Fibroblast growth factor 23 during septic shock and myocardial injury in ICU patients.

Objective: Fibroblast growth factor 23 (FGF23) has been recognized as an important biomarker of cardiovascular disease and is closely related to inflammation over the past decade. This study aimed to assess the relationship between FGF23 and myocardial injury in patients with sepsis. Methods: We sequentially measured serum FGF23, Klotho, biomarkers of inflammation (CRP, IL-6 and WBC), myocardial injury (cTnI and N-terminal B-type natriuretic peptide) and sepsis (procalcitonin) at peak of intercurrent septic shock and after complete resolution or before death in a series of 29 patients with septic shock. 29 healthy adults without infections were used as controls. Results: There was a difference in serum FGF23 level between patients with septic shock and healthy adults (p < 0.0001), and the peak level of FGF23 in septic shock in the survivor group was higher than that after complete remission (p < 0.0001). No statistical difference was found in the level of FGF23 before and after treatment in the death group (p = 0.0947). At the peak of septic shock, FGF23 was significantly correlated with inflammatory markers, CRP (r = 0.8063, p < 0.0001), PCT (r = 0.6091, p = 0.0005) and WBC (r = 0.8312, p < 0.0001), while the correlation with IL-6 was not statistically significant (r = 0.0098, p = 0.9598). At the same time, it was found that FGF23 was significantly correlated with myocardial injury markers, cTNI (r = 0.8475, p < 0.0001) and NTproBNP (r = 0.8505, p < 0.0001). Nevertheless, FGF23 and klotho are not correlated (r = 0.2609, p = 0.1717). Conclusion: In conclusion, in patients with septic shock and myocardial injury, the exacerbation of inflammation in the septic process was accompanied by a abnormal increase of circulating FGF23 level. FGF23 also subsided after the improvement of inflammation, and the opposite was true for patients who did not survive. The up-regulation of FGF23 may be involved in the response of patients to septic shocks, and it is also speculated that FGF23 is involved in the myocardial injury of septic shock.

Cellular and molecular mechanisms of Notch signal in pulmonary microvascular endothelial cells after acute lung injury.

This study focused on the effect and mechanism of Notch signal on pulmonary microvascular endothelial cells (PMVECs) following acute lung injury. PMVECs were cultured in vitro and randomly divided into eight groups. Grouping was based on whether cells were co-cultured with T cells (splenic CD4+T cells were isolated using MACS microbeads) and the level of Notch expression: Normal group and Normal+T cells group, Model group and Model+T cells group, Notch low-expression group and Notch low-expression+T cells group, and Notch overexpression group and Notch overexpression+T cells group. Except for the Normal group and Normal+T cells group, all other groups were treated with 500 µL lipopolysaccharide (1 µg/mL). The expression of VE-cadherin and Zo-1 protein in the Model group (with or without T cells) was lower than that in the normal group (with or without T cells), their expression in the Notch low-expression group (with or without T cells) was significantly increased, and their expression in the Notch overexpression group (with or without T cells) was significantly decreased. Compared with the normal+T cells group, the number of Treg cells in the Notch low-expression+T cells group decreased significantly (P<0.01). The number of Th17 cells in the Notch overexpression+T cells group was higher than that in the Model+T cells group (P<0.01), while the number of Treg cells decreased (P<0.01). Our results demonstrated that activated Notch signal can down-regulate the expression of the tight junction proteins VE-Cadherin and Zo-1 in PMVECs and affect Th17/Treg immune imbalance. Autophagy was discovered to be involved in this process.

The SlWRKY57-SlVQ21/SlVQ16 module regulates salt stress in tomato.

Salt stress is a major abiotic stress which severely hinders crop production. However, the regulatory network controlling tomato resistance to salt remains unclear. Here, we found that the tomato WRKY transcription factor WRKY57 acted as a negative regulator in salt stress response by directly attenuating the transcription of salt-responsive genes (SlRD29B and SlDREB2) and an ion homeostasis gene (SlSOS1). We further identified two VQ-motif containing proteins SlVQ16 and SlVQ21 as SlWRKY57-interacting proteins. SlVQ16 positively, while SlVQ21 negatively modulated tomato resistance to salt stress. SlVQ16 and SlVQ21 competitively interacted with SlWRKY57 and antagonistically regulated the transcriptional repression activity of SlWRKY57. Additionally, the SlWRKY57-SlVQ21/SlVQ16 module was involved in the pathway of phytohormone jasmonates (JAs) by interacting with JA repressors JA-ZIM domain (JAZ) proteins. These results provide new insights into how the SlWRKY57-SlVQ21/SlVQ16 module finely tunes tomato salt tolerance.

Cellular and molecular mechanisms of Notch signal in pulmonary microvascular endothelial cells after acute lung injury

Abstract This study focused on the effect and mechanism of Notch signal on pulmonary microvascular endothelial cells (PMVECs) following acute lung injury. PMVECs were cultured in vitro and randomly divided into eight groups. Grouping was based on whether cells were co-cultured with T cells (splenic CD4+T cells were isolated using MACS microbeads) and the level of Notch expression: Normal group and Normal+T cells group, Model group and Model+T cells group, Notch low-expression group and Notch low-expression+T cells group, and Notch overexpression group and Notch overexpression+T cells group. Except for the Normal group and Normal+T cells group, all other groups were treated with 500 μL lipopolysaccharide (1 μg/mL). The expression of VE-cadherin and Zo-1 protein in the Model group (with or without T cells) was lower than that in the normal group (with or without T cells), their expression in the Notch low-expression group (with or without T cells) was significantly increased, and their expression in the Notch overexpression group (with or without T cells) was significantly decreased. Compared with the normal+T cells group, the number of Treg cells in the Notch low-expression+T cells group decreased significantly (P<0.01). The number of Th17 cells in the Notch overexpression+T cells group was higher than that in the Model+T cells group (P<0.01), while the number of Treg cells decreased (P<0.01). Our results demonstrated that activated Notch signal can down-regulate the expression of the tight junction proteins VE-Cadherin and Zo-1 in PMVECs and affect Th17/Treg immune imbalance. Autophagy was discovered to be involved in this process.

SlWRKY45 interacts with jasmonate-ZIM domain proteins to negatively regulate defense against the root-knot nematode Meloidogyne incognita in tomato.

Parasitic root-knot nematodes (RKNs) cause a severe reduction in crop yield and seriously threaten agricultural production. The phytohormones jasmonates (JAs) are important signals regulating resistance to multiple biotic and abiotic stresses. However, the molecular mechanism for JAs-regulated defense against RKNs in tomato remains largely unclear. In this study, we found that the transcription factor SlWRKY45 interacted with most JA-ZIM domain family proteins (JAZs), key repressors of the JA signaling. After infection by the RKN Meloidogyne incognita, the slwrky45 mutants exhibited lower gall numbers and egg numbers per gram of roots than wild type, whereas overexpression of SlWRKY45 attenuated resistance to Meloidogyne incognita. Under M. incognita infection, the contents of jasmonic acid (JA) and JA-isoleucine (JA-Ile) in roots were repressed by SlWRKY45-overexpression. Furthermore, SlWRKY45 bound to and inhibited the promoter of the JA biosynthesis gene ALLENE OXIDE CYCLASE (AOC), and repressed its expression. Overall, our findings revealed that the SlJAZ-interaction protein SlWRKY45 attenuated RKN-regulated JA biosynthesis and repressed defense against the RKN M. incognita in tomato.

SlVQ15 interacts with jasmonate-ZIM domain proteins and SlWRKY31 to regulate defense response in tomato.

Botrytis cinerea is one of the most widely distributed and harmful pathogens worldwide. Both the phytohormone jasmonate (JA) and the VQ motif-containing proteins play crucial roles in plant resistance to B. cinerea. However, their crosstalk in resistance to B. cinerea is unclear, especially in tomato (Solanum lycopersicum). In this study, we found that the tomato VQ15 was highly induced upon B. cinerea infection and localized in the nucleus. Silencing SlVQ15 using virus-induced gene silencing reduced resistance to B. cinerea. Overexpression of SlVQ15 enhanced resistance to B. cinerea, while disruption of SlVQ15 using clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein9 (Cas9) technology increased susceptibility to B. cinerea. Furthermore, SlVQ15 formed homodimers. Additionally, SlVQ15 interacted with JA-ZIM domain proteins, repressors of the JA signaling pathway, and SlWRKY31. SlJAZ11 interfered with the interaction between SlVQ15 and SlWRKY31 and repressed the SlVQ15-increased transcriptional activation activity of SlWRKY31. SlVQ15 and SlWRKY31 synergistically regulated tomato resistance to B. cinerea, as silencing SlVQ15 enhanced the sensitivity of slwrky31 to B. cinerea. Taken together, our findings showed that the SlJAZ-interacting protein SlVQ15 physically interacts with SlWRKY31 to cooperatively control JA-mediated plant defense against B. cinerea.

Neuropilin-1 Identifies a New Subpopulation of TGF-ß-Induced Foxp3+ Regulatory T Cells With Potent Suppressive Function and Enhanced Stability During Inflammation.

CD4+Foxp3+ regulatory T cells (Tregs) play a crucial role in preventing autoimmunity and inflammation. There are naturally-derived in the thymus (tTreg), generated extrathymically in the periphery (pTreg), and induced in vitro culture (iTreg) with different characteristics of suppressiveness, stability, and plasticity. There is an abundance of published data on neuropilin-1 (Nrp-1) as a tTreg marker, but little data exist on iTreg. The fidelity of Nrp-1 as a tTreg marker and its role in iTreg remains to be explored. This study found that Nrp-1 was expressed by a subset of Foxp3+CD4+T cells in the central and peripheral lymphoid organs in intact mice, as well as in iTreg. Nrp-1+iTreg and Nrp-1-iTreg were adoptively transferred into a T cell-mediated colitis model to determine their ability to suppress inflammation. Differences in gene expression between Nrp-1+ and Nrp-1-iTreg were analyzed by RNA sequencing. We demonstrated that the Nrp-1+ subset of the iTreg exhibited enhanced suppressive function and stability compared to the Nrp-1- counterpart both in vivo and in vitro, partly depending on IL-10. We found that Nrp-1 is not an exclusive marker of tTreg, however, it is a biomarker identifying a new subset of iTreg with enhanced suppressive function, implicating a potential for Nrp-1+iTreg cell therapy for autoimmune and inflammatory diseases.

Multicenter Investigation of the Initial Hemodialysis Vascular Access and Its Related Factors in Hangzhou of China.

OBJECTIVE: To investigate the initial hemodialysis vascular access in Hangzhou and provide evidence for improving the use of autologous arteriovenous fistula by identifying factors associated with the choice of initial vascular access. METHODS: We retrospectively studied the initial hemodialysis vascular access of 257 patients in five hemodialysis units in Hangzhou of China during a 21-month period (January 2018 to September 2019). A logistic regression was used to identify the risk factors of failing to use an arteriovenous fistula at the initiation of hemodialysis. RESULTS: (1) 257 participants with mean age 67.65 ± 13.43 years old were reviewed, including 165 males (64.2%) and 92 females (35.8%). The etiologies of end-stage renal disease included diabetic nephropathy (37.35%), chronic glomerulonephritis (31.13%), hypertensive nephropathy (14.01%), and other diseases (17.51%). Only 51 patients (19.84%) received arteriovenous fistula, whereas the remaining 206 patients (80.16%) initiated dialysis with a central venous catheter. (2) Logistic regression analysis revealed that the independent risk factors for central venous catheter at the initial hemodialysis were age >70 years old (OR = 4.827, p < 0.01 versus ≤70 years old), chronic glomerulonephritis as the primary etiology (OR = 2.565, p < 0.05 versus nonchronic glomerulonephritis) and eGFR <8.5 mL/min/1.73m2 (OR = 2.283, p < 0.05 versus eGFR ≥8.5 mL/min/1.73m2). CONCLUSION: The proportion of patients using arteriovenous fistula as the initial hemodialysis vascular access in Hangzhou was still low. The choice of vascular access for the first hemodialysis was related to age, eGFR, and the primary etiology of end-stage renal disease. Increasing the proportion of planned vascular access and arteriovenous fistula at the initiation of hemodialysis is still our current goal.

Follicular helper T cells and follicular regulatory T cells in the immunopathology of primary Sjögren's syndrome.

Primary Sjögren's syndrome (pSS) is a chronic autoimmune disease, characterized by lymphocytic infiltration into exocrine glands, which causes dry eyes, dry mouth, and systemic damage. Although the precise etiology of pSS is not clear yet, highly activated B cells, abundant anti-SSA/Ro, and anti-SSB/La autoantibodies are the hallmarks of this disease. Follicular helper T cells (Tfh), a subset of CD4+ T cells, with cell surface receptors PD-1 and CXCR5, express ICOS, transcription factor Bcl-6, and a cytokine IL-21. These cells help in the differentiation of B cells into plasma cells and stimulate the formation of germinal center (GC). Previous studies have demonstrated abundant Tfh cells in the peripheral blood and salivary glands (SGs) of the patients with pSS, correlated with extensive lymphocytic infiltration of the SGs and high disease activity scores. Patients with pSS who are treated with abatacept (CTLA-4 Ig) show fewer circulating Tfh cells, reduced expression of ICOS, and lower disease activity scores. Recently identified follicular regulatory T (Tfr) cells, a subset of regulatory T cells, control the function of Tfh cells and the GC reactions. Here, we summarize the observed alterations in Tfh and Tfr cell numbers, activation state, and circulating subset distribution in pSS. Our goal is to improve the understanding of the roles of Tfh and Tfr cells (surface marker expression, cytokine production, and transcription factors) in the pathogenesis of pSS, thus contributing to the identification of candidate therapeutic agents for this disease.

Trib1 Contributes to Recovery From Ischemia/Reperfusion-Induced Acute Kidney Injury by Regulating the Polarization of Renal Macrophages.

Increasing evidence suggests that macrophage polarization is involved in the recovery from ischemia-reperfusion (I/R)-induced acute kidney injury (AKI), implying that the regulation of macrophage polarization homeostasis might mediate AKI recovery. Trib1 is a key regulator of macrophage differentiation, but its role in AKI remains unclear. Here, we aimed to investigate the role of Trib1 and its link with the macrophage phenotype in the process of adaptive recovery from I/R-induced renal injury. Lentiviral vector-mediated RNA interference (RNAi) was used to knock down Trib1 expression in vitro and in vivo, and a mouse model of moderate AKI was established by the induction of I/R injury. Renal function measurements and inflammatory factors were determined by the corresponding kits. Histomorphology was assessed by hematoxylin-eosin, Masson and PAS staining. Western blot and flow cytometry were employed for the analysis of signal transduction, cell apoptosis and macrophage phenotypes. Trib1 knockdown inhibited cell viability of tubular epithelial cells (TECs) by inhibiting proliferation and enhancing apoptosis in vitro. I/R-induced AKI significantly impaired renal function in mice via increasing the levels of BUN, Scr, NGAL and renal tubular damage, leading to renal fibrosis from days 1 to 3. Through the adaptive self-repair mechanism, renal dysfunction recovered over time and returned to almost normal levels on day 28 after I/R intervention. However, Trib1 depletion worsened renal damage on day 3 and blunted the adaptive repair process of the renal tissue. Mechanistically, Trib1 inhibition suppressed renal tubular cell proliferation under adaptive self-repair conditions by affecting the expression of the proliferation-related proteins cyclin D1, cyclin B, p21, and p27, the apoptosis-related proteins Bcl-2 and Bax, and the fibrosis-related proteins collagen I and III. Furthermore, the M1/M2 macrophage ratio increased in the first 3 days and decreased from day 7 to day 28, consistent with changes in the expression of inflammatory factors, including TNFα, IL-6, IL-12, IL-10, and IL-13. Trib1 inhibition blocked macrophage polarization during adaptive recovery from I/R-induced moderate AKI. Our results show that Trib1 plays a role in kidney recovery and regeneration via the regulation of renal tubular cell proliferation by affecting macrophage polarization. Thus, Trib1 might be a viable therapeutic target to improve renal adaptive repair following I/R injury.

PD-1+CXCR5-CD4+T cells are correlated with the severity of systemic lupus erythematosus.

OBJECTIVES: PD-1+CXCR5-CD4+T peripheral helper (Tph) cells, a recently identified T cell subset, are proven to promote B cell responses and antibody production in rheumatoid arthritis, but their role in the pathogenesis of SLE is unknown. We explored the role of Tph in lupus disease development. METHODS: This cohort study included 68 patients with SLE and 41 age- and sex-matched healthy individuals. The frequency of PD-1+CXCR5-CD4+T cells was analysed in peripheral blood by flow cytometry. Inducible T-cell costimulator, CD38, MHC-II, IL-21, CXCR3 and CCR6 expression were measured in Tph cells. Comparisons between the two groups were performed, and correlations between Tph cells and other parameters were investigated. RESULTS: We revealed a markedly expanded population of Tph cells (8.31 ± 5.45 vs 2.86 ± 1.31%, P < 0.0001) in the circulation of patients with SLE (n = 68), compared with healthy controls (n = 41). Tph cells were much higher in the active group than in the inactive group (14.21 ± 5.21 vs 5.49 ± 2.52%, P < 0.0001). Tph cells were significantly associated with SLEDAI score (r = 0.802), ESR (r = 0.415), IgG (r = 0.434), C3 (r = -0.543), C4 (r = -0.518) and IL-21 level (r = 0.628), and ANA titre (r = 0.272). Furthermore, Tph cells were much higher in lupus patients with arthritis, nephritis, rash, alopecia, pleuritis, pericarditis and haematological involvement. Tph cells were associated with CD138+/CD19+ plasma cells (r = 0.518). Furthermore, MHC-II, inducible T-cell costimulator, CD38, and IL-21 expression were all higher in Tph cells from SLE patients compared with healthy controls. CXCR3+CCR6-Tph (Tph1) cells were expanded in the SLE patients. CONCLUSION: Our data show that relative number of Tph cells is correlated with disease measures in patients with SLE, suggesting an important role in lupus disease development.

Mesenchymal Stem Cells in Primary Sjögren's Syndrome: Prospective and Challenges.

Primary Sjögren's syndrome (pSS) is a chronic systemic inflammatory autoimmune disease characterized by lymphocytic infiltrates in exocrine glands. Current approaches do not control harmful autoimmune attacks or prevent irreversible damage and have considerable side effects. Mesenchymal stem cells (MSCs) have been effective in the treatment of several autoimmune diseases. The objective of this review is to illustrate the potential therapeutic role of MSCs in pSS. We summarize the recent advances in what is known about their immunomodulatory function and therapeutic applications in pSS. MSC transfusion can suppress autoimmunity and restore salivary gland secretory function in mouse models and patients with pSS by inducing regulatory T cells, suppressing Th1, Th17, and T follicular helper cell responses. In addition, MSCs can differentiate into salivary epithelial cells, presenting an option as a suitable alternative treatment. We also discuss current bioengineering methods which improve functions of MSCs for pSS. However, there remain many challenges to overcome before their wide clinical application.

Apremilast Ameliorates Experimental Arthritis via Suppression of Th1 and Th17 Cells and Enhancement of CD4+Foxp3+ Regulatory T Cells Differentiation.

Apremilast is a novel phosphodiesterase 4 (PDE4) inhibitor suppressing immune and inflammatory responses. We assessed the anti-inflammatory effects of Apremilast in type II collagen (CII)-induced arthritis (CIA) mouse model. To determine whether Apremilast can ameliorate arthritis onset in this model, Apremilast was given orally at day 14 after CII immunization. Bone erosion was measured by histological and micro-computed tomographic analysis. Anti-mouse CII antibody levels were measured by enzyme-linked immunosorbent assay, and Th17, Th1 cells, and CD4+Foxp3+ regulatory T (Treg) cells were assessed by flow cytometry in the lymph nodes. Human cartilage and rheumatoid arthritis (RA) synovial fibroblasts (RASFs) implantation in the severe combined immunodeficiency mouse model of RA were used to study the role of Apremilast in the suppression of RASF-mediated cartilage destruction in vivo. Compared with untreated and vehicle control groups, we found that Apremilast therapy delayed arthritis onset and reduced arthritis scores in the CIA model. Total serum IgG, IgG1, IgG2a, and IgG2b were all decreased in the Apremilast treatment groups. Moreover, Apremilast markedly prevented the development of bone erosions in CIA mice by CT analysis. Furthermore, in the Apremilast treated group, the frequency of Th17 cells and Th1 cells was significantly decreased while Treg cells' frequency was significantly increased. The high dose of Apremilast (25 mg/kg) was superior to low dose (5 mg/kg) in treating CIA. Apremilast treatment reduced the migratory ability of RASFs and their destructive effect on cartilage. Compared with the model group, Apremilast treatment significantly reduced the RASFs invasion cartilage scores in both primary implant and contralateral implant models. Our data suggest that Apremilast is effective in treating autoimmune arthritis and preventing the bone erosion in the CIA model, implicating its therapeutic potential in patients with RA.

Gut bacteria selectively promoted by dietary fibers alleviate type 2 diabetes.

The gut microbiota benefits humans via short-chain fatty acid (SCFA) production from carbohydrate fermentation, and deficiency in SCFA production is associated with type 2 diabetes mellitus (T2DM). We conducted a randomized clinical study of specifically designed isoenergetic diets, together with fecal shotgun metagenomics, to show that a select group of SCFA-producing strains was promoted by dietary fibers and that most other potential producers were either diminished or unchanged in patients with T2DM. When the fiber-promoted SCFA producers were present in greater diversity and abundance, participants had better improvement in hemoglobin A1c levels, partly via increased glucagon-like peptide-1 production. Promotion of these positive responders diminished producers of metabolically detrimental compounds such as indole and hydrogen sulfide. Targeted restoration of these SCFA producers may present a novel ecological approach for managing T2DM.

Human Gingiva-Derived Mesenchymal Stem Cells Ameliorate Streptozoticin-induced T1DM in mice via Suppression of T effector cells and Up-regulating Treg Subsets.

There is yet no cure for type 1 diabetes (T1DM) so far. A significant body of evidence has demonstrated that bone marrow-derived mesenchymal stem cells (BMSCs) showed great potential in controlling T1DM. But there exists much difficulty in using BMSCs as a clinical therapy. We here test whether a new population of mesenchymal stem cells from human gingiva (GMSCs), which has many advantages over BMSCs, can delay or prevent progress of T1DM. GMSCs were adoptively transferred to multiple low-dose streptozotocin (STZ)-induced T1DM. Blood glucose levels and disease severities were analyzed. T cells subsets in blood, spleen and lymph nodes were detected dynamically by flow cytometry. GMSC distribution was dynamically analyzed. We found that infusion of GMSCs but not fibroblast cells significantly controlled blood glucose levels, delayed diabetes onset, ameliorated pathology scores in pancreas, and down-regulated production of IL-17 and IFN-γ in CD4+ and CD8+ T cells in spleens, pancreatic lymph nodes (pLN) and other lymph nodes. GMSCs also up-regulated the levels of CD4+ Treg induced in the periphery. Mechanismly, GMSCs could migrate to pancreas and local lymph node and function through CD39/CD73 pathway to regulate effector T cells. Thus, GMSCs show a potential promise in treating T1DM in the clinic.

Nonalcoholic Fatty Liver Disease and Associated Metabolic Risks of Hypertension in Type 2 Diabetes: A Cross-Sectional Community-Based Study.

The mechanisms facilitating hypertension in diabetes still remain to be elucidated. Nonalcoholic fatty liver disease (NAFLD), which is a higher risk factor for insulin resistance, shares many predisposing factors with diabetes. However, little work has been performed on the pathogenesis of hypertension in type 2 diabetes (T2DM) with NAFLD. The aim of this study is to investigate the prevalence of hypertension in different glycemic statuses and to analyze relationships between NAFLD, metabolic risks, and hypertension within a large community-based population after informed written consent. A total of 9473 subjects aged over 45 years, including 1648 patients with T2DM, were enrolled in this cross-sectional study. Clinical and biochemical parameters of all participants were determined. The results suggested that the patients with prediabetes or T2DM were with higher risks to have hypertension. T2DM with NAFLD had significantly higher levels of blood pressure, triglyceride, uric acid, and HOMA-IR than those without NAFLD. Data analyses suggested that hypertriglyceridemia [OR = 1.773 (1.396, 2.251)], NAFLD [OR = 2.344 (1.736, 3.165)], hyperuricemia [OR = 1.474 (1.079, 2.012)], and insulin resistance [OR = 1.948 (1.540, 2.465)] were associated with the higher prevalence of hypertension independent of other metabolic risk factors in type 2 diabetes. Further studies are needed to focus on these associations.

The role of all-trans retinoic acid in the biology of Foxp3+ regulatory T cells.

Regulatory T (Treg) cells are necessary for immune system homeostasis and the prevention of autoimmune diseases. Foxp3 is specifically expressed in Treg cells and plays a key role in their differentiation and function. Foxp3(+) Treg cells are consisted of naturally occurring, thymus-derived Treg (nTreg) and peripheral-induced Treg (iTreg) cells that may have different functional characteristics or synergistic roles. All-trans retinoic acid (atRA), a vitamin A metabolite, regulates a wide range of biological processes, including cell differentiation and proliferation. Recent studies demonstrated that atRA also regulates the differentiation of T helper (Th) cells and Treg cells. Moreover, atRA also sustains nTreg stability under inflammatory conditions. In this review, we summarize the significant progress of our understanding of the role(s) and mechanisms of atRA in Treg biology.

Differential role of all-trans retinoic acid in promoting the development of CD4+ and CD8+ regulatory T cells.

It is known that ATRA promotes the development of TGF-ß-induced CD4(+)Foxp3(+) iTregs, which play a vital role in the prevention of autoimmune diseases; however, the role of ATRA in facilitating the differentiation and function of CD8(+)Foxp3(+) iTregs remains elusive. Using a head-to-head comparison, we found that ATRA promoted expression of Foxp3 and development of CD4(+) iTregs, but it did not promote Foxp3 expression on CD8(+) cells. Using a standard in vitro assay, we demonstrated that CD8(+) iTregs induced by TGF-ß and ATRA were not superior to CD8(+) iTregs induced by TGF-ß alone. In cGVHD, in a typical lupus syndrome model where DBA2 spleen cells were transferred to DBA2xC57BL/6 F1 mice, we observed that both CD8(+) iTregs induced by TGF-ß and ATRA and those induced by TGF-ß alone had similar therapeutic effects. ATRA did not boost but, conversely, impaired the differentiation and function of human CD8(+) iTregs. CD8(+) cells expressed the ATRA receptor RAR and responded to ATRA, similar to CD4(+) cells. We have identified the differential role of ATRA in promoting Foxp3(+) Tregs in CD4(+) and CD8(+) cell populations. These results will help to determine a protocol for developing different Treg cell populations and may provide novel insights into clinical cell therapy for patients with autoimmune diseases and those needing organ transplantation.

NADPH oxidase-dependent formation of reactive oxygen species contributes to transforming growth factor ß1-induced epithelial-mesenchymal transition in rat peritoneal mesothelial cells, and the role of astragalus intervention.

OBJECTIVE: To investigate the role of nicotinamide-adenine dinucleotide phosphate (NADPH) oxidasedependent formation of reactive oxygen species (ROS) in the transforming growth factor ß1 (TGF-ß1)-induced epithelial-mesenchymal transition (EMT) in rat peritoneal mesothelial cells (RPMCs), and the effect of Astragalus injection (AGI) intervention. METHODS: Primary RPMCs were cultured to the second generation in vitro. After synchronization for 24 h, the cells were randomly assigned to the following groups: control (Group A), AGI (2 g/mL; Group B), TGF-ß1 (10 ng/mL; Group C), TGF-ß1 (10 ng/mL) + AGI (2 g/mL; Group D; pretreated for 1 h with AGI before TGF-ß1 stimulation). Reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analysis were employed to evaluate the mRNA and protein expression of the NADPH oxidase subunit p67phox, α-smooth muscle actin (α-SMA) and E-cadherin. The dichlorofluorescein-sensitive cellular ROS levels were measured by a fluorometric assay and confocal microscopy. RESULTS: TGF-ß1 significantly induced NADPH oxidase subunit p67phox mRNA and protein expression in RPMCs, as well as inducing the production of intracellular ROS. AGI inhibited this TGF-ß1-induced up-regulation by 39.3% and 47.8%, respectively (P<0.05), as well as inhibiting the TGF-ß1-induced ROS generation by 56.3% (P<0.05). TGF-ß1 also induced α-SMA mRNA and protein expression, and down-regulated E-cadherin mRNA and protein expression (P<0.05). This effect was suppressed by AGI (P<0.05). CONCLUSIONS: NADPH oxidase-dependent formation of ROS may mediate the TGF-ß1-dependent EMT in RPMCs. AGI could inhibit this process, providing a theoretical basis for AGI in the prevention of peritoneal fibrosis.

Phenotypic and functional characteristic of a newly identified CD8+ Foxp3- CD103+ regulatory T cells.

TGF-ß and Foxp3 expressions are crucial for the induction and functional activity of CD4(+)Foxp3(+) regulatory T (iTreg) cells. Here, we demonstrate that although TGF-ß-primed CD8(+) cells display much lower Foxp3 expression, their suppressive capacity is equivalent to that of CD4(+) iTreg cells, and both Foxp3(-) and Foxp3(+) CD8+ subsets have suppressive activities in vitro and in vivo. CD8(+)Foxp3(-) iTreg cells produce little IFN-γ but almost no IL-2, and display a typical anergic phenotype. Among phenotypic markers expressed in CD8(+)Foxp3(-) cells, we identify CD103 expression particularly crucial for the generation and function of this subset. Moreover, IL-10 and TGF-ß signals rather than cytotoxicity mediate the suppressive effect of this novel Treg population. Therefore, TGF-ß can induce both CD8(+)Foxp3(-) and CD8(+)Foxp3(+) iTreg subsets, which may represent the unique immunoregulatory means to treat autoimmune and inflammatory diseases.