An Adaptive Deconvolution Method with Improve Enhanced Envelope Spectrum and Its Application for Bearing Fault Feature Extraction.

To address the problem that complex bearing faults are coupled to each other, and the difficulty of diagnosis increases, an improved envelope spectrum-maximum second-order cyclostationary blind deconvolution (IES-CYCBD) method is proposed to realize the separation of vibration signal fault features. The improved envelope spectrum (IES) is obtained by integrating the part of the frequency axis containing resonance bands in the cyclic spectral coherence function. The resonant bands corresponding to different fault types are accurately located, and the IES with more prominent target characteristic frequency components are separated. Then, a simulation is carried out to prove the ability of this method, which can accurately separate and diagnose fault types under high noise and compound fault conditions. Finally, a compound bearing fault experiment with inner and outer ring faults is designed, and the inner and outer ring fault characteristics are successfully separated by the proposed IES-CYCBD method. Therefore, simulation and experiments demonstrate the strong capability of the proposed method for complex fault separation and diagnosis.

Mixed small cell neuroendocrine carcinoma and squamous cell carcinoma covered by tubulovillous adenoma in the rectum: a case report and detailed molecular analyses.

BACKGROUND: Previously, only six cases of mixed neuroendocrine-non-neuroendocrine neoplasm (MiNENs) with squamous cell carcinoma (SCC) component have been described in the colorectum, and the molecular landscape of MiNENs is also poorly understood. Herein, we present a unique case in which the SCC developed as a component of a MiNEN in the rectum. CASE PRESENTATION: The patient was firstly diagnosed as rectal small cell neuroendocrine carcinoma (SCNEC) covered by tubulovillous adenoma, and then mixed SCNEC and SCC in the same site 6 months later. Representative samples from the three histologic subtypes were then sent for next-generation sequencing (NGS) separately. Multiple liver metastases occurred in the following month after the last surgery. The patient died of ketoacidosis 1 year after initial diagnosis of the tumor. CONCLUSION: This is the first report of this exceedingly rare tumor type to include NGS of the 3 separate morphological entities. Our findings may expedite the understanding of combined tumors in the colorectum.

Sublytic C5b-9 Induces IL-23 and IL-36a Production by Glomerular Mesangial Cells via PCAF-Mediated KLF4 Acetylation in Rat Thy-1 Nephritis.

Sublytic C5b-9 formation on glomerular mesangial cells in rat Thy-1 nephritis (Thy-1N), a model of human mesangioproliferative glomerulonephritis, is accompanied by the production of proinflammatory cytokines, but the relationship between sublytic C5b-9 and cytokine synthesis and the underlying mechanism remains unclear. To explore the problems mentioned above, in this study, we first examined the levels of proinflammatory ILs (e.g., IL-23 and IL-36a) as well as transcription factor (KLF4) and coactivator (PCAF) in the renal tissues of Thy-1N rats and in the glomerular mesangial cell line (HBZY-1) stimulated by sublytic C5b-9. Then, we further determined the role of KLF4 and PCAF in sublytic C5b-9-induced IL-23 and IL-36a production as well as the related mechanism. Our results showed that the levels of KLF4, PCAF, IL-23, and IL-36a were obviously elevated. Mechanistic investigation revealed that sublytic C5b-9 stimulation could increase IL-23 and IL-36a synthesis through KLF4 and PCAF upregulation, and KLF4 and PCAF could form a complex, binding to the IL-23 or IL-36a promoter in a KLF4-dependent manner, causing gene transcription. Importantly, KLF4 acetylation by PCAF contributed to sublytic C5b-9-induced IL-23 and IL-36a transcription. Besides, the KLF4 binding regions on IL-23 or IL-36a promoters and the KLF4 lysine site acetylated by PCAF were identified. Furthermore, silencing renal KLF4 or PCAF gene could significantly inhibit IL-23 or IL-36a secretion and tissue damage of Thy-1N rats. Collectively, these findings implicate that the KLF4/PCAF interaction and KLF4 acetylation by PCAF play a pivotal role in the sublytic C5b-9-mediated IL-23 and IL-36a production of Thy-1N rats.

Sublytic C5b-9 induces proliferation of glomerular mesangial cells via ERK5/MZF1/RGC-32 axis activated by FBXO28-TRAF6 complex.

Mesangioproliferative glomerulonephritis (MsPGN) is characterized by the proliferation of glomerular mesangial cells (GMCs) and accumulation of extracellular matrix (ECM), followed by glomerulosclerosis and renal failure of patients. Although our previous studies have demonstrated that sublytic C5b-9 complex formed on the GMC membrane could trigger GMC proliferation and ECM expansion of rat Thy-1 nephritis (Thy-1N) as an animal model of MsPGN, their mechanisms are still not fully elucidated. In the present studies, we found that the levels of response gene to complement 32 (RGC-32), myeloid zinc finger 1 (MZF1), phosphorylated extracellular signal-regulated kinase 5 (phosphorylated ERK5, p-ERK5), F-box only protein 28 (FBXO28) and TNF receptor-associated factor 6 (TRAF6) were all markedly up-regulated both in the renal tissues of rats with Thy-1N (in vivo) and in the GMCs upon sublytic C5b-9 stimulation (in vitro). Further in vitro experiments revealed that up-regulated FBXO28 and TRAF6 could form protein complex binding to ERK5 and enhance ERK5 K63-ubiquitination and subsequent phosphorylation. Subsequently, ERK5 activation contributed to MZF1 expression and MZF1-dependent RGC-32 up-regulation, finally resulting in GMC proliferative response. Furthermore, the MZF1-binding element within RGC-32 promoter and the functions of FBXO28 domains were identified. Additionally, knockdown of renal FBXO28, TRAF6, ERK5, MZF1 and RGC-32 genes respectively markedly reduced GMC proliferation and ECM production in Thy-1N rats. Together, these findings indicate that sublytic C5b-9 induces GMC proliferative changes in rat Thy-1N through ERK5/MZF1/RGC-32 axis activated by the FBXO28-TRAF6 complex, which might provide a new insight into MsPGN pathogenesis.

Sublytic C5b-9 Induces Glomerular Mesangial Cell Apoptosis through the Cascade Pathway of MEKK2-p38 MAPK-IRF-1-TRADD-Caspase 8 in Rat Thy-1 Nephritis.

The apoptosis of glomerular mesangial cells (GMCs) in the early phase of rat Thy-1 nephritis (Thy-1N), a model of human mesangioproliferative glomerulonephritis (MsPGN), is primarily triggered by sublytic C5b-9. However, the mechanism of GMC apoptosis induced by sublytic C5b-9 remains unclear. In this study, we demonstrate that expressions of TNFR1-associated death domain-containing protein (TRADD) and IFN regulatory factor-1 (IRF-1) were simultaneously upregulated in the renal tissue of Thy-1N rats (in vivo) and in GMCs under sublytic C5b-9 stimulation (in vitro). In vitro, TRADD was confirmed to be a downstream gene of IRF-1, because IRF-1 could bind to TRADD gene promoter to promote its transcription, leading to caspase 8 activation and GMC apoptosis. Increased phosphorylation of p38 MAPK was verified to contribute to IRF-1 and TRADD production and caspase 8 activation, as well as to GMC apoptosis induced by sublytic C5b-9. Furthermore, phosphorylation of MEK kinase 2 (MEKK2) mediated p38 MAPK activation. More importantly, three sites (Ser153/164/239) of MEKK2 phosphorylation were identified and demonstrated to be necessary for p38 MAPK activation. In addition, silencing of renal MEKK2, IRF-1, and TRADD genes or inhibition of p38 MAPK activation in vivo had obvious inhibitory effects on GMC apoptosis, secondary proliferation, and urinary protein secretion in rats with Thy-1N. Collectively, these findings indicate that the cascade axis of MEKK2-p38 MAPK-IRF-1-TRADD-caspase 8 may play an important role in GMC apoptosis following exposure to sublytic C5b-9 in rat Thy-1N.

Sublytic C5b-9 Induces Glomerular Mesangial Cell Apoptosis Through miR-3546/SOX4/Survivin Axis in Rat Thy-1 Nephritis.

BACKGROUND/AIMS: The activation of complement system and the formation of C5b-9 complex have been confirmed in the glomeruli of patients with mesangioproliferative glomerulonephritis (MsPGN). However, the role and mechanism of C5b-9-induced injury in glomerular mesangial cell (GMC) are poorly understood. Rat Thy-1N is an animal model for studying MsPGN. It has been revealed that the attack of C5b-9 to the GMC in rat Thy-1N is sublytic, and sublytic C5b-9 can cause GMC apoptosis, but the underlying mechanism is not fully elucidated. To explore the role and regulatory mechanism of C5b-9 in MsPGN lesion, we used rat Thy-1N model and first detected the change of microRNA (miRNA) profiles both in Thy-1N rat renal tissues (in vivo) and in the cultured GMCs with sublytic C5b-9 stimulation (in vitro). Then we determined the effect of miR-3546, which increased both in vivo and in vitro, on GMC apoptosis upon sublytic C5b-9 as well as the involved mechanism. METHODS: Rat Thy-1N model was established and GMCs were treated with sublytic C5b-9. The rat renal cortex and the stimulated GMCs were obtained for miRNA microarray detection. Subsequently, the increased miRNAs were verified by real-time PCR. Meanwhile, to ascertain the ability of some miRNAs to upregulate cleaved caspase 3 and induce GMC apoptosis, the corresponding miRNA mimics were transfected into GMCs, followed by western blotting (WB) and flow cytometry mesurement. Thereafter, the miR-3546-targeted gene (SOX4) was predicted using bioinformatics approaches, and SOX4 expression in Thy-1N tissues and in the GMCs upon sublytic C5b-9 stimulation or miR-3546 mimic/inhibitor transfection were detected using real-time PCR and WB. To prove that miR-3546 can affect SOX4 gene transcription and SOX4 can regulate survivin expression, dual luciferase reporter assay, real-time PCR, WB and chromatin immunoprecipitation (ChIP) assays were performed. Furthermore, the role of miR-3546/SOX4/survivin axis in the GMC apoptosis induced by sublytic C5b-9 was examined using WB and flow cytometry. RESULTS: Compared with normal renal tissues and untreated GMCs, there were 43 and 62 upregulated miRNAs (> 2-fold) in Thy-1N tissues and sublytic C5b-9-stimulated GMCs respectively. A total of 17 miRNAs were increased both in vivo and in vitro, 11 of which were validated by real-time PCR. Among them, miR-3546 could markedly promote GMC apoptosis and inhibit SOX4 or survivin expression in response to sublytic C5b-9, and either SOX4 or survivin overexpression markedly rescued the GMC apoptosis mediated by miR-3546 mimic. Additionally, SOX4 overexpression could reverse the survivin suppression by miR-3546 mimic, and SOX4 could bind to survivin promoter (-1,278 to -853 nt) and activate survivin gene transcription. CONCLUSION: MiR-3546/ SOX4/survivin axis has a promoting role in the GMC apoptosis triggered by sublytic C5b-9, and our findings may provide a new insight into the pathogenesis of rat Thy-1N and human MsPGN.

Interleukin-17 (IL-17)-induced microRNA 873 (miR-873) contributes to the pathogenesis of experimental autoimmune encephalomyelitis by targeting A20 ubiquitin-editing enzyme.

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.

Sublytic C5b-9 induces IL-6 and TGF-ß1 production by glomerular mesangial cells in rat Thy-1 nephritis through p300-mediated C/EBPß acetylation.

CCAAT/enhancer-binding protein (C/EBPß)-enhanced IL-6 and TGF-ß1 promoter activity and p300-mediated C/EBPß acetylation were involved in up-regulation of IL-6 and TGF-ß1 expression in GMCs attacked by sublytic C5b-9. In detail, the elements of C/EBPß binding to rat IL-6 and TGF-ß1 promoter and 3 acetylated sites of rat C/EBPß protein were first revealed. Furthermore, silencing the p300 or C/EBPß gene in rat kidney significantly reduced the production of IL-6 and TGF-ß1 and renal lesions in Thy-1N rats. Together, these data indicate that the mechanism of IL-6 and TGF-ß1 production in renal tissue of Thy-1N rats is associated with sublytic C5b-9 up-regulated p300 and p300-mediated C/EBPß acetylation as well as C/EBPß-activated IL-6 and TGF-ß1 genes.

Spatial metabolome of biosynthesis and metabolism in Cyclocarya paliurus leaves.

A large number of plant metabolites were discovered, but their biosynthetic and metabolic pathways are still largely unknown. However, the spatial distribution of metabolites and their changes in metabolic pathways can be supplemented by mass spectrometry imaging (MSI) techniques. For this purpose, the combination of desorption electrospray ionization (DESI)-MSI and non-targeted metabolomics was used to obtain the spatial distribution information of metabolites in the leaves of Cyclocarya paliurus (Batal.) Iljinskaja (C. paliurus). The sample pretreatment method was optimized to have higher detection sensitivity in DESI. The changes of metabolites in C. paliurus were analyzed in depth with the integration of the spatial distribution information of metabolites. The main pathways for biosynthesis of flavonoid precursor and the effect of changes in compound structure on the spatial distribution were found. Spatial metabolomics can provide more metabolite information and a platform for the in-depth understanding of the biosynthesis and metabolism in plants.

Sulfated Chinese yam polysaccharide exert anti-inflammatory potential via MAPK/NF-κB signaling pathways in a co-culture system and LPS-induced acute inflammatory mice model.

Our previous study has demonstrated that sulfated Chinese yam polysaccharide (SCYP) can improve immunomodulatory activity in Raw 264.7 cells. However, its anti-inflammatory is little known. In this study, the anti-inflammatory effects of SCYP were systematically investigated via the Lipopolysaccharides (LPS)-induced Raw264.7 cell model, Caco-2/Raw264.7 co-culture system, and acute inflammation mice model. The results suggested SCYP promoted the cell proliferation and have no toxicity in Raw264.7 and Caco-2 cells at the concentration of 200 µg/mL. Moreover, when treated with SCYP, the production of pro-inflammatory cytokines (interleukin [IL]-1ß, IL-6, and tumor necrosis factor-α) reduced significantly in Raw264.7 via the MAPK/NF-κB pathway. In the Caco-2/Raw264.7 co-cultured system, SCYP could regulate inflammation reaction by improving intestinal barrier, which might prevent systemic inflammation. Further, systemic inflammation was alleviated by SCYP in LPS-induced acute inflammation mice through MAPK/NF-κB pathway. PRACTICAL APPLICATION: These results supported that SCYP may be used as an anti-inflammation agent in the functional food field.

Micheliolide ameliorates lipopolysaccharide-induced acute kidney injury through suppression of NLRP3 activation by promoting mitophagy via Nrf2/PINK1/Parkin axis.

BACKGROUND: Sepsis-associated acute kidney injury (SA-AKI) represents a frequent complication of in critically ill patients. The objective of this study is to illuminate the potential protective activity of Micheliolide (MCL) and its behind mechanism against SA-AKI. METHODS: The protective potential of MCL on SA-AKI was investigated in lipopolysaccharide (LPS) treated HK2 cells and SA-AKI mice model. The mitochondrial damage was determined by detection of reactive oxygen species and membrane potential. The Nrf2 silencing was achieved by transfection of Nrf2-shRNA in HK2 cells, and Nrf2 inhibitor, ML385 was employed in SA-AKI mice. The mechanism of MCL against SA-AKI was evaluated through detecting hallmarks related to inflammation, mitophagy and Nrf2 pathway via western blotting, immunohistochemistry, and enzyme linked immunosorbent assay. RESULTS: MCL enhanced viability, suppressed apoptosis, decreased inflammatory cytokine levels and improved mitochondrial damage in LPS-treated HK2 cells, and ameliorated renal injury in SA-AKI mice. Moreover, MCL could reduce the activation of NLRP3 inflammasome via enhancing mitophagy. Additionally, Nrf2 deficiency reduced the suppression effect of MCL on NLRP3 inflammasome activation and blocked the facilitation effect of MCL on mitophagy in LPS-treated HK2 cells, the consistent is true for ML385 treatment in SA-AKI mice. CONCLUSIONS: MCL might target Nrf2 and further reduce the NLRP3 inflammasome activation via enhancing mitophagy, which alleviated SA-AKI.

Integrated clinicopathological and immunohistochemical analysis of gastric adenocarcinoma with hepatoid differentiation: an exploration of histogenesis, molecular characteristics, and prognostic markers.

In addition to hepatoid adenocarcinoma (HAC), gastric adenocarcinoma with enteroblastic differentiation (GAED) and common adenocarcinoma (COM) could also show hepatoid differentiation, which presents a poor prognosis. To elucidate the histogenesis and development of gastric cancer with hepatoid differentiation, we identified 55 cases by histological morphology and a panel of markers, including α-fetoprotein (AFP), Glypican 3 (GPC3) and SALL4, then clinicopathological parameters, pathomorphological characteristics, mucin phenotypes, molecular features, Immunoscore and survival analysis were assessed. A mixture of three types (COM + GAED + HAC) was most commonly observed in the same case, and typical transitions between each histological subtype were frequently seen. Hyaline globule and pink amorphous substance were often present. HER2 was amplified in 21.8% of cases. All the tumors showed intestinal phenotype (69.1%) and mixed gastric/intestinal phenotype (30.9%) and were all defined to chromosomal instable (CIN)/genomically stable (GS) group. Considering that 83.6% cases presented TP53 gene mutation phenotype and 61.8% cases showed ≥10% aberrant E-cadherin expression, the precise molecule classification is ambiguous. Survival analysis showed that patients with high SALL4 expression, high preoperative serum AFP level, or low Immunoscore had a significantly poor overall survival (OS). Moreover, SALL4, HER2, and Immunoscore had an independent influence on OS. In conclusion, we suggest that the development of gastric adenocarcinoma with hepatoid differentiation might a continuously progressive profile: from intestinal-type COM adenocarcinoma to GAED and then HAC. CIN/GS subtypes might be where they belonged. SALL4, HER2, and Immunoscore may be potential therapeutic targets.

Comparison of Effects of Sodium Bicarbonate and Sodium Carbonate on the Hydration and Properties of Portland Cement Paste.

Carbonates and bicarbonates are two groups of accelerators which can be used in sprayed concrete. In this study, the effects of the two accelerators sodium carbonate (Na2CO3) and sodium bicarbonate (NaHCO3) (0%, 1%, 2%, 3%, and 4% by weight of ordinary Portland cement OPC) on the properties of OPC paste were compared. The results show that both of them could accelerate the initial and final setting time of OPC paste, but the effect of the two accelerators on the compressive strength were different. After 1 day, sodium bicarbonate at 3% had the highest strength while sodium carbonate at 1% had the highest strength. After 7 days, both of the two accelerators at 1% had the highest compressive strength. After 28 days, the compressive strength decreased with the increase of the two. The improved strength at 1 and 7 days was caused by the accelerated formation of ettringite and the formation of CaCO3 through the reactions between the two with portlandite. The decrease of strength was caused by the Na⁺ could reduce the adhesion between C-S-H gel by replacing the Ca2+. NaHCO3 was found be a better accelerator than Na2CO3.

C5a induces A549 cell proliferation of non-small cell lung cancer via GDF15 gene activation mediated by GCN5-dependent KLF5 acetylation.

Non-small cell lung cancer (NSCLC) is the most common type of lung cancer, and multiple evidence has confirmed that C5a production is elevated in NSCLC microenvironment. Although NSCLC cell proliferation induced by C5a has been reported, the involved mechanism has not been elucidated. In this study, we examined the proliferation-related genes (i.e., KLF5, GCN5, and GDF15) and C5a receptor (C5aR) expression in tumor tissues as well as C5a concentration in plasma of NSCLC patients, and then determined the roles of KLF5, GCN5, and GDF15 in C5a-triggered NSCLC cell proliferation and the related mechanism both in vitro and in vivo. Our results found that the expression of KLF5, GCN5, GDF15, C5aR, and C5a was significantly upregulated in NSCLC patients. Mechanistic exploration in vitro revealed that C5a could facilitate A549 cell proliferation through increasing KLF5, GCN5, and GDF15 expression. Besides, KLF5 and GCN5 could form a complex, binding to GDF15 promoter in a KLF5-dependent manner and leading to GDF15 gene transcription. More importantly, GCN5-mediated KLF5 acetylation contributing to GDF15 gene transcription and cell proliferation upon C5a stimulation, the region (-103 to +58 nt) of GDF15 promoter which KLF5 could bind to, and two new KLF5 lysine sites (K335 and K391) acetylated by GCN5 were identified for the first time. Furthermore, our experiment in vivo demonstrated that the growth of xenograft tumors in BALB/c nude mice was greatly suppressed by the silence of KLF5, GCN5, or GDF15. Collectively, these findings disclose that C5a-driven KLF5-GCN5-GDF15 axis had a critical role in NSCLC proliferation and might serve as targets for NSCLC therapy.

Biomarkers of DNA Repair and Related Pathways: Significance of Treatment in Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) is a complex heterogeneous disease that lacks the expressions of hormone receptors (HR) and human epidermal growth factor receptor 2 (HER2). Although TNBC make up less than 20% of breast cancer, it accounts for a large number of metastatic cases and deaths. Currently, extensive efforts have been made to look for potentially biomolecular targets for TNBC treatment. Based on the differences of molecular events identified by gene profiling analysis, the TNBC may be divided into some broad categories: basal-like (BL), mesenchymal-like (ML), immune-associated, HER2-enriched and luminal/apocrine breast cancers. Most of these are in the BL-TNBC category. BL-TNBC carries a representative molecular event of DNA-repair deficiency that increases the effectiveness of DNA destabilizers (represented by platinum agents) and DNA-damage response inhibitors (represented by PARP inhibitors). However, the results from clinical and preclinical studies have been inconsistent. Herein, we simply outline the progress of breast cancer classification and the significance of DNA repair deficiency in the clinic treatment for TNBCs. Previous studies have shown that the neoadjuvant therapies with platinum agents are effective for early-stage and metastatic TNBC patients with DNA repair defects. The results indicate that proper biomarkers (such as homologous recombination repair defects, HRD) are necessary for predicting the response to chemotherapy and often sufficient to select patients in early-phase treatment. Furthermore, the combination of chemotherapy drugs and inhibitors of DNA damage response represents a potential therapeutic strategy for TNBCs.

Sublytic C5b-9 triggers glomerular mesangial cell apoptosis in rat Thy-1 nephritis via Gadd45 activation mediated by Egr-1 and p300-dependent ATF3 acetylation.

The apoptosis of glomerular mesangial cells (GMCs) is considered to be an important contributor to the initiation and development of rat Thy-1 nephritis (Thy-1N) and is accompanied by sublytic C5b-9 deposition. However, the mechanism by which sublytic C5b-9 triggers GMC apoptosis has not been elucidated. In this study, functional and histological examinations were performed on GMCs treated with sublytic C5b-9 (in vitro) and renal tissues of Thy-1N rats (in vivo). The in vitro studies found that sublytic C5b-9 could trigger GMC apoptosis through upregulating Egr-1, ATF3, and Gadd45 expression. Egr-1-mediated post-transcriptional modulation of ATF3, Egr-1/ATF3-enhanced Gadd45 promoter activity, and p300-mediated ATF3 acetylation were all involved in GMC apoptosis. More importantly, the effective binding elements for Egr-1 and ATF3 to Gadd45ß/γ promoters and the ATF3 acetylation site were identified. In vivo, silencing renal p300, Egr-1, ATF3, and Gadd45ß/γ significantly decreased GMC apoptosis, secondary GMC proliferation, and urinary protein secretion in Thy-1N rats. Together, these findings implicate that sublytic C5b-9-induced activation of Egr-1/p300-ATF3/Gadd45 axis plays a critical role in GMC apoptosis in Thy-1N rats.

C5a Induces the Synthesis of IL-6 and TNF-α in Rat Glomerular Mesangial Cells through MAPK Signaling Pathways.

Inflammatory response has been reported to contribute to the renal lesions in rat Thy-1 nephritis (Thy-1N) as an animal model of human mesangioproliferative glomerulonephritis (MsPGN). Besides C5b-9 complex, C5a is also a potent pro-inflammatory mediator and correlated to severity of various nephritic diseases. However, the role of C5a in mediating pro-inflammatory cytokine production in rats with Thy-1N is poorly defined. In the present studies, the levels of C5a, interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) were first determined in the renal tissues of rats with Thy-1N. Then, the expression of IL-6 and TNF-α was detected in rat glomerular mesangial cells (GMC) stimulated with our recombinant rat C5a in vitro. Subsequently, the activation of mitogen-activated protein kinase (MAPK) signaling pathways (p38 MAPK, ERK1/2 and JNK) and their roles in the regulation of IL-6 and TNF-α production were examined in the GMC induced by C5a. The results showed that the levels of C5a, IL-6 and TNF-α were markedly increased in the renal tissues of Thy-1N rats. Rat C5a stimulation in vitro could up-regulate the expression of IL-6 and TNF-α in rat GMC, and the activation of MAPK signaling pathways was involved in the induction of IL-6 and TNF-α. Mechanically, p38 MAPK activation promoted IL-6 production, while either ERK1/2 or JNK activation promoted TNF-α production in the GMC with exposure to C5a. Taken together, these data implicate that C5a induces the synthesis of IL-6 and TNF-α in rat GMC through the activation of MAPK signaling pathways.

A novel recombinant DNA vaccine encoding Mycobacterium tuberculosis ESAT-6 and FL protects against Mycobacterium tuberculosis challenge in mice.

Mycobacterium tuberculosis 6-kDa early secretory antigenic target (ESAT-6) is a dominant target antigen for cell-mediated immunity in the early phase of tuberculosis. The fms-like tyrosine kinase 3 ligand (FL) that induces potent immune response has been used as an adjuvant in vaccine development. In this study, a new recombinant plasmid (pIRES-epitope-peptides-FL) encoding three T cell epitopes of ESAT-6 and FL was constructed, and the immunogenicity of the DNA vaccine was assessed in C57BL/6 mice immunized with the plasmid DNA vaccine. Additionally, a strategy of intramuscular injection with the DNA vaccine (prime) and intranasal administration of the epitope peptides (boost) was employed to induce higher immune reaction of the mice. The results showed that mice vaccinated with the recombinant plasmid DNA vaccine and boosted with the peptides not only increased the levels of Th1 cytokines (IFN-γ and IL-12), the number of IFN-γ(+) T cells and activities of cytotoxic T lymphocytes as well as IgG, but also enhanced protection against Mycobacterium tuberculosis challenge. In conclusion, these data indicate that the novel recombinant pIRES-epitope-peptides-FL plasmid is a useful DNA vaccine for preventing Mycobacterium tuberculosis infection.