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Alzheimer’s disease (AD) and Huntington’s disease (HD) are destructive worldwide diseases. Efforts have been made to elucidate the process of these two diseases, yet the pathogenesis remains elusive as it involves a combination of multiple factors, including genetic and environmental ones. To explore the potential role of forkhead box O1 (FOXO1) in the development of AD and HD, we identified 1,853 differentially expressed genes (DEGs) from 19,414 background genes in both the AD&HD/control and FOXO1-low/high groups. Four coexpression modules were predicted by the weighted gene coexpression network analysis (WGCNA), among which blue and turquoise modules had the strongest correlation with AD&HD and high expression of FOXO1. Functional enrichment analysis showed that DEGs in these modules were enriched in phagosome, cytokine-cytokine receptor interaction, cellular senescence, FOXO signaling pathway, pathways of neurodegeneration, GABAergic synapse, and AGE-RAGE signaling pathway in diabetic complications. Furthermore, the cross-talking pathways of FOXO1 in AD and HD were jointly determined in a global regulatory network, such as the FOXO signaling pathway, cellular senescence, and AGE-RAGE signaling pathway in diabetic complications. Based on the performance evaluation of the area under the curve of 85.6%, FOXO1 could accurately predict the onset of AD and HD. We then identified the cross-talking pathways of FOXO1 in AD and HD, respectively. More specifically, FOXO1 was involved in the FOXO signaling pathway and cellular senescence in AD; correspondingly, FOXO1 participated in insulin resistance, insulin, and the FOXO signaling pathways in HD. Next, we use GSEA to validate the biological processes in AD&HD and FOXO1 expression. In GSEA analysis, regulation of protein maturation and regulation of protein processing were both enriched in the AD&HD and FOXO1-high groups, suggesting that FOXO1 may have implications in onset and progression of these two diseases through protein synthesis. Consequently, a high expression of FOXO1 is a potential pathogenic factor in both AD and HD involving mechanisms of the FOXO signaling pathway, AGE-RAGE signaling pathway in diabetic complications, and cellular senescence. Our findings provide a comprehensive perspective on the molecular function of FOXO1 in the pathogenesis of AD and HD.

Aims Deep mining of the molecular mechanisms underlying diabetic retinopathy (DR) is critical for the development of novel therapeutic targets. This study aimed to identify key molecular signatures involved in experimental DR on the basis of integrated bioinformatics analysis. Materials and methods Four datasets consisting of 37 retinal samples were downloaded from the National Center of Biotechnology Information Gene Expression Omnibus (GEO). After batch-effect adjustment, bioinformatics tools such as Networkanalyst, Enrichr, STRING, and Metascape were used to evaluate the differentially expressed genes (DEGs), perform enrichment analysis, and construct protein-protein interaction (PPI) networks. The hub genes were identified using Cytoscape software. The DEGs of interest from the meta-analysis were confirmed by quantitative reverse transcription-polymerase chain reaction (RT-PCR) in diabetic rats and a high glucose-treated retinal cell model, respectively. Results A total of 743 DEGs related to lens differentiation, insulin resistance, and HDL cholesterol metabolism were obtained using the meta-analysis. Alterations of dynamic gene expression in the chloride ion channel, retinol metabolism, and fatty acid metabolism were involved in the course of DR in rats. Importantly, H3K27m3 modifications regulated the expression of most DEGs at the early stage of DR. This article is protected by copyright. All rights reserved. Conclusions Using an integrated bioinformatics approach, novel molecular signatures were obtained for different stages of DR progression, and the findings may represent distinct therapeutic strategies for DR patients.

Epigenome editing methods enable the precise manipulation of epigenetic modifications, such as histone posttranscriptional modifications (PTMs), for uncovering their biological functions. While histone PTMs have been correlated with certain gene expression status, the causalities remain elusive. Histone H3 Lysine 27 acetylation (H3K27ac) and histone H3 Lysine 4 trimethylation (H3K4me3) are both associated with active genes, and located at active promoters and enhancers or around transcriptional start sites (TSSs). Although crosstalk between histone lysine acetylation and H3K4me3 has been reported, relationships between specific epigenetic marks during transcriptional activation remain largely unclear. Here, using clustered regularly interspaced short palindromic repeats (CRISPR)/dCas-based epigenome editing methods, we discovered that the ectopic introduction of H3K27ac in the promoter region lead to H3K4me3 enrichment around TSS and transcriptional activation, while H3K4me3 installation at the promoter cannot induce H3K27ac increase and failed to activate gene expression. Blocking the reading of H3K27ac by BRD proteins using inhibitor JQ1 abolished H3K27ac-induced H3K4me3 installation and downstream gene activation. Furthermore, we uncovered that BRD2, not BRD4, mediated H3K4me3 installation and gene activation upon H3K27ac writing. Our studies revealed the relationships between H3K27ac and H3K4me3 in gene activation process and demonstrated the application of CRISPR/dCas-based epigenome editing methods in elucidating the crosstalk between epigenetic mechanisms.

GPR174 is abundantly expressed in B and T lymphocytes and has a role in restraining T cell responses, but the function of GPR174 in B cells is less clear. Here we report that upon in vitro culture B cells undergo a spontaneous GPR174-dependent activation process that is associated with marked changes in gene expression, including up-regulation of Cd86, Nr4a1, Ccr7, and phosphodiesterases. B cells lacking Gαs show a block in induction of the GPR174-dependent program. Spontaneous up-regulation of CD86 in cultured B cells is dependent on protein kinase A. Both GPR174- and Gαs-deficient B cells show enhanced survival in culture. In vivo, GPR174 contributes to NUR77 expression in follicular B cells and is needed for establishing a marginal zone compartment of normal size. Treatment of mice with lysophosphatidylserine (lysoPS), a GPR174 ligand, is sufficient to promote CD86 up-regulation by follicular B cells. These findings demonstrate that GPR174 can signal via Gαs to modulate B cell gene expression and show this can occur in vivo in response to lysoPS. Additionally, the findings illuminate a pathway that might be targeted to improve systems for the in vitro study of B cell responses.

High-protein diets are known to reduce weight and fat deposition. However, there have been only a few studies on the efficacy of different types of high-protein diets in preventing obesity. Therefore, the emphasis of this study lies in comparing the efficacy of two high-protein diets (milk protein and whey protein) in preventing obesity and exploring specific mechanisms. Eighty Sprague Dawley rats were divided into two groups and fed with milk protein concentrate (MPC) and whey protein concentrate (WPC) for 12 weeks. Each group was divided into four levels: two low-fat regimens with either low or high protein content (L-14%, L-40%) and two high-fat regimens with either low or high protein content (H-14%, H-40%). The studies we have performed showed that rats treated with MPC at the 40% protein level had significantly reduced body weight, fat weight and fat ratio gain induced by a high-fat diet, while the protein level in the WPC group had no effect on body weight or body fat in rats fed with a high-fat diet. What is more, rats fed with MPC at the H-40% energy level showed a significant decrease in plasma triglyceride, total cholesterol and low-density lipoprotein cholesterol levels and a significant increase in plasma high-density lipoprotein cholesterol levels compared with the H-14% energy level group. In contrast, in the WPC groups, increasing the protein content in high-fat diets had no significant influence on plasma lipid levels. The results of the amino acid composition of the two proteins and plasma showed that the MPC diet of 40% protein level increased the transsulfuration pathway in rats, thereby increasing the level of H2S. This research work has shown that not all types of high-protein diets can effectively prevent obesity induced by high-fat diets, as effectiveness depends on the amino acid composition of the protein.

Background Metastasis is the most devastating stage of cancer progression and often shows a preference for specific organs. Methods To reveal the mechanisms underlying organ-specific metastasis, we systematically analyzed gene expression profiles for three common metastasis sites across all available primary origins. A rank-based method was used to detect differentially expressed genes between metastatic tumor tissues and corresponding control tissues. For each metastasis site, the common differentially expressed genes across all primary origins were identified as organ-specific metastasis genes. Results Pathways enriched by these genes reveal an interplay between the molecular characteristics of the cancer cells and those of the target organ. Specifically, the neuroactive ligand-receptor interaction pathway and HIF-1 signaling pathway were found to have prominent roles in adapting to the target organ environment in brain and liver metastases, respectively. Finally, the identified organ-specific metastasis genes and pathways were validated using a primary breast tumor dataset. Survival and cluster analysis showed that organ-specific metastasis genes and pathways tended to be expressed uniquely by a subgroup of patients having metastasis to the target organ, and were associated with the clinical outcome. Conclusions Elucidating the genes and pathways underlying organ-specific metastasis may help to identify drug targets and develop treatment strategies to benefit patients.

In China, the fruits of Physalis alkekengi L. var. franchetii, which are conventionally utilized as edible berry, have attracted wide attention due to its significant biological activities. In the present study, phytochemical studies on the fruits of Physalis plants afforded six compounds, including two new withanolides (1-2) and four known agnologues (3-6). The inhibitory effects of these compounds on the formation of nitric oxide (NO) stimulated by lipopolysaccharide (LPS) in RAW264.7 macrophages were evaluated. Physapubescin M (1), with IC50 value of 1.58 μM, was selected for further study. The protein expression of COX-2 and iNOS, and LPS-induced production of cytokines (IL-6, IL-1β and TNF-α) were reduced by physapubescin M (1) in a dose-dependent way. In addition, transcriptomic analyses were conducted to profile gene expression alterations in LPS-induced RAW264.7 cells upon treatment of physapubescin M (1) and the potential antiinflammatory mechanism of withnolides was mentioned. These results provide broad view to the underlying antiinflammatory mechanism of withnolides, and give a theoretical basis for the utilization of the fruits of P. alkekengi L. var. franchetii.

The purpose of this study was to investigate the potential roles of protein kinase C beta (PRKCB) in the pathogenesis of Alzheimer’s disease (AD). We identified 2,254 differentially expressed genes from 19,245 background genes in AD versus control as well as PRKCB-low versus high group. Five co-expression modules were constructed by weight gene correlation network analysis. Among them, the 1,222 genes of the turquoise module had the strongest relation to AD and those with low PRKCB expression, which were enriched in apoptosis, axon guidance, gap junction, Fc gamma receptor (FcγR)-mediated phagocytosis, mitogen-activated protein kinase (MAPK) and vascular endothelial growth factor (VEGF) signaling pathways. The intersection pathways of PRKCB in AD were determined, including gap junction, FcγR-mediated phagocytosis, MAPK and VEGF signaling pathways. Based on the performance evaluation of the area under the curve of 75.3%, PRKCB could accurately predict the onset of AD. Therefore, low expressions of PRKCB was a potential causative factor of AD, which might be involved in gap junction, FcγR-mediated phagocytosis, MAPK and VEGF signaling pathways.

Glycosylation is a widely occurring posttranslational modification. Here, we applied a quick, convenient and high-throughput strategy (lectin array) to investigate the variation in glycans on different macrophage subtypes derived from THP-1 and RAW264.7 cells. For THP-1 cells, there were more significant differences in the glycan on M2 macrophages compared to the other two subtypes. In contrast, M1 macrophages exhibited more significant glycan remodeling than the other subtypes for the RAW264.7 cell line. The response of the lectins which recogonize the N-glycan and α2,6 sialic acid was higher during polarization into anti-inflammatory phase (THP-1 derived M2 subtypes), and lower in pro-inflammatory phase (RAW264.7 M1 subtypes). The regulation of several α2,6 sialyltransferase genes was coincident with the regulation of the α2,6 sialic acid on the two cell lines. The lectin response and glycosyltranferase gene expression confirmed that α2,6 sialic acid showed higher expression in the anti-inflammatory phase. This indicated that α2,6 sialic acid was a potential indicator for the anti-inflammatory response.

Backgrounds: β-Amyloid (Aβ) is a principal constituent of senile plaques in Alzheimer’s disease (AD) and induces neuronal cell death. The molecular mechanism of how Aβ evokes neuronal cell death remains complicated, which were investigated in the present study. Methods: Using the human neuroblastoma cell line SHSY5Y, we investigated the neurotoxic effects of human β-Amyloid 1–42 (Aβ1–42) aggregates on gene expression profile and protein expression profile by using the Agilent GeneChip Human 1A (V2) Oligo MicroArray, Quantitative Real-time PCR, PF-2D and Western blot analysis. Results: Our results show that Aβ1–42 specifically influences gene and protein expression such as EGR1, eIF5A, PDE8A, ERp57 and ERp5 in pathways associated with apoptotic process, protein translation, cAMP catabolic process and response to endoplasmic reticulum stress. Conclusion: Although Genes with significant changes in transcriptomic analysis matched very few of the proteins identified in proteomics analysis, our findings will strengthen our knowledge concerning the molecular mechanisms underlying AD.

Triggering receptor expressed on myeloid cells 2 (TREM2) is a microglial surface receptor genetically linked to the risk for Alzheimer’s disease (AD). A proteolytic product, soluble TREM2 (sTREM2), is abundant in the cerebrospinal fluid and its levels positively correlate with neuronal injury markers. To gain insights into the pathological roles of sTREM2, we studied sTREM2 in the brain of 5xFAD mice, a model of AD, by direct stereotaxic injection of recombinant sTREM2 protein or by adeno-associated virus (AAV)-mediated expression. We found that sTREM2 reduces amyloid plaque load and rescues functional deficits of spatial memory and long-term potentiation. Importantly, sTREM2 enhances microglial proliferation, migration, clustering in the vicinity of amyloid plaques and the uptake and degradation of Aβ. Depletion of microglia abolishes the neuroprotective effects of sTREM2. Our study demonstrates a protective role of sTREM2 against amyloid pathology and related toxicity and suggests that increasing sTREM2 can be explored for AD therapy., TREM2 is a genetic risk factor for Alzheimer’s disease, and soluble TREM2 (sTREM2) in the CSF correlates with AD progression. Here the authors study the role of sTREM2 in a mouse model of Alzheimer’s disease, and find it reduces amyloid accumulation and increases the numbers of plaque-associated microglia which correlates with improved behavioural function in the mice.

Cancer cells are typically characterized by abnormal quality control of mitochondria, production of reactive oxygen species (ROS), dysregulation of the cell redox state, and the Warburg effect. Mutation or depletion of PTEN-induced kinase 1 (PINK1) or Parkin leads to mitophagy defects and accumulation of malfunctioning mitochondria, and is often detected in a variety of tumors. However, PINK1's role in the progression of gastric cancer (GC) remains unclear, with its main effect being on mitochondrial turnover, metabolic reprogramming, and tumor microenvironment (TME) alteration. To address these issues, we first assessed the expression levels of PINK1, mitophagy-associated molecules, ROS, HIF-1α, glycolysis-associated genes, and macrophage signatures in GC tissues and matched tumor-adjacent normal samples. In addition, GC cell lines (AGS and MKN-45) and xenograft mouse models were used to determine the mechanism by which PINK1 regulates mitophagy, metabolic reprogramming, tumor-associated macrophage (TAM) polarization, and GC progression. We found that PINK1 loss correlated with advanced stage GC and poorer overall survival. GC tissues with lower PINK1 levels showed compromised mitophagy signaling and enhanced glycolytic enzyme expression. In vitro experiments demonstrated that PINK1 deficiency promoted GC cell proliferation and migration through the inhibition of mitophagy, production of mitochondrial ROS, stabilization of HIF-1α, and facilitation of the Warburg effect under both normoxic and hypoxic conditions. Moreover, PINK1 deficiency in GC cells promoted TAM polarization toward the M2-like phenotype. Reintroduction of PINK1 or inhibition of HIF-1α effectively repressed PINK1 deficiency-mediated effects on GC cell growth, metabolic shift, and TAM polarization. Thus, mitophagy defects caused by PINK1 loss conferred a metabolic switch through accumulation of mtROS and stabilization of HIF-1α, thereby facilitating the M2 polarization of TAM to remodel an immunosuppressive microenvironment in GC. Our results clarify the mechanism between PINK1 and GC progression and may provide a novel strategy for the treatment of GC.

Profiling of active gene-regulatory DNA elements in brain cancer tissues illuminates disease biology and therapeutic targets., Molecular profiling of the most aggressive brain tumor glioblastoma (GBM) on the basis of gene expression, DNA methylation, and genomic variations advances both cancer research and clinical diagnosis. The enhancer architectures and regulatory circuitries governing tumor-intrinsic transcriptional diversity and subtype identity are still elusive. Here, by mapping H3K27ac deposition, we analyze the active regulatory landscapes across 95 GBM biopsies, 12 normal brain tissues, and 38 cell line counterparts. Analyses of differentially regulated enhancers and super-enhancers uncovered previously unrecognized layers of intertumor heterogeneity. Integrative analysis of variant enhancer loci and transcriptome identified topographies of transcriptional enhancers and core regulatory circuitries in four molecular subtypes of primary tumors: AC1-mesenchymal, AC1-classical, AC2-proneural, and AC3-proneural. Moreover, this study reveals core oncogenic dependency on super-enhancer–driven transcriptional factors, long noncoding RNAs, and druggable targets in GBM. Through profiling of transcriptional enhancers, we provide clinically relevant insights into molecular classification, pathogenesis, and therapeutic intervention of GBM.

Background Brain metastases explain the majority of mortality associated with lung cancer, which is the leading cause of cancer death. Cytology analysis of the cerebrospinal fluid (CSF) remains the diagnostic gold standard, however, the circulating tumor cells (CTCs) in CSF (CSF‐CTCs) are not well defined at the molecular and transcriptome levels. Methods We established an effective CSF‐CTCs collection procedure and isolated individual CSF cells from five lung adenocarcinoma leptomeningeal metastases (LUAD‐LM) patients and three controls. Three thousand seven hundred ninety‐two single‐cell transcriptomes were sequenced, and single‐cell RNA sequencing (scRNA‐seq) gene expression analysis was used to perform a comprehensive characterization of CSF cells. Results Through clustering and expression analysis, we defined CSF‐CTCs at the transcriptome level based on epithelial markers, proliferation markers, and genes with lung origin. The metastatic‐CTC signature genes are enriched for metabolic pathway and cell adhesion molecule categories, which are crucial for the survival and metastases of tumor cells. We discovered substantial heterogeneity in patient CSF‐CTCs. We quantified the degree of heterogeneity and found significantly greater among‐patient heterogeneity compared to among‐cell heterogeneity within a patient. This observation could be explained by spatial heterogeneity of metastatic sites, cell‐cycle gene, and cancer‐testis antigen (CTA) expression profiles as well as the proportion of CTCs displaying mesenchymal and cancer stem cell properties. In addition, our CSF‐CTCs transcriptome profiling allowed us to determine the biomarkers during the progression of an LM patient with cancer of unknown primary site (CUP). Conclusions Our results will provide candidate genes for an RNA‐based digital detection of CSF‐CTCs from LUAD‐LM and CUP‐LM cases, and shed light on the therapy and mechanism of LUAD‐LM., Our study firstly investigated CSF‐CTCs characteristics of five LUAD‐LM patients and one CUP‐LM patient. CSF‐CTCs enriched metabolism and cell‐adhesion pathways. CSF‐CTCs exhibited heterogeneity manifested by cell cycle, mesenchymal and cancer stem cell, and cancer‐testis antigen properties

A member of the newly discovered RNA family, circular RNA (circRNA) is considered as the intermediate product of by-product splicing or abnormal RNA splicing. With the development of RNA sequencing, circRNA has recently drawn research interest. CircRNA exhibits stability, species conservatism, and tissue cell specificity. It acts as a miRNA sponge in the circRNA-microRNA (miRNA-mRNA axis, which can regulate gene transcription and protein translation. Studies have confirmed that circRNA is ubiquitous in eukaryotic cells, which play an important role in the regulation of human gene expression and participate in the occurrence and development of various human diseases. CircRNA may be closely related to the occurrence and development of female reproductive system diseases. By analyzing the biological functions and mechanism of circRNA, we find that circRNA has certain development prospects as biomarkers of the female reproductive system diseases. The production and degradation of circRNA, biological functions, and their association with the occurrence of diseases of female reproductive system are reviewed in this article.

Reprograming lipid metabolism, one of the major metabolic alterations in cancer, is believed to play an essential role in cancer development, but the exact molecular mechanism remains elusive. Here, we present a computational study of transcriptomic data of HCC with HCV etiology to investigate how lipid metabolism alters during HCC progression. Our analyses reveal that: (1) cancer tissue cells tend to synthesize fatty acids de novo and its phospholipid derivatives; (2) lipid catabolism and fatty acid oxidation are remarkably down-regulated in HCC; (3) the lipid metabolism in HCC is largely independent of lipids in blood circulation; (4) stage-specific co-expression networks for lipid metabolic genes were identified during HCC progression; and (5) the expression levels of several lipid metabolic genes that are differentially expressed or co-expressed specifically at the HCC stage have a strong correlation with cancer survival. Overall, the results provide detailed information about the reprogramed lipid metabolism in HCV-derived HCC. © Copyright © 2020 Liu, Liu, Cui and Xu. This work was supported by grants from the Inner Mongolia Natural Science Foundation of China (2018LH03023) and the Science Foundation for Excellent Youth Scholars of Inner Mongolia University of Science and Technology (2016YQL06).

Numb is a negative regulator of Notch signal pathway. Previous study has demonstrated that Notch signal pathway activation is required for hepatic progenitor cell (HPC) differentiating into cholangiocytes in cholestatic liver fibrosis (CLF), and Huang Qi Decoction (HQD) could prevent CLF through inhibition of the Notch signal pathway. However, the role of Numb in HQD against CLF is yet unclear. Thus, CLF rats transplanted into rat bone marrow-derived mesenchymal stem cells with knocked down Numb gene (BMSCNumb-KD) were treated with HQD. Simultaneously, Numb gene knockdown was also performed in WB-F344 cell line and then treated with refined HQD in vitro. In vivo study revealed that liver fibrosis was inhibited by HQD plus BMSCNumb-KD treatment, while Hyp content in liver tissue, the gene and protein expression of α-SMA, gene expression of Col I, TNF-α, and TGF-β1 were increased compared to that in HQD group. Furthermore, Notch signal pathway was inhibited by HQD plus BMSCNumb-KD, while the protein expression of Numb was decreased and RBP-Jκ and Hes1 was increased compared to that in HQD group. In vitro, HQD reduced the differentiation of WB-F344 cells into cholangiocyte phenotype, while this effect was attenuated after Numb-knockdown. This study highlights that the absence of hepatic stem cell Numb gene decreases effect of HQD against CLF, which give rise the conclusion that Numb might be a potential target for HQD against CLF.

The aim of this study is to explore the genomic mechanism of early death in Huntington’s disease (HD). We identified 10160 and 1511 differentially expressed genes (DEGs) from comparisons of HD versus control and early versus late death, respectively. On the basis of 922 overlapped DEGs among them, six functional modules were established by weight gene correlation network analysis. The turquoise module most strongly related to overall survival of HD was significantly enriched in GABAergic synapse, retrograde endocannabinoid signaling and neuroactive ligand-receptor interaction. Low expression of five DEGs (CA10, WSB2, ACTR3B, PCDH19 and GABRB3) with highest degree of connectivity and non-zero regression coefficients were identified as hub genes, based on which the LASSO model exactly predicted the occurrence of early death in HD. Furthermore, the network analysis revealed indirect interaction of CA10 with the central hub gene GABRB3, which was involved in the pathway of GABAergic synapse. Compared with high expression of hub genes, their low expression appeared shorter overall survival of HD patients according to Kaplan-Meier survival analysis. Consequently, low expression of CA10, WSB2, ACTR3B, PCDH19 and GABRB3 were possibly associated with early death of HD. The likelihood of low expression of GABRB3 regulated by CA10 in GABAergic synapse contributed to the pathogenesis of early death in HD.

Quantitative assessment of the intracellular oxidative stress level is a very important problem since it is the basis for elucidation of the fundamental causes of metabolic changes in diseased human cells, particularly cancer. However, the problem proves to be very challenging to solve in vivo because of the complex nature of the problem. Here a computational method is presented for predicting the quantitative level of the intracellular oxidative stress in cancer tissue cells. The basic premise of the predictor is that the genomic mutation level is strongly associated with the intracellular oxidative stress level. Based on this, a statistical analysis is conducted to identify a set of enzyme-encoding genes, whose combined expression levels can well explain the mutation rates in individual cancer tissues in the TCGA database. We have assessed the validity of the predictor by assessing it against genes that are known to have anti-oxidative functions for specific types of oxidative stressors. Then the applications of the predictor are conducted to illustrate its utility.

Purpose This study aimed to explore the role of the protein kinase A (PKA) pathway in proliferative vitreoretinopathy (PVR) and the effect of the PKA inhibitor H89 on experimental PVR. Methods Epiretinal membranes (ERMs) were acquired from PVR patients and analyzed by frozen-section immunofluorescence. An in vivo model was developed by intravitreal injecting rat eyes with ARPE-19 cells and platelet-rich plasma, and changes in eye structures and vision function were observed. An in vitro epithelial-mesenchymal transition (EMT) cell model was established by stimulating ARPE-19 cells with transforming growth factor (TGF)-β. Alterations in EMT-related genes and cell function were detected. Mechanistically, PKA activation and activity were explored to assess the relationship between TGF-β1 stimulation and the PKA pathway. The effect of H89 on the TGF-β-Smad2/3 pathway was detected. RNA sequencing was used to analyze gene expression profile changes after H89 treatment. Results PKA was activated in human PVR membranes. In vivo, H89 treatment protected against structural changes in the retina and prevented decreases in electroretinogram b-wave amplitudes. In vitro, H89 treatment inhibited EMT-related gene alterations and partially reversed the functions of the cells. TGF-β-induced PKA activation was blocked by H89 pretreatment. H89 did not affect the phosphorylation or nuclear translocation of regulatory Smad2/3 but increased the expression of inhibitory Smad6. Conclusions PKA pathway activation is involved in PVR pathogenesis, and the PKA inhibitor H89 can effectively inhibit PVR, both in vivo and in vitro. Furthermore, the protective effect of H89 is related to an increase in inhibitory Smad6.

A novel method is developed for predicting the stage of a cancer tissue based on the consistency level between the co-expression patterns in the given sample and samples in a specific stage. The basis for the prediction method is that cancer samples of the same stage share common functionalities as reflected by the co-expression patterns, which are distinct from samples in the other stages. Test results reveal that our prediction results are as good or potentially better than manually annotated stages by cancer pathologists. This new co-expression-based capability enables us to study how functionalities of cancer samples change as they evolve from early to the advanced stage. New and exciting results are discovered through such functional analyses, which offer new insights about what functions tend to be lost at what stage compared to the control tissues and similarly what new functions emerge as a cancer advances. To the best of our knowledge, this new capability represents the first computational method for accurately staging a cancer sample. The R source code used in this study is available at GitHub (https://github.com/yxchspring/CECS).

Macrophages play critical roles in wound healing process. They switch from "classically activated" (M1) phenotype in the early inflammatory phase to "alternatively activated" (M2) phenotype in the later healing phase. However, the dynamic process of macrophage phenotype switching in diabetic wounds burdened with bacteria is unclear. In this report, Pseudomonas aeruginosa, frequently detected in diabetic foot ulcers, was inoculated into cutaneous wounds of db/db diabetic mice to mimic bacterium-infected diabetic wound healing. We observed that P. aeruginosa infection impaired diabetic wound healing and quickly promoted the expression of pro-inflammatory genes (M1 macrophage markers) tumor necrosis factor-α (tnf-α), interleukin-1β (il-1β) and il-6 in wounds. The expression of markers of M2 macrophages, including il-10, arginase-1, and ym1 were also upregulated. In addition, similar gene expression patterns were observed in macrophages isolated directly from wounds. Immunostaining showed that P. aeruginosa infection increased both the ratios of M1 and M2 macrophages in wounds compared with that in control groups, which was further confirmed by in vitro culturing macrophages with P. aeruginosa and skin fibroblast conditioned medium. However, the ratios of the expression levels of pro-inflammatory genes to anti-inflammatory gene il-10 was increased markedly in P. aeruginosa infected wounds and macrophages compared with that in control groups, and P. aeruginosa prolonged the presence of M1 macrophages in the wounds. These data demonstrated that P. aeruginosa in diabetic wounds activates a mixed M1/M2 macrophage phenotype with an excessive activation of M1 phenotype or relatively inadequate activation of M2 phenotype.

Chemical examination of an EtOAc extract of cultured Aspergillus versicolor fungus from deep-sea sediments resulted in the isolation of four xanthones, eight anthraquinones and five alkaloids, including a new xanthone, oxisterigmatocystin D (1) and a new alkaloid, aspergillusine A (13). High resolution electron impact mass spectrometry (HR-EI-MS), FT-IR spectroscopy, and NMR techniques were used to elucidate the structures of these compounds, and the absolute configuration of compound 1 was established by its NMR features and coupling constant. Furthermore, the biosynthesis pathway of these xanthones and anthraquinones were deduced, and their antioxidant activity and cytotoxicity in human cancer cell lines (HTC-8, Bel-7420, BGC-823, A549, and A2780) were evaluated. The trolox equivalent antioxidant capacity (TEAC) assay indicated most of the xanthones and anthraquinones possessing moderate antioxidant activities. The Nrf2-dependent luciferase reporter gene assay revealed that compounds 6, 7, 9, and 12 potentially activated the expression of Nrf2-regulated gene. In addition, compounds 5 and 11 showed weak cytotoxicity on A549 with the IC50 values of 25.97 and 25.60 μmol·L-1, respectively.

Di(2-ethylhexyl) phthalate (DEHP), a manufactured chemical, is suitable for large-scale production and has extensive applications. Although restricted for use, DEHP is still ubiquitous in the environment and shows potential to disrupt the structure or function of the thyroid system. However, its toxic mechanism is complex and not clearly understood. In this study, a battery of methods was employed to investigate DEHP-induced thyroid-disrupting effects and their mechanism of action, focusing on a newly discovered membrane receptor-mediated mechanism. The results showed that DEHP promoted rat pituitary tumor (GH3) cell proliferation and c-fos gene expression at environment level concentrations (2 and 5 μmol/L) in a manner similar to that of the natural thyroid hormone 3,3′,5-triiodo-L-thyronine (T3). The macromolecule DEHP-BSA cannot pass through the cell membrane to interact with nuclear receptors but upregulated the c-fos gene expression when administered at concentrations comparable to DEHP concentrations; molecular docking demonstrated that DEHP has affinity for the membrane receptor integrin αvβ3; DEHP at 2 μmol/L upregulated the β3 gene expression in GH3 cells; after the addition of integrin αvβ3-inhibiting RGD peptide, DEHP-induced c-fos gene upregulation decreased. All of these findings support the supposition that DEHP-induced thyroid-disrupting effects might be mediated by the membrane receptor integrin αvβ3. Moreover, DEHP activated the downstream extracellular regulated protein kinase (ERK1/2) pathway, upregulating the gene expression of raf-1, MEK-1 and MAPK1 and increasing the protein levels of p-ERK; interestingly, ERK1/2 activation and c-fos upregulation induced by DEHP were attenuated by PD98059 (an ERK1/2 inhibitor). Taken together, the data suggest that the membrane receptor integrin αvβ3 and the downstream ERK1/2 pathway might be involved in DEHP-induced thyroid-disrupting effects. This study provides new insight into the thyroid-disrupting effect and the underlying mechanism and will advance the effort to construct adverse outcome pathways of DEHP and other thyroid hormone disrupting chemicals.

Objective The aim of the present study was to determine how many sentinel lymph nodes (SLNs) are appropriate for predicting non-SLN metastasis in breast cancer. Methods The association between clinicopathological features and non-SLN metastasis was retrospectively analyzed in 472 patients who underwent axillary lymph node dissection (ALND) following SLN biopsy. Another 251 patients who underwent only SLN biopsy without ALND were analyzed and followed up for 2 years. Results A large tumor size, positive SLN, and HER-2 positivity were independent predictors of non-SLN metastasis. There were significant differences in non-SLN metastasis between patients with one negative SLN and patients with an absence of negative SLNs. There was no significant difference in non-SLN metastasis between patients with one negative SLN and two or more negative SLNs. The recurrence-free survival rate for patients who did not undergo ALND was 99.6% (245/246). Conclusion Surgeons should ensure that the number of SLNs obtained is appropriate. The presence of one negative SLN is enough in SLN biopsy. Considering the invasiveness of the surgery, two or more negative SLNs may be unnecessary.

BACKGROUND Pelvic organ prolapse (POP) brings major health issues for women, affecting 40% of postmenopausal women, and directly affects bladder and bowel function, as well as quality of life. In light of the projected growth in demand for care for pelvic floor disorders, determining the etiology and progression of POP has important public health implications. MATERIAL AND METHODS Uterosacral ligaments (USLs) samples of POP patients and normal controls were enrolled for RNA-Seq, and functional annotation analysis and Protein-Protein interaction (PPI) networks construction were performed for differentially expressed genes (DEGs). RESULTS A total of 81 DEGs were identified between POP and normal control, and distinctly classify all samples into normal and POP group by hierarchical clustering. Sixty-six DEGs demonstrated the same expression pattern among the POP samples with different stages. For those DEGs, canonical Wnt receptor signaling pathway was the most significantly enriched GO term (P value=3.33E-07), and neuroactive ligand-receptor interaction was the most significantly enriched pathway (P value=1.24E-03). In The PPI networks of 81 dysregulated genes, significant hub proteins contained TOP2A (Degree=54), KCNA5 (Degree=22) and PLA2G2A (Degree=19), suggesting their important role in the development of POP. CONCLUSIONS This RNA-seq analysis identified a POP signature of 81 genes, and some ECM-related genes, including COMP, NDP, and SNAI2 might participate in the pathology of POP and be applied as potential therapeutic targets.

Tissue-based gene expression data analyses, while most powerful, represent a significantly more challenging problem compared to cell-based gene expression data analyses, even for the simplest differential gene expression analyses. The result in determining if a gene is differentially expressed in tumor vs. non-tumorous control tissues does not only depend on the two expression values but also on the percentage of the tissue cells being tumor cells, i.e., the tumor purity. We developed a novel matched-pairs feature selection method, which takes into full consideration of the tumor purity when deciding if a gene is differentially expressed in tumor vs. control experiments, which is simple, effective, and accurate. To evaluate the validity and performance of the method, we have compared it with four published methods using both simulated datasets and actual cancer tissue datasets and found that our method achieved better performance with higher sensitivity and specificity than the other methods. Our method was the a matched-pairs feature selection method on gene expression analysis under matched case-control design which takes into consideration the tumor purity information, which can set a foundation for further development of other gene expression analysis needs.

Polycystic ovary syndrome (PCOS) is a complex and heterogeneous endocrine disorder. To understand the pathogenesis of PCOS, we established rat models of PCOS induced by letrozole and employed deep sequencing to screen the differential expression of microRNAs (miRNAs) in PCOS rats and control rats. We observed vaginal smear and detected ovarian pathological alteration and hormone level changes in PCOS rats. Deep sequencing showed that a total of 129 miRNAs were differentially expressed in the ovaries from letrozole-induced rat model compared with the control, including 49 miRNAs upregulated and 80 miRNAs downregulated. Furthermore, the differential expression of miR-201-5p, miR-34b-5p, miR-141-3p, and miR-200a-3p were confirmed by real-time polymerase chain reaction. Bioinformatic analysis revealed that these four miRNAs were predicted to target a large set of genes with different functions. Pathway analysis supported that the miRNAs regulate oocyte meiosis, mitogen-activated protein kinase (MAPK) signaling, phosphoinositide 3-kinase/Akt (PI3K-Akt) signaling, Rap1 signaling, and Notch signaling. These data indicate that miRNAs are differentially expressed in rat PCOS model and the differentially expressed miRNA are involved in the etiology and pathophysiology of PCOS. Our findings will help identify miRNAs as novel diagnostic markers and therapeutic targets for PCOS.

Summary Improving nitrogen (N) remobilization from aboveground to underground organs during yearly shoot senescence is an important goal for sustainable production of switchgrass (Panicum virgatum) as a biofuel crop. Little is known about the genetic control of senescence and N use efficiency in perennial grasses such as switchgrass, which limits our ability to improve the process. Switchgrass aboveground organs (leaves, stems and inflorescences) and underground organs (crowns and roots) were harvested every month over a 3-yr period. Transcriptome analysis was performed to identify genes differentially expressed in various organs during development. Total N content in aboveground organs increased from spring until the end of summer, then decreased concomitant with senescence, while N content in underground organs exhibited an increase roughly matching the decrease in shoot N during fall. Hundreds of senescence-associated genes were identified in leaves and stems. Functional grouping indicated that regulation of transcription and protein degradation play important roles in shoot senescence. Coexpression networks predict important roles for five switchgrass NAC (NAM, ATAF1,2, CUC2) transcription factors (TFs) and other TF family members in orchestrating metabolism of carbohydrates, N and lipids, protein modification/degradation, and transport processes during senescence. This study establishes a molecular basis for understanding and enhancing N remobilization and conservation in switchgrass.

Switchgrass is an important bioenergy crop typically grown in marginal lands, where the plants must often deal with abiotic stresses such as drought and salt. Alamo is known to be more tolerant to both stress types than Dacotah, two ecotypes of switchgrass. Understanding of their stress response and adaptation programs can have important implications to engineering more stress tolerant plants. We present here a computational study by analyzing time-course transcriptomic data of the two ecotypes to elucidate and compare their regulatory systems in response to drought and salt stresses. A total of 1,693 genes (target genes or TGs) are found to be differentially expressed and possibly regulated by 143 transcription factors (TFs) in response to drought stress together in the two ecotypes. Similarly, 1,535 TGs regulated by 110 TFs are identified to be involved in response to salt stress. Two regulatory networks are constructed to predict their regulatory relationships. In addition, a time-dependent hidden Markov model is derived for each ecotype responding to each stress type, to provide a dynamic view of how each regulatory network changes its behavior over time. A few new insights about the response mechanisms are predicted from the regulatory networks and the time-dependent models. Comparative analyses between the network models of the two ecotypes reveal key commonalities and main differences between the two regulatory systems. Overall, our results provide new information about the complex regulatory mechanisms of switchgrass responding to drought and salt stresses. Georgia Research Alliance; Noble Research Institute, LLC The authors thank Georgia Research Alliance for funding support for the presented study here. The commercial affiliation: Noble Research Institute, LLC provided support in the form of salaries for author YT, but did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The specific roles of these authors are articulated in the 'author contributions section'.

MicroRNAs (miRNAs) are a class of noncoding RNAs and function as key regulators of gene expression at the post-transcriptional level. In this study, we found that miR-495 reduces cell growth, induces apoptosis and suppresses the migration of endometrial cancer by directly inhibiting FOXC1 expression. Further analysis revealed that FOXC1 promotes growth and migration and functions as an oncogene in vitro. FOXC1 overexpression reversed the cellular responses mediated by miR-495 in endometrial cancer cells. We also found that miR-495 suppresses the growth of endometrial cancer in vivo. Altogether, these results indicate that miR-495 acts as a tumour suppressor gene by targeting FOXC1 at the post-transcriptional level in endometrial cancer.

SEPALLATA (SEP) MADS-box genes play crucial roles in the regulation of floral growth and development. They are required for the specification of sepals, petals, stamens, and carpels as well as for floral determinacy. SEPs perform their functions through the formation of homo- or hetero-polymers, which are the molecular basis of floral quartets. In vitro assays indicated that SEP3 forms a tetramer after binding to DNA, but it is unclear whether DNA binding induces the tetramer, because SEP3 is often reported to form a dimer. Here, we analyzed the oligomeric status of SEP3 domains in the absence of the DNA-binding MADS-box domain. The truncated SEP3 was constructed as a fusion protein and expressed in prokaryotic cells. The purified protein fragment displayed as a tetramer in the size exclusion chromatographic column, and a glutaraldehyde cross-linking assay demonstrated that the protein contained a dimer unit. Yeast two-hybrid tests further verified that the fragments form homologous polymers in vivo, and that the K domain is involved in tetramer formation. Current results imply that the SEP3 protein regulates the formation of flower meristems using the tetramer as a unit, and that the DNA-binding MADS-box is dispensable for polymer formation. The C-terminal region does not contribute to homo-tetramer formation, but it may be reserved to glue other proteins.

High chilling-susceptibility is becoming the bottleneck for cultivation and commercialization of Jatropha curcas L. For insights to chilling resistance ability of this plant species, a cold response transcription factor, JcCBF2, was cloned and studied. It codes a 26 kDa protein, which contains all conserved motifs unique to the C-repeat binding factor (CBF) family and has high similarity to CBFs of Ricinus communis and Populus. Its transcripts express specifically in leaves of Jatropha at cold temperature. After transmitting the report vector, 35S::JcCBF2-GFP, into Arabidopsis thaliana, JcCBF2 protein is main detected in cell nucleus, being consistent to the nuclear orientation signal in its N-terminal. Compared to the control Arabidopsis, the frozen leaves of JcCBF2-overexpressed seedlings grow stronger with less malondialdehyde, smaller leaf conductivity and activer superoxide dismutase, showing their higher freezing tolerance. RT-PCR tests revealed that JcCBF2 functioned mainly at the early stage (0-6 h) of resistance events in Arabidopsis, and its transcripts reduced after 6 h. In addition, JcCBF2 could quickly regulate transcripts of some cold-responsive (COR) genes such as RD29A, COR105A and COR6.6, also during the early stage of frozen treatment. This study not only proves the chilling resistance roles of JcCBF2, but also presents a candidate gene engineering for improvement of chilling tolerance in J. curcas.

Atherosclerosis (AS) is the leading cause of death in modern societies. Active substance from Traditional Chinese Medicine has been used for the treatment of AS, such as ligustrazine and puerarin. However, the pathogenesis of AS and the curative mechanisms of ligustrazine and puerarin stay unclear. In this work, we attempted to figure out these questions using a rat AS model and digital gene expression (DGE) system. Our results showed that DGE sequencing outcomes were high quality and reproductively. Differentially expressed genes were obtained from different comparisons. The Gene Ontology (GO) analysis revealed that mainly enriched GO terms due to the drug treatment were the same as those obtained from the control group vs. the AS model group. Pathway analysis indicated that metabolic pathways, oxidative phosphorylation, and PPAR single pathways were enriched in all comparisons. Our work provided a comprehensive basis for a better understanding of the pathogenesis of AS and the curative mechanisms of ligustrazine and puerarin.

The oxytocinergic system is crucial for sociality and well-being and is associated with empathy. It is suggested that the oxytocinergic system exerts context- and person-dependent effects. We examined how sexual sadistic contexts influenced the effects of the oxytocinergic system on empathic-related behaviors and brain activity in healthy adults. Combining genetic neuroimaging, pharmacological techniques and a psychological paradigm of empathy, we recorded EEG neural responses in female OXTR rs53756 G/G and A/A carriers and measured subjective empathic ratings after intranasal administration of oxytocin/placebo in healthy male adults during the perception of painful facial expressions in sadistic/general social contexts. The results revealed that sadistic contexts modulate oxytocinergic effects on empathy at both behavioral and neural levels. The oxytocinergic system preferentially modulated empathic responses to sadistic contexts. These effects are moderated by individual’s trait empathy. Our combined genetic-pharmacological-imaging results provide a neurochemical mechanism for sadistic context-dependent effects of the oxytocinergic system on empathy.

The zinc transporter SLC39A4 influences epithelial cell morphology and migration in various cancers; however, its role in regulating cell invasion and chemotherapeutic resistance in human lung cancer is not yet clear. Here, integrated analysis of gene expression in non-small cell lung cancer revealed that SLC39A4 expression is significantly correlated with increased tumour size and regional lymph node spread, as well as shorter overall survival (OS) and disease-free survival (DFS). SLC39A4 silencing by lentivirus-mediated shRNA blocked human lung cancer cell epithelial-mesenchymal transition and metastasis in vitro and in vivo, respectively. Moreover, SLC39A4 knockdown enhanced cancer cell sensitivity to cisplatin-induced death by inhibiting stemness in lung cancer cells. Collectively, these data suggest that SLC39A4 may be a novel therapeutic target and predictive marker of tumour metastasis in non-small cell lung cancer.

Carboxylesterases (CCEs) are one of three large detoxification enzyme families. Some CCEs are active on synthetic insecticides with ester structures. Anopheles sinensis is an important malaria vector in eastern Asia. This study identified and characterized the CCE genes in the A. sinensis genome and determined CCE genes associated with pyrethroid resistance using RNA sequencing (RNA-seq) and quantitative reverse transcription - polymerase chain reaction (qRT-PCR), in A. sinensis from Anhui, Chongqing, and Yunnan in China.Fifty-seven putative CCEs were identified and placed into three classes, 12 subfamilies and 14 clades through phylogenetic and homology analyses. Exon sizes ranged from 31 to 4317 bp, with 49 CCEs having two to five exons and eight having six to 11 exons. A total of 183 introns were recognized with sizes ranging from 31 to 4317 bp. The 57 CCEs were located on 14 scaffolds, with 70% located on four scaffolds. The alpha-esterase subfamily was significantly expanded compared with that of Anopheles gambiae. In a pyrethroid-resistant strain, RNA-seq detected five upregulated CCE genes and qRT-PCR detected 12 upregulated CCE genes. The α-esterase 10 (AsAe10) and acetylcholinesterase 1 (AsAce1) genes were the main CCE genes associated with pyrethroid resistance.This information will be useful for further study of the CCE gene family and pyrethroid resistance mechanisms mediated by CCEs. © 2017 Society of Chemical Industry.

FHL2 is a LIM domain protein that is able to form various protein complexes and regulate gene transcription. Recent findings showed that FHL2 is a potential tumor suppressor gene that was down-regulated in hepatocellular carcinoma. In the present study, microarray profiling of gene expression was performed to identify the genes regulated by FHL2 in mouse livers. The differentially expressed genes were further analyzed by bioinformatics tools including DAVID, KEGG, and STRING. Our data illustrate that FHL2 affects genes involved in various functions including signal transduction, responses to external stimulus, cancer-related pathways, cardiovascular function and regulation of actin cytoskeleton. Moreover, a network of differentially expressed genes identified in this study and known FHL2-interacting proteins was constructed. Then, genes identified by bioinformatics tools and most functional relevant to FHL2 were selected for further validation. Finally, the differential expression of Ar, Id3, Inhbe, Alas1, Bcl6, Pparδ, Angptl4, and Erbb4 were confirmed by quantitative real-time PCR. In summary, we have established a database of genes that are potentially regulated by FHL2 and these genes should be future targets for the elucidation of functional roles of FHL2. J. Cell. Biochem. 115: 744–753, 2014. © 2013 Wiley Periodicals, Inc.

Rad9 protein plays an important role in cell-cycle checkpoint signal transduction in human and yeast cells, but knowledge about Rad9 in plants is limited. This study reports that the Rad9 gene of rice can generate the transcript products OsRad9.1 and OsRad9.2 through alternative splicing. OsRad9.1, with all nine exons, is the main cell-cycle checkpoint protein involved in the response of rice to genotoxic stresses (ultraviolet radiation and antibiotic stress), environmental stresses (drought, salt, and heavy metal stress), and auxin stimuli (2,4-D, IAA, and IBA). Meanwhile, transcript isoform OsRad9.2, which lost exon7 and exon8, showed different preferential stimulation effects on these stresses and pollen development duration. These results might indicat that besides the monitoring and repair of DNA damage, Rad9 might involve in the development of pollen.

Mouse ovarian surface epithelium (OSE) is a single layer of cubodial epithelial cells that covers the ovary surface and is involved in regulating the secretion and transport of 17β-hydroxysteroid dehydrogenase. Recently, OSE cells have attracted particular interest as a major source of ovarian cancer. Death-associated protein DAXX along with PML (promyelocytic leukemia protein) nuclear bodies (PML-NBs) reportedly play roles in transcriptional regulation and apoptosis. However, little is known regarding a role for DAXX in mOSE cells. In this study, we both over-expressed DAXX and depleted DAXX in primary mOSE cells. We found that Daxx deletion accelerated senescence in a p53/p21-dependent manner and promoted DNA damage by interacting with PML bodies without affecting cell cycle progression. These results suggest that DAXX may transform mOSE cells to an ovarian oncogenic phenotype and may be an anti-cancer target.