The core clock gene Period2 (PER2) is associated with mammary gland development and lipid synthesis in rodents and has recently been found to have a diurnal variation in the process of lactation, but has not yet been demonstrated in bovine mammary epithelial cells (BMECs). To explore the regulatory function of PER2 on milk fat synthesis in bovine mammary epithelial cells, we initially assessed the expression of clock genes and milk fat metabolism genes for 24 h using real-time quantitative PCR and fitted the data to a cosine function curve. Subsequently, we overexpressed the PER2 in BMECs using plasmid vector (pcDNA3.1-PER2), with empty vector pcDNA3.1-myc as the control. After transfecting BMECs for 48 h, we assessed the protein abundance related to milk fat synthesis by Western blot, the expression of genes coding for these proteins using real time-quantitative PCR, the production of triacylglycerol, and the fatty acid profile. The findings indicated that a total of nine clock genes (PER1/2, CRY1/2, REV-ERBα, BMAL1, NCOR1, NR2F2, FBXW11), seven fatty acid metabolism genes (CD36, ACSS2, ACACA, SCD, FADS1, DGAT1, ADFP), and six nuclear receptor-related genes (INSIG1, SCAP, SREBF1, C/EBP, PPARG, LXR) exhibited oscillation with a period close to 24 h in non-transfected BMECs (R² ≥ 0.7). Compared to the control group (transfected with empty pcDNA3.1-myc), the triglyceride content significantly increased in the PER2 overexpression group (p < 0.05). The lipogenic genes for fatty acid transport and triglyceride synthesis (ACACA, SCD, LPIN1, DGAT1, and SREBF1) were upregulated after PER2 overexpression, along with the upregulation of related protein abundance (p < 0.05). The contents and ratios of palmitic acid (C16:0), oleic acid (C18:1n9c), and trans-oleic acid (C18:1n9t) were significantly increased in the overexpression group (p < 0.05). Overall, the data supported that PER2 participated in the process of milk fat metabolism and is potentially involved in the de novo synthesis and desaturation of fatty acid in bovine mammary epithelial cells.

Silage is the most important component of a ruminant diet and has important production and health significance in ruminant production. The aim of the research was to investigate how the mixed silage of Chinese cabbage waste and rice straw (mixed silage) impacts the fecal microorganisms and metabolites in Hu sheep using Illumina sequencing and metabolomic analysis. A total of 16 Hu sheep (8 rams and 8 ewes) weighing about 39 kg and 5.5 months old were used as experimental sheep and divided into two groups (4 rams and 4 ewes, n = 8) using the principle of randomized trials: the control group with peanut sprouts, corn husks, and sorghum husks as roughage and the silage group with the mixed silage as roughage. There were no significant differences in the average daily gain (ADG), dry matter intake (DMI), or feed conversion rate (FCR) between the control group and the mixed silage groups (p > 0.05). Microbiome results showed that 15 microorganisms such as Ruminococcaceae UCG 010, Breznakia, Erysipelothrix, Desulfovibrio, Succiniclasticum, and Shuttleworthia were significantly different between the two groups. In addition, metabolomics showed that the mixed silage modulated the concentrations and metabolic pathways of metabolites in the manure. Significantly different metabolites were mainly enriched in amino acid anabolism (“glycine, serine, and threonine metabolism”, “valine, leucine, and isoleucine biosynthesis”, “arginine biosynthesis”, etc.), nucleic acid metabolism (pyrimidine metabolism). In conclusion, the addition of mixed silage to the diet of Hu sheep can alter the structure of the hindgut microflora and regulate the metabolism of amino acids and nucleotides, which affects health performance.

Improper disposal of vegetable waste can cause serious environmental pollution, but because they contain huge water content and organic matter, they are not suitable for disposal by methods such as incineration and landfill. However, vegetable waste contains a large amount of nutrients and have some complementary effects with rice straw in terms of physical structure, nutrients, and moisture. In this experiment, the plant feed (corn husk, peanut shells and sorghum shells) was used as the control group (CON group), and the mixed silage of Chinese cabbage waste and rice straw (mixed silage) was used as the experiment group (TRE group), and its safety performance was evaluated by testing its toxin content, pesticide residues, vitamin contents and feeding experiment of Hu sheep. In the animal experiment, 16 healthy Hu sheep (5.5 months, 39.11 ± 4.16 kg) were randomly divided into two groups of 8 each. The results of the safety performance evaluation showed that the content of mycotoxins, heavy metals, and nitrites as well as pesticide residues in the crude feeds of both groups were within the range of Chinese feed hygiene standards. In addition, the levels of deoxynivalenol (DON) and aflatoxin (AFT) in the CON group were lower, while the content of ochratoxin (OTA) and zearalenone were higher than those in the TRE group (p < 0.05). The levels of plumbum(Pb), chromium (Cr), cadmium (Cd), and nitrite in the CON group were lower than the mixed silage, while the levels of As were higher than the mixed silage (p < 0.05). It is worth noticing that the content of vitamin B₂ (VB₂) and vitamin C (VC) in the TRE group was higher than the CON group (p < 0.05). The results of the feeding experiment showed that the mixed silage did not affect the growth performance, nutrient digestibility, organ index, and intestinal index of Hu sheep (p > 0.05). In addition, the mixed silage reduced the weight of omasum, the proportion of omasum to live weight before slaughter, the amount of compound stomach, and the proportion of compound stomach to live weight before slaughter, which were higher than those in the TRE group (p < 0.05). The thickness of the basal layer of the rumen abdominal sac, the red blood cell count, the content of IL-10, and TNF-α in the blood, and TNF-α content in the rumen of the Hu sheep in the TRE group were higher than the CON group (p < 0.05). In conclusion, the feed safety index content of the mixed silage did not exceed the Chinese feed hygiene and safety standards and did not cause adverse effects on the growth performance of the Hu sheep, and it improved the immune performance of the body and digestive tract of the sheep to a certain extent and promoted the healthy development of the sheep.

Glucose and amino acids are important sources of nutrients in the synthetic milk of dairy cows, and understanding the fate of amino acids is essential to optimize the utilization of amino acids in milk protein synthesis, thereby reducing nutrient inefficiencies during lactation. The purpose of this study was to investigate the effects of LPS and different concentrations of glucose on (1) the expression of inflammatory factors and genes, (2) the glucose metabolism, and (3) amino acid utilization in BMECs. The results showed that there was an interaction (LPS × glucose, p < 0.05) between LPS and glucose content in the inflammatory cytokine genes (IL-6 and TNF-α) and the inflammatory regulatory genes (CXCL2, CXCL8, and CCL5). With the addition of LPS, the HG + LPS group caused downregulated (p < 0.05) expression of IL-6 and TNF-α, compared with the LG + LPS group. Interestingly, compared with the LG + LPS group, the HG + LPS group upregulated (p < 0.05) the expression of CXCL2, CXCL8, and CCL5. LPS supplementation increased (p = 0.056) the consumption of glucose and GLUT1 gene expression (p < 0.05) and tended to increase (p = 0.084) the LDHA gene expression of BMECs under conditions of different concentrations of glucose culture. High glucose content increased (p < 0.001) the consumption of glucose and enhanced (p < 0.05) the GLUT1, HK1, HK2, and LDHA gene expression of BMECs with or without LPS incubation, and there was an interaction (LPS × glucose, p < 0.05) between LPS and glucose concentrations in GLUT1 gene expression. In this study, LPS enhanced (p < 0.05) the consumption of amino acids such as tryptophan, leucine, isoleucine, methionine, valine, histidine, and glutamate, while high levels of glucose decreased (p < 0.01) consumption, except in the case of tyrosine. For histidine, leucine, isoleucine, and valine consumption, there was an interaction (LPS × glucose, p < 0.05) between LPS and glucose levels. Overall, these findings suggest that relatively high glucose concentrations may lessen the LPS-induced BMEC inflammatory response and reduce amino acid consumption, while low glucose concentrations may increase the demand for most amino acids through proinflammatory responses.

The gradual decline in feed resources for livestock needs alternate ways to ensure non-stop feed supply throughout the year. The objective of this study was to evaluate the impact of vegetable waste and rice straw silage (VTRS) on immune response, antioxidant status, and microbial changes in duodenum and caecum in Hu sheep. Eight healthy male Hu sheep were randomly distributed into control (fed farm roughage) and VTRS (fed vegetable waste silage) groups for 35 days. Results had shown that silage had less mycotoxin content (p < 0.05). The VTRS increased butyrate content in duodenal digesta, while acetate, butyrate, total volatile fatty acids (TVFA), and valerate were enhanced in caecal digesta (p < 0.05). The VTRS also increased amylase activity in duodenum and ileum tissues, along with GLUT2 and SGLT1 expressions. In serum, Interleukin-10 (IL-10) concentration and total antioxidant capacity (T-AOC) were increased while malondialdehyde (MDA) was decreased. An increase in T-AOC and GSH-Px activity was also observed, along with increased IL-6, immunoglobulin A (IgA), and catalase in duodenum tissue (p < 0.05). Prevotella was increased in the duodenum and caecum, with Prevotellacae UCG-001 and Christensenellacae R-7 group representing the VTRS group in the duodenum (p < 0.05). KEGG pathway prediction also indicated the enrichment of energy metabolism-related pathways. Significant microbes had shown a significant correlation with immune parameters. It can be concluded that vegetable waste silage has the ability to improve antioxidant status, enhance energy metabolism, and balance intestinal microbiota in Hu sheep.