In mammals, exposure to low temperatures induces white adipose tissue (WAT) browning and alters lipid metabolism to promote thermogenesis, thereby maintaining body temperature. However, this response varies across different adipose depots. In this study, Hezuo pigs were exposed to either room temperature (23 ± 2 °C) or low temperature (−15 ± 2 °C) for periods of 12 h, 24 h, 48 h, 5 d, 10 d, and 15 d. Inguinal fat (IF) and perirenal fat (PF) were collected and analyzed using hematoxylin and eosin (HE) staining, transmission electron microscopy, RT-qPCR, and RNA-seq. Following cryoexposure, our results demonstrated a significant increase in adipocyte number and a corresponding decrease in cross-sectional area in both IF and PF groups from 24 h to 10 d. While adipocyte numbers were elevated at 12 h and 15 d, these changes were not statistically significant. Moreover, lipid droplets and mitochondria were more abundant, and the mRNA expression levels of thermogenic genes UCP3 and PGC-1α were significantly higher compared to the control group during the 24 h-10 d cold exposure period. No significant changes were observed in the other groups. RNA-seq data indicated that the lipid metabolism of IF and PF peaked on day 5 of low-temperature treatment. In IF tissue, lipid metabolism is mainly regulated by genes such as FABP4, WNT10B, PCK1, PLIN1, LEPR, and ADIPOQ. These genes are involved in the classical lipid metabolism pathway and provide energy for cold adaptation. In contrast, in PF tissue, genes like ATP5F1A, ATP5PO, SDHB, NDUFS8, SDHA, and COX5A play roles within the neurodegenerative disease pathway, and PF tissue has a positive impact on the process related to degenerative diseases. Further investigation is needed to clarify the functions of these candidate genes in lipid metabolism in Hezuo pigs and to explore the genetic mechanisms underlying the cold-resistance traits in local pig populations.

The Hezuo pig, an important native Tibetan breed in China, exhibits differences in adult body weight, with females typically heavier than males. The underlying mechanisms for this disparity remain unclear. DNA methylation changes are known to influence animal growth and development and regulate Hezuo pig growth by altering gene expression related to these processes, thus differentially affecting adult body weight between genders. This study conducted DNA methylation analysis and expression profiling using pituitary tissues from male and female Hezuo pigs at 3 and 8 months old (M3M, M3F, M8M, and M8F). In total 346, 795, 371, and 839 differentially methylated genes (DMGs) were identified in the M3M vs. M3F, M3F vs. M8F, M3M vs. M8M, and M8M vs. M8F groups, respectively. The comparative analysis of differentially methylated regions (DMRs) genes and DEGs (differentially expressed regions) revealed that key genes involved in growth, hormone secretion, and the hypothalamic-pituitary-gonadal axis are primarily enriched in signaling pathways such as PI3K-Akt, Hippo, and adrenergic. Further analysis combining methylation and transcriptomics identified five candidate methylated genes (CCL2, MYL2, GST, CTSH, and MCH) linked to adult body weight in Hezuo pigs. Additionally, the correlation analysis suggested that these genes influence growth and development in boars and sows by regulating the secretion and synthesis of related hormones, leading to heavier weights in females. In conclusion, variations in adult body weight between male and female pigs may stem from the impact of DNA methylation on gene expression related to growth and development. These findings offer new insights into the regulatory mechanisms of DNA methylation during weight gain in Hezuo pigs.

Oxidative stress compromises intestinal barrier function and antioxidant defense in weaned piglets, causing substantial economic losses in the pig industry. Probiotic interventions to improve gut health are considered a highly promising solution; research indicates that certain Lactobacillus species can promote the proliferation of Lgr5⁺ stem cells and activate the AhR/STAT3 pathway by modulating microbiota metabolism, thereby enhancing intestinal barrier and antioxidant function. Although Limosilactobacillus mucosae (L. mucosae) has demonstrated antioxidant properties in rodent models, its involvement in modulating intestinal stem cell (ISC) fate in piglets, as well as the associated mechanisms, remains unclear. We hypothesized that L. mucosae could alleviate oxidative stress in weaned piglets by modulating intestinal microbiota and activating the aryl hydrocarbon receptor/signal transducer and activator of transcription 3 (AhR/STAT3) signaling-mediated intestinal epithelial renewal. We compared the gut microbiota composition between Hezuo pigs (a subgroup of the plateau-type Tibetan breed) and Landrace × Yorkshire (LY) pigs, and isolated L. mucosae LM410 from Hezuo pigs. The strain was evaluated for acid and bile tolerance, antioxidant activity, and inhibitory properties. Subsequently, a 14-day animal trial was conducted using Duroc × Landrace × Yorkshire (DLY) weaned piglets orally administered with 1 × 10⁹ CFU/mL L. mucosae LM410. Intestinal morphology, barrier integrity, immune responses, antioxidant capacity and cecal microbiota composition and metabolic pathways were assessed. We found that Hezuo pigs harbored a higher abundance of L. mucosae in the gut compared with LY pigs. The isolate L. mucosae LM410 from Hezuo pigs, demonstrated strong acid and bile salt tolerance, as well as antioxidant and antimicrobial activities, and effectively improved intestinal morphology, barrier integrity, immune responses, and antioxidant capacity in DLY weaned piglets. Mechanistically, L. mucosae LM410 reshaped the cecal microbiota by enriching short-chain fatty acid (SCFA)-producing bacteria and upregulating the tyrosine metabolic pathway. Concomitantly, L. mucosae LM410 activated the AhR/STAT3 signaling axis, which likely promoted ISC proliferation and differentiation into goblet cells. These findings demonstrate for the first time that L. mucosae may enhance piglet antioxidant defense through coordinated cecal microbiome remodeling, upregulation of tyrosine metabolism and AhR/STAT3-driven epithelial renewal, providing a theoretical foundation for developing L. mucosae LM410 as a candidate probiotic to improve piglet health.