Background: The MDCK cell line is the primary cell line used for influenza vaccine production. Using genetic engineering technology to change the expression and activity of genes that regulate virus proliferation to obtain high-yield vaccine cell lines has attracted increasing attention. A comprehensive understanding of the key genes, targets, and molecular mechanisms of viral regulation in cells is critical to achieving this goal, yet the post-transcriptional regulation mechanism involved in virus proliferation—particularly the effect of lncRNA on influenza virus proliferation—is still poorly understood. Therefore, this study used high-throughput RNA-seq technology to identify H1N1 infection-induced lncRNA and mRNA expression changes in MDCK cells and explore the regulatory relationship between these crucial lncRNAs and their target genes. Results: In response to H1N1 infection in MDCK cells 16 h post-infection (hpi) relative to uninfected controls, we used multiple gene function annotation databases and initially identified 31,501 significantly differentially expressed (DE) genes and 39,920 DE lncRNAs (|log2FC| > 1, p < 0.05). Among these, 102 lncRNAs and 577 mRNAs exhibited predicted correlations with viral response mechanisms. Based on the magnitude of significant expression differences, related research, and RT-qPCR expression validation at the transcriptional level, we further focused on 18 DE mRNAs and 32 DE lncRNAs. Among these, the differential expression of the genes RSAD2, CLDN1, HCLS1, and IFIT5 in response to influenza virus infection was further verified at the protein level using Western blot technology, which showed results consistent with the RNA-seq and RT-qPCR findings. We then developed a potential molecular regulatory network between these four genes and their six predicted lncRNAs. Conclusions: The results of this study will contribute to a more comprehensive understanding of the molecular mechanism of host cell non-coding RNA-mediated regulation of influenza virus replication. These results may also identify methods for screening target genes in the development of genetically engineered cell lines capable of high-yield artificial vaccine production.

The aim of this study was to investigate the effects of hypoxia-induced phenotype, glucose metabolism, ROS levels, and the PDK1-mediated regulation of TGF-β/Smad signaling in yellow cattles, yaks, and those overexpressing PDK1 PASMCs using growth curves, flow cytometry, scratch experiments, glucose and lactic acid assays, RT-qPCR, and Western blotting. The results showed that hypoxia significantly promoted proliferation, migration, antiapoptosis, ROS levels, glucose consumption, and lactate production in yellow cattle PASMCs (p < 0.05), and the cells were dedifferentiated from the contractile phenotype; conversely, hypoxia had no significant effect on yak PASMCs (p > 0.05). PDK1 overexpression significantly promoted proliferation, antiapoptosis, glucose consumption, and lactate production in yak PASMCs under normoxia and hypoxia (p < 0.05), decreased their migration levels under hypoxia (p < 0.05), and dedifferentiated the contractile phenotype of the cells. Overexpression of PDK1 in yak PASMCs is detrimental to their adaptation to hypoxic environments. Yak PASMCs adapted to the effects of hypoxia on lung tissue by downregulating the expression of genes related to the PDK1 and TGF-β/Smad signaling pathways. Taken together, the regulation of PDK1-mediated TGF-β/Smad signaling may be involved in the process of yaks’ adaptation to the hypoxic environment of the plateau, reflecting the good adaptive ability of yaks. The present study provides basic information to further elucidate the mechanism of PDK1-mediated TGF-β/Smad signaling induced by hypoxia in the lungs of yaks, as well as target genes for the treatment of plateau diseases in humans and animals.

Background: The Madin–Darby canine kidney (MDCK) cell line constitutes a key component of influenza vaccine production, but its dependence on adherent growth limits cell culture density and hinders vaccine yield. There is evidence that the use of gene editing techniques to inhibit cell adhesion and establish an easily suspended cell line can improve vaccine yield; however, the mechanisms underlying MDCK cell adhesion are unclear. Methods: In this study, we used transcriptomics to analyse differentially expressed mRNAs and miRNAs in adherent and suspension cultures of MDCK cells. Results: We found that claudin-1 (CLDN1) expression was downregulated in the suspension MDCK cells and that CLDN1 promotes MDCK cell–extracellular matrix adhesion. Additionally, microRNA (miR)-175 expression was upregulated in the suspension MDCK cells. Importantly, we demonstrated that miR-175 inhibits MDCK cell adhesion by targeting the CLDN1 3′-untranslated region (UTR). These findings contribute to a more comprehensive understanding of the regulatory mechanisms modulating cell adhesion and provide a basis for establishing suspension-adapted, genetically engineered cell lines. Our work could also facilitate the identification of targets for tumour therapy.