Understanding downstream signaling pathways regulated by miR-34a-5p in articular cartilage by RNA sequencing using miR-34a knock-out mice

The goal of this study was to determine the role of miR-34a in regulating chondrocyte transcript profiles. Next Generation Sequencing of total RNA extracted from mouse KO and WT chondrocytes revealed 175 significantly differentially expressed genes (84 up, 91 down) in miR-34a KO chondrocytes compared to WT cells. We are currently validating potential direct targets of miR-34a from our NGS data and performing computational pathway analysis to identify novel pathways regulated by miR-34a. Overall design: Heterozygous miR-34a KO mice were bred to generate global homozygous KOs. Articular cartilage was harvested and pooled from hips and knees, and enzymatically digested to isolate chondrocytes from 5 week-old homozygous miR-34a KO and WT mice. Chondrocytes were cultured in complete medium for approximately ten days. Total RNA was extracted from miR-34a KO or WT mouse chondrocytes followed by RNA quality assessment, sample library preparation, and library quality assessment. The prepared libraries were sequenced on the Illumina NextSeq 550 sequencer for 75 paired end read-cycles.