Estudo da inflamação e da autoimunidade na doença hepática gordurosa não alcoólica

Pós-graduação em Fisiopatologia em Clínica Médica - FMB

Análise microbiológica e imunologica do fluido gengival e proteoma salivar de individuos com Sindrome de Down com doença periodontal

Pós-graduação em Odontologia - FOAR

Evaluation of clinical and inflammatory markers of subclinical carotid atherosclerosis in postmenopausal women

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Synergistic Anti-Inflammatory Activity of the Antimicrobial Peptides Human Beta-Defensin-3 (hBD-3) and Cathelicidin (LL-37) in a Three-Dimensional Co-Culture Model of Gingival Epithelial Cells and Fibroblasts

Given the spread of antibiotic resistance in bacterial pathogens, antimicrobial peptides that can also modulate the immune response may be a novel approach for effectively controlling periodontal infections. In the present study, we used a three-dimensional (3D) co-culture model of gingival epithelial cells and fibroblasts stimulated with Aggregatibacter actinomycetemcomitans lipopolysaccharide (LPS) to investigate the anti-inflammatory properties of human beta-defensin-3 (hBD-3) and cathelicidin (LL-37) and to determine whether these antimicrobial peptides can act in synergy. The 3D co-culture model composed of gingival fibroblasts embedded in a collagen matrix overlaid with gingival epithelial cells had a synergistic effect with respect to the secretion of IL-6 and IL-8 in response to LPS stimulation compared to fibroblasts and epithelial cells alone. The 3D co-culture model was stimulated with non-cytotoxic concentrations of hBD-3 (10 and 20 mu M) and LL-37 (0.1 and 0.2 mu M) individually and in combination in the presence of A. actinomycetemcomitans LPS. A multiplex ELISA assay was used to quantify the secretion of 41 different cytokines. hBD-3 and LL-37 acted in synergy to reduce the secretion of GRO-alpha, G-CSF, IP-10, IL-6, and MCP-1, but only had an additive effect on reducing the secretion of IL-8 in response to A. actinomycetemcomitans LPS stimulation. The present study showed that hBD-3 acted in synergy with LL-37 to reduce the secretion of cytokines by an LPS-stimulated 3D model of gingival mucosa. This combination of antimicrobial peptides thus shows promising potential as an adjunctive therapy for treating inflammatory periodontitis.

Evaluation of Toll-Like Receptor 2 and 4 RNA Expression and the Cytokine Profile in Postmenopausal Women with Metabolic Syndrome

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Associação entre expressão de receptores TLR2 e TLR4 e produção de TNF-alpha e IL-10 por monócitos de gestantes portadoras de pré-eclâmpsia

Preeclampsia (PE) is a pregnancy specific syndrome characterized by a systemic inflammatory response, with higher intensity than that observed in normal pregnancy. Cells of the immune system, such as monocytes and granulocytes are endogenously activated and secrete high levels of free radicals and inflammatory cytokines. The objective of this study was to assess the activation state of monocytes from pregnant women with preeclampsia by endogenous expression of TLR2 e TLR4 receptors and to correlate the expression of TLR2 and TLR4 on monocytes surface of pregnant women with PE with the production of tumor necrosis factor-alpha (TNF- and interleukin-10 (IL-10) by these cells stimulated or not with peptidoglycan (PG) and lipopolysaccharide (LPS), as agonists agents of TLR2 and TLR4, respectively. We evaluated 15 pregnant women with PE, 15 normotensive pregnant women (NT) and 15 non-pregnant (NP). Peripheral blood monocytes were incubates in the presence or absence of LPS or PG. The supernatant obtained after 18h of culture was aspirated and used for TNF- and IL-10 determination by enzyme immunoassay (ELISA). The endogenous expression of TLR2 and TLR4 receptors was evaluated by flow cytometry. Our results showed significant highly concentrations of TNF- and TLR4 expression in monocytes of preeclamptic women when compared with NT and NP. Normal pregnant women presented higher levels of IL-10 in comparison with PE and NP groups. TLR2 expression was similar in the three groups studied. Therefore, our study highlights the important role of TLR4 in PE and the consequent high production of TNF- by monocytes of these patients, as well as the potential mechanism involving low levels of IL-10 in the pathophysiology of the disease. These observations demonstrate the strong link between the pathology of PE and the immune system of these patients

Antitumor and Anti-Inflammatory Effects of Palladium(II) Complexes on Ehrlich Tumour

Palladium(II) complexes are an important class of cyclopalladated compounds that play a pivotal role in various pharmaceutical applications. Here, we investigated the antitumour, anti-infl ammatory, and mutagenic effects of two complexes: [Pd(dmba)(Cl)tu] (1) and [Pd(dmba)(N3)tu] (2) (dmba = N,N-dimethylbenzylamine and tu = thiourea), on Ehrlich ascites tumour (EAT) cells and peritoneal exudate cells (PECs) from mice bearing solid Ehrlich tumour. The cytotoxic effects of the complexes on EAT cells and PECs were assessed using the 3-(4,5-dimethylthiazol-3-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay. The effects of the complexes on the immune system were assessed based on the production of nitric oxide (NO) (Griess assay) and tumour necrosis factor-Į (TNF-Į), interleukin-12 (IL-12), and interleukin-10 (IL-10) (ELISA). Finally the mutagenic activity was assessed by the Ames test using the Salmonella typhimurium strain TA 98. Cisplatin was used as a standard. The IC50 ranges for the growth inhibition of EAT cells and PECs were found to be (72.8 ± 3.23) µM and (137.65 ± 0.22) µM for 1 and (39.7 ± 0.30) µM and (146.51 ± 2.67) µM for 2, respectively. The production of NO, IL-12, and TNF-Į, but not IL-10, was induced by both complexes and cisplatin. The complexes showed no mutagenicity in vitro, unlike cisplatin, which was mutagenic in the strain. These results indicate that the complexes are not mutagenic and have potential immunological and antitumour activities. These properties make them promising alternatives to cisplatin.

Modulation of Immune Response by RAGE and TLR4 Signalling in PBMCs of Diabetic and Non-Diabetic Patients

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Anti-inflammatory effect of MAPK phosphatase-1 local gene transfer in inflammatory bone loss

Alveolar bone loss associated with periodontal diseases is the result of osteoclastogenesis induced by bacterial pathogens. The mitogen-activated protein kinase (MAPK) phosphatase 1 (MKP-1) is a critical negative regulator of immune response as a key phosphatase capable of dephosphorylating activated MAPKs. In this study, rat macrophages transduced with recombinant adenovirus (Ad.)MKP-1 specifically dephosphorylated activated MAPKs induced by lipopolysaccharide (LPS) compared with control cells. Bone marrow macrophages from MKP-1 knockout (KO) mice exhibited higher interleukin (IL)-6, IL-10, tumor necrosis factor (TNF)-α, and select chemokine compared with wild-type (WT) mice when stimulated by LPS. In addition, bone marrow cultures from MKP-1 KO mice exhibited significantly more osteoclastogenesis induced by LPS than when compared with WT mice. Importantly, MKP-1 gene transfer in bone marrow cells of MKP-1 KO mice significantly decreased IL-6, IL-10, TNF-α and chemokine levels, and formed fewer osteoclasts induced by LPS than compared with control group of cells. Furthermore, MKP-1 gene transfer in an experimental periodontal disease model attenuated bone resorption induced by LPS. Histological analysis confirmed that periodontal tissues transduced with Ad. MKP-1 exhibited less infiltrated inflammatory cells, less osteoclasts and less IL-6 than compared with rats of control groups. These studies indicate that MKP-1 is a key therapeutic target to control of inflammation-induced bone loss.

Silencing mitogen-activated protein kinase-activated protein kinase-2 arrests inflammatory bone loss

p38 mitogen-activated protein kinases (MAPKs) are critical for innate immune signaling and subsequent cytokine expression in periodontal inflammation and bone destruction. In fact, previous studies show that systemic p38 MAPK inhibitors block periodontal disease progression. However, development of p38 MAPK inhibitors with favorable toxicological profiles is difficult. Here, we report our findings regarding the contribution of the downstream p38 MAPK substrate, mitogen-activated protein kinase-activated protein kinase 2 (MK2 or MAPKAPK-2), in immune response modulation in an experimental model of pathogen-derived lipopolysaccharide (LPS)-induced periodontal bone loss. To determine whether small interfering RNA (siRNA) technology has intraoral applications, we initially validated MK2 siRNA specificity. Then, gingival tissue surrounding maxillary molars of rats was injected with MK2 siRNA or scrambled siRNA at the palatal regions of bone loss. Intraoral tissues treated with MK2 siRNA had significantly less MK2 mRNA expression compared with scrambled siRNA-treated tissues. MK2 siRNA delivery arrested LPS-induced inflammatory bone loss, decreased inflammatory infiltrate, and decreased osteoclastogenesis. This proof-of-concept study suggests a novel target using an intraoral RNA interference strategy to control periodontal inflammation.

The effect of conventional mechanical periodontal treatment on red complex microorganisms and clinical parameters in Down syndrome periodontitis patients: a pilot study

Periodontal disease (PD) is induced by a complex microbiota, such as Porphyromonas gingivalis, Tannerella forsythia, and Treponema denticola (together called the red complex), which triggers intense inflammatory reaction. Down syndrome (DS) individuals demonstrate a high prevalence of PD compared with those who are otherwise chromosomally normal (euploids). This pilot study aimed to evaluate the effect of non-surgical periodontal treatment in DS chronic periodontitis patients on clinical and microbiological parameters. Patients with chronic periodontitis, 23 DS and 12 euploids (control group), were submitted to non-surgical mechanical periodontal treatment, followed by maintenance for 45 days. Clinical parameters after periodontal treatment were similar in diseased and healthy sites, independent of the genetic background. Diseased sites of DS and control patients harbored similar levels of P. gingivalis and T. forsythia at baseline, but significantly higher levels of T. denticola were found in DS patients. Increased levels of P. gingivalis at healthy sites were found in DS individuals. Non-surgical periodontal therapy decreased the levels of red complex microorganisms and improved the tested clinical parameters of diseased sites in both groups. However, the levels of red complex bacteria were higher in diseased sites of DS patients after the periodontal treatment. We conclude in this pilot study that, although the mechanical periodontal treatment seemed to be effective in DS subjects over a short-term period, the red complex bacteria levels did not decrease significantly in diseased sites, as occurred in controls. Therefore, for DS patients, it seems that the conventional non-surgical periodontal therapy should be improved by utilizing adjuvants to reduce the presence of periodontopathogens.

Effects of omega-3 on metabolic markers in postmenopausal women with metabolic syndrome

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Modulation of immune response by RAGE and TLR4 signalling in PBMCs of diabetic and non-diabetic patients

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Oxidative stress and inflammatory response biomarkers in dogs with mammary carcinoma

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Anti-inflammatory effects of Brazilian ginseng (Pfaffia paniculata) on TNBS-induced intestinal inflammation: experimental evidence

Inflammatory bowel disease (IBD) is a chronic, relapsing, idiopathic inflammation of the gastrointestinal tract. Clinical studies suggest that the initiation of IBD is multifactorial, involving genetics, the immune system and environmental factors, such as diet, drugs and stress. Pfaffia paniculata is an adaptogenic medicinal plant used in Brazilian folk medicine as an anti-stress agent. Thus, we hypothesised that the P. paniculata enhances the response of animals subjected to colonic inflammation. Our aim was to investigate the intestinal anti-inflammatory activity of P. paniculata in rats before or after induction of intestinal inflammation using trinitrobenzenesulfonic acid (TNBS). The animals were divided into groups that received the vehicle, prednisolone or P. paniculata extract daily starting 14days before or 7days after TNBS induction. At the end of the procedure, the animals were killed and their colons were assessed for the macroscopic damage score (MDS), extent of the lesion (EL) and weight/length ratio, myeloperoxidase (MPO) activity and glutathione (GSH), cytokines and C-reactive protein (CRP) levels. Histological evaluation and ultrastructural analysis of the colonic samples were performed. Treatment with the 200mg/kg dose on the curative schedule was able to reduce the MDS and the EL. In addition, MPO activity was reduced, GSH levels were maintained, and the levels of pro-inflammatory cytokines and CRP were decreased. In conclusion, the protective effect of P. paniculata was related to reduced oxidative stress and CRP colonic levels, and due to immunomodulatory activity as evidenced by reduced levels of IL-1β, INF-γ, TNF-α and IL-6.