Abstract
BACKGROUND: Human hair follicle dermal sheath cup cells (DSCCs) hold promise as a cell source of regenerative medicine treatment for hair loss owing to their ability to secrete growth factors and/or signal pathway activators. The therapeutic effect of autologous DSCCs transplantation for male/female pattern hair loss (PHL) was demonstrated in a phase III equivalent clinical study. Intralesional inflammation has been implicated in the pathophysiology of various hair loss diseases, including PHL. As DSCCs possess mesenchymal stem/stromal cell (MSC)-like properties and MSCs are immunosuppressive, we investigated whether they exhibit immunoregulatory capabilities comparable to MSCs and developed an in vitro morphometric assay to predict this capability. METHODS: DSCCs were isolated via microdissection and propagated in vitro. Their conformity to MSC criteria was assessed based on cell surface antigen expression and differentiation potential. Furthermore, immunoregulatory capabilities were assessed by co-culturing DSCCs with anti-CD3/28 antibody-stimulated peripheral blood mononuclear cells (PBMCs) and examining the suppression mechanisms through pharmacological intervention. Multiple lots of DSCCs derived from various donors and manufacturing conditions were cultured and analyzed by phase-contrast microscopy to obtain their morphometric profiles. Parameters correlating with the expression levels of immunomodulatory factors were used to create a predictive model. Additional DSCC lots were manufactured to validate the predictive model. RESULTS: Similar to MSCs, cell differentiation assays revealed that DSCCs exhibited multipotency, and they did not express co-stimulatory molecules in response to immunogenic stimuli, suggesting low immunogenicity. Moreover, co-culture experiments with allogeneic PBMCs revealed that DSCCs reduced T cell proliferation (from 78 to 5%) and increased the proportion of FOXP3-positive regulatory T cells (from 3 to 10%) relative to the baseline. This capability was indoleamine 2,3-dioxygenase 1 (IDO)-dependent and affected by IDO expression levels. Image analysis of DSCCs revealed morphometric parameters that were highly correlated with IDO expression, and a model established using these parameters was able to predict IDO expression levels in the validation lots. CONCLUSIONS: Results suggest that human DSCCs can help regenerative medicine approaches, not only for supporting hair follicle regeneration but also for suppressing microinflammation that potentially contributes to hair loss, and this can be readily evaluated using a newly developed morphometric analysis.