MiR-29a Family as a Key Regulator of Skeletal Muscle Dysplasia in a Porcine Model of Intrauterine Growth Retardation

Abstract

MicroRNAs (miRNAs) play an essential role in many biological processes. In this study, miRNAs in the skeletal muscle of normal and intrauterine growth retardation (IUGR) neonatal piglets were identified by sequencing, and canonical miRNAs were functionally validated in vitro. A total of 403 miRNAs were identified in neonatal piglet skeletal muscle, among them 30 and 46 miRNAs were upregulated and downregulated in IUGR pigs, respectively. Upregulated miRNAs were mainly enriched in propanoate metabolism, endocytosis, beta-Alanine metabolism, gap junction, and tumor necrosis factor signaling pathway. Down-regulated miRNAs were mainly enriched in chemical carcinogenesis—receptor activation, endocytosis, MAPK signaling pathway, insulin resistance, and EGFR tyrosine kinase inhibitor resistance. Co-expression network analysis of umbilical cord blood and skeletal muscle miRNAs showed that the miR-29 family is an essential regulator of IUGR pigs. The dual-luciferase reporter system showed that IGF1 and CCND1 were target genes of the miR-29 family. Transfection of IUGR pig umbilical cord blood exosomes and miR-29a mimic significantly inhibited cell proliferation and promoted the expression of cellular protein degradation marker genes Fbxo32 and Trim63. In summary, these results enrich the regulatory network of miRNAs involved in skeletal muscle development in IUGR animals.

Characteristics of microRNAs in Skeletal Muscle of Intrauterine Growth-Restricted Pigs

“…miR-1 and miR-206 were the two miRNAs with the highest expression levels, which accounted for more than 94%. These results are consistent with our previous reports [33]. Both miR-1 and miR-206 are members of the miR-1 family and share a common seed sequence.…”

“…Our published study showed that miR-29a was significantly upregulated in IUGR pig skeletal muscle. In C2C12 cells, overexpression of miR-29a can play a functional role in inhibiting myocyte proliferation by targeting the mRNA-level expression of IGF1 and CCND1 [33]. In adult muscle stem cells, miR-29a mediates fibroblast growth factor-2 (FGF-2) to promote myoblast proliferation, and inhibition of miR-29a expression showed the opposite effect [56].…”

“…The quantification of miRNA and mRNA was performed using the methods in our previous study [33]; the small RNA first strand was synthesized using the Mir-X™miRNA First Strand synthesis kit (Takara, Kusatsu, Japan). RT-qPCR was performed in the Bio-Rad CFX96 Real-Time PCR assay system using TB Green Premix Ex Taq II (Takara).…”

“…miRNAs are involved in the regulation of target gene expression at the post-transcriptional level by translational inhibition and mRNA destabilization [30,31], and miRNAs have become potential therapeutic and diagnostic targets for various diseases [32]. Studies on miRNA and IUGR also reported that the miR-29 family identified in skeletal muscle of IUGR pigs inhibits cell proliferation and promotes protein degradation by targeting IGF1 and CCND1 [33]. miR-1227-3p may regulate trophoblast cell proliferation and apoptosis by targeting genes involved in the insulin pathway [34].…”

Identification of tRNA-derived small RNAs and their potential roles in porcine skeletal muscle with intrauterine growth restriction

“…Further screening revealed that IGF1 is likely among tRF-Glu-TTC-047 target genes, with IGF1 at the core of IUGR porcine skeletal muscle regulatory network. IGF1 has also been reported to be involved in the regulation of skeletal muscle development and associated with the occurrence of IUGR ( Zhu et al, 2022 ). The binding relationship between the 3′ UTR of IGF1 and tRF-Glu-TTC-047 was further confirmed by a dual-luciferase reporter system.…”