Quantification of Proliferating Human Antigen-specific CD4&lt;sup&gt;+&lt;/sup&gt; T Cells using Carboxyfluorescein Succinimidyl Ester

Carluccio, Anthony R. Di; Tresoldi, Eleonora; So, Michelle; Mannering, Stuart I.

doi:10.3791/59545

Cited by 6 publications

(1 citation statement)

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“…Double polyclonal stimulation of PBMC-enriched lymphocytes was shown to promote a strong proliferation. Lymphocyte proliferation was measured by using the CFSE dilution method [ 55 ]. To this end, lymphocytes from two unrelated donors were stained for 8 min with 8 µm CFSE in basal media supplemented with 0.5% FBS and then washed twice in culture media before their use in experiments.…”

Section: Methodsmentioning

confidence: 99%

Human Omental Mesothelial Cells Impart an Immunomodulatory Landscape Impeding B- and T-Cell Activation

Gauthier

Rubio-Contreras

Gomez-Rosado

et al. 2022

IJMS

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Mesothelial cells form the mesothelium, a simple epithelium lining the walls of serous cavities and the surface of visceral organs. Although mesothelial cells are phenotypically well characterized, their immunoregulatory properties remain largely unknown, with only two studies reporting their capacity to inhibit T cells through TGF-β and their consumption of L-arginine by arginase-1. Whether human mesothelial cells can suppress other immune cells and possess additional leukosuppressive mechanisms, remain to be addressed to better delineate their therapeutic potential for cell therapy. Herein, we generated secretomes from omental mesothelial cells (OMC) and assess their capacity to inhibit lymphocytes proliferation, suppress activated T and B cells, as well as to modify macrophage activation markers. The secretome from mesenchymal stromal cells (MSC) served as a control of immuno-suppression. Although OMC and MSC were phenotypically divergent, their cytokine secretion patterns as well as expression of inflammatory and immunomodulary genes were similar. As such, OMC- and MSC-derived secretomes (OMC-S and MSC-S) both polarized RAW 264.7 macrophages towards a M2-like anti-inflammatory phenotype and suppressed mouse and human lymphocytes proliferation. OMC-S displayed a strong ability to suppress mouse- and human-activated CD19+/CD25+ B cells as compared to MSC-S. The lymphosuppressive activity of the OMC-S could be significantly counteracted either by SB-431542, an inhibitor of TGFβ and activin signaling pathways, or with a monoclonal antibody against the TGFβ1, β2, and β3 isoforms. A strong blockade of the OMC-S-mediated lymphosuppressive activity was achieved using L-NMMA, a specific inhibitor of nitric oxide synthase (NOS). Taken together, our results suggest that OMC are potent immunomodulators.

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Section: Methodsmentioning

confidence: 99%