Search

The variable diapause features of bivoltine silkworm (Bombyx mori) strains regulated by environmental signals in the embryonic stage are closely related to epigenetics. Previously, we showed that the expression of YTHDF3 is significantly different in the pupae of the bivoltine silkworm Qiufeng developed from eggs incubated at a normal temperature (QFHT, diapause egg producer) compared to those from eggs incubated at a low temperature (QFLT, nondiapause egg producer), indicating that the expression of diapause-associated genes is regulated by the m6A modification level. However, how YTHDF3 regulates the expression of diapause-related genes remains unclear. In this study, we observed that the knockdown of B. mori YTHDF3 resulted in delayed embryo development, while the overexpression of YTHDF3 resulted in the transformation of nondiapause-destined eggs into a mixture of diapause and nondiapause eggs. Further studies showed that YTHDF3, as a reading protein, can recognize the m6A site of Cyp307a1 and Cyp18a1 genes in the ecdysone synthesis pathway (ESP), and the overexpression of YTHDF3 affects the diapause traits of the silkworm by decreasing the stabilities of mRNAs of Cyp307a1 and Cyp18a1 and inhibiting their translation. The above results demonstrate that m6A modification mediates YTHDF3 to affect the expression levels of its target genes, Cyp307a1 and Cyp18a1, in the ESP to regulate diapause in bivoltine B. mori. This is the first report of the m6A methylation regulation mechanism in diapause in B. mori and provides new experimental data for clarifying the diapause regulation network. [ABSTRACT FROM AUTHOR]

Objective. To screen CXC chemokines related to the risk of lung adenocarcinoma (LUAD) using bioinformatics and construct a CXC-based prognostic risk model to improve the diagnosis and treatment of LUAD patients. Methods. The Cancer Genome Atlas (TCGA) database and Gene Expression Omnibus (GEO) database were searched to obtain mRNA expression data and clinicopathological information of LUAD patients. CXC genes differentially expressed in LUAD were screened using the R packages. Further, risk factors significantly associated with the survival of LUAD patients were obtained by the univariate Cox proportional hazard regression, LASSO regression, and multivariate Cox proportional hazard regression analysis, following which a risk prediction model was constructed. The performance of the CXCL13-based model in predicting the prognosis of low-risk and high-risk effect LUAD patients was verified, and the association between the model and the degree of tumor immune cell infiltration was investigated. Results. CXCL13 was significantly highly expressed in the cancer tissues of LUAD patients. The risk of death in patients with highly expressed CXCL13 was about 1.5 times higher than in those with lowly expressed CXCL13 (HR = 1.5153357). CXCL13-based risk scoring showed that the high-risk score of LUAD patients was significantly correlated with poor prognosis, but no relation between the two was found in the GEO validation sets, suggesting that this risk model may not be accurate enough. In addition, activated B cells, CD4+ T cells, CD8+ T cells, and dendritic cells were significantly positively correlated with the high risk of LUAD. Conclusions. Although we found that a high expression of CXCL13 was associated with a high risk of death and immune cell infiltration and activation in LUAD patients, the CXCL13-based risk model was not accurate enough for predicting the prognosis of LUAD patients. [ABSTRACT FROM AUTHOR]

Objective. Red raspberry serves as a proven natural product to produce anti-inflammatory, antioxidant, and anticancer functions, but limited findings are available on its effects on depression. This study, by using a chronic unpredictable mild stress- (CUMS-) induced depression model, thus investigated the effects and underlying mechanism of red raspberry extract (RRE) on depressive behavior, inflammation, and oxidative stress. Methods. Different treatments were given after random grouping of Sprague-Dawley rats, including no intervention (control), CUMS induction, and CUMS+different concentrations of RRE, and subsequently, depression-like behavior tests were performed. HE staining was designed to observe the pathological damage of the hippocampal tissue in rats. The levels of oxidative stress, endocrine hormones, and inflammatory factors were determined by biochemical assay and ELISA, and gene expression (mRNA and protein) in the hippocampal tissue by qRT-PCR and Western blot. Results. On completion of CUMS treatment, the rats showed severe depression-like behavior, with obvious hippocampal tissue damage, oxidative inflammatory response, and endocrine imbalance. Importantly, RRE treatment significantly improved such depression-like behavior and attenuated histopathological damage in CUMS rats when reducing inflammation and oxidative stress and endocrine imbalance with upregulation of glutathione (GSH), superoxide dismutase (SOD), and interleukin- (IL-) 10 and downregulation of adrenocorticotropic hormone (ACTH), corticosterone (CORT), malondialdehyde (MDA), IL-1β, cyclooxygenase- (COX-) 2, and human macrophage chemoattractant protein- (MCP-) 1. In addition, for CUMS rats, RRE was a contributor to increasingly expressed brain-derived neurotrophic factor (BDNF), neurotrophic tyrosine receptor kinase 2 (TrkB), and p-mTOR but inhibited p-GSK-3β expression in the hippocampal tissue. All the above antidepressant effects of RRE were concentration-dependent. Conclusion. By regulating neuroinflammation, oxidative stress response, endocrine level, and BDNF/TrkB level, RRE showed potential efficacy in alleviating depression-like behavior and histopathological damage of hippocampal tissue in CUMS rats by regulating the GSK3β and mTOR signaling pathways. [ABSTRACT FROM AUTHOR]

This study aimed to explore the regulatory mechanisms of age-related cataract (ARC) formation. Cataracts in zebrafish were induced by injecting hydrogen peroxide into the fish anterior chamber. The mRNA and miRNA expression profiles of the lens from H2O2-injected and PBS-injected zebrafishes were detected by RNA sequencing. The LIMMA package was applied to identify differentially expressed genes (DEGs). Gene Ontology categories were enriched by the R "cluster Profiler" package and Kyoto Encyclopedia of Genes and Genomes pathway enrichment was performed based on hypergeometric distribution using the R "phyper" function. The protein-protein interaction network of DEGs was built via the STRING. Target genes of differentially expressed miRNAs (DEmiRs) were predicted by miRanda. Furthermore, DEGs were selected as DEmiR targets and a DEmiR-DEG regulatory network was constructed via Cytoscape. In total, 3689 DEGs (such as opn1mw4, LOC103908930, si:dkeyp-1h4.8, crispld1b, cyp1a, and gdpd3a) including 2478 upregulated and 1211 downregulated genes were identified. 177 DEmiRs (such as dre-miR-96-3p, dre-miR-182-5p, dre-miR-9-7-3p, and dre-miR-124-4-5p) including 108 upregulated and 69 downregulated miRNAs were detected. The DEGs are involved in cell death, DNA repair, and cell development-related pathways. A protein-protein interaction network including 79 node genes was constructed to explore the interactions of DEGs. Furthermore, a DEmiR-DEG regulatory network focusing on the DNA repair process was constructed, including 21 hub DEGs and 15 hub DEmiRs. We identified several DEGs and constructed a miRNA-mRNA regulatory network related to the DNA repair process in a zebrafish cataract model. These genes participate in the oxidative stress response of lens epithelium cells and finally contribute to the formation of zebrafish cataracts. The hub DEGs and hub DEmiRs could be potential therapeutic targets for ARC. [ABSTRACT FROM AUTHOR]

To study the regulatory function of Bombyx mori (B. mori) miRNAs (bmo-miR) on the expression of fibroin light chain gene (BmFib-L), the 3'UTR of BmFib-L mRNA was used as the target for online prediction of miRNAs from miRBase using RNAhybrid Software, and miR-2845 was screened out. First, the expression profiles of miR-2845 and BmFib-L in larvae of the 5th instar were analyzed by Real-time quantitative PCR (RT-qPCR). Then recombinant plasmids (pcDNA3.0-pre-miR-2845 and pGL3.0-BmFib-L) were constructed to use for the expression of miR-2845 and BmFib-L 3'UTR, respectively. Cellular-level functional verification of miR-2845 on BmFib-L was carried out using multiple experimental methods (including dual luciferase reporter vectors, artificially synthesized mimics and inhibitors, and target site mutations). Finally, in vivo functional verification was performed by injecting the recombinant vector in 5th instar larvae. BmFib-L expression levels were detected using RT-qPCR in the posterior silk glands (PSG) of the injected larvae. Results showed that the expression of miR-2845 increased between the 1st and 5th day in 5th instar larvae, but began to decline on the 5th day, while the expression of the target gene BmFib-L increased sharply. This suggests that miR-2845 and BmFib-L expression levels show opposing trends, implying a negative regulatory relationship. In BmN cells, miR-2845 significantly down-regulated the expression of BmFib-L; the inhibitory effect of miR-2845 on BmFib-L was disappeared after mutation of the targeting site on 3'UTR of BmFib-L; in individuals, miR-2845 significantly down-regulated BmFib-L expression levels. Our results provide new experimental data for clarifying the molecular regulation mechanism of silk protein expression. [ABSTRACT FROM AUTHOR]

[Display omitted] • The expression levels of BmINR and BmAC6 are significantly higher in NDEPs compared to DEPs. • Knockdown of BmINR upregulates the 3-hydroxykynurenine synthesis pathway and results in a change in the diapause phenotype. • Overexpression of BmINR or BmAC6 highly upregulates the expression levels of key genes related to carbohydrate metabolism. • BmINR and BmAC6 play a crucial role in regulating embryonic diapause via the insulin/IGF signaling pathway. Diapause of the silkworm (Bombyx mori) is an important ecological adaptation strategy regulated by multiple signaling pathways. As an evolutionarily conserved signaling pathway, the insulin/IGF signaling (IIS) pathway is essential in regulating lifespan, energy accumulation, and stress resistance in diapause insects. However, the regulatory mechanism of IIS on diapause in B. mori is still not fully understood. To investigate the role of the IIS pathway in regulating diapause, we first analyzed the transcription levels of the insulin receptor (BmINR) and its downstream gene adenylate cyclase 6 (BmAC6). The diapause-terminated eggs of a bivoltine strain QiuFeng (V2-QF) were incubated at 25 °C in natural room light for preparing diapause egg producers (DEPs) and at 17 °C in total darkness for preparing non-diapause egg producers (NDEPs), respectively. Then we investigated the effects of BmINR and BmAC6 on diapause phenotype and expression of diapause-related genes by RNA interference (RNAi) and overexpression techniques. The results showed that the mRNA expression levels of BmINR and BmAC6 in the head and ovary of NDEPs were higher than those in DEPs during the early and middle pupal stages. Furthermore, when BmINR was knocked down in the NDEPs, approximately 14.43% of eggs were in light red color and subsequently changed into gray-purple color after 48 h post-oviposition, then stayed in a diapause state. On the other hand, overexpression of BmINR or BmAC6 via recombinant baculoviruses did not cause any obvious phenotypic alterations in NDEPs, but it upregulated the expression of genes related to carbohydrate metabolism, which provides energy for embryonic growth and development. Therefore, it can be concluded that BmINR and BmAC6 genes regulate embryonic diapause in bivoltine B. mori. [ABSTRACT FROM AUTHOR]

Diapause is an important characteristic of insects used to adapt to extreme changes in environmental conditions. Embryonic diapause of the bivoltine silkworm (Bombyx mori) is determined by environmental conditions experienced by the mother while in the embryo stage. If they are incubated at 25°C with natural light, their progenies will be diapause‐destined. If they are incubated at 17°C in darkness, their progenies will be non‐diapause‐destined. The molecular mechanism of diapause remains unknown. In the present study, we analyzed two downregulated genes (BGIBMGA003835, BGIBMGA012335) and two upregulated genes (BGIBMG012996, BGIBMG002426) related to carbohydrate metabolism, verified differentially expressed in ovaries and heads of 1‐day‐old fifth‐instar larvae to 6‐day pupae by quantitative real‐time reverse transcription polymerase chain reaction (qRT‐PCR). In line with published data, the expression level of these genes in larvae were generally lower than in pupae. We further analyzed the expression levels of the four genes in BmN cells that had been treated with various concentrations of diapause hormone (DH). It demonstrated that the expression of these genes was affected by DH. Knockdown of the selected genes in non‐diapause‐destined female pupae changed the fate of the progeny from non‐diapause‐ to daipause‐destined, as seen by the appearance of diapause eggs. Our study provides insight into the molecular mechanism of diapause in B. mori. The expression levels of BGIBMGA003835, BGIBMGA012335, BGIBMG012996 and BGIBMG002426 were different at diverse developmental stages. They can also be influenced by Bmo‐DH. RNAi if several genes can turn over the fate of non‐diapause eggs. [ABSTRACT FROM AUTHOR]

Contemporary therapy for advanced castration-resistant prostate cancer (CRPC) employs reagents such as enzalutamide and abiraterone acetate targeting the androgen receptor (AR) transcription axis only provide a temporary response and rapidly develop resistance. Additionally, neuroendocrine prostate cancer (NEPC) is an AR pathway-independent and lethal-stage prostate cancer with no standard therapy. Qingdai Decoction (QDT), a traditional Chinese medicine formula, has various pharmacological activities and was widely used for the treatment of different diseases including prostatitis which may contribute to prostate cancer development. This study aims to explore the anti-tumor role and potential mechanism of QDT on prostate cancer. CRPC prostate cancer cell models and xenograft mice models were established for research. The effect of TCMs on cancer growth and metastasis were determined by CCK-8, wound-healing assays and the PC3-xenografted mice model. The toxicity of QDT in the major organs was investigated by H&E staining. The compound-target network was analyzed with network pharmacology. The correlation of QDT targets with prostate cancer patient's prognosis was analyzed with multiple prostate cancer patient cohorts. The expression of related proteins and mRNA were detected by western blot and real-time PCR. The gene knockdown was achieved with CRISPR-Cas13 technology. By integrating functional screening, network pharmacology analysis, CRISPR-Cas13 directed RNA targeting, and molecular biology validation in different prostate cancer models and clinical prostate cancer cohorts, we found that Qingdai Decoction (QDT), a Traditional Chinese Medicine, can repress cancer growth in advanced prostate cancer models in vitro and in vivo in an AR independent manner by targeting NOS3, TGFB1 , and NCOA2. This study not only identified QDT as a novel drug for lethal-stage prostate cancer treatment but also provided an extensive Integrative research paradigm for investigating the roles and mechanisms of TCMs for the treatment of other diseases. [Display omitted] • QDT efficiently represses multiple prostate cancer cells growth in vitro and in vivo. • QDT targets are closely related to prostate cancer progression and patient survival. • NOS3, TGFB1 and NCOA2 mediate the antitumor effect of QDT. [ABSTRACT FROM AUTHOR]

Background: Due to experimental batch effects, the application of a quantitative transcriptional signature for disease diagnoses commonly requires inter-sample data normalization, which would be hardly applicable under common clinical settings. Many cancers might have qualitative differences with the non-cancer states in the gene expression pattern. Therefore, it is reasonable to explore the power of qualitative diagnostic signatures which are robust against experimental batch effects and other random factors. Results: Firstly, using data of technical replicate samples from the MicroArray Quality Control (MAQC) project, we demonstrated that the low-throughput PCR-based technologies also exist large measurement variations for gene expression even when the samples were measured in the same test site. Then, we demonstrated the critical limitation of low stability for classifiers based on quantitative transcriptional signatures in applications to individual samples through a case study using a support vector machine and a naïve Bayesian classifier to discriminate colorectal cancer tissues from normal tissues. To address this problem, we identified a signature consisting of three gene pairs for discriminating colorectal cancer tissues from non-cancer (normal and inflammatory bowel disease) tissues based on within-sample relative expression orderings (REOs) of these gene pairs. The signature was well verified using 22 independent datasets measured by different microarray and RNA_seq platforms, obviating the need of inter-sample data normalization. Conclusions: Subtle quantitative information of gene expression measurements tends to be unstable under current technical conditions, which will introduce uncertainty to clinical applications of the quantitative transcriptional diagnostic signatures. For diagnosis of disease states with qualitative transcriptional characteristics, the qualitative REO-based signatures could be robustly applied to individual samples measured by different platforms. [ABSTRACT FROM AUTHOR]

To identify the hub miRNAs and mRNAs contributing to the spontaneous recovery of an H 2 O 2 -induced zebrafish cataract model. Zebrafishes were divided into three groups, i.e., Group A, which included normal control fish (day 0), and Groups B and C, where fish were injected with 2.5% hydrogen peroxide into the anterior chamber and reared for 14 and 30 days, respectively. Fish eyes were examined by stereomicroscope photography and optical coherence tomography (OCT). RNA profiles of fish lenses were detected by RNA sequencing. Differentially expressed genes (DEGs) and differentially expressed miRNAs (DEmiRs) were identified among three groups. The DEGs and DEmiRs, which changed in opposite positions between "B vs. A" and "C vs. B" were defined as ODGs (opposite positions changed DEGs) and ODmiRs (opposite positions changed DEmiRs). Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes pathway (KEGG) analysis were carried out by R language. The protein-protein interaction network (PPI) was constructed using STRING. Potential targets of miRNAs were obtained using miRanda. miRNA-mRNA networks were constructed by Cytoscape. The fish lens opacity formed on day 14 and recovered to transparent on day 30 after injection. Compared to group B, 1366 DEGs and 54 DEmiRs were identified in group C. "C vs. B" DEGs were enriched in gene clusters related to development and oxidative phosphorylation. Target genes of DEmiRs were enriched in clusters such as development and cysteine metabolism. Among three groups, 786 ODGs and 27 ODmiRs were identified, and 480 ODGs were predicted as targets of ODmiRs. Target ODGs were enriched in pathways related to methionine metabolism, ubiquitin, sensory system development, and structural constituents of the eye lens. In addition, we established an ODmiRs-ODGs regulation network. We identified several hub mRNAs and altered miRNAs in the formation and reversal of zebrafish cataracts. These hub miRNAs/mRNAs could be potential targets for the non-surgical treatment of ARC. • Zebrafish cataract formed on day 14 and spontaneously recovered to transparent on day 30 after H 2 O 2 injection. • Oxidative stress and development related pathways involved in the recovery of zebrafish cataract. • Key genes (such as nat16 , ddb2 , cav1) involved in the reversal of zebrafish cataract were regulated by several miRNAs. [ABSTRACT FROM AUTHOR]