This study was to evaluate the fermentation dynamics, structural and nonstructural carbohydrate composition and in vitro gas production of rice straw ensiled with lactic acid bacteria and molasses.

Glucose metabolism plays a key role in thymocyte development. The mammalian target of rapamycin complex 2 (mTORC2) is a critical regulator of cell growth and metabolism, but its role in early thymocyte development and metabolism has not been fully studied. We show here that genetic ablation of Sin1, an essential component of mTORC2, in T lineage cells results in severely impaired thymocyte development at the CD4-CD8- double negative (DN) stages but not at the CD4+CD8+ double positive (DP) or later stages. Notably, Sin1-deficient DN thymocytes show markedly reduced proliferation and glycolysis. Importantly, we discover that the M2 isoform of pyruvate kinase (PKM2) is a novel and crucial Sin1 effector in promoting DN thymocyte development and metabolism. At the molecular level, we show that Sin1-mTORC2 controls PKM2 expression through an AKT-dependent PPAR-γ nuclear translocation. Together, our study unravels a novel mTORC2-PPAR-γ-PKM2 pathway in immune-metabolic regulation of early thymocyte development.

Vascular endothelial (VE)-cadherin forms homotypic adherens junctions (AJs) in the endothelium, whereas N-cadherin forms heterotypic adhesion between endothelial cells and surrounding vascular smooth muscle cells and pericytes. Here we addressed the question whether both cadherin adhesion complexes communicate through intracellular signaling and contribute to the integrity of the endothelial barrier. We demonstrated that deletion of N-cadherin (Cdh2) in either endothelial cells or pericytes increases junctional endothelial permeability in lung and brain secondary to reduced accumulation of VE-cadherin at AJs. N-cadherin functions by increasing the rate of VE-cadherin recruitment to AJs and induces the assembly of VE-cadherin junctions. We identified the dual Rac1/RhoA Rho guanine nucleotide exchange factor (GEF) Trio as a critical component of the N-cadherin adhesion complex, which activates both Rac1 and RhoA signaling pathways at AJs. Trio GEF1-mediated Rac1 activation induces the recruitment of VE-cadherin to AJs, whereas Trio GEF2-mediated RhoA activation increases intracellular tension and reinforces Rac1 activation to promote assembly of VE-cadherin junctions and thereby establish the characteristic restrictive endothelial barrier.

Hepatoma is one of the most common malignant tumors, and most patients have very poor prognosis. Early prediction and intervention of the hepatoma recurrence/metastasis are the most effective way to improve the patients' clinical outcomes. Here, we used isobaric tags for relative and absolute quantitation (iTRAQ) based quantitative phospho-proteomics approach to identify biomarkers associated with hepatoma recurrence/metastasis in hepatoma cell lines with increasing metastasis ability. In total, 75 phosphorylated peptides corresponding to 60 phosphoproteins were significantly dysregulated and the participated biological processes of these phosphoproteins were tightly associated with tumor metastasis. Further signaling pathway analysis revealed that key signaling pathways which play crucial roles in cancer metastasis have been significantly over activated in the highly metastatic cells. Furthermore, the phosphorylation of FLNASer2152 and ANXA2Tyr23 were validated to be significantly up regulated in the high-metastatic cells comparing with the low-metastatic cells. By further investigation the clinical significance of the phosphorylation of FLNASer2152 and ANXA2Tyr23 in large-scale clinical samples, revealed that the over phosphorylation of FLNASer2152 and ANXA2Tyr23 were associated with poor prognosis and might be potential prognostic biomarkers for the primary hepatoma. When FLNASer2152 combined with ANXA2Tyr23, it had a better prognostic value for both OS and TTR.

To analyse the factors predicting uncontrolled seizures in epilepsy with auditory features (EAF).

Traditional Chinese medicine (TCM) has been practiced for thousands of years, although concerns about the efficacy, legality, and safety of TCM continue to be raised. Chromatographic studies have detected the presence of heavy metals and plant toxins within some TCM preparations. However, chromatography is not able to identify all of the compounds of TCM, particularly those items that are not clearly labeled on the packaging. The present study aimed to establish a supplemental method that better assesses the ingredient components of TCM preparations.We established an effective approach to screen the biological and toxical composition of TCM based on high-throughput sequencing (HTS), as well as fast detection and validation of the toxical species by real-time PCR, based on ITS2 DNA barcoding. Ruyi jinhuang powder (RHP), a classical herbal prescription containing the toxical herb Arisaematis rhizoma, was chosen to test the method. This method could determine whether the Arisaematis Rhizoma had been replaced by Pinellia pedatisecta in the RHP. The results were validated by real-time PCR. 90% compositions of RHP were identified by ITS2 DNA barcoding, suggesting that more DNA barcoding markers are needed for TCM identification. The strategy of high-throughput sequencing has the potential for comprehensive ingredient profiling for TCM preparations. Real-time PCR provides a expeditious metehod for monitoring the safety and legality of TCM preparations.

The mammalian central nervous system (CNS) is considered an immune privileged system as it is separated from the periphery by the blood brain barrier (BBB). Yet, immune functions have been postulated to heavily influence the functional state of the CNS, especially after injury or during neurodegeneration. There is controversy regarding whether adaptive immune responses are beneficial or detrimental to CNS injury repair. In this study, we utilized immunocompromised SCID mice and subjected them to spinal cord injury (SCI). We analyzed motor function, electrophysiology, histochemistry, and performed unbiased RNA-sequencing. SCID mice displayed improved CNS functional recovery compared to WT mice after SCI. Weighted gene-coexpression network analysis (WGCNA) of spinal cord transcriptomes revealed that SCID mice had reduced expression of immune function-related genes and heightened expression of neural transmission-related genes after SCI, which was confirmed by immunohistochemical analysis and was consistent with better functional recovery. Transcriptomic analyses also indicated heightened expression of neurotransmission-related genes before injury in SCID mice, suggesting that a steady state of immune-deficiency potentially led to CNS hyper-connectivity. Consequently, SCID mice without injury demonstrated worse performance in Morris water maze test. Taken together, not only reduced inflammation after injury but also dampened steady-state immune function without injury heightened the neurotransmission program, resulting in better or worse behavioral outcomes respectively. This study revealed the intricate relationship between immune and nervous systems, raising the possibility for therapeutic manipulation of neural function via immune modulation.

MicroRNAs (miRNAs) are important regulators of tumor formation, progression and metastasis. The present study characterized a novel miRNA (miR)-888, as a potent oncomiR in human colorectal cancer (CRC). The clinicopathological investigation on 126 cases of CRC patients demonstrated that the expression level of miR-888 was significantly upregulated in tumors compared with adjacent healthy tissue, and was associated with tumor stage and histological differentiation. A Kaplan-Meier analysis and log-rank test demonstrated that CRC patients with increased miR-888 expression exhibited a decreased overall survival (OS) and disease-free survival (DFS) compared with patients with low miR-888 expression. Further univariate and multivariate analyses identified miR-888 as an independent prognostic factor for poor survival outcome in CRC patients. To determine the biological role of miR-888 in human CRC, in vitro Cell Counting kit-8, wound healing and transwell assays were performed and demonstrated that miR-888 contributed greatly to CRC cell proliferation, invasion and metastasis. Furthermore, potential targets of miR-888 were investigated using a luciferase reporter assay, followed by polymerase chain reaction and western blot analysis. The findings revealed that miR-888 directly bound to the 3'-untranslated region of mothers against decapentaplegic-4 and thus inhibited its expression and promoted the tumor growth factor-1-induced cancer metastasis signaling. The results of the present study identified miR-888 as an oncogenic miRNA in CRC and provide a foundation for promising research in the future regarding this predictive and prognostic biomarker.

The present study aimed to investigate the effects of diabetes mellitus (DM) on the generation of experimental corneal neovascularization (CrNV) and choroidal neovascularization (ChNV). Diabetes was induced in mice by intraperitoneal injection of streptozotocin (STZ). Experimental CrNV and ChNV were induced by alkali injury and laser photocoagulation, respectively. CrNV and ChNV were compared between the STZ‑induced diabetic mice and control mice two weeks after injury. Relative expression of angiogenic factors was quantified by reverse transcription‑quantitative polymerase chain reaction, and progenitor cell or macrophage accumulation in the early phase following injury was examined by flow cytometric analysis. Compared with the alkali‑injured normal mice, the alkali‑injured diabetic mice (STZ‑induced) exhibited no significant difference in CrNV occurrence, whereas the laser‑injured diabetic mice exhibited significantly reduced levels of ChNV compared with those of the laser‑injured control animals. The laser‑induced intrachoroidal mRNA expression levels of angiogenic factors, including vascular endothelial growth factor, hypoxia‑induced factor‑1α, chemokine (C‑C motif) ligand 3, and stromal cell‑derived factor‑1α, were reduced in the laser‑injured diabetic mice when compared with laser‑injured control mice. Furthermore, the laser‑induced intrachoroidal infiltration of c‑Kit+ progenitor cells was impaired in the laser‑injured diabetic mice compared with the laser‑injured control mice. Overall, diabetes did not exert a significant effect on the generation of experimental CrNV. However, diabetes reduced laser‑induced ChNV through downregulation of intrachoroidal progenitor cell infiltration and angiogenic factor expression.

The DNA methylation age, a good reflection of human aging process, has been used to predict chronological age of adults and newborns. However, the prediction model for children and adolescents was absent. In this study, we aimed to generate a prediction model of chronological age for children and adolescents aged 6-17 years by using age-specific DNA methylation patterns from 180 Chinese twin individuals. We identified 6,350 age-related CpGs from the epigenome-wide association analysis (N=179). 116 known age-related sites in children were confirmed. 83 novel CpGs were selected as predictors from all age-related loci by elastic net regression and they could accurately predict the chronological age of the pediatric population, with a correlation of 0.99 and the error of 0.23 years in the training dataset (N=90). The predictive accuracy in the testing dataset (N=89) was high (correlation=0.93, error=0.62 years). Among the 83 predictors, 49 sites were novel probes not existing on the Illumina 450K BeadChip. The top two predictors of age were on the PRKCB and REG4 genes, which are associated with diabetes and cancer, respectively. Our results suggest that the chronological age can be accurately predicted among children and adolescents aged 6-17 years by 83 newly identified CpG sites.

Stem cell transplantation has been considered a promising therapeutic approach for premature ovarian failure (POF). However, to date, no quantitative data analysis of stem cell therapy for POF has been performed. Therefore, the present study performed a meta-analysis to assess the efficacy of stem cell transplantation in improving ovarian function in animal models of POF. In addition, a case report of a patient with POF subjected to stem cell treatment was included to demonstrate that stem cell therapy also contributes to the recovery of ovarian function in patients. Published studies were identified by a systematic review of the PubMed, Embase, and Cochrane's library databases, and references cited in associated reviews were also considered. Data regarding follicle-stimulating hormone (FSH), estradiol (E2), ovarian weight, follicle count, the number of pregnancies and other parameters, including delivery route and cell type, were extracted. Pooled analysis, sensitivity analyses, subgroup analyses and meta-regression were performed. In the case of POF, transvaginal ultrasound (TVS), abdominal ultrasound (TAS) and color Doppler flow imaging (CDFI) were performed to observe the endometrial morphology and blood flow signals in the patient. Overall, pooled results from 16 pre-clinical studies demonstrated that stem cell-based therapy significantly improved FSH levels [standardized mean difference (SMD)=-1.330; 95% confidence interval (CI), -(2.095-0.565); P=0.001], E2 levels (SMD=2.334; 95% CI, 1.350-3.319; P<0.001), ovarian weight (SMD=1.310; 95% CI, 0.157-2.463; P=0.026), follicle count (SMD=1.871; 95% CI, 1.226-2.516; P<0.001), and the number of pregnancies (risk ratio=1.715, 95% CI, 1.213-2.424; P=0.002). The results of TVS and TAS demonstrated improved ovarian size and endometrial thickness in the patient with POF after MSC treatment. Of note, a rich blood flow signal in the endometrium was observed on CDFI. It appeared that stem cell-based therapy may be an effective method for the resumption of ovarian function in a patient and in animal models of POF; however, large-scale and high-quality future studies are required to confirm the present findings due to heterogeneity.

γδ T cells recognize a wide variety of ligands in mammals, among them members of the butyrophilin (BTN) family. Nothing is known about γδ T cell ligands in chickens, despite there being many such cells in blood and lymphoid tissues, as well as in mucosal surfaces. The major histocompatibility complex (MHC) of chickens was discovered because of polymorphic BG genes, part of the BTN family. All but two BG genes are located in the BG region, oriented head-to-tail so that unequal crossing-over has led to copy number variation (CNV) as well as hybrid (chimeric) genes, making it difficult to identify true alleles. One approach is to examine BG genes expressed in particular cell types, which likely have the same functions in different BG haplotypes and thus can be considered "functional alleles." We cloned nearly full-length BG transcripts from peripheral T cells of four haplotypes (B2, B15, B19, and B21), and compared them to the BG genes of the B12 haplotype that previously were studied in detail. A dominant BG gene was found in each haplotype, but with significant levels of subdominant transcripts in three haplotypes (B2, B15, and B19). For three haplotypes (B15, B19, and B21), most sequences are closely-related to BG8, BG9, and BG12 from the B12 haplotype. We found that variation in the extracellular immunoglobulin-variable-like (Ig-V) domain is mostly localized to the membrane distal loops but without evidence for selection. However, variation in the cytoplasmic tail composed of many amino acid heptad repeats does appear to be selected (although not obviously localized), consistent with an intriguing clustering of charged and polar residues in an apparent α-helical coiled-coil. By contrast, the dominantly-expressed BG gene in the B2 haplotype is identical to BG13 from the B12 haplotype, and most of the subdominant sequences are from the BG5-BG7-BG11 clade. Moreover, alternative splicing leading to intron read-through results in dramatically truncated cytoplasmic tails, particularly for the dominantly-expressed BG gene of the B2 haplotype. The approach of examining "functional alleles" has yielded interesting data for closely-related genes, but also thrown up unexpected findings for at least one haplotype.

To explore the potential molecular mechanism of low-intensity pulsed ultrasound (LIPUS) in the treatment of temporomandibular joint osteoarthritis (TMJ-OA), and identify the target genes for therapy of TMJ-OA.

Hypertension is the most important risk factor for cardiovascular and cerebrovascular diseases. The study found that CXCL12 and CNNM2 gene affects the risk of coronary heart disease, but the relationship with hypertension is unclear. The aim of this research is to explore the association between CXCL12 and CNNM2 gene and hypertension in Chinese Han population.

YAP, a key effector of Hippo pathway, is activated by its translocation from cytoplasm to nucleus to regulate gene expression and promote tumorigenesis. Although the mechanism by which YAP is suppressed in cytoplasm has been well-studied, how the activated YAP is sequestered in the nucleus remains unknown. Here, we demonstrate that YAP is a nucleocytoplasmic shuttling protein and its nuclear export is controlled by SET1A-mediated mono-methylation of YAP at K342, which disrupts the binding of YAP to CRM1. YAP mimetic methylation knockin mice are more susceptible to colorectal tumorigenesis. Clinically, YAP K342 methylation is reversely correlated with cancer survival. Collectively, our study identifies SET1A-mediated mono-methylation at K342 as an essential regulatory mechanism for regulating YAP activity and tumorigenesis.

Understanding virus assembly mechanisms is important for developing therapeutic interventions. Nipah virus (NiV) is of interest because of its high mortality rate and efficient human-human transmissions. The current model for most enveloped viruses suggests that matrix proteins (M) recruit attachment glycoproteins (G) and fusion glycoproteins (F) to the assembly site at the plasma membrane. Here we report an assembly model that differs in many aspects from the current one. Examining NiV proteins on the cell plasma membrane using super-resolution microscopy reveals that clusters of F and G are randomly distributed on the plasma membrane regardless of the presence or absence of M. Our data suggests a model in which the M molecules assemble at the plasma membrane to form virus-like particles (VLPs), while the incorporation of F and G into the nascent VLPs is stochastic.

Regulatory T (Treg) cells are essential for maintaining immune homeostasis and tolerance, but the mechanisms regulating the stability and function of Treg cells have not been fully elucidated. Here we show SUMO-specific protease 3 (SENP3) is a pivotal regulator of Treg cells that functions by controlling the SUMOylation and nuclear localization of BACH2. Treg cell-specific deletion of Senp3 results in T cell activation, autoimmune symptoms and enhanced antitumor T cell responses. SENP3-mediated BACH2 deSUMOylation prevents the nuclear export of BACH2, thereby repressing the genes associated with CD4+ T effector cell differentiation and stabilizing Treg cell-specific gene signatures. Notably, SENP3 accumulation triggered by reactive oxygen species (ROS) is involved in Treg cell-mediated tumor immunosuppression. Our results not only establish the role of SENP3 in the maintenance of Treg cell stability and function via BACH2 deSUMOylation but also clarify the function of SENP3 in the regulation of ROS-induced immune tolerance.

Multiple myeloma (MM) is an incurable hematologic malignancy due to inevitable relapse and chemoresistance development. Our preliminary data show that MM cells express high levels of PGC1β and LDHA. In this study, we investigated the mechanism behind PGC1β-mediated LDHA expression and its contribution to tumorigenesis, to aid in the development of novel therapeutic approaches for MM. Real-time PCR and western blotting were first used to evaluate gene expression of PGC1β and LDHA in different MM cells, and then, luciferase reporter assay, chromatin immunoprecipitation, LDHA deletion report vectors, and siRNA techniques were used to investigate the mechanism underlying PGC1β-induced LDHA expression. Furthermore, knockdown cell lines and lines stably overexpressing PGC1β or LDHA lentivirus were established to evaluate in vitro glycolysis metabolism, mitochondrial function, reactive oxygen species (ROS) formation, and cell proliferation. In addition, in vivo xenograft tumor development studies were performed to investigate the effect of PGC1β or LDHA expression on tumor growth and mouse survival. We found that PGC1β and LDHA are highly expressed in different MM cells and LDHA is upregulated by PGC1β through the PGC1β/RXRβ axis acting on the LDHA promoter. Overexpression of PGC1β or LDHA significantly potentiated glycolysis metabolism with increased cell proliferation and tumor growth. On the other hand, knockdown of PGC1β or LDHA largely suppressed glycolysis metabolism with increased ROS formation and apoptosis rate, in addition to suppressing tumor growth and enhancing mouse survival. This is the first time the mechanism underlying PGC1β-mediated LDHA expression in multiple myeloma has been identified. We conclude that PGC1β regulates multiple myeloma tumor growth through LDHA-mediated glycolytic metabolism. Targeting the PGC1β/LDHA pathway may be a novel therapeutic strategy for multiple myeloma treatment.

Real-time reverse transcription PCR (RT-qPCR) is a preferred method for rapid and accurate quantification of gene expression studies. Appropriate application of RT-qPCR requires accurate normalization though the use of reference genes. As no single reference gene is universally suitable for all experiments, thus reference gene(s) validation under different experimental conditions is crucial for RT-qPCR analysis. To date, only a few studies on reference genes have been done in other plants but none in papaya. In the present work, we selected 21 candidate reference genes, and evaluated their expression stability in 246 papaya fruit samples using three algorithms, geNorm, NormFinder and RefFinder. The samples consisted of 13 sets collected under different experimental conditions, including various tissues, different storage temperatures, different cultivars, developmental stages, postharvest ripening, modified atmosphere packaging, 1-methylcyclopropene (1-MCP) treatment, hot water treatment, biotic stress and hormone treatment. Our results demonstrated that expression stability varied greatly between reference genes and that different suitable reference gene(s) or combination of reference genes for normalization should be validated according to the experimental conditions. In general, the internal reference genes EIF (Eukaryotic initiation factor 4A), TBP1 (TATA binding protein 1) and TBP2 (TATA binding protein 2) genes had a good performance under most experimental conditions, whereas the most widely present used reference genes, ACTIN (Actin 2), 18S rRNA (18S ribosomal RNA) and GAPDH (Glyceraldehyde-3-phosphate dehydrogenase) were not suitable in many experimental conditions. In addition, two commonly used programs, geNorm and Normfinder, were proved sufficient for the validation. This work provides the first systematic analysis for the selection of superior reference genes for accurate transcript normalization in papaya under different experimental conditions.

Recent studies have demonstrated that photosynthetic cyanobacteria could be an excellent cell factory to produce renewable biofuels and chemicals due to their capability to utilize solar energy and CO2 as the sole energy and carbon sources. Biosynthesis of carbon-neutral biofuel alkanes with good chemical and physical properties has been proposed. However, to make the process economically feasible, one major hurdle to improve the low cell tolerance to alkanes needed to be overcome.