019) (Supplemental Figures S3A,B).LCN2 and Adenosine A2A receptor (A2AR) Gene ID RAB27B Could Act as Biomarkers in Meniscus for OA Severity Predictors and Early OA DiagnosisWe additional examined no matter if meniscus-specific LCN2 and RAB27B possess the prospective of predicting OA severity. qRTPCR H2 Receptor Compound confirmed LCN2 and RAB27B expression patterns, both of which show substantial upregulation in OA degenerative menisci, when also time-dependently upregulated in inflammatory chemokine-stimulated menisci (Figures 5I, 6A,B). Interestingly, LCN2 and RAB27B expression showed robust correlation with patients’ OA severity determined by OARSI Osteoarthritis Cartilage Histopathology Assessment Technique (Waldstein et al., 2016; Figure 6C). LCN2 and RAB27B had been also examined in spontaneous aging C57BL/6J mouse model to validate if meniscus-specific LCN2 and RAB27B could act as biomarkers for early-stage OA. Both of them had been found to become substantially upregulated at the age of 26 weeks, which is about 40 years old in human lifespan (Figure 6D). This suggests that LCN2 and RAB27B may possibly be possible diagnostic biomarkers in meniscus for OA severity prediction and early-stage OA diagnosis.qRT-PCR Validations in Typical and Degenerative Menisci and Screening for Possible Diagnostic Messenger RNA Biomarkers within the Menisci Throughout Early-Stage OsteoarthritisTo further confirm the results of whole-transcriptome sequencing, we chosen prior qRT-PCR-verified DEMs, differentially expressed lncRNAs (DELs), and DECs to validate their expression patterns between regular and degenerative menisci by qRT-PCR (Figure 5A). Consequently, three out of 5 miRNAs, five out of six lncRNAs, and 4 out of six circRNAs had been shown to have concurrent expression trends with all the sequencing final results and qRT-PCR validation among the OA menisci with and devoid of IL-1 stimulation (Figures 5B ). Moreover, the lncRNA LOC107986251-miR-212-5pSESN3 network and hsa_circ_0018069-miR-147b-3p-TJP2 network had been also confirmed (Figures 5E,F). Interestingly,DISCUSSIONWhole-transcriptome sequencing is often a novel bioinformatics evaluation strategy to test the differential expression levels of mRNA, miRNA, lncRNA, and circRNA among regular and pathological tissues. This technique has currently been widely applied in the field of oncology (Zheng et al., 2016). A current study illustrated the comprehensive transcriptome map of typical and OA cartilage and identified four DELs and six DEGs targeted by lncRNAs throughout OA (Li et al., 2019). Prospective OA-associated genes, pathways, competing endogenous RNA networks, and co-expression networks in knee cartilage had been additional identified in lately studies, thereby supplying a superior understanding of OA mechanism (Chen and Chen, 2020; Qi et al., 2020). However, a complete analysis of the expression patterns of mRNA, miRNA, lncRNA, andFrontiers in Genetics | frontiersin.orgOctober 2021 | Volume 12 | ArticleJiang et al.Osteoarthrititc Meniscus Expression ProfilesFIGURE 3 | Differential expression profile of long noncoding RNA (lncRNA) and lncRNA LOC107986251 ceRNA network prediction. (A) Hierarchical clustering illustrates distinguished expression difference of lncRNA amongst the two groups and homogeneity between groups. (B) Volcano plots of differentially expressed lncRNAs. (C) Scatter plots of differentially expressed lncRNAs. (D) Relative expression level of selected lncRNAs in unfavorable manage versus IL-1-treated osteoarthritis (OA) menisci. GAPDH was made use of as the internal reference gene for qRT-PC