Search Results (31)

The scientific evidence regarding the use of plant-derived extracts to alleviate exercise-induced muscle damage in horses remains limited. Mulberry leaf flavonoids (MLFs) are the primary bioactive constituents of a traditional medicinal plant and are potent antioxidants. The aim of this study was to investigate the protective effects of MLFs against exercise-induced muscle damage. In this study, twelve Mongolian horses were used in a 3 × 3 Latin square design to investigate the protective effects of MLFs. Our results showed that high-intensity exercise negatively impacted the immune status, metabolic state, myofibrillar structure, and antioxidant capacity of the horses. Conversely, MLFs significantly reduced blood levels of white blood cells (WBC), monocytes (MON), aspartate aminotransferase (AST), creatine kinase (CK), and malondialdehyde (MDA) across various exercise distances and during recovery. Simultaneously, MLFs increased serum glutathione peroxidase (GPx), superoxide dismutase (SOD), and total antioxidant capacity (T-AOC). Mechanistically, transcriptomic analysis revealed that dietary MLFs upregulated genes associated with myofibrillar structural proteins (MYOZ2, MYOM3), the antioxidant defense system (GPX3, SOD3), and skeletal muscle satellite cell proliferation and differentiation (MYOD1, MRF6). Furthermore, quantitative proteomics indicated the enrichment of the PI3K-Akt and TGF-β signaling pathways, as well as ECM–receptor interactions, suggesting their potential involvement in regulating protein metabolism and facilitating myofibrillar restoration. Overall, MLFs effectively alleviated inflammation, metabolic disorder, and exercise-induced muscle damage. Under the tested conditions, a daily dosage of 10 g MLFs provided superior protective effects. Full article

Severe winter nutritional deficiencies may impair reproductive performance in Mongolian mares, yet optimal protein requirements during late gestation remain undefined. This study aimed to determine the effects of varying protein levels in complementary feed on gestational performance, physiology, and gut health. Seventeen late-gestating mares were assigned to three isocaloric diets differing in crude protein (CP)—high (HCP, 13.25%), medium (MCP, 12.04%), and low (LCP, 10.85%)—for 40 days. Statistical analysis was conducted using one-way ANOVA followed by Tukey’s HSD post hoc test. Mares fed the MCP diet tended to show more favorable growth performance and nutrient digestibility relative to the HCP group (p < 0.05). Blood analysis suggested that MCP mares had comparatively lower serum creatinine and creatine kinase concentrations, along with higher antioxidant capacity (catalase) and interleukin-1β levels (p < 0.001). Fecal microbiota sequencing showed that MCP was associated with comparatively higher microbial diversity, while HCP was characterized by enrichment of Proteobacteria, and LCP by enrichment of Bacteroidetes. Metabolomics identified 533 differential metabolites linked to protein metabolism. The MCP diet may help balance immune function, antioxidant status, and microbial homeostasis. These findings suggest that a complementary feed containing 12.04% CP may be associated with favorable effects on maternal health-related indicators in late-gestating Mongolian mares during winter. Full article

Mongolian horses are an indigenous Chinese breed known for their endurance capacity, yet quantitative descriptions of their gait-related kinematic characteristics remain limited. This pilot exploratory study aimed to describe the kinematics of Mongolian horses during walk, slow trot, and fast trot, and to examine whether selected variables differed between race-result groups in a 12 km endurance race. Forty-six horses were classified into an excellent group and an ordinary group based on the result of a single race. Kinematic data were collected using optical motion capture and three-dimensional skeletal modelling. Separate gait-specific linear mixed-effects models were fitted, with horse identity as a random effect and group and speed as fixed effects. The results showed gait-dependent between-group differences. During walk, the excellent group had significantly greater range of motion of the tarsal, hip, and elbow joints, as well as a greater maximum forelimb retraction angle (all p < 0.001). During slow trot, the excellent group showed significantly greater stride length (p = 0.009), elbow joint range of motion (p < 0.001), minimum hindlimb forward extension angle (p = 0.033), and minimum forelimb forward extension angle (p = 0.004). During fast trot, the between-group differences were most pronounced, with significantly greater stride length (p < 0.001) and range of motion of the tarsal joint (p < 0.001), hip joint (p = 0.015), and elbow joint (p = 0.014), together with greater maximum hindlimb retraction angle (p = 0.001) and minimum forelimb forward extension angle (p = 0.026). Overall, these findings provide preliminary evidence that gait-related kinematic differences may exist between race-result groups in Mongolian horses. However, because this was an exploratory study based on a single race, the findings should be interpreted cautiously and require validation in larger and more diverse cohorts. Full article

(1) Background: The “Bider” marking refers to the symmetrical black stripes distributed on the shoulder blades of Dun Mongolian horses, representing an ancestral trait of significant genetic value. However, the molecular mechanisms underlying its formation remain unclear. This study aims to elucidate the molecular basis of these markings by comparing transcriptomic differences in skin tissues from variously pigmented areas of Mongolian horses’ “Bider” patterns. (2) Methods: Using three Dun Mongolian horses as subjects, skin tissue samples were collected from their shoulders (dark-marked and light-marked areas), dorsal midline, and croup regions for transcriptome sequencing. Differentially expressed genes were identified based on sequencing data, followed by Gene Ontology (GO) functional annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis. Key findings were validated through quantitative reverse transcription polymerase chain reaction (qRT-PCR). (3) Results: The sequencing yielded approximately 893 million high-quality clean reads, with an overall alignment rate exceeding 96%. A total of 140 to 775 differentially expressed genes were identified. GO enrichment analysis revealed that these genes were significantly enriched in biological processes related to pigment metabolism, skin and hair follicle development, signal transduction (including calcium and cyclic guanosine monophosphate (cGMP) signaling), and immune regulation. KEGG analysis further indicated that multiple pathways closely associated with pigment regulation, including the calcium signaling pathway, tyrosine metabolism, cyclic adenosine monophosphate (cAMP) signaling pathway, and melanoma pathway, were significantly enriched across different tissue comparison groups, suggesting their potential key roles in coat color phenotype formation. The reliability of the sequencing data was corroborated by the results of qRT-PCR validation. (4) Conclusions: This study conducted a transcriptome analysis of skin samples from various pigmented regions of the Dun Mongolian horse’s Bider marking, revealing that the formation of this marking is associated with the differential expression of numerous genes and is co-regulated by multiple pigment-related signaling pathways. Full article

(1) Background: The “Bider marking” on the shoulder of the Dun Mongolian horse represents a unique pigmentation pattern, the molecular formation mechanism of which remains incompletely understood. This study investigates the differential expression and protein localization of pigment-related genes—specifically the core transcription factor MITF, as well as EDN3, SLC7A11, and WNT3A—in the skin. The analysis focuses on three distinct regions: the dark-colored area of the ‘Bider marking’ shoulder (BIDC), the light-colored area of the ‘Bider marking’ shoulder (BILC), and the non-Bider-marked shoulder area (NBIS). The aim is to clarify their correlation with the formation of this distinctive pigmentation pattern. (2) Methods: Skin tissue samples from both the “Bider marking” and non-Bider-marked shoulder regions were collected (n ≥ 3). The mRNA expression levels were quantified using RT-qPCR, protein levels were analyzed through Western blotting, and protein localization was assessed via immunohistochemistry and immunofluorescence. (3) Results: Compared to the NBIS group, both the BIDC and BILC groups exhibited significantly elevated protein expression of MITF and WNT3A. Further immunofluorescence showed that the distribution of MITF protein exhibits regional specificity in the epidermis and hair follicles. In the BIDC region, the protein is localized specifically to the stratum corneum of the epidermis, the dermal papilla, and the outer root sheath of hair follicles. In contrast, the mRNA and protein expression levels of SLC7A11 and EDN3 did not display consistent patterns among the three groups, and no specific differences were observed in tissue localization. (4) Conclusions: The findings show that the specific pigmentation in dark “Bider marking” regions is closely linked to the upregulated protein levels and unique spatial patterns of MITF and WNT3A; SLC7A11 and EDN3 may not be primary regulators of this trait. Full article

Ferulic acid is a bioactive phenolic compound with potential benefits for skeletal muscle health. In this study, Mongolian horses were used as experimental subjects and were orally administered ferulic acid at doses of 5, 10, or 15 g per horse per day for 40 consecutive days. Muscle biopsy samples were analyzed using proteomics to assess fiber type composition and regulatory protein expression. Ferulic acid supplementation increased the proportion of fast-twitch fibers and upregulated key differentiation factors such as MUSTN1, while modulating glycolysis, ECM remodeling, and calcium signaling pathways. Although ferulic acid induced moderate oxidative stress, it did not trigger classical ferroptosis. Collectively, these findings demonstrate that ferulic acid promotes fast-twitch fiber transformation in horses through coordinated metabolic and signaling mechanisms, highlighting its potential as a nutritional strategy to enhance skeletal muscle adaptability and athletic performance in horses. Full article

(1) Background: The dun coat color, a wild-type phenotype in horses characterized by pigment dilution and primitive markings, is regulated by TBX3. This study explored the expression and localization of TBX3 in the Bider marking (a primitive mark unique to the shoulder of horses); (2) Methods: We compared skin tissues from Bider-marked and non-Bider dun Mongolian horses. Samples were collected from the Bider area (dark-colored/light-colored shoulder), dorsal midline, and croup. Histological staining, qRT-PCR, and Western blotting were used to analyze pigment distribution and TBX3 expression at mRNA and protein levels; (3) Results: Histology revealed asymmetric pigment deposition in hair shafts from light-colored areas of both Bider and non-Bider horses, whereas dark areas showed symmetric distribution. qRT-PCR and Western blotting showed TBX3 expression was significantly higher in the shoulder of non-Bider horses compared to Bider horses. Conversely, Bider horses exhibited higher TBX3 levels in all other sampled areas. Immunohistochemistry localized TBX3 protein to the epidermis and hair follicle bulbs in both groups; (4) Conclusions: In dun Mongolian horses, TBX3 expression differences between dark and light skin areas correlate with Bider markings. TBX3 is implicated in this specific pigment marking, though its upstream regulation requires further study. These findings provide key insights into the mechanism behind Bider marking formation. Full article

Background: Normal spermatogenesis in Mongolian horses depends on the mitotic division of spermatogonia, two successive meiotic divisions, and the morphological transformation of spermatids into mature spermatozoa. The MEI1 gene is involved in the meiosis cycle and is required for normal chromosome association during meiosis. Previous studies have shown that alternative splicing of MEI1 may promote spermatogenesis in Mongolian horses. In this paper, the regulatory effects of different MEI1 alternative splicing events on Mongolian horse spermatogenesis are investigated. Results: In this study, two overexpressed lentiviral vectors with mutually exclusive exon (MXE) and skipped exon (SE) events of MEI1 were constructed and successfully used to infect Sertoli cells. After 72 h of viral infection, the expression of MEI1 was higher in the SE event than in the MXE event (p < 0.001), as shown by fluorescence quantification; transcriptomics and metabolomics were then used to screen and annotate the differential genes and metabolites, and 193 differentially expressed genes (comprising 109 genes, such as MEI1, and 84 genes with upregulated and downregulated expression, respectively) and 11,360 differentially expressed metabolites (comprising 7494 and 3866 metabolites with upregulated and downregulated expression, respectively) were screened. Differential genes and metabolites were mainly enriched in several metabolic pathways related to spermatogenesis. Differential genes such as IL31RA, ATP2B3, and CASQ2 were highly expressed in SE events, while IL11, PRLR, and CCR7 were highly expressed in MXE events. Metabolites such as folic acid and spermine were highly expressed during SE events, while citric acid and glutathione were highly expressed during MXE events. This suggests that both MXE and SE events of the MEI1 gene can promote the activity of the spermatogenesis signaling pathway. Conclusions: The MXE and SE splicing events of the MEI1 gene may influence spermatogenesis by regulating the expression of spermatogenesis-related genes and metabolites. These findings provide a theoretical foundation for further investigations into the regulatory mechanisms of different alternative splicing events in Mongolian horse spermatogenesis. Full article

This study aimed to generate information for parentage testing in horse breeds using microsatellites (STRs) and single-nucleotide polymorphisms (SNPs). Genotype data were obtained from 189 horse hair root samples, including 38 Thoroughbreds (TBs), 17 Jeju horses (JHs), 20 Quarter horses (QHs), 21 American Miniatures (AMs), and 93 Mongolian horses (MHs), using 15 STR markers and 71 SNP markers. Comparative analysis revealed that the mean expected heterozygosity ranged from 0.468 (AM) to 0.491 (JH) for SNPs and from 0.695 (TB) to 0.791 (MH) for STRs. The mean observed heterozygosity ranged from 0.415 (AM) to 0.487 (MH) for SNPs and from 0.706 (JH) to 0.776 (MH) for STRs. The mean polymorphic information content ranged from 0.349 (AM) to 0.364 (MH) for SNPs and from 0.635 (TB) to 0.761 (MH) for STRs. The inbreeding coefficient ranged from −0.009 (MH) to 0.113 (AM) for SNPs and from −0.058 (TB) to 0.043 (AM) for STRs. The cumulative exclusion probability (PE) for the 71-SNP panel exceeded 0.9999, indicating that SNP markers may be sufficient for parentage testing. In comparison, the STR markers yielded a combined PE of 0.9988 when one parent was known and 0.9999 when both parents were known. These findings highlight the potential of SNPs as alternatives to STRs for routine paternity verification in horses. Full article

Mongolian horses are renowned for their remarkable endurance and ability to adapt to harsh environments. To delve deeper into the molecular mechanisms that underlie these traits, researchers conducted a comprehensive analysis of transcriptomic and proteomic changes in Mongolian horses at three distinct time points: before, immediately after, and 24 h following a 20 km run. The transcriptomic analysis uncovered significant variations in gene expression patterns across these time points. Specifically, 291 differentially expressed genes (DEGs) were identified when comparing pre-exercise to post-exercise conditions, 832 DEGs in the comparison between post-exercise and 24 h post-exercise, and 127 DEGs in the comparison of pre-exercise to 24 h post-exercise. Notably, key genes involved in metabolic activities and cellular proliferation, such as PI3K and LDHA, exhibited significant upregulation immediately after exercise but demonstrated a downward trend 24 h post-exercise. Concurrently, the proteomic analysis revealed 49 differentially expressed proteins (DEPs) in the pre-exercise versus post-exercise comparison, 61 DEPs in the post-exercise versus 24 h post-exercise comparison, and 101 DEPs in the pre-exercise versus 24 h post-exercise comparison. Some proteins, like PDK4 and GLUL, remained upregulated at 24 h post-exercise, whereas others, such as PFKM and LDHA, showed signs of recovery or downregulation. By integrating the transcriptomic and proteomic data, we were able to pinpoint overlapping DEGs/DEPs and implicate crucial signaling pathways, including the HIF-1 signaling pathway and glycolysis, in the molecular response of Mongolian horses to exercise. These findings offer insights into the endurance adaptation mechanisms of the Mongolian horse. Full article

Skeletal muscle satellite cells are muscle stem cells that play an important role in the growth, development, and repair of skeletal muscle as well as in the locomotor performance of the animal body. Lysine is the first limiting amino acid and is involved in multiple metabolic pathways in the organism to maintain overall physiological requirements. In this study, Mongolian horse satellite cells were cultured using lysine culture solution at different concentrations, and the proliferative capacity of satellite cells was detected by the cck-8 assay, and the optimal culture concentration was selected. Then, whole transcriptome sequencing technology was used to determine the differential gene expression and regulatory pathways during the proliferation of lysine-cultured satellite cells after 48 h of culture. Our findings revealed that 0.5 mmol/L lysine is the optimal concentration to increase satellite cell activity in equine muscle. The differential genes involved in satellite cell proliferation were mainly enriched in the cAMPsignaling pathway, calcium signaling pathway, and PPAR signaling pathway. Furthermore, upregulation of PLIN5, ACADL, and FADS2 and downregulation of LOC100052888 regulated the expression of the PPAR signaling pathway. 0.5 mmol/L lysine was the optimal concentration to increase satellite cell activity. Lysine can regulate mitochondrial function and lipid metabolism through the PPAR signaling pathway, and promote the proliferation of equine myosatellite cells. Full article

Speed and endurance are the primary goals in racehorse breeding. The Grassland-Thoroughbred is a newly developed breed in northern China that combines speed, endurance, and environmental adaptability. However, current research on the genetic background of this breed and the genes associated with athletic performance remains limited. We conducted whole-genome resequencing on Mongolian (MG), Thoroughbred (TB), Xilingol (XL), and Grassland-Thoroughbred (CY) horses, generating 3813.74 Gb of clean data after quality control. The number of transitions was significantly higher than that of transversions. The SNPs were mainly located in intergenic regions, followed by intronic regions. Principal component analysis, population structure analysis, and phylogenetic tree results indicated that the CYs had a distinct genetic background from MGs, TBs, and XLs, but based on PCA and phylogenetic clustering, they showed greater genetic similarity to Thoroughbreds. Using fixation index (Fst) and nucleotide diversity ratio (π ratio) analyses between CYs and the other three horse populations, 70, 76, and 80 candidate genes were identified from the intersection of the two methods, respectively. A total of 179 candidate genes were obtained from the union of the three groups. Candidate genes associated with athletic performance (ATF2, NDUFS7, PRKG1, IGFN1, MTOR, TTN) and growth and development (MTOR, IGFN1, COL21A1, NEDD4, PIEZO1) were screened. These genes are related to athletic ability and developmental processes in the CY population. Our study reveals genomic information associated with important traits in Grassland-Thoroughbreds and identifies valuable candidate genes, laying a foundation for future breeding and trait association studies. Full article

The traditional horse industry has undergone a remarkable evolution, with horse racing emerging as a prominent and pivotal economic driver within the sector. Among the various breeds, Mongolian horses, renowned for their exceptional endurance and speed, occupy a significant position in the horse industry. To investigate their homeostasis mechanisms during and after a 20 km endurance exercise and identify novel oxidative-imbalance markers, we selected 12 two-year-old horses and collected blood samples at various time points before, during (at 5, 10, 15, and 20 km), and after the exercise (at 1, 2, 4, and 6 h post-exercise). These samples were analyzed for haematology, blood biochemistry, antioxidant enzyme activities, and liquid chromatography–mass spectrometry (LC-MS) metabolomics. Our results revealed significant changes in heart rate, speed, blood cells, and biochemical markers throughout the exercise. Antioxidant indicators decreased, while malondialdehyde increased, indicating oxidative imbalance post-exercise. Metabolomics analysis identified 122 differential metabolites, including uric acid and L-tyrosine, which were enriched in pathways related to energy metabolism. Uric acid and tyrosine correlated positively with serum creatine kinase, suggesting their potential as markers of oxidative-imbalance injury. These findings elucidate the mechanisms of endurance adaptability in Mongolian horses and provide a theoretical basis for mitigating oxidative imbalance, enhancing horse performance, and promoting the sustainable development of the equine industry. Full article

Muscle fibers, as the fundamental units of muscle tissue, play a crucial role in determining skeletal muscle function through their growth, development, and composition. To investigate changes in muscle fiber types and their regulatory mechanisms in Mongolian horses (MG), Xilingol horses (XL), and Grassland-Thoroughbreds (CY), we conducted histological and bioinformatic analyses on the gluteus medius muscle of these three horse breeds. Immunofluorescence analysis revealed that Grassland-Thoroughbreds had the highest proportion of fast-twitch muscle fibers at 78.63%, while Mongolian horses had the lowest proportion at 57.54%. Whole-transcriptome analysis identified 105 differentially expressed genes (DEGs) in the CY vs. MG comparison and 104 DEGs in the CY vs. XL comparison. Time-series expression profiling grouped the DEGs into eight gene sets, with three sets showing significantly up-regulated or down-regulated expression patterns (p < 0.05). Additionally, 280 differentially expressed long non-coding RNAs (DELs) were identified in CY vs. MG, and 213 DELs were identified in CY vs. XL. A total of 32 differentially expressed microRNAs (DEMIRs) were identified in CY vs. MG, while 44 DEMIRs were found in CY vs. XL. Functional enrichment analysis indicated that the DEGs were significantly enriched in essential biological processes, such as actin filament organization, muscle contraction, and protein phosphorylation. KEGG pathway analysis showed their involvement in key signaling pathways, including the mTOR signaling pathway, FoxO signaling pathway, and HIF-1 signaling pathway. Furthermore, functional variation-based analyses revealed associations between non-coding RNAs and mRNAs, with some non-coding RNAs targeting genes potentially related to muscle function regulation. These findings provide valuable insights into the molecular basis for the environmental adaptability, athletic performance, and muscle characteristics in horses, offering new perspectives for the breeding of Grassland-Thoroughbreds. Full article

The epigenetic regulation of gene expression through the covalent modification of histones is crucial for developing germline cells. To study the regulatory role of alternative splicing (AS) of euchromatic histone lysine methyltransferase 2 (EHMT2/G9A) in spermatogenesis in Mongolian horses, this study first examines the localization of the EHMT2 gene in testicular support cells and then predicts the higher-order structures of sequences with and without AS. Two types of lentiviral vectors for overexpression were subsequently constructed for the EHMT2 gene, one with AS and one without, to infect support cells. The proliferation and activity of infected cells were measured using CCK8, and the differential expression of spermatogenesis-related genes in the two types of support cells was analyzed via qRT–PCR. We analyzed the expression of EHMT2 by immunofluorescence staining. EHMT2 was expressed in the nuclei of Sertoli cells. The expression of spermatogenesis-related genes was measured in the two types of cells. The results reveal that the expression levels of the FSH, Stra8, CCNB2, CDC27, NRG1, PPP2R5C, CCNB2, and YWHAZ genes in the AS group were greater than those in the control group. These results indicate that AS events in EHMT2 affect gene expression and thus affect spermatogenesis. Full article