Ying Wang

Heterogeneity of natural Foxp3+ T cells: A committed regulatory T-cell lineage and an uncommitted minor population retaining plasticity

Natural regulatory T cells (Treg) represent a distinct lineage of T lymphocytes committed to suppressive functions, and expression of the transcription factor Foxp3 is thought to identify this lineage specifically. Here we report that, whereas the majority of natural CD4+Foxp3+ T cells maintain stable Foxp3 expression after adoptive transfer to lymphopenic or lymphoreplete recipients, a minor fraction enriched within the CD25− subset actually lose it. Some of those Foxp3− T cells adopt effector helper T cell (Th) functions, whereas some retain “memory” of previous Foxp3 expression, reacquiring Foxp3 upon activation. This minority “unstable” population exhibits flexible responses to cytokine signals, relying on transforming growth factor-β to maintain Foxp3 expression and responding to other cytokines by differentiating into effector Th in vitro. In contrast, CD4+Foxp3+CD25high T cells are resistant to such conversion to effector Th even after many rounds of cell division. These results demonstrate that natural Foxp3+ T cells are a heterogeneous population consisting of a committed Treg lineage and an uncommitted subpopulation with developmental plasticity.

Plasticity of mesenchymal stem cells in immunomodulation: pathological and therapeutic implications

Mesenchymal stem cells (MSCs) are multipotent stromal cells that exist in many tissues and are capable of differentiating into several different cell types. Exogenously administered MSCs migrate to damaged tissue sites, where they participate in tissue repair. Their communication with the inflammatory microenvironment is an essential part of this process. In recent years, much has been learned about the cellular and molecular mechanisms of the interaction between MSCs and various participants in inflammation. Depending on their type and intensity, inflammatory stimuli confer on MSCs the ability to suppress the immune response in some cases or to enhance it in others. Here we review the current findings on the immunoregulatory plasticity of MSCs in disease pathogenesis and therapy.

Th2 Lymphoproliferative Disorder of LatY136F Mutant Mice Unfolds Independently of TCR-MHC Engagement and Is Insensitive to the Action of Foxp3+ Regulatory T Cells

Mutant mice where tyrosine 136 of linker for activation of T cells (LAT) was replaced with a phenylalanine (LatY136F mice) develop a fast-onset lymphoproliferative disorder involving polyclonal CD4 T cells that produce massive amounts of Th2 cytokines and trigger severe inflammation and autoantibodies. We analyzed whether the LatY136F pathology constitutes a bona fide autoimmune disorder dependent on TCR specificity. Using adoptive transfer experiments, we demonstrated that the expansion and uncontrolled Th2-effector function of LatY136F CD4 cells are not triggered by an MHC class II-driven, autoreactive process. Using Foxp3EGFP reporter mice, we further showed that nonfunctional Foxp3+ regulatory T cells are present in LatY136F mice and that pathogenic LatY136F CD4 T cells were capable of escaping the control of infused wild-type Foxp3+ regulatory T cells. These results argue against a scenario where the LatY136F pathology is primarily due to a lack of functional Foxp3+ regulatory T cells and suggest that a defect intrinsic to LatY136F CD4 T cells leads to a state of TCR-independent hyperactivity. This abnormal status confers LatY136F CD4 T cells with the ability to trigger the production of Abs and of autoantibodies in a TCR-independent, quasi-mitogenic fashion. Therefore, despite the presence of autoantibodies causative of severe systemic disease, the pathological conditions observed in LatY136F mice unfold in an Ag-independent manner and thus do not qualify as a genuine autoimmune disorder.

An Osteopontin‐Integrin Interaction Plays a Critical Role in Directing Adipogenesis and Osteogenesis by Mesenchymal Stem Cells

An imbalance between normal adipogenesis and osteogenesis by mesenchymal stem cells (MSCs) has been shown to be related to various human metabolic diseases, such as obesity and osteoporosis; however, the underlying mechanisms remain elusive. We found that the interaction between osteopontin (OPN), an arginine‐glycine‐aspartate‐containing glycoprotein, and integrin αv/β1 plays a critical role in the lineage determination of MSCs. Although OPN is a well‐established marker during osteogenesis, its role in MSC differentiation is still unknown. Our study reveals that blockade of OPN function promoted robust adipogenic differentiation, while inhibiting osteogenic differentiation. Re‐expression of OPN restored a normal balance between adipogenesis and osteogenesis in OPN−/− MSCs. Retarded bone formation by OPN−/− MSCs was also verified by in vivo implantation with hydroxyapatite‐tricalcium phosphate, a bone‐forming matrix. The role of extracellular OPN in MSC differentiation was further demonstrated by supplementation and neutralization of OPN. Blocking well‐known OPN receptors integrin αv/β1 but not CD44 also affected MSC differentiation. Further studies revealed that OPN inhibits the C/EBPs signaling pathway through integrin αv/β1. Consistent with these in vitro results, OPN−/− mice had a higher fat to total body weight ratio than did wild‐type mice. Therefore, our study demonstrates a novel role for OPN‐integrin αv/β1 in regulating MSC differentiation. Stem Cells 2014;32:327–337

Loss of the LAT Adaptor Converts Antigen-Responsive T Cells into Pathogenic Effectors that Function Independently of the T Cell Receptor

Despite compromised T cell antigen receptor (TCR) signaling, mice in which tyrosine 136 of the adaptor linker for activation of T cells (LAT) was constitutively mutated (Lat(Y136F) mice) accumulate CD4(+) T cells that trigger autoimmunity and inflammation. Here we show that equipping postthymic CD4(+) T cells with LATY136F molecules or rendering them deficient in LAT molecules triggers a lymphoproliferative disorder dependent on prior TCR engagement. Therefore, such disorders required neither faulty thymic T cell maturation nor LATY136F molecules. Unexpectedly, in CD4(+) T cells recently deprived of LAT, the proximal triggering module of the TCR induced a spectrum of protein tyrosine phosphorylation that largely overlapped the one observed in the presence of LAT. The fact that such LAT-independent signals result in lymphoproliferative disorders with excessive cytokine production demonstrates that LAT constitutes a key negative regulator of the triggering module and of the LAT-independent branches of the TCR signaling cassette.

Anti-Inflammatory Properties and Regulatory Mechanism of a Novel Derivative of Artemisinin in Experimental Autoimmune Encephalomyelitis

Ethyl 2-[4-(12-β-artemisininoxy)]phenoxylpropionate (SM933) is a novel derivative of artemisinin, an herbal compound approved for the treatment of malaria. In this study, we show that SM933 has unique anti-inflammatory properties through regulation of signaling pathways, leading to amelioration of experimental autoimmune encephalomyelitis. The anti-inflammatory properties of SM933 were characterized by inhibition of encephalitogenic T cell responses that were altered to exhibit a Th2 immune deviation and reduced activity and concentration of NO and inducible NO synthase. The observed effect of SM933 was mediated through regulatory mechanisms involving the NFκB and the Rig-G/JAB1 signaling pathways. SM933 was found to inhibit the activity of NFκB by up-regulating IκB, which accounted for various down-stream anti-inflammatory actions. Furthermore, it up-regulated Rig-G through the action of IFN-α and prevented JAB1, a master cell cycle regulator, from entering the nucleus to promote p27 degradation, resulting in down-regulation of CDK2 and cyclin A and cell cycle progression. Regulation of the Rig-G/JAB1 pathway by SM933 led to altered cell cycle activity of encephalitogenic T cells as a result of its selective effect on activated, but not resting, T cells. The study indicates that SM933 is a novel anti-inflammatory agent acting through defined signaling mechanisms and provides regulatory mechanisms required for effective drug targeting in treatment of autoimmune disease and inflammation.

Methylation of α-synuclein and leucine-rich repeat kinase 2 in leukocyte DNA of Parkinson's disease patients.

BACKGROUND Recent studies highlight the role of DNA methylation in the pathogenesis of Parkinsons disease (PD). However, there is a paucity of studies exploring the role of blood-based DNA methylation in PD. We aimed to explore identifiable epigenetic biomarkers for PD by analyzing the methylation status of α-synuclein (SNCA) and leucine-rich repeat kinase 2 (LRRK2) in leukocytes. METHODS Bisulfite Specific PCR-based Sequencing method was used for semi-quantitative detection of methylation status of CpG islands in SNCA and LRRK2 promoter regions. Bisulfite Specific Cloning-based Sequencing method was used for further quantitative examination of CpG-2 methylation of SNCA. mRNA level was also detected in leukocytes. RESULTS Semi-quantitative detection showed that the methylation status of SNCA CpG-2 differed between PD patients and normal controls, while there was no difference in CpG-1 of SNCA or in LRRK2 promoter. Further quantitative analysis by clonal assay showed that the CpG-2 of SNCA was hypomethylated in PD patients compared with the normal control (5.90% versus 7.69%, P=0.034). Moreover, among the 14 CpG sites of CpG-2, the 2nd, 4th and 9th CpG sites were significantly hypomethylated in PD patients. In subgroups of PD, the methylation level decreased in the early-onset PD patients (P=0.001). RT-PCR examination showed that SNCA mRNA was increased in PD patients compared with normal control (P=0.003). CONCLUSIONS Our results indicated that the methylation level of SNCA CpG-2, especially that of the 2nd, 4th and 9th CpG sites in leukocytes might have great potential to be a useful and informative biomarker in PD diagnosis and treatment.

Tumour-associated mesenchymal stem/stromal cells: emerging therapeutic targets

Mesenchymal stem cells, also known as mesenchymal stromal cells (MSCs), exist in many tissues and are known to actively migrate to sites of tissue injury, where they participate in wound repair. Tumours can be considered “wounds that never heal” and, in response to cues from a tumour, MSCs are continuously recruited to and become integral components of the tumour microenvironment. Recently, it has become apparent that such tumour-associated MSCs (TA-MSCs) have an active role in tumour initiation, promotion, progression and metastasis. In this Review, we discuss recent advances in our understanding of the pathogenic role of TA-MSCs in regulating the survival, proliferation, migration and drug resistance of tumour cells, as well as the influence of MSCs on the immune status of the tumour microenvironment. Moreover, we discuss therapeutic approaches that target TA-MSC upstream or downstream modulators or use MSCs as vehicles for the delivery of tumoricidal agents. It is anticipated that new insights into the functions of TA-MSCs will lead to the development of novel therapeutic strategies against tumours.

Plasma metabolite profiles of Alzheimer's disease and mild cognitive impairment.

Previous studies have demonstrated altered metabolites in samples of Alzheimers disease (AD) patients. However, the sample size from many of them is relatively small and the metabolites are relatively limited. Here we applied a comprehensive platform using ultraperformance liquid chromatography-time-of-flight mass spectrometry and gas chromatography-time-of-flight mass spectrometry to analyze plasma samples from AD patients, amnestic mild cognitive impairment (aMCI) patients, and normal controls. A biomarker panel consisting of six plasma metabolites (arachidonic acid, N,N-dimethylglycine, thymine, glutamine, glutamic acid, and cytidine) was identified to discriminate AD patients from normal control. Another panel of five plasma metabolites (thymine, arachidonic acid, 2-aminoadipic acid, N,N-dimethylglycine, and 5,8-tetradecadienoic acid) was able to differentiate aMCI patients from control subjects. Both biomarker panels had good agreements with clinical diagnosis. The 2 panels of metabolite markers were all involved in fatty acid metabolism, one-carbon metabolism, amino acid metabolism, and nucleic acid metabolism. Additionally, no altered metabolites were found among the patients at different stages, as well as among those on anticholinesterase medication and those without anticholinesterase medication. These findings provide a comprehensive global plasma metabolite profiling and may contribute to making early diagnosis as well as understanding the pathogenic mechanism of AD and aMCI.

TGF-β Promotes Immune Responses in the Presence of Mesenchymal Stem Cells

Mesenchymal stem cells (MSCs) possess potent immunosuppression capacity and could exert strong therapeutic effects in many diseases, especially inflammatory disorders, in animal models and clinical settings. Although inflammatory cytokines are critical in inducing the immune modulatory properties of MSCs, detailed molecular mechanisms are yet to be fully understood. TGF-β is a well-known anti-inflammatory cytokine and exists in various inflammatory processes; therefore, we investigated whether it could synergize with MSCs in suppressing immune responses. To our surprise, we found that TGF-β actually reversed the immunosuppressive effect of MSCs on anti-CD3 activated splenocytes. Using TGF-β unresponsive MSCs, we demonstrated that the TGF-β directly acted on MSCs. Furthermore, we showed that the effect of TGF-β is exerted through inhibiting inflammatory cytokines induced inducible NO synthase (iNOS) expression in a SMAD3-dependent manner. Interestingly, we found that TGF-β produced by MSCs could act in an autocrine manner to reduce inflammatory cytokine-induced inducible NO synthase expression by MSCs themselves. Therefore, our study revealed a previously unrecognized property of TGF-β in promoting immune responses in the presence of MSCs.