A Molecular Perspective and Role of NAD+ in Ovarian Aging.

The decline in female fecundity is linked to advancing chronological age. The ovarian reserve diminishes in quantity and quality as women age, impacting reproductive efficiency and the aging process in the rest of the body. NAD+ is an essential coenzyme in cellular energy production, metabolism, cell signaling, and survival. It is involved in aging and is linked to various age-related conditions. Hallmarks associated with aging, diseases, and metabolic dysfunctions can significantly affect fertility by disturbing the delicate relationship between energy metabolism and female reproduction. Enzymes such as sirtuins, PARPs, and CD38 play essential roles in NAD+ biology, which actively consume NAD+ in their enzymatic activities. In recent years, NAD+ has gained much attention for its role in aging and age-related diseases like cancer, Alzheimer's, cardiovascular diseases, and neurodegenerative disorders, highlighting its involvement in various pathophysiological processes. However, its impact on female reproduction is not well understood. This review aims to bridge this knowledge gap by comprehensively exploring the complex interplay between NAD+ biology and female reproductive aging and providing valuable information that could help develop plans to improve women's reproductive health and prevent fertility issues.

RNA Sequencing Reveals the Inhibitory Effect of High Levels of Arachidonic Acid and Linoleic Acid on C2C12 Differentiation and Myogenic Biomarkers.

Over the past three decades, studies have shown that consuming polyunsaturated fatty acids (PUFAs) can enhance animal and human health and welfare through biological, biochemical, pathological, and pharmacological impacts. Furthermore, omega-6 plays key roles in the cardiopulmonary system, including promoting airway relaxation and inhibiting atherosclerosis and hypertension. However, findings from investigations of the effects of omega-6 fatty acids on molecular and cellular activity and discussions on their influence on biomarkers are still unclear. Therefore, the present study aimed to evaluate omega-6 fatty acids, the arachidonic acid (AA), and linoleic acid (LA) effects on C2C12 proliferation, myogenesis morphology, and relative myogenic biomarker expression through the Wnt pathway. C2C12 cells were cultured with and without 25, 50, 100, and 150 µM of LA and AA and then subjected to CCK8, Giemsa staining, RT qPCR, Western blotting, and RNA Sequencing. The CCK8 Assay results showed that 25, 50, 100, and 150 µM LA significantly decreased the viability after 72 h for 25, 50, 100, and 150 µM concentrations. Also, AA supplementation decreased cell viability after 24 h for 150 µM, 48 h for 150 µM, and 72 h for 50, 100, and 150 µM concentrations. Moreover, the LA and AA inhibitory effects noticed through Gimesa staining were morphological changes during myoblast differentiation. Both LA and AA showed inhibiting IGF1, Cola1, Col6a2, Col6a1, Itga10, Itga11, SFRP2, DAAM2, and NKD2 effects; however, the depressing effect was higher for AA compared to LA. The previous results were confirmed through Western blotting, which showed that 50 µM LA and AA significantly reduced DAAM2 and SFRP2 protein levels compared to the control. Regarding RNA sequencing results, LA and AA increased the number of differentially expressed (DE) Mt-rRNA and snoRNA; however, the numbers of lncRNA detected decreased compared to the control. Our findings demonstrate that high and moderate LA and AA concentrations reduce primary myoblast proliferation and differentiation. Also, they highlight novel biomarkers and regulatory factors to improve our understanding of how the nutrition of fatty acids can control and modulate the myogenesis and differentiation process through different biomarker families.

Integrating mid-infrared spectroscopy, machine learning, and graphical bias correction for fatty acid prediction in water buffalo milk.

BACKGROUND: Buffalo milk, constituting 15% of global production, has higher fatty acids content than Holstein milk. Fourier-transform mid-infrared (FT-MIR) spectroscopy is widely used for dairy analysis, but its application to buffalo milk, with larger fat globules, remains understudied. The ultimate goal of this study is to develop machine learning models based on FT-MIR spectroscopy for predicting fatty acids in buffalo milk and to assess the accuracy of commercial milk analyzers. This research provides a convenient, fast, and environmentally friendly method for detecting the fatty acid composition in buffalo milk. RESULTS: We employed six machine learning algorithms to establish a detection model for 34 fatty acids in buffalo milk. The predictive models demonstrated robust capabilities for high-content fatty acids [C14:0, C15:0, C16:0, C17:0, C18:0, C18:1, saturated fatty acid (SFA), monounsaturated fatty acid (MUFA)], with errors within a 15% range. Traditional FT6000 detection methods exhibited limitations in measuring SFAs and polyunsaturated fatty acids (PUFA). Implementing a mean difference correction of 0.21 for MUFAs and applying regression equations (SFA × 1.0639 + 0.0705; PUFA × 0.5472 + 0.0047) significantly improved measurement accuracy. CONCLUSION: This study successfully developed a predictive model for fatty acids in Mediterranean buffalo milk based on FT-MIR spectroscopy. Additionally, a correction was applied to the existing measurement device, FT6000, enabling more accurate measurements of fatty acids in buffalo milk. The findings have practical implications for the food industry, offering a faster and more reliable approach to assess and monitor fatty acid composition in buffalo milk, potentially influencing product development and quality control processes. © 2024 Society of Chemical Industry.

Identification of Milk Adulteration in Camel Milk Using FT-Mid-Infrared Spectroscopy and Machine Learning Models.

Camel milk, esteemed for its high nutritional value, has long been a subject of interest. However, the adulteration of camel milk with cow milk poses a significant threat to food quality and safety. Fourier-transform infrared spectroscopy (FT-MIR) has emerged as a rapid method for the detection and quantification of cow milk adulteration. Nevertheless, its effectiveness in conveniently detecting adulteration in camel milk remains to be determined. Camel milk samples were collected from Alxa League, Inner Mongolia, China, and were supplemented with varying concentrations of cow milk samples. Spectra were acquired using the FOSS FT6000 spectrometer, and a diverse set of machine learning models was employed to detect cow milk adulteration in camel milk. Our results demonstrate that the Linear Discriminant Analysis (LDA) model effectively distinguishes pure camel milk from adulterated samples, maintaining a 100% detection rate even at cow milk addition levels of 10 g/100 g. The neural network quantitative model for cow milk adulteration in camel milk exhibited a detection limit of 3.27 g/100 g and a quantification limit of 10.90 g/100 g. The quantitative model demonstrated excellent precision and accuracy within the range of 10-90 g/100 g of adulteration. This study highlights the potential of FT-MIR spectroscopy in conjunction with machine learning techniques for ensuring the authenticity and quality of camel milk, thus addressing concerns related to food integrity and consumer safety.

Establishment and Validation of Fourier Transform Infrared Spectroscopy (FT-MIR) Methodology for the Detection of Linoleic Acid in Buffalo Milk.

Buffalo milk is a dairy product that is considered to have a higher nutritional value compared to cow's milk. Linoleic acid (LA) is an essential fatty acid that is important for human health. This study aimed to investigate and validate the use of Fourier transform mid-infrared spectroscopy (FT-MIR) for the quantification of the linoleic acid in buffalo milk. Three machine learning models were used to predict linoleic acid content, and random forest was employed to select the most important subset of spectra for improved model performance. The validity of the FT-MIR methods was evaluated in accordance with ICH Q2 (R1) guidelines using the accuracy profile method, and the precision, the accuracy, and the limit of quantification were determined. The results showed that Fourier transform infrared spectroscopy is a suitable technique for the analysis of linoleic acid, with a lower limit of quantification of 0.15 mg/mL milk. Our results showed that FT-MIR spectroscopy is a viable method for LA concentration analysis.

Evaluation of Parity Effect on Characteristics and Minerals in Buffalo (Bubalus Bubalis) Colostrum and Mature Milk.

Colostrum is a vital performance for buffaloes and potentially functional foods in the future. Therefore, this study aimed to evaluate the difference between the parity of buffalo colostrum and mature milk. Twenty pregnant buffaloes (primiparous = 10; multiparous = 10) were assigned to the same diet prepartum and milking routine postpartum. Calves were separated from the dams immediately after birth and colostrum was harvested within 2 h, whilst mature milk was harvested at 7 days postpartum. The colostrum was analyzed for immunoglobulin G and milk composition as the mature milk. The results showed that there was a higher level of protein, solid not fat, and milk urea nitrogen (p < 0.05), with a tendency for higher total solids (p = 0.08) in primiparous buffaloes' colostrum compared with multiparous. No parity effect was observed in colostrum immunoglobulin G, fat, lactose, and yields of colostrum and composition (p > 0.05). There was no difference in mature milk composition and yield by parity affected (p > 0.05). Compared with mature milk composition, colostrum had a higher content protein, total solids, solid not fat, and milk urea nitrogen (p < 0.05); however, fat and lactose were lower than that of mature milk (p < 0.05). For minerals, multiparous buffaloes' colostrum had a higher concentration of Fe (p = 0.05), while the mature milk had higher concentrations of K and P compared with primiparous. Buffalo colostrum had higher concentrations of Na, Mg, Co, Fe, and K with a lower concentration of Ca relative to mature milk (p < 0.05). It was observed that parity affected colostrum characteristics rather than mature milk and caused subtle variations in minerals in colostrum and mature milk of buffaloes. As lactation proceeded, both milk composition and minerals in the milk changed drastically.

Comparative Genomic Analysis of the Thiolase Family and Functional Characterization of the Acetyl-Coenzyme A Acyltransferase-1 Gene for Milk Biosynthesis and Production of Buffalo and Cattle.

Cattle and buffalo served as the first and second largest dairy animals, respectively, providing 96% milk products worldwide. Understanding the mechanisms underlying milk synthesis is critical to develop the technique to improve milk production. Thiolases, also known as acetyl-coenzyme A acetyltransferases (ACAT), are an enzyme family that plays vital roles in lipid metabolism, including ACAT1, ACAT2, ACAA1, ACAA2, and HADHB. Our present study showed that these five members were orthologous in six livestock species including buffalo and cattle. Transcriptomic data analyses derived from different lactations stages showed that ACAA1 displayed different expression patterns between buffalo and cattle. Immunohistochemistry staining revealed that ACAA1 were dominantly located in the mammary epithelial cells of these two dairy animals. Knockdown of ACAA1 inhibited mammary epithelial cell proliferation and triglyceride and ß-casein secretion by regulating related gene expressions in cattle and buffalo. In contrast, ACAA1 overexpression promoted cell proliferation and triglyceride secretion. Finally, three novel SNPs (g.-681A>T, g.-23117C>T, and g.-24348G>T) were detected and showed significant association with milk production traits of Mediterranean buffaloes. In addition, g.-681A>T mutation located in the promoter region changed transcriptional activity significantly. Our findings suggested that ACAA1 play a key role in regulating buffalo and cattle milk synthesis and provided basic information to further understand the dairy animal lactation physiology.

Summer Buffalo Milk Produced in China: A Desirable Diet Enriched in Polyunsaturated Fatty Acids and Amino Acids.

The objective of the study was to compare and reveal differences in basic chemical parameters, fatty acids, amino acids, and lipid quality indices of crossbred buffalo (swamp x river type) milk produced in summer and winter. The buffalo milk samples were collected in summer (Jul-Aug) and winter (Dec-Jan) from Hubei province, China. The samples were detected by using CombiFoss apparatus, gas chromatography, and an automated specialized amino acid analyzer. The results showed that the basic chemical parameters, fatty acid profiles, lipid quality indices, and amino acid profiles of crossbred buffalo milk differed between summer and winter. Specifically, summer buffalo milk exhibited a higher content of MUFA (monounsaturated fatty acids) and PUFA (polyunsaturated fatty acids) than winter buffalo milk. Summer buffalo milk had a lower content of major SFA (saturated fatty acids), a higher content of ω-3 and DFA (hypocholesterolemic fatty acids), a lower ω-6/ω-3 ratio, a higher value of 3 unsaturated fatty acid indices (C14, C16, C18), and a lower value of IA (index of atherogenicity) and IT (index of thrombogenicity) than winter buffalo milk. Additionally, 17 amino acids, including 8 EAA (essential amino acids) and 9 NEAA (non-essential amino acids) were higher in summer buffalo milk. These results indicated that summer buffalo milk was more health-beneficial than winter buffalo milk. Therefore, summer buffalo milk might be a desirable diet option for human nutrition and health. Our findings provide valuable information for the research and development of buffalo dairy products in China or other Asian countries.

Effects of capsicum oleoresin supplementation on rumen fermentation and microbial abundance under different temperature and dietary conditions in vitro.

This study aimed to determine the effect of capsicum oleoresin (CAP) on rumen fermentation and microbial abundance under different temperature and dietary conditions in vitro. The experimental design was arranged in a 2 × 2 × 3 factorial format together with two temperatures (normal: 39°C; hyperthermal: 42°C), two forage/concentrate ratios (30:70; 70:30), and two CAP concentrations in the incubation fluid at 20 and 200 mg/L with a control group. Regarding the fermentation characteristics, high temperature reduced short-chain fatty acids (SCFA) production except for molar percentages of butyrate while increasing acetate-to-propionate ratio and ammonia concentration. The diets increased total SCFA, propionate, and ammonia concentrations while decreasing acetate percentage and acetate-to-propionate ratio. CAP reduced acetate percentage and acetate-to-propionate ratio. Under hyperthermal condition, CAP could reduce acetate percentage and increase acetate-to-propionate ratio, lessening the negative effect of high heat on SCFA. Hyperthermal condition and diet altered the relative abundance of microbial abundance in cellulose-degrading bacteria. CAP showed little effect on the microbial abundance which only increased Butyrivibrio fibrisolvens. Thus, CAP could improve rumen fermentation under different conditions, with plasticity in response to the ramp of different temperature and dietary conditions, although hardly affecting rumen microbial abundance.

Icariin Alleviates Escherichia coli Lipopolysaccharide-Mediated Endometritis in Mice by Inhibiting Inflammation and Oxidative Stress.

Icariin (ICA) is a naturally occurring phytochemical agent primarily extracted from Epimedium Brevicornum Maxim (Family Berberidaceae) with a broad spectrum of bioactivities. Endometritis is a uterine disease that causes enormous losses in the dairy industry worldwide. In this study, anti-inflammatory and anti-oxidant properties of ICA were investigated against lipopolysaccharide (LPS)-induced endometritis in mice to investigate possible underlying molecular mechanisms. Sixty heathy female Kunming mice were randomly assigned to four groups (n = 15), namely control, LPS, LPS + ICA, and ICA groups. The endometritis was induced by intrauterine infusion of 50 µL of LPS (1 mg/mL). After 24 h of onset of LPS-induced endometritis, ICA groups were injected thrice by ICA intraperitoneally six hours apart. Histopathological examination, enzyme linked immunosorbent assay (ELISA), real time quantitative polymerase chain reaction (RT-qPCR), western blotting, and immunohistochemistry were used in this study. Histological alterations revealed that ICA markedly mitigated uterine tissue injury caused by LPS. The results showed that the ICA inhibited the production of pro-inflammatory cytokines (IL-1ß, IL-6, and TNF-α) and boosted the production of anti-inflammatory cytokines (IL-10). Additionally, ICA modulated the expression of malondialdehyde (MDA), reactive oxygen species (ROS), superoxide dismutase 1 (SOD1), catalase (CAT), and glutathione peroxidase 1 (Gpx1) induced by LPS. The administration of ICA significantly (p < 0.05) improved the mRNA and protein expression of Toll-like receptor (TLR) 4. The western blotting and ELISA finding revealed that the ICA repressed LPS-triggered NF-κB pathway activation. Moreover, ICA improved the antioxidant defense system via activation of the Nrf2 pathway. The results revealed that ICA up-regulated the mRNA and protein expression of Nuclear erythroid-2-related factor (Nrf2), NAD(P)H: quinone oxidoreductase 1 (NQO1), heme oxygenase-1 (HO-1), and glutamate-cysteine ligase catalytic subunit (GCLC) under LPS exposure. Conclusively, our findings strongly suggested that ICA protects endometritis caused by LPS by suppressing TLR4-associated NF-κB and Nrf2 pathways. Altogether, these innovative findings may pave the way for future studies into the therapeutic application of ICA to protect humans and animals against endometritis.

Evaluating the Effect of Forage Rape (Brassica napus) Ensiling Kinetics on Degradability and Milk Performance as Non-conventional Forage for Dairy Buffalo.

The recent increase in demand for animal protein sources has led to the urgency to introduce non-conventional feed sources and opened the space to study feed management and its effects on animal productivity. Forage rape (Brassica napus L.) is a high-quality forage crop with a remarkable nutritional value and productive and fast growth capacity; however, studies on processing methods are limited. This study evaluates the effect of an ensiling process on rape silage quality kinetics, in situ degradability, and milk responses in dairy buffaloes. Firstly, the whole-plant forage rape was ensiled, and silage samples were collected 30, 60, and 90 days after ensiling to determine pH, evaluation of sensory characteristics, and chemical composition. Then, samples were taken for further chemical analysis at days 30, 60, and 90. After that, the degradability of the dry matter (DM) and crude protein (CP) of the silage was evaluated by an in situ degradability experiment using three fistulated buffalos (550 ± 20 kg body weight, 4.7 ± 0.76 years). Finally, whole-plant rape silage (after 60 days) was included in a 10, 20, and 30% of DM dairy buffalo diet in the lactating buffalo ration. The results showed that silage pH did not change significantly during the ensiling process (p > 0.05); however, the silage achieved the optimal comprehensive sensory characteristic score from days 30 to 60. There was also a significant change in neutral detergent fiber (NDF) content and acid detergent fiber content, which decreased significantly (p = 0.001 and p < 0.001, respectively). Ensiling of the whole-plant rape significantly reduced effective DM degradability (p < 0.05) without altering CP degradability (p > 0.05). Furthermore, the inclusion of forage rape silage linearly (p = 0.03) increased milk fat and protein contents and did not affect milk yield, lactose, and urea nitrogen contents in raw buffalo milk. In conclusion, whole-plant rape silage could significantly maintain the optimal ether extract (EE) protein content without affecting CP degradability, in addition to improving milk fat and milk protein. Therefore, ensiling may be an efficient method of forage rape utilization, and forage rape silage can be recommended as a good forage source for dairy buffaloes.

Nutritional Modulation, Gut, and Omics Crosstalk in Ruminants.

Ruminant nutrition has significantly revolutionized a new and prodigious molecular approach in livestock sciences over the last decade. Wide-spectrum advances in DNA and RNA technologies and analysis have produced a wealth of data that have shifted the research threshold scheme to a more affluent level. Recently, the published literature has pointed out the nutrient roles in different cellular genomic alterations among different ruminant species, besides the interactions with other factors, such as age, type, and breed. Additionally, it has addressed rumen microbes within the gut health and productivity context, which has made interpreting homogenous evidence more complicated. As a more systematic approach, nutrigenomics can identify how genomics interacts with nutrition and other variables linked to animal performance. Such findings should contribute to crystallizing powerful interpretations correlating feeding management with ruminant production and health through genomics. This review will present a road-mapping discussion of promising trends in ruminant nutrigenomics as a reference for phenotype expression through multi-level omics changes.

Analysis of miRNA expression changes in bovine endometrial stromal cells treated with lipopolysaccharide.

After parturition, bovine uterine stromal cells are often exposed to complex bacterial and viral stimuli owing to epithelial cell rupture, resulting in an inflammatory response. In this study, we used an in vitro model to study the response of bovine endometrial stromal cells to inflammatory mediators and the associated regulated microRNAs in response to lipopolysaccharide. Lipopolysaccharide (LPS) is a bacterial wall component in gram-negative bacteria that causes inflammation upon immune recognition, which is used to create in vitro inflammation models. Thus, we used high-throughput RNA sequencing to identify miRNAs that may have an anti-inflammatory role in the LPS-induced inflammatory response. Two groups of bovine uterine cells were treated with phosphate buffer saline (PBS) and LPS, respectively. Compared with the control (PBS) group, the LPS-treated group had 219 differentially expressed miRNAs, of which 113 were upregulated, and 106 were downregulated. Gene ontology enrichment analysis revealed that the target genes of differentially expressed miRNAs were significantly enriched in several activities, such as transferase activity, small molecule binding, and protein binding. Kyoto Encyclopedia of Genes and Genomes pathway analysis indicated that the target genes of differential miRNAs were significantly enriched in fluid shear stress and atherosclerosis, MAPK signaling pathway, TNF signaling pathway. By analyzing differentially expressed miRNAs, we found that miR-200c, miR-1247-3p, and let-7b are directly related to the inflammatory response. For instance, miR-200c target genes (MAP3K1, MAP4K3, MAPKAPK5, MAP3K8, MAP3K5) and let-7b target genes (CASP3, IL13, MAPK8, CXCL10) were significantly enriched in the MAPK and IL-17 signaling pathways, respectively. In summary, our research provides insight into the molecular mechanism underlying LPS-induced inflammation in vitro, which may unveil new targets for the treatment of endometritis.

Role of MicroRNAs in Protective Effects of Forsythoside A Against Lipopolysaccharide-Induced Inflammation in Bovine Endometrial Stromal Cells.

Bovine endometrial stromal cells (bESCs) are exposed to a complex environment of bacteria and viruses due to the rupture of epithelial cells after delivery. Inflammatory responses are elicited by the activation of host pattern recognition receptors through pathogen-related molecules such as lipopolysaccharides (LPS) on the cell membrane. Forsythoside A (FTA) is a major active constituent of Forsythia suspensa (Thunb.) Vahl. is a flowering plant widely employed as a traditional Chinese herbal medicine to treat various inflammatory diseases such as nephritis, eye swelling, scabies, ulcers, and mastitis; however, the molecular mechanisms underlying its therapeutic effects on bovine endometritis are still unclear. The aim of this study was to explore the role of miRNA and the mechanisms underlying the protective activity of FTA on the inflammation of bovine endometrial stromal cells induced by LPS. Based on previous research, we isolated and cultured bESCs in vitro and categorized them into LPS and LPS+FTA groups with three replicates. Upon reaching 80% confluence, the bESCs were treated with 0.5 µg/mL of LPS or 0.5 µg/mL of LPS + 100 µg/mL of FTA. We, then, performed high-throughput sequencing (RNA-Seq) to investigate the effects of FTA on LPS-stimulated primary bESCs and their underlying mechanisms. We identified 167 miRNAs differentially expressed in the LPS groups; 72 miRNAs were up-regulated, and 95 were down-regulated. Gene ontology enrichment analysis revealed that differentially expressed microRNA (DEGs) were most enriched during the cellular metabolic process; they were mostly located intracellularly and participated in protein, enzyme, and ion binding. Kyoto Encyclopedia of Genes and Genomes pathway analysis revealed that the DEGs were most enriched in the mitogen-activated protein kinase, tumor necrosis factor, and Interleukin-17 signaling pathways. These results reveal the complex molecular mechanism involved in the FTA and provide a basis for future studies of bovine endometritis treatment with traditional Chinese medicine monomer.

Analysis of Transcriptomic Changes in Bovine Endometrial Stromal Cells Treated With Lipopolysaccharide.

Endometritis adversely affects the ability of cattle to reproduce and significantly reduces milk production. The is mainly composed of epithelial and stromal cells, and they produce the first immune response to invading pathogens. However, most of the epithelial cells are disrupted, and stromal cells are exposed to an inflammatory environment when endometritis occurs, especially postpartum. Many bacteria and toxins start attacking stromal cell due to loss of epithelium, which stimulates Toll-like receptor (TLRs) on stromal cells and causes upregulated expression of cytokines. Understanding the genome-wide characterization of bovine endometritis will be beneficial for prevention and treatment of endometritis. In this study, whole-transcriptomic gene changes in bovine endometrial stromal cells (BESCs) treated with LPS were compared with those treated with PBS (control group) and were analyzed by RNA sequencing. Compared with the control group, a total of 366 differentially expressed genes (DEGs) were identified in the LPS-induced group (234 upregulated and 132 downregulated genes), with an adjusted P < 0.05 by DESeq. Gene Ontology (GO) enrichment analysis revealed that DEGs were most enriched in interleukin-1 receptor binding, regulation of cell activation, and lymphocyte-activated interleukin-12 production. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis revealed DEGs were most enriched in the TNF signaling pathway, Toll-like receptor signaling pathway, cytokine-cytokine receptor interaction, NF-κB signaling pathway, and chemokine signaling pathway. The results of this study unraveled BESCs affected with LPS transcriptome profile alterations, which may have a significant effect on treatment inflammation by comprehending molecular mechanisms and authenticating unique genes related to endometritis.