Organometallic Gold(III) Complex [Au(Hdamp)(L14)]+ (L1 = SNS-Donating Thiosemicarbazone) as a Candidate to New Formulations against Chagas Disease.

Chagas disease remains a serious public health concern with unsatisfactory treatment outcomes due to strain-specific drug resistance and various side effects. To identify new therapeutic drugs against Trypanosoma cruzi, we evaluated both the in vitro and in vivo activity of the organometallic gold(III) complex [Au(III)(Hdamp)(L14)]Cl (L1 = SNS-donating thiosemicarbazone), henceforth denoted 4-Cl. Our results demonstrated that 4-Cl was more effective than benznidazole (Bz) in eliminating both the extracellular trypomastigote and intracellular amastigote forms of the parasite without cytotoxic effects on mammalian cells. In in vivo assays, 4-Cl in PBS solution loses the protonation and becomes the 4-neutral. 4-Neutral reduced parasitaemia and tissue parasitism in addition to protecting the liver and heart from tissue damage at 2.8 mg/kg/day. All these changes resulted in the survival of 100% of the mice treated with the gold complex during the acute phase. Analyzing the surviving animals of the acute infection, the parasite load after 150 days of infection was equivalent to those treated with the standard dose of Bz without demonstrating the hepatotoxicity of the latter. In addition, we identified a modulation of interferon gamma (IFN-γ) levels that may be targeting the disease's positive outcome. To the best of our knowledge, this is the first gold organometallic study that shows promise in an in vivo experimental model against Chagas disease.

The lack of PI3Kγ favors M1 macrophage polarization and does not prevent kidney diseases progression.

Acute kidney injury (AKI) and chronic kidney disease (CKD) are major concerns in worldwide public health, and their pathophysiology involves immune cells activation, being macrophages one of the main players of both processes. It is suggested that metabolic pathways could contribute to macrophage modulation and phosphatidylinositol­3 kinase (PI3K) pathway was shown to be activated in kidneys subjected to ischemia and reperfusion as well as unilateral ureteral obstruction (UUO). Although PI3K inhibition is mostly associated with anti-inflammatory response, its use in kidney injuries has been shown controversial results, which indicates the need for further studies. Our aim was to unveil the role of PI3Kγ in macrophage polarization and in kidney diseases development. We analyzed bone-marrow macrophages polarization from wild-type (WT) and PI3Kγ knockout (PI3K KO) animals. We observed increased expression of M1 (CD86, CCR7, iNOS, TNF, CXCL9, CXCL10, IL-12 and IL-23) and decreased of M2 (CD206, Arg-1, FIZZ1 and YM1) markers in the lack of PI3Kγ. And this modulation was accompanied by higher levels of inflammatory cytokines in PI3K KO M1 cells. PI3K KO mice had increased M1 in steady state kidneys, and no protection was observed in these mice after acute and chronic kidney insults. On the contrary, they presented higher levels of protein-to-creatinine ratio and Kim-1 expression and increased tubular injury. In conclusion, our findings demonstrated that the lack of PI3Kγ favors M1 macrophages polarization providing an inflammatory-prone environment, which does not prevent kidney diseases progression.

Canonical PI3Kγ signaling in myeloid cells restricts Trypanosoma cruzi infection and dampens chagasic myocarditis.

Chagas disease is caused by infection with the protozoan Trypanosoma cruzi (T. cruzi) and is an important cause of severe inflammatory heart disease. However, the mechanisms driving Chagas disease cardiomyopathy have not been completely elucidated. Here, we show that the canonical PI3Kγ pathway is upregulated in both human chagasic hearts and hearts of acutely infected mice. PI3Kγ-deficient mice and mutant mice carrying catalytically inactive PI3Kγ are more susceptible to T. cruzi infection. The canonical PI3Kγ signaling in myeloid cells is essential to restrict T. cruzi heart parasitism and ultimately to avoid myocarditis, heart damage, and death of mice. Furthermore, high PIK3CG expression correlates with low parasitism in human Chagas' hearts. In conclusion, these results indicate an essential role of the canonical PI3Kγ signaling pathway in the control of T. cruzi infection, providing further insight into the molecular mechanisms involved in the pathophysiology of chagasic heart disease.

Mast Cells and Serotonin Synthesis Modulate Chagas Disease in the Colon: Clinical and Experimental Evidence.

BACKGROUND: Trypanosoma cruzi (T. cruzi) infects millions of Latin Americans each year and can induce chagasic megacolon. Little is known about how serotonin (5-HT) modulates this condition. Aim We investigated whether 5-HT synthesis alters T. cruzi infection in the colon. MATERIALS AND METHODS: Forty-eight paraffin-embedded samples from normal colon and chagasic megacolon were histopathologically analyzed (173/2009). Tryptophan hydroxylase 1 (Tph1) knockout (KO) mice and c-KitW-sh mice underwent T. cruzi infection together with their wild-type counterparts. Also, mice underwent different drug treatments (16.1.1064.60.3). RESULTS: In both humans and experimental mouse models, the serotonergic system was activated by T. cruzi infection (p < 0.05). While treating Tph1KO mice with 5-HT did not significantly increase parasitemia in the colon (p > 0.05), rescuing its synthesis promoted trypanosomiasis (p < 0.01). T. cruzi-related 5-HT release (p < 0.05) seemed not only to increase inflammatory signaling, but also to enlarge the pericryptal macrophage and mast cell populations (p < 0.01). Knocking out mast cells reduced trypanosomiasis (p < 0.01), although it did not further alter the neuroendocrine cell number and Tph1 expression (p > 0.05). Further experimentation revealed that pharmacologically inhibiting mast cell activity reduced colonic infection (p < 0.01). A similar finding was achieved when 5-HT synthesis was blocked in c-KitW-sh mice (p > 0.05). However, inhibiting mast cell activity in Tph1KO mice increased colonic trypanosomiasis (p < 0.01). CONCLUSION: We show that mast cells may modulate the T. cruzi-related increase of 5-HT synthesis in the intestinal colon.

Emerging Role for the PERK/eIF2α/ATF4 in Human Cutaneous Leishmaniasis.

Leishmania parasites utilize adaptive evasion mechanisms in infected macrophages to overcome host defenses and proliferate. We report here that the PERK/eIF2α/ATF4 signaling branch of the integrated endoplasmic reticulum stress response (IERSR) is activated by Leishmania and this pathway is important for Leishmania amazonensis infection. Knocking down PERK or ATF4 expression or inhibiting PERK kinase activity diminished L. amazonensis infection. Knocking down ATF4 decreased NRF2 expression and its nuclear translocation, reduced HO-1 expression and increased nitric oxide production. Meanwhile, the increased expression of ATF4 and HO-1 mRNAs were observed in lesions derived from patients infected with the prevalent related species L.(V.) braziliensis. Our data demonstrates that Leishmania parasites activate the PERK/eIF2α/ATF-4 pathway in cultured macrophages and infected human tissue and that this pathway is important for parasite survival and progression of the infection.

Leishmania infantum Parasites Subvert the Host Inflammatory Response through the Adenosine A2A Receptor to Promote the Establishment of Infection.

Adenosine is an endogenously released purine nucleoside that signals through four widely expressed G protein-coupled receptors: A1, A2A, A2B, and A3. Of these, A2AR is recognized as mediating major adenosine anti-inflammatory activity. During cutaneous leishmaniasis, adenosine induces immunosuppression, which promotes the establishment of infection. Herein, we demonstrated that A2AR signaling is exploited by Leishmania infantum parasites, the etiologic agent that causes Visceral Leishmaniasis, to successfully colonize the vertebrate host. A2AR gene-deleted mice exhibited a well-developed cellular reaction with a strong Th1 immune response in the parasitized organs. An intense infiltration of activated neutrophils into the disease-target organs was observed in A2AR-/- mice. These cells were characterized by high expression of CXCR2 and CD69 on their cell surfaces and increased cxcl1 expression. Interestingly, this phenotype was mediated by IFN-γ on the basis that a neutralizing antibody specific to this cytokine prevented neutrophilic influx into parasitized organs. In evaluating the immunosuppressive effects, we identified a decreased number of CD4+ FOXP3+ T cells and reduced il10 expression in A2AR-/- infected mice. During ex vivo cell culture, A2AR-/- splenocytes produced smaller amounts of IL-10. In conclusion, we demonstrated that the A2AR signaling pathway is detrimental to development of Th1-type adaptive immunity and that this pathway could be associated with the regulatory process. In particular, it promotes parasite surveillance.

Nucleotide-binding oligomerization domain-containing protein 2 prompts potent inflammatory stimuli during Neospora caninum infection.

Neospora caninum is an apicomplexan parasite responsible for major economic losses due to abortions in cattle. Innate immune responses are crucial for host resistance against the infection, however the molecules involved in parasite recognition are still poorly understood. Nod2 is a cytosolic receptor that recognizes several pathogens and its role during N. caninum infection has not yet been described. In that sense, we evaluated the role of Nod2 in host response against this parasite. We found that infection of macrophages induced increased expression of Nod2, which colocalized with the parasites' vacuoles. Nod2-deficient macrophages showed an impaired induction of pro-inflammatory cytokines, increased production of modulatory molecules, and failure to restrict parasite replication. In vivo, Nod2-knockout mice showed a reduction of MAPK phosphorylation and proinflammatory cytokines, followed by decreased inflammation in target organs and increment in parasite burden. Surprisingly, these mice were partially resistant to lethal doses of tachyzoites. In addition, these phenomena were not observed in Rip2-/- mice. In conclusion, our study indicates that Nod2-dependent responses account for N. caninum elimination. On the other hand, the inflammatory milieu induced by this innate receptor provoked pathogenesis and death in severe experimental neosporosis.

GITR Activation Positively Regulates Immune Responses against Toxoplasma gondii.

Toxoplasma gondii is a widespread parasite responsible for causing clinical diseases especially in pregnant and immunosuppressed individuals. Glucocorticoid-induced TNF receptor (GITR), which is also known as TNFRS18 and belongs to the TNF receptor superfamily, is found to be expressed in various cell types of the immune system and provides an important costimulatory signal for T cells and myeloid cells. However, the precise role of this receptor in the context of T. gondii infection remains elusive. Therefore, the current study investigated the role of GITR activation in the immunoregulation mechanisms induced during the experimental infection of mice with T. gondii. Our data show that T. gondii infection slightly upregulates GITR expression in Treg cells and B cells, but the most robust increment in expression was observed in macrophages and dendritic cells. Interestingly, mice infected and treated with an agonistic antibody anti-GITR (DTA-1) presented a robust increase in pro-inflammatory cytokine production at preferential sites of parasite replication, which was associated with the decrease in latent brain parasitism of mice under treatment with DTA-1. Several in vivo and in vitro analysis were performed to identify the cellular mechanisms involved in GITR activation upon infection, however no clear alterations were detected in the phenotype/function of macrophages, Tregs and B cells under treatment with DTA-1. Therefore, GITR appears as a potential target for intervention during infection by the parasite Toxoplasma gondii, even though further studies are still necessary to better characterize the immune response triggered by GITR activation during T. gondii infection.

CCR2 signaling contributes to the differentiation of protective inflammatory dendritic cells in Leishmania braziliensis infection.

In vertebrate hosts, Leishmania braziliensis parasites infect mainly mononuclear phagocytic system cells, which when activated by T helper cell type 1 cytokines produce nitric oxide and kill the pathogens. Chemokine (C-C motif) receptor 2 is a chemokine receptor that binds primarily chemokine (C-C motif) ligand 2 and has an important role in the recruitment of monocytic phagocytes. Although it has been reported that Leishmania braziliensis infection induces CCR2 expression in the lesions, the role of CCR2 during Leishmania braziliensis infection remains unknown. Here, we showed that CCR2 has a role in mediating protection against Leishmania braziliensis infection in mice. The absence of CCR2 resulted in increased susceptibility to infection and was associated with low amounts of Ly6C(+) inflammatory dendritic cells in the lesions, which we found to be the major sources of tumor necrosis factor production and induced nitric oxide synthase expression in C57BL/6 mice lesions. Consequently, CCR2(-/-) mice showed decreased tumor necrosis factor production and induced nitric oxide synthase expression, resulting in impaired parasite elimination. We also demonstrated that CCR2 has a role in directly mediating the differentiation of monocytes into inflammatory dendritic cells at the infection sites, contributing to the accumulation of inflammatory dendritic cells in Leishmania braziliensis lesions and subsequent control of parasite replication. Therefore, these data provide new information on the role of chemokines during the immune response to infections and identify a potential target for therapeutic interventions in cutaneous leishmaniasis.

IL17 Promotes Mammary Tumor Progression by Changing the Behavior of Tumor Cells and Eliciting Tumorigenic Neutrophils Recruitment.

The aggressiveness of invasive ductal carcinoma (IDC) of the breast is associated with increased IL17 levels. Studying the role of IL17 in invasive breast tumor pathogenesis, we found that metastatic primary tumor-infiltrating T lymphocytes produced elevated levels of IL17, whereas IL17 neutralization inhibited tumor growth and prevented the migration of neutrophils and tumor cells to secondary disease sites. Tumorigenic neutrophils promote disease progression, producing CXCL1, MMP9, VEGF, and TNFα, and their depletion suppressed tumor growth. IL17A also induced IL6 and CCL20 production in metastatic tumor cells, favoring the recruitment and differentiation of Th17. In addition, IL17A changed the gene-expression profile and the behavior of nonmetastatic tumor cells, causing tumor growth in vivo, confirming the protumor role of IL17. Furthermore, high IL17 expression was associated with lower disease-free survival and worse prognosis in IDC patients. Thus, IL17 blockade represents an attractive approach for the control of invasive breast tumors.

Nucleosides present on phlebotomine saliva induce immunossuppression and promote the infection establishment.

BACKGROUND: Sand fly saliva plays a crucial role in establishing Leishmania infection. We identified adenosine (ADO) and adenosine monophosphate (AMP) as active pharmacologic compounds present in Phlebotomus papatasi saliva that inhibit dendritic cell (DC) functions through a PGE2/IL 10-dependent mechanism. METHODOLOGY/PRINCIPAL FINDINGS: Herein, we prepared a mixture of ADO and AMP in equimolar amounts similar to those present in the salivary-gland extract (SGE) form one pair of salivary glands of P. papatasi and co-injected it with Leishmania amazonensis or L. major into mouse ears. ADO+AMP mimicked exacerbative effects of P. papatasi saliva in leishmaniasis, increasing parasite burden and cutaneous lesions. Enzymatic catabolism of salivary nucleosides reversed the SGE-induced immunosuppressive effect associated with IL-10 enhancement. Immunosuppressive factors COX2 and IL-10 were upregulated and failed to enhance ear lesion and parasite burden in IL 10-/- infected mice. Furthermore, nucleosides increased regulatory T cell (Treg) marker expression on CD4+CD25- cells, suggesting induction of Tregs on effector T cells (T eff). Treg induction (iTreg) was associated with nucleoside-induced tolerogenic dendritic cells (tDCs) expressing higher levels of COX2 and IL-10. In vitro generation of Tregs was more efficient in DCs treated with nucleosides. Suppressive effects of nucleosides during cutaneous leishmaniasis were mediated through an A2AR-dependent mechanism. Using BALB/c mice deficient in A2A ADO receptor (A2AR-/-), we showed that co-inoculated mice controlled infection, displaying lower parasite numbers at infection sites and reduced iTreg generation. CONCLUSION/SIGNIFICANCE: We have demonstrated that ADO and AMP in P. papatasi saliva mediate exacerbative effects of Leishmania infection by acting preferentially on DCs promoting a tolerogenic profile in DCs and by generating iTregs in inflammatory foci through an A2AR mechanism.

Tr-1-like CD4+CD25-CD127-/lowFOXP3- cells are the main source of interleukin 10 in patients with cutaneous leishmaniasis due to Leishmania braziliensis.

CD4(+)CD25(+)FOXP3(+) regulatory T cells have long been shown to mediate susceptibility to Leishmania infection, mainly via interleukin 10 production. In this work, we showed that the main sources of interleukin 10 in peripheral blood mononuclear cells (PBMCs) from patients with cutaneous leishmaniasis due to Leishmania braziliensis are CD4(+)CD25(-)CD127(-/low)FOXP3(-) cells. Compared with uninfected controls, patients with CL had increased frequencies of circulating interleukin 10-producing CD4(+)CD25(-)CD127(-/low) cells, which efficiently suppressed tumor necrosis factor α production by the total PBMC population. Also, in CL lesions, interleukin 10 was mainly produced by CD4(+)CD25(-) cells, and interleukin 10 messenger RNA expression was associated with interleukin 27, interleukin 21, and interferon γ expression, rather than with FOXP3 or transforming growth factor ß expressions. Active production of both interleukin 27 and interleukin 21, together with production of interferon γ and interleukin 10, was also detected in the lesions. Since these cytokines are associated with the differentiation and activity of Tr-1 cells, our results suggest that this cell population may play an important role in the immunomodulation of CL. Therefore, development of treatments that interfere with this pathway may lead to faster parasite elimination.

Relevance of the myeloid differentiation factor 88 (MyD88) on RANKL, OPG, and nod expressions induced by TLR and IL-1R signaling in bone marrow stromal cells.

The myeloid differentiation factor 88 (MyD88) plays a pivotal role in Toll-like receptor (TLR)- and interleukin-1 receptor (IL-1R)-induced osteoclastogenesis. We examined the role of MyD88 on p38 mitogen-activated protein kinase (MAPK) and nuclear factor kappa-light-chain-enhancer of activated B cell (NF-κB) activation and nucleotide-binding oligomerization domain (Nod) induction by lipopolysaccharide (LPS) and IL-1 beta, and their effect on receptor activator of NF-κB ligand (RANKL) and osteoprotegerin (OPG) production in bone marrow stromal cell (BMSC). RANKL, Nod1, Nod2, NF-κB, and p38 protein levels were determined by Western blot. Nod2 was stimulated with muramyl dipeptide (MDP) prior to TLR4 stimulation with LPS. MyD88 deficiency markedly inhibited RANKL expression after LPS stimulation and increased OPG messenger RNA (mRNA) production. Also, MyD88 was necessary for NF-κB and p38 MAPK activation. MDP alone did not induce RANKL and OPG expressions; however, when combined with LPS, their expressions were significantly increased (p < 0.05). Our results support that MyD88 signaling has a pivotal role in osteoclastogenesis thought NF-κB and p38 activation. Nod2 and especially Nod1 levels were influenced by MyD88.

Role of dystrophin in acute Trypanosoma cruzi infection.

Previous studies have demonstrated loss/reduction of dystrophin in cardiomyocytes in both acute and chronic stages of experimental Trypanosoma cruzi (T. cruzi) infection in mice. The mechanisms responsible for dystrophin disruption in the hearts of mice acutely infected with T. cruzi are not completely understood. The present in vivo and in vitro studies were undertaken to evaluate the role of inflammation in dystrophin disruption and its correlation with the high mortality rate during acute infection. C57BL/6 mice were infected with T. cruzi and killed 14, 20 and 26 days post infection (dpi). The intensity of inflammation, cardiac expression of dystrophin, calpain-1, NF-κB, TNF-α, and sarcolemmal permeability were evaluated. Cultured neonatal murine cardiomyocytes were incubated with serum, collected at the peak of cytokine production and free of parasites, from T. cruzi-infected mice and dystrophin, calpain-1, and NF-κB expression analyzed. Dystrophin disruption occurs at the peak of mortality and inflammation and is associated with increased expression of calpain-1, TNF-α, NF-κB, and increased sarcolemmal permeability in the heart of T. cruzi-infected mice at 20 dpi confirmed by in vitro studies. The peak of mortality occurred only when significant loss of dystrophin in the hearts of infected animals occurred, highlighting the correlation between inflammation, dystrophin loss and mortality.

Expression of apoptosis mediators p53 and caspase 3, 7, and 9 in chronic rhinosinusitis with nasal polyposis.

BACKGROUND: The causal factor for the perpetuation of the inflammatory process in chronic rhinosinusitis with nasal polyps (CRSwNPs) has been extensively studied. However, little is known about the influence of cell death in this disease. Thus, the molecular assessment of mechanisms involved in apoptosis might shed light on the pathogenesis of CRSwNPs. This study was designed to evaluate the gene expression of different apoptotic factors in patients with NPs compared with control patients. METHODS: The mRNA expression of the apoptosis mediators caspase 3, 7, and 9 and of p53 protein was analyzed using quantitative reverse transcription-polymerase chain reaction in 25 NPs and 18 control samples. RESULTS: We observed significantly lower expression of p53 and caspase 3 and 9 in patients with CRSwNPs compared with the controls, whereas caspase 7 expression was not significantly different from the controls. CONCLUSION: The reduced expression of these apoptosis factors in CRSwNPs might be related to higher proliferation and the perpetuation of inflammatory cells hindering the control of the disease. A better understanding of the possible influence of apoptosis factors on CRSwNPs could provide rationale for future therapies.

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.

Influence of the CCR-5/MIP-1 α axis in the pathogenesis of Rocio virus encephalitis in a mouse model.

Rocio virus (ROCV) caused an outbreak of human encephalitis during the 1970s in Brazil and its immunopathogenesis remains poorly understood. CC-chemokine receptor 5 (CCR5) is a chemokine receptor that binds to macrophage inflammatory protein (MIP-1 α). Both molecules are associated with inflammatory cells migration during infections. In this study, we demonstrated the importance of the CCR5 and MIP-1 α, in the outcome of viral encephalitis of ROCV-infected mice. CCR5 and MIP-1 α knockout mice survived longer than wild-type (WT) ROCV-infected animals. In addition, knockout mice had reduced inflammation in the brain. Assessment of brain viral load showed mice virus detection five days post-infection in wild-type and CCR5-/- mice, while MIP-1 α-/- mice had lower viral loads seven days post-infection. Knockout mice required a higher lethal dose than wild-type mice as well. The CCR5/MIP-1 α axis may contribute to migration of infected cells to the brain and consequently affect the pathogenesis during ROCV infection.

Non-peptidic cruzain inhibitors with trypanocidal activity discovered by virtual screening and in vitro assay.

A multi-step cascade strategy using integrated ligand- and target-based virtual screening methods was developed to select a small number of compounds from the ZINC database to be evaluated for trypanocidal activity. Winnowing the database to 23 selected compounds, 12 non-covalent binding cruzain inhibitors with affinity values (K i) in the low micromolar range (3-60 µM) acting through a competitive inhibition mechanism were identified. This mechanism has been confirmed by determining the binding mode of the cruzain inhibitor Nequimed176 through X-ray crystallographic studies. Cruzain, a validated therapeutic target for new chemotherapy for Chagas disease, also shares high similarity with the mammalian homolog cathepsin L. Because increased activity of cathepsin L is related to invasive properties and has been linked to metastatic cancer cells, cruzain inhibitors from the same library were assayed against it. Affinity values were in a similar range (4-80 µM), yielding poor selectivity towards cruzain but raising the possibility of investigating such inhibitors for their effect on cell proliferation. In order to select the most promising enzyme inhibitors retaining trypanocidal activity for structure-activity relationship (SAR) studies, the most potent cruzain inhibitors were assayed against T. cruzi-infected cells. Two compounds were found to have trypanocidal activity. Using compound Nequimed42 as precursor, an SAR was established in which the 2-acetamidothiophene-3-carboxamide group was identified as essential for enzyme and parasite inhibition activities. The IC50 value for compound Nequimed42 acting against the trypomastigote form of the Tulahuen lacZ strain was found to be 10.6±0.1 µM, tenfold lower than that obtained for benznidazole, which was taken as positive control. In addition, by employing the strategy of molecular simplification, a smaller compound derived from Nequimed42 with a ligand efficiency (LE) of 0.33 kcal mol(-1) atom(-1) (compound Nequimed176) is highlighted as a novel non-peptidic, non-covalent cruzain inhibitor as a trypanocidal agent candidate for optimization.

Adjunct effect of the antimicrobial photodynamic therapy to an association of non-surgical and surgical periodontal treatment in modulation of gene expression: a human study.

BACKGROUND: This study has evaluated the effect of antimicrobial photodynamic therapy (aPDT) used in conjunction with non-surgical and surgical periodontal treatment (PT) in modulating gene expression during periodontal wound healing. METHODS: Fifteen patients with chronic periodontitis, presenting bilaterally lower molars with class III furcation lesions and scheduled for extraction, were selected. In initial therapy, scaling and root planing (SRP) was performed in the Control Group (CG), while SRP+aPDT were performed in the Test Group (TG). 45days later, flap surgery plus SRP, and flap surgery plus SRP+aPDT were performed in the CG and TG, respectively. At 21days post-surgery, the newly formed granulation tissue was collected, and Real-time PCR evaluated the expression of the genes: tumor necrosis factor-α, interleukin-1ß, interleukin-4, interleukin-10, matrix metalloproteinase-2 (MMP-2), tissue inhibitor of metalloproteinase-2 (TIMP-2), osteoprotegerin (OPG), receptor activator of nuclear factor-κB ligand (RANKL), type I collagen, alkaline phosphatase, osteopontin, osteocalcin, and bone sialoprotein. RESULTS: There were statistically significant differences between the groups in relation to mRNA levels for MMP-2 (TG=3.26±0.89; CG=4.23±0.97; p=0.01), TIMP-2/MMP-2 ratio (TG=0.91±0.34; CG=0.73±0.32; p=0.04), OPG (TG=0.84±0.45; CG=0.30±0.26; p=0.001), and OPG/RANKL ratio (TG=0.60±0.86; CG=0.23±0.16; p=0.04), favoring the TG. CONCLUSION: The present data suggest that the aPDT associated to nonsurgical and surgical periodontal therapy may modulate the extracellular matrix and bone remodeling by up regulating the TIMP- 2/MMP-2 and OPG/RANKL mRNA ratio, but the clinical relevance needs to be evaluated in further studies.