GPR120 induces regulatory dendritic cells by inhibiting HK2-dependent glycolysis to alleviate fulminant hepatic failure.

Fulminant hepatic failure (FHF) is a potentially fatal liver disease that is associated with intrahepatic infiltration of inflammatory cells. As the receptor of polyunsaturated long chain fatty acids, GPR120 can regulate cell differentiation, proliferation, metabolism, and immune response. However, whether GPR120 is involved in FHF remains unknown. Using Propionibacterium acnes (P. acnes)-primed, LPS-induced FHF in mice, we found that interference with GPR120 activity using pharmacological agonist attenuated the severity of the liver injury and mortality of FHF in mice, while a lack of GPR120 exacerbated the disease. GPR120 activation potently alleviated FHF and led to decreased T helper (Th) 1 cell response and expansion of regulatory T cells (Tregs). Interestingly, GPR120 agonist didn't directly target T cells, but dramatically induced a distinct population of CD11c+MHC IIlowCD80lowCD86low regulatory DCs in the livers of FHF mice. GPR120 was found to restrict HIF-1α-dependent glycolysis. The augmented HIF-1α stabilization caused by GPR120 antagonism or deletion could be attenuated by the inhibition of ERK or by the activation of AMPK. Through the analysis of the clinical FHF, we further confirmed the activation of GPR120 was negatively associated with the severity in patients. Our findings indicated that GPR120 activation has therapeutic potential in FHF. Strategies to target GPR120 using agonists or free fatty acids (FFAs) may represent a novel approach to FHF treatment.

miR-99a regulates CD4+ T cell differentiation and attenuates experimental autoimmune encephalomyelitis by mTOR-mediated glycolysis.

Multiple microRNAs exhibit diverse functions to regulate inflammatory and autoimmune diseases. MicroRNA-99a (miR-99a) has been shown to be involved in adipose tissue inflammation and to be downregulated in the inflammatory lesions of autoimmune diseases rheumatoid arthritis and systemic lupus erythematosus. In this study, we found that miR-99a was downregulated in CD4+ T cells from experimental autoimmune encephalomyelitis (EAE) mice, an animal model of multiple sclerosis. Overexpression of miR-99a alleviated EAE development by promoting regulator T cells and inhibiting T helper type 1 (Th1) cell differentiation. Bioinformatics and functional analyses further revealed that the anti-inflammatory effects of miR-99a was attributable to its role in negatively regulating glycolysis reprogramming of CD4+ T cells by targeting the mTOR pathway. Additionally, miR-99a expression was induced by transforming growth factor ß (TGF-ß) to regulate CD4+ T cell glycolysis and differentiation. Taken together, our results characterize a pivotal role of miR-99a in regulating CD4+ T cell differentiation and glycolysis reprogramming during EAE development, which may indicate that miR-99a is a promising therapeutic target for the amelioration of multiple sclerosis and possibly other autoimmune diseases.

Inhibition of Dot1L Alleviates Fulminant Hepatitis Through Myeloid-Derived Suppressor Cells.

BACKGROUND & AIMS: Fulminant hepatitis (FH) is a clinical syndrome characterized by sudden and severe liver dysfunction. Dot1L, a histone methyltransferase, is implicated in various physiologic and pathologic processes, including transcription regulation and leukemia. However, the role of Dot1L in regulating inflammatory responses during FH remains elusive. METHODS: Propionibacterium acnes (P. acnes)-primed, lipopolysaccharides (LPS)-induced FH was established in C57BL/6 mice and was treated with the Dot1L inhibitor EPZ-5676. Myeloid derived suppressor cells (MDSCs) were depleted by anti-Gr-1 antibody to evaluate their therapeutic roles in Dot1L treatment of FH. Moreover, peripheral blood of patients suffered with FH and healthy controls was collected to determine the expression profile of Dot1L-SOCS1-iNOS axis in their MDSCs. RESULTS: Here we identified that EPZ-5676, pharmacological inhibitor of Dot1L, attenuated the liver injury of mice subjected to FH. Dot1L inhibition led to decreased T helper 1 cell response and expansion of regulatory T cells (Tregs) during FH. Interestingly, Dot1L inhibition didn't directly target T cells, but dramatically enhanced the immunosuppressive function of MDSCs. Mechanistically, Dot1L inhibition epigenetically suppressed SOCS1 expression, thus inducing inducible nitric oxide synthase (iNOS) expression in a STAT1-dependent manner. Moreover, in human samples, the levels of Dot1L and SOCS1 expression were upregulated in MDSCs, accompanied by decreased expression of iNOS in patients with FH, compared with healthy controls. CONCLUSIONS: Altogether, our findings established Dot1L as a critical regulator of MDSC immunosuppressive function for the first time, and highlighted the therapeutic potential of Dot1L inhibitor for FH treatment.

Inflammation-induced inhibition of chaperone-mediated autophagy maintains the immunosuppressive function of murine mesenchymal stromal cells.

Macroautophagy has been implicated in modulating the therapeutic function of mesenchymal stromal cells (MSCs). However, the biological function of chaperone-mediated autophagy (CMA) in MSCs remains elusive. Here, we found that CMA was inhibited in MSCs in response to the proinflammatory cytokines interferon-γ (IFN-γ) and tumor necrosis factor-α (TNF-α). In addition, suppression of CMA by knocking down the CMA-related lysosomal receptor lysosomal-associated membrane protein 2 (LAMP-2A) in MSCs significantly enhanced the immunosuppressive effect of MSCs on T cell proliferation, and as expected, LAMP-2A overexpression in MSCs exerted the opposite effect on T cell proliferation. This effect of CMA on the immunosuppressive function of MSCs was attributed to its negative regulation of the expression of chemokine C-X-C motif ligand 10 (CXCL10), which recruits inflammatory cells, especially T cells, to MSCs, and inducible nitric oxide synthase (iNOS), which leads to the subsequent inhibition of T cell proliferation via nitric oxide (NO). Mechanistically, CMA inhibition dramatically promoted IFN-γ plus TNF-α-induced activation of NF-κB and STAT1, leading to the enhanced expression of CXCL10 and iNOS in MSCs. Furthermore, we found that IFN-γ plus TNF-α-induced AKT activation contributed to CMA inhibition in MSCs. More interestingly, CMA-deficient MSCs exhibited improved therapeutic efficacy in inflammatory liver injury. Taken together, our findings established CMA inhibition as a critical contributor to the immunosuppressive function of MSCs induced by inflammatory cytokines and highlighted a previously unknown function of CMA.

Graft-versus-host disease depletes plasmacytoid dendritic cell progenitors to impair tolerance induction.

Graft-versus-host disease (GVHD) causes failed reconstitution of donor plasmacytoid dendritic cells (pDCs) that are critical for immune protection and tolerance. We used both murine and human systems to uncover the mechanisms whereby GVHD induces donor pDC defects. GVHD depleted Flt3-expressing donor multipotent progenitors (MPPs) that sustained pDCs, leading to impaired generation of pDCs. MPP loss was associated with decreased amounts of MPP-producing hematopoietic stem cells (HSCs) and oxidative stress-induced death of proliferating MPPs. Additionally, alloreactive T cells produced GM-CSF to inhibit MPP expression of Tcf4, the transcription factor essential for pDC development, subverting MPP production of pDCs. GM-CSF did not affect the maturation of pDC precursors. Notably, enhanced recovery of donor pDCs upon adoptive transfer early after allogeneic HSC transplantation repressed GVHD and restored the de novo generation of donor pDCs in recipient mice. pDCs suppressed the proliferation and expansion of activated autologous T cells via a type I IFN signaling-dependent mechanism. They also produced PD-L1 and LILRB4 to inhibit T cell production of IFN-γ. We thus demonstrate that GVHD impairs the reconstitution of tolerogenic donor pDCs by depleting DC progenitors rather than by preventing pDC maturation. MPPs are an important target to effectively bolster pDC reconstitution for controlling GVHD.

Inhibition of EZH2 ameliorates bacteria-induced liver injury by repressing RUNX1 in dendritic cells.

Fulminant hepatic failure (FHF) is a clinical syndrome characterized by a sudden and severe impairment in liver function. However, the precise mechanism of immune dysregulation that is significant to FHF pathogenesis remains unclear. Enhancer of zeste homolog 2 (EZH2) has been implicated in inflammation as a regulator of immune cell function. In this study, we investigated the role of EZH2 in an animal model of human FHF induced by Propionibacterium acnes (P. acnes) and lipopolysaccharide (LPS). We demonstrated that EZH2 depletion in dendritic cells (DCs) and pharmacological inhibition of EZH2 using GSK126 both significantly ameliorated liver injury and improved the survival rates of mice with P. acnes plus LPS-induced FHF, which could be attributed to the decreased infiltration and activation of CD4+ T cells in the liver, inhibition of T helper 1 cells and induction of regulatory T cells. The expression of EZH2 in DCs was increased after P. acnes administration, and EZH2 deficiency in DCs suppressed DC maturation and prevented DCs from efficiently stimulating CD4+ T-cell proliferation. Further mechanistic analyses indicated that EZH2 deficiency directly increased the expression of the transcription factor RUNX1 and thereby suppressed the immune functions of DCs. The functional dependence of EZH2 on RUNX1 was further illustrated in DC-specific Ezh2-deficient mice. Taken together, our findings establish that EZH2 exhibits anti-inflammatory effects through inhibition of RUNX1 to regulate DC functions and that inhibition of EZH2 alleviates P. acnes plus LPS-induced FHF, probably by inhibiting DC-induced adaptive immune responses. These results highlight the effect of EZH2 on DCs, serving as a guide for the development of a promising immunotherapeutic strategy for FHF.

Dendritic Cell Regulation of Graft-Vs.-Host Disease: Immunostimulation and Tolerance.

Graft-vs.-host disease (GVHD) remains a significant cause of morbidity and mortality after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Significant progresses have been made in defining the dichotomous role of dendritic cells (DCs) in the development of GVHD. Host-derived DCs are important to elicit allogeneic T cell responses, whereas certain donor-types of DCs derived from newly engrafted hematopoietic stem/progenitor cells (HSPCs) can amply this graft-vs.-host reaction. In contrast, some DCs also play non-redundant roles in mediating immune tolerance. They induce apoptotic deletion of host-reactive donor T cells while promoting expansion and function of regulatory T cells (Treg). Unfortunately, this tolerogenic effect of DCs is impaired during GVHD. Severe GVHD in patients subject to allo-HSCT is associated with significantly decreased number of circulating peripheral blood DCs during engraftment. Existing studies reveal that GVHD causes delayed reconstitution of donor DCs from engrafted HSPCs, impairs the antigen presentation function of newly generated DCs and reduces the capacity of DCs to regulate Treg. The present review will discuss the importance of DCs in alloimmunity and the mechanism underlying DC reconstitution after allo-HSCT.

Protective role of GPR120 in the maintenance of pregnancy by promoting decidualization via regulation of glucose metabolism.

BACKGROUND: Intake of ω-3 PUFAs have been demonstrated to have positive effects on pregnancy outcome, whose receptor, GPR120, regulates several cellular functions including differentiation, metabolism and immune reaction. However, whether GPR120 is involved in decidualization and pregnancy remains unknown. METHODS: Decidua tissue from women with normal pregnancy and spontaneous abortion were collected to determine the expression profile of GPR120. Abortion mouse models and artificially induced deciduoma in mice were established to evaluate the effect of GPR120 on pregnancy outcome and in vivo decidualization. HESCs and primary DSCs were used to explore the roles of GPR120 in decidualization and mechanisms involved. FINDINGS: We found that GPR120 functioned to promote decidualization by upregulating glucose uptake and pentose-phosphate pathway (PPP) of human endometrial stromal cells. Firstly, the expression of GPR120 in decidua of spontaneous abortion was downregulated compared to normal decidua. Lack of GPR120 predisposed mice to LPS or RU486 induced abortion. Decidualization was augmented by GPR120 via improving GLUT1-mediated glucose uptake and G6PD- mediated PPP. FOXO1 was upregulated by GPR120 via activation of ERK1/2 and AMPK signaling and increased the expression of GLUT1. Furthermore, the expression of chemokines and cytokines in decidual stromal cells was enhanced by GPR120. Lastly, GPR120 agonist ameliorated LPS-induced abortion in the mice. INTERPRETATION: GPR120 plays significant roles in decidualization and the maintenance of pregnancy, which might be a potential target for diagnosis and treatment of spontaneous abortion. FUND: Ministry of Science and Technology of China, National Natural Science Foundation of China, the Program of Science and Technology Commission of Shanghai Municipality.

Reduced production of laminin by hepatic stellate cells contributes to impairment in oval cell response to liver injury in aged mice.

Aged liver is usually impaired in response to hepatic injury. Tissue-specific stem cells participate in the repair of tissue injury. However, how oval cells (OCs) respond to injury and how the process is regulated by tissue microenvironment in aged mice have not been fully understood. In this study, taking advantage of well-established murine OC activation model, we demonstrated that OCs were less activated upon injury in aged mice and the impairment was mainly attributed to dysfunction in their niche. Through analyzing global gene expression, we found that the genes differentially expressed in damaged young and aged mouse liver tissues were predominantly those required for the formation and remodeling of extracellular matrix. As one of the most important extracellular matrix components in the OC niche, laminin was shown to promote the proliferation of OCs. Not surprisingly, laminin was downregulated with aging. Consistent with the downregulation of genes encoding DNA-dependent protein kinase (DNA-PK) proteins in aged hepatic stellate cells (HSCs), inhibition of DNA-PK also led to reduced expression of laminin in HSCs. Moreover, impairment in OC activation caused by less supporting from DNA-damaged HSCs could be rescued by laminin. This study reveals a new cellular mechanism underlying impaired OCs functionality during aging.

The deubiquitinating enzyme UCHL1 negatively regulates the immunosuppressive capacity and survival of multipotent mesenchymal stromal cells.

It is known that proinflammatory cytokines empower multipotent mesenchymal stromal cells (MSCs) the immunosuppressive capacity to treat various inflammatory diseases. Nevertheless, how the proinflammatory cytokines modulate the immunosuppressive capacity of MSCs is poorly understood. In the present study, we identified that the deubiquitinating enzyme ubiquitin C-terminal hydrolase 1 (UCHL1) was upregulated in MSCs upon stimulation of proinflammatory cytokines IFN-γ plus TNF-α. Interestingly, through intervening UCHL1 by shRNA knockdown or its inhibitor LDN57444 or overexpression, we found that UCHL1 played a critical role in suppressing cytokines-induced inducible nitric oxide synthase expression in murine MSCs and indoleamine 2,3-dioxygenase expression in human MSCs, thereby restrained their immunosuppressive capacity. This effect of UCHL1 was attributed to the negative role in regulating NF-κB and STAT1 signaling, as exhibited by promoting NF-κB and STAT1 activation upon inhibition of UCHL1. Besides, inhibition of UCHL1 suppressed cytokines-induced MSC apoptosis via upregulation of Bcl-2. As a consequence, UCHL1-inhibited MSCs effectively alleviated concanavalin A-induced inflammatory liver injury. Therefore, our study demonstrates a novel role of UCHL1 in regulating the immunosuppressive capacity and survival of MSCs, which further affects their immunotherapy for inflammatory diseases.

Hsp90 inhibition destabilizes Ezh2 protein in alloreactive T cells and reduces graft-versus-host disease in mice.

Modulating T-cell alloreactivity has been a main strategy to reduce graft-versus-host disease (GVHD), a life-threatening complication after allogeneic hematopoietic stem-cell transplantation (HSCT). Genetic deletion of T-cell Ezh2, which catalyzes trimethylation of histone H3 at lysine 27 (H3K27me3), inhibits GVHD. Therefore, reducing Ezh2-mediated H3K27me3 is thought to be essential for inhibiting GVHD. We tested this hypothesis in mouse GVHD models. Unexpectedly, administration of the Ezh2 inhibitor GSK126, which specifically decreases H3K27me3 without affecting Ezh2 protein, failed to prevent the disease. In contrast, destabilizing T-cell Ezh2 protein by inhibiting Hsp90 using its specific inhibitor AUY922 reduced GVHD in mice undergoing allogeneic HSCT. In vivo administration of AUY922 selectively induced apoptosis of activated T cells and decreased the production of effector cells producing interferon γ and tumor necrosis factor α, similar to genetic deletion of Ezh2. Introduction of Ezh2 into alloreactive T cells restored their expansion and production of effector cytokines upon AUY922 treatment, suggesting that impaired T-cell alloreactivity by inhibiting Hsp90 is achieved mainly through depleting Ezh2. Mechanistic analysis revealed that the enzymatic SET domain of Ezh2 directly interacted with Hsp90 to prevent Ezh2 from rapid degradation in activated T cells. Importantly, pharmacological inhibition of Hsp90 preserved antileukemia activity of donor T cells, leading to improved overall survival of recipient mice after allogeneic HSCT. Our findings identify the Ezh2-Hsp90 interaction as a previously unrecognized mechanism essential for T-cell responses and an effective target for controlling GVHD.

The combination of aricept with a traditional Chinese medicine formula, smart soup, may be a novel way to treat Alzheimer's disease.

Alzheimer's disease (AD) is a common neurodegenerative disease affecting cognitive function in the elderly, which is characterized by the presence of extracellular deposits of insoluble amyloid-ß plaques and neuronal loss. Modern pharmacology and drug development usually follow a single-target principle, which might contribute to the failure of most compounds in clinical trials against AD. Considering AD is a multifactorial disease, a combination therapeutic strategy that applies drugs with different mechanisms would be an alternative way. Smart Soup (SS), a Traditional Chinese Medicine formula, is composed of three herbaceous plants and has been applied in the treatment of amnesia in China for hundreds of years. In this work, we studied the clinical potency of the combination of SS and Aricept in AD therapy. In the in vivo model, both longevity and locomotive activity of AD transgenic Drosophila were improved remarkably in the combined medicine treated group. We also observed less amyloid-ß deposition and retarded neuronal loss following the combined drug treatment. In the retrospective cohort study, we found the combination therapy exerted better therapeutic effect on AD patients. Our study revealed that combination therapy with multiple drug targets did have a better therapeutic outcome. It provides a new strategy to develop an optimum pharmaceutical approach against AD.