Search

Primary open-angle glaucoma (POAG) is characterized by irreversible neurodegeneration accompanied by visual field defects and high intraocular pressure. Currently, an effective treatment is not available to prevent the progression of POAG, other than treatments to decrease the high intraocular pressure. We performed proteomic analysis of aqueous humour (AH) samples from patients with POAG combined with cataract and patients with cataract to obtain a better understanding of the pathogenesis of POAG and explore potential treatment targets for this condition. Samples were collected from 10 patients with POAG combined with cataract and 10 patients with cataract. Samples from each group were pooled. A high-resolution, label-free, liquid chromatography-tandem mass spectrometry-based quantitative proteomic analysis was performed. In total, 610 proteins were identified in human AH samples from the two groups. A total of 48 up-regulated proteins and 49 down-regulated proteins were identified in the POAG combined with cataract group compared with the control group. Gene Ontology (GO) analysis revealed key roles for these proteins in inflammation, immune responses, growth and development, cellular movement and vesicle-mediated transport in the biological process category. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated the down-regulated expression of glutathione S-transferase P (GSTP1) in the glutathione metabolism signalling pathway in the POAG combined with cataract group. Additionally, certain significantly differentially expressed proteins in the proteomic profile were verified by enzyme-linked immunosorbent assay (ELISA). GSTP1 levels were reduced in the human AH samples from the POAG combined with cataract group, based on the results of ELISA and proteomic profiling. Therefore, GSTP1, a redox-related marker, may be involved in the pathological process of POAG and may become a treatment target in the future., (© 2021 The Authors. Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd.)

Zinc finger E-box binding homeobox factor 1 (ZEB1), as a crucial mediator of "epithelial-mesenchymal transition," contributes to malignant progression of various epithelial tumors. However, its involvement in human esophageal squamous cell carcinoma (ESCC) remains unclear. In order to investigate the expression pattern of ZEB1 in ESCC tissues and evaluate its associations with tumor progression and patients' prognosis, 100 pairs of formalin-fixed and paraffin-embedded cancerous and adjacent noncancerous tissues from patients with ESCC were used to detect the expression pattern of ZEB1 by immunohistochemistry. Then, the association between ZEB1 expression, clinicopathological parameters, and prognosis of ESCC was examined. We also performed migration and invasion assays of small interfering RNA (siRNA)-targeted ZEB1-transfected cells in vitro. As a result, expression level of ZEB1 was significantly higher in ESCC tissues compared to that in adjacent noncancerous tissues (P < 0.001). High expression of ZEB1 was observed in 55.00 % (55/100) of ESCCs. In addition, high ZEB1 expression was found to be closely correlated with advanced tumor stage (P = 0.001), positive lymph node metastasis (P = 0.001), great tumor depth (P = 0.03), and high histologic grade (P = 0.008). Moreover, multivariate analysis showed that the status of ZEB1 expression was an independent predictor for overall survival in ESCC. Furthermore, knockdown of ZEB1 by transfection of siRNA-ZEB1 abrogated the migration and invasion of ESCC cells in vitro. Taken together, our data offer the convincing evidence that ZEB1 may play a crucial role in promoting aggressive ESCC progression. ZEB1 may serve as an effective prognostic marker and a potential target for therapeutic intervention of ESCC.

Wheat end-use quality mainly derives from two interrelated characteristics: the compositions of gluten proteins and grain hardness. The composition of gluten proteins determines dough rheological properties and thus confers the unique viscoelastic property on dough. One group of gluten proteins, high molecular weight glutenin subunits (HMW-GS), plays an important role in dough functional properties. On the other hand, grain hardness, which influences the milling process of flour, is controlled by Puroindoline a (Pina) and Puroindoline b (Pinb) genes. However, little is known about the combined effects of HMW-GS and PINs on dough functional properties. In this study, we crossed a Pina-expressing transgenic line with a 1Ax1-expressing line of durum wheat and screened out lines coexpressing 1Ax1 and Pina or lines expressing either 1Ax1 or Pina. Dough mixing analysis of these lines demonstrated that expression of 1Ax1 improved both dough strength and over-mixing tolerance, while expression of PINA detrimentally affected the dough resistance to extension. In lines coexpressing 1Ax1 and Pina, faster hydration of flour during mixing was observed possibly due to the lower water absorption and damaged starch caused by PINA expression. In addition, expression of 1Ax1 appeared to compensate the detrimental effect of PINA on dough resistance to extension. Consequently, coexpression of 1Ax1 and PINA in durum wheat had combined effects on dough mixing behaviors with a better dough strength and resistance to extension than those from lines expressing either 1Ax1 or Pina. The results in our study suggest that simultaneous modulation of dough strength and grain hardness in durum wheat could significantly improve its breadmaking quality and may not even impair its pastamaking potential. Therefore, coexpression of 1Ax1 and PINA in durum wheat has useful implications for breeding durum wheat with dual functionality (for pasta and bread) and may improve the economic values of durum wheat.

Objective: To explore the possible mechanism of scalp acupuncture (SA) in relieving cerebral ischemia reperfusion (CI-R) injury., Methods: Seventy male SD rats were randomized into sham-operation (sham, n = 10), model (n = 30) and SA (n = 30) groups. The later 2 groups were further divided into 24 h, 48 h and 72 h subgroups respectively, with 10 cases in each. CI-R model was established by middle cerebral artery occlusion (MCAO) and reperfusion. Electroacupuncture (2 mA, 2 Hz/100 Hz) was applied to "Dingnie Houxiexian" (MS 7) and "Dingnie Qianxiexian" (MS 6) for 30 min, once every 24 h. Changes of the animal behavior were observed by using neurological severity score (NSS), nuclear factor-kappa B (NF-kappaB) and cyclooxygenase (COX-2) contents and their mRNA expression were detected with enzyme-linked immunosorbent assay (ELISA) and polymerase chain reaction (PCR) techniques respectively., Results: Following modeling, the NSS at 24 h, 48 h and 72 h after MCAO increased significantly, while compared with model group, NSS of SA group at the 3 time-points decreased considerably (P < 0.05, P < 0.01), suggesting an improvement of the neurological functions after SA treatment. In comparison with sham group, NF-kappaB mRNA and COX-2 mRNA, and NF-kappaB and COX-2 protein expression of model group in the infarcted cerebral tissue were significantly upregulated at the 3 time-points (P < 0.01) except NF-KB mRNA at 72 h (no significant change), while compared with the 3 time-points of model group, NF-kappaB mRNA and COX-2 mRNA, and NF-KB and COX-2 protein expression of SA group were downregulated obviously (P < 0.01) except COX-2 mRNA at 72 h., Conclusion: Scalp acupuncture can suppress cerebral ischemia-induced upregulation of NF-kappaB mRNA and COX-2 mRNA, and NF-kappaB and COX-2 protein expression, which may contribute to its effect in promoting neurofunctional rehabilitation of CI-R rats by reducing cytokines-mediated inflammatory reaction.

AML1-ETO fusion protein, a product of leukemia-related chromosomal translocation t(8;21), was reported to upregulate expression of connexin-43 (Cx43), a member of gap junction-constituted connexin family. However, its mechanism(s) remains unclear. By bioinformatic analysis, here we showed that there are two putative AML1-binding consensus sequences followed by two activated protein (AP)1 sites in the 5'-flanking region upstream to Cx43 gene. AML1-ETO could directly bind to these two AML1-binding sites in electrophoretic mobility shift assay, but luciferase reporter assay revealed that the AML1 binding sites were not indispensable for Cx43 induction by AML1-ETO protein. Conversely, AP1 sites exerted an important role in this event. In agreement, AML1-ETO overexpression in leukemic U937 cells activated c-Jun N-terminal kinase (JNK), while its specific inhibitor SP600125 effectively abrogated AML1-ETO-induced Cx43 expression, indicating that JNK signaling pathway contributes to AML1-ETO induced Cx43 expression. These results would shed new insights for understanding mechanisms of AML1-ETO-associated leukemogenesis.

Leukotriene B4 acts through its receptors, BLT(1) and BLT(2), however, their expression in rheumatoid arthritis is unknown. In this experiment, BLT(1) and BLT(2) mRNA expressions in the synovium of rats with collagen-induced arthritis (CIA) at days 1, 3, 7 and 14 after CIA onset were analyzed by RT-PCR. The expression of two immunological and inflammatory factors, S100A8 and S100A9, in the synovium of the arthritic rats was also determined at the indicated time. At d14, the differential expressions of BLT(1) and BLT(2) in the synovium, spleen, peripheral blood mononuclear cells (PBMC) and thymus of CIA rats were analyzed. The results showed that, in the synovium of the arthritic rats, the BLT(1) mRNA expression increased after CIA onset, reached the highest value between d1 and d3, and declined afterwards while the BLT(2) expression increased with time and reached its peak at d14. Both S100A8 and S100A9 expression reached the peak levels between d1 and d3, and decreased to lower levels between d7 and d14. For the analyzed tissues from CIA rats at d14, BLT(1) mRNA was expressed in the thymus with the highest level, followed by the spleen, PBMC and synovium. BLT(2) mRNA was expressed in the thymus the highest as well, but followed by the synovium, spleen and PBMC. Since BLT(1) and BLT(2) play distinct roles during CIA, this study may provide basis for new therapies targeting BLT(1) and BLT(2), respectively, for the treatment of arthritic inflammation at different stages.

Ganoderma has received much attention for its medicinal value, but the manipulation of multiple genes remains a challenge, hindering the genetic engineering of this species for the development of cell factories. Here, we first showed that the presence of an intron is necessary for the efficient expression of the endogenous cDNA of carboxin-resistant gene (cbx) in G. lucidum. Then, the self-cleaving function of 2 A peptide was investigated in G. lucidum by linking cbx cDNA to the codon-optimized hygromycin B-resistant gene (op hph) using the 2A-peptide sequence. The results showed that cbx cDNA and op hph can be successfully expressed in G. lucidum in a bicistronic manner from a single transcript. Moreover, the expression of both genes was not affected by the order within the 2 A cassette. In addition, simultaneous expression of cbx cDNA, op hph , and codon-optimized yellow fluorescent protein gene (op yfp) was conducted for the first time in G. lucidum using the 2 A peptide-based approach. The developed method was successfully applied to express both cDNA of the 3-hydroxy-3-methylglutaryl coenzyme A reductase (hmgr) and squalene epoxidase gene (se) for enhanced production of ganoderic acids (GAs) in G. lucidum. The engineered strain produced the maximum content of GA-Mk, GA-T, GA-S, and GA-Me were 26.56±3.53,39.58±3.75, 16.54±2.16, and 19.1±1.87 μg/100 mg dry weight, respectively. These values were 3.85-, 4.74-, 3.65-, and 3.23-fold higher than those produced by the control strain. The developed method will be useful for the manipulation of complex metabolic or regulatory pathways involving multiple genes in Ganoderma. • The intron is required for the efficient expression of cbx cDNA in G. lucidum. • A 2A peptide-based multigene expression system was developed in G. lucidum. • Co-expression of hmgr and se cDNA improved production of ganoderic acids in G. lucidum. [ABSTRACT FROM AUTHOR]

Vitellogenin (Vtg) is a precursor of yolk proteins in egg-laying vertebrates and invertebrates and plays an important role in vitellogenesis and embryonic development. However, the Vtg family remains poorly characterized in Exopalaemon carinicauda, a major commercial mariculture species found along the coasts of the Yellow and Bohai Seas. In this study, 10 Vtg genes from the genomes of E. carinicauda were identified and characterized. Phylogenetic analyses showed that the Vtg genes in crustaceans could be classified into four groups: Astacidea, Brachyra, Penaeidae, and Palaemonidae. EcVtg genes were unevenly distributed on the chromosomes of E. carinicauda, and a molecular evolutionary analysis showed that the EcVtg genes were primarily constrained by purifying selection during evolution. All putative EcVtg proteins were characterized by the presence of three conserved functional domains: a lipoprotein N-terminal domain (LPD_N), a domain of unknown function (DUF1943), and a von Willebrand factor type D domain (vWD). All EcVtg genes exhibited higher expression in the female hepatopancreas than in other tissues, and EcVtg gene expression during ovarian development suggested that the hepatopancreas is the main synthesis site in E. carinicauda. EcVtg1a, EcVtg2, and EcVtg3 play major roles in exogenous vitellogenesis, and EcVtg3 also plays a major role in endogenous vitellogenesis. Bilateral ablation of the eyestalk significantly upregulates EcVtg mRNA expression in the female hepatopancreas, indicating that the X-organ/sinus gland complex plays an important role in ovarian development, mostly by inducing Vtg synthesis. These results could improve our understanding of the function of multiple Vtg genes in crustaceans and aid future studies on the function of EcVtg genes during ovarian development in E. carinicauda. [ABSTRACT FROM AUTHOR]

Background: Renal cell carcinoma (RCC) is one of the most common malignant tumor worldwide. Metastasis is a leading case of cancer-related deaths of RCC. Circular RNAs (circRNAs), a class of noncoding RNAs, have emerged as important regulators in cancer metastasis. However, the functional effects and regulatory mechanisms of circRNAs on RCC metastasis remain largely unknown. Methods: High-throughput RNA sequencing techniques were performed to analyze the expression profiles of circRNAs and mRNAs in highly and poorly invasive clear cell renal cell carcinoma (ccRCC) cell lines. Functional experiments were performed to unveil the regulatory role of circPPAP2B in the proliferation and metastatic capabilities of ccRCC cells. RNA pulldown, Mass spectrometry analysis, RNA methylation immunoprecipitation (MeRIP), RNA immunoprecipitation (RIP), co-immunoprecipitation (CoIP), next-generation RNA-sequencing and double luciferase experiments were employed to clarify the molecular mechanisms by which circPPAP2B promotes ccRCC metastasis. Results: In this study, we describe a newly identified circular RNA called circPPAP2B, which is overexpressed in highly invasive ccRCC cells, as determined through advanced high-throughput RNA sequencing techniques. Furthermore, we observed elevated circPPAP2B in ccRCC tissues, particularly in metastatic ccRCC tissues, and found it to be associated with poor prognosis. Functional experiments unveiled that circPPAP2B actively stimulates the proliferation and metastatic capabilities of ccRCC cells. Mechanistically, circPPAP2B interacts with HNRNPC in a m6A-dependent manner to facilitate HNRNPC nuclear translocation. Subcellular relocalization was dependent upon nondegradable ubiquitination of HNRNPC and stabilization of an HNRNPC/Vimentin/Importin α7 ternary complex. Moreover, we found that circPPAP2B modulates the interaction between HNRNPC and splicing factors, PTBP1 and HNPNPK, and regulates pre-mRNA alternative splicing. Finally, our studies demonstrate that circPPAP2B functions as a miRNA sponge to directly bind to miR-182-5p and increase CYP1B1 expression in ccRCC. Conclusions: Collectively, our study provides comprehensive evidence that circPPAP2B promotes proliferation and metastasis of ccRCC via HNRNPC-dependent alternative splicing and miR-182-5p/CYP1B1 axis and highlights circPPAP2B as a potential therapeutic target for ccRCC intervention. [ABSTRACT FROM AUTHOR]

Simple Summary: This study aimed to investigate the influencing mechanisms of the intestinal microbiota and gene expression of Fenneropenaeus chinensis under low-salinity stress. The 16S rDNA results suggest that the relative abundances of Photobacterium and Vibrio decreased significantly, whereas some bacteria, Shewanella, Pseudomonas, and Lactobacillus, became the predominant communities. Transcriptome sequencing identified numerous differentially expressed genes (DEGs) through various types of N-glycan biosynthesis, amino acid sugar and nucleotide sugar metabolism, and lysosomes to adapt to stress. Correlation analysis between microbiota and DEGs showed that significant changes in Pseudomonas, Ralstonia, Colwellia, and Cohaesibacter were positively correlated with immune-related genes such as peritrophin-1-like and mucin-2-like, and negatively correlated with caspase-1-like. Salinity is an important environmental stress factor in mariculture. Shrimp intestines harbor dense and diverse microbial communities that maintain host health and anti-pathogen capabilities under salinity stress. In this study, 16s amplicon and transcriptome sequencing were used to analyze the intestine of Fenneropenaeus chinensis under low-salinity stress (15 ppt). This study aimed to investigate the response mechanisms of the intestinal microbiota and gene expression to acute low-salinity stress. The intestinal tissues of F. chinensis were analyzed using 16S microbiota and transcriptome sequencing. The microbiota analysis demonstrated that the relative abundances of Photobacterium and Vibrio decreased significantly, whereas Shewanella, Pseudomonas, Lactobacillus, Ralstonia, Colwellia, Cohaesibacter, Fusibacter, and Lachnospiraceae_NK4A136_group became the predominant communities. Transcriptome sequencing identified numerous differentially expressed genes (DEGs). The DEGs were clustered into many Gene Ontology terms and further enriched in some immunity- or metabolism-related Kyoto Encyclopedia of Genes and Genomes pathways, including various types of N-glycan biosynthesis, amino acid sugar and nucleotide sugar metabolism, and lysosome and fatty acid metabolism. Correlation analysis between microbiota and DEGs showed that changes in Pseudomonas, Ralstonia, Colwellia, and Cohaesibacter were positively correlated with immune-related genes such as peritrophin-1-like and mucin-2-like, and negatively correlated with caspase-1-like genes. Low-salinity stress caused changes in intestinal microorganisms and their gene expression, with a close correlation between them. [ABSTRACT FROM AUTHOR]

Objective: To evaluate the relation between the expression of PD‐1, PD‐L1, CD3, CD8, Foxp3 and clinicopathological features in patients with oral leukoplakia (OLK) and oral squamous cell carcinomas (OSCC) as well as the malignant outcome in OLK patients, and to study the effect of PD‐1 and PD‐L1 on immune microenvironment in the progression of oral carcinogenesis. Methods: We evaluated the expression of PD‐1/PD‐L1 and composition of CD3+, CD8+and Foxp3+ T lymphocytes in OLK and OSCC samples by immunohistochemical (IHC) staining and analyzed their relation with clinical information and malignant transformation in OLK patients. Results: IHC staining demonstrated that the expression of PD‐1 was significantly increased in the high‐grade OLK group than in the low‐grade OLK group, while PD‐L1 was detected mainly in OSCC. The expression of CD3, CD8, and Foxp3 was found higher in the high‐grade OLK group than in the low‐grade OLK group, and the Foxp3+ cells were found more in the OSCC group than in the high‐grade OLK group. PD‐1 was significantly correlated with CD3 (p < 0.05, R = 0.52), CD8 (p < 0.05, R = 0.46), and Foxp3 (p < 0.05, R = 0.46), and the low PD‐1‐expression group showed a better malignant‐free survival than high PD‐1 expression group in the OLK (p < 0.05). Conclusion: The PD‐1/PD‐L1 may induce immune suppression in OLK and accelerate the progress of malignant transformation. [ABSTRACT FROM AUTHOR]

Background: Diabetic nephropathy (DN) is a serious microvascular complication of diabetes that may lead to chronic renal failure and end-stage renal disease. Circular RNAs (circRNAs) play important roles in DN progression. However, the action of circRNA ADP ribosylation factor 3 (circ_Arf3) in high glucose (HG)-induced change is still unclear.Methods: Mouse mesangial cells (MCs) were treated with 30 mM HG as a DN cell model in vitro. Quantitative real-time polymerase chain reaction (qRT-PCR) was employed to examine the expression levels of circ_Arf3, microRNA (miR)-452-5p and muscleblind like splicing regulator 1 (Mbnl1). The proliferation of HG-treated MCs was assessed using 5 Ethynyl 2′ deoxyuridine (EdU) and cell counting kit-8 (CCK-8) assays, and the levels of proliferation and fibrosis-related proteins and Mbnl1 were detected by Western blot. Dual-luciferase reporter and RNA pull-down assays were utilized to determine the relationship between miR-452-5p and circ_Arf3 or Mbnl1.Results: Our results discovered that circ_Arf3 and Mbnl1 were lowly expressed in HG-treated MCs, while miR-452-5p expression was up-regulated. Moreover, circ_Arf3 was mainly located in the cytoplasm and had a ring-like stable structure. Functional assays demonstrated that overexpression of circ_Arf3 prevented cell proliferation and fibrous formation in HG-treated MCs. Circ_Arf3 could sponge miR-452-5p, and the effect of circ_Arf3 overexpression was reversed by enhanced expression of miR-452-5p. Mbnl1 was a direct target of miR-452-5p. Knockdown of Mbnl1 abolished the suppressive effects of miR-452-5p inhibitor on proliferation and fibrosis-related protein expression in HG-treated MCs. Moreover, circ_Arf3 regulated Mbnl1 through miR-452-5p.Conclusion: Overexpression of circ_Arf3 prevents cell proliferation and fibrous formation in HG-treated MCs by regulating the expression of Mbnl1 via miR-452-5p. [ABSTRACT FROM AUTHOR]

Bovine respiratory disease (BRD) is one of the major health issues in the cattle industry, resulting in significant financial crises globally. There is currently no good treatment, and cattle are made resistant to pneumonia through disease-resistant breeding. The serial blood samples from six Xinjiang brown (XJB) calves were collected for the RNA sequencing (RNA-seq). The obtained six samples were grouped into two groups, in each group as infected with BRD and healthy calves, respectively. In our study, the differential expression mRNAs were detected by using RNA-seq and constructed a protein–protein interaction (PPI) network related to the immunity in cattle. The key genes were identified by protein interaction network analysis, and the results from RNA-seq were verified using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). A total of 488 differentially expressed (DE) mRNAs were identified. Importantly, the enrichment analysis of these identified DEGs classified them as mainly enriched in the regulation and immune response processes. The 16 hub genes were found to be related to immune pathways categorized by PPIs analysis. Results revealed that many hub genes were related to the immune response to respiratory disease. These results will provide the basis for a better understanding of the molecular mechanism of bovine resistance to BRD. [ABSTRACT FROM AUTHOR]

Simple Summary: Exopalaemon carinicauda is a typical representative of crustaceans. However, the mechanism of ovary early development remains obscure. The purpose of this study is to identify and verify important genes related to ovary development by transcriptome sequencing and two-color fluorescent in situ hybridization, respectively. The result would provide clues for further study of ovary development in crustaceans. Eyestalk ablation is an effective method to promote ovarian development in crustaceans. Herein, we performed transcriptome sequencing of ovary and hepatopancreas tissues after eyestalk ablation in Exopalaemon carinicauda to identify genes related to ovarian development. Our analyses led to the identification of 97,383 unigenes and 190,757 transcripts, with an average N50 length of 1757 bp. In the ovary, four pathways related to oogenesis and three related to oocyte rapid growth were enriched. In the hepatopancreas, two vitellogenesis-associated transcripts were identified. Furthermore, short time-series expression miner (STEM) and gene ontology (GO) enrichment analyses revealed five terms related to gamete generation. In addition, two-color fluorescent in situ hybridization results suggested that dmrt1 might play a vital role in oogenesis during the early stage of ovarian development. Overall, our insights should support future studies focusing on investigating oogenesis and ovarian development in E. carinicauda. [ABSTRACT FROM AUTHOR]

This research attempted to screen potential signatures associated with KIRC progression and overall survival by weighted gene co‐expression network analysis (WGCNA) and Cox regression. The KIRC‐associated mRNA expression and clinical data were accessed from The Cancer Genome Atlas (TCGA) database. Differentially expressed genes (DEGs) were screened by differential analysis. A co‐expression network was constructed by "WGCNA". Based on WGCNA module, GO and KEGG analyses were performed. Protein–protein interaction (PPI) network was constructed. Prognostic signatures were screened by Lasso‐Cox regression. Prognostic model was evaluated by Receiver Operating Characteristic (ROC) and Kaplan‐Meier (K‐M) curves. Multivariate Cox and nomogram were introduced to examine whether risk score could be an independent marker. qRT‐PCR was introduced to determine expression of 9 hub genes in KIRC clinical tumor tissues and adjacent tissues, respectively. Genes in the green module were highly associated with clinical status, and green module genes were significantly enriched in mitotic nuclear division, cell cycle, and p53 signaling pathway. Twenty‐six candidates were subsequently screened out from the green module. Next, a 9‐gene prognostic model (DLGAP5, NUF2, TOP2A, RRM2, HJURP, PLK1, AURKB, KIF18A, CCNB2) was constructed. The predicting ability of the model was optimal. Some cancer‐related signaling pathways were differently activated between two risk score groups. Additionally, under‐expression of some signature genes (AURKB, CCNB2, PLK1, RRM2, TOP2A) was associated with better survival rate for KIRC patients. Meanwhile, all 9 hub genes were substantially overexpressed in KIRC patients. A KIRC prognostic signature was screened in this study, contributing valuable findings to KIRC biomarker development. [ABSTRACT FROM AUTHOR]

Objective: To explore the effects of CUMS combined with CRS on mouse hippocampal glial cells and synaptic plasticity-related proteins. Methods: Forty mice were randomly divided into normal group (n=20) and model group (n=20). The model group used CUMS combined with CRS to prepare a mouse model of depression for 7 weeks. The behavioral evaluation of the mice at 3 weeks and 7 weeks after modeling was performed by sugar water preference test, open field test and tail suspension test. After the experiment, the samples were collected, and the content of TNF-a in the hippocampus of mice was detected by enzyme-linked immunosorbent assay. Immunohistochemical method was used to detect the Iba-1 and GFAP MOD values of mouse hippocampal CA1 area, CA3 area and DG area. Western blot was used to detect the protein expression of Iba-1, GFAP, SYN1 and PSD-95 in the hippocampus. fluorescence quantitative PCR method was used to detect the expression of SYN1, PSD-95 mRNA in hippocampus. Results: At the 3rd week after modeling, the body weight, sugar water preference rate, total distance moved, number of standing uprights, and stay time in the central area of the mice in the model group were all lower than those in the normal group (P<0.05), and the tail suspension immobility time was longer than that in the normal group (P<0.01). After 7 weeks of modeling, the body weight, sugar water preference rate, total distance moved, number of erection times, central area residence time, and average movement speed of the mice in the model group were lower than those in the normal group (P< 0.05), the tail suspension immobility time was longer than that in the normal group (P<0.01). The contents of TNF-a in the hippocampus were higher than those in the normal group (P<0.05). The GFAP MOD value and the relative expression of GFAP protein in hippocampal CA1, CA3 and DG regions were significantly lower than those in the normal group (P<0.05). The Iba-1 MOD value and the relative expression of Iba-1 protein in hippocampal CA1, CA3 and DG regions were significantly higher than those in the normal group (P<0.05). The relative expression of SYN1 and PSD-95 protein and the relative expression of SYN1 and PSD-95 mRNA in the hippocampus were significantly lower than those in the normal group (P<0.05).Conclusion: After 3 weeks of CUMS and CRS modeling, the depression-like behavior of mice appeared, and the depression of mice was more obvious after 7 weeks of modeling. The depression mouse model made by CUMS combined with CRS method may be related to increased hippocampal inflammation, excessive activation of microglia, decreased number of astrocytes and decreased synaptic plasticity.objective: To explore the effects of CUMS combined with CRS on mouse hippocampal glial cells and synaptic plasticity-related proteins. [ABSTRACT FROM AUTHOR]

Simple Summary: Long non-coding RNAs have been demonstrated to play important roles in regulating tumor development and progression in breast cancer, which is tumor type dependent and cellular localization dependent. However, the regulatory mechanisms remain unclear. Herein, we found a dual function of long non-coding RNA CCAT2 in the luminal subtype of breast cancer, depending on its subcellular distribution. CCAT2 showed an overall downregulation in the tumor tissues from the luminal breast cancer patients. Cytoplasmic CCAT2 in the luminal subtype of breast cancer cell MCF-7 or T47D significantly suppressed cell proliferation and cancer cell stemness in vitro. It inhibited tumor growth in vivo, which was mediated with miR-221-p27 signaling. In contrast, nuclear overexpression of CCAT2 led to upregulation of OCT4-PG1 and the induction of cancer cell stemness. In summary, for the first time, the current study revealed a dual function of lncRNA CCAT2 as a tumor suppressor or oncogene depending upon its subcellular distribution. Breast cancer is the most common cancer in women around the world. Emerging evidence has indicated the important roles that non-coding RNAs play in regulating tumor development and progression in breast cancer. Herein, we found a dual function of long non-coding RNA (LncRNA) CCAT2 in the luminal subtype of breast cancer, depending on its subcellular distribution. CCAT2 showed an overall downregulation in the tumor tissues from luminal breast cancer patients. Transient overexpression of CCAT2 in the luminal subtype of breast cancer cell MCF-7 or T47D significantly suppressed cell proliferation in vitro and inhibited tumor growth in vivo. Gene expression analysis of cancer stem cell markers including OCT4, NANOG, h-TERT, SOX2 and KLF4; flow cytometry analysis of breast cancer stem cell population, and mammosphere formation assay demonstrated inhibition of cancer cell stemness with transient transfection of CCAT2 in which exogenous CCAT2 mainly distributed in the cytoplasm and regulated miR-221-p27 signaling via RNA sequence interaction. However, overexpression of CCAT2 in MCF-7 cells through pMX retroviral nuclear expression vector accumulated CCAT2 in the nucleus, leading to upregulation of OCT4-PG1, a pseudogene of stem gene OCT4, thereby promoting the cancer cell stemness. In conclusion, the current study, for the first time, revealed a dual function of lncRNA CCAT2 as a tumor suppressor or oncogene depending upon its subcellular distribution. It also demonstrated the regulatory mechanism of cytoplasmic CCAT2 in suppressing tumorigenesis in the luminal subtype of breast cancer. [ABSTRACT FROM AUTHOR]

Objective: The methylation of DNA promoter region mediates the low expression of many tumor suppressor genes and plays an essential part in cancer progression. We investigated methylation and expression of ZNF582 in clear cell renal cell carcinoma (ccRCC), and to study the function of ZNF582 in ccRCC cells. Methods: Methylation data and mRNA expression data of TCGA-KIRC were obtained from TCGA database to screen methylation-driven genes. Survival analysis and gene set enrichment analysis (GSEA) were done for the target gene. The methylation degree and mRNA level of ZNF582 in ccRCC cell line were detected by methylation-specific PCR (MSP) and qRT-PCR, respectively. Effects of overexpression of ZNF582 on ccRCC cells were assessed via CCK-8, flow cytometry, wound healing, Transwell, and cell adhesion assays. Results: Eighteen methylation-driven genes were identified via bioinformatics methods. Among them, ZNF582 was noticeably hypermethylated and lowly expressed in tumor tissue, and ZNF582 methylation and expression levels were pronouncedly associated with prognosis and clinical stage. MSP also displayed that the ZNF582 DNA promoter region was hypermethylated in ccRCC cells, and the mRNA expression of ZNF582 was dramatically elevated after demethylation. In vitro cell experiments disclosed that overexpression of ZNF582 markedly hindered cell proliferation, invasion, migration, and fostered cell apoptosis and adhesion of ccRCC. Conclusion: ZNF582 was hypermethylated in ccRCC, which mediated its low level. Overexpression of ZNF582 inhibited tumor cell proliferation, migration and invasion. This study generates novel ideas for ccRCC diagnosis and treatment. [ABSTRACT FROM AUTHOR]

Macrophages in atherosclerotic patients are notably plastic and heterogeneous. Single-cell RNA sequencing (Sc RNA-seq) can provide information about all the RNAs in individual cells, and it is used to identify cell subpopulations in atherosclerosis (AS) and reveal the heterogeneity of these cells. Recently, some findings from Sc RNA-seq experiments have suggested the existence of multiple macrophage subsets in atherosclerotic plaque lesions, and these subsets exhibit significant differences in their gene expression levels and functions. These cells affect various aspects of plaque lesion development, stabilization, and regression, as well as plaque rupture. This article aims to review the content and results of current studies that used RNA-seq to explore the different types of macrophages in AS and the related molecular mechanisms as well as to identify the potential roles of these macrophage types in the pathogenesis of atherosclerotic plaques. Also, this review listed some new therapeutic targets for delaying atherosclerotic lesion progression and treatment based on the experimental results. [ABSTRACT FROM AUTHOR]

Objective. The aims of this study were to screen the gene mutations that are able to predict the risk of cigarette smoking-related lung adenocarcinoma (LUAD) and to evaluate its prognostic significance. Methods. Clinical data and genetic information were retrieved from the TCGA database, and the patients with LUAD were divided into three groups including never smoking, light smoking, and heavy smoking according to cigarette smoking dose. Differentially mutated genes (DMGs) of each group were analyzed. At the same time, the function of DMGs in three smoking groups was evaluated by GO function and KEGG pathway analysis. The driver genes and protein variation effect of DMGs were performed to further screen key genes. The survival characteristics of the gene expression and mutation of those genes were analyzed and plotted to visualize by the Kaplan-Meier model. Result. The DMGs for different smoking doses were identified. The driver and deleterious mutation in the DMGs were screened and gene interaction network was constructed. The DMGs with driver mutations and deleterious mutations that were associated with the overall survival in the heavy smoking patients were considered as the candidate genes for novel markers of smoking-related LUAD. The final novel risk factor gene was identified as MYH7 and the high express of MYH7 in LUAD correlation with patients' gender, lymph node metastasis, T stage, and clinical stage. Conclusions. In summary, it can be concluded that MYH7 is a novel biomarker for heavy smoking-related LUAD and it is significantly correlated with the prognosis of lung cancer and is related to the clinical characteristics of lung cancer. [ABSTRACT FROM AUTHOR]

Triptolide (TP), a major active component of the herb Tripterygium wilfordii Hook F (TwHF), has been shown to exert therapeutic potential against rheumatoid arthritis (RA). However, its molecular mechanism of action has not been fully elucidated. This study aimed to analyze the potential target of TP based on the discovery of differentially methylated and expressed genes (DMEGs) in RA using methylated RNA immunoprecipitation sequencing (MeRIP-seq) and RNA sequencing (RNA-seq). Five RA samples and ten control samples were obtained from China-Japan Friendship Hospital. The various levels of m6A methylation and genes expressed in the RA and control groups were compared by MeRIP-seq and RNA-seq. Bioinformatics explorations were also performed to explore the enriched biological roles and paths of the differentially expressed m6A methylation and genes. Molecular networks between TP target proteins and DMEGs were performed using Ingenuity Pathway Analysis (IPA) software. Potential target of TP was determined with Gene Expression Omnibus (GEO) database mining, molecular docking, and in vitro experiment validation. In total, 583 dysregulated m6A peaks, of which 295 were greatly upregulated and 288 were greatly downregulated, were identified. Similarly, 1,570 differentially expressed genes were identified by RNA-seq, including 539 upregulated and 1,031 downregulated genes. According to the deeper joint exploration, the m6A methylation and mRNA expression degrees of 35 genes varied greatly. Molecular networks between TP target proteins and DMEGs were constructed, and the results revealed that tubulin beta-2A chain (TUBB2A), insulin-like growth factor 2 mRNA-binding protein 3 (IGF2BP3), cytoplasmic dynein 1 intermediate chain 1 (DYNC1I1), and FOS-like 1 (FOSL1) were the most relevant genes that correlated with the target proteins of TP. The results of the GEO database showed that the gene expression of IGF2BP3 was increased in RA synovial tissue and consistent with the trend of our sequencing results of RA PBMCs. Molecular docking and in vitro experiment suggested that TP and IGF2BP3 had a high binding affinity and TP could decrease the mRNA expression of IGF2BP3 in PBMCs and MH7A.This research established a transcriptional map of m6A in RA PBMCs and displayed the hidden association between RNA methylation alterations and associated genes in RA. IGF2BP3 might be a potential therapeutic target of TP during RA treatment. [ABSTRACT FROM AUTHOR]

Background: Tyrophagus putresecentiae is an important mite species in rural and urban environments, causing sensitization and allergic disease. While evidence suggests that microRNAs (miRNAs) may regulate the expression of allergen‐encoding genes, no study has directly investigated this possibility. Here, this gap was addressed by profiling miRNAs and elucidating their target allergen messenger RNAs (mRNAs) in this mite species. Methods: Small RNA and transcriptome libraries were constructed for eggs, larvae, nymphs, and adults. After deep miRNA and whole‐transcriptome sequencing were performed, the miRNA and allergen‐encoding mRNA regulatory networks were explored. Results: A total of 540 miRNAs were identified, including 155 with expression levels differing significantly across the four mite developmental stages (p <.01), 59 of which were novel. The mRNA expression for allergens was higher for Tyr p 1 in adults than in other developmental stages; Tyr p 2–5, 7, 10, 13, 33, and 34 in immature stages; and Tyr p 28, 35, and 36 in eggs and adults. A combined miRNA and transcriptome bioinformatics analysis showed that allergen Tyr p 3 was regulated by miRNA PC‐5p‐5698441_1, Tyr p 4 was regulated by PC‐5p‐7050653_1, and Tyr p 34 was regulated by PC‐5p‐5534223_1 and PC‐5p‐5698441_1. These three allergen mRNA and three miRNAs were identified using qRT‐PCR, and their regulatory roles were confirmed by double‐fluorescent reporter gene system and site‐directed mutagenesis technology. Conclusions: For the first time, allergen mRNA expression and miRNAs were profiled throughout the life cycle for an allergen‐producing mite, and the results showed that miRNAs bind to target allergen mRNAs to regulate their expression. [ABSTRACT FROM AUTHOR]

Background. Vascular endothelial cell injury is not only the initiating factor of cardiovascular and cerebrovascular diseases but also the essence of blood stasis. Levistilide A (LA), a natural component isolated from the traditional Chinese herb, Ligusticum chuanxiong Hort, has traditional effects on improving blood circulation and removing stasis. In this study, the effects and potential mechanisms of LA in the rat model of blood stasis and the mechanism in endothelial cell injury have been explored. Materials and Methods. In this experiment, the effects of LA on the model of acute blood stasis in rats were explored. The blood samples were collected for the measurement of coagulation and hemorheological indices, and the carotid arteries were also excised from rats for hematoxylin-eosin (HE) staining and immunohistochemistry (IHC). In addition, the improvement effects of LA on the H2O2-induced human umbilical vein endothelial cell (HUVEC) injury model were evaluated. And the cell viability detection was conducted by the CCK8 assay, and the pathway-related protein expressions were detected by western blotting. Results. In vivo, compared with the model group, the treatment of LA (10 mg/kg) could reduce the FIB (fibrinogen) content (P < 0.01), increase the INR (international normalized ratio) and PT (prothrombin time) (P < 0.01), and reduce the plasma viscosity (P < 0.05) and whole blood viscosities of low, medium, and high shear rates in the blood of blood stasis model rats (P < 0.01). In vitro, the cell viability in the LA-pretreated group was higher than that of the model group (P < 0.05). The expression levels of PI3K, AKT, and eNOs in the LA-pretreated group were increased (P < 0.01) as compared to the model group. Conclusion. These findings demonstrated that LA has the ability to improve blood hypercoagulation and blood viscosity, and enhance the viability of cells. It is more likely that it exerts a protective effect on the endothelial cell through the PI3K-AKT-eNOs pathway. These results indicate LA will be a potential candidate to cure blood stasis with endothelial cell injury. [ABSTRACT FROM AUTHOR]

Objective. To identify significant pathways and genes in intervertebral disc degeneration (IDD) based on bioinformatics analysis. Design. The GEO database was used to download the GSE124272 dataset. Differentially expressed genes (DEGs) were analyzed using Limma package in R language. Then, gene ontologies (GO), Kyoto encyclopedia of genes and genomes (KEGG), and protein-protein interaction (PPI) networks were used to further identify hub genes. The mRNA expression levels of top six hub genes were verified. Results. We found 563 DEGs, of which 214 were upregulated and 349 were downregulated. The top 5 GO terms and pathways were shown including immune response, cell cycle, and p53 pathway. Based on the PPI analysis, we verified the mRNA expression levels of 6 hub genes. The mRNA levels of CHEK1, CDCA2, SKA3, and KIF20A were upregulated in degenerative NP tissue than in healthy NP tissue. However, the mRNA level of BUB1 and SPC25 was downregulated. Conclusions. This study may provide new biomarkers for the IDD and treatments to repair IDD related to CHEK1, CDCA2, SKA3, BUB1, KIF20A, and SPC25. [ABSTRACT FROM AUTHOR]

Objective. To investigate the role of curcumin in regulating pathogenesis of pulmonary arterial smooth muscle cells (PASMCs) derived from pulmonary arterial hypertension (PAH) model. Methods. Male Sprague Dawley rats were injected with monocrotaline (MCT) to establish the PAH experimental model. The rats were divided into control group, MCT group, and curcumin group. At the end of the study, hemodynamic data were measured to determine pulmonary hypertension. Proliferation ability of PASMCs, a remodeling indicator of pulmonary artery and right ventricle, was detected. In addition, the morphology and function of mitochondria, antiglycolysis and antiproliferation pathways, and genes were also analyzed. Results. Curcumin may function by reversing MCT-mediated pulmonary vascular remodeling in rats. Curcumin effectively improved pulmonary vascular remodeling, promoted PASMC apoptosis, and protected mitochondrial function. In addition, curcumin treatment suppressed the PI3K/AKT pathway in PASMCs and regulated the expression of antiproliferative genes. Conclusion. Curcumin can improve energy metabolism and reverse the process of PAHS. However, there were side effects of curcumin in MCT-induced rats, suggesting that the dosage should be treated with caution and its toxicological mechanism should be further studied and evaluated. [ABSTRACT FROM AUTHOR]

Background: Impaired hepatic fatty acid metabolism and persistent mitochondrial dysfunction are phenomena commonly associated with liver failure. Decreased serum levels of L-carnitine, a amino acid derivative involved in fatty-acid and energy metabolism, have been reported in severe burn patients. The current study aimed to evaluate the effects of L-carnitine supplementation on mitochondrial damage and other hepatocyte injuries following severe burns and the related mechanisms. Methods: Serum carnitine and other indicators of hepatocytic injury, including AST, ALT, LDH, TG, and OCT, were analyzed in severe burn patients and healthy controls. A burn model was established on the back skin of rats; thereafter, carnitine was administered, and serum levels of the above indicators were evaluated along with Oil Red O and TUNEL staining, transmission electron microscopy, and assessment of mitochondrial membrane potential and carnitine palmitoyltransferase 1 (CPT1) activity and expression levels in the liver. HepG2 cells pretreated with the CPT1 inhibitor etomoxir were treated with or without carnitine for 24 h. Next, the above indicators were examined, and apoptotic cells were analyzed via flow cytometry. High-throughput sequencing of rat liver tissues identified several differentially expressed genes (Fabp4, Acacb, Acsm5, and Pnpla3) were confirmed using RT-qPCR. Results: Substantially decreased serum levels of carnitine and increased levels of AST, ALT, LDH, and OCT were detected in severe burn patients and the burn model rats. Accumulation of TG, evident mitochondrial shrinkage, altered mitochondrial membrane potential, decreased ketogenesis, and reduced CPT1 activity were detected in the liver tissue of the burned rats. Carnitine administration recovered CPT1 activity and improved all indicators related to cellular and fatty acid metabolism and mitochondrial injury. Inhibition of CPT1 activity with etomoxir induced hepatocyte injuries similar to those in burn patients and burned rats; carnitine supplementation restored CPT1 activity and ameliorated these injuries. The expression levels of the differentially expressed genes Fabp4, Acacb, Acsm5, and Pnpla3 in the liver tissue from burned rats and etomoxir-treated hepatocytes were also restored by treatment with exogenous carnitine. Conclusion: Exogenous carnitine exerts protective effects against severe burn-induced cellular, fatty-acid metabolism, and mitochondrial dysfunction of hepatocytes by restoring CPT1 activity. [ABSTRACT FROM AUTHOR]

HMGN5 regulates biological function and molecular transcription via combining with a nucleosome. There has been growing evidence that aberrant expression of HMGN5 is associated with malignant neoplasm development and progression. In the present study, we found that the expression of HMGN5 is significantly higher in high-grade glioblastoma tissues than in low-grade samples. To clarify the function of HMGN5 in glioblastoma, we knocked down HMGN5 in U87 and U251 glioblastoma cells via siRNA. The results demonstrated that HMGN5 was involved in the regulation of proliferation and apoptosis, migration, and invasion of glioblastoma cells. These outcomes also indicated that silencing HMGN5 possibly suppressed the expression of p-AKT and p-ERK1/2. Taken together, our research reveals that HMGN5 might be an efficient target for glioblastoma-targeted therapy. [ABSTRACT FROM AUTHOR]

The Fuling (Poria cocos)-Guizhi (Cinnamomi ramulus) herb pair (FGHP) is a commonly used traditional Chinese herbal formula with coronary heart disease (CHD) treatment potential. However, the mechanism of FGHP in the treatment of CHD was still unclear. In this study, the action targets and underlying mechanism of FGHP against CHD were successfully achieved by combined network pharmacology prediction with experimental verification. 76 common targets were screened out by overlapping the chemical-protein data of FGHP and CHD-related targets. Then, two key targets were further selected for verification by using western blot analysis after analyzing PPI, GO function, and KEGG pathway. Results indicated FGHP could alleviate CHD syndromes and regulate inflammatory responses in acute myocardial ischemia rats, and the reduction of expression of TNF-α and IL-6 in myocardial tissue would be one of its possible underlying mechanisms. Our work demonstrated that network pharmacology combined with experimental verification provides a credible method to elucidate the pharmacological mechanism of FGHP against CHD. [ABSTRACT FROM AUTHOR]

Background. Dehydrocostus lactone (DL), one of the main active constituents in Aucklandia lappa Decne. (Muxiang), reported to have anti-inflammatory, antiulcer, and immunomodulatory properties. However, the effect of DL on ulcerative colitis (UC) has not been reported. To analyze the anti-inflammatory potential role of DL in UC, we provide a mechanism for the pharmacological action of DL. Methods. The experimental model of UC was induced by using oral administration of 2% dextran sulfate sodium (DSS) with drinking water in BALB/c mice. Mesalazine (Mes, 0.52 g/kg/d), DL-high doses (DL-H, 20 mg/kg/d), DL-middle doses (DL-M, 15 mg/kg/d), DL-low doses (DL-L, 10 mg/kg/d) were gavaged once a day from day 4 to day 17. Disease activity index (DAI) was calculated daily. On day 18, mice were rapidly dissected and the colorectal tissues were used to detect the levels of UC-related inflammatory cytokines (TNF-α, IL-1β, MCP-1, MPO, SOD, IL-6, IL-17, and IL-23), IL-6/STAT3 inflammatory signaling pathway (iNOS, COX2, IL-6, GP130, L-17, and IL-23), and colorectal mucosal barrier-related regulatory factors (MUC2, XBP1s, and α-defensins) by ELISA or qRT-PCR. Results. DL reduced the colorectal inflammation histological assessment, decreased UC-related inflammatory cytokines (TNF-α, IL-1β, MCP-1, MPO, SOD, IL-6, IL-17, and IL-23), downregulated IL-6/STAT3 inflammatory signaling pathway (iNOS, COX2, IL-6, GP130, L-17, and IL-23), repaired the key colorectal mucosal barrier protein-MUC2, and inhibited the downstream pathway (XBP1s and α-defensin). Conclusions. DL possessed the potential of anti-inflammatory effect to treated colitis. The protective mechanism of DL may involve in reducing inflammation and improving colorectal barrier function via downregulating the IL-6/STAT3 signaling. [ABSTRACT FROM AUTHOR]

• ush2a gene knockout caused abnormal accumulation of vesicles in zebrafish photoreceptors. • ush2a gene defects affected the expression and localization of Rab8 protein in the retina of zebrafish, and USH2A may interact with Rab8. • USH2A mutations might cause vesicle transport defects by causing functional abnormalities of Rab8 and other vesicle transport-related factors, which could not maintain the survival of photoreceptors, and ultimately lead to RP. USH2A (Usher syndrome type 2A) gene mutations are the predominant cause of Usher syndrome type 2, characterized by sensorineural hearing loss and retinitis pigmentosa (RP), and also significant contributors to non-syndromic RP. To date, there is a lack of definitive therapeutic interventions to mitigate the associated disorders caused by USH2A mutations, and the precise pathogenic mechanisms underlying their onset remain unclear. In the present study, we utilized the ush2a knockout zebrafish model to investigate the pathological mechanisms of RP. In late-stage ush2a-/- zebrafish, the outer segments of rods displayed shortened length and decreased number. Anomalous vesicle accumulation was observed at the junction between the inner and outer segments, accompanied by reduced expression and structural damage of actin filaments in the photoreceptor cells. Furthermore, we discovered that Rab8 expression was downregulated and exhibited aberrant localization in ush2a-/- zebrafish. Additionally, we identified an interaction between USH2A and Rab8. Therefore, the knockout of ush2a may potentially affect vesicle transport through the regulation of Rab8, providing a novel target for maintaining the survival of photoreceptor cells. These findings also contribute to our understanding of the potential molecular pathogenesis underlying RP caused by USH2A gene mutations. [ABSTRACT FROM AUTHOR]

The purpose of this study was to set up a beagle dog model, for radiation-induced lung injury, that would be able to supply fresh lung tissues in the different injury phases for research into oxidative stress levels and mitochondrial gene expression. Blood serum and tissues were collected via CT-guided core needle biopsies from dogs in the various phases of the radiation response over a 40-week period. Levels of reactive oxygen species (ROS) and manganese superoxide dismutase 2 (MnSOD) protein expression in radiation-induced lung injury were determined by in situ immunocytochemistry; malondialdehyde (MDA) content and reductase activity in the peripheral blood were also tested; in addition, the copy number of the mitochondrial DNA and the level of function of the respiratory chain in the lung tissues were assessed. ROS showed dynamic changes and peaked at 4 weeks; MnSOD was mainly expressed in the Type II alveolar epithelium at 8 weeks; the MDA content and reductase activity in the peripheral blood presented no changes; the copy numbers of most mitochondrial genes peaked at 8 weeks, similarly to the level of function of the corresponding respiratory chain complexes; the level of function of the respiratory chain complex III did not peak until 24 weeks, similarly to the level of function of the corresponding gene Cytb. Radiation-induced lung injury was found to be a dynamically changing process, mainly related to interactions between local ROS, and it was not associated with the levels of oxidative stress in the peripheral blood. Mitochondrial genes and their corresponding respiratory chain complexes were found to be involved in the overall process. [ABSTRACT FROM AUTHOR]

The Penaeus vannamei is an important shrimp species with enormous commercial and ecological values. In production process, the air exposure resistance is vital for live transportation without water. We tested the air exposure resistant ability of P. vannamei , and carried out gill histological observation and gene expression analysis. The physiology and molecular response to the air exposure stress of P. vannamei was revealed. We found that body weight could affect the air exposure tolerance. Air exposure caused epithelial cell of gill filament shrinking and tissue fluid exudation within half of hour, and triggered oxidative stress response. After retrieved to water, epithelial cell shrinking and tissue fluid exudation recovered gradually, but oxidative and antioxidant response is still going on. Organisms reduced oxidative stress by regulating levels of antioxidants and antioxidant enzymes that remove reactive oxygen species (ROS) and RNA and DNA processing to repair tissue damage, and expression of apoptosis associated-genes altered. Furthermore, the survive shrimps could live steadily more than 5 days, and their gill filament recovered to normal state, proving that the damage of air exposure is reversible. These findings could be considered in the waterless live transportation of P. vannamei. • Air exposure caused epithelial cell of the gill shrinking within half of hour. • After retrieve to water, the oxidative and antioxidant response is still going on. • Expression of apoptosis associated-genes altered. • After re-water for several days, the gill filament returned to normal state. • The damage of air exposure is reversible. [ABSTRACT FROM AUTHOR]

Lidocaine has been recognized to induce neurotoxicity. However, the molecular mechanism underlying this effect, especially the critical molecules in cells that mediated the lidocaine-induced apoptosis were unclear. In the present study, PC12 cells were administrated with lidocaine for 48 h. Using MTT assay and flow cytometry, we found lidocaine significantly decreased the cell proliferation and S phases in PC12 cells with treatment concentrations, and significantly enhanced cell apoptosis with treatment concentrations. Two-dimensional gel electrophoresis (2-DE) analysis and LC–MS/MS were used to identification of protein biomarkers. Six proteins were identified. Among them, three were up-expressed including ANXA6, GNB2 and STMN1, other three were down-expressed including ubiquitin-linke protein 7 (UBL7), DDAH2 and BLVRB. Using qRT-PCR, we confirmed that lidocaine up-regulated the mRNA expression of STMN1, GNB2, ANXA6 and DDAH2, and found that the GNB2 had the largest change (about increased by 6.4 folds). The up-regulation of GNB2 by lidocaine was also validated by western blot. After transfected with 100 μM GNB2-Rat-453 siRNA, the expression of GNB2 in PC12 cells was almost completely inhibited; and the cell proliferation and cells in S phases were significantly enhanced, cell apoptosis including both early apoptosis and later apoptosis were significantly reduced in the presence of 0.5 mM lidocaine for 48 h. Therefore, neuronal apoptosis was induced by lidocaine and this effect was mediated by GNB2. Further research is needed to assess the clinical relevance and exact mechanism of neuronal apoptosis caused by lidocaine. [ABSTRACT FROM AUTHOR]

Genomic imprinting is a phenomenon characterized by parent-of-origin-specific gene expression. While widely documented in viviparous mammals and plants, imprinting in oviparous birds remains controversial. Because genomic imprinting is temporal- and tissue-specific, we investigated this phenomenon only in the brain tissues of 1-day-old chickens (Gallus gallus). We used next-generation sequencing technology to compare four transcriptomes pooled from 11 chickens, generated from reciprocally crossed families, to the DNA sequences of their parents. Candidate imprinted genes were then selected from these sequence alignments and subjected to verification experiments that excluded all but one SNP. Subsequent experiments performed with two new sets of reciprocally crossed families resulted in the exclusion of that candidate SNP as well. Attempts to find evidence of genomic imprinting from long non-coding RNAs yielded negative results. We therefore conclude that genomic imprinting is absent in the brains of 1-day-old chickens. However, due to the temporal and tissue specificity of imprinting, our results cannot be extended to all growth stages and tissue types. [ABSTRACT FROM AUTHOR]

Precartilaginous stem cells (PSCs) are adult stem cells which could initiate chondrocytes and bone growth. In the current study, we purified PSCs from the neonate mice' perichondrial mesenchyme through immunomagnetic beads with the fibroblast growth factor receptor-3 (FGFR-3) antibody. Mouse PSCs were seeded and cultured, and their phenotype was confirmed by FGFR-3 over-expression. Transforming growth factor-β (TGF-β) was added to induce PSCs differentiation. TGF-β increased mRNA expression of chondrogenesis-related genes (collagen type II, Sox 9, and aggrecan) in the cultured PSCs, which was abolished by TGF-β receptor II (TGFRII) lentiviral shRNA depletion. TGF-β induced AKT activation in mouse PSCs, while the PI3K/AKT inhibitor (LY294002) and the AKT specific inhibitors (perifosine and MK-2206) largely suppressed TGF-β-induced collagen II, Sox 9, and aggrecan mRNA expression. Meanwhile, the mTOR complex 1 (mTORC1) blocker RAD001 or the mTORC1/2 dual inhibitor AZD-2014 also alleviated TGF-β-induced chondrogenesis-associated genes expression. Further, lentiviral shRNA depletion of SIN1 (a mTORC2 component) or mTOR inhibited TGF-β's effect in the mouse PSCs. In conclusion, our evidence suggests that TGF-β induces the expression of chondrogenesis-related genes through TGFRII-AKT-mTOR signaling in cultured mouse PSCs. [ABSTRACT FROM AUTHOR]

Durum wheat is the second-most widely grown wheat species, and is primarily used in the production of pasta and couscous. The grain utilization of durum wheat is partly related to its very hard kernel texture because of the lack of the D genome and consequentially the Puroindoline genes. Our previous study reported the transformation of durum wheat with the Puroindoline a ( Pina) gene. Here, we characterized the transgenic durum wheat lines expressing the Pina gene, and studied the effects of PINA on grain texture and other kernel characteristics. SDS-PAGE and Western blotting results demonstrated that starch-bound PINA levels of Pina-overexpressing lines were lower than that of Pina-positive control, common wheat cv. Chinese Spring, suggesting a weak association of PINA protein with starch granules in the absence of Pinb. Grain hardness analysis and flour milling tests indicated that the overexpression of PINA resulted in decreased grain hardness and increased flour yield in transgenic durum wheat lines. The agronomic performance of the transgenic and control lines was also examined and it was found that no significant differences in measured traits were observed between Pina-overexpressing and control lines in the 2-year field trials. Since grain hardness strongly affects milling and end-use qualities, the development of medium-hard-textured durum wheat lines is not only of significance for our knowledge of grain hardness and Puroindolines, but also has practical implications for plant breeders and food technologists for the expansion of utilization of durum wheat. [ABSTRACT FROM AUTHOR]

Cell survival signaling is important for the malignant phenotypes of cancer cells. Although the role of receptor-interacting protein 1 (RIP1) in cell survival signaling is well documented, whether RIP1 is directly involved in cancer development has never been studied. In this report, we found that RIP1 expression is substantially increased in human non-small cell lung cancer and mouse lung tumor tissues. RIP1 expression was remarkably increased in cigarette smoke-exposed mouse lung. In human bronchial epithelial cells (HBECs), RIP1 was significantly induced by cigarette smoke extract or benzo[a]pyrene diol epoxide (BPDE), the active form of the tobacco-specific carcinogen benzo(a)pyrene. In RIP1 knockdown HBECs, BPDE-induced cytotoxicity was significantly increased, which was associated with induction of cellular reactive oxygen species (ROS) and activation of mitogen-activated protein kinases (MAPKs), including c-jun N-terminal kinase (JNK), extracellular signal-regulated kinase (ERK) and p38. Scavenging ROS suppressed BPDE-induced MAPK activation and inhibiting ROS or MAPKs substantially blocked BPDE-induced cytotoxicity, suggesting ROS-mediated MAPK activation is involved in BPDE-induced cell death. The ROS-reducing enzyme catalase is destabilized in an ERK- and JNK-dependent manner in RIP1 knockdown HBECs and application of catalase effectively blocked BPDE-induced ROS accumulation and cytotoxicity. Importantly, BPDE-induced transformation of HBECs was significantly reduced when RIP1 expression was suppressed. Altogether, these results strongly suggest an oncogenic role for RIP1, which promotes malignant transformation through protecting DNA-damaged cells against carcinogen-induced cytotoxicity associated with excessive ROS production. [ABSTRACT FROM PUBLISHER]

Improper adjustments of autophagy and silent information regulator 1 (Sirt-1) expression were reported to be closely associated with metabolic disorders. In this study, we examined the roles of Sirt-1 and autophagy in streptozotocin-induced diabetes mellitus, assessed the relationship between autophagy and Sirt-1, and investigated the protective mechanism of silibinin. Diabetes was induced in 6-week-old mice by intravenous injection of streptozotocin (150 mg/kg/day, for 2 weeks). In the treatment groups, silibinin (50 mg/kg/day, intramuscular injection, for 8 weeks) or inhibitors (50 mg/kg/day, subcutaneous injection, for 8 weeks) were given. Diabetic control animals received vehicle for the same time. Compared with diabetic controls, silibinin or autophagy inhibitor, 3-methyladenine, treated mice showed decreased levels of glycosylated hemoglobin A1C (P < 0.01), serum triglyceride (P < 0.01), cholesterol (P < 0.01), blood glucose (P < 0.05), autophagy (P < 0.05), and apoptosis ratio (P < 0.05) of pancreatic β-cells. Systemic administration of silibinin reversed streptozotocin-induced downregulation of Sirt-1 expression. Sirt-1 may play a role in regulating the physiological level of autophagy and is associated with loss of pancreatic β-cells and metabolic biochemical disorders. Through promoting Sirt-1 expression and recovering autophagy physiologically, silibinin may reverse hyperglycemia and repair damaged pancreatic β-cells. [ABSTRACT FROM PUBLISHER]

Aim:To investigate the anticancer effect of crocetin, a major ingredient in saffron, and its underlying mechanisms.Methods:Cervical cancer cell line HeLa, non-small cell lung cancer cell line A549 and ovarian cancer cell line SKOV3 were treated with crocetin alone or in combination with vincristine. Cell proliferation was examined using MTT assay. Cell cycle distribution and sub-G1 fraction were analyzed using flow cytometric analysis after propidium iodide staining. Apoptosis was detected using the Annexin V-FITC Apoptosis Detection Kit with flow cytometry. Cell death was measured based on the release of lactate dehydrogenase (LDH). The expression levels of p53 and p21WAF1/Cip1 as well as caspase activation were examined using Western blot analysis.Results:Treatment of the 3 types of cancer cells with crocetin (60-240 μmol/L) for 48 h significantly inhibited their proliferation in a concentration-dependent manner. Crocetin (240 μmol/L) significantly induced cell cycle arrest through p53-dependent and -independent mechanisms accompanied with p21WAF1/Cip1 induction. Crocetin (120-240 μmol/L) caused cytotoxicity in the 3 types of cancer cells by enhancing apoptosis in a time-dependent manner. In the 3 types of cancer cells, crocetin (60 μmol/L) significantly enhanced the cytotoxicity induced by vincristine (1 μmol/L). Furthermore, this synergistic effect was also detected in the vincristine-resistant breast cancer cell line MCF-7/VCR.Conclusion:Ccrocetin is a potential anticancer agent, which may be used as a chemotherapeutic drug or as a chemosensitizer for vincristine. [ABSTRACT FROM AUTHOR]

Background: Gene expression variation is a key underlying factor influencing phenotypic variation, and can occur via cis- or trans-regulation. To understand the role of cis- and trans-regulatory variation on population divergence in chicken, we developed reciprocal crosses of two chicken breeds, White Leghorn and Cornish Game, which exhibit major differences in body size and reproductive traits, and used them to determine the degree of cis versus trans variation in the brain, liver, and muscle tissue of male and female 1-day-old specimens. Results: We provided an overview of how transcriptomes are regulated in hybrid progenies of two contrasting breeds based on allele specific expression analysis. Compared with cis-regulatory divergence, trans-acting genes were more extensive in the chicken genome. In addition, considerable compensatory cis- and trans-regulatory changes exist in the chicken genome. Most importantly, stronger purifying selection was observed on genes regulated by trans-variations than in genes regulated by the cis elements. Conclusions: We present a pipeline to explore allele-specific expression in hybrid progenies of inbred lines without a specific reference genome. Our research is the first study to describe the regulatory divergence between two contrasting breeds. The results suggest that artificial selection associated with domestication in chicken could have acted more on trans-regulatory divergence than on cis-regulatory divergence. [ABSTRACT FROM AUTHOR]

A stepwise partially overlapping primer-based PCR (SWPOP-PCR) method for isolating flanking unknown DNA regions was developed, which comprises three rounds of nested PCRs sequentially driven by SWPOP primer-nested specific primer pairs. SWPOP primer set is characterized by a partial overlap of 10 bp with 3′-part of the latter primer is identical to 5′-part of the former one, which makes the SWPOP primer in use anneal to SWPOP site of the prior PCR product only at relatively low temperature. For each PCR, target single-stranded DNA primed by the SWPOP primer in the exclusive one low-stringency cycle is converted into double-stranded form in the following high-stringency cycle due to the presence of a perfect annealing site for the specific primer. This double-stranded DNA bounded by the specific primer and the SWPOP primer is exponentially amplified in the remaining high-stringency cycles. Non-target single-stranded DNA, however, cannot be amplified given the lack of perfect complementary sequences for any primers. Therefore, the partial overlap of a SWPOP primer set preferentially synthesizes target products but inhibits nonspecific amplification. We successfully exploited SWPOP-PCR to obtain the DNA sequences flanking glutamate decarboxylase gene (gadA) locus in Lactobacillus brevis NCL912 and hygromycin gene (hyg) integrated in rice. [ABSTRACT FROM AUTHOR]

Accumulating evidence has revealed that aberrant abundance of 5-hydroxymethylcytosine (5hmC) is critically involved in tumorigenesis. The aim of the present study was to investigate the level of 5hmC in primary oral squamous cell carcinoma (OSCC) and determine its clinical significance as well as prognostic value in predicting patients’ outcomes. The expression levels of 5hmC in 95 human OSCC samples and 24 normal oral mucosa were evaluated by immunohistochemical staining. Moreover, the associations between the expression status of 5hmC and several clinicopathological parameters as well as patients’ survival were further statistically assessed. Our immunohistochemical results revealed that 5hmC was significantly downregulated in a significant fraction of OSCC as compared their normal counterparts. However, elevated 5hmC level was found to be significantly associated with pathological grade and cervical node metastasis with P -values of 0.0239 and 0.0041, respectively. Results from Kaplan-Meier cumulative survival analyses indicated that high expression of 5hmC in OSCC was significantly associated with decreased overall survival, disease-free and disease-specific survival as compared to those with low 5hmC (Log-rank, P = 0.0210, 0.0313, 0.0415, respectively). Furthermore, the univariate and multivariate survival analyses further identified the expression status of 5hmC as an independent prognostic factor affecting patients’ survival. Taken together, our results reveal a significant decrease of 5hmC level in a large subset of OSCC. However, high level of 5hmC associates with tumor aggressive features and unfavorable prognosis in a fraction of OSCC patients. [ABSTRACT FROM AUTHOR]

A hydroponic experiment was conducted to verify the effects of inoculation with endophytic bacteria Sphingomonas SaMR12 on root growth, cadmium (Cd) uptake, reactive oxygen species (ROS), antioxidases, glutathione (GSH) and the related gene expression of Sedum alfredii Hance under different levels of Cd such as 0, 10, 25, 100 and 400 μM. The results showed that inoculation of SaMR12 improved Cd accumulation and upregulated glutathione synthase ( GS ) expression, but slightly reduced malondialdehyde (MDA) concentration and alleviated Cd-induced damage in roots. However it didn't alter the activities of antioxidant enzymes. When Cd concentration exceeded 25 μM, SaMR12 increased the concentration of GSH and the expression level of GSH1 . At high Cd treatment levels (100 and 400 μM), SaMR12 significantly reduced H 2 O 2 concentration and enhanced expression level of 1-Cys peroxiredoxin PER1 and ATPS genes. These results indicate that although SaMR12 has no significant effects on antioxidases activities, it reduces H 2 O 2 concentration by enhancing GSH concentration and relevant genes expression, and subsequently improves Cd tolerance and accumulation. [ABSTRACT FROM AUTHOR]

In this research, we examined which genes are involved in N-linked sialylation in Chinese Hamster Ovary (CHO) cells using siRNA knockdown approaches. Three genes from the sialyltransferase family (ST3GAL3, ST3GAL4 and ST3GAL6) were chosen as knockdown targets with siRNA applied to reduce their expression. Single, double and triple gene knockdowns were investigated, and the reduction levels of sialylation on the total cell lysate were monitored by enzyme-linked lectin absorption assays (ELLA) and sialic acid quantification with high performance liquid chromatography (HPLC). All transfection groups showed effective reduction in 2,3-linked sialylation whereas the trend of reduction levels of triple siRNA transfection outweighed both the dual siRNA groups and single siRNA transfection groups. Next, this transfection approach was applied to CHO cells producing erythropoietin (EPO). Quantification of EPO sialylation showed similar result to total cell lysate except that the ST3GAL4 siRNA transfection exhibited the largest reduction according to the HPLC analysis as compared with other single siRNA transfections. Finally, the N-glycan released from the EPO transfected with ST3GAL4 siRNA showed a prominent reduction in sialyation level among the single siRNA transfections. From these experiments, we concluded that each of these three genes were involved in N-linked sialylation and ST3GAL4 may play the critical role in glycoprotein sialylation of recombinant proteins such as EPO. [ABSTRACT FROM AUTHOR]

Gonadal soma-derived factor (Gsdf) is a new member of the transforming growth factor beta superfamily. As a teleost- and gonad-specific growth factor, several studies indicate that Gsdf plays an important role in early germ cell development. In the present study, for the first time, a 1700-bp long gsdf gene was cloned from a protandrous species, Acanthopagrus latus . We further analyzed the cellular localization and the expression patterns of gsdf in respective testicular and ovarian zones during the first maleness reproductive phase. The results showed that gsdf transcripts were highly expressed in the ovotestis during sex differentiation, and the somatic cells of the testicular zone expressed many more gsdf transcripts than those of the ovarian zone. At the onset of puberty, the gsdf expression levels decreased gradually during spermatogenesis. Conversely, the ovarian zone exhibited a stable increase pattern which was similar to the plasma 17β-estradiol (E 2 ) levels. These results suggested that Gsdf may participates in early germ cell development, e.g. proliferation and differentiation of spermatogonia and oogonia in A. latus . [ABSTRACT FROM AUTHOR]

Abstract Background NK cells are presented in tumor microenvironments and acts as an essential defense line against multiple malignancies. Recently, miRNAs are reported to involve in the development of natural killer (NK) cells via negatively regulating gene expression. Here, we aim to explore the function and mechanism underlying how miR-20a modulated the killing effect of NK cells to cervical cancer cells. Methods Abundances of miR-20a and runt-related transcription factor 1 (RUNX1) in NK cells from cervical cancer patients and healthy donors were detected by qRT-PCR and western blot. The releases of IFN-γ and TNF-α were determined by ELISA. The cytotoxicity of NK cells against cervical cancer cells was measured by CytoTox 96 non-radioactive cytotoxicity assay. Luciferase reporter, western blot, and RNA immunoprecipitation (RIP) assays were performed to assess the interaction between miR-20a and RUNX1. Result miR-20a was upregulated while RUNX1 was downregulated in NK cells from cervical cancer patients compared to healthy donors. IL-2 stimulated the releases of IFN-γ and TNF-α, and the killing effect of NK cells to cervical cancer cells, which was overturned by miR-20a introduction. RUNX1 was identified to be a target of miR-20a. Restoration of RUNX1 abolished the inhibitory effects of miR-20a on the secretions of IFN-γ and TNF-α, as well as the killing effect of NK cells to colorectal cancer cells. Conclusion miR-20a attenuated the killing effect of NK cells to cervical cancer cells by directly targeting RUNX1. Highlights • MiR-20a was upregulated while RUNX1 was downregulated in NK cells from cervical cancer patients. • MiR-20a attenuated the natural killing effect of NK cells to cervical cancer cells. • RUNX1 was targeted by miR-20a. • MiR-20a inhibited the natural killing effect of NK cells to cervical cancer cells via targeting RUNX1. [ABSTRACT FROM AUTHOR]