The essence of sepsis is the body’s inflammatory response to pathogenic microbial infection. In addition to the direct damage of pathogens and their toxins, the immune function of the body also plays a crucial role in the process of sepsis. Treg cells are classic immune regulatory cells. Previous studies have shown that Treg cells can mediate changes in the TH1/TH2 balance during the pathophysiological process of sepsis, thereby affecting the outcome of inflammatory responses. Kinsey et al[14]used antibodies (PC61) to eliminate Treg cells in mice 5 days before generating an ischemia-reperfusion injury-induced AKI model. As a result, the local infiltration of inflammatory cells significantly increased in the kidneys, and the secretion of IL-6, TNF-α and TGF- β (but not IL-10) increased, thereby aggravating renal function damage and renal tubular necrosis. To further verify the relationship between Treg cells and AKI, Kinsey injected Treg cells from wild-type mice and Foxp3-deficient mice into RAG-1 mice (lacking T and B cells) before AKI modeling. The former alleviated AKI while the latter increased the infiltration of local neutrophils and macrophages in the kidneys, thereby aggravating AKI. Gandolfo et al[15]used PC61 to eliminate Treg cells in mice 24 h after AKI injury and found that it could still lead to increased Cr, while the transfer of Treg cells from wild-type mice 24 h after injury significantly reduced tubular necrosis scores and promoted tubular epithelial cell regeneration, suggesting that Treg cells are involved in AKI repair. Our study also confirmed that although the local Treg cells in septic kidney tissues were slightly increased, the damage was too severe to reverse the changes with respect to the TH1/TH2 balance, resulting in kidney injury.
TH17 cells are a newly identified T cell subpopulation that mainly secretes IL-17. IL-17 secreted by TH17 cells recruits neutrophils and promotes inflammatory mediator release[16]. Studies have shown that in the peripheral blood of septic mice, IL-17 increased in a time-dependent manner. The injection of antibodies to neutralize IL-17 before modeling significantly improved the survival of mice and reduced the expression of inflammatory factors and chemokines. Even if the neutralization of IL-17 is delayed until 12 h after modeling, it can still result in the improvement of sepsis[17].
In addition to an increase in IL-17 in an animal model of sepsis, Brunialti et al[18] also detected significantly increased IL-17 and significantly increased IL-17-secreting TH17 cells in the peripheral blood of patients with sepsis. In addition, in recent years, studies have made progress in establishing the role of IL-17 in kidney diseases. Elevated IL-17 expression was detected by immunofluorescence in kidney tissues from patients with focal segmental sclerosing glomerulonephritis, membranoproliferative glomerulonephritis, and microscopic lesions[19].
Li et al[20] found that IL-17 was significantly increased in an IRI-AKI model and that IL-17 or IL-17 receptor-deficient mice and IL-17 neutralization all alleviated AKI injury, indicating that IL-17 was involved in the pathogenesis of sepsis and AKI. Our study found a significant increase in the percentage of local TH17 cells in the kidney of septic mice. Although the percentage of Treg cells also increased slightly, it was much less than the increase in TH17 cells, suggesting a role of the TH17-mediated TH1 inflammatory response in the pathogenesis of SA-AKI.
Recent studies have found that in addition to the regulation of TH1 cells, the Gal-9/Tim-3 signaling pathway is also involved in the regulation of TH17 and Treg cells[21, 22]. When the Tim-3 pathway was blocked by a Tim-3 antibody, the levels of TH1- and TH17-related cytokines (IFN-γ, IL-17, IL-2, and IL-6) increased, and the immunosuppressive function of Treg cells significantly decreased; furthermore, the expression of cytokine IL-10 also significantly decreased[23].
Tim-3 expression is dysregulated during the pathological process of sepsis. The expression of Tim-3 in patients with severe sepsis is significantly lower than that in patients with mild sepsis. Consistent with this, in a mouse model of sepsis, Tim-3 expression was significantly reduced in the acute/severe phase, and blockade of the Tim-3 pathway by Tim-3 antibodies and Tim-3 fusion proteins reduced the survival rate, increased proinflammatory factor release, and exacerbated septic injury[24].
In an in vitro study of sepsis, lipopolysaccharide (LPS) caused the upregulation of the expression of inflammatory factors such as IFN-γ, TNF-α and IL-17, indicative of excessive proliferation and activation of TH1 and TH17 cells[25], while Gal-9 inhibited the upregulation of these factors and promoted the production of the TH2 and Treg cell-related cytokines IL-4 and IL-10[26].
These results indicate that the Gal-9/Tim-3 pathway can regulate the secretion of TH17/Treg cells and related factors, IL-17/IL-10, in sepsis and kidney diseases. Our previous study found that IL-17 knockout improved SA-AKI and increased IL-10 while reducing IL-6 levels[27]. This finding indicates that IL-17 is involved in the pathogenesis of SA-AKI. In addition, in an IRI-AKI model, increasing the percentage of Treg cells and the secretion of IL-10 significantly improved AKI, while clearing Treg cells accelerated the deterioration of AKI and reduced IL-10 levels[28]. This study further confirmed that the number of IL-10-secreting Treg cells in SA-AKI was significantly reduced, while the number of IL-17-secreting TH17 cells was significantly increased, and after exogenous Gal-9 activation of the Gal-9/Tim-3 pathway, the number of IL-17-secreting TH17 cells also decreased, while the number of IL-10-secreting Treg cells increased. In addition, blockade of the Gal-9/Tim-3 pathway with Tim-3 further aggravated the local TH17/Treg imbalance in the kidneys of septic mice, resulting in an increase in TH17 cells and a decrease in the number of IL-10-secreting Treg cells. In summary, TH17/Treg cells and IL-17/IL-10 secreted by TH17/Treg cells are important pathogenic factors involved in SA-AKI. Inducing TH17/Treg cells to establish a new balance may become a new target for SA-AKI treatment. The Gal-9/Tim-3 pathway may be an important mechanism affecting the TH17/Treg ratio. Further regulation of the TH17/Treg balance by manipulating the Gal-9/Tim-3 pathway may become a breakthrough strategy.
Previous studies have found that MSCs can induce IL-10 secretion by Treg cells in an IRI-AKI model[28]. Our previous studies demonstrated that MSCs inhibited the secretion of IL-17 by TH17 cells in SA-AKI mouse and that MSCs increased IL-10 expression in SA-AKI mice.
Recent studies have found that MSCs can inhibit T cell function through the secretion of Gal-1 and Gal-3. Silencing the expression of Gal-1 and/or Gal-3 in MSCs with small interfering RNA can partially or even completely eliminate the proliferative effect of MSCs on T cells[29].
The present study found that the administration of MSCs in SA-AKI mice significantly reversed the TH1/TH2 balance, resulting in a significant decrease in the percentages of IL-17-secreting TH17 cells and a significant increase in IL-10-secreting Treg cells. In addition, the activation of the Gal-9/Tim-3 pathway by exogenous Gal-9 reduced the number of IL-17-secreting TH17 cells and increased the number of IL-10-secreting Treg cells. Blocking the Gal-9/Tim-3 pathway with Tim-3 further aggravated the local TH17/Treg imbalance in the kidneys of septic mice, resulting in an increase in TH17 cells and a decrease in Il-10-secreting Treg cells. Therefore, we speculate that reestablishing the TH17/Treg balance by MSCs through the Gal-9/Tim-3 pathway might be an important mechanism for protection against SA-AKI.
Our experiment has some limitations. For example, we did not further verify how MSCs affect the Gal-9/Tim-3 pathway. In the future, using in vitro experiments, we will further verify whether MSCs affect the Gal-9/Tim-3 pathway through the secretion of Gal-9 or through other mechanisms.