Due to their suppressive capacity, regulatory T cells (Tregs) have attracted growing interest as an adoptive cellular therapy for the prevention of allograft rejection, but limited Treg recovery and lower quality of adult-derived Tregs could represent an obstacle to success. To address this challenge, we developed a new approach that provides large quantities of Tregs with high purity and excellent features, sourced from thymic tissue routinely removed during pediatric cardiac surgeries (thyTregs). We report on a 2-year follow-up of the first patient treated worldwide with thyTregs, included in a phase I/II clinical trial evaluating the administration of autologous thyTreg in infants undergoing heart transplantation. In addition to observing no adverse effects that could be attributed to thyTreg administration, we report that the Treg frequency in the periphery was preserved during the 2-year follow-up period. These initial results are consistent with the trial objective, which is to confirm safety of the autologous thyTreg administration and its capacity to restore the Treg pool.

Regulatory T cells (Tregs), which are characterized by the expression of the transcription factor forkhead box P3 (FOXP3), are the main immune cells that induce tolerance and are regulators of immune homeostasis. Natural Treg cells (nTregs), described as CD4⁺CD25⁺FOXP3⁺, are generated in the thymus via activation and cytokine signaling. Transforming growth factor beta type 1 (TGF-β1) is pivotal to the generation of the nTreg lineage, its maintenance in the thymus, and to generating induced Treg cells (iTregs) in the periphery or in vitro arising from conventional T cells (Tconvs). Here, we tested whether TGF-β1 treatment, associated with interleukin-2 (IL-2) and CD3/CD28 stimulation, could generate functional Treg-like cells from human thymocytes in vitro, as it does from Tconvs. Additionally, we genetically manipulated the cells for ectopic FOXP3 expression, along with the TGF-β1 treatment. We demonstrated that TGF-β1 and ectopic FOXP3, combined with IL-2 and through CD3/CD28 activation, transformed human thymocytes into cells that expressed high levels of Treg-associated markers. However, these cells also presented a lack of homogeneous suppressive function and an unstable proinflammatory cytokine profile. Therefore, thymocyte-derived cells, activated with the same stimuli as Tconvs, were not an appropriate alternative for inducing cells with a Treg-like phenotype and function.

Optical tissue clearing (OTC) techniques have transformed three‐dimensional (3D) biological imaging, yet existing protocols often face limitations related to pigment persistence, reagent viscosity, or restricted applicability across tissues and species. Here, we introduce White and Clear (WaC), a tunable hydrophilic tissue clearing strategy developed from the CUBIC (clear, unobstructed brain imaging cocktails) workflow that integrates efficient delipidation, hydrogen peroxide‐based pigment removal, and customizable refractive index matching using 2,2′‐thiodiethanol (TDE). WaC provides rapid, versatile, and reproducible clearing across multiple organs and species while preserving morphology and immunoreactivity, and enabling deep 3D imaging with confocal and light‐sheet microscopy. WaC improves ease of use, versatility and clearing efficiency across samples from multiple species, including rodents, pigs, and humans; it also includes the capability for whole‐body clearing of small animals like mice; compatibility with 3D imaging techniques such as selective plane illumination microscopy (SPIM)/light‐sheet fluorescence microscopy (LSFM); the possibility of fine‐tuning clearing parameters for optimal results in different situations; and the avoidance of cumbersome synthesis or application procedures as well as the use of toxic reagents.

Background: The pathophysiology of non-IgE-mediated cow’s milk allergy is mostly unknown. Previous studies suggested a mechanism mediated by T cells, but this was not confirmed in subsequent studies. The aim of this study was to investigate the immunological mechanisms, especially the role of regulatory T cells (Tregs), in the pathophysiology of allergic proctocolitis (FPIAP). Methods: A prospective observational study was conducted on infants with FPIAP and a control group of healthy infants with similar ages. The main variables were lymphocyte populations, included Tregs, which were extracted from peripheral blood and processed immediately by flow cytometry at two time points: in the acute phase (“T0”) and after clinical resolution (“Tres”). Results: A total of 32 patients with FPIAP and 10 healthy infants were enrolled. There was a higher T-CD4 memory cell count, increased numbers of regulatory B cells and a higher percentage of Tregs (p < 0.01) in patients with acute FPIAP in contrast to the healthy group. The levels of granulocytes (mainly eosinophils), dendritic cells (mDC2) and NK16+56- cells were also significantly higher in the FPIAP group. NK16+56- cells and the number of granulocytes appeared to be the best markers for distinguishing between the healthy and FPIAP infants based on the ROC curves. Conclusions: FPIAP does not appear to have an immune mechanism mediated by T cells, but it may be associated with innate immunity responses characterized by an increase in NK16+56- cells, eosinophils and dendritic cells. These cells could be evaluated in future studies as possible markers of non-IgE-mediated cow’s milk protein allergy.