Document Type : Research Paper
Author
Department of Animal Science, Faculty of Agriculture, Yasouj University, Yasouj, Iran
Abstract
Keywords
Main Subjects
Extended Abstract
Introduction
Mastitis is one of the most prevalent and economically significant diseases affecting dairy cattle worldwide. It is characterized by inflammation of the mammary gland, which adversely impacts milk production and quality, compromises animal health, and leads to substantial financial losses within the dairy industry. The disease is primarily caused by bacterial infections; however, its pathogenesis is complex and involves intricate immune and inflammatory responses. Early diagnosis and timely management of mastitis are crucial for controlling disease spread, minimizing tissue damage, and improving recovery rates. Traditional diagnostic methods, such as somatic cell count and microbial culture, although useful, have limitations including time lags and variable sensitivity. Therefore, the exploration of molecular biomarkers that can enable rapid and sensitive detection of mastitis is essential. MicroRNAs (miRNAs), a class of small non-coding RNA molecules approximately 20-22 nucleotides in length, have emerged as pivotal post-transcriptional regulators of gene expression. By binding to complementary sequences on target messenger RNAs (mRNAs), miRNAs can inhibit translation or promote mRNA degradation, thereby finely tuning various biological processes, including immune responses and inflammation. This study employed an in silico bioinformatics approach aimed at investigating the regulatory roles of specific miRNAs in the pathogenesis of bovine mastitis. The objective was to identify key miRNAs associated with mastitis and their corresponding target genes, and to explore the biological pathways they influence using reliable public databases and computational tools such as miRTarBase, TargetScan, DAVID, and NCBI.
Method
The study leveraged a comprehensive bioinformatics pipeline to identify and analyze miRNAs and their target genes associated with bovine mastitis. Candidate miRNAs were initially selected based on literature reports and database screening for their known involvement in inflammatory and immune-related pathways pertinent to mastitis. Five key miRNAs—bta-mir-146a, bta-mir-16a, bta-mir-181, bta-mir-21-5p, and bta-mir-223—were prioritized due to their strong association with immune regulation and prior evidence of differential expression in mastitis-affected tissues. Target genes for these miRNAs were extracted using experimentally validated interactions in miRTarBase and predicted targets from TargetScan. Subsequent functional annotation and pathway enrichment analyses of these target genes were conducted using DAVID and related bioinformatics tools to reveal their involvement in signaling cascades critical to mastitis pathology. Special emphasis was placed on pathways such as Toll-like receptor 4 (TLR4)/nuclear factor kappa B (NF-κB), mitogen-activated protein kinase (MAPK), phosphatidylinositol 3-kinase/protein kinase B (PI3K-AKT), transforming growth factor beta (TGFβ), and hormonal signaling pathways involving estrogen and progesterone. Additionally, the study assessed the potential application of these miRNAs as non-invasive biomarkers by reviewing their detectability and stability in milk and blood serum samples.
Results
The results of these analyses identified five key miRNAs, including bta-mir-146a, bta-mir-16a, bta-mir-181, bta-mir-21-5p, and bta-mir-223, each of which plays a role in critical biological pathways such as TLR4/NF-κB, MAPK, PI3K-AKT, TGFβ, and estrogen and progesterone hormone signaling. Furthermore, the findings—based on a context++ score ≤ –0.2 and conservation ≥ 0.7—revealed that bta-mir-146a, bta-mir-21-5p, and bta-mir-223 influence 7, 9, and 11 key proteins, respectively, within these biological pathways. For instance, bta-mir-146a, bta-mir-21-5p, and bta-mir-223 were found to regulate a substantial number of key proteins—7, 9, and 11 respectively—that play essential roles in TLR4/NF-κB and MAPK signaling, pivotal pathways that mediate innate immune activation and inflammatory responses during mastitis. The PI3K-AKT pathway, implicated in cell survival and proliferation, along with the TGFβ signaling pathway, which modulates immune tolerance and tissue repair, were also influenced by these miRNAs. Moreover, hormonal pathways involving estrogen and progesterone, known for their immunomodulatory effects in the mammary gland, were shown to be modulated by these miRNAs, suggesting their broader involvement in mastitis pathogenesis. The multifunctional roles of these miRNAs underline their importance as molecular switches orchestrating responses to pathogenic challenge. Importantly, their stability and presence in easily accessible biological fluids such as milk and serum make them attractive candidates for development as sensitive and specific biomarkers. Early detection through miRNA profiling could enable timely interventions, reducing the severity of infection and associated economic losses. These findings also open avenues for therapeutic strategies targeting miRNA expression to modulate inflammatory and immune responses in mastitis.
Conclusion
The present study underscores the pivotal role of miRNAs in the complex pathophysiology of bovine mastitis. The identification of five key miRNAs—bta-mir-146a, bta-mir-16a, bta-mir-181, bta-mir-21-5p, and bta-mir-223—and their involvement in crucial signaling pathways provides valuable insights into the molecular mechanisms driving mastitis development. Their detectability in milk and blood serum supports their potential utility as non-invasive biomarkers for early disease diagnosis and health monitoring in dairy cattle. Furthermore, understanding miRNA-mediated regulation offers promising perspectives for the design of targeted therapeutic interventions aimed at mitigating inflammatory damage and enhancing host defense mechanisms. Ultimately, applying these findings in clinical settings could improve animal welfare, sustain milk production, and alleviate the economic burdens posed by mastitis in dairy farming.
All authors contributed equally to the conceptualization of the article and writing of the original and subsequent drafts.
I would like to extend my heartfelt appreciation to Zahra Beiranvand for her invaluable contributions to our research efforts. Her expertise and dedication have significantly enhanced our work.
In this study, we did not utilize any biological samples, as we relied solely on data deposited in biological databases.
The authors declare no conflict of interest.
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