Respiratory infections are a real threat for humans, and therefore the pig model is of interest for studies. As one of a case for studies, Actinobacillus pleuropneumoniae (APP) caused infections and still worries many pig breeders around the world. To better understand the influence of pathogenic effect of APP on a respiratory system-lungs and tracheobronchial lymph nodes (TBLN), we aimed to employ matrix-assisted laser desorption/ionization time-of-flight mass spectrometry imaging (MALDI-TOF MSI). In this study, six pigs were intranasally infected by APP and two were used as non-infected control, and 48 cryosections have been obtained. MALDI-TOF MSI and immunohistochemistry (IHC) were used to study spatial distribution of infectious markers, especially interleukins, in cryosections of porcine tissues of lungs (necrotic area, marginal zone) and tracheobronchial lymph nodes (TBLN) from pigs infected by APP. CD163, interleukin 1β (IL‑1β) and a protegrin-4 precursor were successfully detected based on their tryptic fragments. CD163 and IL‑1β were confirmed also by IHC. The protegrin-4 precursor was identified by MALDI-TOF/TOF directly on the tissue cryosections. CD163, IL‑1β and protegrin‑4 precursor were all significantly (p < 0.001) more expressed in necrotic areas of lungs infected by APP than in marginal zone, TBLN and in control lungs.

• Keywords
• Actinobacillus pleuropneumoniae, CD163, MALDI MSI, interleukin 1β, lungs infection, pig model, protegrin‑4 precursor,
• MeSH
• Actinobacillus pleuropneumoniae pathogenicity   MeSH
• CD163 Antigen MeSH
• Antigens, Differentiation, Myelomonocytic metabolism   MeSH
• Biomarkers metabolism   MeSH
• Bronchi metabolism   MeSH
• Antigens, CD metabolism   MeSH
• Actinobacillus Infections metabolism   microbiology   MeSH
• Respiratory Tract Infections metabolism   microbiology   MeSH
• Interleukin-1beta metabolism   MeSH
• Antimicrobial Cationic Peptides metabolism   MeSH
• Lymph Nodes metabolism   MeSH
• Lung metabolism   MeSH
• Swine MeSH
• Receptors, Cell Surface metabolism   MeSH
• Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization methods   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Names of Substances
• CD163 Antigen MeSH
• Antigens, Differentiation, Myelomonocytic MeSH
• Biomarkers MeSH
• Antigens, CD MeSH
• Interleukin-1beta MeSH
• Antimicrobial Cationic Peptides MeSH
• protegrin-4 MeSH Browser
• Receptors, Cell Surface MeSH

These guidelines are a consensus work of a considerable number of members of the immunology and flow cytometry community. They provide the theory and key practical aspects of flow cytometry enabling immunologists to avoid the common errors that often undermine immunological data. Notably, there are comprehensive sections of all major immune cell types with helpful Tables detailing phenotypes in murine and human cells. The latest flow cytometry techniques and applications are also described, featuring examples of the data that can be generated and, importantly, how the data can be analysed. Furthermore, there are sections detailing tips, tricks and pitfalls to avoid, all written and peer-reviewed by leading experts in the field, making this an essential research companion.

• MeSH
• Allergy and Immunology standards   MeSH
• Phenotype MeSH
• Consensus MeSH
• Humans MeSH
• Flow Cytometry methods   standards   MeSH
• Cell Separation methods   standards   MeSH
• Check Tag
• Humans MeSH
• Publication type
• Journal Article MeSH

Monocytes play an essential role in the defense against bacterial pathogens. Bone marrow (BM) and peripheral blood (PB) monocytes in pigs consist of the main "steady-state" subpopulations: CD14 hi/CD163-/SLA-DR- and CD14 low/CD163+/SLA-DR+. During inflammation, the subpopulation of "inflammatory" monocytes expressing very high levels of CD163, but lacking the SLA-DR molecule (being CD14 low/CD163+/SLA-DR-) appears in the BM and PB and replaces the CD14 low/CD163+/SLA-DR+ subpopulation. However, current knowledge of monocyte migration into inflamed tissues in pigs is limited. The aim of the present study was to evaluate the distribution of "inflammatory" CD14 low/CD163+/SLA-DR- monocytes during experimental inflammation induced by Actinobacillus pleuropneumoniae (APP) and a possible role for chemokines in attracting "inflammatory" CD14 low/CD163+/SLA-DR- monocytes into the tissues. Monocyte subpopulations were detected by flow cytometry. Chemokines and chemokine receptors were detected by RT-qPCR. The "steady-state" monocytes were found in the BM, PB, spleen and lungs of control pigs. After APP-infection, "inflammatory" monocytes replaced the "steady-state" subpopulation in BM, PB, spleen and moreover, they appeared in an unaffected area, demarcation zone and necrotic area of the lungs and in tracheobronchial lymph nodes. They did not appear in mesenteric lymph nodes. Levels of mRNA for various chemokines with their appropriate receptors were found to be elevated in BM (CCL3-CCR1/CCR5, CCL8-CCR2/CCR5, CCL19-CCR7), necrotic area of the lungs (CCL3-CCR1, CCL5-CCR1/CCR3, CCL11-CCR3, CCL22/CCR4) and tracheobronchial lymph nodes (CCL3-CCR1) and therefore they could play a role in attracting monocytes into inflamed tissues. In conclusion, "inflammatory" monocytes appear in different lymphoid tissues and the lungs after APP infection in pigs. Various chemokines could drive this process.

• MeSH
• Actinobacillus pleuropneumoniae physiology   MeSH
• CD163 Antigen MeSH
• Antigens, Differentiation, Myelomonocytic metabolism   MeSH
• Antigens, CD metabolism   MeSH
• Chemokines genetics   metabolism   MeSH
• Actinobacillus Infections immunology   microbiology   veterinary   MeSH
• Lymphoid Tissue metabolism   MeSH
• RNA, Messenger genetics   MeSH
• Monocytes cytology   metabolism   MeSH
• Swine Diseases immunology   microbiology   MeSH
• Lung metabolism   MeSH
• Swine MeSH
• Flow Cytometry veterinary   MeSH
• Receptors, Cell Surface metabolism   MeSH
• Receptors, Chemokine genetics   metabolism   MeSH
• Inflammation microbiology   MeSH
• Animals MeSH
• Check Tag
• Animals MeSH
• Publication type
• Journal Article MeSH
• Research Support, Non-U.S. Gov't MeSH
• Names of Substances
• CD163 Antigen MeSH
• Antigens, Differentiation, Myelomonocytic MeSH
• Antigens, CD MeSH
• Chemokines MeSH
• RNA, Messenger MeSH
• Receptors, Cell Surface MeSH
• Receptors, Chemokine MeSH