Adriana Lino-dos-Santos-Franco

Formaldehyde induces lung inflammation by an oxidant and antioxidant enzymes mediated mechanism in the lung tissue

Formaldehyde (FA) is an indoor and outdoor pollutant widely used by many industries, and its exposure is associated with inflammation and oxidative stress in the airways. Our previous studies have demonstrated the role of reactive oxygen species (ROS) in lung inflammation induced by FA inhalation but did not identify source of the ROS. In the present study, we investigate the effects of FA on the activities and gene expression of glutathione peroxidase (GPX), glutathione reductase (GR), glutathione S-transferase (GST), superoxide dismutase (SOD) 1 and 2, catalase (CAT), nitric oxide synthase (iNOS and cNOS) and cyclooxygenase (COX) 1 and 2. The hypothesized link between NADPH-oxidase, nitric oxide synthase and cyclooxygenase, the lung inflammation after FA inhalation was also investigated. For experiments, male Wistar rats were submitted to FA inhalation (1%, 90 min daily) for 3 consecutive days. The treatments with apocynin and indomethacin before the FA exposure reduced the number of neutrophils recruited into the lung. Moreover, the treatments with apocynin and indomethacin blunted the effect of FA on the generation of IL-1β, while the treatments with L-NAME and apocynin reduced the generation of IL-6 by lung explants when compared to the untreated group. FA inhalation increased the levels of NO and hydrogen peroxide by BAL cells cultured and the treatments with apocynin and l-NAME reduced these generations. FA inhalation did not modify the activities of GPX, GR, GST and CAT but reduced the activity of SOD when compared to the naïve group. Significant increases in SOD-1 and -2, CAT, iNOS, cNOS and COX-1 expression were observed in the FA group compared to the naïve group. The treatments with apocynin, indomethacin and L-NAME reduced the gene expression of antioxidant and oxidant enzymes. In conclusion, our results indicate that FA causes a disruption of the physiological balance between oxidant and antioxidant enzymes in lung tissue, most likely favoring the oxidant pathways and thus positively modulating lung inflammation.

Exposure to low doses of formaldehyde during pregnancy suppresses the development of allergic lung inflammation in offspring.

Formaldehyde (FA) is an environmental and occupational pollutant, and its toxic effects on the immune system have been shown. Nevertheless, no data are available regarding the programming mechanisms after FA exposure and its repercussions for the immune systems of offspring. In this study, our objective was to investigate the effects of low-dose exposure of FA on pregnant rats and its repercussion for the development of allergic lung inflammation in offspring. Pregnant Wistar rats were assigned in 3 groups: P (rats exposed to FA (0.75 ppm, 1 h/day, 5 days/week, for 21 days)), C (rats exposed to vehicle of FA (distillated water)) and B (rats non-manipulated). After 30 days of age, the offspring was sensitised with ovalbumin (OVA)-alum and challenged with aerosolized OVA (1%, 15 min, 3 days). After 24 h the OVA challenge the parameters were evaluated. Our data showed that low-dose exposure to FA during pregnancy induced low birth weight and suppressed the development of allergic lung inflammation and tracheal hyperresponsiveness in offspring by mechanisms mediated by reduced anaphylactic antibodies synthesis, IL-6 and TNF-alpha secretion. Elevated levels of IL-10 were found. Any systemic alteration was detected in the exposed pregnant rats, although oxidative stress in the uterine environment was evident at the moment of the delivery based on elevated COX-1 expression and reduced cNOS and SOD-2 in the uterus. Therefore, we show the putative programming mechanisms induced by FA on the immune system for the first time and the mechanisms involved may be related to oxidative stress in the foetal microenvironment.

Amphetamine modulates cellular recruitment and airway reactivity in a rat model of allergic lung inflammation.

Asthma is characterized by pulmonary cellular infiltration, vascular exudation and airway hyperresponsiveness. Several drugs that modify central nervous system (CNS) activity can modulate the course of asthma. Amphetamine (AMPH) is a highly abused drug that presents potent stimulating effects on the CNS and has been shown to induce behavioral, biochemical and immunological effects. The purpose of this study was to investigate the effects of AMPH on pulmonary cellular influx, vascular permeability and airway reactivity. AMPH effects on adhesion molecule expression, IL-10 and IL-4 release and mast cell degranulation were also studied. Male Wistar rats were sensitized with ovalbumin (OVA) plus alum via subcutaneous injection. One week later, the rats received another injection of OVA-alum (booster). Two weeks after this booster, the rats were subjected to AMPH treatment 12 h prior to the OVA airway challenge. In rats treated with AMPH, the OVA challenge reduced cell recruitment into the lung, the vascular permeability and the cellular expression of ICAM-1 and Mac-1. Additionally, elevated levels of IL-10 and IL-4 were found in samples of lung explants from allergic rats. AMPH treatment, in comparison, increased IL-10 levels but reduced those of IL-4 in the lung explants. Moreover, the tracheal responsiveness to methacholine (MCh), as well as to an in vitro OVA challenge, was reduced by AMPH treatment, and levels of PCA titers were not modified by the drug. Our findings suggest that single AMPH treatment down-regulates several parameters of lung inflammation, such as cellular migration, vascular permeability and tracheal responsiveness. These results also indicate that AMPH actions on allergic lung inflammation include endothelium-leukocyte interaction mechanisms, cytokine release and mast cell degranulation.

Low Level Laser Therapy Reduces the Development of Lung Inflammation Induced by Formaldehyde Exposure.

Lung diseases constitute an important public health problem and its growing level of concern has led to efforts for the development of new therapies, particularly for the control of lung inflammation. Low Level Laser Therapy (LLLT) has been highlighted as a non-invasive therapy with few side effects, but its mechanisms need to be better understood and explored. Considering that pollution causes several harmful effects on human health, including lung inflammation, in this study, we have used formaldehyde (FA), an environmental and occupational pollutant, for the induction of neutrophilic lung inflammation. Our objective was to investigate the local and systemic effects of LLLT after FA exposure. Male Wistar rats were exposed to FA (1%) or vehicle (distillated water) during 3 consecutive days and treated or not with LLLT (1 and 5 hours after each FA exposure). Non-manipulated rats were used as control. 24 h after the last FA exposure, we analyzed the local and systemic effects of LLLT. The treatment with LLLT reduced the development of neutrophilic lung inflammation induced by FA, as observed by the reduced number of leukocytes, mast cells degranulated, and a decreased myeloperoxidase activity in the lung. Moreover, LLLT also reduced the microvascular lung permeability in the parenchyma and the intrapulmonary bronchi. Alterations on the profile of inflammatory cytokines were evidenced by the reduced levels of IL-6 and TNF-α and the elevated levels of IL-10 in the lung. Together, our results showed that LLLT abolishes FA-induced neutrophilic lung inflammation by a reduction of the inflammatory cytokines and mast cell degranulation. This study may provide important information about the mechanisms of LLLT in lung inflammation induced by a pollutant.

In vivo hydroquinone exposure causes tracheal hyperresponsiveness due to TNF secretion by epithelial cells

Hydroquinone (HQ) is the main oxidative substance in cigarette smoke and a toxic product of benzene biotransformation. Although the respiratory tract is an inlet pathway of HQ exposure, its effect on airway muscle responsiveness has not been assessed. We thus investigated the effects of low dose in vivo HQ-exposure on tracheal responsiveness to a muscarinic receptor agonist. Male Swiss mice were exposed to aerosolised 5% ethanol/saline solution (HQ vehicle; control) or 0.04 ppm HQ (1h/day for 5 days) and tracheal rings were collected 1h after the last exposure. HQ exposure caused tracheal hyperresponsiveness to methacholine (MCh), which was abolished by mechanical removal of the epithelium. This hyperresponsiveness was not dependent on neutrophil infiltration, but on tumour necrosis factor (TNF) secretion by epithelial cells. This conclusion was based on the following data: (1) trachea from HQ-exposed mice presented a higher amount of TNF, which was abrogated following removal of the epithelium; (2) the trachea hyperresponsiveness and TNF levels were attenuated by in vivo chlorpromazine (CPZ) treatment, an inhibitor of TNF synthesis. The involvement of HQ-induced TNF secretion in trachea mast cell degranulation was also demonstrated by the partial reversion of tracheal hyperresponsiveness in sodium cromoglicate-treated animals, and the in vivo HQ-exposure-induced degranulation of trachea connective tissue and mucosal mast cells, which was reversed by CPZ treatment. Our data show that in vivo HQ exposure indirectly exacerbates the parasympathetic-induced contraction of airway smooth muscle cells, mediated by TNF secreted by tracheal epithelial cells, clearly showing the link between environmental HQ exposure and the reactivity of airways.

Formaldehyde inhalation during pregnancy abolishes the development of acute innate inflammation in offspring

Formaldehyde (FA) is an environmental and occupational pollutant that induces programming mechanisms on the acquired immune host defense in offspring when exposed during the prenatal period. Hence, here we investigated whether the exposure of FA on pregnant rats could affect the development of an innate acute lung injury in offspring induced by lipopolissacaride (LPS) injection. Pregnant Wistar rats were exposed to FA (0.92 mg/m(3)) or vehicle (distillated water), both 1 h/day, 5 days/week, from 1 to 21 days of pregnancy. Non-manipulated rats were used as control. After 30 days of birth, the offspring was submitted to injection of LPS (Salmonella abortus equi, 5 mg/kg, i.p.). Systemic and lung inflammatory parameters were evaluated 24 h later. Exposure to FA during gestation abolished the development of acute lung injury in offspring, as observed by reduced number of leukocytes in the bronchoalveolar fluid (BAL), in the blood and in the bone marrow, and decreased myeloperoxidase activity in the lung. Moreover, phagocytes from BAL presented normal phagocytosis, but reduced oxidative burst. Alterations on the profile of inflammatory cytokines were evidenced by reduced mRNA levels of IL-6 and elevated levels of IL-10 and IFN gamma in the lung tissue. Indeed, mRNA levels of toll-likereceptor-4 and nuclear factor-kappa B translocation into the nucleus were also reduced. Additionally, hyperresponsiveness to methacholine was blunted in the trachea of offspring of FA exposed mothers. Together, our data clearly show that FA exposure in the prenatal period modifies the programming mechanisms of the innate defense in the offspring leading to impaired defense against infections.

Intestinal Lymph-Borne Factors Induce Lung Release of Inflammatory Mediators and Expression of Adhesion Molecules After an Intestinal Ischemic Insult

BACKGROUND Intestinal ischemia and reperfusion (I/R) is a documented cause of acute lung injury (ALI) and systemic inflammation. We previously reported that obstruction of thoracic lymphatic flow during intestinal I/R blunts pulmonary neutrophil recruitment and microvascular injury and decreases the systemic levels of tumor necrosis factor. Here, we consider the existence of a gut-lung axis promoting the induction of systemic inflammation, whereby drained intestinal lymph stimulates lung expression of adhesion molecules and matrix components and generation of inflammatory mediators. MATERIAL AND METHODS Upon administration of anesthesia, male Wistar rats were subjected to occlusion of the superior mesenteric artery for 45 min, followed by 2 h of intestinal reperfusion (I/R); groups of rats were subjected to I/R with or without thoracic lymphatic duct ligation immediately before the procedure. The non-manipulated rats were used to investigate basal parameters. RESULTS Obstruction of thoracic lymphatic flow before intestinal I/R decreased the ability of cultured lung tissue explants to release IL-1β, IL-10, and VEGF. In contrast, lymphatic obstruction normalized the elevated lung expression of PECAM-1 caused by intestinal I/R. On the other hand, lung E-selectin expression was significantly reduced, whereas fibronectin expression and collagen synthesis were not affected. Lymph levels of LTB(4) and TXB(2) were found to be significantly increased. CONCLUSIONS These data suggest that lymph factors drained from the intestine during ischemic trauma stimulate the lung to generate inflammatory mediators and alter the expression of adhesion molecules. Disturbances in lung homeostasis mediated by lymph might contribute to the spread of inflammatory processes, thereby accounting for the systemic inflammation induced by intestinal I/R.

Effects of MK-801 and amphetamine treatments on allergic lung inflammatory response in mice

Glutamate acts as a neurotransmitter within the Central Nervous System (CNS) and modifies immune cell activity. In lymphocytes, NMDA glutamate receptors regulate intracellular calcium, the production of reactive oxygen species and cytokine synthesis. MK-801, a NMDA receptor open-channel blocker, inhibits calcium entry into mast cells, thereby preventing mast cell degranulation. Several lines of evidence have shown the involvement of NMDA glutamate receptors in amphetamine (AMPH)-induced effects. AMPH treatment has been reported to modify allergic lung inflammation. This study evaluated the effects of MK-801 (0.25mg/kg) and AMPH (2.0mg/kg), given alone or in combination, on allergic lung inflammation in mice and the possible involvement of NMDA receptors in this process. In OVA-sensitized and challenged mice, AMPH and MK-801 given alone decreased cellular migration into the lung, reduced IL-13 and IL10 levels in BAL supernatant, reduced ICAM-1 and L-selectin expression in granulocytes in the BAL and decreased mast cell degranulation. AMPH treatment also decreased IL-5 levels. When both drugs were administered, treatment with MK-801 reversed the decrease in the number of eosinophils and neutrophils induced by AMPH in the BAL of OVA-sensitized and challenged mice as well as the effects on the expression of L-selectin and ICAM-1 in granulocytes, the IL-10, IL-5 and IL-13 levels in BAL supernatants and increased mast cell degranulation. At the same time, treatment with MK-801, AMPH or with MK-801+AMPH increased corticosterone serum levels in allergic mice. These results are discussed in light of possible indirect effects of AMPH and MK-801 via endocrine outflow from the CNS (i.e., HPA-axis activity) to the periphery and/or as a consequence of the direct action of these drugs on immune cell activity, with emphasis given to mast cell participation in the allergic lung response of mice.

Long-term amphetamine treatment exacerbates inflammatory lung reaction while decreases airway hyper-responsiveness after allergic stimulus in rats.

Asthma is an allergic lung disease can be modulated by drugs that modify the activity of central nervous system (CNS) such as amphetamine (AMPH). AMPH is a highly abused drug that exerts potent effects on behavior and immunity. In this study we investigated the mechanism involved in the effects of long-term AMPH treatment on the increased magnitude of allergic lung response. We evaluated mast cells degranulation, cytokines release, airways responsiveness and, expression of adhesion molecules. Male Wistar rats were treated with AMPH or vehicle (PBS) for 21 days and sensitized with ovalbumin (OVA) one week after the first injection of vehicle or AMPH. Fourteen days after the sensitization, the rats were challenged with an OVA aerosol, and 24h later their parameters were analyzed. In allergic rats, the treatment with AMPH exacerbated the lung cell recruitment due increased expression of ICAM-1, PECAM-1 and Mac-1 in granulocytes and macrophages recovered from bronchoalveolar lavage. Elevated levels of IL-4, but decreased levels of IL-10 were also found in samples of lung explants after AMPH treatment. Conversely, the ex-vivo tracheal hyper-responsiveness to methacholine (MCh) was reduced by AMPH treatment, whereas the force contraction of tracheal segments due to in vitro antigen challenge remained unaltered. Our findings suggest that lung inflammation and airway hyper-responsiveness due to OVA challenge are under the distinct control of AMPH during long-term treatment. Our data strongly indicate that AMPH positively modulates allergic lung inflammation via the increase of ICAM-1, PECAM-1, Mac-1 and IL-4. AMPH also abrogates the release of the anti-inflammatory cytokine IL-10.

Connective tissue mast cells are the target of formaldehyde to induce tracheal hyperresponsiveness in rats: Putative role of leukotriene B4 and nitric oxide

Formaldehyde (FA) exposure induces upper airways irritation and respiratory abnormalities, but its mechanisms are not understood. Since mast cells are widely distributed in the airways, we hypothesized that FA might modify the airways reactivity by mechanism involving their activation. Tracheal rings of rats were incubated with Dulbeccos modified medium culture containing FA (0.1 ppm) in 96-well plastic microplates in a humid atmosphere. After 30 min, 6h, and 24-72 h, the rings were suspended in an organ bath and dose-response curve to methacholine (MCh) were determined. Incubation with FA caused a transient tracheal hyperresponsiveness to MCh that was independent from tracheal epithelium integrity. Connective tissue mast cell depletion caused by compound 48/80 or mast cell activation by the allergic reaction, before exposure of tracheal rings to FA prevented the increased responsiveness to MCh. LTB(4) concentrations were increased in the culture medium of tracheas incubated with FA for 48 h, whereas the LTB(4)-receptor antagonist MK886 (1 microM) added before FA exposure rendered the tracheal rings normoreactive to MCh. In addition, FA exposure did not cause hyperresponsiveness in tracheal segments incubated with l-arginine (1 microM). We suggest that airway connective tissue mast cells constitute the target and may provide the increased LTB(4) generation as well as an elevated consumption of NO leading to tracheal hyperresponsiveness to MCh.