Detection and consistency of mucosal fluid T lymphocyte phenotypes and their relationship with blood, age and gender

Innate and adaptive immune responses at mucosal surfaces play a role in protection against most infectious diseases. However, the relative importance either of mucosal versus systemic, or of cellular versus humoral immunity in protection against such infections remains unclear. We aimed to determine...

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Published in:	Journal of immunological methods Vol. 532; p. 113731
Main Authors:	Nasab, Shervin Dokht Sadeghi, Eniya, Muruganantham Lillimary, Judith, Albert, Clasen, Frederick, Faith, Beulah, Poongulali, Selvamuthu, Gita, Jayaraman Bhagavad, Ashok, Chakrapani, Raghavi, Velmurugan, Vedavalli, Subramanian, Lavanya, Chandra, Ranganathan, Kannan, Rajan, Gunaseelan, Kumarasamy, Nagalingeswaran, Moyes, David, Ide, Mark, Shoaie, Saeed, Kurushima, Yuko, Jagdev, Daljit, Pun, Mina, Johnson, Newell, Kannian, Priya, Challacombe, Stephen
Format:	Journal Article
Language:	English
Published:	Netherlands Elsevier B.V 01-09-2024
Subjects:	Consistency Mucosal immunity Peripheral blood mononuclear cells (PBMCs) Reproducibility Saliva Stimulated whole mouth fluid (SWMF) T lymphocytes T lymphocytes PBS CD Mucosal immunity Consistency Peripheral blood mononuclear cells (PBMCs) FACS PBMC COVID-19 SARS-CoV2 SD SE HIV Reproducibility SWMF Stimulated whole mouth fluid (SWMF) Saliva
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Summary:	Innate and adaptive immune responses at mucosal surfaces play a role in protection against most infectious diseases. However, the relative importance either of mucosal versus systemic, or of cellular versus humoral immunity in protection against such infections remains unclear. We aimed to determine the relative percentages and reproducibility of detection of five major T lymphocyte phenotypes in stimulated whole mouth fluid (SWMF); to compare matched mucosal and blood phenotypes; to evaluate the consistency of phenotypes in SWMF over time; and to determine any associations with age or gender. Peripheral blood and SWMF samples were collected from 194 participants and sequential concomitant samples were collected from 27 of those and from 12 subjects living with HIV. CD3, CD4, CD8, Th1 and Th2 T lymphocyte phenotypes were determined by FACS. All the five T lymphocyte phenotypes were detected consistently by FACS in PBMC and SWMF with experimental replicates (N = 10; PBMC CV: 3–30%; SWMF CV: 12–36%). In longitudinal samples detection rates were reproducible in both fluids but variations were higher in SWMF (CV: 23–79.6%) than PBMC (CV: 9.7–75%). Statistically significant correlations of the percentages of all the T lymphocyte phenotypes except CD8 was seen between the two fluids. In PBMCs a negative correlation with age was found with CD3, CD4 and CD8 phenotypes, whilst a positive correlation was found in both SWMF and PBMC with the Th2 phenotype. CD3, CD4 and CD8 phenotypes in SWMF and PBMCs from an HIV-positive cohort were not significantly correlated in contrast with the HIV-negative controls. Our study provides a robust FACS protocol for the detection of the five major T lymphocyte phenotypes in SWMF which should prove useful for research with other mucosal fluids. •Five major T lymphocyte phenotypes can be detected consistently by flow cytometry in mucosal fluid samples.•Biological variation of T lymphocyte phenotype percentages in longitudinal SWMF samples was greater than in blood.•CD3, CD4, Th1 and Th2 but not CD8 T lymphocyte phenotype percentages in the SWMF and PBMCs were positively correlated.•T lymphocyte phenotype distribution in the SWMF expressed as a percentage of CD3 positive cells differed from PBMCs.•The percentages of the T lymphocyte phenotypes in the SWMF apart from Th1 did not differ by gender.•Age was negatively correlated with CD3, CD4, and CD8 values in PBMCs, but positively correlated with Th2 in both SWMF and PBMCs.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	0022-1759 1872-7905 1872-7905
DOI:	10.1016/j.jim.2024.113731