Functional expression of aryl hydrocarbon receptor on mast cells populating human endometriotic tissues.

Endometriosis is an inflammatory disease characterized by the presence of ectopic endometrial tissue outside the uterus. A diffuse infiltration of mast cells (MCs) is observed throughout endometriotic lesions, but little is known about how these cells contribute to the network of molecules that modulate the growth of ectopic endometrial implants and promote endometriosis-associated inflammation. The aryl hydrocarbon receptor (AhR), a transcription factor known to respond to environmental toxins and endogenous compounds, is present in MCs. In response to AhR activation, MCs produce IL-17 and reactive oxygen species, highlighting the potential impact of AhR ligands on inflammation via MCs. Here, we investigated the possibility that endometrial MCs promote an inflammatory microenvironment by sensing AhR ligands, thus sustaining endometriosis development. Using human endometriotic tissue (ET) samples, we performed the following experiments: (i) examined the cytokine expression profile; (ii) counted AhR-expressing MCs; (iii) verified the phenotype of AhR-expressing MCs to establish whether MCs have a tolerogenic (IL-10-positive) or inflammatory (IL-17-positive) phenotype; (iv) measured the presence of AhR ligands (tryptophan-derived kynurenine) and tryptophan-metabolizing enzymes (indoleamine 2,3-dioxygenase 1 (IDO1)); (v) treated ET organ cultures with an AhR antagonist in vitro to measure changes in the cytokine milieu; and (vi) measured the growth of endometrial stromal cells cultured with AhR-activated MC-conditioned medium. We found that ET tissue was conducive to cytokine production, orchestrating chronic inflammation and a population of AhR-expressing MCs that are both IL-17 and IL-10-positive. ET was rich in IDO1 and the AhR-ligand kynurenine compared with control tissue, possibly promoting MC activation through AhR. ET was susceptible to treatment with an AhR antagonist, and endometrial stromal cell growth was improved in the presence of soluble factors released by MCs on AhR activation. These results suggest a new mechanistic role of MCs in the pathogenesis of endometriosis.

Variants of beta-microglobulin cleaved at lysine-58 retain the main conformational features of the native protein but are more conformationally heterogeneous and unstable at physiological temperature.

Cleavage of the small amyloidogenic protein beta2-microglobulin after lysine-58 renders it more prone to unfolding and aggregation. This is important for dialysis-related beta2-microglobulin amyloidosis, since elevated levels of cleaved beta2-microglobulin may be found in the circulation of dialysis patients. However, the solution structures of these cleaved beta2-microglobulin variants have not yet been assessed using single-residue techniques. We here use such methods to examine beta2-microglobulin cleaved after lysine-58 and the further processed variant (found in vivo) from which lysine-58 is removed. We find that the solution stability of both variants, especially of beta2-microglobulin from which lysine-58 is removed, is much reduced compared to wild-type beta2-microglobulin and is strongly dependent on temperature and protein concentration. 1H-NMR spectroscopy and amide hydrogen (1H/2H) exchange monitored by MS show that the overall three-dimensional structure of the variants is similar to that of wild-type beta2-microglobulin at subphysiological temperatures. However, deviations do occur, especially in the arrangement of the B, D and E beta-strands close to the D-E loop cleavage site at lysine-58, and the experiments suggest conformational heterogeneity of the two variants. Two-dimensional NMR spectroscopy indicates that this heterogeneity involves an equilibrium between the native-like fold and at least one conformational intermediate resembling intermediates found in other structurally altered beta2-microglobulin molecules. This is the first single-residue resolution study of a specific beta2-microglobulin variant that has been found circulating in dialysis patients. The instability and conformational heterogeneity of this variant suggest its involvement in beta2-microglobulin amyloidogenicity in vivo.