Nanoparticles increase the efficacy of cancer chemopreventive agents in cells exposed to cigarette smoke condensate

Lung cancer is a leading cause of death in developed countries. Although smoking cessation is a primary strategy for preventing lung cancer, former smokers remain at high risk of cancer. Accordingly, there is a need to increase the efficacy of lung cancer prevention. Poor bioavailability is the main factor limiting the efficacy of chemopreventive agents. The aim of this study was to increase the efficacy of cancer chemopreventive agents by using lipid nanoparticles (NPs) as a carrier. This study evaluated the ability of lipid NPs to modify the pharmacodynamics of chemopreventive agents including N-acetyl-L-cysteine, phenethyl isothiocyanate and resveratrol (RES). The chemopreventive efficacy of these drugs was determined by evaluating their abilities to counteract cytotoxic damage (DNA fragmentation) induced by cigarette smoke condensate (CSC) and to activate protective apoptosis (annexin-V cytofluorimetric staining) in bronchial epithelial cells both in vitro and in ex vivo experiment in mice. NPs decreased the toxicity of RES and increased its ability to counteract CSC cytotoxicity. NPs significantly increased the ability of phenethyl isothiocyanate to attenuate CSC-induced DNA fragmentation at the highest tested dose. In contrast, this potentiating effect was observed at all tested doses of RES, NPs dramatically increasing RES-induced apoptosis in CSC-treated cells. These results provide evidence that NPs are highly effective at increasing the efficacy of lipophilic drugs (RES) but are not effective towards hydrophilic agents (N-acetyl-L-cysteine), which already possess remarkable bioavailability. Intermediate effects were observed for phenethyl isothiocyanate. These findings are relevant to the identification of cancer chemopreventive agents that would benefit from lipid NP delivery.

Cigarette smoke exposure impairs respiratory epithelial ciliogenesis

Background: Cigarette smoke exposure is considered an important negative prognostic factor for chronic rhinosinusitis (CRS) patients. However, there is no clear mechanistic evidence implicating cigarette smoke exposure in the poor clinical evolution of the disease or in the maintenance of the inflammatory state characterizing CRS. This study aimed to evaluate the effects of cigarette smoke exposure on respiratory cilia differentiation. Methods: Monse nasal septal epithelium cultures grown at an air-liquid interface were used as a model of respiratory epithelium. After 5 days of cell growth, cultures were exposed to air on the apical surface. Additionally, cigarette smoke condensate (CSC; the particulate phase of tobacco smoke) or cigarette smoke extract (CSE; the volatile phase) Were diluted in the basolateral compartment in different concentrations. After 15 days of continuous exposure, scanning electron microscopy and immunofluorescence for type IV tubulin were used to determine presence and maturation of cilia. Transepithelial resistance was also recorded to evaluate confluence and physiological barrier integrity. Results: CSC and CSE impair ciliogenesis in a dose-dependent manner with notable effects in concentrations higher than 30 mu g/mL, yielding >70% nonciliation and shorter cilia compared With control. No statistical difference on transepithelial resistance was evident. Conclusion: CSC and CSE exposure negatively impacts ciliogenesis of respiratory cells at concentrations not effecting transepithelial resistance. The impairment on ciliogenesis reduce the mucociliary clearance apparatuts after injury and/or infection and may explain the poor response to therapy for CRS patients exposed to tobacco smoke.

Effects of sidestream cigarette smoke exposure on baroreflex components in spontaneously hypertensive rats

We evaluated short-term effects of sidestream cigarette smoke (SSCS) exposure on baroreflex function in spontaneously hypertensive rats (SHR) and Wistar Kyoto (WKY) normotensive rats. Rats were exposed to SSCS during three weeks, 180min, five days per week, in a concentration of carbon monoxide (CO) between 100 and 300ppm. We observed that SSCS exposure increased tachycardic peak and heart rate range while it attenuated bradycardic reflex in WKY. In respect to SHR, SSCS also increased tachycardic peak. Taken together, our data suggests that three weeks of exposure to SSCS affects the sympathetic and parasympathetic component of the baroreflex in normotensive WKY while it tended to affect the sympathetic component in SHR.

Cigarette smoke inhalation modulates gene expression in sites of bone healing: a study in rats

Objective. The aim of this study was to assess the effect of cigarette smoke inhalation (CSI) on gene expression in alveolar bone healing sites. Study design. Wistar rats were randomly assigned to the groups: control [animals not exposed to CSI (n = 20)] and test [animals exposed to CSI, starting 3 days before teeth extraction and maintained until killing them (n = 20)]. First mandibular molars were bilaterally extracted, and the expression of alkaline phosphatase, bone morphogenetic protein (BMP) 2 and 7, receptor activator of nuclear factor kappa B ligand, osteoprotegerin, and d2 isoform of vacuolar adenosine triphosphatase V(o) domain were assessed by quantitative polymerase chain reaction in the newly formed tissue in the sockets. Results. Overall, data analysis demonstrated that CSI significantly affected the expression pattern of all of the studied genes except BMP-7. Conclusion. The expression of key genes for bone healing may be affected by CSI in tooth extraction sites. (Oral Surg Oral Med Oral Pathol Oral Radiol Endod 2010;110:447-452)

Expression of placental glutathione S-transferase in rat tongue mucosa exposed to cigarette smoke

Glutatione S-transferases (GSTs) are a family of enzymes involved in detoxification of xenobiotics. Placental GST, known as GST-P, has been detected in tissues following exposure to carcinogenic agents being regarded a reliable biomarker of exposure and susceptibility in early phases of carcinogenesis. The aim of this study was to investigate the expressivity of GST-P positive foci in the rat tongue mucosa exposed to cigarette smoke by means of immunohistochemistry. A total of twelve male Wistar rats were distributed into two groups: negative control and experimental group exposed to cigarette smoke during 75 days. After experimental period, no histopathological changes in the tongue mucosa were evidenced in the negative control and the experimental group. However, a total of five GST-P positive foci were detected in two out of six animals exposed to cigarrette smoke. None control animals were noticed GST-P positive foci. These data indicate that expression of GST-P may reflect the carcinogenic effect of cigarette smoke as well as the genetic susceptibility of animals in relation to continuous carcinogens exposure.

Deposition of Lead and Cadmium Released by Cigarette Smoke in Dental Structures and Resin Composite

Cigarette smoke is a significant source of cadmium, lead, and toxic elements, which are absorbed into the human organism. In this context, the aim of this study was to investigate in vitro the presence of toxic elements, cadmium, and lead deriving from cigarette smoke in the resin composite, dentine, and dental enamel. Eight cylindrical specimens were fabricated from resin composite, bovine enamel, and root dentin fragments that were wet ground and polished with abrasive paper to obtain sections with 6-mm diameter and 2-mm thickness. All specimens were exposed to the smoke of 10 cigarettes/day during 8 days. After the simulation of the cigarette smoke, the specimens were examined with scanning electron microscopy (SEM) and the energy-dispersive X-ray analysis. In the photomicrographic analysis in SEM, no morphological alterations were found; however, the microanalysis identified the presence of cadmium, arsenic, and lead in the different specimens. These findings suggest that the deposition of these elements derived from cigarette smoke could be favored by dental structures and resin composite. Microsc. Res. Tech. 74:287-291, 2011. (C) 2010 Wiley-Liss, Inc.

Behavior of cardiac variables in animals exposed to cigarette smoke

OBJECTIVE: To assess the behavior of cardiac variables in animals exposed to cigarette smoke. METHODS: Two groups of Wistar rats were studied as follows: control group (C), comprising 28 animals; and smoking group (S), comprising 23 animals exposed to cigarette smoke for 30 days. Left ventricular cardiac function was assessed in vivo with transthoracic echocardiography, and myocardial performance was analyzed in vitro in preparations of isolated left ventricular papillary muscle. The cardiac muscle was assessed in isometric contractions with an extracellular calcium concentration of 2.5 mmol/L. RESULTS: No statistical difference was observed in the values of the body variables of the rats and in the mechanical data obtained from the papillary muscle between the control and smoking groups. The values of left ventricular systolic diameter were significantly greater in the smoking animals than in the control animals (C= 3.39 ± 0.4 mm and S= 3.71 ± 0.51 mm, P=0.02). A significant reduction was observed in systolic shortening fraction (C= 56.7 ± 4.2% and S= 53.5 ± 5.3%, P=0.02) and in ejection fraction (C= 0.92 ± 0.02 and S= 0.89 ± 0.04, P=0.01). CONCLUSION: The rats exposed to cigarette smoke had a reduction in left ventricular systolic function, although their myocardial function was preserved.

Short-term exposure of mice to cigarette smoke and/or residual oil fly ash produces proximal airspace enlargements and airway epithelium remodeling

Chronic obstructive pulmonary disease (COPD) is associated with inflammatory cell reactions, tissue destruction and lung remodeling. Many signaling pathways for these phenomena are still to be identified. We developed a mouse model of COPD to evaluate some pathophysiological mechanisms acting during the initial stage of the disease. Forty-seven 6- to 8-week-old female C57/BL6 mice (approximately 22 g) were exposed for 2 months to cigarette smoke and/or residual oil fly ash (ROFA), a concentrate of air pollution. We measured lung mechanics, airspace enlargement, airway wall thickness, epithelial cell profile, elastic and collagen fiber deposition, and by immunohistochemistry transforming growth factor-β1 (TGF-β1), macrophage elastase (MMP12), neutrophils and macrophages. We observed regional airspace enlargements near terminal bronchioles associated with the exposure to smoke or ROFA. There were also increases in airway resistance and thickening of airway walls in animals exposed to smoke. In the epithelium, we noted a decrease in the ciliated cell area of animals exposed to smoke and an increase in the total cell area associated with exposure to both smoke and ROFA. There was also an increase in the expression of TGF-β1 both in the airways and parenchyma of animals exposed to smoke. However, we could not detect inflammatory cell recruitment, increases in MMP12 or elastic and collagen fiber deposition. After 2 months of exposure to cigarette smoke and/or ROFA, mice developed regional airspace enlargements and airway epithelium remodeling, although no inflammation or increases in fiber deposition were detected. Some of these phenomena may have been mediated by TGF-β1.

Electron spin resonance study of puff-resolved free radical formation in mainstream cigarette smoke

Puff-by-puff resolved gas phase free radicals were measured in mainstream smoke from Kentucky 2R4F reference cigarettes using ESR spectroscopy. Three spin-trapping reagents were evaluated: PBN, DMPO and DEPMPO. Two procedures were used to collect gas phase smoke on a puff-resolved basis: i) the accumulative mode, in which all the gas phase smoke up to a particular puff was bubbled into the trap (i.e., the 5th puff corresponded to the total smoke from the 1st to 5th puffs). In this case, after a specified puff, an aliquot of the spin trap was taken and analysed; or, ii) the individual mode, in which the spin trap was analysed and then replaced after each puff. Spin concentrations were determined by double-integration of the first derivative of the ESR signal. This was compared with the integrals of known standards using the TEMPO free radical. The radicals trapped with PBN were mainly carbon-centred, whilst the oxygen-centred radicals were identified with DMPO and DEPMPO. With each spin trap, the puff-resolved radical concentrations showed a characteristic pattern as a function of the puff number. Based on the spin concentrations, the DMPO and DEPMPO spin traps showed better trapping efficiencies than PBN. The implication for gas phase free radical analysis is that a range of different spin traps should be used to probe complex free radical reactions in cigarette smoke.

Electron paramagnetic resonance investigation of the free radicals in the gas and particulate phases of cigarette smoke using spin-trapping techniques

Free radicals in cigarette smoke have been studied using spin trapping EPR techniques. 2R4F reference cigarettes were smoked using 35 ml puff volumes of 2 seconds duration, once every 60 seconds. The particulate phase of the smoke was separated from the gas phase by passing the smoke through a Cambridge filter pad. For both phases, free radicals were measured and identified. A range of spin-traps was employed: PBN, DMPO, DEPMPO, and DPPH-PBN. In the gas-phase, short-lived carbon- and oxygen- centered radicals were identified; the ratios between them changed during the smoking runs. For the first puffs, C-centered radicals predominated while for the later puffs, O-centered radicals were mainly observed. The particulate phase and the ‘tar’ were studied as well.

Cigarette smoke-induced neurogenic inflammation is mediated by α,β-unsaturated aldehydes and the TRPA1 receptor in rodents

Cigarette smoke (CS) inhalation causes an early inflammatory response in rodent airways by stimulating capsaicin-sensitive sensory neurons that express transient receptor potential cation channel, subfamily V, member 1 (TRPV1) through an unknown mechanism that does not involve TRPV1. We hypothesized that 2 alpha,beta-unsaturated aldehydes present in CS, crotonaldehyde and acrolein, induce neurogenic inflammation by stimulating TRPA1, an excitatory ion channel coexpressed with TRPV1 on capsaicin-sensitive nociceptors. We found that CS aqueous extract (CSE), crotonaldehyde, and acrolein mobilized Ca2+ in cultured guinea pig jugular ganglia neurons and promoted contraction of isolated guinea pig bronchi. These responses were abolished by a TRPA1-selective antagonist and by the aldehyde scavenger glutathione but not by the TRPV1 antagonist capsazepine or by ROS scavengers. Treatment with CSE or aldehydes increased Ca2+ influx in TRPA1-transfected cells, but not in control HEK293 cells, and promoted neuropeptide release from isolated guinea pig airway tissue. Furthermore, the effect of CSE and aldehydes on Ca2+ influx in dorsal root ganglion neurons was abolished in TRPA1-deficient mice. These data identify alpha,beta-unsaturated aldehydes as the main causative agents in CS that via TRPA1 stimulation mediate airway neurogenic inflammation and suggest a role for TRPA1 in the pathogenesis of CS-induced diseases.

Cigarette smoke exposure severely reduces peripheral insulin sensitivity without changing GLUT4 expression in oxidative muscle of Wistar rats

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

Sidestream cigarette smoke effects on cardiovascular responses in conscious rats: involvement of oxidative stress in the fourth cerebral ventricle

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

Association of diabetes and cigarette smoke exposure on the glycemia and liver glycogen of pregnant Wistar rats

Purpose: To evaluate cigarette smoke exposure and/or diabetes association effects on the glycemia and liver glycogen levels of pregnant Wistar rats. Methods: 60 adult rats were randomly distributed into (n= 10/group): non-diabetic exposed to filtered air (G1); non-diabetic exposed to cigarette smoke only before pregnancy (G2); non-diabetic exposed to cigarette smoke before and during pregnancy (G3); diabetic exposed to filtered air (G4); diabetic exposed to cigarette smoke only before pregnancy (G5), and diabetic exposed to cigarette smoke before and during pregnancy (G6). Glycemia was determined at days 0 and 21 of pregnancy. Liver samples were collected for liver glycogen determinations. Results: At day 21 of pregnancy, glycemia was higher in G5 and G6 compared to G4 group. G2 (2.43 +/- 0.43), G3 (3.20 +/- 0.49), G4 (2.62 +/- 0.34), G5 (2.65 +/- 0.27) and G6 groups (1.94 +/- 0.35) presented decreased liver glycogen concentrations compared to G1 (4.20 +/- 0.18 mg/100mg liver tissue) (p<0.05). G5 and G6 groups presented decreased maternal weight gain and litter weight. Conclusions: Severe diabetes and cigarette smoke exposure, alone or associated, caused impairment in liver glycogen storage at term pregnancy. Due to the fact that liver glycogen storages were considered determinant for glucose tolerance, it is relevant to point out a rigid clinical glycemic control and to stop smoking so earlier in pregnancy programming.

Could zinc prevent reproductive alterations caused by cigarette smoke in male rats?

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)