Guinea Pig Lung Research Articles

Pathogenicity of Bordetella bronchiseptica isolated from apparently healthy rabbits in guinea pig, rat, and mouse.

Bordetella bronchiseptica (B. bronchiseptica) is associated with respiratory tract infections in laboratory animals. In our laboratory animal facility, B. bronchiseptica was isolated from 21 of 27 apparently healthy rabbits obtained from a breeding farm contaminated with B. bronchiseptica. Restriction fragment length polymorphism (RFLP) analysis showed that the flagellin genotype of isolates from the laboratory animal facility and breeding farm was type A, which is seen relatively frequently in rabbits in Europe. To examine its pathogenicity, guinea pigs, rats, and mice were inoculated intranasally with a representative strain isolated in the laboratory animal facility. Following inoculation of 107 colony forming unit (cfu), severe inflammation was observed in the lungs of guinea pig and mice, although the inflammation was less severe in rats. The strain was recovered from the trachea and lungs of these species after inoculation with lower dose such as 103 or 104 cfu. These results suggest that the isolated strain causes respiratory tract infection in guinea pigs, rats, and mice, and that its pathogenicity higher in mice than in rats. This study extends our knowledge of interpreting the microbiologic status of laboratory animals, which will contribute to the development of reliable and reproducible animal experiments.

Dynamics of quantitative changes of diffuse lymphoid tissue cells of bronchi and lungs of guinea pigs sensitized with ovalbumin

Background. There is a progressive increase of respiratory allergic diseases nowadays in the world, made it necessary to study the participation of the components of the immune system in these processes. From the point of view of modern aspects about the organs of the immune system, interesting are the patterns of morphogenesis and function of bronchus associated lymphoid tissue, occupied a special place in the immunological protection of the body due to the large area of contact with various antigens. Morphogenesis and reactive changes in the local immunity in respiratory system in conditions of allergic inflammation remain urgent issue of modern morphology. Objective. To study the changes in diffuse lymphoid tissue of bronchi and lungs of guinea pigs sensitized with ovalbumin. Methods. We have studied the lung of 48 guinea pigs, using histological, immunohistochemical, morphometric, statistical methods, under conditions of experimental ovalbumin-induced allergic inflammation, assessed the average number of lymphocytes, macrophages and plasma cells in the diffuse lymphoid tissue. Results. The average number of lymphocytes in diffuse lymphoid tissue of bronchi and lungs increased from the 23rd day of observation and remained at a high level until the end of the experiment, the maximum was during the early period of the development of allergic inflammation, the increasing coefficient was 4.7. The average number of plasma cells also acquired maximum elevation in the early period of allergic process, the increasing coefficient was 2.0. The most significant average number of macrophages was on the 23rd day of observation with same increasing coefficient. Among all types of immunocompetent cells of diffuse lymphoid tissue in bronchi and lungs, T-lymphocytes prevailed during the experiment elevated almost by 5 times. Conclusions. In the early period of development of experimental ovalbumin-induced allergic inflammation, the specific resistance of the respiratory system manifests itself in the form of activation of local links of cellular and humoral adaptive immunity, as evidenced by the dynamics of changes in the average number of lymphocytes (the maximum increasing coefficient 4.7 in the 1st experimental group), macrophages and plasma cells (maximum increasing coefficient 2.0 in the 1st experimental group) of diffuse lymphoid tissue of bronchi and lungs of guinea pigs.

DISTRIBUTION AND QUANTITATIVE CHANGES OF MAST CELLS IN GUINEA PIGS LUNG IN OVALBUMIN-INDUCED ALLERGIC INFLAMMATION

Background. One of the most important cells in local immunity in lung are mast cells. They are involved in both innate and adaptive immune responses to inhaled allergens. The question of distribution of these both cell types in guinea pig lung in case of experimental allergic inflammation in most aspects remains open. Objective. The aim of this research is to study the distribution and quantitative changes of mast cells in lung of guinea pigs in ovalbumin-induced allergic inflammation. Methods. the lungs of 48 male guinea pigs have been studied using histological, morphometric and statistical methods in cases of experimental ovalbumin-induced allergic inflammation. The total number of mucosa related must cells and perivascular mast cells in guinea pig lungs were counted. Results. It has been established that mucosa related mast cells are normally more abundant in guinea pigs lung than perivascular ones. Maximum increase in a number of mucosa related mast cells was revealed in the early period of allergic inflammation, as evidenced by maximum increase coefficient of 1.4 in the 1st experimental group, compare to the control (P*/**<0.05). However, maximum increase in number of perivascular mast cells in 5 times was found during the late period of allergic inflammation in the 4th experimental group (P*/**<0.05). Conclusion. Experimental sensitization and challenge with ovalbumin leads to statistically significant increase in average number of both types of mast cells but predominantly the latter ones. It has been proved that cells dynamics is multidirectional.

Peculiarities of changes in a functional condition of pro-oxidant and antioxidant systems in guinea pigs’ lungs in experimental allergic alveolitis under the conditions of immobilization stress

We have analyzed the results of investigation of alterations in indices of pro-oxidant (conjugated diene and malondialdehyde) and antioxidant (superoxide dismutase, ceruloplasmin, catalase) systems in guinea pigs’ lungs in experimental allergic alveolitis in the dynamics of EAA development under the conditions of immobilization stress. The investigation was conducted on 62 female guinea pigs weighing 180-220 g, divided into 4 groups: I – intact guinea pigs (n=20), II – guinea pigs (n=14) with EAA under the conditions of IS (1st day); III – guinea pigs (n=14) with EAA under the conditions of IS (2nd day); IV – guinea pigs (n=14) with EAA under the conditions of IS (64th day). The results of experimental investigation showed that a significant increase in conjugated diene level in animals’ lungs was observed at all stages of EAA development under the conditions of immobilization stress as compared with control group, indicating activation of this marker. The same changes occurred with MDA content, indicating excessive accumulation of this lipid peroxidation product in lung tissue. Intensive synthesis of free radical compounds caused activation of some components of enzymatic system of antioxidant defense. In particular, a moderate decrease in superoxide dismutase activity in lung tissue is observed in response to increased level of LOPs at early stages of EAA and immobilization stress development as compared with the indices in intact animals. The same situation is observed with catalase and ceruloplasmin activity in the lungs of guinea pigs with modeled AA and IS.

Reaction of eosinophilic granulocytes of guinea pigs airways to the experimental allergic inflammation according to the morphometric study

Background. After creating an animal model of experimental airways allergic inflammation using ovalbumin, scientists mainly studied the reaction of the cellular and humoral links of the acquired specific immunity. At the same time, it is known that the development of allergic inflammation of the airways to the effect of the environmental chemoattractants is primarily the result of the local innate immune system response. Pulmonary neuroendocrine cells of the airway epithelium with the help of neuropeptides activate the secretion of IL-5 and IL-13 by type 2 innate lymphoid cells IL-5 determines the eosinophilic inflammation. The question of the reaction of the components of innate immunity of the respiratory tract to the allergic inflammation in most aspects remains open. open. Objective. The purpose of the current study was to define the reaction of eosinophilic granulocytes in guinea pigs lungs as an important component of the innate immunity of the respiratory tract to experimental ovalbumin-induced allergic inflammation according to the data of a morphometric study. Methods. Histological, morphometric, statistical methods were used to study the topographic features and the number of eosinophils in the lungs of male 48 guinea pigs on the 23rd, 30th, 36th and 44th days after initiation of the experimental ovalbumin-induced allergic airway inflammation. Results. Our study demonstrates a significant reaction occurs on the part of the cellular link of innate immunity, which consists in the activation of eosinophils, in the experimental model of ovalbumin-induced airways allergic inflammation. The most significant changes were observed in the distal parts of the intrapulmonary airways and in the structures of the pulmonary acinus in the early period of the development of the inflammatory process (23rd and 30th days after the start of the experiment). The late (36th and 44th days after the start of the experiment) period of the development of an allergic inflammatory process in the lungs is accompanied by a gradual decrease in the activity of eosinophilic inflammation. At the same time, the quantitative indicators remain statistically significantly higher in the intact and control groups, which indicates the continuation of allergic inflammation in the absence of the action of the allergen and is a manifestation of the violation of the recovery and adaptation processes in the local immune system of the lung. Conclusion. After ovalbumin-sensitization and aeroallergization eosinophilic inflammation develops in the lungs of a guinea pig, as a result of the reaction of the innate immunity to the action of an allergen. Actively and first of all, allergic eosinophilic inflammation develops in the distal airways (terminal bronchioles) and in the connective tissue stroma of the pulmonary acinus.

Vaccination with BCG\u0394BCG1419c protects against pulmonary and extrapulmonary TB and is safer than BCG

A single intradermal vaccination with an antibiotic-less version of BCGΔBCG1419c given to guinea pigs conferred a significant improvement in outcome following a low dose aerosol exposure to M. tuberculosis compared to that provided by a single dose of BCG Pasteur. BCGΔBCG1419c was more attenuated than BCG in murine macrophages, athymic, BALB/c, and C57BL/6 mice. In guinea pigs, BCGΔBCG1419c was at least as attenuated as BCG and induced similar dermal reactivity to that of BCG. Vaccination of guinea pigs with BCGΔBCG1419c resulted in increased anti-PPD IgG compared with those receiving BCG. Guinea pigs vaccinated with BCGΔBCG1419c showed a significant reduction of M. tuberculosis replication in lungs and spleens compared with BCG, as well as a significant reduction of pulmonary and extrapulmonary tuberculosis (TB) pathology measured using pathology scores recorded at necropsy. Evaluation of cytokines produced in lungs of infected guinea pigs showed that BCGΔBCG1419c significantly reduced TNF-α and IL-17 compared with BCG-vaccinated animals, with no changes in IL-10. This work demonstrates a significantly improved protection against pulmonary and extrapulmonary TB provided by BCGΔBCG1419c in susceptible guinea pigs together with an increased safety compared with BCG in several models. These results support the continued development of BCGΔBCG1419c as an effective vaccine for TB.

Morphological features of histogenic differon cells in connective tissue of guinea pigs’ lungs after sensitization with ovalbumin

An urgent issue of modern morphology is establishing a number of patterns of morphological changes and reactivity of connective tissue components of lungs in case of experimental sensitization with allergens. The aim is to estimate morphological features of histogenic differon cells in connective tissue of guinea pigs’ lungs after sensitization with ovalbumin. Materials and methods. Using morphometric and histological method, we have estimated the lung connective tissue of 48 male guinea pigs with experimental ovalbumin-induced allergic inflammation, simulated by subcutaneous sensitization and aeroallergization with ovalbumin. The number of fibrocytes, fibroblasts per 5000 μm2 and their ratio – fibroblast/fibrocyte coefficient were determined. Results. We have established the regularity of morphological changes dynamics in the cellular elements of pulmonary connective tissue. Experimental sensitization and inhalated allergization with ovalbumin leads to a statistically significant increase in the average number of fibroblasts and fibrocytes throughout the observation period in all experimental groups. It has been proved that the dynamics of cells has a multidirectional character, demonstrated by indicators of the fibroblast/fibrocyte coefficient, which shows the disproportion in the fibroblast/fibrocyte ratio and proves the tendency to the development of fibrosis in guinea pigs’ pulmonary connective tissue in case of experimental sensitization with ovalbumin. Conclusions. A gradual increase in the number of fibrocytes, against the background of a decrease in the number of fibroblasts is observed from the 23rd day to the completion of experimental sensitization with ovalbumin in the lungs of guinea pigs, compared with control group. A decrease of fibroblast/fibrocyte coefficient from 1.37 ± 0.03 in the early period to 0.82 ± 0.03 in the late period of the allergic inflammation demonstrates multidirectional nature of the dynamics in the number of connective tissue cells and indicates a tendency towards the development of fibrosis in pulmonary connective tissue.

Morphological rearrangement of the metabolic link of the microcirculatory bed of guinea pigs lungs after sensitization with ovalbumin

The reaction of the lung microvessels is an urgent issue of morphology and medicine in general, as well as one of the insufficiently investigated points in the study of morphological changes in chronic allergic diseases of the respiratory system.
 The aim is to study the morphological changes in the vessels of the metabolic link of the microcirculation in the lungs of guinea pigs sensitized with ovalbumin.
 Materials and methods. We have studied the lungs of 48 guinea pigs, using histological and morphometric methods, under conditions of experimental ovalbumin-induced allergic inflammation, simulated by three-time subcutaneous sensitization and subsequent 8-day intranasal inhalation of ovalbumin. To estimate the structural and functional state of capillaries, the inner diameter of the vessels was determined.
 Results. A general regularity in the reactivity of the hemomicrocirculatory bed of guinea pig lungs in experimental ovalbumin-induced allergic inflammation was established, which consists in a significant structural and functional restructuring of the exchange vessels of the microcirculatory bed. Dysfunction of the capillaries endothelium is manifested by a change of vasodilatation to vasospasm, as evidenced by morphometric changes in the diameter of the capillaries lumen in the experimental groups, and an increase in the permeability of the capillaries wall, which is confirmed by edema and disorganization of the connective tissue component.
 Conclusions. Allergic inflammation leads to structural and functional reorganization of the metabolic link of the hemomicrocirculatory bed of guinea pig lungs, which has a multidirectional staging character depending on the duration of the experiment and is a manifestation of a violation of the compensatory-adaptive processes of hemomicrocirculation. The most pronounced changes in the form of a decrease in the diameter of the lumen of the blood capillaries of the lungs of guinea pigs by 23 % compared to the control are observed during the late period of the development of the allergic inflammatory process.

The novel TRPA1 antagonist BI01305834 inhibits ovalbumin-induced bronchoconstriction in guinea pigs

BackgroundAsthma is a chronic respiratory disease in which the nervous system plays a central role. Sensory nerve activation, amongst others via Transient Receptor Potential Ankyrin 1 (TRPA1) channels, contributes to asthma characteristics including cough, bronchoconstriction, mucus secretion, airway hyperresponsiveness (AHR) and inflammation. In the current study, we evaluated the efficacy of the novel TRPA1 antagonist BI01305834 against AHR and inflammation in guinea-pig models of asthma.MethodsFirst, a pilot study was performed in a guinea-pig model of allergic asthma to find the optimal dose of BI01305834. Next, the effect of BI01305834 on (1) AHR to inhaled histamine after the early and late asthmatic reaction (EAR and LAR), (2) magnitude of EAR and LAR and (3) airway inflammation was assessed. Precision-cut lung slices and trachea strips were used to investigate the bronchoprotective and bronchodilating-effect of BI01305834. Statistical evaluation of differences of in vivo data was performed using a Mann–Whitney U test or One-way nonparametric Kruskal–Wallis ANOVA, for ex vivo data One- or Two-way ANOVA was used, all with Dunnett’s post-hoc test where appropriate.ResultsA dose of 1 mg/kg BI01305834 was selected based on AHR and exposure data in blood samples from the pilot study. In the subsequent study, 1 mg/kg BI01305834 inhibited AHR after the EAR, and the development of EAR and LAR elicited by ovalbumin in ovalbumin-sensitized guinea pigs. BI01305834 did not inhibit allergen-induced total and differential cells in the lavage fluid and interleukin-13 gene expression in lung homogenates. Furthermore, BI01305834 was able to inhibit allergen and histamine-induced airway narrowing in guinea-pig lung slices, without affecting histamine release, and reverse allergen-induced bronchoconstriction in guinea-pig trachea strips.ConclusionsTRPA1 inhibition protects against AHR and the EAR and LAR in vivo and allergen and histamine-induced airway narrowing ex vivo, and reverses allergen-induced bronchoconstriction independently of inflammation. This effect was partially dependent upon histamine, suggesting a neuronal and possible non-neuronal role for TRPA1 in allergen-induced bronchoconstriction.

The potential of microdialysis to estimate rifampicin concentrations in the lung of guinea pigs.

Optimised pre-clinical models are required for TB drug development to better predict the pharmacokinetics of anti-tuberculosis (anti-TB) drugs to shorten the time taken for novel drugs and combinations to be approved for clinical trial. Microdialysis can be used to measure unbound drug concentrations in awake freely moving animals in order to describe the pharmacokinetics of drugs in the organs as a continuous sampling technique. The aim of this work was to develop and optimise the microdialysis methodology in guinea pigs to better understand the pharmacokinetics of rifampicin in the lung. In vitro experiments were performed before progressing into in vivo studies because the recovery (concentration of the drug in the tissue fluid related to that in the collected dialysate) of rifampicin was dependent on a variety of experimental conditions. Mass spectrometry of the dialysate was used to determine the impact of flow rate, perfusion fluid and the molecular weight cut-off and membrane length of probes on the recovery of rifampicin at physiologically relevant concentrations. Following determination of probe efficiency and identification of a correlation between rifampicin concentrations in the lung and skeletal muscle, experiments were conducted to measure rifampicin in the sacrospinalis of guinea pigs using microdialysis. Lung concentrations of rifampicin were estimated from the rifampicin concentrations measured in the sacrospinalis. These studies suggest the potential usefulness of the microdialysis methodology to determine drug concentrations of selected anti-TB drugs to support new TB drug development.

Early Life Deficiencies in Ascorbate and Cysteine Impact the Long-term Glutathione Metabolism in Liver and Lungs of Guinea Pigs

Early Life Deficiencies in Ascorbate and Cysteine Impact the Long-term Glutathione Metabolism in Liver and Lungs of Guinea Pigs

Modeling rapidly adapting pulmonary stretch receptor activity to step-wise and constant pressure inflation of the lungs

Rapidly-adapting pulmonary stretch receptors (RAPSRs) provide the central nervous system with information regarding the rate of lung inflation, lung compliance and the sensation of dyspnea. Other than satisfying parameters of an adaptation index to constant pressure lung inflation for identification, no mathematical model has been ascribed to the stimulus-response relationship of lung volume-pressure to RAPSR activity. Herein, linear, power, polynomial and non-linear (four parameters logistic) models are tested for the best “goodness of fit” line of RAPSR activity to step-wise lung inflation to four times tidal volume and constant pressure inflation to 10, 20, 30 and 40 cm H2O of the lungs of guinea pigs and dogs. Goodness of fit was determined by evaluating coefficient of determination (R2) and visual inspection. The best “goodness of fit” is one of a non-linear symmetrical, stimulus-response function.

ФУНКЦІОНАЛЬНИЙ СТАН ПРООКСИДАНТНОЇ ТА АНТИОКСИДАНТНОЇ СИСТЕМ У ЛЕГЕНЯХ ЗА УМОВ ЕКСПЕРИМЕНТАЛЬНОГО АЛЬВЕОЛІТУ І СТРЕСУ

We have analysed the results of the study of alterations in indices of pro-oxidant (conjugated diene and malondialdehyde) and antioxidant (superoxide dismutase, ceruloplasmin, catalase) systems in guinea pigs’lungs in experimental allergic alveolitis in the dynamics of EAA development under the conditions of immobilization stress. The study included 62 female guinea pigs weighing 180-220 g, divided into 4 groups: 1 – intact guinea pigs (n=20); 2 – guinea pigs (n=14) with EAA under conditions of IS (1st day); 3 – guinea pigs (n=14) with EAA under conditions of IS (2nd day); 4 – guinea pigs (n=14) with EAA under conditions of IS (34th day). The results of the experimental study showed that a signifi cant increase in conjugated diene level in animal lungs was observed at all stages of EAA development under conditions of immobilization stress as compared to a control group, indicating the activation of this marker. The same changes occurred with MDA content, indicating excessive accumulation of this lipid peroxidation product in lung tissue. Intensive synthesis of free radical compounds caused activation of some components of the enzymatic system of antioxidant defence. In particular, a moderate decrease in superoxide dismutase activity in lung tissue is observed in response to an increased level of LOPs at early stages of EAA and immobilization stress development as compared to these indices in intact animals. The same situation is observed with catalase and ceruloplasmin activity in the lungs of guinea pigs with modelled AA and IS.

Induction of autophagy through CLEC4E in combination with TLR4: an innovative strategy to restrict the survival of Mycobacterium tuberculosis

ABSTRACT Host-directed therapies are gaining considerable impetus because of the emergence of drug-resistant strains of pathogens due to antibiotic therapy. Therefore, there is an urgent need to exploit alternative and novel strategies directed at host molecules to successfully restrict infections. The C-type lectin receptor CLEC4E and Toll-like receptor TLR4 expressed by host cells are among the first line of defense in encountering pathogens. Therefore, we exploited signaling of macrophages through CLEC4E in association with TLR4 agonists (C4.T4) to control the growth of Mycobacterium tuberculosis (Mtb). We observed significant improvement in host immunity and reduced bacterial load in the lungs of Mtb-infected mice and guinea pigs treated with C4.T4 agonists. Further, intracellular killing of Mtb was achieved with a 10-fold lower dose of isoniazid or rifampicin in conjunction with C4.T4 than the drugs alone. C4.T4 activated MYD88, PtdIns3K, STAT1 and RELA/NFKB, increased lysosome biogenesis, decreased Il10 and Il4 gene expression and enhanced macroautophagy/autophagy. Macrophages from autophagy-deficient (atg5 knockout or Becn1 knockdown) mice showed elevated survival of Mtb. The present findings also unveiled the novel role of CLEC4E in inducing autophagy through MYD88, which is required for control of Mtb growth. This study suggests a unique immunotherapeutic approach involving CLEC4E in conjunction with TLR4 to restrict the survival of Mtb through autophagy. Abbreviations 3MA: 3 methyladenine; AO: acridine orange; Atg5: autophagy related 5; AVOs: acidic vesicular organelles; BECN1: beclin 1, autophagy related; BMDMs: bone marrow derived macrophages; bw: body weight; C4.T4: agonists of CLEC4E (C4/TDB) and TLR4 (T4/ultra-pure-LPS); CFU: colony forming unit; CLEC4E/Mincle: C-type lectin domain family 4, member e; CLR: c-type lectin receptor; INH: isoniazid; LAMP1: lysosomal-associated membrane protein 1; MφC4.T4: Mtb-infected C4.T4 stimulated macrophages; MAP1LC3/LC3: microtubule-associated protein 1 light chain 3; MDC: monodansylcadaverine; MTOR: mechanistic target of rapamycin kinase; MYD88: myeloid differentiation primary response 88; NFKB: nuclear factor of kappa light polypeptide gene enhance in B cells; NLR: NOD (nucleotide-binding oligomerization domain)-like receptors; PFA: paraformaldehyde; PPD: purified protein derivative; PtdIns3K: class III phosphatidylinositol 3-kinase; RELA: v-rel reticuloendotheliosis viral oncogene homolog A (avian); RIF: rifampicin; RLR: retinoic acid-inducible gene-I-like receptors; TDB: trehalose-6,6´-dibehenate; TLR4: toll-like receptor 4; Ultra-pure-LPS: ultra-pure lipopolysaccharide-EK; V-ATPase: vacuolar-type H+ ATPase.

OXIDATIVE STRESS MARKERS IN THE LUNGS UNDER EXPERIMENTAL HYPEROXIA WITH INHALED LIPOSOMES BASED ON EGG LECITHIN

Oxidative stress is considered to be a factor leading to lung damage in premature infants. The aim of this study was to investigate the effect of inhaled antioxidants incorporated into egg lecithin liposomes on the indicators of oxidative stress in the lungs of newborn guinea pigs under experimental hyperoxia (3 days). Bronchoalveolar lavage fl uid (BALF) was used as a material for the study. Under hyperoxia exposure, inhalation of liposomes containing N-acetylcysteine and alpha-tocopherol contributed to the suppression of the reactive oxygen species production by cells, normalization of glutathione peroxidase activity and carbonyls content, while not affecting the level of diene conjugates in BALF. The introduction of retinoid-containing liposomes (retinol and retinoic acid) under hyperoxia was accompanied by normalization of glutathione peroxidase activity as well as the content of protein oxidation products in BALF, while the generation of reactive oxygen species remained enhanced, and the diene conjugates and thiobarbituric acid reactive products exceeded the levels in animals exposed to hyperoxia alone. Thus, the inhaled liposomes containing retinoids and egg lecithin exhibit not only anti-, but also a prooxidant effect in the lungs under hyperoxia exposure, unlike the liposomal forms of N-acetylcysteine and alpha-tocopherol.

Effect of dexmedetomidine on pyroptosis during lung ischemia-reperfusion in rats: an in vitro experiment

Objective To evaluate the effect of dexmedetomidine on pyroptosis during lung ischemia-reperfusion (I/R) in rats. Methods Adult male Sprague-Dawley rats, weighing 250-320 g, were used in this study.The model of isolated lung perfusion was established using an IL-2 Isolated Perfused Rat or Guinea Pig Lung System after the rats were anesthetized.Thirty lungs in which an ex vivo lung perfusion model was successfully established were divided into 3 groups (n=10 each) by a random number table method: control group (C group), I/R group and dexmedetomidine group (DEX group). The lungs were continuously perfused with K-H solution for 150 min in C group.After 15 min of perfusion, lungs were subjected to 60-min suspension of ventilation and perfusion, followed by 75 min of reperfusion and ventilation to establish the model of lung I/R injury in I/R and DEX groups.In DEX group, dexmedetomidine 10 nmol/L was added to K-H solution at the beginning of reperfusion.Lung tissues were obtained for determination of wet/dry weight ratio (W/D ratio) and for examination of pathological changes (with a light microscope) and ultrastructure (using an electron microscope), and the alveolar damage rate (IAR) was calculated.The expression of pyroptosis-related factors including NOD-like receptor family protein 3 (NLRP3), caspase-1, interleukin-1β (IL-1β), IL-18 and gasdermin D (GSDMD) protein and mRNA was detected by Western blot or by real-time polymerase chain reaction. Results Compared with C group, the W/D ratio and IAR in lung tissues were significantly increased, and the expression of NLRP3, caspase-1, IL-1β, IL-18 and GSDMD protein and mRNA was up-regulated in I/R and DEX groups (P<0.05). Compared with I/R group, the W/D ratio and IAR in lung tissues were significantly decreased, the expression of NLRP3, caspase-1, IL-1β, IL-18 and GSDMD protein and mRNA was down-regulated (P<0.05), and the pathological changes were significantly attenuated in DEX group. Conclusion The mechanism by which dexmedetomidine reduces isolated rat lung I/R injury may be related to inhibiting pyroptosis. Key words: Dexmedetomidine; Reperfusion injury; Lung; Pyroptosis

Histopathological changes of brucellosis in experimentally infected guinea pig

Background: Brucellosis, a chronic infectious and zoonotic disease, is endemic in many countries of the world including Bangladesh, that affects humans and animals, leading to significant impact on public health and animal industry. There are several reports on seroprevalence, risk factors, molecular, epidemiological and review of brucellosis in human and animals but least of histopathological reports in Bangladesh. As laboratory animal species guinea pigs are the most susceptible to Brucella infection in comparison to mice, monkeys, rats and sheep. Hence, this study was undertaken to determine the histopathology of brucellosis in guinea pig. Methods: After collection of aborted bovine foetal membranes from the Central Cattle Breeding and Dairy Farm (CCBDF), Savar, screened with modified Ziehl-Neelsen staining method and inoculum (10 CFU/2ml) prepared from positive samples were inoculated into each of ten guinea pig. The guinea pigs were sacrificed after showing positive results from the serum in Rose Bengal test and from blood in rapid kit test 3 weeks post inoculation. Then the gross and histopathological lesions were observed in the liver, spleen, heart, lungs, and kidneys. Results: Liver showed congestion, haemorrhage and fatty changes and granuloma formation with infiltration of macrophages. However, only caseous necrosis of splenic follicle was observed in spleen. The major findings in lungs of guinea pig were granuloma, haemorrhage and necrosis of lung parenchyma and mononuclear cellular infiltration. The heart revealed haemorrhagic endocarditis and monocytes infiltration. A variable degree of multifocal accumulation of mononuclear cells infiltration and congestion were found in kidneys. Conclusions: It is concluded that the histopathological changes caused by Brucella spp. are similar to those observed in humans with brucellosis. The animal models, particularly the guinea pig, can be used to study the detail pathogenesis of this organism.

DT-0111: a novel drug-candidate for the treatment of COPD and chronic cough.

Background:Extracellular adenosine 5′-triphosphate (ATP) plays important mechanistic roles in pulmonary disorders in general and chronic obstructive pulmonary disease (COPD) and cough in particular. The effects of ATP in the lungs are mediated to a large extent by P2X2/3 receptors (P2X2/3R) localized on vagal sensory nerve terminals (both C and Aδ fibers). The activation of these receptors by ATP triggers a pulmonary-pulmonary central reflex, which results in bronchoconstriction and cough, and is also proinflammatory due to the release of neuropeptides from these nerve terminals via the axon reflex. These actions of ATP in the lungs constitute a strong rationale for the development of a new class of drugs targeting P2X2/3R. DT-0111 is a novel, small, water-soluble molecule that acts as an antagonist at P2X2/3R sites.Methods:Experiments using receptor-binding functional assays, rat nodose ganglionic cells, perfused innervated guinea pig lung preparation ex vivo, and anesthetized and conscious guinea pigs in vivo were performed.Results:DT-0111 acted as a selective and effective antagonist at P2X2/3R, that is, it did not activate or block P2YR; markedly inhibited the activation by ATP of nodose pulmonary vagal afferents in vitro; and, given as an aerosol, inhibited aerosolized ATP-induced bronchoconstriction and cough in vivo.Conclusions:These results indicate that DT-0111 is an attractive drug-candidate for the treatment of COPD and chronic cough, both of which still constitute major unmet clinical needs. The reviews of this paper are available via the supplementary material section.

PPE11 of Mycobacterium tuberculosis can alter host inflammatory response and trigger cell death

Tuberculosis (TB), which is caused by Mycobacterium tuberculosis (Mtb), remains a serious global health problem. The PE/PPE family, featuring unique sequences, structures and expression in Mtb, is reported to interfere with the macrophage response to the pathogen and facilitate its infection. PPE11 (Rv0453) existed in pathogenic mycobacteria and was persistently expressed in the infected guinea pig lungs. However, the role it played in the pathogenesis remains unclear. Here, to investigate the interaction and potential mechanism of PPE11 between pathogens and hosts, we heterologously expressed PPE11 in non-pathogenic, rapidly growing Mycobacterium smegmatis strains. We found that the overexpression of the cell wall-associated protein, PPE11, can improve the viability of bacteria in the presence of lysozyme, hydrogen peroxide and acid stress. Expression of PPE11 enhanced the early survival of M. smegmatis in macrophages and sustained a higher bacterial load in mouse tissues that showed exacerbated organ pathology. Macrophages infected with recombinant M. smegmatis produced significantly greater amounts of interleukin (IL)-1β, IL-6, tumour necrosis factor (TNF)-α and an early decrease in IL-10 along with higher levels of host cell death. Similar cytokines changes were observed in the sera of infected mice. Accordingly, PPE11 protein causes histopathological changes by disrupting the dynamic balance of the inflammatory factors and promoting host-cell death, indicating a potential role in the virulence of Mtb.

The ethanol extract of Involcucrum castaneae ameliorated ovalbumin-induced airway inflammation and smooth muscle thickening in guinea pigs

Ethnopharmacological relevanceInvolucrum castaneae(IC)is used in Chinese folk medicine to treat various lung diseases, as well as for its reducing phlegm and anti-inflammatory properties. Aim of the studyThe purpose of this experiment is to verify the effect of IC on airway inflammation, responsiveness in ovalbumin (OVA)-induced asthmatic guinea pigs. The main chemical components of IC were also analyzed. Materials and methodsThe potential of the ethanol extract of Involucrum castaneae (EEIC) to protect against OVA-induced allergic airway response in guinea pigs was investigated. The latency of asthma in guinea pigs were recorded after the allergic asthma induced. Enzyme-linked immunosorbent assay (ELISA) was used to measure the levels of immunoglobulin E (IgE), interleukin-5 (IL-5), nerve growth factor (NGF) and interferon-γ (IFN-γ) in asthma allergy. Reverse transcription-PCR (RT-PCR) was used to detect the expression of IL-5 mRNA in asthmatic guinea pig lungs. Paraffin sections of lung tissue were used to analyze pathological changes. The total flavonoid content was determined and the chemical components were analyzed by LC-MS/MS. ResultsIt was found that EEIC was able to reduce the number of eosinophil (EOS) in bronchoalveolar lavage fluid (BALF) and peripheral blood (PB) in the guinea pig model of OVA -induced asthma. Meanwhile, it also significantly reduced the levels of inflammation-related factors IgE and IL-5, decreased the expression of IL-5 mRNA in lung tissue, and increased the level of IFN-γ. Pathological examination of paraffin section of lung tissue showed that EEIC can reduce the thickening of bronchial smooth muscle and reduce the infiltration damage of tissues by various inflammatory cells. The presence of flavonoids, terpenoids and phenolic compounds in EEIC might be responsible for these activities. ConclusionIC alleviated airway inflammation and smooth muscle thickening in guinea pigs with OVA-sensitized allergic asthma. The paper explains the traditional efficacy and material basis of IC and lays a foundation for further development.