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Altered microRNA expression in stenoses of native arteriovenous fistulas in hemodialysis patients  Lei Lv, MD, Weibin Huang, MD, Jiwei Zhang, MD, Yaxue.

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Presentation on theme: "Altered microRNA expression in stenoses of native arteriovenous fistulas in hemodialysis patients  Lei Lv, MD, Weibin Huang, MD, Jiwei Zhang, MD, Yaxue."— Presentation transcript:

1 Altered microRNA expression in stenoses of native arteriovenous fistulas in hemodialysis patients 
Lei Lv, MD, Weibin Huang, MD, Jiwei Zhang, MD, Yaxue Shi, MD, Lan Zhang, MD  Journal of Vascular Surgery  Volume 63, Issue 4, Pages e3 (April 2016) DOI: /j.jvs Copyright © 2016 Society for Vascular Surgery Terms and Conditions

2 Fig 1 MicroRNA (miRNA) profiles differentiate the stenosed vein segments of primary arteriovenous fistulas (AVFs) from the control segments. A, Heatmap of aberrantly expressed miRNAs in the stenosed vein segments of primary AVF and the controls. Colors indicate relative signal intensities: red and green colors indicate upregulated and downregulated miRNAs, respectively. B, Volcano plot demonstrates the differences in expression levels of miRNAs between stenosed vein segments of primary AVFs and the control veins. The log-fold change was plotted against the log odds of differential expression using the R package limna (The R Foundation for Statistical Computing, The vertical lines correspond to twofold up and down, respectively, and the horizontal line represents a P value = .05. The red point in the plot represents the differentially expressed miRNAs with statistical significance. A volcano plot provided further information about the significance and magnitude of expressive alteration of selected miRNAs. Journal of Vascular Surgery  , e3DOI: ( /j.jvs ) Copyright © 2016 Society for Vascular Surgery Terms and Conditions

3 Fig 2 Verification of selected microarray data by quantitative reverse-transcription polymerase chain reaction (qRT-PCR). Triplicate assays were done for each RNA sample, and the relative amount of each microRNA (miRNA) was normalized to U6 small nuclear RNA. Values are expressed as mean ± standard deviation. *P < .05 indicates statistically significant difference between the two groups. A, Upregulated miRNAs in the stenosed vein segments of primary arteriovenous fistulas (AVFs). B, Downregulated miRNAs in the stenosed vein segments of primary AVFs. Journal of Vascular Surgery  , e3DOI: ( /j.jvs ) Copyright © 2016 Society for Vascular Surgery Terms and Conditions

4 Fig 3 The microRNA gene ontology (GO) network. GO function enrichment was performed by mapping the predicted target genes from the microRNAs (miRNAs) to the GO databases. By enrichment analyses on the predicted targets predicted from the significantly validated most differentially expressed miRNAs, we determined that the target genes could be categorized into a large number of significant GO terms. The major target genes in the biologic process categories were involved in cellular process, protein binding, regulation of biologic process, RNA biosynthetic process, regulation of metabolic process, and cell proliferation, among others. The circles represent genes and the squares represent miRNAs. The center of the network was represented by degree. The term degree refers to the contribution of one miRNA to the GOs around or the contribution one GO to the miRNAs around. There were just a few predicted target genes for hsa-miR-3960, the miRNA therefore was absent in the network. Journal of Vascular Surgery  , e3DOI: ( /j.jvs ) Copyright © 2016 Society for Vascular Surgery Terms and Conditions

5 Fig 4 The pathway network. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment was performed by mapping the predicted target genes from the microRNAs (miRNAs) to the KEGG databases. By enrichment analyses on the targets predicted from the significantly validated most differentially expressed miRNAs, we determined that the target genes could be categorized into many regulatory pathways. Enrichment ranking of KEGG pathways include mitogen-activated protein kinase (MAPK) signaling pathway, Wnt signaling pathway, phosphatidylinositol 3-kinase-Akt signaling pathway, gap junction, focal adhesion, p53 signaling pathway, apoptosis, and pathways in cancer, among others. The red circle represents pathways. The relationship of the pathway is represented by an arrow. The center of the network is represented by degree. The term degree refers to the number of interaction relationship of pathways. The pathway network could summarize the pathways interaction of differential expression genes under diseases. Journal of Vascular Surgery  , e3DOI: ( /j.jvs ) Copyright © 2016 Society for Vascular Surgery Terms and Conditions

6 Fig 5 Validation for the expression of c-Jun NH2-terminal kinase (JNK2), extracellular signal-regulated kinase 5 (ERK5), growth-factor receptor-bound protein 2 (GRB2), and protein phosphatase 3 calcineurin A-α (PPP3CA) in the vessels by Western blot analysis. Representative blots are shown in A, C, E, and G. Bands were quantified by densitometric analysis, and the results are shown as relative density compared with control in B, D, F, and H. Values expressed as mean ± standard deviation from three independent experiments. *P < .05 indicates statistically significant difference between the two groups. JNK2, ERK5, GRB2, and PPP3CA were overexpressed in the stenotic arteriovenous fistula (AVF) group compared with control veins. Journal of Vascular Surgery  , e3DOI: ( /j.jvs ) Copyright © 2016 Society for Vascular Surgery Terms and Conditions

7 Fig 6 Validation for the expression of c-Jun NH2-terminal kinase (JNK2), extracellular signal-regulated kinase 5 (ERK5), growth-factor receptor-bound protein 2 (GRB2), and protein phosphatase 3 calcineurin A-α (PPP3CA) in the vessels by immunohistochemistry analysis. Representative photomicrographs are shown at original magnification ×200. The brown color staining represents cellular JNK2, ERK5, GRB2, and PPP3CA expression. Enhanced JNK2, ERK5, GRB2, and PPP3CA staining was observed in the stenotic AVF group. Journal of Vascular Surgery  , e3DOI: ( /j.jvs ) Copyright © 2016 Society for Vascular Surgery Terms and Conditions

8 Supplementary Fig 1 (online only)
The micro RNA (miRNA)-gene ontology (GO) network. The circles represent genes and the squares represent miRNAs. Their relationship is represented by one edge. The center of the network represents the degree, the interaction of one miRNA with the GOs around and the interaction of one GO with the miRNAs around. The key miRNAs and genes in the network always have the highest degrees. There were just a few predicted target genes for hsa-miR-3960, the miRNA therefore was absent in the network. Journal of Vascular Surgery  , e3DOI: ( /j.jvs ) Copyright © 2016 Society for Vascular Surgery Terms and Conditions

9 Supplementary Fig 2 (online only)
Expression of c-Jun NH2-terminal kinase (JNK2), extracellular signal-regulated kinase 5 (ERK5), growth-factor receptor-bound protein 2 (GRB2), and protein phosphatase 3 calcineurin A-α (PPP3CA) in the vessels by real-time polymerase chain reaction (PCR). Values are expressed as mean ± standard deviation from three independent experiments. *P < .05 indicates statistically significant difference between the two groups. In the stenotic arteriovenous fistula (AVF) group compared with controls, the messenger RNA (mRNA) expression of JNK2, ERK5, GRB2, and PPP3CA was significantly increased. Journal of Vascular Surgery  , e3DOI: ( /j.jvs ) Copyright © 2016 Society for Vascular Surgery Terms and Conditions

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