The aim of this study was to determine the dynamic changes of dendritic cell (DC) pheno-types and T cell in response to Echinococcus multilocularis (Em) infection in BALB/c mice. Mice comprised the control and Em-infected group. At day 0, 2, 7, 30, 60, 90 and 120 after infection, the size of larval cysts, the phenotype of DC and Th in splenocytes and the expression of CD40, CD86, TLR2 and TLR4, on DCs sulfur were examined. The results show that after 60 days' infection, larval cysts grow on the surface of liver, and they become larger over time. Compared with the control mice, MHC I and MHC II expressions on DC were significantly increased at day 7 (p < 0.05). At the same time, CD40, CD86, TLR2 and TLR4 increased rapidly, but after that they decreased gradually. At day 120, those markers were lower than in the control group. The ratio of CD4/CD8 was normal during 90 days of infection, while at day 120, a decline in CD4 T cell and increase in CD8 were foundleading to the inversion of the CD4/CD8 ratio. Our findings suggest that within the 120 days of Em infection, the major function of DC is to present antigens. Immune response is provided predominantly by Th1 cells, inducing host immune response against Em. However, after 120 days, DC matured and the function was suppressed. Furthermore, inversion of the CD4/CD8 ratio is beneficial to the growth of Em, thus favoring its immune evasion.

To evaluated the effect and mechanism of leflunomide (LEF) in murine model of collage-induced arthritis (CIA) on modulating the balance of Th17/Treg cells. DBA mice were divided into: control, CIA, CIA + LEF and CIA + Ibuprofen group. After 14 days immunization with bovine type II collagen (CII), except control group, mice were orally administered saline (CIA group), LEF or ibuprofen daily for 14 days. The severity of arthritis joint was assessed by using arthritis score. The numbers of CD3 + CD4 + interleukin 17 (IL-17) + Th17 cells and CD4 + CD25 + Foxp3+ Tregs were determined by flow cytometry, the expression of ROR-γt, Foxp3 and IL-21 mRNA in spleen were quantified by RT-PCR and IL-17, IL-21 and transforming growth factor β (TGF-β) level in serum was measured by ELISA. In comparison with CIA group, CIA + LEF and CIA + Ibuprofen group reduced the disease severity obviously (p < 0.05). Leflunomide reduced Th17 cells, the expression of ROR-γt and IL-21 mRNA (p < 0.05) and the levels of IL-17 and IL-21 in serum. In addition, Ibuprofen but not LEF has effect on Treg cells number, Foxp3 mRNA expression and TGF-β secretion. These results indicate that LEF effect on the balance od Th17/Treg through suppressing Th17 cells, but not stimulating Treg cells. Leflunomide may act as a potential immunomodulator for the treatment of rheumatoid arthritis (RA).

Live attenuated influenza vaccine (LAIV) candidates of the H7 subtype, A/Netherlands/219/03 (H7N7, NL03 ca) and A/chicken/British Columbia/CN-6/2004 (H7N3, BC04 ca), were evaluated for their receptor binding specificity and immunogenicity in ferrets. The BC04 ca virus exhibited α2,3-SA and α2,6-SA dual receptor binding preference while the NL03 ca virus preferentially bound to α2,3-SA. Substitution of the Q226 and G228 (Q-G) by the L226 and S228 (L-S) residues in the HA improved binding to α2,6-SA for NL03 ca. The vaccine viruses with L-S retained the attenuation phenotype. NL03 L-S ca replicated more efficiently than the original NL03 ca virus in the upper respiratory tract of ferrets, and induced higher levels of humoral and cellular immune responses. Prior vaccination with seasonal LAIV reduced H7-specific antibody responses, but did not reduce the H7N7 vaccine mediated protection against a heterologous H7N3 BC04 wt virus infection in ferrets. In addition, the H7N3 and H7N7 vaccine immunized ferret sera cross reacted with the newly emerged H7N9 virus. These data, in combination with the safety data from previously conducted Phase 1 studies, suggest that these vaccines may have a role in responding to the threat posed by the H7N9 virus.

The anaerobic, thermophilic bacterium, Clostridium thermocellum, secretes multi-protein enzyme complexes, termed cellulosomes, which synergistically interact with the microbial cell surface and efficiently disassemble plant cell wall biomass. C. thermocellum has also been considered a potential consolidated bioprocessing (CBP) organism due to its ability to produce the biofuel products, hydrogen, and ethanol. We found that C. thermocellum fermentation of pretreated yellow poplar (PYP) produced 30 and 39% of ethanol and hydrogen product concentrations, respectively, compared to fermentation of cellobiose. RNA-seq was used to analyze the transcriptional profiles of these cells. The PYP-grown cells taken for analysis at the late stationary phase showed 1211 genes up-regulated and 314 down-regulated by more than two-fold compared to the cellobiose-grown cells. These affected genes cover a broad spectrum of specific functional categories. The transcriptional analysis was further validated by sub-proteomics data taken from the literature; as well as by quantitative reverse transcription-PCR (qRT-PCR) analyses of selected genes. Specifically, 47 cellulosomal protein-encoding genes, genes for 4 pairs of SigI-RsgI for polysaccharide sensing, 7 cellodextrin ABC transporter genes, and a set of NAD(P)H hydogenase and alcohol dehydrogenase genes were up-regulated for cells growing on PYP compared to cellobiose. These genes could be potential candidates for future studies aimed at gaining insight into the regulatory mechanism of this organism as well as for improvement of C. thermocellum in its role as a CBP organism.

In order to assess whether N-Myc downstream regulated gene 4 (NDRG4) methylation was associated with the diagnosis and prognosis of gastric cancer, we measured the methylation of NDRG4 promoter and gene body regions among 110 gastric cancer patients using quantitative methods (MethyLight and pyrosequencing). Both NDRG4 promoter and gene body methylation levels were increased in tumor tissues than paired adjacent normal tissues (P < 0.001). NDRG4 gene body methylation was found to be significantly associated with age and tumor differentiation. NDRG4 promoter hypermethylation was proved to be a predictor of poor overall survival. However, opposite result was observed among The Cancer Genome Atlas (TCGA) cohort. The findings from gastric cell lines and public databases have suggested that NDRG4 methylation level was inversely associated with NDRG4 transcription level. Subsequent luciferase reporter gene assay showed that promoter CpG island but not gene body CpG island was able to upregulate gene expression. Collectively, NDRG4 promoter hypermethylation contributed to the risk of gastric cancer and predicted a poor prognosis in Chinese gastric cancer patients. Moreover, the combined methylation levels of NDRG4 promoter and gene body served as diagnostic biomarkers in gastric cancer.

Long interspersed nuclear element-1 (LINE-1 or L1) is a type of retrotransposons comprising 17% of the human and mouse genome, and has been found to be associated with several types of neurological disorders. Previous post-mortem brain studies reveal increased L1 copy number in the prefrontal cortex from schizophrenia patients. However, whether L1 retrotransposition occurs similarly in major depressive disorder (MDD) is unknown. Here, L1 copy number was measured by quantitative PCR analysis in peripheral blood of MDD patients (n = 105) and healthy controls (n = 105). The results showed that L1 copy number was increased in MDD patients possibly due to its hypomethylation. Furthermore, L1 copy number in peripheral blood and five brain regions (prefrontal cortex, hippocampus, amygdala, nucleus accumbens and paraventricular hypothalamic nucleus) was measured in the chronic unpredictable mild stress (CUMS) model of depression in mice. Intriguingly, increased L1 copy number in blood and the decreased L1 copy number in the prefrontal cortex were observed in stressed mice, while no change was found in other brain regions. Our results suggest that the changes of L1 may be associated with the pathophysiology of MDD, but the biological mechanism behind dysfunction of L1 retrotransposition in MDD remains to be further investigated.

Multidrug resistance remains a major obstacle to effective chemotherapy of colon cancer. ABCG2, as a half-transporter of the G subfamily of ATP-binding cassette transporter genes (ABC transporters), is known to play a crucial role in multidrug resistance. However, the molecular mechanism of controlling ABCG2 expression in drug resistance of colon cancer is unclear and scarcely reported. In the present study, we systematically investigate the potential role of the c-Jun NH2-terminal kinase (JNK) signal pathway in ABCG2-induced multidrug resistance in colon cancer. In the hydroxycamptothecin (HCPT) resistant cell line SW1116/HCPT from human colon cancer cell line SW1116, ABCG2 is the major factor for multidrug resistance, other than well-studied ABCB1 or ABCC1. Our findings indicate that blocking the JNK pathway by pathway inhibitor SP600125 reduces the expression level and transport function of ABCG2 in drug-resistant cells SW116/HCPT. Notably, the experiments of small interfering RNA directed against JNK1 and JNK2 show that only silence of JNK1 gene has the equal effect as SP600125 on dephosphorylation of transcription factor c-Jun and the expression of ABCG2 protein, while the corresponding phenomena were not observed after silence of JNK2 gene. Meanwhile, SP600125 induces the apoptosis of SW116/HCPT cells by promoting the cleavage of PARP and suppressing the anti-apoptotic protein survivin and bcl-2, and increases the sensitivity of SW1116/HCPT to HCPT. Taken together, our work demonstrated that JNK1/c-jun signaling pathway was involved in ABCG2-mediated multidrug resistance in colon cancer cells. Definitely, inhibition of the JNK1/c-jun pathway is useful for reversing ABCG2-mediated drug resistance in HCPT-resistant colon cancer cells.

Microorganisms employ a multiplicity of enzymes to efficiently degrade the composite structure of plant cell wall cellulosic polysaccharides. These remarkable enzyme systems include glycoside hydrolases (cellulases, hemicellulases), polysaccharide lyases, and the carbohydrate esterases. To accomplish this challenging task, several strategies are commonly observed either separately or in combination. These include free enzyme systems, multifunctional enzymes, and multi-enzyme self-assembled designer cellulosome complexes.

Retinoic acid-inducible gene I (RIG-I)-like receptors (RLRs) have recently been identified as cytoplasmic sensors for RNA virus. Recent research has shown that RIG-I, a member of this family, play an important role in innate immunity. In this study, we cloned the RIG-I gene from Jinding duck by reverse transcription polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE). We determined that the cDNA of duRIG-I contains a 14-bp 5' UTR, a 2802-bp open reading frame, and alternative 3' UTRs (295-bp and 927-bp) and encodes a polypeptide of 933 amino acids. Based on this sequence, the duRIG-I protein is predicted to have conserved domains typical of RLRs. In addition, duRIG-I was found to be distributed throughout DF1 cells by indirect immunofluorescence, as predicted. duRIG-I mRNA was scarcely detected in healthy tissues by semi-quantitative RT-PCR (sqRT-PCR). To study the role of RIG-I in innate immunity, we used synthetic double-stranded RNA to mimic viral infection in vivo and detected duRIG-I transcripts in spleen and liver by quantitative real-time PCR (qRT-PCR). The expression of duRIG-I mRNA was significantly elevated at 8h post-injection (P < 0.05) and was indistinguishable from control levels at other time points (P > 0.05). These results suggest that duRIG-I plays an important role in innate immune responses to double-stranded RNA viruses and warrant further studies to reveal the possible mechanism.

Although previous studies confirmed that steaming and the fermentation process could significantly improve the cognitive-enhancement and neuroprotective effects of Codonopsis lanceolata, the anti-tumor efficacy of steamed C. lanceolata (SCL) and what mechanisms are involved remain largely unknown. The present study was designed to evaluate the anti-tumor effect in vivo of SCL in H22 tumor-bearing mice. The results clearly indicated that SCL could not only inhibit the tumor growth, but also prolong the survival time of H22 tumor-bearing mice. Besides, the serum levels of cytokines, such as interferon gamma (IFN-γ), tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6) and interleukin-2 (IL-2), were enhanced by SCL administration. The observations of Hoechst 33258 staining demonstrated that SCL was able to induce tumor cell apoptosis. Finally, immunohistochemical analysis revealed that SCL treatment significantly increased Bax expression and decreased Bcl-2 and vascular endothelial growth factor (VEGF) expression of H22 tumor tissues in a dose-dependent manner. Moreover, LC/MS analysis of SCL indicated that it mainly contained lobetyolin and six saponins. Taken all together, the findings in the present study clearly demonstrated that SCL inhibited the H22 tumor growth in vivo at least partly via improving the immune functions, inducing apoptosis and inhibiting angiogenesis.

The Müllerian duct (MD) is the anlage of the oviduct, uterus and upper part of the vagina, the main parts of the female reproductive tract. Several wingless-type mouse mammary tumor virus (MMTV) integration site family member (Wnt) genes, including Wnt4, Wnt5a and Wnt7a, are involved in the development of MD and its derivatives, with Wnt4 particularly critical, since the MD fails to develop in its absence. We use, here, Wnt4(EGFPCre)-based fate mapping to demonstrate that the MD tip cells and the subsequent MD cells are derived from Wnt4+ lineage cells. Moreover, Wnt4 is required for the initiation of MD-forming cell migration. Application of anti-Wnt4 function-blocking antibodies after the initiation of MD elongation indicated that Wnt4 is necessary for the elongation as well, and consistent with this, cell culture wound-healing assays with NIH3T3 cells overexpressing Wnt4 promoted cell migration by comparison with controls. In contrast to the Wnt4 null embryos, some Wnt4(monomeric cherry/monomeric cherry) (Wnt4(mCh/mCh)) hypomorphic mice survived to adulthood and formed MD in ∼45% of cases. Nevertheless, the MD of the Wnt4(mCh/mCh) females had altered cell polarization and basement membrane deposition relative to the controls. Examination of the reproductive tract of the Wnt4(mCh/mCh) females indicated a poorly coiled oviduct, absence of the endometrial glands and an undifferentiated myometrium, and these mice were prone to develop a hydro-uterus. In conclusion, the results suggest that the Wnt4 gene encodes signals that are important for various aspects of female reproductive tract development.

A genome-wide association study of Han Chinese samples identified three single-nucleotide polymorphisms in the tetraspanin 18 (TSPAN18) gene to be associated with schizophrenia. However, the replication of the TSPAN18 association was inconsistent across studies. To explore the possible reason for poor replication, we conducted a case-control study to validate the TSPAN18 finding in an independent Chinese sample. The frequency of rs11038167 minor allele (A) was significantly higher only in female patients with thought disorder. Our result suggested that the TSPAN18 gene may be involved in the development of psychotic symptoms and contribute to clinical heterogeneity of schizophrenia.

Piwi-interacting RNAs (piRNAs) play a key role in spermatogenesis. Here, we describe the piRNAs profiling of primordial germ cells (PGCs), spermatogonial stem cells (SSCs), and the spermatogonium (Sp) during early-stage spermatogenesis in chicken. We obtained 31,361,989 reads from PGCs, 31,757,666 reads from SSCs, and 46,448,327 reads from Sp cells. The length distribution of piRNAs in the three samples showed peaks at 33 nt. The resulting genes were subsequently annotated against the Gene Ontology (GO) database. Five genes (RPL7A, HSPA8, Pum1, CPXM2, and PRKCA) were found to be involved in cellular processes. Interactive pathway analysis (IPA) further revealed three important pathways in early-stage spermatogenesis including the FGF, Wnt, and EGF receptor signaling pathways. The gene Pum1 was found to promote germline stem cell proliferation, but it also plays a role in spermatogenesis. In conclusion, we revealed characteristics of piRNAs during early spermatogonial development in chicken and provided the basis for future research.

The geese have strong broodiness and poor egg performance. These characteristics are the key issues that hinder the goose industry development. Yet little is known about the mechanisms responsible for follicle development due to lack of genomic resources. Hence, studies based on high-throughput sequencing technologies are needed to produce a comprehensive and integrated genomic resource and to better understand the biological mechanisms of goose follicle development.

Background: Fibroblast proliferation is a critical feature during heart failure development. Previous studies reported regulatory T-lymphocytes (Tregs)' protective role against myocardial fibrosis. However, notably, Tregs also secrete fibrogenic cytokine TGF-β when activated. This study aimed to clarify the intriguing link between Tregs and fibrosis, the role of Tregs Kv1.3 potassium channel (regulating T-lymphocytes activation) in the fibrosis process, and how selective aldosterone receptor antagonist Eplerenone affects Tregs and fibrosis through its action on Kv1.3 channel. Methods and Results: After co-incubation with Tregs, cardiac fibroblast proliferation (CCK-8 assay) and levels of collagen I, III, and Matrix metalloproteinase2 (ELISA) significantly elevated. Cell viability assays, Kv1.3 channel mRNA (RT-qPCR), and protein expression (In-Cell Western Blotting) revealed Tregs were activated/proliferated when co-cultured with fibroblasts. Treg intracellular TGF-β level increased by 5.8-fold, far more than that of intracellular IL-10, extracellular TGF-β and IL-10 (ELISA). And 30 μM eplerenone suppressed Tregs proliferation by 82.77% and furthermore, suppressed intracellular TGF-β level to a significantly greater extent than that of intracellular IL-10, extracellular TGF-β and IL-10. Moreover, the Kv1.3 current (whole-cell patch clamp) of Tregs in congestive heart failure patients and rats (induced by coronary artery ligation and exhaustive exercise) elevated by >4-fold than that of healthy volunteers and control rats, whereas 30 μM eplerenone suppressed the current by >60% in control Tregs. In addition, docking calculations (AutoDock software 4.0 suite) showed eplerenone has higher H-bond energy with Kv1.3 channel than other selective blockers. Conclusion: Immuno-regulation in the late stage of CHF activates Tregs proliferation via the upregulation of Kv1.3 channels, which promotes cardiac fibrosis by primarily secreting TGF-β. Taken together, eplerenone's high affinity to Kv1.3 channel enables it to antagonize the Kv1.3 channels directly to suppress Tregs proliferation, which in turn may play an immuno-regulatory role during CHF.

The RNA polymerase complex (RNApc) in influenza A viruses (IVs) is composed of the PB2, PB1 and PA subunits, which are encoded by the three longest genome segments (Seg1-3) and are responsible for the replication of vRNAs and transcription of viral mRNAs. However, the co-evolutionary relationships of the three segments from the known 126 subtypes IVs are unclear. In this study, we performed a detailed analysis based on a total number of 121,191 nucleotide sequences. Three segment sequences were aligned before the repeated, incomplete and mixed sequences were removed for homologous and phylogenetic analyses. Subsequently, the estimated substitution rates and TMRCAs (Times for Most Recent Common Ancestor) were calculated by 175 representative IVs. Tracing the cladistic distribution of three segments from these IVs, co-evolutionary patterns and trajectories could be inferred. The further correlation analysis of six internal protein coding segments reflect the RNApc segments have the closer correlation than others during continuous reassortments. This global approach facilitates the establishment of a fast antiviral strategy and monitoring of viral variation.

Diabetes is a systemic disease that can cause brain damage such as synaptic impairments in the hippocampus, which is partly because of neuroinflammation induced by hyperglycemia. Brain-derived neurotrophic factor (BDNF) is essential in modulating neuroplasticity. Its role in anti-inflammation in diabetes is largely unknown. In the present study, we investigated the effects of BDNF overexpression on reducing neuroinflammation and the underlying mechanism in mice with type 1 diabetes induced by streptozotocin (STZ). Animals were stereotactically microinjected in the hippocampus with recombinant adeno-associated virus (AAV) expressing BDNF or EGFP. After virus infection, four groups of mice, the EGFP+STZ, BDNF+STZ, EGFP Control and BDNF Control groups, received STZ or vehicle treatment as indicated. Three weeks later brain tissues were collected. We found that BDNF overexpression in the hippocampus significantly rescued STZ-induced decreases in mRNA and protein expression of two synaptic plasticity markers, spinophilin and synaptophysin. More interestingly, BDNF inhibited hyperglycemia-induced microglial activation and reduced elevated levels of inflammatory factors (TNF-α, IL-6). BDNF blocked the increase in HMGB1 levels and specifically, in levels of one of the HMGB1 receptors, RAGE. Downstream of HMGB1/RAGE, the increase in the protein level of phosphorylated NF-κB was also reversed by BDNF in STZ-treated mice. These results show that BDNF overexpression reduces neuroinflammation in the hippocampus of type 1 diabetic mice and suggest that the HMGB1/RAGE/NF-κB signaling pathway may contribute to alleviation of neuroinflammation by BDNF in diabetic mice.

Non-coding RNAs have attracted considerable attention for their vital role in cancer. The purpose of this study was to determine the effects of non-coding RNAs on hepatocellular carcinoma (HCC) and reveal their regulatory mechanism in the pathophysiological process.

Moderate exercise can alleviate symptoms of osteoarthritis (OA) such as pain, stiffness, and joint deformities that are associated with progressive cartilaginous degeneration, osteophyte formation, subchondral bone changes, and synovial inflammation. Irisin is an exercise-related myokine that reportedly plays a crucial role in bone remodeling. However, its role in OA remains unknown. This study aimed to determine whether irisin can attenuate OA progression and the mechanism of its therapeutic effect. Three-month-old male C57BL/6J mice were randomized to groups that underwent sham operation, and anterior cruciate ligament transection (ACLT) intraperitoneally injected with vehicle or irisin in vivo. Apoptosis was induced by stretching murine osteocyte-like MLO-Y4 cells in vitro. Irisin reduced wear, maintained the proportion of hyaline cartilage, a more complete cartilage structure, and lower Osteoarthritis Research Society International (OARSI) scores at 4 weeks after ACLT. Irisin reduced the expression of matrix metalloproteinase (MMP)-13 in cartilage and caspase 3 in the subchondral bone. Irisin exerted rescue effects in microstructural parameters of subchondral trabecular bone including bone volume fraction (BV/TV), trabecular number (Tb.N), connection density (Conn. D), and the structure model index (SMI) compared with ACLT-vehicle group. Bone histomorphometry showed that irisin increased subchondral bone remodeling. The decreasing ratio (%) of the eroded surface (ES/BS) was reversed by irisin in the ACLT+vehicle group. Staining with tartrate-resistant acid phosphatase showed a decreased number of osteoclasts. Irisin significantly increased the proliferation of osteocytes, protected them from apoptosis, and maintained cellular activity by regulating the expression of Bax, Bcl-2, and osteoprotegerin/receptor activator of nuclear factor (NF)-kB-ligand (OPG/Rankl). Irisin activated serine/threonine-selective protein kinases (Erk) and p38 signaling, and its anti-apoptosis function depended on the Erk signaling pathway. Irisin attenuated OA progression by decreasing osteocyte apoptosis and improving the microarchitecture of subchondral bone. Activation of the Erk pathway by irisin plays an important role in reducing osteocyte apoptosis in vitro.

Glycoside hydrolase (GH) family 48 is an understudied and increasingly important exoglucanase family found in the majority of bacterial cellulase systems. Moreover, many thermophilic enzyme systems contain GH48 enzymes. Deletion of GH48 enzymes in these microorganisms results in drastic reduction in biomass deconstruction. Surprisingly, given their importance for these microorganisms, GH48s have intrinsically low cellulolytic activity but even in low ratios synergize greatly with GH9 endoglucanases. In this study, we explore the structural and enzymatic diversity of these enzymes across a wide range of temperature optima. We have crystallized one new GH48 module from Bacillus pumilus in a complex with cellobiose and cellohexaose (BpumGH48). We compare this structure to other known GH48 enzymes in an attempt to understand GH48 structure/function relationships and draw general rules correlating amino acid sequences and secondary structures to thermostability in this GH family.