Exploring fibroblast heterogeneity in the formation of TLS in the context of primary Sjögrens syndrome

The immune system is vital in the protection and maintenance of the body in a healthy state against extracellular pathogens, tissue injury and self-antigens (autoimmunity). Tertiary lymphoid structures (TLS) are non-encapsulated immune cell aggregates which can arise in response to pathogenic insult, cancer and tissue transplant in order to support antigen-specific immune responses in non-immune organs. Contrastingly, in the context of inflammation and autoimmunity, TLS can form within permissive tissues and cause tissue damage and destruction. The Barone group has explored the lymphoid function and phenotype of the immunofibroblast which contribute to the formation and organisation of these lymphocytic aggregates in the formation and organisation of TLS. Originally grouped coarsely, fibroblasts have been more recently recognized as a largely heterogeneous population defined by uniquely assigned phenotypes and functions. In this thesis, we deconvolute the stromal cell heterogeneity using scRNAseq in human primary Sjogren’s syndrome (pSS) salivary glands (SG) and inflamed murine SG and explore how these cell populations possibly underpin TLS diversity and organisation. We apply bioinformatic tools to explore mRNA gene expression patterns within cell populations, identifying a CCL21+ CCL19+ “immunopericyte” population which has not yet been explored in the context of TLS prior. We also explore the possible signalling mechanisms engaged within the immunofibroblast and “immunopericyte” populations using the inducible TLS mouse model. This work has created a cellular atlas of the human and mouse pSS SG and the different cell populations present within these primary tissues. This allows for future work to validate the mRNA dataset at the protein level and generate potential novel therapeutic targets at a cell specific level for patients

Immunofibroblasts are pivotal drivers of tertiary lymphoid structure formation and local pathology.

Resident fibroblasts at sites of infection, chronic inflammation, or cancer undergo phenotypic and functional changes to support leukocyte migration and, in some cases, aggregation into tertiary lymphoid structures (TLS). The molecular programming that shapes these changes and the functional requirements of this population in TLS development are unclear. Here, we demonstrate that external triggers at mucosal sites are able to induce the progressive differentiation of a population of podoplanin (pdpn)-positive stromal cells into a network of immunofibroblasts that are able to support the earliest phases of TLS establishment. This program of events, that precedes lymphocyte infiltration in the tissue, is mediated by paracrine and autocrine signals mainly regulated by IL13. This initial fibroblast network is expanded and stabilized, once lymphocytes are recruited, by the local production of the cytokines IL22 and lymphotoxin. Interfering with this regulated program of events or depleting the immunofibroblasts in vivo results in abrogation of local pathology, demonstrating the functional role of immunofibroblasts in supporting TLS maintenance in the tissue and suggesting novel therapeutic targets in TLS-associated diseases

N-(4-Methoxyphenyl)benzenesulfonamide

In the title compound, C13H13NO3S, the benzene ring of the benzenesulfonamide moiety is disordered with an occupancy ratio of 0.56 (3):0.44 (3), the disorder components being twisted at and angle of 21 (1)° to each other. The methoxybenzene group is roughly planar (r.m.s. deviation = 0.0144 Å) and the amide N atom is displaced from this plane by 0.090 (6) Å. The dihedral angles between the methoxybenzene group and the major and minor occupancy components of the disordered benzene ring are 54.6 (4) and 62.9 (5)°, respectively. In the crystal, infinite polymeric chains are formed along [100] due to intermolecular N—H...O hydrogen bonding. Weak C—H...π interactions are also present in the crystal

Immunofibroblasts regulate LTα3 expression in tertiary lymphoid structures in a pathway dependent on ICOS/ICOSL interaction.

Immunofibroblasts have been described within tertiary lymphoid structures (TLS) that regulate lymphocyte aggregation at sites of chronic inflammation. Here we report, for the first time, an immunoregulatory property of this population, dependent on inducible T-cell co-stimulator ligand and its ligand (ICOS/ICOS-L). During inflammation, immunofibroblasts, alongside other antigen presenting cells, like dendritic cells (DCs), upregulate ICOSL, binding incoming ICOS + T cells and inducing LTα3 production that, in turn, drives the chemokine production required for TLS assembly via TNFRI/II engagement. Pharmacological or genetic blocking of ICOS/ICOS-L interaction results in defective LTα expression, abrogating both lymphoid chemokine production and TLS formation. These data provide evidence of a previously unknown function for ICOSL-ICOS interaction, unveil a novel immunomodulatory function for immunofibroblasts, and reveal a key regulatory function of LTα3, both as biomarker of TLS establishment and as first driver of TLS formation and maintenance in mice and humans