokines.

Mucosal leishmaniasis (ML) is characterised by severe tissue destruction. Herein, we evaluated the involvement of the IL-17-type response in the inflammatory infiltrate of biopsy specimens from 17 ML patients. IL-17 and IL-17-inducing cytokines (IL-1ß, IL-23, IL-6 and TGF-ß) were detected by immunohistochemistry in ML patients. IL-17(+) cells exhibited CD4(+), CD8(+) or CD14(+) phenotypes, and numerous IL-17(+) cells co-expressed the CC chemokine receptor 6 (CCR6). Neutrophils, a hallmark of Th17-mediated inflammation, were regularly detected in necrotic and perinecrotic areas and stained positive for neutrophil elastase, myeloperoxidase and MMP-9. Taken together, these observations demonstrate the existence of Th17 cells in ML lesions associated with neutrophils in areas of tissue injury and suggest that IL-17 is involved in ML pathogenesis.

Regulatory T cells attenuate experimental periodontitis progression in mice.

AIMS: The aim of this study was to identify the presence and characterize the function of regulatory T cells (Tregs) in experimental periodontitis in mice. MATERIAL AND METHODS: C57Bl/6 mice infected with Actinobacillus actinomycetemcomitans, treated or not with anti-glucocorticoid-inducible tumour necrosis factor receptor (anti-GITR) to inhibit Tregs function, were analysed regarding inflammatory cell and Tregs influx, alveolar bone loss and cytokine expression/production (analysed by real-time polymerase chain reaction and ELISA) throughout experimental periodontitis. RESULTS: A. actinomycetemcomitans inoculation in mice resulted in periodontal disease characterized by marked alveolar bone loss and an influx of inflammatory cells. Flow cytometry evaluation of inflammatory cells demonstrated an increased number of CD4(+)CD25(+) and CD4(+)FOXp3(+) cells, characterizing the presence of Tregs in the periodontal environment in a late stage after infection. Tregs-associated cytokines interleukin-10 (IL-10), cytotoxic T lymphocyte-associated molecule 4 (CTLA-4) and transforming growth factor-beta (TGF-beta) were found to be expressed/produced in a kinetics that resembles Tregs migration. Treatment with anti-GITR, which inhibits Tregs function, showed increased alveolar bone loss and inflammatory cell migration. A reduction in IL-10, CTLA-4 and TGF-beta levels was also observed, while interferon-gamma, tumour necrosis factor-alpha and receptor activator for nuclear factor kappaB ligand levels were increased. However, bacterial load and C-reactive protein serum did not show any differences. CONCLUSION: Taken together, our results showed that the presence of Treg cells attenuates the severity of experimental periodontitis without impairment in the control of infection.

Down-regulation of expression of osteoblast and osteocyte markers in periodontal tissues associated with the spontaneous alveolar bone loss of interleukin-10 knockout mice.

The aim of this study was to unravel the mechanisms by which interleukin (IL)-10, a potent pleiotropic cytokine, modulates alveolar bone homeostasis in C57BL/6 wild-type (WT) and IL-10 knockout (IL-10 KO) mice, evaluated at 8, 24, and 48 wk of age. Interleukin-10 KO mice presented significant alveolar bone loss when compared with WT mice, and this was not associated with changes in leukocyte counts or bacterial load. The levels of expression of messenger RNA (mRNA) for tumor necrosis factor-alpha (TNF-alpha), IL-1beta, IL-6, transforming growth factor-beta (TGF-beta), receptor activator of nuclear factor kappaB ligand (RANKL), osteoprotegerin (OPG), and matrix metalloproteinase 13 (MMP13) were similar between both strains, whereas a significant decrease of tissue inhibitor of metalloproteinase 1 (TIMP1) mRNA expression was found at 48 wk in IL-10 KO mice. The osteoblast markers core binding factor alpha1 (CBFA1) and type I collagen (COL-I) were expressed at similar levels in both strains, whereas the levels of alkaline phosphatase (ALP) and osteocalcin (OCN), and those of the osteocyte markers phosphate-regulating gene endopeptidases (PHEX) and dentin matrix protein 1 (DMP1) were significantly lower in IL-10 KO mice. Our results demonstrate that the alveolar bone loss in the absence of IL-10 was associated with a reduced expression of osteoblast and osteocyte markers, an effect independent of microbial, inflammatory or bone-resorptive pathways.

Strong and persistent microbial and inflammatory stimuli overcome the genetic predisposition to higher matrix metalloproteinase-1 (MMP-1) expression: a mechanistic explanation for the lack of association of MMP1-1607 single-nucleotide polymorphism genotypes with MMP-1 expression in chronic periodontitis lesions.

AIMS: Our objective was to evaluate the association between the MMP1-1607 single-nucleotide polymorphism (SNP), periodontopathogens and inflammatory cytokines with matrix metalloproteinase-1 (MMP-1) mRNA levels in vitro and in vivo. MATERIALS AND METHODS: This study investigated the influence of genetic (MMP1-1607 SNP), microbial (Porphyromonas gingivalis, Treponema denticola, Tannerella forsythia, Actinobacillus actinomycetemcomitans) and inflammatory [tumour necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta)] factors on the determination of MMP-1 mRNA levels in periodontal tissues of non-smoker chronic periodontitis (CP, N=178) and control (C, N=190) groups. The effects of single and repeated lipopolysaccharide (LPS) and inflammatory cytokine stimulation of macrophages with distinct MMP1-1607 SNP genotypes were also investigated. RESULTS: In healthy tissues, the MMP1-1607 2G allele was associated with higher MMP-1 levels while in CP MMP-1 levels were associated with the presence and load of periodontopathogens, and also with TNF-alpha and IL-1beta expression irrespective of the MMP1-1607 genotype. In vitro data demonstrate that in 2G macrophages low- and intermediate-dose LPS and TNF-alpha+IL-1beta stimulation was associated with increased MMP-1 expression, while strong and repeated stimulation resulted in higher MMP-1 levels irrespective of the MMP1-1607 genotype. CONCLUSION: Our data demonstrate a limited role for MMP1-1607 SNP in periodontitis, where the extensive chronic antigenic challenge exposure overcomes the genetic control and plays a major role in the determination of MMP-1 expression.

IL-4 regulates susceptibility to intestinal inflammation in murine food allergy.

Allergies involve a state of immediate hypersensitivity to antigens, including food proteins. The mechanism underlying the initiation and development of allergic responses involves IL-4 that directly induces the differentiation of committed effector Th2 lymphocytes. Although it is clear that Th2 responses play a pivotal role in the development of allergic responses, it remains unclear which mechanisms are involved in the development of the intestinal damages observed in food allergy. Accordingly, this work aimed to study the role of Th2/IL-4-dependent responses in the development of food allergy and intestinal pathology. C57BL/6 wild-type (WT) and IL-4-/- mice were sensitized with peanut proteins, challenged with peanut seeds, and followed for the development of food allergy and intestinal inflammation. Results demonstrated that exposure to peanut seeds led to weight loss in WT but not in IL-4-/- mice that preserved gut integrity with no signs of mucosal inflammation. These animals presented increased levels of IgG2a in sera, suggesting a role for allergic antibodies in the pathogenesis of WT animals. Most importantly, results also showed that lack of IL-4 modulated gut mucosal response in food allergy through diminished expression of TNF-alpha mRNA, increased Th1 IFN-gamma, IL-12p40, regulatory cytokines, and Foxp3, demonstrating their relevance in the control of allergic inflammatory processes, especially in the intestine. Finally, this study highlighted some of the complex mechanisms involved in the pathogenesis of allergic responses to food antigens in the gut, thereby providing valuable tools for directing novel therapeutic or preventive strategies to the control of allergic enteropathy.

Chlorhexidine-induced apoptosis or necrosis in L929 fibroblasts: A role for endoplasmic reticulum stress.

Chlorhexidine (CHX), widely used as antiseptic and therapeutic agent in medicine and dentistry, has a toxic effect both in vivo and in vitro. The intrinsic mechanism underlying CHX-induced cytotoxicity in eukaryotic cells is, however, still unknown. A recent study from our laboratory has suggested that CHX may induce death in cultured L929 fibroblasts via endoplasmic reticulum (ER) stress. This hypothesis was further tested by means of light and electron microscopy, quantification of apoptosis and necrosis by flow cytometry, fluorescence visualization of the cytoskeleton and endoplasmic reticulum, and evaluation of the expression of 78-kDa glucose-regulated protein 78 (Grp78), a marker of activation of the unfolded protein response (UPR) in cultured L929 fibroblasts. Our finding showing increased Grp 78 expression in CHX-treated cells and the results of flow cytometry, cytoskeleton and endoplasmic reticulum fluorescence visualization, and scanning and transmission electron microscopy allowed us to suggest that CHX elicits accumulation of proteins in the endoplasmic reticulum, which causes ER overload, resulting in ER stress and cell death either by necrosis or apoptosis. It must be pointed out, however, that this does not necessarily mean that ER stress is the only way that CHX kills L929 fibroblasts, but rather that ER stress is an important target or indicator of cell death induced by this drug.

The broad effects of the functional IL-10 promoter-592 polymorphism: modulation of IL-10, TIMP-3, and OPG expression and their association with periodontal disease outcome.

Periodontal diseases are infectious diseases, in which periodontopathogens trigger chronic inflammatory and immune responses that lead to tissue destruction. It occurs through the generation of metalloproteinases and the activation of bone resorption mechanisms. Anti-inflammatory cytokines such as IL-10 seem to attenuate periodontal tissue destruction through the induction of tissue inhibitors of metalloproteinases (TIMPs) and the inhibitor of osteoclastogenesis osteoprotegerin (OPG). A high individual variation in levels of IL-10 mRNA is verified in periodontitis patients, which is possibly determined by genetic polymorphisms. In this study, the IL-10 promoter -592C/A single nucleotide polymorphism (SNP), which is associated with a decrease in IL-10 production, was analyzed by RFLP in 116 chronic periodontitis (CP) patients and 173 control (C) subjects, and the IL-10, TIMPs, and OPG mRNA expression levels in diseased gingival tissues were determined by real-time-PCR. The IL-10-592 SNP CA (P=0.0012/OR=2.4/CI:1.4-4.1), AA (P=0.0458/OR=2.3/CI:1.1-4.9), and CA+AA (P=0.0006/OR=2.4/CI:1.4-3.4) genotypes and the allele A (P=0.0036/OR=1.7/CI:1.2-2.4) were found to be significantly more prevalent in the CP group when compared with control subjects. Both CA and AA genotypes were associated with lower levels of IL-10, TIMP-3, and OPG mRNA expression in diseased periodontal tissues and were also associated with disease severity as mean pocket depth. Taken together, the results presented here demonstrate that IL10-592 SNP is functional in CP, being associated with lower levels of IL-10 mRNA expression, which is supposed to consequently decrease the expression of the downstream genes TIMP-3 and OPG, and influence periodontal disease outcome.

The involvement of CD4+CD25+ T cells in the acute phase of Trypanosoma cruzi infection.

The infection with Trypanosoma cruzi leads to a vigorous and apparently uncontrolled inflammatory response in the heart. Although the parasites trigger specific immune response, the infection is not completely cleared out, a phenomenon that in other parasitic infections has been attributed to CD4+CD25+ T cells (Tregs). Then, we examined the role of natural Tregs and its signaling through CD25 and GITR in the resistance against infection with T. cruzi. Mice were treated with mAb against CD25 and GITR and the parasitemia, mortality and heart pathology analyzed. First, we demonstrated that CD4+CD25+GITR+Foxp3+ T cells migrate to the heart of infected mice. The treatment with anti-CD25 or anti-GITR resulted in increased mortality of these infected animals. Moreover, the treatment with anti-GITR enhanced the myocarditis, with increased migration of CD4+, CD8+, and CCR5+ leukocytes, TNF-alpha production, and tissue parasitism, although it did not change the systemic nitric oxide synthesis. These data showed a limited role for CD25 signaling in controlling the inflammatory response during this protozoan infection. Also, the data suggested that signaling through GITR is determinant to control of the heart inflammation, parasite replication, and host resistance against the infection.