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by Keyword: Animal model


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Menal, M. J., Jorba, I., Torres, M., Montserrat, J. M., Gozal, D., Colell, A., Piñol-Ripoll, G., Navajas, D., Almendros, I., Farré, R., (2018). Alzheimer's disease mutant mice exhibit reduced brain tissue stiffness compared to wild-type mice in both normoxia and following intermittent hypoxia mimicking sleep apnea Frontiers in Neurology 9, Article 1

Background: Evidence from patients and animal models suggests that obstructive sleep apnea (OSA) may increase the risk of Alzheimer’s disease (AD) and that AD is associated with reduced brain tissue stiffness. Aim: To investigate whether intermittent hypoxia (IH) alters brain cortex tissue stiffness in AD mutant mice exposed to IH mimicking OSA. Methods: Six-eight month old (B6C3-Tg(APPswe,PSEN1dE9)85Dbo/J) AD mutant mice and wild-type (WT) littermates were subjected to IH (21% O2 40 s to 5% O2 20 s; 6 h/day) or normoxia for 8 weeks. After euthanasia, the stiffness (E) of 200-μm brain cortex slices was measured by atomic force microscopy. Results: Two-way ANOVA indicated significant cortical softening and weight increase in AD mice compared to WT littermates, but no significant effects of IH on cortical stiffness and weight were detected. In addition, reduced myelin was apparent in AD (vs. WT), but no significant differences emerged in the cortex extracellular matrix components laminin and glycosaminoglycans when comparing baseline AD and WT mice. Conclusion: AD mutant mice exhibit reduced brain tissue stiffness following both normoxia and IH mimicking sleep apnea, and such differences are commensurate with increased edema and demyelination in AD.

Keywords: Animal model, Atomic force microscopy, Brain mechanics, Cortex stiffness, Neurodegenerative disease


Noguera-Ortega, Estela, Secanella-Fandos, Silvia, Eraña, Hasier, Gasión, Jofre, Rabanal, Rosa M., Luquin, Marina, Torrents, Eduard, Julián, Esther, (2016). Nonpathogenic Mycobacterium brumae inhibits bladder cancer growth in vitro, ex vivo, and in vivo European Urology Focus , 2, (1), 67-76

Background Bacillus Calmette-Guérin (BCG) prevents tumour recurrence and progression in non–muscle-invasive bladder cancer (BC). However, common adverse events occur, including BCG infections. Objective To find a mycobacterium with similar or superior antitumour activity to BCG but with greater safety. Design In vitro, ex vivo, and in vivo comparisons of the antitumour efficacy of nonpathogenic mycobacteria and BCG. Intervention The in vitro antitumour activity of a broad set of mycobacteria was studied in seven different BC cell lines. The most efficacious was selected and its ex vivo capacity to activate immune cells and its in vivo antitumour activity in an orthotopic murine model of BC were investigated. Outcome measurements and statistical analysis Growth inhibition of BC cells was the primary outcome measurement. Parametric and nonparametric tests were use to analyse the in vitro results, and a Kaplan-Meier test was applied to measure survival in mycobacteria-treated tumour-bearing mice. Results and limitations Mycobacterium brumae is superior to BCG in inhibiting low-grade BC cell growth, and has similar effects to BCG against high-grade cells. M. brumae triggers an indirect antitumour response by activating macrophages and the cytotoxic activity of peripheral blood cells against BC cells. Although no significant differences were observed between BCG and M. brumae treatments in mice, M. brumae treatment prolonged survival in comparison to BCG treatment in tumour-bearing mice. In contrast to BCG, M. brumae does not persist intracellularly or in tumour-bearing mice, so the risk of infection is lower. Conclusions Our preclinical data suggest that M. brumae represents a safe and efficacious candidate as a therapeutic agent for non–muscle-invasive BC. Patient summary We investigated the antitumour activity of nonpathogenic mycobacteria in in vitro and in vivo models of non–muscle-invasive bladder cancer. We found that Mycobacterium brumae effectively inhibits bladder cancer growth and helps the host immune system to eradicate cancer cells, and is a promising agent for antitumour immunotherapy.

Keywords: Animal models, Bacillus Calmette-Guérin, Cytokines, Immunomodulation, Immunotherapy, Mycobacteria, Urothelial cell line


Torres, M., Rojas, M., Campillo, N., Cardenes, N., Montserrat, J. M., Navajas, D., Farré, R., (2015). Parabiotic model for differentiating local and systemic effects of continuous and intermittent hypoxia Journal of Applied Physiology , 118, (1), 42-47

Hypoxia can be damaging either because cells are directly sensitive to low oxygen pressure in their local microenvironment and/or because they are exposed to circulating factors systemically secreted in response to hypoxia. The conventional hypoxia model, breathing hypoxic air, does not allow one to distinguish between these local and systemic effects. Here we propose and validate a model for differentially applying local and systemic hypoxic challenges in an animal. We used parabiosis, two mice sharing circulation by surgical union through the skin, and tested the hypothesis that when one of the parabionts breathes room air and the other one is subjected to hypoxic air, both mice share systemic circulation but remain normoxic and hypoxic, respectively. We tested two common hypoxic paradigms in 10 parabiotic pairs: continuous hypoxia (10% O2) mimicking chronic lung diseases, and intermittent hypoxia (40 s, 21% O2; 20 s, 5% O2) simulating sleep apnea. Arterial oxygen saturation and oxygen partial pressure at muscle tissue were measured in both parabionts. Effective cross-circulation was assessed by intraperitoneally injecting a dye in one of the parabionts and measuring blood dye concentration in both animals after 2 h. The results confirmed the hypothesis that tissues of the parabiont under room air were perfused with normally oxygenated blood and, at the same time, were exposed to all of the systemic mediators secreted by the other parabiont actually subjected to hypoxia. In conclusion, combination of parabiosis and hypoxic/normoxic air breathing is a novel approach to investigate the effects of local and systemic hypoxia in respiratory diseases.

Keywords: Animal model, Local hypoxia, Parabiosis, Systemic hypoxia


Dalmases, M., Torres, M., Márquez-Kisinousky, L., Almendros, I., Planas, A. M., Embid, C., Martínez-Garcia, M. A., Navajas, D., Farré, R., Montserrat, J. M., (2014). Brain tissue hypoxia and oxidative stress induced by obstructive apneas is different in young and aged rats Sleep , 37, (7), 1249-1256

Study Objectives: To test the hypotheses that brain oxygen partial pressure (PtO2) in response to obstructive apneas changes with age and that it might lead to different levels of cerebral tissue oxidative stress. Design: Prospective controlled animal study. Setting: University laboratory. Participants: Sixty-four male Wistar rats: 32 young (3 mo old) and 32 aged (18 mo). Interventions: Protocol 1: Twenty-four animals were subjected to obstructive apneas (50 apneas/h, lasting 15 sec each) or to sham procedure for 50 min. Protocol 2: Forty rats were subjected to obstructive apneas or sham procedure for 4 h. Measurements and Results: Protocol 1: Real-time PtO2 measurements were performed using a fast-response oxygen microelectrode. During successive apneas cerebral cortex PtO2 presented a different pattern in the two age groups; there was a fast increase in young rats, whereas it remained without significant changes between the beginning and the end of the protocol in the aged group. Protocol 2: Brain oxidative stress assessed by lipid peroxidation increased after apneas in young rats (1.34 ± 0.17 nmol/mg of protein) compared to old ones (0.63 ± 0.03 nmol/mg), where a higher expression of antioxidant enzymes was observed. Conclusions: The results suggest that brain oxidative stress in aged rats is lower than in young rats in response to recurrent apneas, mimicking obstructive sleep apnea. This could be due to the different PtO2 response observed between age groups and the increased antioxidant expression in aged rats.

Keywords: Aging, Animal model, Obstructive apnea, Oxidative stress, Tissue oxygenation, antioxidant, glutathione disulfide, aged, animal experiment, animal model, animal tissue, apnea, arterial oxygen saturation, article, brain cortex, brain oxygen tension, brain tissue, controlled study, groups by age, hypoxia, lipid peroxidation, male, nonhuman, oxidative stress, pressure, priority journal, rat


Melo, E., Cárdenes, N., Garreta, E., Luque, T., Rojas, M., Navajas, D., Farré, R., (2014). Inhomogeneity of local stiffness in the extracellular matrix scaffold of fibrotic mouse lungs Journal of the Mechanical Behavior of Biomedical Materials , 37, 186-195

Lung disease models are useful to study how cell engraftment, proliferation and differentiation are modulated in lung bioengineering. The aim of this work was to characterize the local stiffness of decellularized lungs in aged and fibrotic mice. Mice (2- and 24-month old; 14 of each) with lung fibrosis (N=20) and healthy controls (N=8) were euthanized after 11 days of intratracheal bleomycin (fibrosis) or saline (controls) infusion. The lungs were excised, decellularized by a conventional detergent-based (sodium-dodecyl sulfate) procedure and slices of the acellular lungs were prepared to measure the local stiffness by means of atomic force microscopy. The local stiffness of the different sites in acellular fibrotic lungs was very inhomogeneous within the lung and increased according to the degree of the structural fibrotic lesion. Local stiffness of the acellular lungs did not show statistically significant differences caused by age. The group of mice most affected by fibrosis exhibited local stiffness that were ~2-fold higher than in the control mice: from 27.2±1.64 to 64.8±7.1. kPa in the alveolar septa, from 56.6±4.6 to 99.9±11.7. kPa in the visceral pleura, from 41.1±8.0 to 105.2±13.6. kPa in the tunica adventitia, and from 79.3±7.2 to 146.6±28.8. kPa in the tunica intima. Since acellular lungs from mice with bleomycin-induced fibrosis present considerable micromechanical inhomogeneity, this model can be a useful tool to better investigate how different degrees of extracellular matrix lesion modulate cell fate in the process of organ bioengineering from decellularized lungs.

Keywords: Ageing, Atomic force microscopy, Decellularization, Lung fibrosis, Tissue engineering, Atomic force microscopy, Biological organs, Peptides, Sodium dodecyl sulfate, Sodium sulfate, Tissue engineering, Ageing, Decellularization, Extracellular matrices, Healthy controls, Inhomogeneities, Lung fibrosis, Micro-mechanical, Statistically significant difference, Mammals, bleomycin, adventitia, animal experiment, animal model, article, atomic force microscopy, bleomycin-induced pulmonary fibrosis, cell fate, controlled study, extracellular matrix, female, intima, lung alveolus, lung fibrosis, lung mechanics, mechanical probe, microenvironment, mouse, nonhuman, pleura, priority journal, rigidity, tissue engineering


Almendros, I., Montserrat, J. M., Torres, M., Bonsignore, M. R., Chimenti, L., Navajas, D., Farre, R., (2012). Obesity and intermittent hypoxia increase tumor growth in a mouse model of sleep apnea Sleep Medicine , 13, (10), 1254-1260

Background: Intermittent hypoxia and obesity which are two pathological conditions commonly found in patients with obstructive sleep apnea (OSA), potentially enhance cancer progression. Objective: To investigate whether obesity and/or intermittent hypoxia (IH) mimicking OSA affect tumor growth. Methods: A subcutaneous melanoma was induced in 40 mice [22 obese (40-45 g) and 18 lean (20-25 g)] by injecting 10(6) B16F10 cells in the flank. Nineteen mice (10 obese/9 lean) were subjected to IH (6 h/day for 17 days). A group of 21 mice (12 obese/9 lean) were kept under normoxia. At day 17, tumors were excised, weighed and processed to quantify necrosis and endothelial expression of vascular endothelial growth factor (VEGF) and CD-31. VEGF in plasma was also assessed. Results: In lean animals, IH enhanced tumor growth from 0.81 +/- 0.17 to 1.95 +/- 0.32 g. In obese animals, a similar increase in tumor growth (1.94 +/- 0.18 g) was observed under normoxia, while adding IH had no further effect (1.69 +/- 0.23 g). IH only promoted an increase in tumoral necrosis in lean animals. However, obesity under normoxic conditions increased necrosis, VEGF and CD-31 expression in tumoral tissue. Plasma VEGF strongly correlated with tumor weight (rho = 0.76, p < 0.001) in the whole sample; it increased in lean IH-treated animals from 66.40 +/- 3.47 to 108.37 +/- 9.48 pg/mL, p < 0.001), while the high baseline value in obese mice (106.90 +/- 4.32 pg/mL) was unaffected by IH. Conclusions: Obesity and IH increased tumor growth, but did not appear to exert any synergistic effects. Circulating VEGF appeared as a crucial mediator of tumor growth in both situations.

Keywords: Intermittent hypoxia, Obesity, Cancer, Sleep apnea, Animal model


Almendros, Isaac, Farre, Ramon, Planas, Anna M., Torres, Marta, Bonsignore, Maria R., Navajas, Daniel, Montserrat, Josep M., (2011). Tissue oxygenation in brain, muscle, and fat in a rat model of sleep apnea: Differential effect of obstructive apneas and intermittent hypoxia Sleep , 34, (8), 1127-1133

Study Objectives: To test the hypotheses that the dynamic changes in brain oxygen partial pressure (PtO(2)) in response to obstructive apneas or to intermittent hypoxia differ from those in other organs and that the changes in brain PtO(2) in response to obstructive apneas is a source of oxidative stress. Design: Prospective controlled animal study. Setting: University laboratory. Participants: 98 Sprague-Dawley rats. Interventions: Cerebral cortex, skeletal muscle, or visceral fat tissues were exposed in anesthetized animals subjected to either obstructive apneas or intermittent hypoxia (apneic and hypoxic events of 15 s each and 60 events/h) for 1 h. Measurements and Results: Arterial oxygen saturation (spO(2)) presented a stable pattern, with similar desaturations during both stimuli. The PtO(2) was measured by a microelectrode. During obstructive apneas, a fast increase in cerebral PtO(2) was observed (38.2 +/- 3.4 vs. 54.8 +/- 5.9 mm Hg) but not in the rest of tissues. This particular cerebral response was not found during intermittent hypoxia. The cerebral content of reduced glutathione was decreased after obstructive apneas (46.2% +/- 15.2%) compared to controls (100.0% +/- 14.7%), but not after intermittent hypoxia. This antioxidant consumption after obstructive apneas was accompanied by increased cerebral lipid peroxidation under this condition. No changes were observed for these markers in the other tissues. Conclusions: These results suggest the cerebral cortex could be protected in some way from hypoxic periods caused by obstructive apneas. The increased cerebral PtO(2) during obstructive apneas may, however, cause harmful effects (oxidative stress). The obstructive apnea model appears to be more adequate than the intermittent hypoxia model for studying brain changes associated with OSA.

Keywords: Tissue oxygenation, Obstructive apnea, Intermittent hypoxia, Animal model, Oxidative stress


Almendros, I., Farré, R., Torres, M., Bonsignore, M. R., Dalmases, M., Ramírez, J., Navajas, D., Montserrat, J. M., (2011). Early and mid-term effects of obstructive apneas in myocardial injury and inflammation Sleep Medicine , 12, (10), 1037-1040

Background: Obstructive sleep apnea (OSA) is associated with cardiovascular disorders, but the different comorbidities in OSA patients make it difficult to know their specific effects on the development of cardiovascular injury. The aim of the present study was to investigate whether recurrent obstructive apneas could lead to myocardial injury. Methods: Thirty-six male Sprague-Dawley rats (300-350. g) were either acutely (3. h) or sustainably (5. h/day, for 10. days) subjected to obstructive apneas with a pattern of 15. s each, 60. apneas/h. Corresponding control groups were formed for the acute and sustained models. To assess the induction of systemic inflammation, IL1-β was measured in plasma. Ventricular tissue injury was evaluated by histological techniques (presence of inflammatory cell infiltration, eosin autofluorescence, and detection of apoptosis). Results: After 3. h of obstructive apneas, a significant increase in IL1-β (64.9. ±. 29.6. ng/μl) were observed with respect to the controls (7.3. ±. 1.0. ng/μl), but no myocardial injury was present. Conversely to the acute model, the systemic inflammation triggered by obstructive apneas for 10. days was reduced. However, the percentage of area with enhanced eosin autofluorescence and of apoptotic cells (1.83. ±. 0.35% and 24.4. ±. 1.5%, respectively) was increased when compared to the control group (0.72. ±. 0.20% and 5.0. ±. 2.8%, respectively). Conclusions: This study suggests that obstructive apneas are a potential source of early systemic and ventricular inflammation and myocardial cell injury after sustained apneas application, which could represent an initial phase in the progression of heart disease associated with OSA.

Keywords: Animal models, Inflammation, Myocardial injury, Obstructive sleep apnea


Cagido, Viviane Ramos, Zin, Walter Araujo, Ramirez, Jose, Navajas, Daniel, Farre, Ramon, (2011). Alternating ventilation in a rat model of increased abdominal pressure Respiratory Physiology & Neurobiology , 175, (3), 310-315

During alternating ventilation (AV) one lung is inflating while the other is deflating. Considering the possible respiratory and hemodynamic advantages of AV, we investigated its effects during increased intra-abdominal pressure (IAP = 10 mmHg). In Sprague-Dawley rats (n = 6, 270–375 g) the main bronchi were independently cannulated, and respiratory mechanics determined while animals underwent different ventilatory patterns: synchronic ventilation without increased IAP (SV-0), elevated IAP during SV (SV-10), and AV with elevated IAP (AV-10). Thirty-three other animals (SV-0, n = 10; SV-10, n = 11 and AV-10, n = 12) were ventilated during 3 h. Mean arterial pressure (MAP), and lung histology were assessed. Increased IAP resulted in significantly higher elastances (p < 0.001), being AV-10 lower than SV-10 (p < 0.020). SV-10 showed higher central venous pressure (p < 0.003) than S-0; no change was observed in AV-10. Wet/dry lung weight ratio was lower in AV-10 than SV-10 (p = 0.009). Application of AV reduced hemodynamic and lung impairments induced by increased IAP during SV.

Keywords: Alternating ventilation, Respiratory mechanics, Intra-abdominal pressure, Hemodynamic, Mechanical ventilation, Animal model


Carreras, Alba, Wang, Yang, Gozal, David, Montserrat, Josep M., Navajas, Daniel, Farre, Ramon, (2011). Non-invasive system for applying airway obstructions to model obstructive sleep apnea in mice Respiratory Physiology & Neurobiology , 175, (1), 164-168

Obstructive sleep apnea (OSA) is characterized by recurrent upper airway obstructions during sleep. The most common animal model of OSA is based on subjecting rodents to intermittent hypoxic exposures and does not mimic important OSA features, such as recurrent hypercapnia and increased inspiratory efforts. To circumvent some of these issues, a novel murine model involving non-invasive application of recurrent airway obstructions was developed. An electronically controlled airbag system is placed in front of the mouse's snout, whereby inflating the airbag leads to obstructed breathing and spontaneous breathing occurs with the airbag deflated. The device was tested on 29 anesthetized mice by measuring inspiratory effort and arterial oxygen saturation (SaO(2)). Application of recurrent obstructive apneas (6s each, 120/h) for 6h resulted in SaO(2) oscillations to values reaching 84.4 +/- 2.5% nadir, with swings mimicking OSA patients. This novel system, capable of applying controlled recurrent airway obstructions in mice, is an easy-to-use tool for investigating pertinent aspects of OSA.

Keywords: Animal model, Upper airway Obstruction, Mouse model, Non-invasive system, Model sleep apnea, Respiratory disease


Carreras, Alba, Almendros, Isaac, Montserrat, Josep M., Navajas, Daniel, Farre, Ramon, (2010). Mesenchymal stem cells reduce inflammation in a rat model of obstructive sleep apnea Respiratory Physiology & Neurobiology , 172, (3), 210-212

The aim was to test the hypothesis that mesenchymal stem cells (MSC) could reduce the inflammation induced by recurrent airway occlusions in an animal model of obstructive sleep apnea (OSA). A nasal mask was applied to 30 anesthetized rats. Twenty rats were subjected to a pattern of recurrent obstructive apneas mimicking OSA (60/h, lasting 15 s each) for 5h. MSC (5x10(6) cells) were intravenously injected into 10 of these rats. Ten rats not subjected to apneas or MSC injection were used as controls. The rat blood serum concentrations of pro-inflammatory cytokine IL-1beta were measured by ELISA. IL-1beta was significantly greater in the rats subjected to recurrent apneas (66.7+/-41.2 pg/mL; m+/-SEM) than in controls (1.9+/-1.0 pg/mL; p<0.05). In the group of apneic rats subjected to MSC injection, IL-1beta was significantly reduced (6.1+/-3.8 pg/mL; p<0.05). In conclusion, MSC triggered an early anti-inflammatory response in rats subjected to recurrent obstructive apneas, suggesting that these stem cells could play a role in the physiological response to counterbalance inflammation in OSA.

Keywords: Obstructive sleep apnea, Animal model, Airway obstruction, Inflammation


Farre, R., Montserrat, J. M., Navajas, D., (2008). Morbidity due to obstructive sleep apnea: insights from animal models Current Opinion in Pulmonary Medicine , 14, (6), 530-536

PURPOSE OF REVIEW: Obstructive sleep apnea (OSA) is a prevalent disorder with clinically well known mid-term and long-term consequences. It is difficult, however, to investigate the mechanisms causing morbidity in OSA from human studies, owing to confounding factors in patients. Animal research is useful to analyze the various injurious stimuli--intermittent hypoxia/hypercapnia, mechanical stress and sleep disruption--that potentially cause OSA morbidity. This review is focused on the most recent advances in our understanding of the consequences of OSA, achieved as a result of animal models. RECENT FINDINGS: Animal research has improved our knowledge of various aspects of the cardiovascular consequences of OSA: myocardial damage, left ventricular dysfunction, vasoconstriction, hypertension and atherosclerosis. The systemic and metabolic consequences of OSA--inflammation, insulin resistance, alterations in lipid metabolism and hepatic morbidity--have also been investigated with animal models. Our understanding of the mechanisms involved in the neurocognitive consequences of OSA--neuronal and brain alterations and cognitive dysfunctions--has also been improved through animal research. Moreover, animal models have recently been used to investigate the mechanisms of upper airway inflammation and dysfunction. SUMMARY: The simple experimental models used to investigate OSA morbidity are useful for investigating isolated mechanisms. However, more complex and realistic models incorporating the various injurious challenges characterizing OSA are required to more precisely translate the results of animal research to patients and to design potentially preventive and therapeutic strategies.

Keywords: Animal model, Morbidity, Sleep apnea, Translational research


Farre, R., Nacher, M., Serrano-Mollar, A., Galdiz, J. B., Alvarez, F. J., Navajas, D., Montserrat, J. M., (2007). Rat model of chronic recurrent airway obstructions to study the sleep apnea syndrome Sleep , 30, (7), 930-933

Study Objectives: To implement a chronic rat model of recurrent airway obstructions to study the obstructive sleep apnea (OSA) syndrome. Design: Prospective controlled animal study. Setting: University laboratory. Patients or Participants: 24 male Sprague-Dawley rats (250-300 g). Interventions: The rats were placed in a setup consisting of a body chamber and a head chamber separated by a neck collar specially designed to apply recurrent airway obstructions with OSA patterns. Rats in the Obstruction group (n=8) were subjected to 5-s obstructions at a rate of 60 per hour, 6 h/day during 4 weeks. Sham rats (n=8) were placed in the setup but no obstructions were applied. Naive rats (n=8) were subjected to no intervention. Measurements and Results: Breathing flow, pressure, CO2 air concentration, and SpO(2) showed that the model mimicked OSA respiratory events (obstructive apneas, increased respiratory efforts, and oxygen saturation dips). Animal stress, assessed by body weight and plasma corticosterone, was not significantly different across Obstruction and Sham groups. This supports the concept that this novel model does not introduce a significant burden of stress in the rat after acclimatization to the chamber. Thromboxane-B2/6-keto-Prostaglandin-F1a ratio in plasma, which is an index of vasoconstriction, was significantly increased in the rats subjected to obstructions. Conclusions: The designed animal model of chronic recurrent airway obstructions is feasible and potentially useful to study the mechanisms involved in the cardiovascular consequences of OSA.

Keywords: Obstructive sleep apnea, Animal model, Airway obstruction