Search Results (15)

The aim of this study is to investigate the physiological characteristics and regulatory mechanisms of porcine intramuscular fat (IMF), subcutaneous fat (take back fat (BF), for example), and visceral fat (take perienteric fat (PF), for example) to address the challenge of optimizing meat quality without excessive fat deposition. Many improved breed pigs have fast growth rates, high lean meat rates, and low subcutaneous fat deposits, but they also have low IMF content, resulting in poor meat quality. There is usually a positive correlation between intramuscular fat and subcutaneous fat deposits. This study selected eight-month-old female Tibetan pigs as experimental subjects. After slaughter, fat samples were collected. Histological differences in adipocyte morphology were observed via hematoxylin–eosin (HE) staining of tissue sections, and phenotypic characteristics of different adipose tissues were analyzed through fatty acid composition determination. Transcriptome sequencing and untargeted metabolomics were employed to perform pairwise comparisons between different fatty tissues to identify differentially expressed genes and metabolites. A siRNA interference model was constructed and combined with Oil Red O staining and lipid droplet optical density measurement to investigate the regulatory role of WNT16 in adipocyte differentiation. Comparative analysis of phenotypic and fatty acid composition differences in adipocytes from different locations revealed that IMF adipocytes have significantly smaller areas and diameters compared to other fat depots and contain higher levels of monounsaturated fatty acids. Integrated transcriptomic and metabolomic analyses identified differential expression of WNT16 and L-tyrosine, both of which are involved in the melanogenesis pathway. Functional validation showed that inhibiting WNT16 in porcine preadipocytes downregulated adipogenic regulators and reduced lipid droplet accumulation. This cross-level regulatory mechanism of “phenotype detection–multi-omics analysis–gene function research” highlighted WNT16 as a potential key regulator of site-specific fat deposition, providing new molecular targets for optimizing meat quality through nutritional regulation and genetic modification. Full article

Porcine latissimus dorsi muscle (LDM) is a crucial source of pork products. Meat quality indicators, such as the proportion of muscle fibers and intramuscular fat (IMF) deposition, vary during the growth and development of pigs. Numerous studies have highlighted the heterogeneous nature of skeletal muscle, with phenotypic differences reflecting variations in cellular composition and transcriptional profiles. This study investigates the cellular-level transcriptional characteristics of LDM in large white pigs at two growth stages (170 days vs. 245 days) using single-nucleus RNA sequencing (snRNA-seq). We identified 56,072 cells across 12 clusters, including myofibers, fibro/adipogenic progenitor (FAP) cells, muscle satellite cells (MUSCs), and other resident cell types. The same cell types were present in the LDM at both growth stages, but their proportions and states differed. A higher proportion of FAPs was observed in the skeletal muscle of 245-day-old pigs. Additionally, these cells exhibited more active communication with other cell types compared to 170-day-old pigs. For instance, more interactions were found between FAPs and pericytes or endothelial cells in 245-day-old pigs, including collagen and integrin family signaling. Three subclasses of FAPs was identified, comprising FAPs_COL3A1⁺, FAPs_PDE4D⁺, and FAPs_EBF1⁺, while adipocytes were categorized into Ad_PDE4D⁺ and Ad_DGAT2⁺ subclasses. The proportions of these subclasses differed between the two age groups. We also constructed differentiation trajectories for FAPs and adipocytes, revealing that FAPs in 245-day-old pigs differentiated more toward fibrosis, a characteristic reminiscent of the high prevalence of skeletal muscle fibrosis in aging humans. Furthermore, the Ad_PDE4D⁺ adipocyte subclass, predominant in 245-day-old pigs, originated from FAPs_PDE4D⁺ expressing the same gene, while the Ad_DGAT2⁺ subclass stemmed from FAPs_EBF1⁺. In conclusion, our study elucidates transcriptional differences in skeletal muscle between two growth stages of pigs and provides insights into mechanisms relevant to pork meat quality and skeletal muscle diseases. Full article

Porcine skeletal muscle development is closely linked to meat production efficiency and quality. The accumulation of porcine intramuscular fat is influenced by the hyperplasia and hypertrophy of adipocytes within the muscle. However, the cellular profiles corresponding to the two stages of muscle development remain undetermined. Single-nucleus RNA sequencing (snRNA-seq) can elucidate cell subsets in tissues, capture gene expression at the individual cell level, and provide innovative perspectives for studying muscle and intramuscular fat formation. In this study, a total of 78,302 nuclei and 9 clusters of cells, which included fibro/adipogenic progenitor (FAP), myonuclei, adipocytes, and other cell types, of Xidu black pigs, were identified on Day 1 and Day 180. The pattern of cell clustering varied between the two developmental stages. Notably, the percentage of adipocytes in the Day 180 group was higher than in the Day 1 group (0.51% vs. 0.15%). Pseudo-time sequence analysis indicated that FAPs could differentiate into adipocytes and myonuclei cells, respectively. The THRSP gene was identified as a biomarker for swine intramuscular fat cells, and its down-regulation resulted in significant reduction in lipid droplet formation in porcine preadipocytes. Our research provides new insights into the cellular characteristics of intramuscular fat formation, which may facilitate the development of novel strategies to enhance intramuscular fat deposition and improve pork quality. Full article

Intramuscular fat (IMF) content plays a crucial role in determining pork quality. Recent studies have highlighted transcriptional mechanisms controlling adipogenesis in porcine IMF. However, the changes in chromatin accessibility during adipogenic differentiation are still not well understood. In this study, we performed the assay for transposase-accessible chromatin with high-throughput sequencing (ATAC-seq) and transcriptome sequencing (RNA-Seq) analyses on porcine intramuscular preadipocytes to explore their adipogenic differentiation into mature adipocytes. We identified a total of 56,374 differentially accessible chromatin peaks and 4226 differentially expressed genes at day 0 and day 4 during adipogenic differentiation. A combined analysis of the ATAC-seq and RNA-seq data revealed that 1750 genes exhibited both differential chromatin accessibility and differential RNA expression during this process, including selenium-binding protein 1 (SELENBP1), PLIN1, ADIPOQ, and FASN. Furthermore, we found that vitamin D receptor (VDR) could bind to the promoter region of the SELENBP1 gene, activate SELENBP1 transcription, and ultimately promote lipid accumulation during adipogenic differentiation. This study provides a detailed overview of chromatin accessibility and gene expression changes during the adipogenic differentiation of porcine intramuscular preadipocytes. Moreover, we propose a novel regulatory mechanism involving the VDR–SELENBP1 signaling axis in adipogenic differentiation. Full article

Obesity and its associated metabolic disease do serious harm to human health. The transcriptional cascade network with transcription factors as the core is the focus of current research on adipogenesis and its mechanism. Previous studies have found that HMG domain protein 20A (HMG20A) is highly expressed in the early stage of adipogenic differentiation of porcine intramuscular fat (IMF), which may be involved in regulating adipogenesis. In this study, HMG20A was found to play a key negative regulatory role in adipogenesis. Gain- and loss-of-function studies revealed that HMG20A inhibited the differentiation of SVF cells and C3H10T1/2 cells into mature adipocytes. RNA-seq was used to screen differentially expressed genes after HMG20A knockdown. qRT-PCR and ChIP-PCR confirmed that MEF2C was the real target of HMG20A, and HMG20A played a negative regulatory role through MEF2C. HMG20A binding protein LSD1 was found to alleviate the inhibitory effect of HMG20A on adipogenesis. Further studies showed that HMG20A could cooperate with LSD1 to increase the H3K4me2 of the MEF2C promoter and then increase the expression of MEF2C. Collectively, these findings highlight a role for HMG20A-dependent transcriptional and epigenetic regulation in adipogenesis. Full article

The intramuscular fat is a major quality trait of meat, affecting sensory attributes such as flavor and texture. Several previous GWAS studies identified Acyl-CoA Synthetase Long Chain Family Member 4 (ACSL4) gene as the candidate gene to regulate intramuscular fat content in different pig populations, but the underlying molecular function of ACSL4 in adipogenesis within pig skeletal muscle is not fully investigated. In this study, we isolated porcine endogenous intramuscular adipocyte progenitors and performed ACSL4 loss- and gain-of-function experiments during adipogenic differentiation. Our data showed that ACSL4 is a positive regulator of adipogenesis in intramuscular fat cells isolated from pigs. More interestingly, the enhanced expression of ACSL4 in pig intramuscular adipocytes could increase the cellular content of monounsaturated and polyunsaturated fatty acids, such as gamma-L eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA). The above results not only confirmed the function of ACSL4 in pig intramuscular adipogenesis and meat quality attributes, but also provided new clues for the improvement of the nutritional value of pork for human health. Full article

miRNA is a small non-coding RNA, which plays an important role in diverse biological processes. In the present study, we explore the effect of ssc-miR-451 on porcine adipose development and meat quality. We observed that ssc-miR-451 was downregulated during porcine primary adipocyte differentiation. Overexpression of ssc-miR-451 inhibited adipogenic differentiation, while inhibition of ssc-miR-451 promoted adipogenic differentiation. The dual luciferase reporter system indicated Acetyl-CoA carboxylase alpha (ACACA) as a target gene of ssc-miR-451. Correlation analysis negatively correlated miR-451 expression with intramuscular fat content (IMF) and positively correlated ACACA expression with IMF. Further analysis of fatty acid composition revealed that pigs with high expression of ssc-miR-451 had higher monounsaturated fatty acid (MUFA) and lower polyunsaturated fatty acid (PUFA). Taken together, our study suggests that ssc-miR-451 regulates lipid deposition and fatty acid composition by targeting ACACA, and ssc-miR-451 may serve as a potential genetic marker to improve pork quality. Full article

The degree of fat accumulation and adipokine production are two major indicators of obesity that are correlated with increased adipose tissue mass and chronic inflammatory responses. Adipocytes have been considered effector cells for the inflammatory responses due to their capacity to express Toll-like receptors (TLRs). In this study, we evaluated the degree of fat accumulation and adipokine production in porcine intramuscular preadipocyte (PIP) cells maintained for in vitro differentiation over a long period without or with stimulation of either TNF-α or TLR2-, TLR3-, or TLR4-ligands. The cytosolic fat accumulation was measured by liquid chromatography and the expression of adipokines (CCL2, IL-6, IL-8 and IL-10) were quantified by RT-qPCR and ELISA at several time points (0 to 20 days) of PIP cells differentiation. Long-term adipogenic differentiation (LTAD) induced a progressive fat accumulation in the adipocytes over time. Activation of TLR3 and TLR4 resulted in an increased rate of fat accumulation into the adipocytes over the LTAD. The production of CCL2, IL-8 and IL-6 were significantly increased in unstimulated adipocytes during the LTAD, while IL-10 expression remained stable over the studied period. An increasing trend of adiponectin and leptin production was also observed during the LTAD. On the other hand, the stimulation of adipocytes with TLRs agonists or TNF-α resulted in an increasing trend of CCL2, IL-6 and IL-8 production while IL-10 remained stable in all four treatments during the LTAD. We also examined the influences of several immunoregulatory probiotic strains (immunobiotics) on the modulation of the fat accumulation and adipokine production using supernatants of immunobiotic-treated intestinal immune cells and the LTAD of PIP cells. Immunobiotics have shown a strain-specific ability to modulate the fat accumulation and adipokine production, and differentiation of adipocytes. Here, we expanded the utility and potential application of our in vitro PIP cells model by evaluating an LTAD period (20 days) in order to elucidate further insights of chronic inflammatory pathobiology of adipocytes associated with obesity as well as to explore the prospects of immunomodulatory intervention for obesity such as immunobiotics. Full article

Intramuscular fat (IMF) is implicated in juiciness, tenderness, and flavor of pork. Meat quality of Chinese fat-type pig is much better than that of lean-type pig because of its higher IMF content. LncRNA is a vital regulator that contributes to adipogenesis. However, it is unknown about the regulation of lncRNA on IMF content. Here, by RNA sequence analysis of intramuscular adipocyte from Bamei pig (fat-type) and Yorkshire pig (lean-type), we found that a novel lncRNA, lncIMF4, was associated with adipogenesis. LncIMF4, abundant in adipose, differently expressed along with intramuscular preadipocyte proliferation and differentiation. Meanwhile, it is located both in cytoplasm and nucleus. Besides, lncIMF4 knockdown promoted proliferation and differentiation of porcine intramuscular preadipocytes, whereas inhibited autophagy. Moreover, lncIMF4 knockdown facilitated intramuscular adipogenesis through attenuating autophagy to repress the lipolysis. Our findings will contribute to understand better the mechanism of lncRNA controlling intramuscular adipogenesis for promoting pork quality. Full article

The meat quality of animal products is closely related to the intramuscular fat content. Aquaglyceroporin (AQP) defines a class of water/glycerol channels that primarily facilitate the passive transport of glycerol and water across biological membranes. In this study, the AQP3 protein of the AQP family was mainly studied in the adipogenic function of intramuscular adipocytes in pigs. Here, we found that AQP3 was increased at both mRNA and protein levels upon adipogenic stimuli in porcine intramuscular adipocytes in vitro. Western blot results showed knockdown of AQP3 by siRNA significantly suppressed the expression of adipogenic genes (PPARγ, aP2, etc.), repressed Akt phosphorylation, as well as reducing lipid accumulation. Furthermore, deletion of AQP3 by siRNA significantly downregulated expression of cell cycle genes (cyclin D, E), and decreased the number of EdU-positive cells as well as cell viability. Collectively, our data indicate that AQP3 is of great importance in both adipogenic differentiation and proliferation in intramuscular adipocytes, providing a potential target for modulating fat infiltration in skeletal muscles. Full article

Adipocytes are dynamic cells that have critical functions to maintain body energy homeostasis. Adipocyte physiology is affected by the adipogenic differentiation, cell program, as well as by the exogenous stimulation of biochemical factors, such as serotonin and TNF-α. In this work, we investigated the global transcriptome modifications when porcine intramuscular preadipocyte (PIP) was differentiated into porcine mature adipocyte (pMA). Moreover, we studied transcriptome changes in pMA after stimulation with serotonin or TNF-α by using a microarray approach. Transcriptome analysis revealed that the expression of 270, 261, and 249 genes were modified after differentiation, or after serotonin and TNF-α stimulation, respectively. Expression changes in APP, HNF4A, ESR1, EGR1, SRC, HNF1A, FN1, ALB, STAT3, CBL, CEBPB, AR, FOS, CFTR, PAN2, PTPN6, VDR, PPARG, STAT5A and NCOA3 genes which are enriched in the ‘PPAR signaling’ and ‘insulin resistance’ pathways were found in adipocytes during the differentiation process. Dose-dependent serotonin stimulation resulted in a decreased fat accumulation in pMAs. Serotonin-induced differentially expressed genes in pMAs were found to be involved in the significant enrichment of ′GPCR ligand-binding′, ‘cell chemotaxis’, ‘blood coagulation and complement’, ‘metabolism of lipid and lipoproteins’, ‘regulation of lipid metabolism by PPARA’, and ‘lipid digestion, mobilization and transport’ pathways. TNF-α stimulation also resulted in transcriptome modifications linked with proinflammatory responses in the pMA of intramuscular origin. Our results provide a landscape of transcriptome modifications and their linked-biological pathways in response to adipogenesis, and exogenous stimulation of serotonin- and TNF-α to the pMA of intramuscular origin. Full article

Intramuscular fat (IMF) content is an important indicator for meat quality evaluation. However, the key genes and molecular regulatory mechanisms affecting IMF deposition remain unclear. In the present study, we identified 75 differentially expressed genes (DEGs) between the higher (H) and lower (L) IMF content of pigs using transcriptome analysis, of which 27 were upregulated and 48 were downregulated. Notably, Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis indicated that the DEG perilipin-1 (PLIN1) was significantly enriched in the fat metabolism-related peroxisome proliferator-activated receptor (PPAR) signaling pathway. Furthermore, we determined the expression patterns and functional role of porcine PLIN1. Our results indicate that PLIN1 was highly expressed in porcine adipose tissue, and its expression level was significantly higher in the H IMF content group when compared with the L IMF content group, and expression was increased during adipocyte differentiation. Additionally, our results confirm that PLIN1 knockdown decreases the triglyceride (TG) level and lipid droplet (LD) size in porcine adipocytes. Overall, our data identify novel candidate genes affecting IMF content and provide new insight into PLIN1 in porcine IMF deposition and adipocyte differentiation. Full article

Intramuscular fat (IMF) content and composition are considered crucial indicators of porcine meat quality. However, the molecular mechanism of porcine IMF development is still mostly unclear. Recently, new evidence suggested that microRNA (miRNAs) play important roles in porcine intramuscular adipogenesis. Previously, microRNA-125a-5p (miR-125a-5p) was identified as an important regulator of adipogenesis. In the present study, we found that the expression of miR-125a-5p is dynamically regulated during porcine intramuscular preadipocytes differentiation and that its expression levels in different porcine muscle tissues were negatively involved with IMF content. To investigate the potential function role of miR-125a-5p in IMF development, porcine intramuscular preadipocytes were collected and transfected with miR-125a-5p mimics, inhibitors, or a negative control (NC), respectively. The results showed that overexpression of miR-125a-5p promoted proliferation and inhibited differentiation of porcine intramuscular preadipocytes while inhibition of miR-125a-5p had the opposite effects. Furthermore, a luciferase reporter assay demonstrated that porcine kruppel like factor 3 (KLF13) is a target gene of miR-125a-5p during porcine intramuscular preadipocytes differentiation. Interestingly, porcine ELOVL fatty acid elongase 6 (ELOVL6), a regulator of fatty acid composition, was also identified as a target gene of miR-125a-5p during porcine intramuscular adipogenesis. Further studies show that miR-125a-5p overexpression reduced total saturated fatty acids (SFA) content and monounsaturated fatty acids (MUFA)/SFA ratios while having no significant impact on polyunsaturated fatty acids (PUFA)/SFA and n-6/n-3 ratios. Taken together, our results identified that miR-125a-5p may be a novel regulator of porcine intramuscular adipogenesis and the fatty acid composition of porcine IMF. Full article

Suitable intramuscular fat (IMF) content improves porcine meat quality. The vital genes regulating IMF deposition are necessary for the selection and breeding of an IMF trait. However, the effect and mechanism of PDGFRα on IMF deposition are still unclear. Here, PDGFRα is moderately expressed in porcine longissimus dorsi muscle (LD), whereas it highly expressed in white adipose tissue (WAT). Moreover, PDGFRα-positive cells were located in the gaps of LD fibers which there were IMF adipocytes. Compared with 180-day-old and lean-type pigs, the levels of PDGFRα were much higher in one-day-old and fat-type pigs. Meanwhile the levels of PDGFRα gradually decreased during IMF preadipocyte differentiation. Furthermore, PDGFRα promoted adipogenic differentiation through activating Erk signaling pathway. Based on PDGFRα upstream regulation analysis, we found that the knockdown of FoxO1 repressed lipogenesis by downregulating PDGFRα, and miR-34a inhibited adipogenesis through targeting PDGFRα. Collectively, PDGFRα is a positive regulator of IMF deposition. Therefore, we suggest that PDGFRα is a possible target to improve meat quality. Full article

Due to the paracrine effects of skeletal muscle, the lipid metabolism of porcine intramuscular (i.m.) preadipocytes was different from that of subcutaneous (s.c.) preadipocytes. To investigate the development of i.m. preadipocytes in vivo, the s.c. preadipocytes were cultured with muscle conditional cultured medium (MCM) for approximating extracellular micro-environment of the i.m. preadipocytes. Insulin signaling plays a fundamental role in porcine adipocyte differentiation. The expression levels of insulin receptor (INSR) and insulin-like growth factor 1 receptor (IGF-1R) in i.m. Preadipocytes were higher than that in s.c. preadipocytes. The effects of MCM on adipocyte differentiation, lipid metabolism and insulin signaling transdution were verified. MCM induced the apoptosis of s.c. preadipocytes but not of s.c. adipocytes. Moreover, MCM inhibited adipocyte differentiation at pre-differentiation and early stages of differentiation, while the expression levels of INSR and IGF-1R were increased. Furthermore, MCM treatment increased adipocyte lipolysis and fatty acid oxidation through induction of genes involved in lipolysis, thermogenesis, and fatty acid oxidation in mitochondria. Consistent with the above, treatment of s.c. adipocytes with MCM upregulated mitochondrial biogenesis. Taken together, MCM can approximate the muscle micro-environment and reduce intramuscular adipocyte differentiation and lipid accumulation via regulating insulin signaling. Full article