Quercetin Alleviates Lipopolysaccharide-Induced Cell Damage and Inflammation via Regulation of the TLR4/NF-κB Pathway in Bovine Intestinal Epithelial Cells.

Acute diarrhoea and intestinal inflammation represent one of the most prevalent clinical disorders of milk production, resulting in enormous annual financial damage for the dairy sector. In the context of an unsatisfactory therapeutic effect of antibiotics, the natural products of plants have been the focus of research. Quercetin is an important flavonoid found in a variety of plants, including fruits and vegetables, and has strong anti-inflammatory effects, so it has received extensive attention as a potential anti-inflammatory antioxidant. However, the underlying basis of quercetin on inflammatory reactions and oxidative tension generated by lipopolysaccharide (LPS) in bovine intestinal epithelial cells (BIECs) is currently unexplained. This research aimed to determine the influence of quercetin on LPS-induced inflammatory reactions, oxidative tension, and the barrier role of BIECs. Our findings demonstrated that BIEC viability was significantly improved in LPS-treated BIEC with 80 µg/mL quercetin compared with the control group. Indicators of oxidative overload and genes involved in barrier role revealed that 80 µg/mL quercetin efficiently rescued BIECs from oxidative and barrier impairment triggered by 5 µg/mL LPS. In addition, the mRNA expression of pro-inflammatory cytokines TNF-α, IL-1ß, and IL-6, as well as chemokines CXCL2, CXCL5, CCL5, and CXCL8, was diminished in LPS-treated BIECs with 80 µg/mL quercetin compared with LPS alone. Furthermore, the mRNA expression of toll-like receptor 4 (TLR4), CD14, myeloid differential protein-2 (MD2), and myeloid differentiation primary response protein (MyD88) genes associated with the TLR4 signal mechanism was markedly reduced by the addition of quercetin to LPS-modulated BIECs, indicating that quercetin can suppress the TLR4 signal mechanism. We performed Western blotting on the NF-κB signalling mechanism and compared it with immunofluorescence to further corroborate this conclusion. The LPS treatment enhanced the proportions of p-IκBα/GAPDH and p-p65/GAPDH. Compared with the LPS-treated group, quercetin administration decreased the proportions of p-IκBα/GAPDH and p-p65/GAPDH. In addition, immunofluorescence demonstrated that quercetin greatly reduced the LPS-induced nuclear translocation of NF-κB p65 in BIECs. The benefits of quercetin on inflammatory reactions in LPS-induced BIECs may be a result of its capacity to inhibit the TLR4-mediated NF-κB signalling mechanism. These findings suggest that quercetin can be used as an anti-inflammatory reagent to treat intestinal inflammation induced by LPS release.

An Extract of Artemisia argyi Leaves Rich in Organic Acids and Flavonoids Promotes Growth in BALB/c Mice by Regulating Intestinal Flora.

In the context of global restrictions on the use of antibiotics, there has been increased research on natural plant-based ingredients as additives. It has been proved that many natural active ingredients contained in plants have positive effects on animal growth regulation. Artemisia argyi (A. argyi) is a traditional Chinese herbal medicine, and its extracts have been reported to have a variety of biological activities. Therefore, in order to explore the potential of the active extract of Artemisia argyi leaves (ALE) as a plant source additive, mice were fed with ALE at different concentrations for 60 days. Finally, the effects of ALE were evaluated by the growth indexes, blood indexes, and intestinal microflora changes of the mice. It was found that a medium concentration of ALE (150 mg/kg) could promote growth, and especially improved the feed efficiency of the mice. However, high concentrations of ALE (300 mg/kg) had some negative effects on the growth of mice, especially liver damage, which significantly increased AST and ALT levels in the blood. Therefore, the 150 mg/kg ALE treatment group was selected for 16S rDNA analysis. It was found that ALE could play a positive role by regulating the proportion of Bacteroidetes and Firmicutes in the intestinal tract. In particular, it can significantly up-regulate the quantities of Akkermansia and Bifidobacterium. These results suggest that ALE at appropriate concentrations can positively regulate animal growth.

Phloretin Protects Bovine Rumen Epithelial Cells from LPS-Induced Injury.

Lipopolysaccharide (LPS) is an endotoxin that induces immune and inflammatory responses in the rumen epithelium of dairy cows. It is well-known that flavonoid phloretin (PT) exhibits anti-oxidative, anti-inflammatory and antibacterial activity. The aim of this research was to explore whether PT could decrease LPS-induced damage to bovine rumen epithelial cells (BRECs) and its molecular mechanisms of potential protective efficacy. BRECs were pretreated with PT for 2 h and then stimulated with LPS for the assessment of various response indicators. The results showed that 100 µM PT had no significant effect on the viability of 10 µg/mL LPS-induced BRECs, and this dose was used in follow-up studies. The results showed that PT pre-relieved the decline in LPS-induced antioxidant indicators (T-AOC and GSH-PX). PT pretreatment resulted in decreased interleukin-1ß (IL-1ß), IL-6, IL-8, tumor necrosis factor-α (TNF-α) and chemokines (CCL2, CCL5, CCL20) expression. The underlying mechanisms explored reveal that PT may contribute to inflammatory responses by regulating Toll-like receptor 4 (TLR4), nuclear transcription factor-κB p65 (NF-κB p65), and ERK1/2 (p42/44) signaling pathways. Moreover, further studies found that LPS-induced BRECs showed decreased expression of claudin-related genes (ZO-1, Occludin); these were attenuated by pretreatment with PT. These results suggest that PT enhances the antioxidant properties of BRECs during inflammation, reduces gene expression of pro-inflammatory cytokines and chemokines, and enhances barrier function. Overall, the results suggest that PT (at least in vitro) offers some protective effect against LPS-induced ruminal epithelial inflammation. Further in vivo studies should be conducted to identify strategies for the prevention and amelioration of short acute rumen acidosis (SARA) in dairy cows using PT.

Effects of Water Extract from Artemisia argyi Leaves on LPS-Induced Mastitis in Mice.

In the context of the unsatisfactory therapeutic effect of antibiotics, the natural products of plants have become a research hotspot. Artemisia argyi (A. argyi) is known as a traditional medicine in China, and its extracts have been reported to have a variety of active functions, including anti-inflammatory. Therefore, after establishing the mouse mastitis model by lipopolysaccharide (LPS), the effects of A. argyi leaves extract (ALE) were evaluated by pathological morphology of the mammary gland tissue, gene expression, and serum oxidation index. Studies have shown that ALE has a restorative effect on LPS-induced mammary gland lesions and significantly down-regulated the rise of myeloperoxidase (MPO) induced by LPS stimulation. In addition, ALE played a positive role in LPS-induced oxidative imbalance by restoring the activities of glutathione peroxidase (GSH-PX) and superoxide dismutase (SOD) and preventing the increase in nitric oxide (NO) concentration caused by the over-activation of total nitric oxide synthase (T-NOS). Further analysis of gene expression in the mammary gland showed that ALE significantly down-regulated LPS-induced up-regulation of inflammatory factors IL6, TNFα, and IL1ß. ALE also regulated the expression of MyD88, a key gene for toll-like receptors (TLRs) signaling, which, in turn, regulated TLR2 and TLR4. The effect of ALE on iNOS expression was similar to the effect of T-NOS activity and NO content, which also played a positive role. The IκB gene is closely related to the NF-κB signaling pathway, and ALE was found to significantly alleviate the LPS-induced increase in IκB. All of these results indicated that ALE may be considered a potential active substance for mastitis.

Genomics Analysis of Bacillus megaterium 1259 as a Probiotic and Its Effects on Performance in Lactating Dairy Cows.

In this study, we isolated a novel bacterium, Bacillus megaterium 1259 (BM1259), from chicken manure. Whole-genome sequencing analysis showed that the BM1259 complete genome is composed of a 5,043,095 bp circular chromosome and three circular plasmids, and it encodes 5379 coding genes and 182 RNA genes. Among these genes, a series of nitrate assimilation-related genes and pathways were identified, implying a potential role of BM1259 in nitrate metabolism. In addition, 24 lactating Holstein dairy cows were randomly assigned to four groups that were fed a total mixed ration (TMR) diet only (C), a TMR diet supplemented with 5 g/day of BM1259 (T1), a TMR diet supplemented with 10 g/day of BM1259 (T2), or a TMR diet supplemented with 15 g/day of BM1259 (T3). The results showed that supplementing dairy cows with 15 g/day of BM1259 increased 4% fat-corrected milk production. The molar proportion of propionate (C3) was significantly higher in T2 than in C. The C2:C3 ratio of T3 was higher than those of C and T2. No negative effect of BM1259 on blood indicators was detected. This study demonstrates BM1259 can be applied as a potential probiotic to improve nitrogen utilization and milk production in lactating dairy cows.