Hypertonic saline solution reduces the inflammatory response in endotoxemic rats

OBJECTIVE: Volume replacement in septic patients improves hemodynamic stability. This effect can reduce the inflammatory response. The objective of this study was to evaluate the effect of 7.5% hypertonic saline solution versus 0.9% normal saline solution for volume replacement during an inflammatory response in endotoxemic rats. METHODS: We measured cytokines (serum and gut), nitrite, and lipid peroxidation (TBARS) as indicators of oxidative stress in the gut. Rats were divided into four groups: control group (C) that did not receive lipopolysaccharide; lipopolysaccharide injection without treatment (LPS); lipopolysaccharide injection with saline treatment (LPS + S); and lipopolysaccharide injection with hypertonic saline treatment (LPS + H). Serum and intestine were collected. Measurements were taken at 1.5, 8, and 24 h after lipopolysaccharide administration. RESULTS: Of the four groups, the LPS + H group had the highest survival rate. Hypertonic saline solution treatment led to lower levels of IL-6, IL-10, nitric oxide, and thiobarbituric acid reactive substances compared to 0.9% normal saline. In addition, hypertonic saline treatment resulted in a lower mortality compared to 0.9% normal saline treatment in endotoxemic rats. Volume replacement reduced levels of inflammatory mediators in the plasma and gut. CONCLUSION: Hypertonic saline treatment reduced mortality and lowered levels of inflammatory mediators in endotoxemic rats. Hypertonic saline also has the advantage of requiring less volume replacement.FAPESP-09/03338-7FAPESP-06/00443-6CNPQ-470744/2004-

Caryocar brasiliense camb protects against genomic and oxidative damage in urethane-induced lung carcinogenesis

The antioxidant effects of Caryocar brasiliense Camb, commonly known as the pequi fruit, have not been evaluated to determine their protective effects against oxidative damage in lung carcinogenesis. In the present study, we evaluated the role of pequi fruit against urethane-induced DNA damage and oxidative stress in forty 8-12 week old male BALB/C mice. An in vivo comet assay was performed to assess DNA damage in lung tissues and changes in lipid peroxidation and redox cycle antioxidants were monitored for oxidative stress. Prior supplementation with pequi oil or its extract (15 µL, 60 days) significantly reduced urethane-induced oxidative stress. A protective effect against DNA damage was associated with the modulation of lipid peroxidation and low protein and gene expression of nitric oxide synthase. These findings suggest that the intake of pequi fruit might protect against in vivo genotoxicity and oxidative stress

Gastrin-Releasing Peptide Receptor Antagonism Induces Protection from Lethal Sepsis: Involvement of Toll-like Receptor 4 Signaling

In sepsis, toll-like receptor (TLR)-4 modulates the migration of neutrophils to infectious foci, favoring bacteremia and mortality. In experimental sepsis, organ dysfunction and cytokines released by activated macrophages can be reduced by gastrin-releasing peptide (GRP) receptor (GRPR) antagonist RC-3095. Here we report a link between GRPR and TLR-4 in experimental models and in sepsis patients. RAW 264.7 culture cells were exposed to lipopolysaccharide (LPS) or tumor necrosis factor (TNF)-alpha and RC-3095 (10 ng/mL), Male Wistar rats were subjected to cecal ligation and puncture (CLP), and RC-3095 was administered (3 mg/kg, subcutaneously); after 6 h, we removed the blood, bronchoalveolar lavage, peritoneal lavage and lung. Human patients with a clinical diagnosis of sepsis received a continuous infusion with RC-3095 (3 mg/kg, intravenous) over a period of 12 h, and plasma was collected before and after RC-3095 administration and, in a different set of patients with systemic inflammatory response syndrome (SIRS) or sepsis. GRP plasma levels were determined. RC-3095 inhibited TLR-4, extracellular-signal-related kinase (ERK)-1/2, Jun NH2-terminal kinase (JNK) and Akt and decreased activation of activator protein 1 (AP-1), nuclear factor (NF)-kappa B and interleukin (IL)-6 in macrophages stimulated by LPS. It also decreased IL-6 release from macrophages stimulated by TNF-alpha. RC-3095 treatment in CLP rats decreased lung TLR-4, reduced the migration of cells to the lung and reduced systemic cytokines and bacterial dissemination. Patients with sepsis and systemic inflammatory response syndrome have elevated plasma levels of GRP which associates with clinical outcome in the sepsis patients. These findings highlight the role of GRPR signaling in sepsis outcome and the beneficial action of GRPR antagonists in controlling the inflammatory response in sepsis through a mechanism involving at least inhibition of TLR-4 signaling. Online address: http://www.molmed.org doi: 10.2119/molmed.2012.0008

Role of B-1 cell in inflammatory response to lipopolysaccharid

A sepse é a Síndrome da Resposta Inflamatória Sistêmica decorrente de uma infecção por gram positivos/negativos, fungos ou vírus. É caracterizada por alta liberação de mediadores inflamatórios podendo levar à morte. As células B-1 são encontradas em cavidades peritoneal e pleural de camundongos e sua origem e função ainda não são completamente conhecidas. Apresentam marcadores de superfície de linhagem mielóide e linfóide e migram para focos inflamatórios comportando-se como macrófagos. Objetivo: investigar o papel da célula B-1 na resposta inflamatória após estímulo com lipopolissacáride (LPS) in vitro e in vivo. Métodos: TNF-, IL-6, IL-10 (ELISA) e nitrito (Griess) foram dosados em sobrenadante de cultura celular (106 cel./ml). As células em cultura receberam por 24h de estímulo com 10 g/mL de LPS de Escherichia coli (026:B6 Sigma®). Foram realizados os seguintes grupos cultura de célula B-1 (Balb/c), cultura de macrófagos de linhagem (RAW 264.7) coculturas (macrófagos de linhagem RAW 264.7 e células B-1 (Balb/c, C57BL/6 e C57BL/6 IL-10 -/-), e células peritoneais de camundongos Balb/c e Balb/Xid (imunodeficiente em célula B-1) A endotoxemia foi induzida com injeção de LPS 15 mg/kg (i.p.) em camundongos Balb/c e Balb/Xid. Foram quantificados, TNF-, IL-6, IL-10 e nitrito em soro, pulmão e intestino dos animais após 1,5, 4 e 6 horas após a injeção de LPS. Ensaios de inoculação de células B-1 (Balb/c) em camundongos Balb/Xid foram realizados, e curva de sobrevida foi analisada após indução de endotoxemia. Resultados: Após o estímulo com LPS, células B-1 produziram IL-10 e a presença destas células em cocultura com macrófago promoveu a diminuição na produção de TNF-, IL-6, Nitrito e aumento de IL-10. Contudo, célula B-1 (IL-10 -/-) em cocultura com macrófagos, não inibem a produção de mediadores pro inflamatórios. Análise com macrófagos peritoneais de camundongo Balb/Xid e Balb/c após estímulo com LPS em cultura mostrou reprodução do fenômeno encontrado com os experimentos com cultura de célula imortalizada, isto é, maior produção de TNF-, IL-6 e NO em Balb/Xid (B-1 deficiente). Os estudos in vivo mostraram 60% de mortalidade em camundongo Balb/Xid comparando com Balb/c (0%) após 16 horas de injeção de LPS. Nos animais Balb/Xid encontramos padrão pro inflamatório exacerbado com maiores concentrações de TNF-, IL-6 e menores concentrações de IL-10 no plasma e tecidos quando comparamos com Balb/c. Conclusões: Nossos dados mostraram que a presença de células B-1 promoveram diminuição de mediadores pro inflamatórios e aumento de IL-10 em coculturas com macrófagos e que a modulação da resposta inflamatória pode ser devida a secreção de IL-10 pela célula B-1. Este padrão de resposta pro inflamatória se repete in vivo e é a possível causadora da maior taxa de mortalidade em camundongos da linhagem Balb/Xid.Sepsis syndrome is caused by inappropriate immune activation due to bacteria and bacterial components released during infection. This syndrome is the leading cause of death in intensive care units. Specialized B-lymphocytes located in the peritoneal and pleural cavities are known as B-1 cells. These cells produce IgM and IL-10, both of which are potent regulators of cell-mediated immunity. It has been suggested that B-1 cells modulate the systemic inflammatory response in sepsis. In this study, we conducted in vitro and in vivo experiments in order to investigate a putative role of B-1 cells in a murine model of LPS-induced sepsis. Macrophages and B-1 cells were studied in monocultures and in co-cultures. The B-1 cells produced the anti-inflammatory cytokine IL-10 in response to LPS. In the B-1 cell-macrophage co-cultures, production of proinflammatory mediators (TNF-, IL-6 and nitrite) was lower than in the macrophage monocultures, whereas that of IL-10 was higher in the co-cultures. Co-culture of B-1 IL-10/ cells and macrophages did not reduce the production of the proinflammatory mediators (TNF-, IL-6 and nitrite). After LPS injection, the mortality rate was higher among Balb/Xid mice, which are B-1 cell deficient, than among wild-type mice (65.0% vs. 0.0%). The Balb/Xid mice also presented a proinflammatory profile of TNF-, IL-6 and nitrite, as well as lower levels of IL-10. In the early phase of LPS stimulation, B-1 cells modulate the macrophage inflammatory response, and the main molecular pathway of that modulation is based on IL-10-mediated intracellular signaling

Cytokine and chemokine levels in the heart tissue of aged rats following severe acute pancreatitis

Severe acute pancreatitis (AP) is a disease associated with high mortality and characterized by overwhelming systemic inflammation. Older people have a prolonged hospital stay and worst prognosis, when affected by this disease. Our group hypothesized, thus, that the systemic inflammatory response in the elderly would promote more organ damage when compared to the young. We sought to investigate the effect of systemic inflammation on the gene expression of cytokines, chemokines, and growth factors in the hearts of older and younger rats in an animal model of AP. AP was induced in all rats by injection of 0.5 mL of 2.5% taurocholate. There were two healthy age-matched control groups. An array of 79 cytokines, chemokines, and growth factors was measured in samples of cardiac tissue taken from the AP rats after 10 h, and from control rats. Older healthy rats had significantly higher levels of interleukin-10 (IL-10) and CCL1 gene expression than younger ones ( P  < 0.05), but all other measurements were similar among the study groups. This study indicates the systemic inflammation may show unique features for different organs in the body, but older animals with systemic inflammation are similar to the young regarding the cardiac inflammatory response

Effect of phototherapy with low intensity laser on local and systemic immunomodulation following focal brain damage in rat

Brain injury is responsible for significant morbidity and mortality in trauma patients, but controversy still exists over therapeutic management for these patients. The objective of this study was to analyze the effect of phototherapy with low intensity lasers on local and systemic immunomodulation following cryogenic brain injury. Laser phototherapy was applied (or not-controls) immediately after cryogenic brain injury performed in 51 adult male Wistar rats. The animals were irradiated twice (3 h interval), with continuous diode laser (gallium-aluminum-arsenide (GaAlAs), 780 nm, or indium-gallium-aluminum-phosphide (InGaAlP), 660 nm) in two points and contact mode, 40 mW, spot size 0.042 cm(2), 3 J/cm(2) and 5 J/cm(2) (3 s and 5 s, respectively). The experimental groups were: Control (non-irradiated), RL3 (visible red laser/ 3 J/cm(2)), RL5 (visible red laser/5 J/cm(2)), IRL3 (infrared laser/ 3 J/cm(2)), IRL5 (infrared laser/5 J/cm(2)). The production of interleukin-1IL-1 beta (IL-1 beta), interleukin6 (IL-6), interleukin-10 (IL-10), and tumor necrosis factor-alpha (TNF-alpha) was analyzed by enzyme immunoassay technique (ELISA) test in brain and blood samples. The IL-1 beta concentration in brain of the control group ;was significantly reduced in 24 h (p < 0.01). This reduction was also observed in the RL5 and IRL3 groups. The TNF-alpha and IL-6 concentrations increased significantly (p < 0.01 and p < 0.05, respectively) in the blood of all groups, except by the IRL3 group. The IL-6 levels in RL3 group were significantly smaller than in control group in both experimental times. IL-10 concentration was maintained stable in all groups in brain and blood. Under the conditions of this study, it is possible to conclude that the laser phototherapy can affect TNF-alpha, IL-1 beta and IL-6 levels in the brain and in circulation in the first 24 h following cryogenic brain injury. (C) 2009 Elsevier B.V. All rights reserved.Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP), Sao Paulo, BrazilCoordenacao de Aperfeicoamento de Pessoal de Nivel Superior (CAPES), Brasilia, DF, Brazi

Angiotensin II modulates CD40 expression in vascular smooth muscle cells

The signalling pathway CD40/CD40L (CD40 ligand) plays an important role in atherosclerotic plaque formation and rupture. AngII (angiotensin II), which induces oxidative stress and inflammation, is also implicated in the progression of atherosclerosis. In the present study, we tested the hypothesis that AngII increases CD40/CD40L activity in vascular cells and that ROS (reactive oxygen species) are part of the signalling cascade that controls CD40/CD40L expression. Human CASMCs (coronary artery smooth muscle cells) in culture exposed to IL (interleukin)-1 beta or TNF-alpha (tumour necrosis factor-a) had increased superoxide generation and enhanced CD40 expression, detected by EPR (electron paramagnetic resonance) and immunoblotting respectively. Both phenomena were abolished by previous incubation with membrane-permeant antioxidants or cell transfection with P22(phox) antisense. AngII (50-200 nmol/l) induced an early and sustained increase in CD40 mRNA and protein expression in CASMCs, which was blocked by treatment with antioxidants. Increased CD40 expression led to enhanced activity of the pathway, as AngII-treated cells stimulated with recombinant CD40L released higher amounts of IL-8 and had increased COX-2 (cyclo-oxygenase-2) expression. We conclude that AngII stimulation of vascular cells leads to a ROS-dependent increase in CD40/CD40L signalling pathway activity. This phenomenon may be an important mechanism modulating the arterial injury observed in atherosclerosis-related vasculopathy.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)FAPESP Fundaqao de Amparo a Pesquisa do Estado de Sao Paulo[02/02930-0]Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)CNPq Conselho Nacional de Desenvolvimento Cientifico e TecnologicoFundacao Faculdade de Medicina e DIREX-LIMFundacao Faculdade de Medicina e DIREX-LI