Long-Term Stress and Concomitant Marijuana Smoke Exposure Affect Physiology, Behavior and Adult Hippocampal Neurogenesis

Marijuana is a widely used recreational drug with increasing legalization worldwide for medical purposes. Most experimental studies use either synthetic or plant-derived cannabinoids to investigate the effect of cannabinoids on anxiety and cognitive functions. The aim of this study was to mimic real life situations where young people smoke cannabis regularly to relax from everyday stress. Therefore, we exposed young adult male NMRI mice to daily stress and concomitant marijuana smoke for 2 months and investigated the consequences on physiology, behavior and adult hippocampal neurogenesis. Animals were restrained for 6-h/day for 5-days a week. During the stress, mice were exposed to cannabis smoke for 2 × 30 min/day. We burned 2 “joints” (2 × 0.8 g marijuana) per occasion in a whole body smoking chamber. Cannabinoid content of the smoke and urine samples was measured by HPLC and SFC-MS/MS. Body weight gain was recorded daily and we did unrestrained, whole body plethysmography to investigate pulmonary functions. The cognitive performance of the animals was evaluated by the novel object recognition and Y maze tests. Anxietyrelated spontaneous locomotor activity and self-grooming were assessed in the open field test (OFT). Adult neurogenesis was quantified post mortem in the hippocampal dentate gyrus. The proliferative activity of the precursor cells was detected by the use of the exogenous marker 5-bromo-20-deoxyuridine. Treatment effects on maturing neurons were studied by the examination of doublecortin-positive neurons. Both stress and cannabis exposure significantly reduced body weight gain. Cannabis smoke had no effect on pulmonary functions, but stress delayed the maturation of several lung functions. Neither stress, nor cannabis smoke affected the cognitive functioning of the animals. Results of the OFT revealed that cannabis had a mild anxiolytic effect and markedly increased self-grooming behavior. Stress blocked cell proliferation in the dentate gyrus, but cannabis had no effect on this parameter. Marijuana smoke however had a pronounced impact on doublecortin-positive neurons influencing their number, morphology and migration. In summary, we report here that long-term stress in combination with cannabis smoke exposure can alter several health-related measures, but the present experimental design could not reveal any interaction between these two treatment factors except for body weight gain

Effects of Thymus vulgaris L., Cinnamomum verum J.Presl and Cymbopogon nardus (L.) Rendle Essential Oils in the Endotoxin-induced Acute Airway Inflammation Mouse Model

Thyme (TO), cinnamon (CO), and Ceylon type lemongrass (LO) essential oils (EOs) are commonly used for inhalation. However, their effects and mechanisms on inflammatory processes are not well-documented, and the number of in vivo data that would be important to determine their potential benefits or risks is low. Therefore, we analyzed the chemical composition and investigated the activity of TO, CO, and LO on airway functions and inflammatory parameters in an acute pneumonitis mouse model. The components of commercially available EOs were measured by gas chromatography–mass spectrometry. Airway inflammation was induced by intratracheal endotoxin administration in mice. EOs were inhaled during the experiments. Airway function and hyperresponsiveness were determined by unrestrained whole-body plethysmography on conscious animals. Myeloperoxidase (MPO) activity was measured by spectrophotometry from lung tissue homogenates, from which semiquantitative histopathological scores were assessed. The main components of TO, CO, and LO were thymol, cinnamaldehyde, and citronellal, respectively. We provide here the first evidence that TO and CO reduce inflammatory airway hyperresponsiveness and certain cellular inflammatory parameters, so they can potentially be considered as adjuvant treatments in respiratory inflammatory conditions. In contrast, Ceylon type LO inhalation might have an irritant e�ect (e.g., increased airway hyperresponsiveness and MPO activity) on the inflamed airways, and therefore should be avoided

Complex Regulatory Role of the TRPA1 Receptor in Acute and Chronic Airway Inflammation Mouse Models

The Transient Receptor Potential Ankyrin 1 (TRPA1) cation channel expressed on capsaicin-sensitive afferents, immune and endothelial cells is activated by inflammatory mediators and exogenous irritants, e.g., endotoxins, nicotine, crotonaldehyde and acrolein. We investigated its involvement in acute and chronic pulmonary inflammation using Trpa1 gene-deleted (Trpa1-/-) mice. Acute pneumonitis was evoked by intranasal Escherichia coli endotoxin (lipopolysaccharide: LPS) administration, chronic bronchitis by daily cigarette smoke exposure (CSE) for 4 months. Frequency, peak inspiratory/expiratory flows, minute ventilation determined by unrestrained whole-body plethysmography were significantly greater, while tidal volume, inspiratory/expiratory/relaxation times were smaller in Trpa1-/- mice. LPS-induced bronchial hyperreactivity, myeloperoxidase activity, frequency-decrease were significantly greater in Trpa1-/- mice. CSE significantly decreased tidal volume, minute ventilation, peak inspiratory/expiratory flows in wildtypes, but not in Trpa1-/- mice. CSE remarkably increased the mean linear intercept (histopathology), as an emphysema indicator after 2 months in wildtypes, but only after 4 months in Trpa1-/- mice. Semiquantitative histopathological scores were not different between strains in either models. TRPA1 has a complex role in basal airway function regulation and inflammatory mechanisms. It protects against LPS-induced acute pneumonitis and hyperresponsiveness, but is required for CSE-evoked emphysema and respiratory deterioration. Further research is needed to determine TRPA1 as a potential pharmacological target in the lung

Pinus sylvestris L. and Syzygium aromaticum (L.) Merr. & L. M. Perry Essential Oils Inhibit Endotoxin-Induced Airway Hyperreactivity despite Aggravated Inflammatory Mechanisms in Mice.

Scots pine (SO) and clove (CO) essential oils (EOs) are commonly used by inhalation, and their main components are shown to reduce inflammatory mediator production. The aim of our research was to investigate the chemical composition of commercially available SO and CO by gas chromatography-mass spectrometry and study their effects on airway functions and inflammation in an acute pneumonitis mouse model. Inflammation was evoked by intratracheal endotoxin and EOs were inhaled three times during the 24 h experimental period. Respiratory function was analyzed by unrestrained whole-body plethysmography, lung inflammation by semiquantitative histopathological scoring, myeloperoxidase (MPO) activity and cytokine measurements. α-Pinene (39.4%) was the main component in SO, and eugenol (88.6%) in CO. Both SO and CO significantly reduced airway hyperresponsiveness, and prevented peak expiratory flow, tidal volume increases and perivascular edema formation. Meanwhile, inflammatory cell infiltration was not remarkably affected. In contrast, MPO activity and several inflammatory cytokines (IL-1β, KC, MCP-1, MIP-2, TNF-α) were aggravated by both EOs. This is the first evidence that SO and CO inhalation improve airway function, but enhance certain inflammatory parameters. These results suggest that these EOs should be used with caution in cases of inflammation-associated respiratory diseases

The effects of Scots pine (Pinus sylvestris L.) essential oil in an endotoxin-induced acute airway inflammation mouse model

Inflammatory lung diseases affect a large population at every age worldwide. Essential oils (EOs) can easily reach the respiratory tract via inhalation due to their volatility. The antiinflammatory effect of EOs is poorly studied and there are only a few in vivo data. Therefore, we investigated the chemical composition and effects of Scots pine EO in the endotoxin-induced acute airway inflammation model in mice. The EO was selected on the basis of its potent antibacterial activity.                The chemical composition of the EO was determined by gas chromatography-mass spectrometry. Airway inflammation was evoked by 100 µg intratracheal endotoxin administration (Escherichia coli 083 lipopolysaccharide: LPS) to female C57BL/6 mice (n=8/group). There were three 30-min long inhalation sessions of the EO for during the 24-h period of the experiments. Airway function was measured by unrestrained whole-body plethysmography in conscious, awake animals. Myeloperoxidase (MPO) activity was determined by spectrophotometry from lung homogenates, while the semiquantitative histopathological score was evaluated from hematoxylin-eosin stained lung sections.                α-Pinene (39.4%) was the main component of the tested Scots pine oil. The inhalation of the EO significantly reduced peak inspiratory and expiratory flow and LPS-induced airway hyperresponsiveness compared to the paraffin oil-treated controls. Scots pine oil significantly reduced the perivascular edema formation. However, the EO significantly increased MPO activity. Scots pine oil or its components may be considered as a potential adjuvant in the treatment of respiratory symptoms

Longitudinal Analysis of Amplitude-Integrated Electroencephalography for Outcome Prediction in Hypoxic-Ischemic Encephalopathy

Objective: To investigate the prognostic accuracy of longitudinal analysis of amplitude-integrated electroencephalography (aEEG) background activity to predict long-term neurodevelopmental outcome in neonates with hypoxic-ischemic encephalopathy (HIE) receiving therapeutic hypothermia.Study design This single-center observational study included 149 neonates for derivation and 55 neonates for validation with moderate-severe HIE and of gestational age 35 weeks at a tertiary neonatal intensive care unit. Single-channel aEEG background pattern, sleep-wake cycling, and seizure activity were monitored over 84 hours during therapeutic hypothermia and rewarming, then scored for each 6-hour interval. Neurodevelopmental outcome was assessed using the Bayley Scales of Infant Development, Second Edition. Favorable outcome was defined as having both a Mental Development Index (MDI) score and Psychomotor Development Index (PDI) score >= 70, and adverse outcome was defined as either an MDI or a PDI <70 or death. Regression modeling for longitudinal analysis of repeatedly measured data was applied, and area under the receiver operating characteristic curve (AUC) was calculated.Resuits Longitudinal aEEG background analysis combined with sleep-wake cycling score had excellent predictive value (AUC, 0.90; 95% CI, 0.85-0.95), better than single aEEG scores at any individual time point. The model performed well in the independent validation cohort (AUC, 0.87; 95% CI, 0.62-1.00). The reclassification rate of this model compared with the conventional analysis of aEEG background at 48 hours was 18% (24 patients); 14% (18 patients) were reclassified correctly. Our results were used to develop a user-friendly online outcome prediction tool.Conclusions Longitudinal analysis of aEEG background activity and sleep-wake cycling is a valuable and accurate prognostic tool

Applicability of cinnamon bark essential oil in respiratory tract diseases – from in vitro to in vivo experiments

Introduction: Because respiratory tract diseases affect every age group and antibiotic resistance is an increasing problem in healthcare, there is a need for additional therapies. Essential oils (EOs) are used via inhalation for the treatment of respiratory tract diseases for a long time. Similarly to other plant extracts, their efficacy should also be proved in several test systems. Therefore, our aim was to study the antibacterial and antiinflammatory effects of cinnamon bark oil in in vitro and in vivo models.                Methods: The chemical composition of cinnamon EO was determined by GC-FID/MS and SPME-GC-MS methods. The antibacterial effect of the EO was tested by macrodilution and vapor-phase methods against respiratory tract pathogens. The emulsion of the EO prepared by nanotechnology was also used for the examination of the biofilm inhibitory effect. The antiinflammatory effect was studied in an LPS-induced acute airway inflammation mouse model.                Results and conclusion: The main component of the EO was trans-cinnamaldehyde (74.0%, 46.0%) in both analytical systems. In the liquid medium, cinnamon EO exhibited antibacterial activity against Streptococcus pyogenes, S. pneumoniae, S. mutans, Haemophilus influenzae and H. parainfluenzae (MIC: 0.06 mg/mL). In the vapor-phase test, the EO was the most effective against Haemophilus strains (MIC: 15.6 μL/L). The biofilm formation of S. mutans was more effectively reduced by the emulsion of cinnamon oil prepared by nanotechnology compared to the emulsions prepared with Tween80 or alcohol. In the animal model, cinnamon oil inhalation reduced airway hyperreactivity, macrophage accumulation in histological images, but did not affect MPO activity. Therefore, cinnamon oil may be a potential antibacterial and antiinflammatory agent for the treatment of respiratory tract diseases

Inhibition of NHE-1 Increases Smoke-Induced Proliferative Activity of Barrett’s Esophageal Cell Line

Several clinical studies indicate that smoking predisposes its consumers to esophageal inflammatory and malignant diseases, but the cellular mechanism is not clear. Ion transporters protect esophageal epithelial cells by maintaining intracellular pH at normal levels. In this study, we hypothesized that smoking affects the function of ion transporters, thus playing a role in the development of smoking-induced esophageal diseases. Esophageal cell lines were treated with cigarettesmoke extract (CSE), and the viability and proliferation of the cells, as well as the activity, mRNA and protein expression of the Na(+)/H(+) exchanger-1 (NHE-1), were studied. NHE-1 expression was also investigated in human samples. For chronic treatment, guinea pigs were exposed to tobacco smoke, and NHE-1 activity was measured. Silencing of NHE-1 was performed by using specific siRNA. CSE treatment increased the activity and protein expression of NHE-1 in the metaplastic cells and decreased the rate of proliferation in a NHE-1-dependent manner. In contrast, CSE increased the proliferation of dysplastic cells independently of NHE-1. In the normal cells, the expression and activity of NHE-1 decreased due to in vitro and in vivo smoke exposure. Smoking enhances the function of NHE-1 in Barrett’s esophagus, and this is presumably a compensatory mechanism against this toxic agent