Lactoferrin (LF), a member of the transferrin family, is widely present in mammalian milk and other secretions, exhibiting anti-inflammatory, antibacterial, and anti-infective properties. Although the biological functions of LF have been extensively studied, there are few reports on its effects and molecular mechanisms concerning bovine mastitis caused by bacterial infection. This study used bovine mammary epithelial cells (BMECs) cultured in vitro as the research model. An inflammatory injury model was established by stimulating BMECs with LPS to investigate whether LF at different concentrations (10, 50, 100, and 200 μg·mL⁻¹) could inhibit the inflammatory response before and after the onset of inflammation. The expression of inflammatory cytokines IL-1β, IL-6, IL-8, and TNF-α at both the gene and protein levels was detected using RT-qPCR and ELISA. Western blotting was employed to evaluate the phosphorylation levels in the inflammatory signaling pathways MAPK/P38/ERK and NF-κB/P65, while RT-qPCR was used to examine the impact on TLR4 receptor gene expression. The results display that pretreatment with LF prior to LPS-induced inflammation in BMECs reduced the expression of inflammatory cytokines IL-1β, IL-6, IL-8, and TNF-α at both the gene and protein levels (p < 0.05). LF also inhibited the phosphorylation of NF-κB/P65 and MAPK/P38/ERK signaling pathways and downregulated TLR4 receptor gene expression (p < 0.05). However, when LF was added after the onset of LPS-induced inflammation, inflammatory cytokine expression and phosphorylation levels in the NF-κB/P65 and MAPK/P38/ERK pathways remained elevated, along with high expression of the TLR4 receptor gene (p < 0.05). These findings show that LF can antagonize LPS-induced inflammatory responses in BMECs and reduce cytokine expression, exhibiting anti-inflammatory effects when administered before inflammation. Conversely, when LF is added post-inflammation, it appears to enhance cytokine expression, potentially promoting the recruitment of more cells or factors to resolve inflammation rapidly. Both effects are mediated through the TLR4 receptor and the NF-κB/P65 and MAPK/P38/ERK signaling pathways.

Spermatogenesis is a highly organized process by which undifferentiated spermatogonia self‐renew and differentiate into spermatocytes and spermatids. The entire developmental process from spermatogonia to sperm occurs within the seminiferous tubules. Spermatogenesis is supported by the close interaction of germ cells with Sertoli cells. In this study, testicular tissues were collected from Hu sheep at 8 timepoints after birth: 0, 30, 90, 180, 270, 360, 540, and 720 days. Immunofluorescence staining and histological analysis were used to explore the development of male germ cells and Sertoli cells in the Hu sheep testes at these timepoints. The changes in seminiferous tubule diameter and male germ cells in the Hu sheep testes at these different developmental stages were analyzed. Then, specific molecular markers were used to study the proliferation and differentiation of spermatogonia, the timepoint of spermatocyte appearance, and the maturation and proliferation of Sertoli cells in the seminiferous tubules. Finally, the formation of the blood–testes barrier was studied using antibodies against the main components of the blood–testes barrier, β‐catenin, and ZO‐1. These findings not only increased the understanding of the development of the Hu sheep testes, but also laid a solid theoretical foundation for Hu sheep breeding.

The number of vertebrae is a crucial economic trait that can significantly impact the carcass length and meat production in animals. However, our understanding of the quantitative trait loci (QTLs) and candidate genes associated with the vertebral number in sheep (Ovis aries) remains limited. To identify these candidate genes and QTLs, we collected 73 Ujimqin sheep with increased numbers of vertebrae (T13L7, T14L6, and T14L7) and 23 sheep with normal numbers of vertebrae (T13L6). Through high-throughput genome resequencing, we obtained a total of 24,130,801 effective single-nucleotide polymorphisms (SNPs). By conducting a selective-sweep analysis, we discovered that the most significantly selective region was located on chromosome 7. Within this region, we identified several genes, including VRTN, SYNDIG1L, LTBP2, and ABCD4, known to regulate the spinal development and morphology. Further, a genome-wide association study (GWAS) performed on sheep with increased and normal vertebral numbers confirmed that ABCD4 is a candidate gene for determining the number of vertebrae in sheep. Additionally, the most significant SNP on chromosome 7 was identified as a candidate QTL. Moreover, we detected two missense mutations in the ABCD4 gene; one of these mutations (Chr7: 89393414, C > T) at position 22 leads to the conversion of arginine (Arg) to glutamine (Gln), which is expected to negatively affect the protein’s function. Notably, a transcriptome expression profile in mouse embryonic development revealed that ABCD4 is highly expressed during the critical period of vertebral formation (4.5–7.5 days). Our study highlights ABCD4 as a potential major gene influencing the number of vertebrae in Ujimqin sheep, with promising prospects for future genome-assisted breeding improvements in sheep.

The aim of this study is to explore the mechanism of IL-17A infection in the development of bacterial mastitis in dairy cows. In this study, RT-qPCR and ELISA were used to measure the promoting effect of IL-17A on the generation of pro-inflammatory cytokines (TNF-α, IL-1β, and IL-6) and chemokine (IL-8). In addition, Western blot (WB) was applied to measure the influences of IL-17A on the inflammation-related ERK and p38 proteins in the MAPK pathways. The results show that under the stimulation of LPS on cow mammary epithelial cells (CMECs), cytokines IL-1β, IL-6, IL-8, TNF-α, and IL-17A will exhibit significantly increased expression levels (p < 0.05). With inhibited endogenous expression of IL-17A, cytokines IL-1β, IL-6, IL-8, and TNF-α will present reduced genetic expression (p < 0.01), with reduced phosphorylation levels of ERK and p38 proteins in the MAPK signaling pathways (p < 0.001). Upon the exogenous addition of the IL-17A cytokine, the genetic expression of cytokines IL-1β, IL-6, IL-8, and TNF-α will increase (p < 0.05), with increased phosphorylation levels of the ERK and p38 proteins in the MAPK signaling pathways (p < 0.001). These results indicate that under the stimulation of CMECs with LPS, IL-17A can be expressed together with relevant inflammatory cytokines. Meanwhile, the inflammatory responses of mammary epithelial cells are directly proportional to the expression levels of IL-17A inhibited alone or exogenously added. In summary, this study shows that IL-17A could be used as an important indicator for assessing the bacterial infections of mammary glands, indicating that IL-17A could be regarded as one potential therapeutic target for mastitis.

Dosage compensation is a mechanism first proposed by Susumu Ohno, whereby X inactivation balances X gene output between males (XY) and females (XX), while X upregulation balances X genes with autosomal gene output. These mechanisms have been actively studied in Drosophila and mice, but research regarding them lags behind in domestic species. It is unclear how the X chromosome is regulated in the sheep male germline. To address this, using single-cell RNA sequencing, we analyzed testes in three important developmental stages of sheep. We observed that the total RNA per cell from X and autosomes peaked in SSCs and spermatogonia and was then reduced in early spermatocytes. Furthermore, we counted the detected reads per gene in each cell type for X and autosomes. In cells experiencing dose compensation, close proximity to MSL (male-specific lethal), which is regulated the active X chromosome and was observed. Our results suggest that there is no dose compensation in the pre-meiotic germ cells of sheep testes and, in addition, MSL1 and MSL2 are expressed in early germ cells and involved in regulating mammalian X-chromosome inactivation and activation.

With the change in diet structure, individuals prefer to consume mutton with less fat. However, sheep tail has a lot of fat. We identified a breed of low-fat short-tailed sheep (i.e., Hulunbuir short-tailed sheep). It is necessary to develop an animal model that can promote research on the potential mechanisms of the short-tail phenotype in sheep, which results from the TBXT gene c.G334T mutation. To create animal models, we selected mice as experimental animals. Mouse embryos lacking the TBXT protein, which crucially regulates mouse embryonic development, cannot develop normally. We utilized CRISPR/Cas9 gene editing technology to generate site-specific mutation (c.G334T) in the TBXT gene of mice, and found that the mouse TBXT mutation (c.G334T) leads to a short-tail phenotype. Furthermore, we investigated the interaction between TBXT and Wnt signaling pathways. The expressions of TBXT, Axin2, Dkk1, Wnt3, Wnt3a, and Wnt5a were discovered to be significantly different between mutant embryos and wild embryos by obtaining mouse embryos at various developmental stages and examining the expression relationship between the TBXT and Wnt signaling pathway-related components in all of these embryos. Therefore, as a transcription factor, TBXT regulates the expression of the aforementioned Wnt signaling pathway components by forming a regulatory network for the normal development of mouse embryos. This study enriches the research on the functional role of the TBXT in the development of mouse embryos and the mechanism by which the short-tailed phenotype in sheep develops.