T Helper Cell Lineage-Defining Transcription Factors: Potent Targets for Specific GVHD Therapy?

Allogenic hematopoietic stem cell transplantation (allo-HSCT) represents a potent and potentially curative treatment for many hematopoietic malignancies and hematologic disorders in adults and children. The donor-derived immunity, elicited by the stem cell transplant, can prevent disease relapse but is also responsible for the induction of graft-versus-host disease (GVHD). The pathophysiology of acute GVHD is not completely understood yet. In general, acute GVHD is driven by the inflammatory and cytotoxic effect of alloreactive donor T cells. Since several experimental approaches indicate that CD4 T cells play an important role in initiation and progression of acute GVHD, the contribution of the different CD4 T helper (Th) cell subtypes in the pathomechanism and regulation of the disease is a central point of current research. Th lineages derive from naïve CD4 T cell progenitors and lineage commitment is initiated by the surrounding cytokine milieu and subsequent changes in the transcription factor (TF) profile. Each T cell subtype has its own effector characteristics, immunologic function, and lineage specific cytokine profile, leading to the association with different immune responses and diseases. Acute GVHD is thought to be mainly driven by the Th1/Th17 axis, whereas Treg cells are attributed to attenuate GVHD effects. As the differentiation of each Th subset highly depends on the specific composition of activating and repressing TFs, these present a potent target to alter the Th cell landscape towards a GVHD-ameliorating direction, e.g. by inhibiting Th1 and Th17 differentiation. The finding, that targeting of Th1 and Th17 differentiation appears more effective for GVHD-prevention than a strategy to inhibit Th1 and Th17 cytokines supports this concept. In this review, we shed light on the current advances of potent TF inhibitors to alter Th cell differentiation and consecutively attenuate GVHD. We will focus especially on preclinical studies and outcomes of TF inhibition in murine GVHD models. Finally, we will point out the possible impact of a Th cell subset-specific immune modulation in context of GVHD.

The Tyrosine Kinase Tec Regulates Effector Th17 Differentiation, Pathogenicity, and Plasticity in T-Cell-Driven Intestinal Inflammation

T helper (Th) 17 cells are not only key in controlling infections mediated by extracellular bacteria and fungi but are also triggering autoimmune responses. Th17 cells comprise heterogeneous subsets, some with pathogenic functions. They can cease to secrete their hallmark cytokine IL-17A and even convert to other T helper lineages, a process known as transdifferentiation relying on plasticity. Both pathogenicity and plasticity are tightly linked to IL-23 signaling. Here, we show that the protein tyrosine kinase Tec is highly induced in Th17 cells. Th17 differentiation was enhanced at low interleukin-6 (IL-6) concentrations in absence of Tec, which correlates with increased STAT3 phosphorylation and higher Il23r expression. Therefore, we uncovered a function for Tec in the IL-6 sensing via STAT3 by CD4+ T cells, defining Tec as a fine-tuning negative regulator of Th17 differentiation. Subsequently, by using the IL-17A fate mapping mouse combined with in vivo adoptive transfer models, we demonstrated that Tec not only restrained effector Th17 differentiation but also pathogenicity and plasticity in a T-cell intrinsic manner. Our data further suggest that Tec regulates inflammatory Th17-driven immune responses directly impacting disease severity in a T-cell-driven colitis model. Notably, consistent with the in vitro findings, elevated levels of the IL-23 receptor (IL-23R) were observed on intestinal pre- and postconversion Th17 cells isolated from diseased Tec−/− mice subjected to adoptive transfer colitis, highlighting a fundamental role of Tec in restraining IL-23R expression, likely via the IL-6-STAT3 signaling axis. Taken together, these findings identify Tec as a negative regulator of Th17 differentiation, pathogenicity, and plasticity, contributing to the mechanisms which help T cells to orchestrate optimal immune protection and to restrain immunopathology.

Expressed Sequence Tags-1-Targeted MiRNA-326 in Gastric Cancer Induces Local Immune Inflammatory Microenvironment by Inducing T Helper Cell 17 Cell Differentiation

This study intends to assess miRNA-326’s effect on the immune-inflammatory microenvironment and its mechanism in gastric cancer (GC). GC adjacent tissues and tumor tissues were collected to analyze inflammatory factors by immunohistochemistry and ELISA, Est-1 and miRNA-326 level by Western blot or PCR, Th17 cells by flow cytometry. CD4+ T cells were transfected with Est-1 inhibitor, Est-1 mimics, or miR-326 mimics followed by measurement of Th17 differentiation-related genes via gene chips and inflammatory factor release. Inflammatory factors in serum of GC patients were significantly increased and miR-326 was upregulated with decreased Est-1 and unbalanced Th17/Treg cell ratio. miR-326 targeted Est-1 to inhibit its expression. After transfection with Est-1 inhibitor, Th17 differentiation-related genes were upregulated. After transfection with miR-326 mimics, Est-1 level was reduced and inflammation was enhanced with maturation of Th17 cells. In conclusion, miRNA-326 induces Th17 cell differentiation by targeting Est-1, thereby promoting the release of inflammatory factors and inducing immune inflammatory microenvironment.

Increased HIF-1α Expression in T Cells and Associated with Enhanced Th17 Pathway in Systemic Lupus Erythematosus

Recent emerging evidences indicate that dysfunction of metabolic remodeling underlies aberrant T cell immune responses in systemic lupus erythematosus (SLE). However, how these aberrant immune activation and metabolic dysfunction interact in lupus patients are not fully understood. This study was undertaken to investigate the expression of HIF-1α, a regulator of metabolic reprogramming, in T cells from SLE. Our results demonstrated that HIF-1α expression is increased in CD4 T cells from SLE patients both in intracellular staining and quantitative real-time PCR analysis. In addition, there is enhanced HIF-1α expression in Th17-skewing murine T cells, and lentivirus-mediated HIF-1α overexpression promotes Th17 differentiation. Moreover, HIF-1α gene expression is positively correlated with the expression of glycolysis- and IL-17-associated genes in SLE patients. These results indicate that HIF-1α expression is increased in T cells from SLE patients, and is associated with enhanced Th17 pathway, implicating HIF-1α contributes to the activation of Th17 cells in SLE, and represents a potential novel therapeutic target.

Clinical efficacy and target signaling pathways of natural products in psoriasis treatment: a systematic review

Background: Psoriasis is a common chronic inflammatory skin disorder, which has adverse effects on patients’ quality of life. Natural products exhibit significant therapeutic capacities with small side effects and might be preferable alternative treatments for patients with psoriasis. Aim of the review: This study summarizes the clinical efficacy of natural products and signaling pathways with the potential targets for psoriasis treatment. Methods: The literature for this article was acquired from PubMed and Web of Science, from Jan 2010 to Dec 2020. The keywords for searching included “psoriasis” and “natural product”, “herbal medicine”, “herbal therapy”, “medicinal plant”, “medicinal herb”, or “pharmaceutical plant”. Results: The anti-psoriatic effect of natural products in clinical studies was summarized. Herbal extracts, natural compounds, and herbal prescriptions could regulate the signaling pathways to alleviate psoriasis symptoms, such as Th17 differentiation, JAK/STAT, NF-κB, MAPK, PI3K/Akt/mTOR, and other signaling pathways, which are involved in the inflammatory response and keratinocyte hyperproliferation. Conclusion: Natural products exerted the anti-psoriatic effect by targeting multiple signaling pathways, providing evidence for the investigation of novel drugs. Further experimental research should be performed to screen and characterize the therapeutic targets of natural products for application in psoriasis treatment.

Overexpressing Kallistatin Aggravates Experimental Autoimmune Uveitis Through Promoting Th17 Differentiation

Kallistatin or kallikrein-binding protein (KBP) has been reported to regulate angiogenesis, inflammation and tumor progression. Autoimmune uveitis is a common, sight-threatening inflammatory intraocular disease. However, the roles of kallistatin in autoimmunity and autoreactive T cells are poorly investigated. Compared to non-uveitis controls, we found that plasma levels of kallistatin were significantly upregulated in patients with Vogt-Koyanagi-Harada (VKH) disease, one of the non-infectious uveitis. Using an experimental autoimmune uveitis (EAU) model induced by human interphotoreceptor retinoid-binding protein peptide 651-670 (hIRBP651-670), we examined the effects of kallistatin on the pathogenesis of autoimmune diseases. Compared to wild type (WT) mice, kallistatin transgenic (KS) mice developed severe uveitis with dominant Th17 infiltrates in the eye. In addition, the proliferative antigen-specific T cells isolated from KS EAU mice produced increased levels of IL-17A, but not IFN-γ or IL-10 cytokines. Moreover, splenic CD4+ T cells from naïve KS mice expressed higher levels of Il17a mRNA compared to WT naïve mice. Under Th17 polarization conditions, KS mice exhibited enhanced differentiation of naïve CD4+ T cells into Th17 cells compared to WT controls. Together, our results indicate that kallistatin promotes Th17 differentiation and is a key regulator of aggravating autoinflammation in EAU. Targeting kallistatin might be a potential to treat autoimmune disease.

Inhibition of TGF-β Signaling Suppresses Th17 Differentiation and Promotes Treg Numbers but does not Reduces Experimental Arthritis

ObjectivesTGF-β is an important growth factor to promote the differentiation of T helper 17 (Th17) and regulatory T cells (Treg). The potential of TGF-β as therapeutic target in T cell-mediated diseases like rheumatoid arthritis (RA) is unclear. We investigated the effect of TGF-β inhibition on murine Th17 differentiation in vitro, on human RA synovial explants ex vivo, and on the development of experimental arthritis in vivo. MethodsMurine splenocytes were differentiated into Th17 cells, and the effect of the TGF-βRI inhibitor SB-505124 was studied. Synovial biopsies were cultured in the presence or absence of SB-505124. Experimental arthritis was induced in C57Bl6 mice and treated daily with SB-505124. FACS analysis was performed to measure different T cell subsets. Histological sections were analysed to determine joint inflammation and destruction.ResultsSB-505124 potently reduced murine Th17 differentiation by decreasing Il7a and Rorc gene expression and IL-17 protein production. SB-505124 significantly suppressed IL-6 production by synovial explants. In vivo, SB-505124 reduced Th17 levels, while increased levels of Tregs were observed. Despite this skewed Th17/Treg balance, SB-505124 treatment did not result in suppression of joint inflammation and destruction.ConclusionsBlocking TGF-β signalling suppresses Th17 differentiation and improves the Th17/Treg balance. However, SB-505124 treatment does not suppress experimental arthritis.

KLF4 Promotes Diabetic Chronic Wound Healing by Suppressing Th17 Cell Differentiation in an MDSC-Dependent Manner

Objectives. Diabetic wound inflammation deficiencies lead to ulcer development and eventual amputation and disability. Our previous research demonstrates that myeloid-derived suppressor cells (MDSCs) accumulate during inflammation and promote chronic wound healing via the regulation of Kruppel-like factor 4 (KLF4). In this study, we aimed to investigate the potential roles of MDSCs and KLF4 in diabetic wound healing. Methods. An ob/ob mouse pressure ulcer (PU) model was used to evaluate the process of wound healing. The expression levels of KLF4 and IL-17A were measured by real-time PCR, and the population of MDSCs and Th17 cells was measured by flow cytometry. The levels of cytokines were determined by an immunosuppression assay. Results. KLF4 deficiency in the diabetic PU model resulted in decreased accumulation of MDSCs, increased expansion of Th17 cells, and significantly delayed wound healing. Conversely, KLF4 activation by APTO-253 accelerated wound healing accompanied by increased MDSC populations and decreased numbers of Th17 cells. MDSCs have been proven to mediate Th17 differentiation via cytokines, and our in vitro data showed that elevated KLF4 expression in MDSCs resulted in reduced Th17 cell numbers and, thus, decreased levels of cytokines indispensable for Th17 differentiation. Conclusions. Our study revealed a previously unreported function of KLF4-regulated MDSCs in diabetic wound healing and identified APTO-253 as a potential agent to improve the healing of pressure ulcers.

Inhibition of TGF-β Signaling Suppresses Th17 Differentiation and Promotes Treg Numbers but Does Not Reduce Experimental Arthritis.

ObjectivesTGF-β is an important growth factor to promote the differentiation of T helper 17 (Th17) as well as regulatory T cells (Treg). Due to its dual role, the potential of TGF-β as therapeutic target in T cell-mediated diseases like rheumatoid arthritis (RA) is unclear. In this study, we investigated the effect of TGF-β inhibition on murine Th17 differentiation in vitro, on human RA synovial explants ex vivo, and on the development of experimental arthritis in vivo. MethodsMurine splenocytes were differentiated into Th17 cells, and the effect of the TGF-βRI inhibitor SB-505124 on Th17 differentiation was studied. RA synovial biopsies were cultured for 24h in the presence or absence of SB-505124. Experimental arthritis models were induced in C57Bl6 mice, and were treated daily with SB-505124. FACS analysis was performed to measure different T cell subsets. Histological sections were analysed to determine joint inflammation and destruction.ResultsSB-505124 potently reduced murine Th17 differentiation by decreasing Il7a and Rorc gene expression and IL-17 protein production. SB-505124 significantly suppressed IL-6 production by RA synovial explants. In the Th17-driven arthritis model, SB-505124 reduced Th17 levels, while increased levels of Tregs were observed. Despite this skewed Th17/Treg balance, SB-505124 treatment did not result in suppression of joint inflammation and destruction in this model.ConclusionsBlocking TGF-β signalling suppresses Th17 differentiation and improves the Th17/Treg balance. However, SB-505124 treatment does not suppress experimental arthritis, and is therefore not an adequate way to target Th17-driven inflammation.

Abstract MP05: The Mechanism Of The Pvat Pro-inflammatory Micro-environment Formation During The Development Of High Fat Diet-induced Hypertension

There is considerable evidence for a causative role for T cells in hypertension, including studies with immunosuppressive drugs and T cell-deficient models. Our previous studies showed that soluble mediators from mesenteric perivascular adipose tissue (mPVAT) modulate T cell function. Specifically, conditioned media from mPVAT (mPVAT-CM) from Dahl S rats on a high fat diet (HFD) promoted expression of the pro-inflammatory cytokines, IFNg, IL-17a and GM-CSF, by activated T cells. Furthermore, the Dahl S rats on HFD will later develop hypertension. Hypothesis: mPVAT is stimulated to produce immunomodulatory mediators that promotes Th1/17 differentiation preceding the development of HFD-induced hypertension. We conducted bulk RNA-seq on activated splenocytes cultured in mPVAT-CM from Dahl S rats on either control or HFD for 10 weeks. In accordance with our previous studies, PVAT-CM from HFD-fed rats significantly upregulated many genes associated with IFNg/IL-17 induction, including Mpeg1, Lyz2 and Tnfsf4 (5.0±1.78, 3.70±0.53 and 1.78±0.42 fold over Control diet, respectively). In contrast, Th2/Treg-associated genes, such as Ctla2a (-0.27±0.02) and Ccr4 (-0.41±0.03) were downregulated. We also performed single cell (sc) RNA-seq on the PVAT stromal vascular fraction (SVF) and found that acute inflammatory genes were enriched in the HFD group. Together with the bulk RNA-seq on mPVAT, these data strongly suggest that the pro-inflammatory mPVAT micro-environment may promote Th1/Th17 differentiation. To identify mediators in PVAT-CM that may induce Th1/Th17 differentiation, we compared the bulk RNA-seq on splenocytes cultured in PVAT-CM with bulk RNA-seq conducted on the whole mPVAT itself. We found that a T cell co-stimulatory receptor DPP4 (CD26), which is closely associated with T cell activation was significantly increased in mPVAT from HFD-fed rats (33.4±2.3 HFD vs. 15.3±1.8 Control diet). We also observed an increase in DPP4 global expression from mPVAT SVF in HFD-fed rats, as determined by scRNA-seq. Conclusion: The data suggest that HFD promotes the IFNg and IL-17a pathways in PVAT, which precedes hypertension in Dahl S rats and correlates with an increase in expression of DPP-4, a gene that promotes T cell activation. (NIH P01 HL070687).