scholarly journals Screening and Identification of Key Genes for Activation of Islet Stellate Cell

2021 ◽  
Vol 12 ◽  
Author(s):  
Xiaohang Wang ◽  
Vladmir Carvalho ◽  
Qianqian Wang ◽  
Jinbang Wang ◽  
Tingting Li ◽  
...  
Keyword(s):  
Real Time ◽  
Differentially Expressed Genes ◽  
Western Blotting ◽  
Real Time Pcr ◽  
Stellate Cells ◽  
Differentially Expressed ◽  
Mrna Levels ◽  
Gk Rats ◽  
A Genome ◽  
Key Genes

BackgroundIt has been demonstrated that activated islet stellate cells (ISCs) play a critical role in islet fibrogenesis and significantly contribute to the progression of type 2 diabetes mellitus. However, the key molecules responsible for ISCs activation have not yet been determined. This study aimed to identify the potential key genes involved in diabetes-induced activation of ISCs.MethodStellate cells were isolated from three 10-week-old healthy male Wistar rats and three Goto-Kakizaki (GK) rats. Cells from each rat were primary cultured under the same condition. A Genome-wide transcriptional sequence of stellate cells was generated using the Hiseq3000 platform. The identified differentially expressed genes were validated using quantitative real-time PCR and western blotting in GK rats, high fat diet (HFD) rats, and their controls.ResultsA total of 204 differentially expressed genes (DEGs) between GK. ISCs and Wistar ISCs (W.ISCs) were identified, accounting for 0.58% of all the 35,362 genes detected. After the Gene Ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) enrichment analyses, the mRNA levels of these genes were further confirmed by real-time PCR in cultured ISCs. We then selected Fos, Pdpn, Bad as the potential key genes for diabetes-induced activation of ISCs. Finally, we confirmed the protein expression levels of FOS, podoplanin, and Bad by western blotting and immunofluorescence in GK rats, HFD rats, and their controls. The results showed that the expression level of FOS was significantly decreased, while podoplanin and Bad were significantly increased in GK.ISCs and HFD rats compared with controls, which were consistent with the expression of α-smooth muscle actin.ConclusionsA total of 204 DEGs were found between the GK.ISCs and W.ISCs. After validating the expression of potential key genes from GK rats and HFD rats, Fos, Pdpn, and Bad might be potential key genes involved in diabetes-induced activation of ISCs.

Increased relaxation of immature airways to β2-adrenoceptor agonists is related to attenuated expression of postjunctional smooth muscle muscarinic M2 receptors

2005 ◽  
Vol 98 (4) ◽  
pp. 1526-1533 ◽  
Author(s):  
Michael Fayon ◽  
Eric Dumas De La Roque ◽  
Patrick Berger ◽  
Hugues Begueret ◽  
Olga Ousova ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Real Time ◽  
Airway Smooth Muscle ◽  
Western Blotting ◽  
Real Time Pcr ◽  
Molecular Techniques ◽  
Receptor Expression ◽  
Smooth Muscle Relaxation ◽  
Fetal Life ◽  
Mrna Levels

Spontaneous or agonist-induced contraction of airway smooth muscle can be observed very early in fetal life, thus explaining the possible occurrence of bronchospasm in very low birth weight infants within the first days of life. In an attempt to better manage such bronchospasms, the aim of the present study was to investigate the age-specific modifications in airway smooth muscle relaxation to β2-agonists and muscarinic antagonists using a combination of functional and molecular techniques. In the rat, isometric relaxation to the β2-agonist salbutamol was examined in tracheae; we also examined muscarinic receptor expression (M2R and M3R mRNA levels) in airway smooth muscle by immunochemistry, Western blotting, and real-time PCR. Compared with adults, salbutamol-induced relaxation was twofold greater in immature rat isolated tracheae preconstricted by carbachol. This effect was associated with a lower expression of M2R in the smooth muscle of immature animals (sixfold and almost twofold as assessed by immunochemistry and Western blotting, respectively). Real-time PCR data indicate that changes in M2R expression according to age occurred at a posttranscriptional level. In adult airways, there was a significantly greater functional efficacy of M2R blockade by methoctramine compared with that shown in immature rats. Because of the limited availability of human neonate lung tissue, only the molecular part of the study was performed, and we observed a qualitatively similar effect, i.e., a lower M2R expression in the neonatal airway smooth muscle, although this was quantitatively smaller. We conclude that β2-agonist-induced relaxation is enhanced in immature compared with adult airways as a result of greater postjunctional M2R expression in adult airway smooth muscle. This finding may be of importance in the clinical management of bronchoconstriction in neonates.

Sepsis stimulates calpain activity in skeletal muscle by decreasing calpastatin activity but does not activate caspase-3

2005 ◽  
Vol 288 (3) ◽  
pp. R580-R590 ◽  
Author(s):  
Wei Wei ◽  
Moin U. Fareed ◽  
Amy Evenson ◽  
Michael J. Menconi ◽  
Hongmei Yang ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Real Time ◽  
Western Blotting ◽  
Real Time Pcr ◽  
Caspase 3 ◽  
Calpain Inhibitor ◽  
Mrna Levels ◽  
Protein Breakdown ◽  
Calpain Activity ◽  
Hydrolysis Of

We examined the influence of sepsis on the expression and activity of the calpain and caspase systems in skeletal muscle. Sepsis was induced in rats by cecal ligation and puncture (CLP). Control rats were sham operated. Calpain activity was determined by measuring the calcium-dependent hydrolysis of casein and by casein zymography. The activity of the endogenous calpain inhibitor calpastatin was measured by determining the inhibitory effect on calpain activity in muscle extracts. Protein levels of μ- and m-calpain and calpastatin were determined by Western blotting, and calpastatin mRNA was measured by real-time PCR. Caspase-3 activity was determined by measuring the hydrolysis of the fluorogenic caspase-3 substrate Ac-DEVD-AMC and by determining protein and mRNA expression for caspase-3 by Western blotting and real-time PCR, respectively. In addition, the role of calpains and caspase-3 in sepsis-induced muscle protein breakdown was determined by measuring protein breakdown rates in the presence of specific inhibitors. Sepsis resulted in increased muscle calpain activity caused by reduced calpastatin activity. In contrast, caspase-3 activity, mRNA levels, and activated caspase-3 29-kDa fragment were not altered in muscle from septic rats. Sepsis-induced muscle proteolysis was blocked by the calpain inhibitor calpeptin but was not influenced by the caspase-3 inhibitor Ac-DEVD-CHO. The results suggest that sepsis-induced muscle wasting is associated with increased calpain activity, secondary to reduced calpastatin activity, and that caspase-3 activity is not involved in the catabolic response to sepsis.

scholarly journals Identification of key genes of papillary thyroid carcinoma by integrated bioinformatics analysis

2020 ◽  
Vol 40 (8) ◽  
Author(s):  
Gang Xue ◽  
Xu Lin ◽  
Jing-Fang Wu ◽  
Da Pei ◽  
Dong-Mei Wang ◽  
...  
Keyword(s):  
Papillary Thyroid Carcinoma ◽  
Thyroid Carcinoma ◽  
Differentially Expressed Genes ◽  
Disease Free Survival ◽  
Differentially Expressed ◽  
Mrna Levels ◽  
Papillary Thyroid ◽  
Positive Role ◽  
Immune Infiltration ◽  
Key Genes

Abstract Background: Papillary thyroid carcinoma (PTC) is one of the fastest-growing malignant tumor types of thyroid cancer. Therefore, identifying the interaction of genes in PTC is crucial for elucidating its pathogenesis and finding more specific molecular biomarkers. Methods: Four pairs of PTC tissues and adjacent tissues were sequenced using RNA-Seq, and 3745 differentially expressed genes were screened (P<0.05, |logFC|>1). The enrichment analysis indicated that the vast majority of differentially expressed genes (DEGs) may play a positive role in the development of cancer. Then, the significant modules were analyzed using Cytoscape software in the protein–protein interaction network. Survival analysis, TNM analysis, and immune infiltration analysis of key genes were analyzed. And the expression of ADORA1, APOE, and LPAR5 genes were verified by qPCR in PTC compared with matching adjacent tissues. Results: Twenty-five genes were identified as hub genes with nodes greater than 10. The expression of 25 genes were verified by the GEPIA database, and the overall survival and disease-free survival analyses were conducted with Kaplan–Meier plotter. We found only three genes were confirmed with our validation and were statistically significant in PTC, namely ADORA1, APOE, and LPAR5. Further analysis found that the mRNA levels and methylation degree of these three genes were significantly correlated with the TNM staging of PTC. And these three genes were related to PTC immune infiltration. Verification of the expression of these three genes by RT-qPCR and Western blot further confirmed the reliability of our results. Conclusion: Our study identified three genes that may play key regulatory roles in the development, metastasis, and immune infiltration of papillary thyroid carcinoma.

scholarly journals Endometriosis-Specific Genes Identified by Real-Time Reverse Transcription-Polymerase Chain Reaction Expression Profiling of Endometriosis Versus Autologous Uterine Endometrium

2006 ◽  
Vol 91 (1) ◽  
pp. 228-238 ◽  
Author(s):  
Wei-Ping Hu ◽  
Sun Kuie Tay ◽  
Yi Zhao
Keyword(s):  
Gene Expression ◽  
Real Time ◽  
Differentially Expressed Genes ◽  
Expression Profiling ◽  
Differentially Expressed ◽  
Cdna Libraries ◽  
Eutopic Endometrium ◽  
Tissue Samples ◽  
Uterine Endometrium ◽  
A Genome

Abstract Context: The etiology and molecular pathogenesis of endometriosis, a prevalent estrogen-dependent gynecologic disease, are poorly understood. Objective: The objective of the study was to identify the differentially expressed genes between autologous ectopic and eutopic endometrium. Design: Subtractive hybridization was used for a genome-wide search for differentially expressed genes between autologous ectopic and eutopic endometrium. Real-time RT-PCR was used for gene expression profiling in the paired tissue samples taken from multiple subjects. Patients: The paired pelvic endometriosis and uterine endometrium tissue biopsies were procured from 15 patients undergoing laparoscopy or hysterectomy for endometriosis. Results: Seventy-eight candidate genes were identified from the subtractive cDNA libraries. Seventy-six of these genes were investigated in approximately 8000 real-time PCR for their differential expression in 30 paired tissue biopsies from 15 patients affected by endometriosis. Cluster analysis on gene expression revealed highly consistent profiles in two groups of genes, despite the clinical heterogeneity of the 15 cases. Thirty-four genes specific to early disease point to their potential roles in establishment and evolution of endometriosis. Most interestingly, 14 genes were consistently dysregulated in the paired samples from the majority of the patients. Of these, there were two uncharacterized transcripts and two novel genes, and 10 were matched to known genes: IGFBP5, PIM2, RPL41, PSAP, FBLN1, SIPL, DLX5, HSD11B2, SET, and RHOE. Conclusions: Dysregulation of 14 genes was found to be overtly associated with endometriosis. Some of these genes, known to participate in estrogen activities and antiapoptosis, may play a role in the pathogenesis of endometriosis and may represent potential diagnostic markers or therapeutic targets for endometriosis.

scholarly journals AB0005 IDENTIFICATION OF KEY GENES AND PATHWAYS FOR PSORIASIS BASED ON GEO DATABASES BY BIOINFORMATICS ANALYSIS

2021 ◽  
Vol 80 (Suppl 1) ◽  
pp. 1037.2-1038
Author(s):  
X. Sun ◽  
S. X. Zhang ◽  
S. Song ◽  
T. Kong ◽  
C. Zheng ◽  
...  
Keyword(s):  
Gene Expression ◽  
Signaling Pathways ◽  
Differentially Expressed Genes ◽  
Enrichment Analysis ◽  
Differentially Expressed ◽  
Shanxi Province ◽  
Receptor Interaction ◽  
Term Therapy ◽  
Pathway Enrichment Analysis ◽  
Key Genes

Background:Psoriasis is an immune-mediated, genetic disease manifesting in the skin or joints or both, and also has a strong genetic predisposition and autoimmune pathogenic traits1. The hallmark of psoriasis is sustained inflammation that leads to uncontrolled keratinocyte proliferation and dysfunctional differentiation. And it’s also a chronic relapsing disease, which often necessitates a long-term therapy2.Objectives:To investigate the molecular mechanisms of psoriasis and find the potential gene targets for diagnosis and treating psoriasis.Methods:Total 334 gene expression data of patients with psoriasis research (GSE13355 GSE14905 and GSE30999) were obtained from the Gene Expression Omnibus database. After data preprocessing and screening of differentially expressed genes (DEGs) by R software. Online toll Metascape3 was used to analyze Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of DEGs. Interactions of proteins encoded by DEGs were discovered by Protein-protein interaction network (PPI) using STRING online software. Cytoscape software was utilized to visualize PPI and the degree of each DEGs was obtained by analyzing the topological structure of the PPI network.Results:A total of 611 DEGs were found to be differentially expressed in psoriasis. GO analysis revealed that up-regulated DEGs were mostly associated with defense and response to external stimulus while down-regulated DEGs were mostly associated with metabolism and synthesis of lipids. KEGG enrichment analysis suggested they were mainly enriched in IL-17 signaling, Toll-like receptor signaling and PPAR signaling pathways, Cytokine-cytokine receptor interaction and lipid metabolism. In addition, top 9 key genes (CXCL10, OASL, IFIT1, IFIT3, RSAD2, MX1, OAS1, IFI44 and OAS2) were identified through Cytoscape.Conclusion:DEGs of psoriasis may play an essential role in disease development and may be potential pathogeneses of psoriasis.References:[1]Boehncke WH, Schon MP. Psoriasis. Lancet 2015;386(9997):983-94. doi: 10.1016/S0140-6736(14)61909-7 [published Online First: 2015/05/31].[2]Zhang YJ, Sun YZ, Gao XH, et al. Integrated bioinformatic analysis of differentially expressed genes and signaling pathways in plaque psoriasis. Mol Med Rep 2019;20(1):225-35. doi: 10.3892/mmr.2019.10241 [published Online First: 2019/05/23].[3]Zhou Y, Zhou B, Pache L, et al. Metascape provides a biologist-oriented resource for the analysis of systems-level datasets. Nat Commun 2019;10(1):1523. doi: 10.1038/s41467-019-09234-6 [published Online First: 2019/04/05].Acknowledgements:This project was supported by National Science Foundation of China (82001740), Open Fund from the Key Laboratory of Cellular Physiology (Shanxi Medical University) (KLCP2019) and Innovation Plan for Postgraduate Education in Shanxi Province (2020BY078).Disclosure of Interests:None declared

scholarly journals Key Genes and Prognostic Analysis in HER2+ Breast Cancer

2021 ◽  
Vol 20 ◽  
pp. 153303382098329
Author(s):  
Yujie Weng ◽  
Wei Liang ◽  
Yucheng Ji ◽  
Zhongxian Li ◽  
Rong Jia ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Gene Expression ◽  
Differentially Expressed Genes ◽  
Protein Interaction ◽  
Differentially Expressed ◽  
Protein Kinase Activity ◽  
Protein Protein Interaction ◽  
Risk Of Death ◽  
Her2 Breast Cancer ◽  
Key Genes

Human epidermal growth factor 2 (HER2)+ breast cancer is considered the most dangerous type of breast cancers. Herein, we used bioinformatics methods to identify potential key genes in HER2+ breast cancer to enable its diagnosis, treatment, and prognosis prediction. Datasets of HER2+ breast cancer and normal tissue samples retrieved from Gene Expression Omnibus and The Cancer Genome Atlas databases were subjected to analysis for differentially expressed genes using R software. The identified differentially expressed genes were subjected to gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses followed by construction of protein-protein interaction networks using the STRING database to identify key genes. The genes were further validated via survival and differential gene expression analyses. We identified 97 upregulated and 106 downregulated genes that were primarily associated with processes such as mitosis, protein kinase activity, cell cycle, and the p53 signaling pathway. Visualization of the protein-protein interaction network identified 10 key genes ( CCNA2, CDK1, CDC20, CCNB1, DLGAP5, AURKA, BUB1B, RRM2, TPX2, and MAD2L1), all of which were upregulated. Survival analysis using PROGgeneV2 showed that CDC20, CCNA2, DLGAP5, RRM2, and TPX2 are prognosis-related key genes in HER2+ breast cancer. A nomogram showed that high expression of RRM2, DLGAP5, and TPX2 was positively associated with the risk of death. TPX2, which has not previously been reported in HER2+ breast cancer, was associated with breast cancer development, progression, and prognosis and is therefore a potential key gene. It is hoped that this study can provide a new method for the diagnosis and treatment of HER2 + breast cancer.

Increased Expression of TXNIP Facilitates Oxidative Stress in Nasal Epithelial Cells of Patients With Chronic Rhinosinusitis With Nasal Polyps

2020 ◽  
pp. 194589242098241
Author(s):  
Hai Lin ◽  
Guangyi Ba ◽  
Ru Tang ◽  
Mingxian Li ◽  
Zhipeng Li ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Epithelial Cells ◽  
Real Time ◽  
Chronic Rhinosinusitis ◽  
Western Blotting ◽  
Real Time Pcr ◽  
Nasal Polyps ◽  
Sod Activity ◽  
Nasal Tissue ◽  
Sod Assay

Background Oxidative stress plays crucial roles in the pathogenesis of chronic rhinosinusitis with nasal polyps (CRSwNP). Thioredoxin-interacting protein (TXNIP) is essential in the process of triggering oxidative stress. However, its role and mechanism in CRSwNP remain unclear. The present study sought to explore the role and mechanism of TXNIP in the pathogenesis of CRSwNP. Methods Western blotting, real-time PCR and immunohistochemistry (IHC) were employed to assess TXNIP, thioredoxin (TRX) expression in nasal tissue samples from patients with CRSwNP and control subjects. MDA level and SOD activity in nasal tissue homogenates were measured using MDA and SOD Assay Kit. To evaluate the role and mechanism of TXNIP in CRSwNP, human nasal epithelial cells (HNECs) were cultured and stimulated using TXNIP siRNA, with or without N-acetylcysteine (NAC, an ROS scavenger). Western blotting, real-time PCR, ROS detecting dye DCFH-DA, MDA and SOD Assay Kit were performed to assess the effects and mechanisms of stimulators on the cells. Results We found significantly increased levels of TXNIP and decreased levels of TRX protein, mRNA, positive cells, increased MDA level and decreased SOD activity in CRSwNP patients compared with control subjects. In vitro study, significantly altered levels of TXNIP, TRX, MDA, SOD and ROS in HNECs were found following treatment of TXNIP siRNA with or without NAC on HNECs. Conclusion TXNIP expression was increased and TRX expression was decreased in CRSwNP at both protein and mRNA levels. MDA levels were increased and SOD activities were decreased in CRSwNP. TXNIP may have negative association with TRX, and then decrease SOD activities and increase MDA levels, resulting in the upregulation of ROS and oxidative stress in HNECs, which may play a pivotal role in the pathogenesis of CRSwNP. Future studies are expected to further explore the role and mechanism of TXNIP in CRSwNP.

DEF8 and Autophagy-Associated Genes are Altered in Mild Cognitive Impairment, Probable Alzheimer’s Disease Patients and a Transgenic Model of the Disease

2021 ◽  
pp. 1-16
Author(s):  
Esteban Leyton ◽  
Diego Matus ◽  
Sandra Espinoza ◽  
José Matías Benitez ◽  
Bastián I. Cortes ◽  
...  
Keyword(s):  
Alzheimer’S Disease ◽  
Alzheimer's Disease ◽  
Cognitive Impairment ◽  
Mild Cognitive Impairment ◽  
Real Time ◽  
Real Time Pcr ◽  
Mononuclear Cells ◽  
Differentially Expressed ◽  
Protein Levels ◽  
5Xfad Mice

Background: Disturbances in the autophagy/endolysosomal systems are proposed as early signatures of Alzheimer’s disease (AD). However, few studies are available concerning autophagy gene expression in AD patients. Objective: To explore the differential expression of classical genes involved in the autophagy pathway, among them a less characterized one, DEF8 (Differentially expressed in FDCP 8), initially considered a Rubicon family member, in peripheral blood mononuclear cells (PBMCs) from individuals with mild cognitive impairment (MCI) and probable AD (pAD) and correlate the results with the expression of DEF8 in the brain of 5xFAD mice. Method: By real-time PCR and flow cytometry, we evaluated autophagy genes levels in PBMCs from MCI and pAD patients. We evaluated DEF8 levels and its localization in brain samples of the 5xFAD mice by real-time PCR, western blot, and immunofluorescence. Results: Transcriptional levels of DEF8 were significantly reduced in PBMCs of MCI and pAD patients compared with healthy donors, correlating with the MoCA and MoCA-MIS cognitive tests scores. DEF8 protein levels were increased in lymphocytes from MCI but not pAD, compared to controls. In the case of brain samples from 5xFAD mice, we observed a reduced mRNA expression and augmented protein levels in 5xFAD compared to age-matched wild-type mice. DEF8 presented a neuronal localization. Conclusion: DEF8, a protein proposed to act at the final step of the autophagy/endolysosomal pathway, is differentially expressed in PBMCs of MCI and pAD and neurons of 5xFAD mice. These results suggest a potential role for DEF8 in the pathophysiology of AD.

Genome-wide screening of m6A-modified transcript in the Wilms’ tumor

2021 ◽  
Author(s):  
Zhuo Liu ◽  
Feng He ◽  
Jing Liu ◽  
Shengrong OuYang ◽  
Zexi Li ◽  
...  
Keyword(s):  
Gene Ontology ◽  
Real Time ◽  
Real Time Pcr ◽  
Wilms Tumor ◽  
Expression Patterns ◽  
Mrna Levels ◽  
Gene Ontology Analysis ◽  
Quantitative Real Time Pcr ◽  
Related Factors ◽  
Significant Difference

Abstract Background Wilms’ tumor, also called nephroblastoma, is the most common pediatric renal malignancy. The pathogenesis of Wilms’ tumor has been attributed to several genetic and epigenetic factors. However, the most pervasive internal mRNA modification that affects almost every process of RNA metabolism, RNA N6-Methyladenosine (m6A) methylation, has not been characterized in Wilms’ tumor. Methods Wilms’ tumor (WT) and adjacent non-cancerous (NC) tissue samples were obtained from 23 children with nephroblastoma, and the global m6A levels were measured by mass spectrometry. Analyses by m6A-mRNA epitranscriptomic microarray and mRNA microarray were performed, and m6A-related mRNAs were validated by quantitative real-time PCR for input and m6A-immunoprecipitated RNA samples from WT and NC tissues. Gene ontology analysis and KEGG pathway analysis were performed for differentially expressed genes, and expression of RNA methylation-related factors was measured by quantitative real-time PCR. Results The total m6A methylation levels in total RNA of WT samples and NC samples were (0.21 ± 0.01)% and (0.22 ± 0.01)%, respectively, with no statistically significant difference. Fifty-nine transcripts were differentially m6A-methylated between the WT and NC groups, which showed distinct m6A modification patterns. Gene ontology analysis indicated that m6A-modified genes were enriched in cancer-associated pathways, including the mTOR pathway, and conjoint analysis of the unique methylation and gene expression patterns in WT samples suggested an association with metabolic pathways.The mRNA levels of the m6A-related “reader” genes, YTHDF1, YTHDF2 and IGF2BP3, were statistically higher in WT samples than in NC samples. Conclusion This is the first study to determine the m6A modification profiles in Wilms’ tumor. Our data provide novel information regarding patterns of m6A modification that correlate with carcinogenesis in Wilms’ tumor.

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