Mesenchymal stem cells derived from adipose tissue (ASC) have immune regulatory function, which makes them interesting candidates for cellular therapy. ASC cultures are however heterogeneous in phenotype. It is unclear whether all ASC contribute equally to immunomodulatory processes. ASC are also responsive to cytokine stimulation, which may affect the ratio between more and less potent ASC populations. In the present study, we determined IL-6 receptor (CD126 and CD130 subunits) and IFN-γ receptor (CD119) expression on ASC by flow cytometry. The production of IL-6 and IFN-γ was measured by ELISA and the frequency of IL-6 and IFN-γ secreting cells by ELISPOT. The results showed that ASC did not express CD126, and only 10–20% of ASC expressed CD130 on their surface, whereas 18–31% of ASC expressed CD119. ASC produced high levels of IL-6 and 100% of ASC were capable of secreting IL-6. Stimulation by IFN-γ or TGF-β had no effect on IL-6 secretion by ASC. IFN-γ was produced by only 1.4% of ASC, and TGF-β significantly increased the frequency to 2.7%. These results demonstrate that ASC cultures are heterogeneous in their cytokine secretion and receptor expression profiles. This knowledge can be employed for selection of potent, cytokine-producing, or responsive ASC subsets for cellular immunotherapy.

Estimates of precursor frequency and assessment of functional characteristics of alloreactive CD4+ T cells are all biased by the need for long-term culture. In this study, direct visualization of human alloreactive CD4+ T cells on the single-cell level was achieved using cell surface expression of CD154 as a tool for identification. The average frequency of alloreactive CD154+CD4+ T cells among peripheral blood CD4+ T cells was 0.1%, with half of the cells displaying a naive phenotype. The proliferation capacity and expression of cytokines after allogeneic stimulation resided in these CD154+CD4+ T cells. The repertoire of alloreactive CD4+ T cells was biased to a Th17 response, and on average 24% of alloreactive CD154+CD4+ memory T cells produced interleukin-17 (IL-17) after polyclonal stimulation. Unexpectedly, mixed cell cultures from human leukocyte antigen (HLA)–identical donors also generated alloreactive CD154+CD4+ T cells and yielded the highest frequency compared with HLA-nonidentical combinations. Therefore, reactivity to minor histocompatibility antigens between HLA-identical subjects appears to be relatively common. Alloreactive HLA-identical T cells did not proliferate or express cytokines, but were driven to proliferation in the presence of exogenous IL-2.

Introduction: De novo donor-specific antibodies (dnDSA) directed against HLA are a major contributing factor to the chronic deterioration of renal allograft function. Several factors, including the degree of HLA matching, younger recipient age and past sensitization events have been shown to increase the risk for the development of dnDSA. The development of dnDSA is also strongly associated with modifications in the immunosuppressive regimen, non-adherence and under-immunosuppression. Areas covered: Tacrolimus is widely used after solid organ transplantation (SOT) and in recent years, both a high intra-patient variability in tacrolimus exposure as well as low tacrolimus exposure have been found to be associated with a higher risk of dnDSA development in kidney transplant recipients. This article provides an overview of current findings published in the recent five years regarding the relationship between tacrolimus exposure and variation therein and the development of dnDSA. Expert opinion: In this review we describe how combining data on tacrolimus intra-patient variability and mean pre-dose concentration may be an effective tool to identify kidney transplant recipients who are at higher risk of developing dnDSA.