KELLY YOSHIZAKI
Projetos de Pesquisa
Unidades Organizacionais
LIM/05 - Laboratório de Poluição Atmosférica Experimental, Hospital das Clínicas, Faculdade de Medicina
10 resultados
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- Effects of different mechanical ventilation strategies on the mucociliary system(2011) PICCIN, Vivien S.; CALCIOLARI, Christiane; YOSHIZAKI, Kelly; GOMES, Susimeire; ALBERTINI-YAGI, Claudia; DOLHNIKOFF, Marisa; MACCHIONE, Mariangela; CALDINI, Elia G.; SALDIVA, Paulo H. N.; NEGRI, Elnara M.To evaluate the effects of different mechanical ventilation (MV) strategies on the mucociliary system. Experimental study. Twenty-seven male New Zealand rabbits. After anesthesia, animals were tracheotomized and ventilated with standard ventilation [tidal volume (Vt) 8 ml/kg, positive end expiratory pressure (PEEP) 5 cmH(2)O, flow 3 L/min, FiO(2) 0.4] for 30 min. Next, animals were randomized into three groups and ventilated for 3 h with low volume (LV): Vt 8 ml/kg, PEEP 5 cmH(2)O, flow 3 L/min (n = 6); high volume (HV): Vt 16 ml/kg, PEEP 5 cmH(2)O, flow 5 L/min (n = 7); or high pressure (HP): Ppeak 30 cmH(2)O, PEEP 12 cmH(2)O (n = 8). Six animals (controls) were ventilated for 10 min with standard ventilation. Vital signals, blood lactate, and respiratory system mechanics were verified. Tracheal tissue was collected before and after MV. Lung and tracheal tissue sections were stained to analyze inflammation and mucosubstances by the point-counting method. Electron microscopy verified tracheal cell ultrastructure. In situ tracheal ciliary beating frequency (CBF), determined using a videoscopic technique, and tracheal mucociliary transport (TMCT), assessed by stereoscopic microscope, were evaluated before and after MV. Respiratory compliance decreased in the HP group. The HV and HP groups showed higher lactate levels after MV. Macroscopy showed areas of atelectasis and congestion on HV and HP lungs. Lung inflammatory infiltrate increased in all ventilated groups. Compared to the control, ventilated animals also showed a reduction of total and acid mucus on tracheal epithelium. Under electron microscopy, injury was observed in the ciliated cells of the HP group. CBF decreased significantly after MV only in the HP group. TMCT did not change significantly in the ventilated groups. Different MV strategies induce not only distal lung alterations but also morphological and physiological tracheal alterations leading to mucociliary system dysfunction.
- Acute exposure to diesel and sewage biodiesel exhaust causes pulmonary and systemic inflammation in mice(2018) BRITO, Jose Mara de; MAUAD, Thais; CAVALHEIRO, Guilherme Franco; YOSHIZAKI, Kelly; ANDRE, Paulo Afonso de; LICHTENFELS, Ana Julia F. C.; GUIMARAES, Eliane Tigre; RIVERO, Dolores Helena Rodriguez Ferreira; ANTONANGELO, Leila; OLIVEIRA, Luciano Basto; PEDROSO, Luiz Roberto Martins; MACCHIONE, Mariangela; SALDIVA, Paulo Hilario NascimentoBiodiesel is a renewable energy source that reduces particle emission, but few studies have assessed its effects. To assess the effects of acute inhalation of two doses (600 and 1200 mu g/m(3)) of diesel (DE) and biodiesel (BD) fuels on the inflammatory pulmonary and systemic profile of mice. Animals were exposed for 2 h in an inhalation chamber inside the Container Laboratory for Fuels. Heart rate, heart rate variability (HRV) and blood pressure were determined 30 min after exposure. After 24 h. we analyzed the lung inflammation using bronchoalveolar lavage fluid (BALF); neutrophil and macrophage quantification in the lung parenchyma was performed, and blood and bone marrow biomarkers as well as receptor of endothelin-A (ET-Ar), receptor of endothelin-B (ET-Br), vascular cell adhesion molecule 1 (VCAM-1), inducible nitric oxide synthase (iNOs) and isoprostane (ISO) levels in the pulmonary vessels and bronchial epithelium were evaluated. HRV increased for BD600, D600 and D1200 compared to filtered air (FA). Both fuels (DE and BD) produced alterations in red blood cells independent of the dose. BALI from the BD600 and BD1200 groups showed an increase in neutrophils compared to those of the FA group. Numeric density of the polyrnorphonuclear and mononudear cells was elevated with BD600 compared to FA. In the peribronchiolar vessels, there was an increase in ET-Ar and ET-Br expression following BD600 compared to IA; and there was a reduction in the iNOs expression for BD1200 and the VCAM-1 for D1200 compared to FA. In the bronchial epithelium, there was an increase in ETAr at BD600, ET-Br at two doses (600 and 1200 mu g/m(3)) of DE and BD, iNOs at D600 and VCAM-1 at BD1200 and D600; all groups were compared to the FA group. Acute exposure to DE and BD derived from sewage methyl esters triggered pulmonary and cardiovascular inflammatory alterations in mice.
- Vesicular acetylcholine transport deficiency potentiates some inflammatory responses induced by diesel exhaust particles(2019) SANTANA, Fernanda P. R.; PINHEIRO, Nathalia M.; BITTENCOURT-MERNAK, Marcia I.; PERINI, Adenir; YOSHIZAKI, Kelly; MACCHIONE, Mariangela; SALDIVA, Paulo H. N.; MARTINS, Milton A.; TIBERIO, Iolanda F. L. C.; PRADO, Marco Antonio M.; PRADO, Vania F.; PRADO, Carla M.Endogenous acetylcholine (ACh), which depends of the levels of vesicular ACh transport (VAChT) to be released, is the central mediator of the cholinergic anti-inflammatory system. ACh controls the release of cytokine in different models of inflammation. Diesel exhaust particles (DEP) are one of the major environmental pollutants produced in large quantity by automotive engines in urban center. DEP bind the lung parenchyma and induce inflammation. We evaluated whether cholinergic dysfunction worsens DEP-induced lung inflammation. Male mice with decreased ACh release due to reduced expression of VAChT (VAChT-KD mice) were submitted to DEP exposure for 30 days (3 mg/mL of DEP, once a day, five days a week) or saline. Pulmonary function and inflammation as well as extracellular matrix fiber deposition were evaluated. Additionally, airway and nasal epithelial mucus production were quantified. We found that DEP instillation worsened lung function and increased lung inflammation. Higher levels of mononuclear cells were observed in the peripheral blood of both wild-type (WT) and VAChT-KD mice. Also, both wild-type (WT) and VAChT-KD mice showed an increase in macrophages in bronchoalveolar lavage fluid (BALF) as well as increased expression of IL-4, IL-6, IL-13, TNF-alpha, and NF-kappa B in lung cells. The collagen fiber content in alveolar septa was also increased in both genotypes. On the other hand, we observed that granulocytes were increased only in VAChT-KD peripheral blood. Likewise, increased BALF lymphocytes and neutrophils as well as increased elastic fibers in alveolar septa, airway neutral mucus, and nasal epithelia acid mucus were observed only in VAChT-KD mice. The cytokines IL-4 and TNF-alpha were also higher in VAChT-KD mice compared with WT mice. In conclusion, decreased ability to release ACh exacerbates some of the lung alterations induced by DEP in mice, suggesting that VAChT-KD animals are more vulnerable to the effects of DEP in the lung.
- The effects of urban particulate matter on the nasal epithelium by gender: An experimental study in mice(2016) YOSHIZAKI, K.; FUZIWARA, C. S.; BRITO, J. M.; SANTOS, T. M. N.; KIMURA, E. T.; CORREIA, A. T.; AMATO-LOURENCO, L. F.; VASCONCELLOS, P.; SILVA, L. F.; BRENTANI, M. M.; MAUAD, T.; SALDIVA, P. H. N.; MACCHIONE, M.Nose is the first portion of the respiratory system into contact with air pollution particles, including organic compounds that could act as endocrine releasers. The objective was to identify and quantify estrogenic receptor-beta (ER beta), aryl hydrocarbon receptor (AhR), the cytochrome P450 enzymes CYP1A1, 1A2, 1B1, and mucus profile in the nasal epithelium of mice. BALB/c mice male (n = 32) and female (n = 82) in proestrus, estrus and diestrus were divided into two groups: 1) exposed to ambient air; 2) concentrated ambient particles (CAPs) to achieve an accumulated dose (concentration vs. time product) of 600 mu g/m(3), the time of the exposure was controlled to ensure the same concentration for all groups (5 days per week for 40-51 days). RT-PCR (Er beta-1, Er beta-2, Ahr, Cyp1a1, Cyp1a2, Cyp1b1), immunohistochemistry and morphometry (ER(3, AhR) were used to analyze. The mucus profiles were examined using acid (Aldan Blue) and neutral (periodic acid Schiff's) stains. Exposed females had significantly lower levels of Er beta-2 mRNA than exposed males (p = 0.036). Cyp1b1 mRNA in diestrus females was significantly lower in the CAP-exposed group compared with the ambient air group (p <= 0.05). ER beta expression in the epithelium and submucosa nucleus was lower in estrus exposed to CAPs compared with ambient air. CAPs increases AhR in the epithelium (p = 0.044) and submucosa (p = 0.001) nucleus of female when compared with male mice. Exposure to CAPs, also led to relatively increased acidic content in the mucus of males (p = 0.048), but decreased acidic content in that of females (p = 0.04). This study revealed sex dependent responses to air pollution in the nasal epithelium that may partially explain the predisposition of females to airway respiratory diseases.
- Chronic exposure of diesel exhaust particles induces alveolar enlargement in mice(2015) YOSHIZAKI, Kelly; BRITO, Jose Mara; MORIYA, Henrique T.; TOLEDO, Alessandra C.; FERZILAN, Sandra; OLIVEIRA, Ana Paula Ligeiro de; MACHADO, Isabel D.; FARSKY, Sandra H. P.; SILVA, Luiz F. F.; MARTINS, Milton A.; SALDIVA, Paulo H. N.; MAUAD, Thais; MACCHIONE, MariangelaBackground: Diesel exhaust particles (DEPs) are deposited into the respiratory tract and are thought to be a risk factor for the development of diseases of the respiratory system. In healthy individuals, the timing and mechanisms of respiratory tract injuries caused by chronic exposure to air pollution remain to be clarified. Methods: We evaluated the effects of chronic exposure to DEP at doses below those found in a typical bus corridor in Sao Paulo (150 mu g/m(3)). Male BALB/c mice were divided into mice receiving a nasal instillation: saline (saline; n = 30) and 30 mu g/10 mu L of DEP (DEP; n = 30). Nasal instillations were performed five days a week, over a period of 90 days. Bronchoalveolar lavage (BAL) was performed, and the concentrations of interleukin (IL)-4, IL-10, IL-13 and interferon-gamma (INF-gamma) were determined by ELISA-immunoassay. Assessment of respiratory mechanics was performed. The gene expression of Muc5ac in lung was evaluated by RT-PCR. The presence of IL-13, MAC2+ macrophages, CD3+, CD4+, CD8+ T cells and CD20+ B cells in tissues was analysed by immunohistochemistry. Bronchial thickness and the collagen/elastic fibers density were evaluated by morphometry. We measured the mean linear intercept (Lm), a measure of alveolar distension, and the mean airspace diameter (D0) and statistical distribution (D2). Results: DEP decreased IFN-gamma levels in BAL (p = 0.03), but did not significantly alter IL-4, IL-10 and IL-13 levels. MAC2+ macrophage, CD4+ T cell and CD20+ B cell numbers were not altered; however, numbers of CD3+ T cells (p <= 0.001) and CD8+ T cells (p <= 0.001) increased in the parenchyma. Although IL-13 (p = 0.008) expression decreased in the bronchiolar epithelium, Muc5ac gene expression was not altered in the lung of DEP-exposed animals. Although respiratory mechanics, elastic and collagen density were not modified, the mean linear intercept (Lm) was increased in the DEP-exposed animals (p <= 0.001), and the index D2 was statistically different (p = 0.038) from the control animals. Conclusion: Our data suggest that nasal instillation of low doses of DEP over a period of 90 days results in alveolar enlargement in the pulmonary parenchyma of healthy mice.
- Fragrances as a trigger of immune responses in different environments(2024) MACCHIONE, M.; YOSHIZAKI, K.; FRIAS, D. P.; MAIER, K.; SMELAN, J.; PRADO, C. M.; MAUAD, T.Fragrances can cause allergic skin reactions, expressed as allergic contact dermatitis and reactions in the respiratory tract that range from acute temporary upper airway irritation to obstructive lung disease. These adverse health effects may result from the stimulation of a specific (adaptive) immune response. Th1 cells, which essentially produce interleukin-2 (IL-2) and interferon-gamma (IFN-gamma), play a key role in allergic contact dermatitis and also on allergic sensitization to common allergens (e.g., nickel and fragrance). It has been shown that fragrance allergy leads to Th2/Th22 production of IL-4, IL-5 and IL-13, controlling the development of IgE and mediating hypersensitivity reactions in the lung, such as asthma. Cytokines released during immune response modulate the expression of cytochrome P450 (CYPs) proteins, which can result in alterations of the pharmacological effects of substances in inflammatory diseases. The mechanisms linking environment and immunity are still not completely understood but it is known that aryl hydrocarbon receptor (AhR) is a sensor with conserved ligand-activated transcription factor, highly expressed in cells that controls complex transcriptional programs which are ligand and cell type specific, with CYPs as targeted genes. This review focuses on these important aspects of immune responses of the skin and respiratory tract cells, describing some in vitro models applied to evaluate the mechanisms involved in fragrance-induced allergy.
- Nrf2 positively regulates autophagy antioxidant response in human bronchial epithelial cells exposed to diesel exhaust particles(2020) FRIAS, Daniela Perroni; GOMES, Raquel Labiapari Nunes; YOSHIZAKI, Kelly; CARVALHO-OLIVEIRA, Regiani; MATSUDA, Monique; JUNQUEIRA, Mara de Souza; TEODORO, Walcy Rosolia; VASCONCELLOS, Perola de Castro; PEREIRA, Daniela Cristina de Almeida; CONCEICAO, Paulo Roberto da; SALDIVA, Paulo Hilario Nascimento; MAUAD, Thais; MACCHIONE, MariangelaDiesel exhaust particles (DEP) are known to generate reactive oxygen species in the respiratory system, triggering cells to activate antioxidant defence mechanisms, such as Keap1-Nrf2 signalling and autophagy. The aim of this study was to investigate the relationship between the Keap1-Nrf2 signalling and autophagy pathways after DEP exposure. BEAS-2B cells were transfected with silencing RNA (siRNA) specific to Nrf2 and exposed to DEP. The relative levels of mRNA for Nrf2, NQO1, HO-1, LC3B, p62 and Atg5 were determined using RT-PCR, while the levels of LCB3, Nrf2, and p62 protein were determined using Western blotting. The autophagy inhibitor bafilomycin caused a significant decrease in the production of Nrf2, HO-1 and NQO1 compared to DEPs treatment, whereas the Nrf2 activator sulforaphane increased the LC3B (p = 0.020) levels. BEAS-2B cells exposed to DEP at a concentration of 50 mu g/mL for 2 h showed a significant increase in the expression of LC3B (p = 0.001), p62 (p = 0.008), Nrf2 (p = 0.003), HO-1 (p = 0.001) and NQO1 (p = 0.015) genes compared to control. In siRNA-transfected cells, the LC3B (p < 0.001), p62 (p = 0.001) and Atg5 (p = 0.024) mRNA levels and the p62 and LC3II protein levels were decreased, indicating that Nrf2 modulated the expression of autophagy markers (R < 1). These results imply that, in bronchial cells exposed to DEP, the Nrf2 system positively regulates autophagy to maintain cellular homeostasis.
- The effects of particulate matter on inflammation of respiratory system: Differences between male and female(2017) YOSHIZAKI, Kelly; BRITO, Jose Mara; SILVA, Luiz Fernando; LINO-DOS-SANTOS-FRANCO, Adriana; FRIAS, Daniela Perroni; SILVA, Renata Calciolari Rossi e; AMATO-LOURENCO, Luis Fernando; SALDIVA, Paulo Hilario Nascimento; TIBERIO, Iolanda de Fatima Lopes Calvo; MAUAD, Thais; MACCHIONE, MariangelaAir pollution is known to exacerbate respiratory diseases and epidemiological studies have shown that women present more chronic respiratory symptoms than man exposed to traffic pollution, however, the reason why is unclear. This study evaluated the inflammatory differences in BALB/c mouse males (n = 34) and females (n = 111) in three phases of the estrous cycle that were exposed to ambient air (AA) or concentrated ambient particles (CAPs). Tracheal hyperreactivity to methacholine, bronchoalveolar lavage fluid (BALF) and immunohistochemical of airways and lung parenchyma were studied. Hyperreactivity increased in CAPs-exposed female mice compared with AA-exposed mice in estrus (p < 0.05) and proestrus phases (p < 0.05) and decreased in CAPs-exposed males compared with those exposed to AA (p < 0.05). Males had increased numbers of total cells (p = 0.037) and macrophages (p = 0.028) compared to females. BALF levels of cyclooxygenase-2(COX-2) (p = 0.000), transforming growth factor alpha (TGF-alpha) (p = 0.001) and IL-8 receptor alpha (IL-8R alpha) (p = 0.014) were increased in males compared with proestrus, estrus and diestrus females, independent of exposure. Proestrus females exhibited significantly higher cadherin expression in lung parenchyma than did males (p = 0.005). CAPs exposure increased matrix metalloproteinase-9 (MMP-9) (p = 0.024) and isoprostane (p = 0.003) expression in the airways of both, males and females. The level of substance P (SP) (p = 0.001) increased in lung parenchyma in males compared with females, while IL-17 levels in airways (p = 0.042) and in lung parenchyma (p = 0.008) increased in females. MMP-9 levels (p = 0.024) were significantly lower in the lung parenchyma of CAPs-exposed females. TGF-alpha (p = 0.007) levels increased in the lung parenchyma of CAPs-exposed females compared to AA-exposed females. These results suggest that inflammatory markers differentially expressed in male mice were mostly linked to acute inflammation (IL-1 beta, IL-8R alpha, COX-2), whereas in females, markers that may lead to a chronic inflammatory process such as IL-17 and remodeling (MMP-9) were increased.
- Acute cardiopulmonary effects induced by the inhalation of concentrated ambient particles during seasonal variation in the city of Sao Paulo(2014) BRITO, Jose Mara de; MACCHIONE, Mariangela; YOSHIZAKI, Kelly; TOLEDO-ARRUDA, Alessandra Choqueta; SARAIVA-ROMANHOLO, Beatriz Mangueira; ANDRADE, Maria de Fatima; MAUAD, Thais; RIVERO, Dolores Helena Rodriguez Ferreira; SALDIVA, Paulo Hilario NascimentoAmbient particles may undergo modifications to their chemical composition as a consequence of climatic variability. The determination of whether these changes modify the toxicity of the particles is important for the understanding of the health effects associated with particle exposure. The objectives were to determine whether low levels of particles promote cardiopulmonary effects, and to assess if the observed alterations are influenced by season. Mice were exposed to 200 mu g/m(3) concentrated ambient particles (CAPs) and filtered air (FA) in cold/dry and warm/humid periods. Lung hyperresponsiveness, heart rate, heart rate variability, and blood pressure were evaluated 30 min after each exposure. After 24 h, blood and tissue samples were collected. During both periods (warm/humid and cold/dry), CAPs induced alterations in red blood cells and lung inflammation. During the cold/dry period, CAPs reduced the mean corpuscular volume levels and increased erythrocytes, hemoglobin, mean corpuscular hemoglobin concentration, and red cell distribution width coefficient variation levels compared with the FA group. Similarly, CAPs during the warm/humid period decreased mean corpuscular volume levels and increased erythrocytes, hemoglobin, hematocrit, and red cell distribution width coefficient variation levels compared with the FA group. CAPs during the cold/dry period increased the influx of neutrophils in the alveolar parenchyma. Short-term exposure to low concentrations of CAPs elicited modest but significant pulmonary inflammation and, to a lesser extent, changes in blood parameters. In addition, our data support the concept that changes in climate conditions slightly modify particle toxicity because equivalent doses of CAPs in the cold/dry period produced a more exacerbated response.
- Inhibition of MAPK and STAT3-SOCS3 by Sakuranetin Attenuated Chronic Allergic Airway Inflammation in Mice(2019) SANTANA, Fernanda P. R.; SILVA, Rafael C. da; GRECCO, Simone dos S.; PINHEIRO, Aruana J. M. C. R.; CAPERUTO, Luciana C.; ARANTES-COSTA, Fernanda M.; CLAUDIO, Samuel R.; YOSHIZAKI, Kelly; MACCHIONE, Mariangela; RIBEIRO, Daniel A.; TIBERIO, Iolanda F. L. C.; LIMA-NETO, Lidio G.; LAGO, Joao H. G.; PRADO, Carla M.Asthma allergic disease is caused by airway chronic inflammation. Some intracellular signaling pathways, such as MAPK and STAT3-SOCS3, are involved in the control of airway inflammation in asthma. The flavonoid sakuranetin demonstrated an anti-inflammatory effect in different asthma models. Our aim was to clarify how sakuranetin treatment affects MAPK and STAT3-SOCS3 pathways in a murine experimental asthma model. Mice were submitted to an asthma ovalbumin-induction protocol and were treated with vehicle, sakuranetin, or dexamethasone. We assayed the inflammatory profile, mucus production, and serum antibody, STAT3-SOCS3, and MAPK levels in the lungs. Morphological alterations were also evaluated in the liver. LPS-stimulated RAW 264.7 cells were used to evaluate the effects of sakuranetin on nitric oxide (NO) and cytokine production. In vivo, sakuranetin treatment reduced serum IgE levels, lung inflammation (eosinophils, neutrophils, and Th2/Th17 cytokines), and respiratory epithelial mucus production in ovalbumin-sensitized animals. Considering possible mechanisms, sakuranetin inhibits the activation of ERK1/2, JNK, p38, and STAT3 in the lungs. No alterations were found in the liver for treated animals. Sakuranetin did not modify in vitro cell viability in RAW 264.7 and reduced NO release and gene expression of IL-1 beta and IL-6 induced by LPS in these cells. In conclusion, our data showed that the inhibitory effects of sakuranetin on eosinophilic lung inflammation can be due to the inhibition of Th2 and Th17 cytokines and the inhibition of MAPK and STAT3 pathways, reinforcing the idea that sakuranetin can be considered a relevant candidate for the treatment of inflammatory allergic airway disease.