Saif Adil Abbood Al-Janabi

MicroRNAs (miRNAs) act as key gene expression regulators, influencing intracellular biological and pathological processes. They are of significant interest in animal genetics as potential biomarkers for animal selection and health. This study aimed to unravel the complex miRNA signature involved in mastitis in in vitro cell culture. For this purpose, we constructed a control and treatment model in ovarian mammary epithelial cells to analyze miRNA responses upon Staphylococcus aureus (S. aureus) stimulation. The high-throughput Illumina Small RNA protocol was employed, generating an average of 7.75 million single-end reads per sample, totaling 46.54 million reads. Standard bioinformatics analysis, including cleaning, filtering, miRNA quantification, and differential expression was performed using the miRbase database as a reference for ovine miRNAs. The results indicated differential expression of 63 miRNAs, including 33 up-regulated and 30 down-regulated compared to the control group. Notably, miR-10a, miR-10b, miR-21, and miR-99a displayed a significant differential expression (p ≤ 0.05) associated to signal transduction, transcriptional pathways, diseases of signal transduction by growth factor receptors and second messengers, MAPK signaling pathway, NF-κB pathway, TNFα, Toll Like Receptor 4 (TLR4) cascade, and breast cancer. This study contributes expanding miRNA databases, especially for sheep miRNAs, and identifies potential miRNA candidates for further study in biomarker identification for mastitis resistance and diagnosis.

Alveolar macrophages (AMs) respond to these infections as the first line of defense and trigger the lung's immune response. Knowing the expression dynamics of key immune-related genes in alveolar macrophages against M. haemolytica challenge will help deepen the understanding of disease immunopathogenesis. For this purpose, first time an in vitro obtained from bronchoalveolar lavage (BAL) fluid and treated with three doses (T1: 1800, T2: 2700, T3: 5400 CFU) of M. haemolytica inoculum, while keeping one untreated as a control. Then, total RNA was extracted, and cDNA was synthesized. The real-time quantification data indicated that the expression of IL-1β, IL-6, IL-8, IL-10, and NF-κB genes were significantly changed as compared to control. Our study revealed that the exposure to M. haemolytica stimulates the immune response in the sheep alveolar macrophages in a dose and time-dependent manner.

The alveolar macrophages (AMs) are frontier of defense against foreign materials that initiate immune response in lungs. Knowledge of the expression dynamics of major immune-related genes in the alveolar macrophages in response to lipopolysaccharide (LPS) and lipoteichoic acid (LTA) challenge can help to understand disease mechanism involved in several respiratory diseases. The aim of this study was to investigate the mRNA expression of selected immune-related genes in response to lipopolysaccharide (LPS) and lipoteichoic acid (LTA) challenge in sheep alveolar macrophages in vivo. Results revealed that Romanov lambs exhibited higher mRNA expression of TLR2, TLR4, NF-ĸβ, TNFα, IL-1β, IL-6, IL-8, and IL-10 genes as compared to Akkaraman lambs along with the control of all treatments. Moreover, the expression of TLR2, TLR4, NF-ĸβ, TNFα, IL-1β, IL-6, IL-8, and IL-10 genes was higher in combine treatment of LPS and LTA as compared to separate treatments of LPS and LTA in both breeds. The results showed that the mRNA expression of immune-related genes was significantly increased in the sheep AMs in response to LPS and LTA treatment whereas a synergistic effect was observed in LPS+LTS treatment. Also, breed comparison showed that the native Akkaraman was more resistant to disease compared to exotic Romanov.