TLR-Mediated Cytokine Gene Expression in Chicken Peripheral Blood Mononuclear Cells as a Measure to Characterize Immunobiotics
Immunobiotics are probiotics that promote intestinal health by modulating immune responses. Immunobiotics are recognized by Toll-like receptors (TLRs) and activate cytokine gene expression. This study aimed to characterize cytokine gene expression in the chicken peripheral blood mononuclear cells (P...
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MDPI AG
2021-01-01
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| Online Access: | https://www.mdpi.com/2073-4425/12/2/195 |
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| author | Anna Slawinska Aleksandra Dunislawska Arkadiusz Plowiec José Gonçalves Maria Siwek |
| author_facet | Anna Slawinska Aleksandra Dunislawska Arkadiusz Plowiec José Gonçalves Maria Siwek |
| author_sort | Anna Slawinska |
| collection | DOAJ |
| description | Immunobiotics are probiotics that promote intestinal health by modulating immune responses. Immunobiotics are recognized by Toll-like receptors (TLRs) and activate cytokine gene expression. This study aimed to characterize cytokine gene expression in the chicken peripheral blood mononuclear cells (PBMC) stimulated with purified TLR ligands and live probiotics. PBMC were isolated from the whole blood. PBMC were stimulated with: lipopolysaccharide (LPS), CpG ODN, Pam3CSK4, Zymosan, galactooligosaccharides (GOS), <i>Lactococcus</i><i>lactis</i> subsp. <i>cremoris</i> (<i>L. lactis</i>), and <i>Saccharomyces cerevisiae</i> at 42.5 °C and 5% CO<sub>2</sub> for 3 h, 6 h, and 9 h. After each time-point, PBMC were harvested for RNA isolation. Relative gene expression was analyzed with RT-qPCR for cytokine genes (<i>IL-1β</i>, <i>IL-2</i>, <i>IL-3</i>, <i>IL-4</i>, <i>IL-6</i>, <i>IL-8</i>, <i>IL-10</i>, <i>IL-12p40</i>, and <i>IFN-ɣ</i>) and reference genes (<i>ACTB</i> and <i>G6PDH</i>). Genes were clustered into pro-inflammatory genes, Th1/Th2 genes, and Th1-regulators. The gene expression differed between treatments in <i>IL1-β</i>, <i>IL-6</i>, <i>IL-8</i>, <i>IL-10</i>, and <i>IL-12p40</i> (<i>p</i> < 0.001). The genes <i>IL-1β</i>, <i>IL-6</i>, and <i>IL-8</i> had the highest fold change of mRNA expression at 3 h in response to TLR ligands. <i>L. lactis</i> up-regulated the pro-inflammatory genes at the 6 h time-point. <i>L. lactis</i> did not activate the anti-inflammatory <i>IL-10</i> gene, but activated <i>IL-12p40</i> at 6 h. Hereby, <i>L. lactis</i> was proven to exert immunostimulatory properties in PBMC. |
| first_indexed | 2024-03-09T03:21:57Z |
| format | Article |
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| language | English |
| last_indexed | 2024-03-09T03:21:57Z |
| publishDate | 2021-01-01 |
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| spelling | doaj.art-dcfcebe7281f496daee0697e2d269f7d2023-12-03T15:09:44ZengMDPI AGGenes2073-44252021-01-0112219510.3390/genes12020195TLR-Mediated Cytokine Gene Expression in Chicken Peripheral Blood Mononuclear Cells as a Measure to Characterize ImmunobioticsAnna Slawinska0Aleksandra Dunislawska1Arkadiusz Plowiec2José Gonçalves3Maria Siwek4Department of Animal Biotechnology and Genetics, UTP University of Science and Technology, 85-084 Bydgoszcz, PolandDepartment of Animal Biotechnology and Genetics, UTP University of Science and Technology, 85-084 Bydgoszcz, PolandDepartment of Animal Biotechnology and Genetics, UTP University of Science and Technology, 85-084 Bydgoszcz, PolandDepartment of Animal Biotechnology and Genetics, UTP University of Science and Technology, 85-084 Bydgoszcz, PolandDepartment of Animal Biotechnology and Genetics, UTP University of Science and Technology, 85-084 Bydgoszcz, PolandImmunobiotics are probiotics that promote intestinal health by modulating immune responses. Immunobiotics are recognized by Toll-like receptors (TLRs) and activate cytokine gene expression. This study aimed to characterize cytokine gene expression in the chicken peripheral blood mononuclear cells (PBMC) stimulated with purified TLR ligands and live probiotics. PBMC were isolated from the whole blood. PBMC were stimulated with: lipopolysaccharide (LPS), CpG ODN, Pam3CSK4, Zymosan, galactooligosaccharides (GOS), <i>Lactococcus</i><i>lactis</i> subsp. <i>cremoris</i> (<i>L. lactis</i>), and <i>Saccharomyces cerevisiae</i> at 42.5 °C and 5% CO<sub>2</sub> for 3 h, 6 h, and 9 h. After each time-point, PBMC were harvested for RNA isolation. Relative gene expression was analyzed with RT-qPCR for cytokine genes (<i>IL-1β</i>, <i>IL-2</i>, <i>IL-3</i>, <i>IL-4</i>, <i>IL-6</i>, <i>IL-8</i>, <i>IL-10</i>, <i>IL-12p40</i>, and <i>IFN-ɣ</i>) and reference genes (<i>ACTB</i> and <i>G6PDH</i>). Genes were clustered into pro-inflammatory genes, Th1/Th2 genes, and Th1-regulators. The gene expression differed between treatments in <i>IL1-β</i>, <i>IL-6</i>, <i>IL-8</i>, <i>IL-10</i>, and <i>IL-12p40</i> (<i>p</i> < 0.001). The genes <i>IL-1β</i>, <i>IL-6</i>, and <i>IL-8</i> had the highest fold change of mRNA expression at 3 h in response to TLR ligands. <i>L. lactis</i> up-regulated the pro-inflammatory genes at the 6 h time-point. <i>L. lactis</i> did not activate the anti-inflammatory <i>IL-10</i> gene, but activated <i>IL-12p40</i> at 6 h. Hereby, <i>L. lactis</i> was proven to exert immunostimulatory properties in PBMC.https://www.mdpi.com/2073-4425/12/2/195<i>Lactococcus lactis</i><i>Saccharomyces cerevisiae</i>GalactooligosaccharidesTh1/Th2 immune responsespro-inflammatory |
| spellingShingle | Anna Slawinska Aleksandra Dunislawska Arkadiusz Plowiec José Gonçalves Maria Siwek TLR-Mediated Cytokine Gene Expression in Chicken Peripheral Blood Mononuclear Cells as a Measure to Characterize Immunobiotics Genes <i>Lactococcus lactis</i> <i>Saccharomyces cerevisiae</i> Galactooligosaccharides Th1/Th2 immune responses pro-inflammatory |
| title | TLR-Mediated Cytokine Gene Expression in Chicken Peripheral Blood Mononuclear Cells as a Measure to Characterize Immunobiotics |
| title_full | TLR-Mediated Cytokine Gene Expression in Chicken Peripheral Blood Mononuclear Cells as a Measure to Characterize Immunobiotics |
| title_fullStr | TLR-Mediated Cytokine Gene Expression in Chicken Peripheral Blood Mononuclear Cells as a Measure to Characterize Immunobiotics |
| title_full_unstemmed | TLR-Mediated Cytokine Gene Expression in Chicken Peripheral Blood Mononuclear Cells as a Measure to Characterize Immunobiotics |
| title_short | TLR-Mediated Cytokine Gene Expression in Chicken Peripheral Blood Mononuclear Cells as a Measure to Characterize Immunobiotics |
| title_sort | tlr mediated cytokine gene expression in chicken peripheral blood mononuclear cells as a measure to characterize immunobiotics |
| topic | <i>Lactococcus lactis</i> <i>Saccharomyces cerevisiae</i> Galactooligosaccharides Th1/Th2 immune responses pro-inflammatory |
| url | https://www.mdpi.com/2073-4425/12/2/195 |
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