Development of a novel and effective vaccine against Mycobacterium tuberculosis (M.tb) is a challenging for preventing TB infection. In this study, a novel nanoparticle-based recombinant DNA vaccine was developed, which contains Esat-6 three T cell epitopes (Esat-6/3e) and fms-like tyrosine kinase 3 ligand (FL) genes (termed Esat-6/3e-FL), and was enveloped with chitosan (CS) nanoparticles (nano-chitosan). The immunologic and protective efficacy of the nano-chitosan-based DNA vaccine (termed nano-Esat-6/3e-FL) was assessed in C57BL/6 mice after intramuscular prime vaccination with the plasmids DNA and nasal boost with the Esat-6/3e peptides. The results showed that the immunized mice remarkably elicited enhanced T cell responses and protection against M.tb H37Rv challenge. These findings indicate that the nano-chitosan can significantly elevate the immunologic and protective effects of the DNA vaccine, and the nano-Esat-6/3e-FL is a useful vaccine for preventing M.tb infection in mice.

Interleukin 17 (IL-17), produced mainly by T helper 17 (Th17) cells, is increasingly recognized as a key regulator in various autoimmune diseases, including human multiple sclerosis (MS) and its animal model, experimental autoimmune encephalomyelitis (EAE). Although several microRNAs (miRNAs) with aberrant expression have been shown to contribute to the pathogenesis of MS and EAE, the mechanisms underlying the regulation of abnormal miRNA expression in astrocytes upon IL-17 stimulation remain unclear. In the present study, we detected the changes of miRNA expression profiles both in the brain tissue of EAE mice and in cultured mouse primary astrocytes stimulated with IL-17 and identified miR-873 as one of the co-up-regulated miRNAs in vivo and in vitro. The overexpression of miR-873, demonstrated by targeting A20 (TNFα-induced protein 3, TNFAIP3), remarkably reduced the A20 level and promoted NF-κB activation in vivo and in vitro as well as increasing the production of inflammatory cytokines and chemokines (i.e. IL-6, TNF-α, MIP-2, and MCP-1/5). More importantly, silencing the endogenous miR-873 or A20 gene with lentiviral vector of miR-873 sponge (LV-miR-873 sponge) or short hairpin RNA (shRNA) of A20 (LV-A20 shRNA) in vivo significantly lessened or aggravated inflammation and demyelination in the central nervous system (CNS) of EAE mice, respectively. Taken together, these findings indicate that miR-873 induced by IL-17 stimulation promotes the production of inflammatory cytokines and aggravates the pathological process of EAE mice through the A20/NF-κB pathway, which provides a new insight into the mechanism of inflammatory damage in MS.

The increased production of interleukin 6 (IL‐6) and transforming growth factor β1 (TGF‐β1) by glomerular mesangial cells (GMCs) in rat Thy‐1 nephritis (Thy‐1N), a model of human mesangioproliferative glomerulonephritis, is accompanied by sublytic C5b‐9 deposition, but the mechanism of IL‐6 and TGF‐β1 upregulation in rat GMCs triggered by sublytic C5b‐9 in this model has not been elucidated. In the present study, molecular biological methods, along with functional and histological studies, were performed. We found that sublytic C5b‐9 (here, <5% lactate dehydrogenase release from cells is regarded as a sublytic effect) could induce the production of IL‐6 and TGF‐β1 through upregulation of p300 and C/EBPβ expression in GMCs, which might contribute to renal Th17/IL‐17 production in Thy‐1N rats. In addition, CCAAT/enhancer‐binding protein (C/EBPβ)‐enhanced IL‐6 and TGF ‐β 1 promoter activity and p300‐mediated C/EBPβ acetylation were involved in upregulation of IL‐6 and TGF‐β1 expression in GMCs attacked by sublytic C5b‐9. In detail, the elements of C/EBPβ binding to rat IL‐6 and TGF ‐β 1 promoter and 3 acetylated sites of rat C/EBPβ protein were first revealed. Furthermore, silencing the p300 or C/EBPβ gene in rat kidney significantly reduced the production of IL‐6 and TGF‐β1 and renal lesions in Thy‐1N rats. Together, these data indicate that the mechanism of IL‐6 and TGF‐β1 production in renal tissue of Thy‐1N rats is associated with sublytic C5b‐9 upregulated p300 and p300‐mediated C/EBPβ acetylation as well as C/EBPβ‐activated IL‐6 and TGF‐β1 genes.—Zhang, J., Li, Y., Shan, K., Wang, L., Qiu, W., Lu, Y., Zhao, D., Zhu, G., He, F., Wang, Y. Sublytic C5b‐9 induces IL‐6 and TGF‐β1 production by glomerular mesangial cells in rat Thy‐1 nephritis through p300‐mediated C/EBPβ acetylation. FASEB J. 28, 1511–1525 (2014). www.fasebj.org

The apoptosis of glomerular mesangial cells (GMCs) in rat Thy-1 nephritis (Thy-1N), a model of human mesangioproliferative glomerulonephritis (MsPGN), is accompanied by sublytic C5b-9 deposition. However, the mechanism by which sublytic C5b-9 induces GMC apoptosis is unclear. In the present studies, the effect of X-linked inhibitor of apoptosis-associated factor 1 (XAF1) expression on GMC apoptosis and the role of p300 and interferon regulatory factor-1 (IRF-1) in mediating XAF1 gene activation were determined, both in the GMCs induced by sublytic C5b-9 (in vitro) and in the renal tissues of rats with Thy-1N (in vivo). The in vitro studies demonstrated that IRF-1-enhanced XAF1 gene activation and its regulation by p300-mediated IRF-1 acetylation were involved in GMC apoptosis induced by sublytic C5b-9. The element of IRF-1 binding to XAF1 promoter and two acetylated sites of IRF-1 protein were also revealed. In vivo, silence of p300, IRF-1 or XAF1 genes in the renal tissues diminished GMC apoptosis and secondary GMC proliferation as well as urinary protein secretion in Thy-1N rats. Together, these data implicate that sublytic C5b-9 induces the expression of both p300 and IRF-1, as well as p300-dependent IRF-1 acetylation that may contribute to XAF1 gene activation and subsequent GMC apoptosis in Thy-1N rats.