Nobiletin enhances mitochondrial function by regulating SIRT1/PGC-1α signaling in porcine oocytes during in vitro maturation.

Nobiletin is a natural flavonoid found in citrus fruits with beneficial effects, including anti-inflammatory, anti-cancer and anti-oxidation effects. The aim of this study was to investigate whether nobiletin improves mitochondrial function in porcine oocytes and examine the underlying mechanism. Oocytes enclosed by cumulus cells were cultured in TCM-199 for 44 h with 0.1% dimethyl sulfoxide (control), or supplemented with 5, 10, 25, and 50 µM of nobiletin (Nob5, Nob10, Nob25, and Nob50, respectively). Oocyte maturation rate was significantly enhanced in Nob10 (70.26 ± 0.45%) compared to the other groups (control: 60.12 ± 0.47%; Nob5: 59.44 ± 1.63%; Nob25: 63.15 ± 1.38%; Nob50: 46.57 ± 1.19%). The addition of nobiletin reduced the levels of reactive oxygen species and increased glutathione levels. Moreover, Nob10 promoted mitochondrial biogenesis by upregulating the protein levels of sirtuin 1 (SIRT1) and peroxisome proliferator-activated receptor-gamma coactivator 1α (PGC-1α). This resulted in an increase in the number of active mitochondria, mitochondrial DNA copy number, mitochondrial membrane potential, and ATP production, thereby enhancing mitochondrial function. The protein level of p53 decreased, followed by the phosphorylation of B-cell lymphoma 2, suggesting a reduction in mitochondria-mediated apoptosis in the Nob10 group. Additionally, the release of cytochrome c from the mitochondria was significantly diminished along with a decrease in the protein expression of caspase 3. Thus, nobiletin has a great potential to promote the in vitro maturation of porcine oocytes by suppressing oxidative stress and promoting mitochondrial function through the upregulation of the SIRT1/PGC-1α signaling pathway.

Molecular mechanisms in the miR-33a/LPPR4 pathway regulating unsaturated fatty acid synthesis in bovine mammary epithelial cells.

The regulatory mechanisms governing metabolism of fatty acids in cow mammary gland are crucial for establishing relationships between milk quality and fatty acid content. Both, microRNAs (miRNAs) and protein-coding genes are important factors involved in the regulation of milk fat synthesis. In this study, high-throughput sequencing of miRNAs and mRNAs in bovine mammary gland tissue was performed during peak lactation (3 samples) and late lactation (3 samples) periods to characterize expression profiles. Differential expression (DE) analyses of miRNA and mRNA and miRNA-mRNA regulatory pathway screening were performed. Two-hundred eighty regulatory miRNA-mRNA pairs were identified, including the miR-33a-lipid phosphate phosphatase-related protein type 4 (LPPR4) pathway. Bioinformatics prediction, dual-luciferase reporter system detection, qRT-PCR, and Western blotting revealed that miR-33a can directly target LPPR4 and inhibit its expression. Experiments also revealed that miR-33a promotes the synthesis of triglycerides and increases the content of unsaturated fatty acids (UFAs) in bovine mammary epithelial cells (BMECs). These results indicate that miR-33a via LPPR4 plays an important role in the regulation of milk fat synthesis and UFA levels.

Lead exposure promotes the inflammation via the circRNA-05280/miR-146a/IRAK1 axis in mammary gland.

Lead, the most widely used heavy metal in industry, is detrimental to human health if exposed to living and occupational environment. Although several studies have been conducted on lead exposure, little has been reported on its harm to mammary gland and its mechanisms. In view of this, our study is the first to verify that lead exposure could promote apoptosis and inflammation in mouse mammary tissue (in vivo) and cow mammary epithelial cells (in vitro). After establishing a lead exposed mouse model, the expression profile of mammary gland tissue was constructed by high-throughput sequencing technology. In the profile, 917 differentially expressed genes were screened, of which IRAK1 was up-regulated by 4.33 times. Then, from qRT-PCR, Western blot and Luciferase report, IRAK1 was found to promote the release of inflammatory factors and tissue apoptosis and could be a specific target of miR-146a. On the other hand, double luciferase reporter system and qRT-PCR predicted the existence of a binding site between circRNA-05280 and miR-146a sequence. Experiments such as immunohistochemistry, apoptosis and EdU demonstrated that circRNA-05280 could promote not only cell apoptosis but also the expression level of inflammatory genes. Nevertheless, the function of miR-146a is opposite to that of circRNA-05280. Specifically, circRNA-05280 can regulate the level of apoptosis and inflammation of mammary gland by binding miR-146a and releasing the expression of miR-146a on target gene IRAK1. This study concludes that circRNA-05280/ miR-146a/ IRAK1 signaling pathway could mediate the mammary gland damage resulting from lead exposure. Accordingly, it sheds new light on further exploration of molecular mechanisms of mammary gland tissue damage caused by lead exposure, the risk assessment of lead, and the mechanism of lead mammary gland toxicity.

Cadmium promotes apoptosis and inflammation via the circ08409/miR-133a/TGFB2 axis in bovine mammary epithelial cells and mouse mammary gland.

Cadmium is a common environmental heavy metal pollutant that can accumulate over long periods of time and cause disease. Thus, analysis of the molecular mechanisms affected by cadmium in the body could be of great significance for the prevention and treatment of cadmium-related diseases. In this study, flow cytometry, immunofluorescence, transmission electron microscopy, H&E (Hematoxylin Eosin) staining and TUNEL (TdT-mediated dUTP Nick-End Labeling) assays were used to verify that cadmium induced apoptosis and immune responses in bovine mammary epithelial cells (BMECs) and in mouse mammary gland. Isolated BMECs cultured with or without cadmium were collected to screen miRNA (microRNA) using high-throughput sequencing. There were 42 differentially-expressed miRNAs among which 27 were upregulated and 15 downregulated including bta-miR-133a, bta-miR-23b-5p, bta-miR-29e, bta-miR-365-5p, bta-miR-615, bta-miR-7, bta-miR-11975, bta-miR-127, and bta-miR-411a. Among those, miR-133a (which can specifically target TGFB2 (Recombinant Transforming Growth Factor Beta 2) was the most significantly downregulated with a fold-change of 5.27 in BMECs cultured with cadmium. Application of the double luciferase reporter system, western blotting, and qRT-PCR (Quantitative Real-time PCR) revealed that circ08409 can directly bind to miR-133a. Experiments demonstrated that circRNA-08409 could adsorb bta-miR-133a. Both circ08409 and TGFB2 significantly increased apoptosis and altered expression level of a series of inflammatory factors in BMECs. In contrast, miR-133a decreased significantly apoptosis and inflammation in the cells. Compared with cultures receiving only cadmium, the miR-133a+cadmium cultures exhibited significant reductions in the occurrence of late apoptosis. Overall, results indicated that circ08409 could relieve the inhibitory effect of miR-133a on TGFB2 expression by combining with miR-133a and subsequently modulating cell proliferation, apoptosis and inflammation. Overall, the data suggested that the circ08409/miR-133a/TGFB2 axis might play a role in mediating the effect of cadmium on BMECs. As such, data provide novel insights into controlling hazards that cadmium could induce in the mammary gland.

The cooperative regulatory effect of the miRNA-130 family on milk fat metabolism in dairy cows.

OBJECTIVE: There is a strong relationship between the content of beneficial fatty acids in milk and milk fat metabolic activity in the mammary gland. To improve milk quality, it is therefore necessary to study fatty acid metabolism in bovine mammary gland tissue. In adipose tissue, peroxisome proliferator-activated receptor gamma (PPARG), the core transcription factor, regulates the fatty acid metabolism gene network and determines fatty acid deposition. However, its regulatory effects on mammary gland fatty acid metabolism during lactation have rarely been reported. METHODS: Transcriptome sequencing was performed during the prelactation period and the peak lactation period to examine mRNA expression. The significant upregulation of PPARG drew our attention and led us to conduct further research. RESULTS: According to bioinformatics prediction, dual-luciferase reporter system detection, real-time quantitative reverse transcription polymerase chain reaction and Western blotting, miR-130a and miR-130b could directly target PPARG and inhibit its expression. Furthermore, triglyceride and oil red O staining proved that miR-130a and miR-130b inhibited milk fat metabolism in bovine mammary epithelial cells (BMECs), while PPARG promoted this metabolism. In addition, we also found that the coexpression of miR-130a and miR-130b significantly enhanced their ability to regulate milk fat metabolism. CONCLUSION: In conclusion, our findings indicated that miR-130a and miR-130b could target and repress PPARG and that they also have a functional superposition effect. miR-130a and miR-130b seem to synergistically regulate lipid catabolism via the control of PPARG in BMECs. In the long-term, these findings might be helpful in developing practical means to improve high-quality milk.

The function of omega-3 polyunsaturated fatty acids in response to cadmium exposure.

Throughout history, pollution has become a part of our daily life with the improvement of life quality and the advancement of industry and heavy industry. In recent years, the adverse effects of heavy metals, such as cadmium (Cd), on human health have been widely discussed, particularly on the immune system. Here, this review summarizes the available evidence on how Cd exposure may affect health. By analyzing the general manifestations of inflammation caused by Cd exposure, we find that the role of omega-3 (n-3) polyunsaturated fatty acids (PUFAs) in vivo can counteract Cd-induced harm. Additionally, we elucidate the effects of n-3 PUFAs on the immune system, and analyze their prophylactic and therapeutic effects on Cd exposure. Overall, this review highlights the role of n-3 PUFAs in the pathological changes induced by Cd exposure. Although n-3 PUFAs remain to be verified whether they can be used as therapeutic agents, as rehabilitation therapy, supplementation with n-3 PUFAs is reliable and effective.

Nanomaterials in Animal Husbandry: Research and Prospects.

Anti-inflammatory, antiviral, and anti-cancer treatments are potential applications of nanomaterials in biology. To explore the latest discoveries in nanotechnology, we reviewed the published literature, focusing on co-assembled nanoparticles for anti-inflammatory and anti-tumor properties, and their applications in animal husbandry. The results show that nanoparticles have significant anti-inflammation and anti-tumor effects, demonstrating broad application prospects in animal breeding. Furthermore, pooled evidence suggests that the mechanism is to have a positive impact on inflammation and tumors through the specific drug loading by indirectly or directly targeting the disease sites. Because the precise regulatory mechanism remains unclear, most studies have focused on regulating particular sites or even specific genes in the nucleus by targeting functional co-assembled nanoparticles. Hence, despite the intriguing scenarios for nanotechnology in farmed animals, most results cannot yet be translated into field applications. Overall, nanomaterials outperformed similar materials in terms of anti-inflammatory and anti-tumor. Nanotechnology also has promising applications in animal husbandry and veterinary care, and its application and development in animal husbandry remain an exciting area of research.

Circ007071 Inhibits Unsaturated Fatty Acid Synthesis by Interacting with miR-103-5p to Enhance PPARγ Expression in the Dairy Goat Mammary Gland.

Understanding more precisely the mechanisms controlling the metabolism of fatty acid in the mammary gland of dairy goats is essential for future improvements in milk quality. Particularly since recent data have underscored a key role for circular RNAs (circRNAs) in the mammary gland function, high-throughput sequencing technology was used to identify expression levels of circRNAs in the mammary tissue of dairy goats during early and peak lactation in the present study. Compared with early lactation, results demonstrated that the expression level of circ007071 during peak lactation was 12.02-fold up-regulated. Subsequent studies in goat mammary epithelial cells (GMECs) revealed that circ007071 stimulated the synthesis of triglycerides (TAG) and cholesterol, as well as increased the content of saturated fatty acids (C16:0 and C18:0). More importantly, using a double luciferase reporting system allowed us to detect the circ007071 sequence at a binding site of miR-103-5p, indicating that it targeted this miRNA. Overexpression of circ007071 significantly decreased the abundance of miR-103-5p and led to inhibition of TAG synthesis. In contrast, the abundance of peroxisome proliferator-activated receptor γ (PPARγ), a target gene of miR-103-5p, was reinforced with the overexpression of circ007071. Thus, we conclude that one key function of circ007071 in the regulation of milk fat synthesis is to attenuate the inhibitory effect of miR-103-5p on PPARγ via direct interactions with miRNA. As a result, the process of TAG and saturated fatty acid is able to proceed.

Lactoferrin: A glycoprotein that plays an active role in human health.

Lactoferrin (Lf), existing widely in human and mammalian milk, is a multifunctional glycoprotein with many functions, such as immune regulation, anti-inflammation, antibacterial, antiviral, and antioxidant. These extensive functions largely attribute to its ability to chelate iron and interfere with the cellular receptors of pathogenic microorganisms and their hosts. Moreover, it is non-toxic and has good compatibility with other supplements. Thus, Lf has been widely used in food nutrition, drug carriers, biotechnology, and feed development. Although Lf has been continuously explored and studied, a more comprehensive and systematic compendium is still required. This review presents the recent advances in the structure and physicochemical properties of Lf as well as clinical studies on human diseases, with the aim of providing a reference for further research of Lf and the development of its related functional products.

Emerging roles of noncoding micro RNAs and circular RNAs in bovine mastitis: Regulation, breeding, diagnosis, and therapy.

Bovine mastitis is one of the most troublesome and costly problems in the modern dairy industry, which is not only difficult to monitor, but can also cause economic losses while having significant implications on public health. However, efficacious preventative methods and therapy are still lacking. Moreover, new drugs and therapeutic targets are in increasing demand due to antibiotic restrictions. In recent years, noncoding RNAs have gained popularity as a topic in pathological and genetic studies. Meanwhile, there is growing evidence that they play a role in regulating various biological processes and developing novel treatment platforms. In light of this, this review focuses on two types of noncoding RNAs, micro RNAs and circular RNAs, and summarizes their characterizations, relationships, potential applications as selection markers, diagnostic or treatment targets and potential applications in RNA-based therapy, in order to shed new light on further research.

Poly(caffeic acid)-coated molecularly imprinted magnetic nanoparticles for specific and ultrasensitive detection of glycoprotein.

Molecularly imprinted polymers (MIPs) are artificial chemical receptors, and can recognize template molecules with a high selectivity and affinity. As "antibody mimics", MIPs have been widely studied in various fields. However, the general applicability of MIPs is limited by the type of functional monomers. Herein, we developed caffeic acid (CA, a natural polyphenol) as novel a functional monomer. An innovative poly(caffeic acid)-coated molecularly imprinted magnetic nanoparticles (PCA-MIMN) with transferrin (TRF) as a model glycoprotein template was fabricated by autoxidation of CA with hexamethylenediamine (HMDA) in an aerobic environment as imprinted layer. The successful fabrication of PCA-MIMN was proved in detail by diversified characterization. The PCA-MIMN exhibited not only outstanding binding affinity and specificity for target glycoprotein, but also excellent hydrophilicity due to the externally generous hydrophilic groups. To evaluate the preeminent performance, the PCA-MIMN was linked with pH-triggered allochroic-graphene oxide (AGO), which was used for determination of TRF in real samples. The proposed PCA-MIMN linked AGO strategy exhibited ultrahigh sensitivity with limit of detection of 0.38 pg mL-1 for TRF. Finally, the proposed strategy was successfully applied in determination of TRF in spiked human serum sample with recovery and relative standard deviation in the range of 97.2%-103.9% and 4.6%-5.8%, respectively. This work demonstrates that the "autoxidation of CA with HMDA" may be a universal tool for synthesis of highly specific MIPs, and the type of functional monomers will increase exponentially due to the presence of numerous polyphenols in nature.

Progress on the Regulation of Ruminant Milk Fat by Noncoding RNAs and ceRNAs.

Milk fat is not only a key factor affecting the quality of fresh milk but also a major target trait forbreeding. The regulation of milk fat involves multiple genes, network regulation and signal transduction. To explore recent discoveries of pathway regulation, we reviewed the published literature with a focus on functional noncoding RNAs and epigenetic regulation in ruminants. Results indicate that miRNAs play key roles in the regulation of milk fat synthesis and catabolism in ruminants. Although few data are available, merging evidence indicates that lncRNAs and circRNAs act on milk fat related genes through indirect action with microRNAs or RNAs in the ceRNA network to elicit positive effects on transcription. Although precise regulatory mechanisms remain unclear, most studies have focused on the regulation of the function of target genes through functional noncoding RNAs. Data to help identify factors that can regulate their own expression and function or to determine whether self-regulation involves positive and/or negative feedback are needed. Despite the growing body of research on the role of functional noncoding RNA in the control of ruminant milk fat, most data are still not translatable for field applications. Overall, the understanding of mechanisms whereby miRNA, lncRNA, circRNA, and ceRNA regulate ruminant milk fat remains an exciting area of research.

Circ11103 Interacts with miR-128/PPARGC1A to Regulate Milk Fat Metabolism in Dairy Cows.

The quality of milk is inseparable from its milk components, and fatty acid content is a key factor affecting the quality of milk. In this study, the miRNA and mRNA profiles of the bovine mammary gland tissue during the dry period and the peak lactation period were determined through high-throughput sequencing. In total, 72 miRNA-mRNA regulatory pathways were screened, including miR-128/PPARGC1A regulatory pathways. miR-128 can directly target PPARGC1A and inhibit its expression. In addition, the study also observed that there was a miR-128 binding site in the sequence of the circular RNA circ11103, and circ11103 significantly reduced the expression of miR-128. circ11103 upregulated the triglyceride levels in bovine mammary epithelial cells (BMECs) and increased the contents of unsaturated fatty acids. However, miR-128 decreased triglyceride and cholesterol levels in BMECs. This study aims to analyze the mechanism governing the regulatory effect of circ11103 on milk fat metabolism, which provides new insights into improving milk quality.

circ01592 regulates unsaturated fatty acid metabolism through adsorbing miR-218 in bovine mammary epithelial cells.

The composition of fatty acids plays a key role in regulating milk flavor and quality. Therefore, to improve the quality of milk, it is particularly important to study the regulatory mechanism of fatty acid metabolism in dairy cows. In this study, the expression profiles at non-lactation, peak-lactation, mid-lactation and late-lactation were constructed by high-throughput sequencing. Considering non-lactation as the control group and the other points as the experimental groups, the differentially expressed genes were screened. ELOVL5 was significantly upregulated and was selected for subsequent analyses. Bioinformatics prediction, a dual-luciferase assay, qPCR analysis and western blot analysis were used for verification. The results showed that ELOVL5 was a downstream target gene of miR-218 that regulated milk fat metabolism. A dual-luciferase assay and expression level analysis showed that circ01592 can directly bind to miR-218 and that overexpression of circ01592 (pcDNA-circ01592) significantly reduced the expression of miR-218 and enhanced the expression of ELOVL5, the target gene of miR-218 in BMECs. A functional study of BMECs showed that circ01592 promoted the synthesis of TAG and increased the content of UFA. The function of miR-218 was opposite to that of circ01592. Overall, the data showed that circ01592 promoted TAG synthesis and fatty acid composition by binding miR-218, alleviating the inhibitory effect of miR-218 on ELOVL5 expression. These mechanisms provide a new research approach and theoretical basis for improving milk quality.