Impact of cannabidiol treatment on regulatory T-17 cells and neutrophil polarization in acute kidney injury

Abstract

Hallmark features of acute kidney injury (AKI) include mobilization of immune and inflammatory mechanisms culminating in tissue injury. Emerging information indicates heterogeneity of neutrophils with pro- and anti-inflammatory functions (N1 and N2, respectively). Also, regulatory T-17 (Treg17) cells curtail T helper 17 (Th-17)-mediated proinflammatory responses. However, the status of Treg17 cells and neutrophil phenotypes in AKI are not established. Furthermore, cannabidiol exerts immunoregulatory effects, but its impact on Treg17 cells and neutrophil subtypes is not established. Thus, we examined the status of Treg17 cells and neutrophil subtypes in AKI and determined whether cannabidiol favors regulatory neutrophils and T cells accompanied with renoprotection. Accordingly, mice were subjected to bilateral renal ischemia-reperfusion injury (IRI), without or with cannabidiol treatment; thereafter, kidneys were processed for flow cytometry analyses. Renal IRI increased N1 and Th-17 but reduced N2 and Treg17 cells accompanied with disruption of mitochondrial membrane potential (ψ) and increased apoptosis/necrosis and kidney injury molecule-1 (KIM-1) immunostaining compared with their sham controls. Importantly, cannabidiol treatment preserved ψ and reduced cell death and KIM-1 accompanied by restoration of N1 and N2 imbalance and preservation of Treg17 cells while decreasing Th-17 cells. The ability of cannabidiol to favor development of Treg17 cells was further established using functional mixed lymphocytic reaction. Subsequent studies showed higher renal blood flow and reduced serum creatinine in cannabidiol-treated IRI animals. Collectively, our novel observations establish that renal IRI causes neutrophil polarization in favor of N1 and also reduces Treg17 cells in favor of Th-17, effects that are reversed by cannabidiol treatment accompanied with significant renoprotection.

Glucocorticoid-Induced Leucine Zipper Promotes Neutrophil and T-Cell Polarization with Protective Effects in Acute Kidney Injury

“…Flow Cytometry. Single-cell suspension was prepared for each mouse kidney and subjected to flow cytometry-based assays (Baban et al, 2009(Baban et al, , 2012(Baban et al, , 2018 to identify and analyze the numbers of total and subtypes of neutrophils and T cells as follows: neutrophils were initially identified as Ly-6G 1 /CD11b 1 cells and were presented as a percentage of whole kidney cells. Neutrophils were further identified as N1 (proinflammatory neutrophils) and N2 (regulatory neutrophils) using combinations of surface and intracellular markers, including Ly-6G 1 /CD11b 1 and tumor necrosis factor-a (TNF-a) for N1s and Ly-6G 1 /CD11b 1 /CD206 1 , and interleukin (IL)-10, for N2s.…”

“…The assessment of c m used the JC-1 assay, whereas apoptotic/ necrotic cell death in whole kidney cell preparations was determined using 7-aminoactinomycin D/caspase-3 assay as described previously (Baban et al, 2012(Baban et al, , 2018.…”

“…Neutrophils are recruited into the kidney very early post-IRI (Lauriat and Linas, 1998;Bolisetty and Agarwal, 2009). Recent studies Baban et al, 2018) indicate the capacity of neutrophils to assume proinflammatory and antiinflammatory functional phenotypes [i.e., type 1 neutrophil (N1) and type 2 neutrophil (N2), respectively], depending on microenvironment cues and via the expression of specific cell markers. Similarly, T cells can undergo polarization to their suppressive/ regulatory phenotypes, regulatory T cells (Tregs), which curtail effector T lymphocytes (Teffs) (Fan and Morand, 2012;Makris and Spanou, 2016).…”

Immune Responses Regulated by Cannabidiol

“…86 A confirmation of in vivo induced Tregs by CBD was noted in an ischemia-reperfusion injury model in the kidney, in which CBD returned the disease-induced reduction in CD3 + Foxp3 + cells to baseline. 152 Interestingly, in the ischemia-reperfusion kidney model, CBD also induced ''TReg17 cells,'' which were defined as CD3 + Foxp3 + CCR6 + STAT3 + . 152 It has been suggested that Treg17 cells help control a TH17 response.…”

“…152 Interestingly, in the ischemia-reperfusion kidney model, CBD also induced ''TReg17 cells,'' which were defined as CD3 + Foxp3 + CCR6 + STAT3 + . 152 It has been suggested that Treg17 cells help control a TH17 response. In vitro, CBD (5 lM) induced a CD69 + LAG + population in CD4 + CD25 À cells, which were identified as one type of regulatory cell, and induced Il10 mRNA expression.…”