Inflammatory, anti-inflammatory and regulatory cytokines in relatively healthy lung tissue as an essential part of the local immune system

Publisher Copyright: © 2017, PALACKY UNIV. All rights reserved.Background. The innate and adaptive immune systems in lungs are maintained not only by immune cells but also by non-immune tissue structures, locally providing wide intercellular communication networks and regulating the local tissue immune response. Aims. The aim of this study was to determine the appearance and distribution of inflammatory, anti-inflammatory and regulatory cytokines in relatively healthy lung tissue samples. Material and Methods. We evaluated lung tissue specimens obtained from 49 patients aged 9–95 years in relatively healthy study subjects. Tissue samples were examined by hematoxylin and eosin staining. Interleukin-1 (IL-1), interleu-kin-4 (IL-4), interleukin-6 (IL-6), interleukin-7 (IL-7), and interleukin-10 (IL-10) were detected by an immunohistochemistry (IMH) method. The number of positive structures was counted semiquantitatively by microscopy. Non-parametric tests were used to analyse the data. Results. IL-1-positive cells were mostly found in the bronchial cartilage and alveolar epithelium. Immunoreactive lung macrophages were also found. The numbers of IL-4, IL-6, IL-7, and IL-10 containing cells were also found in the bronchial epithelium (in addition to those previously listed). The number of positive structures varied from occasional to moderate, but was graded higher in cartilage. Overall, fewer IL-1-positive cells and more IL-10-positive cells were found. Almost no positive structures for all examined cytokines were found in connective tissue and bronchial glands. Conclusions. Relatively healthy lung tissue exhibits anti-inflammatory response patterns. The cytokine distribution and appearance suggest persistent stimulation of cytokine expression in lung tissue and indicate the presence of local regulatory and modulating patterns. The pronounced cytokine distribution in bronchial cartilage suggests the involvement of a compensatory local immune response in the supporting tissue.publishersversionPeer reviewe

An insight into COPD morphopathogenesis : Chronic inflammation, remodeling, and antimicrobial defense

Publisher Copyright: © 2019 by the authors. Licensee MDPI, Basel, Switzerland.Background and Objectives: Intercellular signaling networks with high complexity cause a spectrum of mechanisms achieving chronic obstructive pulmonary disease (COPD) that still question many uncertainties. Materials and Methods: Immunoreactive cells in bronchial tissue obtained from 40 COPD patients and 49 healthy control subjects were detected by biotin-streptavidin immunohistochemistry method for the following markers of IL-1α, IL-4, IL-6, IL-7, IL-8, IL-10, IL-12, TNF-α, MMP-2, TIMP-2, TGF-β1, Hsp-70, hBD-2, hBD-3, hBD-4. Results: Overall the highest numbers (from mostly moderate (++) to abundance (++++)) of IL-1α, IL-4, IL-7, IL-8, IL-10, IL-12, MMP-2, TIMP-2, TGF-β1 immunoreactive cells were marked increasingly in the blood vessel wall, connective tissue, and bronchial epithelium of COPD-affected lung, respectively. We found statistically significant (p < 0.05) higher numbers of immunoreactive cells positive for all of examined interleukins, TNF-α, MMP-2, TIMP-2, TGF-β1, hBD-2, and hBD-3 in the COPD-affected lung compared to the control group, but not for Hsp-70 and hBD-4. Conclusions: COPD-affected lung tissue exhibits mostly inflammatory response patterns of increased IL-1α, IL-4, IL-8, IL-12, and TNF-α, especially in the airway epithelium. Increased MMP-2 and TGF-β1, but decreased Hsp-70, proposes pronounced tissue damage and remodeling in COPD. High numbers of hBD-2 and hBD-3 immunoreactive cells may highlight antimicrobial activity in COPD within stable regulation of local immunity.publishersversionPeer reviewe

COPD affected lung tissue remodeling due to the local distribution of MMP-2, TIMP-2, TGF-β1 and HSP-70

Chronic obstructive pulmonary disease (COPD) is strongly associated with progressive airway limitation where the key mechanism is abnormal airway remodelling of bronchial mucosa maintained by complex crosstalk of tissue remodelling and regulatory factors. The aim of this study was to determine the appearance and local distribution of tissue remodelling factors in COPD affected lung tissue and to compare the findings with the control group. In this study, lung tissue specimens were obtained from 27 patients with COPD and 49 control patients. Tissue samples were examined by routine hematoxylin and eosin staining. Remodelling and regulatory factors matrix metalloproteinase-2 (MMP-2), tissue inhibitor of metalloproteinase-2 (TIMP-2), transforming growth factor-β1 (TGF-β1), as well as heat shock protein-70 (Hsp-70) were detected by immunohistochemistry in airway mucosa. The numbers of positive structures were evaluated semiquantitatively. Non-parametrical statistical analysis was performed. Overall COPD-affected lung tissue presented chronic inflammation and tissue remodelling in routine histological analysis. Compared to the control group, statistically significant (P<0.05) difference was calculated between COPD affected lung tissue and control group with overall more immunoreactive cells containing MMP-2, TIMP-2 and TGF-β1 and less Hsp-70 immunoreactive bronchial epithelial cells, subepithelial connective tissue fibroblasts, bronchial smooth muscle cells and secretory cells of bronchial glands with more pronounced findings of TGF-β1, however, less Hsp-70. Study findings suggest pronounced tissue damage and remodelling with the local regulatory environment of up-regulated TGF-β1. Increased numbers of MMP-2, TIMP-2 and TGF-β1 immunoreactive cells suggest the key role of these remodelling factors in COPD pathogenesis. Decrease of Hsp-70 proves increased cell damage in COPD-affected airway mucosa.publishersversionPeer reviewe

Expression Analysis of FGF/FGFR and FOX Family Proteins in Mucosal Tissue Obtained from Orofacial Cleft-Affected Children

Funding Information: Funding: The present study was funded by Riga Stradinš University (RSU), Project Nb. 5-1/384/2020 (dated 10 September 2020). Publisher Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland.Orofacial clefts affect hundreds of thousands of children worldwide annually and are usually corrected by a series of surgeries extending to childhood. The underlying mechanisms that lead to clefts are still unknown, mainly because of the multifactorial etiology and the myriad of interactions between genes and environmental factors. In the present study, we investigated the role and expression of candidate genes belonging to the FGF/FGFR signaling pathway and FOX family in tissue material obtained from 12 pediatric patients undergoing cleft correction surgery. The expression was investigated using immunohistochemistry (IHC) and chromogenic in-situ hybridization (CISH) in three cell/tissue types—epithelial cells, connective tissue, and endothelial cells. We found elevated expression of FGFR1 in epithelial cells while no expression was observed in endothelial cells. Further, our results elucidate the potential pathogenetic role of FGFR1 in cellular proliferation, local site inflammation, and fibrosis in cleft patients. Along with bFGF (also called FGF2), FGFR1 could play a pro-inflammatory role in clefts. Over-amplification of FGFR2 in some patients, along with bFGF, could potentially suggest roles for these genes in angiogenesis. Additionally, increased expression of FOXE1 (also called TTF2) contributes to local site inflammation. Finally, zero to low amplification of FOXO1 could suggest its potential role in inducing oxidative stress in the endothelium along with reduced epithelial apoptosis.publishersversionPeer reviewe

Cytokines and Defensins in Tissue Biopsies Obtained by Bronchoscopy from Patients with Post-Intubation Tracheal Stenosis

Aims: The objective of our study was to perform the routine analysis of bronchoscopically obtained tracheal samples to determine the appearance and relative distribution of cytokines and antimicrobial proteins in patients with post-intubation tracheal stenosis (PITS). Study Design: Retrospective. Place and Duration of Study: Rīga Stradiņš University, Institute of Anatomy and Anthropology, Pauls Stradiņš Clinical University Hospital, between May 2014 and May 2015. Methodology: Five patients with PITS were involved in this study. Tissue samples were obtained by bronchoscopy from the upper part of trachea, then proceeded for routine histological staining with hematoxylin and eosin. Interleukine-1 (IL-1), interleukine-10 (IL-10) and tumor necrosis factor alpha (TNFα), as well as beta defensin-2 (β def-2) were detected by use of immunohistochemistry (IMH) method. The number of immunoreactive (positive) structures was graded semi-quantitatively. Results: Squamous metaplasia, inflammatory cell infiltration and formation of granulation tissue were observed in all cases. Significant expression of IL-10 and β def-2 was seen as various number of immunoreactive structures in tracheal tissue. Only few scattered IL-1 and TNFα positive macrophages were found in part of cases. Conclusions: The leading role in pathogenesis of post-intubation tracheal stenosis is assumed to be the chronic inflammation, fibrous scarring, as well as the remodeling of tracheal wall due to the ischemia. Compensatory expression of antimicrobial peptide β def-2 and anti-inflammatory cytokine IL-10 indicates the intense local tissue defense reactions. TNFα and IL-1 are not among the most significant factors in pathogenesis of PITS.authorsversionPeer reviewe

Local Defence System in Healthy Lungs

Background: The respiratory system is one of the main entrance gates for infection. The aim of this work was to compare the appearance of specific mucosal pro-inflammatory and common anti-microbial defence factors in healthy lung tissue, from an ontogenetic point of view. Materials and methods: Healthy lung tissues were collected from 15 patients (three females and 12 males) in the age range from 18 to 86. Immunohistochemistry to human β defensin 2 (HBD-2), human β defensin 3 (HBD-3), human β defensin 4 (HBD-4), cathelicidine (LL-37) and interleukine 17A (IL-17A) were performed. Results: The lung tissue material contained bronchial and lung parenchyma material in which no histological changes, connected with the inflammatory process, were detected. During the study, various statistically significant differences were detected in immunoreactive expression between different factors in all lung tissue structures. Conclusion: All healthy lung structures, but especially the cartilage, alveolar epithelium and the alveolar macrophages, are the main locations for the baseline synthesis of antimicrobial proteins and IL-17A. Cartilage shows high functional plasticity of this structure, including significant antimicrobial activity and participation in local lung protection response. Interrelated changes between antimicrobial proteins in different tissue confirm baseline synergistical cooperation of all these factors in healthy lung host defence.publishersversionPeer reviewe

Expression of anti-inflammatory markers IL-2, IL-10, TGF-β1, βDEF-2, βDEF-3 and Cathelicidin LL37 in dairy cattle milk with different health status of the udder

Funding Information: The project is implemented under ERA-NET ICRAD (International Coordination of Research on Infectious Diseases) and co-financing by the Spanish State Research Agency (AEI/10.13039/501100011033; PCI2020-120693-2), Hungarian National Food Chain Safety Office, Polish National Centre for Research and Development, and Latvian State Education Development Agency. This project has received funding from the European Union’s Horizon 2020 research and innovation program under grant agreement no. 862605. Funding Information: We would like to express our gratitude to the farm owners, Mirosław and Janina Gontar, who take care of the animals and help to collect milk samples for performing our research, and as well as to our Laboratory Assistant Natālija Moroza, who prepared and processed milk smear samples for further analysis. The project is implemented under ERA-NET ICRAD (International Coordination of Research on Infectious Diseases) and co-financing by the Spanish State Research Agency (AEI/10.13039/501100011033; PCI2020-120693-2), Hungarian National Food Chain Safety Office, Polish National Centre for Research and Development, and Latvian State Education Development Agency. This project has received funding from the European Union's Horizon 2020 research and innovation program under grant agreement no. 862605. Publisher Copyright: © 2022 Polish Academy of Sciences. All rights reserved.Great economic losses to the dairy industry are associated with bovine mastitis, which results in poor milk quality and high treatment costs. Anti-inflammatory proteins play an important role in the suppression of the immune response against invading pathogenic microorganisms and are therefore being studied for possible use in the early diagnosis of mastitis. In our study, we used milk samples from 15 cows of Holstein Friesian breed with different health status (5 healthy, 5 subclinical, and 5 clinical animals), and tested them using immunohistochemical (IHC) analysis to evaluate the presence of IL-2, IL-10, TGF-β1, βDEF-2, DEF-3, and Cathelicidin LL37 proteins. The calculation of positively and negatively stained cells for each biomarker was performed using the semiquantitative counting method. We found the presence of all factors with the exception of Cathelicidin LL37, which was almost absent in milk samples of all animal groups. The significant decrease of IL-10, β-def2, and β-def3 expression levels within the 3 days of sampling, found in the milk of animals with sub- and clinical mastitis, indicates the loss of antiinflammatory protection of the affected cow’s udder. In contrast, the stable increase of IL-2 and TGF-β1 positive cells observed in the milk of mastitis-affected cows, and the similar expression of these factors in the milk of healthy animals, indicate the possible lack of involvement of these cytokines at an early stage of udder inflammation.publishersversionPeer reviewe

Identification of Inflammatory and Regulatory Cytokines IL-1α-, IL-4-, IL-6-, IL-12-, IL-13-, IL-17A-, TNF-α-, and IFN-γ-Producing Cells in the Milk of Dairy Cows with Subclinical and Clinical Mastitis

Funding Information: The project is implemented under ERA-NET ICRAD (International Coordination of Research on Infectious Diseases) and co-financing of Spanish State Research Agency, Hungarian National Food Chain Safety Office, Polish National Centre for Research and Development, and Latvian State Education Development Agency. This project has received funding from the European Union?s Horizon 2020 research and innovation program under grant agreement no. 862605. Funding Information: Funding: The project is implemented under ERA-NET ICRAD (International Coordination of Research on Infectious Diseases) and co-financing of Spanish State Research Agency, Hungarian National Food Chain Safety Office, Polish National Centre for Research and Development, and Latvian State Education Development Agency. This project has received funding from the European Union’s Horizon 2020 research and innovation program under grant agreement no. 862605. Publisher Copyright: © 2022 by the authors. Licensee MDPI, Basel, Switzerland.In naturally occurring bovine mastitis, effects of infection depend on the host inflammatory response, including the effects of secreted cytokines. Knowledge about the inflammatory and regulatory cytokines in milk cells of free-stall barn dairy cows and in naturally occurring mastitis is lacking as most studies focus on induced mastitis. Hereby, the aim of the study was to determine inflammatory and regulatory cytokines in the milk of dairy cows with subclinical and clinical mastitis. The following examinations of milk samples were performed: differential counting of somatic cells (SCC), bacteriological examination, and immunocytochemical analysis. Mean SCC increased in subclinical and clinical mastitis cases. The number of pathogenic mastitis-causing bacteria on plates increased in subclinical mastitis cases but decreased in clinical mastitis. The inflammatory and regulatory markers in the milk cells of healthy cows showed the highest mean cell numbers (%). In mastitis cases, immunoreactivity was more pronounced for IL-4, IL-6, IL-12, IL-13, IL-17A, TNF-, and IFN-y. Data about subclinical and clinical mastitis demonstrate inflammatory responses to intramammary infection driven by IL-1, IL-4, and IL-17A. Moreover, the host defense response in mastitis is characterized by continuation or resolution of initial inflammation. IL-12 and INF-immunoreactivity was recognized to differ mastitis cases from the relative health status.publishersversionPeer reviewe

Hroniskas obstruktīvas plaušu slimības (HOPS) skartu audu morfoloģisks raksturojums. Promocijas darba kopsavilkums

The Doctoral Thesis was developed at the Department of Morphology, Institute of Anatomy and Anthropology, Rīga Stradiņš University, Latvia. Defence: at the public session of the Promotion Council of the Basic Sciences of Medicine, including Pharmacology on 28 December 2020 at 10.00 in Hippocrates Lecture Theatre, 16 Dzirciema Street, Rīga Stradiņš University and using the Zoom platform.Chronic obstructive pulmonary disease (COPD) is a potentially preventable and treatable disease characterized by progressive airway disorders. Under the influence of various risk factors, oxidative stress and chronic inflammation develop, which determines the formation and course of practically all other processes (for example, local fibrosis). Even with the maximal elimination of risk factors, inflammation in COPD persists and even continues. In general, COPD morphopathogenesis is characterized by oxidative stress, chronic inflammation, remodeling, as well as impaired antimicrobial and other locally realized tissue protective responses. The aim of the study was to determine and evaluate the morphological characteristics of bronchial tissues affected by COPD and analyze the tissue distribution of markers relevant to chronic inflammation, changes in tissue remodeling and local protective mechanisms. Also, these markers were evaluated in ontogenesis perspective. The study included 40 COPD and 49 relatively healthy control patients. Tissue material from COPD patients was collected during bronchoscopy. Tissue material from the control group was obtained by autopsy from individuals of different ages who died of accidents and from diseases not affecting respiratory system, as well as from / or during lung surgery. IL-1α (interleukin-1 alpha), IL-4 (interleukin-4), IL-6 (interleukin-6), IL-7 (interleukin-7), IL-8 (interleukin-8), IL-10 (interleukin-10), IL-12 (interleukin-12), TNF-α (tumour necrosis factor-alpha), TGF-β1 (transforming growth factor-beta 1), MMP-2 (matrix metalloproteinase-2), TIMP-2 (matrix metalloproteinase-2 tissue inhibitor), Hsp-70 (heat shock protein-70), hBD-2 (human beta defensin-2), hBD-3 (human beta defensin-3), hBD-4 ( human beta defensin-4) immunoreactive cell counts were determined by immunohistochemistry in eight different tissue groups and lung localizations in control and COPD patients, and also compared. The immunohistochemical findings were also analyzed in terms of ontogenesis or aging. In the lung tissues of the control group, we found a mild to moderate increase in the number of immunoreactive cells of all studied factors at different localizations, indicating low, persistent, continuous and adaptive “baseline” level for various factors. Lung tissue in COPD patients is characterized by variable and various localized changes with chronic inflammation and tissue remodeling including inflammatory cell infiltration, granulation tissue, thickened basement membrane, bronchial gland and smooth muscle hypeplasia and hypertrophy, also thickened vascular fibrosis, moreover, fibrosis and epithelial metaplasia. The most pronounced finding of the studied tissue factors was in bronchial epithelium, mucosal connective tissue and blood vessels of COPD patients, which proves the significant involvement and role of these structures in COPD morphopathogenesis. Overall, COPD morphopathogenesis is characterized by increased IL-1α, IL-4, IL-6, IL-8, TNF-α, IL-7, IL-10, IL-12, TGF-β1, MMP-2, TIMP-2, hBD-2 and hBD-3 immunoreactive cell counts, as well as reduced Hsp-70 and hBD-4-containing cell counts, indicating high activity of persistent complex cytokine pleiade, tissue remodeling and total tissue antimicrobial protection. Ontogenesis dependent changes, including in terms of aging, within the lungs of relatively healthy individuals are associated with mild inflammation, aging of immune system cells and disorder of immune system regulation. Age changes in the relatively healthy lungs are individual, even different for individuals in different age groups, as well as mostly characterized by a lack to maintain inflammation, also, reduced number of immune cells are found. In elderly COPD patients within ontogenesis, we detected an increase in the numbers of IL-1α, IL-4, IL-6, IL-10, IL-12, MMP-2, hBD-4 immunoreactive cells, but decreased numbers of IL-7, IL-8, TNF-α, TIMP-2, TGF-β1, Hsp-70, hBD-2, hBD-3 immunoreactive cells with age, as well as chronic bronchitis, epithelial metaplasia, granulation tissue and poorer functional parameters. In the case of COPD, aging is associated with the worsening of the disease, as well as persistence and altered remodeling of inflammatory cytokines otherwise characteristic of COPD