To study the structure-activity relationship (SAR) of Toll-like receptor 7 (TLR-7) agonists based on 8-oxoadenines, a novel subset of C9-substituted 8-hydroxy-2-(2-methoxyethoxy)-adenines and their antigen conjugates were synthesized. In vitro, the ability of cytokines (IL-12p70 and IFN-γ) induction of ligands with alkyl acid at C9-position were very weak compared with benzoic acid counter parts. Unexpectedly, its antigen conjugates that conjugated with proteins or peptides with weak immunogenicity, showed enhanced activity of cytokines induction. After administered systemically in mice in vivo, all conjugates induced prolonged increase in pro-inflammatory cytokines and antigen-specific IgG levels in serum compared with free compounds. Results from molecular dynamics (MD) simulations further confirmed the conclusion and provided the details of interaction to explain the phenomenon of experiment. In conclusion, we discovered that TLR-7 could be activated via some conjugates of weak ligand and weak antigen, which could be safer adjuvant candidates for vaccines in the future.

Major depression is a highly prevalent disorder with no effective medical treatments available. Recent evidence has shown that sirtuins (SIRTs) signaling has been implicated to play an essential in the pathogenesis of depression. Here in this study, we aimed to investigate the potential role of the phencynonate hydrochloride (PHH) in rat models of chronic unpredictable mild stress (CUMS)-induced depression. SIRT6 expression was up-regulated by PHH via increasing NAD+/NADH ratio in the prefrontal cortex. PHH was able to suppress CUMS-induced oxidative stress and enhance the antioxidant capacity and antioxidant proteins activity, such as superoxide dismutase 2 (SOD2) and peroxiredoxin 6 (Prdx6). In vitro study, we found that SIRT6 directly bound to SOD2 and Prdx6 and deacetylated them at Lys68/122 and Lys63/209, which were acetylated by p300/CBP-associated factor (PCAF). Finally, we showed that PHH ameliorated CUMS-induced depressive phenotypes by up-regulating SIRT6 deacetylation activity. In summary, PHH-mediating SIRT6 pathway is required for antidepressant response and PHH can be used as a novel therapeutic to effectively treat depression.

The aim of this study is to explore the potential neuroprotective effects of Ginkgolide B (GB), a main terpene lactone and active component in Ginkgo biloba, in hypoxia-induced neuronal damage, and to further investigate its possible mechanisms.

The aim of this protocol is to provide the methods used to assess the effectiveness and safety of acupuncture for the treatment of patients with Alzheimer's disease.

The expression of microRNA-206 (miR-206) is high in skeletal muscle but low in most other tissues. The expression of miR-206 is increased in muscular dystrophy, suggesting its involvement in the pathogenesis of muscle diseases. To determine the role of miR-206 in muscle cell differentiation and explore a possible gene therapy vector, we constructed a miR-206 adenoviral expression vector (AdvmiR-206) and tested for transfection into C2C12 stem cells.

Here, two temperature sensitive promoters, P2 and P7, isolated from Bacillus subtilis, were characterized. The production of beta-galactosidase driven by these promoters was much higher at 45 degrees C than that at 37 degrees C both in Escherichia coli and B. subtilis and that the P2 promoter showed higher expression strength in B. subtilis at 45 degrees C. Thereby, an efficient temperature-inducible expression system was constructed by using P2 promoter in B. subtilis. Thus, we isolated and characterized a newly temperature inducible promoter and exploited it as a potential expression element in B. subtilis.

Combination therapy with inhibitors of cytotoxic T lymphocyte-associated protein (CTLA)4 and programmed death (PD)-1 has demonstrated efficacy in cancer patients. However, there is little information on CTLA4 and PD-1 expression levels and their clinical significance across diverse cancers. In this study, we addressed this question by analyzing PD-1 and CTLA4 levels in 33 different types of cancer along with their prognostic significance using The Cancer Genome Atlas (TCGA) and Cancer Cell Line Encyclopedia datasets. Liver hepatocellular carcinoma (LIHC) patients receiving cytokine-induced killer cell (CIK) immunotherapy at Sun Yat-sen University cancer center were enrolled for survival analysis. The correlation between PD-1/CTLA4 expression and cancer immunity was also analyzed. The results showed that PD-1 and CTLA4 transcript levels varied across cancer cell lines, with aberrant expression detected in certain cancer types; Kaplan-Meier analysis with the Cox proportional hazards model showed that this was closely related to overall survival in breast invasive carcinoma, glioblastoma multiforme, head and neck squamous cell carcinoma, acute myeloid leukemialymphoma, uterine corpus endometrial carcinoma, and uveal melanoma in TCGA. High serum PD-1 and CTLA4 levels predicted better survival in LIHC patients receiving CIK therapy. PD-1 and CTLA4 levels were found to be significantly correlated with the degree of tumor cell infiltration using Tumor Immune Estimation Resource, Estimating Relative Subsets of RNA Transcripts, and Estimation of Stromal and immune Cells in Malignant Tumor Tissues Using Expression Data as well as with tumor-infiltrating lymphocyte marker expression; they were also related to tumor mutation burden, microsatellite instability, mismatch repair, and the expression of DNA methyltransferases in some cancer types. Gene set enrichment analysis of 33 cancer types provided further evidence for associations between PD-1/CTLA4 levels and cancer development and immunocyte infiltration. Thus, PD-1 and CTLA4 play important roles in tumorigenesis and tumor immunity and can serve as prognostic biomarkers in different cancer types.

Basic processes of the fatty acid metabolism have an important impact on the function of intestinal epithelial cells (IEC). However, while the role of cellular fatty acid oxidation is well appreciated, it is not clear how de novo fatty acid synthesis (FAS) influences the biology of IECs. We report here that interfering with de novo FAS by deletion of the enzyme Acetyl-CoA-Carboxylase (ACC)1 in IECs results in the loss of epithelial crypt structures and a specific decline in Lgr5+ intestinal epithelial stem cells (ISC). Mechanistically, ACC1-mediated de novo FAS supports the formation of intestinal organoids and the differentiation of complex crypt structures by sustaining the nuclear accumulation of PPARδ/β-catenin in ISCs. The dependency of ISCs on cellular de novo FAS is tuned by the availability of environmental lipids, as an excess delivery of external fatty acids is sufficient to rescue the defect in crypt formation. Finally, inhibition of ACC1 reduces the formation of tumors in colitis-associated colon cancer, together highlighting the importance of cellular lipogenesis for sustaining ISC function and providing a potential perspective to colon cancer therapy.

Ulcerative colitis (UC), one of the typical inflammatory bowel diseases caused by dysregulated immunity, still requires novel therapeutic medicine with high efficacy and low toxicity. Hericium erinaceus has been widely used to treat different health problems especially gastrointestinal sickness in China for thousands of years. Here, we isolated, purified, and characterized a novel low weight polysaccharide (HEP10, Mw: 9.9 kDa) from the mycelia of H. erinaceus in submerged culture. We explored the therapeutic effect of HEP10 on UC and explored its underlying mechanisms. On one hand, HEP10 suppressed the production of TNF-α, IL-1β, IL-6, inducible iNOS, and COX-2 in LPS challenged murine macrophage RAW264.7 cells, as well as in colons from DSS-induced colitis mice. On the other hand, HEP10 treatment markedly suppressed the activation of NLRP3 inflammasome, NF-κB, AKT, and MAPK pathways. Moreover, HEP10 reversed DSS-induced alternation of the gut community composition and structure by significantly increasing Akkermansia muciniphila and also promoting functional shifts in gut microbiota. Structural equation modeling also highlighted that HEP10 can change widely through gut microbiota. In conclusion, HEP10 has a better prebiotic effect than the crude polysaccharides of H. erinaceus, which can be used as a novel dietary supplement and prebiotic to ameliorate colitis.

Interferon-gamma (IFN-γ) plays a complex role in modulating tumor microenvironment during lung adenocarcinoma (LUAD) development. In order to define the role of IFN-γ response genes in LUAD progression, we characterized the gene expression, mutation profile, protein-protein interaction of 24 IFN-γ response genes, which exhibited significant hazard ratio in overall survival. Two subgroups of LUAD from the TCGA cohort, which showed significant difference in the survival rate, were identified based on the expression of these genes. Furthermore, LASSO penalized cox regression model was used to derive a risk signature comprising seven IFN-γ response genes, including CD74, CSF2RB, PTPN6, MT2A, NMI, LATS2, and PFKP, which can serve as an independent prognostic predictor of LUAD. The risk signature was validated in an independent LUAD cohort. The high risk group is enriched with genes regulating cell cycle and DNA replication, as well as a high level of pro-tumor immune cells. In addition, the risk score is negatively correlated with the expression of immune metagenes, but positively correlated with DNA damage repair genes. Our findings reveal that seven-gene risk signature can be a valuable prognostic predictor for LUAD, and they are crucial participants in tumor microenvironment of LUAD.

Rationale: Epithelial-mesenchymal transition (EMT) has been recognized as an important step toward high invasion and metastasis of many cancers including hepatocellular carcinoma (HCC), while the mechanism for EMT promotion is still ambiguous. Methods: The dynamic alterations of site-specific glycosylation during HGF/TGF-β1-induced EMT process of three HCC cell lines were systematically investigated using precision glycoproteomic methods. The possible roles of EMT-related glycoproteins and site-specific glycans were further confirmed by various molecular biological approaches. Results: Using mass spectrometry-based glycoproteomic methods, we totally identified 2306 unique intact glycopeptides from SMMC-7721 and HepG2 cell lines, and found that core-fucosylated glycans were accounted for the largest proportion of complex N-glycans. Through quantification analysis of intact glycopeptides, we found that the majority of core-fucosylated intact glycopeptides from folate receptor α (FOLR1) were up-regulated in the three HGF-treated cell lines. Similarly, core-fucosylation of FOLR1 were up-regulated in SMMC-7721 and Hep3B cells with TGF-β1 treatment. Using molecular approaches, we further demonstrated that FUT8 was a driver for HGF/TGF-β1-induced EMT. The silencing of FUT8 reduced core-fucosylation and partially blocked the progress of HGF-induced EMT. Finally, we confirmed that the level of core-fucosylation on FOLR1 especially at the glycosite Asn-201 positively regulated the cellular uptake capacity of folates, and enhanced uptake of folates could promote the EMT of HCC cells. Conclusions: Based on the results, we proposed a potential pathway for HGF or TGF-β1-induced EMT of HCC cells: HGF or TGF-β1 treatment of HCC cells can increase the expression of glycosyltransferase FUT8 to up-regulate the core-fucosylation of N-glycans on glycoproteins including the FOLR1; core-fucosylation on FOLR1 can then enhance the folate uptake capacity to finally promote the EMT progress of HCC cells.

Although embryonic stem cells or induced pluripotent stem cells are able to differentiate into inner ear hair cells (HCs), they have drawbacks limiting their clinical application, including a potential risk of tumourigenicity. Direct reprogramming of fibroblasts to inner ear HCs could offer an alternative solution to this problem. Here, we present a stepwise guidance protocol to induce mouse embryonic fibroblasts to differentiate into inner ear HC-like cells (HCLs) via mesenchymal-to-epithelial transition and then acquisition of otic sensory epithelial cell traits by overexpression of three key transcription factors. These induced HCLs express multiple HC-specific proteins, display protrusions reminiscent of ciliary bundle structures, respond to voltage stimulation, form functional mechanotransduction channels, and exhibit a transcriptional profile of HC signature. Together, our work provides a new method to produce functional HCLs in vitro, which may have important implications for studies of HC development, drug discovery, and cell replacement therapy for hearing loss.

Interleukin-17A (IL-17A) produced by Th17 cells, contributes to the pathogenesis of various autoimmune diseases by stimulating the release of cytokines and chemokines and its regulation. Anti-IL-17A antibody which blocks the function of IL-17A has been proved to be an effective treatment of autoimmune disease. The aim of our study was to generate a potential humanized anti-IL-17A therapeutic monoclonal antibody (mAb) through a comprehensive panel of in vitro and in vivo biological activity studies, as well as physicochemical characterization. HZD37-5, a humanized monoclonal antibody specifically recognizing N78 loci of IL-17A, binds to human and rhesus monkeys, blocks IL-17 induced signal transduction and the release of IL-6, IL-8, CXCL-1 and G-GSF. In an in vivo efficacy mouse model, HZD37-5 significantly inhibited human IL-17A induced-keratinocyte chemoattractant (KC) secretion in a dose-dependent manner. The pharmacokinetics (PK) study result of HZD37-5 in rhesus monkeys indicated that HZD37-5 had favorable PK characteristics with limited distribution (78.0-78.8 ml/kg), slow elimination (5.00-6.45 ml/day/kg), long half-life (9.1-10.7 days) and high bioavailability (103%) following a single IV or SC dose at 1.5 mg/kg. These findings provided a comprehensive preclinical characterization of HZD37-5 and supported that it may be developed as a potential therapeutic for the treatment of autoimmune diseases, including psoriasis, psoriatic arthritis, axial spondyloarthritis, etc.

Wilm's tumor is a common renal malignancy in childhood with unsatisfactory prognosis. microRNA-215-5p (miR-215-5p) has been reported as a tumor-suppressive miRNA in different types of human cancers, but rarely in the Wilm's tumor. In light of this, we tried to investigate the regulatory role and underlying mechanism of miR-215-5p in the Wilm's tumor.

Sepsis is one of the possible outcomes of severe trauma, and it poses a dire threat to human life, particularly in immunocompromised people. The most prevalent pathogens are bacteria and fungi, but viruses should not be overlooked. For viral metagenomic analysis, we collected blood samples from eight patients with post-traumatic sepsis before and seven days after treatment. The results demonstrated that Anellovirus predominated the viral community, followed by Siphoviridae and Myoviridae, and that the variations in viral community and viral load before and after treatment were not statistically significant. This study allows us to investigate methods for establishing NGS-based viral diagnostic instruments for detecting viral infections in the blood of sepsis patients so that antiviral therapy can be administered quickly.

Conventional chemotherapy and radiotherapy for the treatment of lymphoma have notable drawbacks, and passive immunotherapy using a monoclonal antibody is restricted to CD20-positive B cell lymphoma. Therefore, new treatment types are urgently required, especially for T cell lymphoma. One type of new antitumour therapy is the use of active immunotherapeutic agents, such as agonists of the Toll-like receptors (TLRs), which facilitate the induction of prolonged antitumour immune responses.

The aim of the study was to examine the association among advanced glycation end products (AGEs), extracellular matrix metalloproteinase inducer (EMMPRIN) and matrix metalloproteinase (MMPs), and investigate whether AGEs affect type I collagen (COL-I) through EMMPRIN or MMPs. A co-culture system with the osteoblast-like cells (MC3T3E1) and mouse RAW264.7 cells was employed to examine the effects of AGE-bovine serum albumin (BSA) (50 mg/l), EMMPRIN antibody (5 mg/l) and AGE-BSA+EMMPRIN antibody separately on COL-I expression for 24 h. Culture media were analyzed for the content of COL-I by ELISA. The effect of different concentrations of AGE-BSA (0, 50, 100, 200 and 400 mg/l) for 24 h was assessed on COL-I levels. Finally, semiquantitative RT-PCR was used to detect the osteoblast COL-I mRNA expression and MMP-2 and MMP-9's PMAO were also measured in the culture medium. COL-I content in the culture medium decreased significantly following treatment with AGE-BSA (P<0.05). EMMPRIN antibody increased COL-I content (P<0.05). EMMPRIN antibody+AGE-BSA increased COL-I significantly (P<0.05). Different concentrations of AGE-BSA increased COL-I mRNA expression significantly compared with the control group (P<0.05), and were enhanced with increasing AGE-BSA concentration (P<0.05). Also MMP-2 and MMP-9 secretion increased significantly (P<0.05), with the increasing AGE-BSA concentration. In conclusion, an increase in AGE levels in vitro stimulates the secretion of EMMPRIN/MMPs, promotes the degradation of COL-I and reduces bone strength.

MiRNAs are a class of non-coding small RNAs that play important roles in the regulation of gene expression. Although plant miRNAs have been extensively studied in model systems, less is known in other plants with limited genome sequence data, including eggplant (Solanum melongena L.). To identify miRNAs in eggplant and their response to Verticillium dahliae infection, a fungal pathogen for which clear understanding of infection mechanisms and effective cure methods are currently lacking, we deep-sequenced two small RNA (sRNA) libraries prepared from mock-infected and infected seedlings of eggplants. Specifically, 30,830,792 reads produced 7,716,328 unique miRNAs representing 99 known miRNA families that have been identified in other plant species. Two novel putative miRNAs were predicted with eggplant ESTs. The potential targets of the identified known and novel miRNAs were also predicted based on sequence homology search. It was observed that the length distribution of obtained sRNAs and the expression of 6 miRNA families were obviously different between the two libraries. These results provide a framework for further analysis of miRNAs and their role in regulating plant response to fungal infection and Verticillium wilt in particular.

Recurrence and metastasis are the main causes of death for prostate cancer patients and cancer stem cells (CSCs) are proposed to play important roles in cancer recurrence and metastasis. It is generally thought that genes upregulated in recurrent/metastatic disease are likely biomarkers of CSCs. Hence we analyzed multiple microarray datasets on prostate tumor tissues to identify upregulated genes associated with cancer recurrence/metastasis, and tried to explore whether those genes were true biomarkers of prostate CSCs. Our results indicated that TOP2A was the most highly upregulated gene in recurrent/metastatic prostate cancer, and its high expression was positively correlated with poor prognosis in patients. Using a promoter reporter system, we unexpectedly discovered enrichment of CSCs in TOP2Aneg cells. Compared to TOP2Ahigh cells, TOP2Aneg cells formed spheres and tumors more efficiently, and became enriched in the presence of stresses. Analysis of cell divisions by time lapse imaging indicated that more slow-cycling cells were observed in TOP2Aneg cells while the proportion of abnormal divisions was higher in TOP2Ahigh cells. Our studies demonstrate that TOP2Ahigh is the phenotype of recurrence/metastasis but TOP2Aneg cells show slow cycling and have CSCs properties in prostate cancer, which has significant implications for prostate cancer therapy.

Although human umbilical cord mesenchymal stem cells (hUC-MSCs) have been identified as a new source of MSCs for potential application in regenerative medicine, their full potential of differentiation has not been determined. In particular, whether they have the capability to differentiate into epithelial cells of endodermal origin such as the prostate epithelial cells is unknown. Here we report that when hUC-MSCs were combined with rat urogenital sinus stromal cells (rUGSSs) and transplanted into the renal capsule in vivo, they could differentiate into prostate epithelial-like cells that could be verified by prostate epithelial cell-specific markers including the prostate specific antigen. The prostatic glandular structures formed in vivo displayed similar cellular architecture with lumens and branching features as seen for a normal prostate. In addition, the human origin of the hUC-MSCs was confirmed by immunocytochemistry for human nuclear antigen. These findings together indicate that hUC-MSCs have the capability to differentiate into epithelial-like cells that are normally derived from the endoderm, implicating their potential applications in tissue repair and regeneration of many endoderm-derived internal organs.