Immunological Landscapes in Lung Transplantation: Insights from T Cell Profiling in BAL and PBMC
Abstract
:1. Introduction
2. Results
2.1. Multi-Colour Flow Cytometric Panel to Identify T Cell Subsets Comparing PBMC and BAL
2.2. CD4 and CD8 T Cells Comparing PBMC and BAL
2.3. Effector T Cells
2.4. Memory T Cell Populations Comparing PBMC and BAL
2.5. Non-Tregs Expressing CD39 Comparing PBMC and BAL
2.6. Regulatory T Cells Comparing PBMC and BAL
2.7. Subpopulation of Tregs Comparing PBMC and BAL
2.8. Teff and Treg Balance (Granzyme B:Treg Ratio) Comparing PBMC and BAL
2.9. T Cell Subsets and Association with CLAD
2.9.1. Effector T Cells (Teffs) and Memory T Cells (Tmems) Comparing PBMC and BAL
2.9.2. Treg Populations Comparing PBMC and BAL
2.9.3. Teffs and Tregs Balance (Granzyme B–Treg Ratio) Comparing PBMC and BAL
3. Discussion
4. Methods
4.1. Patient Demographics
4.2. Ethics Approval
4.3. Isolation of PBMCs from Peripheral PBMC
4.4. Isolation of Immune Cells from BAL
4.5. Flow Cytometric Analysis
4.6. Lung Function Parameters
4.7. Statistical Analysis
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
References
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Transplant (n = 20) | Non-Transplant (n = 4) | |
---|---|---|
Gender | ||
Male | 10 | 3 |
Female | 10 | 1 |
Initial disease condition for transplant | ||
Cystic Fibrosis | 11 | N/A |
COPD | 4 | N/A |
IPF | 4 | N/A |
Histiocytosis | 1 | N/A |
Type of transplant | ||
Bilateral | 19 | N/A |
Heart and Lung | 1 | N/A |
Time post-transplant (months) | 52.83 (SEM ± 10.84) | N/A |
Age at bronchoscopy (years) | 43.66 (SEM ± 3.78) | 46.96 (SEM ± 9.38) |
Marker | Purpose | Antibody Used |
---|---|---|
CD3 | Marker for T lymphocytes | anti-CD3-AF700 (BioLegend) |
CD4 | Marker for CD4 T cells | anti-CD4-BV510 (BioLegend) |
CD8 | Marker for CD8 T cells | anti-CD8-APC-H7 (BD Biosciences) |
CD14 | Marker of macrophages and granulocytes. | anti-CD14-PE-Cy 7 (BD Biosciences) |
CD25 | Regulatory T cells express high CD25 | anti-CD25-BV421 (BD Biosciences) |
CD127 | Regulatory T cells express low CD127 | anti-CD127-FITC (BD Biosciences) |
CD39 | Immunosuppressive ecto-enzyme converts ATP to ADP | anti-CD39-BV711 (BioLegend) |
CD73 | Immunosuppressive ecto-enzyme coverts ADP to AMP | anti-CD73-BV605 (BD Biosciences) |
CD194 | Marker of cell migration, also known as CCR4 | anti-CD194-PE-Cy 7 (BD Biosciences) |
CD45RO | Marker of memory cells | anti-CD45RO-BV650 (BD Biosciences) |
HLA-DR | Marker of MHC II and upregulated in activated Treg | anti-HLA-DR-BV785 (BioLegend) |
Ki67 | Marker of cellular proliferation | anti-Ki67-PerCp- Cy 5.5 (BioLegend) |
FOXP3 | Transcription factor specific to regulatory T cells | anti-FoxP3-PE (BD Biosciences) |
Granzyme B | Serine protease commonly found in the granules of cytotoxic T cells | anti-granzyme B-APC (Invitrogen) |
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de Silva, T.A.; Apte, S.; Voisey, J.; Spann, K.; Tan, M.; Chambers, D.; O’Sullivan, B. Immunological Landscapes in Lung Transplantation: Insights from T Cell Profiling in BAL and PBMC. Int. J. Mol. Sci. 2024, 25, 2476. https://doi.org/10.3390/ijms25052476
de Silva TA, Apte S, Voisey J, Spann K, Tan M, Chambers D, O’Sullivan B. Immunological Landscapes in Lung Transplantation: Insights from T Cell Profiling in BAL and PBMC. International Journal of Molecular Sciences. 2024; 25(5):2476. https://doi.org/10.3390/ijms25052476
Chicago/Turabian Stylede Silva, Tharushi Ayanthika, Simon Apte, Joanne Voisey, Kirsten Spann, Maxine Tan, Daniel Chambers, and Brendan O’Sullivan. 2024. "Immunological Landscapes in Lung Transplantation: Insights from T Cell Profiling in BAL and PBMC" International Journal of Molecular Sciences 25, no. 5: 2476. https://doi.org/10.3390/ijms25052476