Human lymph node fibroblastic reticular cells maintain heterogeneous characteristics in culture

Summary

Fibroblastic reticular cells (FRCs) are mesenchymal stromal cells in human lymph nodes (LNs) playing a pivotal role in adaptive immunity. Several FRC subsets have been identified, yet it remains to be elucidated if their heterogeneity is maintained upon culture. Here, we established a protocol to preserve and culture FRCs from human LNs and characterized their phenotypic profile in fresh LN suspensions and upon culture using multispectral flow cytometry. We found nine FRC subsets in fresh human LNs, independent of donor, of which four persisted in culture throughout several passages. Interestingly, the historically FRC-defining marker podoplanin (PDPN) was not present on all FRC subsets. Therefore, we propose that CD45 ^negCD31 ^neg human FRCs are not restricted by PDPN expression, as we found CD90, BST1, and CD146/MCAM to be more widely expressed. Together, our data provide insight into FRC heterogeneity in human LNs, enabling further investigation into the function of individual FRC subsets.

Graphical abstract

Highlights

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  Improved isolation and culture of human lymph node-derived FRCs

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  Nine heterogeneous clusters of FRCs found in ex vivo CD45 ^neg CD31 ^neg cells

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  Four out of nine clusters are preserved in vitro, while four new clusters emerge

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  Donor homogeneity indicates that not all FRCs are PDPN ⁺

Abstract

Biological sciences; Molecular biology; Cell biology