The Effects of Adoptively Transferred IL-23/IL-18-Polarized Neutrophils on Tumor and Collagen-Induced Arthritis in Mice

Abstract

Background: Neutrophils present great diverse phenotypes in various microenvironments and play different immune regulatory functions. Neutrophils generally classified into inflammatory phenotype N1 and anti-informatory phenotype N2. Our recent studies showed that IL-23 alone stimulated neutrophils to express IL-17A, IL-17F and IL-22 and displayed a gene transcriptional profile similar to Th17 cells. In the present study, we tried to identify potential cytokines to promote IL-23-induced neutrophil polarization. Methods: Mouse bone marrow-derived neutrophils and human peripheral blood neutrophils were treated with IL-23 (10 ng/mL) plus IL-18 (25 ng/mL) to induce Th17-like subset in vitro and detected by real-time PCR, flow cytometry, ELISA, immunofluorescence and RNA-seq assays. In vivo, collagen-induced arthritis (CIA) mouse model and EL4 tumorbearing mouse model were used to characterize the potential roles of N(IL-23+IL-18) in inflammation and tumor. Results: Real-time PCR, ELISA and flow cytometry assays showed that IL-18 could significantly enhance IL-23-induced IL-17A, IL-17F and IL-22 expressions in mouse and human neutrophils in a synergistic way, although IL-18 alone failed to induce these cytokines expression. RNA-seq and molecular studies showed that the polarization of N(IL-23+IL-18) is mainly mediated by the JNK/p38-STAT3-BATF signaling pathway. Adoptive transfer of the induced N(IL-23+IL-18) neutrophils significantly accelerated the tumor growth in EL4 tumor-bearing mice and enhanced disease progression in the CIA mouse model. IL-17Adeficient N(IL-23+IL-18) neutrophils failed to enhance the CIA pathogenesis in this model, suggesting that IL-17A may be involved in the N(IL-23+IL-18) neutrophils-promoted arthritis in mice. Conclusion:The Th17-type subpopulation N(IL-23+IL-18) has pro-tumor and proinflammatory properties. Recognizing the different functional polarization of neutrophils would significantly help us to understand the distinctive protective/pathological roles of neutrophils in physiological and different pathological situations.

Trappc1 deficiency impairs thymic epithelial cell development by breaking endoplasmic reticulum homeostasis

“…Next, cDNA was reversely transcribed with the SuperScript III Reverse Transcriptase Kit (Invitrogen, 18080–093). A quantitative PCR of target genes was performed using SYBR Premix Ex TaqTM (TaKaRa, RR420) on a CFX96 apparatus (Bio‐Rad Laboratories) [99, 100]. For the RT‐qPCR analysis, the expression of target genes was normalized by Hprt expression using the 2 –ΔΔCt methods.…”

Sulforaphene Inhibits Periodontitis through Regulating Macrophage Polarization via Upregulating Dendritic Cell Immunoreceptor

“…Moreover, some studies have reported the crucial role of JNK and P38 phosphorylation in macrophage polarization. , Taking this into consideration, we decided to focus on the effect of SFE on JNK and P38 phosphorylation for further investigation. In order to further determine whether the suppression of M1 macrophage polarization by SFE is mediated by the inhibition of JNK and P38 phosphorylation, the p-JNK activator AM (10 nM) or the p-P38 activator DHC (1 μM) was added at the same time when RAW 264.7 cells were pretreated with SFE (5 μM). , Figure B,C shows that the coprecipitation of DHC and SFE partially reversed the suppressive effect of SFE on the expression of genes and proteins (IL-1β and INOS) related to M1 macrophage polarization. Similarly, the p-JNK activator AM also partly reversed SFE’s inhibitory impact on gene and protein expression associated with M1 macrophage polarization (Figure D,E).…”

“…In order to further determine whether the suppression of M1 macrophage polarization by SFE is mediated by the inhibition of JNK and P38 phosphorylation, the p-JNK activator AM (10 nM) or the p-P38 activator DHC (1 μM) was added at the same time when RAW 264.7 cells were pretreated with SFE (5 μM). 31,32 Figure 3B,C shows that the coprecipitation of DHC and SFE partially reversed the suppressive effect of SFE on the expression of genes and proteins (IL-1β and INOS) related to M1 macrophage polarization. Similarly, the p-JNK activator AM also partly…”

Tuberous Sclerosis Complex 1 Deficiency in Macrophages Promotes Unclassical Inflammatory Response to Lipopolysaccharide In Vitro and Dextran Sodium Sulfate-Induced Colitis in Mice

“…Cell culture supernatants were examined for IL-17A with Mouse IL-17A ELISA Kits (BioLegend, USA) according to the manufacturer’s instructions [ 35 ]. The 96-well plate coated with diluted capture antibody solution was prepared in advance on day 1.…”