Biomarker Identification in Membranous Nephropathy Using a Long Non-coding RNA-Mediated Competitive Endogenous RNA Network.

PURPOSE: This study was aimed to identify biomarker associated with membranous nephropathy (MN) progression by integration of expression profiles and competitive endogenous RNA (ceRNA) network analysis. METHODS: The gene (GSE108113) and microRNAs (miRNAs) expression profiles (GSE64306) were downloaded to identify the differentially expressed mRNAs, miRNAs and long non-coding RNAs (lncRNAs) between MN and control groups. The functions and pathways enriched by the differentially expressed mRNAs were analyzed. The mRNA-lncRNA co-expression network was constructed followed by and the ceRNA network construction. RESULTS: Total 264 upregulated and 196 downregulated differentially expressed mRNAs, 79 upregulated and 4 downregulated lncRNAs, as well as 115 upregulated and 93 downregulated miRNAs were obtained between MN and control groups. After analysis, the differential mRNAs were significantly involved in multiple immune-related processes, and cell proliferation, apoptosis and differentiation processes, as well as pathways of taste transduction and lysosome. Finally, a ceRNA network consisting of 4 mRNAs (EPB41L5, FAM43A, PRKG1 and TTC14), 3 lncRNAs (LINC00052, LINC00641 and N4BP2L2-IT2) and 5 miRNAs (hsa-miR-145-5p, hsa-miR-3605-5p, hsa-miR-148a-3p, hsa-miR-497-5p and hsa-miR-148b-3p) was constructed. CONCLUSION: Our study indicated dysregulation of immune- and apoptosis-associated functions and taste transduction and lysosome pathways may play important roles in MN progression. Deregulated ceRNAs, such as LINC00052-hsa-miR-145-5p-EPB41L5, LINC00052-hsa-miR-148a-3p-FAM43A and LINC00641-hsa-497-5p-PRKG1, may be associated with MN development.

Construction and Validation of Predictive Model to Identify Critical Genes Associated with Advanced Kidney Disease.

BACKGROUND: Chronic kidney disease (CKD) is characterized by progressive renal function loss, which may finally lead to end-stage renal disease (ESRD). The study is aimed at identifying crucial genes related to CKD progressive and constructing a disease prediction model to investigate risk factors. METHODS: GSE97709 and GSE37171 datasets were downloaded from the GEO database including peripheral blood samples from subjects with CKD, ESRD, and healthy controls. Differential expressed genes (DEGs) were identified and functional enrichment analysis. Machine learning algorithm-based prediction model was constructed to identify crucial functional feature genes related to ESRD. RESULTS: A total of 76 DEGs were screened from CDK vs. normal samples while 10,114 DEGs were identified from ESRD vs. CDK samples. For numerous genes related to ESRD, several GO biological terms and 141 signaling pathways were identified including markedly upregulated olfactory transduction and downregulated platelet activation pathway. The DEGs were clustering in three modules according to WGCNA access, namely, ME1, ME2, and ME3. By construction of the XGBoost model and dataset validation, we screened cohorts of genes associated with progressive CKD, such as FZD10, FOXD4, and FAM215A. FZD10 represented the highest score (F score = 21) in predictive model. CONCLUSION: Our results demonstrated that FZD10, FOXD4, PPP3R1, and UCP2 might be critical genes in CKD progression.

Long non-coding RNA HCP5 serves as a ceRNA sponging miR-17-5p and miR-27a/b to regulate the pathogenesis of childhood obesity via the MAPK signaling pathway.

Background This study aimed to investigate the completing endogenous RNA (ceRNA) network involved in childhood obesity. Methods The microarray dataset GSE9624 was downloaded from the Gene Expression Omnibus (GEO) database. Differentially expressed long non-coding RNAs (lncRNAs) (DELs) and messenger RNAs (DEMs) were isolated between the childhood obesity and non-obesity tissue samples. Then, Gene Ontology (GO) functional and the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses of isolated DEMs were performed. DELs and DEMs targeted miRNAs were predicted to construct a ceRNA regulatory network. Finally, critical lncRNAs were validated in another dataset. Results A total of 1257 differentially expressed RNAs were screened, including 28 lncRNAs and 1229 mRNAs. In addition, these RNAs were mainly involved in defense response, cell cycle, mitogen-activated protein kinase (MAPK) signaling pathway, apoptosis, etc. Three lncRNAs (human leukocyte antigen complex 5 [HCP5], long intergenic non-protein coding RNA 839 [LINC00839] and receptor activity modifying protein 2 [RAMP2-AS1]) and two related miRNAs (hsa-miR-17-5p and hsa-miR-27a/b-3p) were identified as key RNAs in childhood obesity. Specifically, lncRNA HCP5 interacted with miR-17-5p and miR-27a/b to regulate nemo-like kinase (NLK) and Ras-related protein 2 (RRAS2) via the MAPK signaling pathway. Finally, four genes (RRAS2, NLK, bcl2/adenovirus E1B protein-interacting protein 3 [BNIP3] and phorbol-12-myristate-13-acetate-induced protein 1 [PMAIP1]) targeted by miRNAs were predicted as critical genes and might be novel diagnostic biomarkers of childhood obesity. Conclusions lncRNA HCP5 could serve as a ceRNA sponging miR-17-5p and miR-27a/b to regulate the pathogenesis of childhood obesity via NLK and RRAS2 in the MAPK signaling pathway.

Green synthesis of silver nanoparticles from Alpinia officinarum mitigates cisplatin-induced nephrotoxicity via down-regulating apoptotic pathway in rats.

The rhizome of A. officinarum possesses immense pharmaceutical properties like antioxidant, anti-inflammatory, antiapoptotic, anticancer activities. The foremost downside of herbal-based drugs is their poor bioavailability, to trounce this we synthesized a herbal based silver nanodrug with Alpinia officinarum rhizome extract and assessed its effect against the cisplatin-induced nephrotoxicity in in vivo model. The A. officinarum biosynthesized silver nanoparticles (AG-AO) were characterized using UV-Spec, FTIR, XRD, TEM and SAED analysis. The antioxidant and the nephroprotective property of biosynthesized AG-AO nanoparticles were assessed by estimating the levels of kidney biomarkers, cytokine, inflammatory markers, free radicals and antioxidants induced in control and experimental. Antiapoptotic effect of AG-AO nanoparticles were evaluated by measuring the levels of apoptotic proteins in control and experimental rats. Further, it is confirmed with histopathological analysis of kidney tissue with haematoxylin and eosin staining. Our physical analysis confirms the biosynthesized silver nanoparticles by A. officinarum and it satisfies the qualities of potent nanoparticles to be used for medication. Our biochemical, molecular and histopathological results confirm the antioxidant, antiapoptotic, anti-inflammatory properties of AG-AO. Overall our results authentically confirm AG-AO is a potent nephroprotective drug, which can be a supplementary drug to prevent cisplatin-induced nephrotoxicity.

Key Genes and Signaling Pathways Contribute to the Pathogensis of Diabetic Nephropathy.

INTRODUCTION: Diabetic nephropathy (DN) is a serious complication of diabetes mellitus involving damage to the capillaries in the glomerulus. This study aimed to explore key genes and signaling pathways participate in the progression of DN. METHODS: Two gene expression profile datasets GSE1009 and GSE30528 downloaded from Gene Expression Omnibus (GEO) were used to analyze the differentially expressed genes (DEGs) between DN samples and controls. Coupled two-way clustering (CTWC) and correspondence analysis were performed to explore the potential functions of DEGs. Then, Gene Ontology (GO) terms and pathways associated with DEGs were identified, followed by constructing of the co-expressed gene network and module. Ultimately, the regulatory network based on the DEGs, miRNAs and transcription factors (TFs) was established. RESULTS: Total 283 common DEGs were identified from the two datasets, including 219 down-regulated ones (bone morphogenetic protein 7 (BMP7), decay accelerating factor (CD55) and coagulation Factor V (F5) etc.) and 64 up-regulated ones (inhibin beta c subunit (INHBC) and colony stimulating factor 1 receptor (CSF1R) etc.). The miRNA-TF regulatory network was established with three miRNAs, 8 TFs and 58 DEGs. Besides, three significant pathways including cytokine-cytokine receptor interaction, complement and coagulation cascades and TGF-beta signaling pathways were identified. CONCLUSION: BMP7, CD55, CSF1R, INHBC and F5 are likely to take crucial roles in the pathogenesis of DN.

Both Peripheral Blood and Urinary miR-195-5p, miR-192-3p, miR-328-5p and Their Target Genes PPM1A, RAB1A and BRSK1 May Be Potential Biomarkers for Membranous Nephropathy.

BACKGROUND To identify noninvasive diagnostic biomarkers for membranous nephropathy (MN). MATERIAL AND METHODS The mRNA microarray datasets GSE73953 using peripheral blood mononuclear cells (PBMCs) of 8 membranous nephropathy patients and 2 control patients; and microRNAs (miRNA) microarray dataset GSE64306 using urine sediments of 4 membranous nephropathy patients and 6 control patients were downloaded from the Gene Expression Omnibus database. The differentially expressed genes (DEGs) and differentially expressed miRNAs (DEMs) were respectively identified from PBMCs and urine sediments of membranous nephropathy patients, followed with functional enrichment analysis, protein-protein interaction (PPI) analysis, and miRNA-target gene analysis. Finally, the DEGs and the target genes of DEMs were overlapped to obtain crucial miRNA-mRNA interaction pairs for membranous nephropathy. RESULTS A total of 1246 DEGs were identified from PBMCs samples, among them upregulated CCL5 was found to be involved in the chemokine signaling pathway, and BAX was found to be apoptosis related; while downregulated PPM1A and CDK1 were associated with the MAPK signaling pathway and the p53 signaling pathway, respectively. The hub role of CDK1 (degree=18) and CCL5 (degree=12) were confirmed after protein-protein interaction network analysis in which CKD1 could interact with RAB1A. A total of 28 DEMs were identified in urine sediments. The 276 target genes of DEMs were involved in cell cycle arrest (PPM1A) and intracellular signal transduction (BRSK1). Thirteen genes were shared between the DEGs in PMBCs and the target genes of DEMs in urine sediments, but only hsa-miR-192-3p-RAB1A, hsa-miR-195-5p-PPM1A, and hsa-miR-328-5p-BRSK1 were negatively related in their expression level. CONCLUSIONS Both peripheral blood and urinary miR-195-5p, miR-192-3p, miR-328-5p, and their target genes PPM1A, RAB1A, and BRSK1 may be potential biomarkers for membranous nephropathy by participating in inflammation and apoptosis.

Biomarker prediction for membranous nephropathy prognosis by microarray analysis.

AIM: This study aimed to explore biomarkers for membranous nephropathy (MN) diagnosis and to provide novel insights into the pathogenesis of this disease. METHODS: A microarray data set, GSE73953, was used, which contained 15 immunoglobulin A nephropathy (IgAN) samples, 8 MN samples and 2 healthy controls. Pretreatments were performed for the raw data, and then, the differentially expressed genes (DEG) were screened out using the limma package. The function and pathways of these genes were demonstrated through enrichment analysis. Protein-protein interaction (PPI) network analysis was performed to uncover interactions of DEG from the protein level. MN-related genes were further selected, integrating the Comparative Toxicogenomics Database (CTD). RESULTS: In total, 446 and 231 DEG were identified in the comparisons of MN versus control and MN versus IgAN, respectively. JUN, NFKB1, TGFB1 and PPBP were the predominant DEG, and the latter two were especially differentially expressed between the MN and IgAN groups. UBL4A and EIF4G1 were the two most important DEG for MN because they were downregulated compared with both control and IgAN groups. The above-mentioned genes were highlighted in the PPI networks and mainly enriched the ribosome- and platelet-related function/pathways. CONCLUSION: Several potential biomarkers were identified in MN, and some of them could well distinguish the MN from IgAN. Disruption of ribosome- and platelet-related function or pathways might contribute to MN progression. EIF4F and UBL4A might be two novel biomarkers for MN prognosis. Nevertheless, more experimental validations are needed.

miR-452 promotes cell metastasis and the epithelial to mesenchymal by targeting SOX7 in clear-cell renal-cell carcinoma.

Clear-cell renal-cell carcinoma (ccRCC) is the most common renal cell carcinoma (RCC), representing 75%-80% of the cases of RCC, and characterized by a high recurrence rate and poor prognosis. miR-452 acts as a tumor promoter in several tumors, including ccRCC. The purpose of this study was to determine the role of miR-452 in ccRCC. miR-452 and SOX7 messenger RNA and protein levels were calculated by quantitative reverse transcription polymerase chain reaction and Western blot analysis. MTT and Transwell assays were utilized to measure proliferative and invasive abilities. The Kaplan-Meier method was used to evaluate the association between the expression of miR-452 or SOX7 and the overall survival of ccRCC patients. Our results showed that miR-452 was overexpressed in ccRCC tissues and cells, and upregulation of miR-452 predicted a poor 5-year survival in ccRCC patients. In contrast, expression of SOX7 was low and downregulation of SOX7 predicted poor prognosis in ccRCC. In addition, miR-452 promoted cell proliferation, invasion, and the EMT, while SOX7 reversed the function of miR-452 on cell proliferation and invasion in 786-O cells. In conclusion, miR-452 was shown to inhibit cell proliferation, invasion, and the EMT through SOX7 in ccRCC, and the newly identified miR-452/SOX7 axis provided novel insight into the pathogenesis of ccRCC.

Serum exosomal miR-328, miR-575, miR-134 and miR-671-5p as potential biomarkers for the diagnosis of Kawasaki disease and the prediction of therapeutic outcomes of intravenous immunoglobulin therapy.

The present study was conducted to screen serum exosomal microRNAs (miRNAs) for the early diagnosis of Kawasaki disease (KD) and to investigate their underlying mechanisms by analyzing microarray data under accession numbers GSE60965 [exosomal miRNA, including three pooled serum samples from 5 healthy children, 5 patients with KD and 5 patients with KD following intravenous immunoglobulin (IVIG) therapy] and GSE73577 (mRNA, including peripheral blood mononuclear cell samples from 19 patients with KD prior to and following IVIG treatment) from the Gene Expression Omnibus database. Differentially expressed miRNAs (DE-miRNAs) and genes (DEGs) were identified using the Linear Models for Microarray data method, and the mRNA targets of DE-miRNAs were predicted using the miRWalk 2.0 database. The functions of the target genes were analyzed using the Database for Annotation, Visualization and Integrated Discovery (DAVID). As a result, 65 DE-miRNAs were identified with different expression patterns between the healthy children and patients with KD and between patients with KD and patients with KD following IVIG therapy. The target genes of 15 common DE-miRNAs were predicted. Following overlapping the target genes of DE-miRNAs with 355 DEGs, 28 common genes were identified and further screened to construct a network containing 30 miRNA-mRNA regulatory associations. Of these associations, only miR-328-spectrin α, erythrocytic 1, miR-575-cyclic AMP-responsive element-binding protein 5/b-1,4-galactosyltransferase 5/WD repeat and FYVE domain-containing 3/cystatin-A/C-X-C motif chemokine receptor 1/protein phosphatase 1 regulatory subunit 3B, miR-134-acyl-CoA synthetase long chain family member 1/C-type lectin domain family 1 member A and miR-671-5p-tripartite motif containing 25/leucine rich repeat kinase 2/kinesin family member 1B/leucine rich repeat neuronal 1 were involved in the negative regulation of gene expression. Functional analysis indicated that the identified target genes may be associated with inflammation. Accordingly, serum exosomal miR-328, miR-575, miR-134 and miR-671-5p may act as potential biomarkers for the diagnosis of KD and the prediction of outcomes of the IVIG therapy by influencing the expression of inflammatory genes.

Candidate gene and mechanism investigations in congenital obstructive nephropathy based on bioinformatics analysis.

The aim of the present study was to explore the candidate genes, chemicals and mechanisms of congenital obstructive nephropathy (CON). The gene expression profiles of GSE48041, including 24 kidney tissue samples from megabladder (mgb­/­) mouse were downloaded from the Gene Expression Omnibus database. Samples were divided into 4 groups: Control, mild, moderate and severe. Differentially expressed genes (DEGs), protein­protein interaction network, Kyoto Encyclopedia of Genes and Genomes pathways and transcription factor (TF)­target gene analyses were performed on Set 1 (mild, moderate and severe groups), while Gene Ontology (GO) function enrichment analysis and chemical investigation were performed on Set 2 (severe group). A total of 187 and 139 DEGs were obtained in Set 1 and Set 2, respectively. Chemical carcinogenesis [enriched by genes such as Carbonyl reductase 1 (CBR1)] was one of the most prominent pathways in Set 1. GO analysis for Set 2 revealed that DEGs were mainly assembled in functions such as cellular response to interleukin­1 and cellular response to tumor necrosis. Furthermore, genes such as Fos Proto­Oncogene (FOS) were co­regulated by TFs including RNA polymerase II subunit A (Polr2a) and serum response factor (Srf). Chemical cyclosporine served the most important role in Set 2 by targeting several DEGs in Set 2. DEGs such as CBR1 and FOS, TFs including Polr2a and Srf, and pathways such as chemical carcinogenesis may serve important roles in the process of CON. Interleukin­1 and tumor necrosis function may be novel targets for CON gene therapy. Furthermore, cyclosporine may be a promising option for future CON therapy.

Investigation of mechanisms of mesenchymal stem cells for treatment of diabetic nephropathy via construction of a miRNA-TF-mRNA network.

BACKGROUND: Recent studies have reported that mesenchymal stem cells (MSCs) exert therapeutic effects on the treatment of diabetic nephropathy (DN), but the underlying mechanisms remain unclear. METHODS: A dataset GSE65561 was obtained from Gene Expression Omnibus (GEO) database, which contained four healthy control samples (group 1), four healthy controls samples co-cultured with MSCs (group 2), five DN samples (group 3) and five DN samples co-cultured with MSCs (group 4). The differentially expressed genes (DEGs) between group 3 vs. group 1 and group 4 vs. group 2 were constructed using Linear Models for Microarray (LIMMA) package package. Then, DAVID was used to analyze the functional enrichment of DEGs. Based on STRING database the protein-protein interaction (PPI) network was visualized by the Cytoscape plug-in CytoNCA. Besides, the hub miRNAs and transcription factors (TFs) regulating DEGs were predicted using Webgestalt. RESULTS: Totally, 303 up-regulated and 88 down-regulated DEGs were shared in group 3 vs. group 1 and group 4 vs. group 2. Besides, the up-regulated DEGs were mainly enriched in 'translation' and 'translational elongation', while the down-regulated genes were only enriched in 'protein kinase activity'. RPS27A and RPLP0 had a higher degree in the PPI network and they were regulated by EIF3M. In addition, ETF1 was predicted to be an important gene, which was regulated by miR-150, miR-134 and EIF2S1. CONCLUSIONS: RPS27A, RPLP0 and ETF1 may be potential targets for MSCs on the treatment of DN. Highlights RPS27A and RPLP0 may be important genes in the treatment of MSCs for DN. TF EIF3M may play a key role in the treatment of MSCs for DN. MiR-150 and miR-134 may be essential microRNAs in the treatment of MSCs for DN.

Identification of key microRNAs, transcription factors and genes associated with congenital obstructive nephropathy in a mouse model of megabladder.

OBJECTIVE: The present study aimed to investigate the molecular mechanism underlying congenital obstructive nephropathy (CON). METHODS: The microarray dataset GSE70879 was downloaded from the Gene Expression Omnibus, including 3 kidney samples of megabladder mice and 4 control kidneys. Using this dataset, differentially expressed miRNAs (DEMs) were identified between the kidney samples from megabladder mice and controls, followed by identification of the target genes for these DEMs and construction of a DEM and target gene interaction network. Additionally, the target genes were subjected to Gene Ontology and pathway enrichment analyses, and were used for construction of a protein-protein interaction (PPI) network. Finally, regulatory networks were constructed to analyze transcription factors for the key miRNAs. RESULTS: From 17 DEMs identified between kidney samples of megabladder mice and controls, 3 key miRNAs were screened, including mmu-miR-150-5p, mmu-miR-374b-5p and mmu-miR-126a-5p. The regulatory networks identified vascular endothelial growth factor A (Vegfa) as the common target gene of mmu-miR-150-5p and five transcription factors, including nuclear receptor subfamily 4, group A, member 2 (Nr4a2), Jun dimerisation protein 2 (Jdp2), Kruppel-like factor 6 (Klf6), Neurexophilin-3 (Nxph3) and RNA binding motif protein 17 (Rbm17). The gene encoding phosphatase and tensin homolog (Pten) was found to be co-regulated by mmu-miR-374b-5p and high mobility group protein A1 (Hmga1), whereas the kirsten rat sarcoma viral oncogene (Kras) was identified as a common target gene of mmu-miR-126a-5p and paired box 6 (Pax6). CONCLUSIONS: In summary, the above-listed key miRNAs, transcription factors and key genes may be involved in the development of CON.

Potential role of upregulated microRNA­146b and ­21 in renal fibrosis.

The aim of the present study was to identify candidate microRNAs (miRNAs) involved in the progression of renal fibrosis. Dataset GSE42716 of miRNAs extracted from kidneys from mice with unilateral ureteral obstruction (UUO), and mice without UUO was downloaded from the Gene Expression Omnibus database. The Limma package was used to identify differential expression between mice with and without UUO. Renal disease­related miRNAs were predicted based on the miRWalk database. Thereafter, candidate miRNAs were screened by taking the intersection of differentially expressed miRNAs and predicted miRNAs, followed by screening of target genes using miRWalk and transcription factors using the TransmiR database. An integrative regulatory network was constructed using Cytoscape software. Enrichment analysis of target genes was also performed. In total, 76 differentially expressed miRNAs were identified in kidneys with UUO compared with the normal samples based on dataset and 9 miRNAs were identified as related to renal disease from the miRWalk database. A Venn diagram revealed two overlapping upregulated miRNAs; miR­146b and miR­21. Transcription factor NFKB1 may activate miR­146b and AKT may activate miR­21. In addition, miR­21 had a regulatory effect on IFNG expression and miR­146b may regulate the expression of BCL2, PTEN and IFNG. Furthermore, target genes of miR­146b and miR­21 were significantly enriched in 14 Gene Ontology terms including regulation of cell proliferation (e.g., BCL2, PTEN and IFNG). Overexpression of miR­21 may be activated by AKT2 and contribute to renal fibrosis by negatively regulating IFNG expression. Furthermore, miR­146b may be activated by NFKB1 and subsequently reduce the expression of BCL2, PTEN and IFNG in the progression of renal fibrosis.

Sex hormone affects the severity of non-alcoholic steatohepatitis through the MyD88-dependent IL-6 signaling pathway.

Recent research has shown that the occurrence of gender disparity in liver cancer associated with sex differences in MyD88-dependent IL-6 production, but the role of this signaling pathway in sex differences of non-alcoholic steatohepatitis (NASH) remains unknown. To investigate the effects of sex hormone-specific intervention on pathology and progression of NASH, and on the inflammatory TLR-MyD88-IL-6 signaling pathway NASH was modeled in C57/BL6 mice by feeding a methionine and choline-deficient (MCD) diet for 4 weeks. Male mice were subjected to sex hormone-related interventions such as orchidectomy, and orchidectomy combined with administration of either testosterone propionate or estradiol benzoate. Next, the degree of non-alcoholic fatty liver disease activity score (NAS), serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), and the expression level of MyD88 and IL-6, were compared between these groups. Males developed more serious inflammatory problems and had a higher NAS than the females. Sex-specific intervention in male mice by orchidectomy reduced NAS, ALT, and AST, and the expression level of MyD88 and IL-6. But administration of exogenous androgen had no influence on either NAS or the expression of ALT, AST, MyD88, and IL-6. On the other hand, exogenous estrogen could alleviate the pathological damage caused by NASH, as well as reduce NAS, ALT and AST, and the expression of MyD88 and IL-6. The result show different sex hormone-related interventions affected the severity of NASH, possibly by modulating the level of sex hormones and regulating the TLR-MyD88-IL-6 signaling pathway.