Human Ad-MSCs possess extraordinary properties of attenuating effector T cell responses, which have been demonstrated in diverse immune disorders. The present study found that human Ad-MSCs specifically inhibited IL-17-producing CD4+ T cell differentiation under the Th17 polarizing condition, which was accompanied by suppressing the expression of the transcription factor RORγt. They induced a functional CD4+CD25+Foxp3+ Treg cell phenotype evidenced by the secretion of IL-10 under the Treg polarizing condition. These findings are in accordance with expectations of a large number of studies searching for innovative therapeutics for PD patients [12, 20, 31, 36–38].
Treg abnormalities in PD patients have been shown either by its reduced number or by its functional deficit [5, 6, 18, 20]. This might be due to the increased number of Th17 cells as well as the increased secretion of IL-17 in the serum of PD patients. Blockage of IL-17 or neutralizing it with an IL-17 antibody rescued neuronal death [6, 7, 9–11]. Animal studies have revealed that an increased number of Th17 cells lead to tissue damage by producing a neurotoxic cytokine-interleukin IL-17[11–14, 39], which is attenuated by Treg cells . Animal studies and clinical trials have demonstrated that Ad-MSCs are a potential therapeutic strategy due to their capability of suppressing Th17 responses and inducing functional Treg cells [25–30, 40–42]. Based on this, it was postulated that Ad-MSCs would modulate immune responses in PD patients in the same way as it did in animals. We found that Ad-MSCs inhibited the differentiation of peripheral CD4+ T cells into Th17 cells and induced their differentiation into functional Treg cells evidenced by the secretion of IL-10. Our results confirmed the hypothesis and demonstrated the beneficial role of Ad-MSCs in regulating immune responses in vitro.
Based on our previous findings that Ad-MSCs mediated immunosuppression of Th17 in MS patients and a mouse model of MS [32, 33], we hypothesized that LIF was the key player in mediating the suppression of Th17 cell differentiation. In the co-cultured Ad-MSCs-CD4+ T cell experiment, we found that Ad-MSCs accelerated the differentiation of CD4+ T cells towards Treg, evidenced by the increased secretion of IL-10, and suppressed differentiation towards IL-17-secreting Th17 cells, which was supported by the decreased expression of RORγt. We also found that LIF protein was significantly increased when CD4+ T cells were co-cultured with Ad-MSCs than when T cells were cultured alone under both Th17 and Treg polarizing conditions. In line with the increased protein level, the level of LIFR mRNA was also increased. Furthermore, expression of LIFR mRNA was increased under both Th17 and Treg polarizing conditions.
Both IL-23R and IL-6R are involved in the signaling pathways that lead to Th17 cell differentiation [21–23]. Consistent with our hypothesis, levels of IL-6R mRNA were downregulated under both Th17 and Treg polarizing conditions, and IL-23R mRNA was decreased under the Th17 polarizing condition. This was supported by the increased expression of RORγt and IL-6R mRNA after neutralizing LIF with an antibody as shown in our previous study . These suggest that LIF is the key mediator of this process, which is in line with previous findings in that TGF-β and PGE2 are involved in Treg upregulation and Th17 downregulation in humans .
LIF belongs to the IL-6-type cytokine family and it is important for T-cell maturation. It is different from IL-6 in that it binds to gp130/gp190 hetero-dimers, whereas IL-6 binds to gp130 homodimers . Gp130, therefore, is the shared subunit of cytokine receptors of the IL-6 family. IL-6 is known to inhibit the expression of the LIFR subunit, opposing the physiological effect of LIF [44, 45]. This might be the explanation for the increased LIFR level observed in our study.
LIF suppresses expression of Th17 specific genes such as the transcription factor RORγt, and simultaneously increases expression of a Treg specific transcription factor-Foxp3 [34, 35, 45]. In contrast, IL-6 decreased the level of Foxp3 and increased the level of RORγt [21, 45]. Therefore, LIF/IL-6 balance is critical for T-cell lineage maturation . These results suggest that LIF, in combination with IL-6 signal pathway blockers, might become promising therapeutics for PD and other immune disorders  though the underlying mechanism remains largely unknown.
In conclusion, human Ad-MSCs suppress the differentiation of CD4+T cells isolated from PD patients into Th17 cells in vitro. This suppressive effect was mainly associated with an increase in functional CD4+CD25+Foxp3+ Treg cells and IL-10 secretion. Ad-MSCs might be a promising candidate in regulating inflammatory responses and in managing PD.