Fig 1: Network construction of lncRNA-miRNA-mRNA. A The mRNA-miRNA network was constructed by using the miRWalk website (The five miRNAs most associated with the common hub gene are highlighted in light blue). B Meaningful signal pathways about the 5 miRNAs by utilizing the DIANA-miRPath. C The lncRNA–miRNA–mRNA regulatory network (The orange nodes represent the lncRNAs, the yellow nodes represent key miRNAs, and the red nodes represent the hub genes). RRM2 ribonucleotide reductase regulatory subunit M2; AURKB aurora kinase B; CDK1 cyclin dependent kinase 1; TIMP1 tissue inhibitor of metalloproteinases 1
Fig 2: The PPI network and Hub genes analysis. A PPI network of all DEGs (The red codes represent upregulated genes in the elderly, the blue codes represent downregulated genes in the elderly, and the size of the nodes represents the correlation between them). Hub gene networks obtained from the PPI network relying on B the Maximal Clique Centrality (MCC) algorithm, C the Maximum Neighborhood Component (MNC) algorithm, D the Degree algorithm, and E the Edge Percolated Component (EPC) algorithm of the Cytoscape plugin cytoHubba. F The Venn diagram showed the identified hub genes (RRM2, AURKB, CDK1, and TIMP1). G The heatmap of common hub genes identified by Metascape was colored by –log10 (P-value). PPI protein–protein interaction; DEGs differentially expressed genes; RRM2 ribonucleotide reductase regulatory subunit M2; AURKB aurora kinase B; CDK1 cyclin dependent kinase 1; TIMP1 tissue inhibitor of metalloproteinases 1
Fig 3: The expressions of Hub genes in the mouse meniscus cells. A SA-ß-gal staining of mouse senescent meniscus cells induced by IL-1ß (10 ng/ml) for 48 h. Scale bar: 500 µm. B, D Apoptotic analysis detected by Annexin V/7AAD after IL-1ß disposition. C Detection of the expressions of hub genes in senescent meniscus cells via qRT-PCR. We normalized the gene expression levels to GAPDH. E Immunofluorescence analysis of hub genes after treating meniscus cells with10ng/ml IL-1ß for 48 h. Scale bars, 50 µm. RRM2 ribonucleotide reductase regulatory subunit M2; AURKB aurora kinase B; CDK1 cyclin dependent kinase 1; TIMP1 tissue inhibitor of metalloproteinases 1; GAPDH Glyceraldehyde 3-phosphate dehydrogenase. n = 6 per group, mean ± S.E.M., *P < 0.05, **P < 0.01 in comparison with the control group
Fig 4: The expression levels of Hub genes in the meniscus tissues. A SafraninO-Fast green and Toloniumchloride Blue staining results of meniscus in 18-month-old mice (aging) and 3-month-old mice (young). Scale bar: 250 µm. Scale bar (enlarged): 100 µm. B Hub genes immunochemical staining of meniscus in young mice and older mice. Scale bar: 100 µm. C Quantification of Hub genes immunostaining (mean optical density). D Immunostaining of Hub genes in young and gerontic human menisci. Scale bar: 100 µm. E Quantification of Hub genes immunostaining (mean optical density). F The mRNA expression levels of Hub genes in human and mouse menisci, respectively. We normalized the gene expression levels to GAPDH. RRM2 ribonucleotide reductase regulatory subunit M2; AURKB aurora kinase B; CDK1 cyclin dependent kinase 1; TIMP1 tissue inhibitor of metalloproteinases 1; GAPDH Glyceraldehyde 3-phosphate dehydrogenase. n = 6 per group, mean ± S.E.M., *P < 0.05 compared with the young group
Fig 5: Human glycoproteomic analysis showed that ApoA1, IL-17E, TIMP-1, and haptoglobin all signaled primarily through cytokine-cytokine receptor signaling after MI. A Protein–protein interactions among each identified plasma marker and the 999 other glycoproteins measured in the array showed enrichment of FOXM1 (ApoA1), STAT1 (IL-17E), STAT3 (haptoglobin), and USF2 (TIMP-1) transcription factors. B Cytokine-cytokine receptor signaling was the most enriched pathway as shown in the bubble chart
Supplier Page from Abcam for Anti-TIMP1 antibody