Long non-coding RNAs (lncRNAs) are emerging as important regulators of plant gene expression, affecting developmental processes, stress tolerance, and organ growth. Their involvement in the determination of seed size is yet to be explored in litchi (Litchi chinensis). An in-depth lncRNA profiling of developing seeds of large- and small-seeded litchi cultivars was conducted and 1,014 lncRNAs, of which 88 were found to be differentially expressed, was discovered. A few lncRNAs, such as XLOC_030992, XLOC_021376, and XLOC_025154, exhibited remarkable regulatory capacity through co-expression or anti-correlation with genes of hormone signalling, intracellular transport, proteostasis, lipid metabolism, and cell cycle regulation. Interestingly, XLOC_030992 appeared to be a hub trans-acting regulator of more than 180 target genes, some of which are implicated in ABA signaling, mitotic progression, and seed viability. Others, including XLOC_009460 and XLOC_006988, showed stage- and genotype-specific expression, indicating dynamic functions in early embryogenesis. This study’s results suggested a regulatory model in which lncRNAs regulate seed development by cis- and trans-regulating important genes. This research provided the first evidence for the role of lncRNAs in litchi seed size determination and provided a basis for functional studies to improve seed quality in fruit crops.
Nodal stem explants of Epidendrum radicans Pav. ex Lindl. showed two distinct in vitro regeneration routes under different plant growth regulators. This study compared these routes, characterized the 2,4-D-induced structures histologically, and evaluated their use in a two-stage micropropagation strategy. Stable aseptic cultures were obtained most effectively with 0.1
Diabetic vascular complications are a leading cause of disability in diabetic. This study aimed to investigate the functional improvement effects and molecular mechanisms of internalizing RGD (iRGD) peptide-modified ferulic acid-targeted liposomes (Lp-iRGD@FA) on high glucose-induced injured endothelial progenitor cells (EPCs). Lp-iRGD@FA was prepared using the thin-film hydration method and subsequently characterized. In vitro experiments were conducted using human peripheral blood-derived EPCs, in which a high glucose injury model was established. Cells were treated with free ferulic acid, non-targeted liposomes, and targeted liposomes, respectively. In vivo, a diabetic hindlimb ischemia mouse model was established, and Lp-iRGD@FA, non-targeted liposomes, or Lp-iRGD were intramuscularly injected into the ischemic muscles. The high glucose environment significantly inhibited EPC proliferation, promoted apoptosis, and impaired tube formation ability, accompanied by inhibited AMPK phosphorylation and downregulated expression of KLF4 and FAM3A. Intervention with the targeted liposomes effectively reversed these changes. However, the AMPK inhibitor Compound C blocked the Lp-iRGD@FA-induced upregulation of KLF4 and FAM3A and the associated functional improvements, whereas KLF4 supplementation partially rescued the decreased FAM3A expression and impaired tube formation caused by AMPK inhibition. In vivo, Lp-iRGD@FA treatment in diabetic mice enhanced angiogenesis in a hindlimb ischemia model and activated the AMPK/KLF4/FAM3A pathway in ischemic muscles. This study demonstrates that Lp-iRGD@FA efficiently delivers ferulic acid to restore FAM3A expression by activating the AMPK/KLF4/FAM3A signaling axis, subsequently upregulating VEGFA/VEGFR2 levels and improving the function of EPCs under high glucose stress. These findings provide a novel nanomedicine strategy and potential molecular targets for the treatment of diabetic vascular complications.
Given the involvement of long non-coding RNA-differentiation antagonizing non-protein coding RNA (lncRNA DANCR) in osteoporosis (OP), this study aimed to investigate its role in osteogenic differentiation (OD) of bone marrow mesenchymal stem cells (BMSCs) and underlying mechanisms. BMSCs were intervened with sh-DANCR, oe-DANCR, sh-phosphatase and tensin homolog (PTEN) lentivirus, miR-19a-3p agomir, or MK-2206 dihydrochlorode. The number of mature osteoblasts, osteogenic ability, and alkaline phosphatase (ALP) activity were assessed by ALP staining, alizarin red staining, and kit assay. Bioinformatics prediction, dual-luciferase assay, and RNA pull-down assay were conducted to assess the binding between lncRNA DANCR and miR-19a-3p and that between PTEN and miR-19a-3p. The levels of lncRNA DANCR, osteogenesis-related proteins (osteocalcin (OCN), runt-related transcription factor 2 (RUNX2), osteopontin (OPN)), PTEN, phosphorylated-protein kinase B (p-AKT), AKT, phosphorylated-mammalian target of rapamycin (p-mTOR), and mTOR were determined by RNA quantitation and western blot techniques. Silencing of lncRNA DANCR potentiated the OD capability of BMSCs, as evidenced by increased ALP activity, calcium nodule formation, and elevated expression of OPN, RUNX2, and OCN. In contrast, lncRNA DANCR overexpression inhibited the OD function by downregulating miR-19a-3p. Mechanistically, lncRNA DANCR regulated PTEN expression by competitively binding to miR-19a-3p. Additionally, PTEN silencing partially reversed the anti-OD effect of lncRNA DANCR. LncRNA DANCR knockdown activated the AKT/mTOR pathway, whereas lncRNA DANCR overexpression or MK-2206 treatment disrupted this activation. Silencing of lncRNA DANCR facilitates BMSC OD by binding to miR-19a-3p and further regulating the PTEN/AKT/mTOR pathway.
Ginsenoside Rc (G-Rc), a major active ingredient of Panax ginseng, has protective effects against various pathological alterations associated with diseases. Nonetheless, the potential benefits of G-Rc in treating cerebral ischemia–reperfusion (CIR) injury remain inadequately explored. In the present study, we evaluated the potential effects of G-Rc on CIR injury via cellular and animal models and explored the underlying mechanisms. Oxygen–glucose deprivation and reoxygenation (OGD/R)-induced neurons treated with G-Rc exhibited a marked reduction in apoptosis, oxidative stress, and inflammation. Rats treated with G-Rc presented significant reductions in neurological deficits, cerebral infarction, and pathological alterations resulting from CIR injury. Moreover, neuronal death, oxidative stress damage, and the inflammatory response in the brains of rats with CIR injury were markedly ameliorated by G-Rc treatment. Molecular docking, molecular dynamics simulations, and relevant experiments validated Sirtuin 1 (Sirt1) as a target of G-Rc. G-Rc increased Sirt1 expression and augmented its enzymatic activity. Notably, G-Rc promoted the activation of nuclear factor-erythroid 2-related factor 2 (Nrf2) by targeting Sirt1. G-Rc failed to induce Nrf2 activation in Sirt1-silenced neurons. Furthermore, the benefits of G-Rc in defending against OGD/R damage were strongly negated by pharmacologically suppressing Nrf2. These observations indicate that G-Rc offers considerable benefits in treating CIR injury in experimental models. These beneficial effects are achieved by enhancing Nrf2 activation via the targeting of Sirt1. This study emphasizes the potential utilization of G-Rc as a pharmaceutical candidate for treating CIR damage.
Meconopsis aculeata Royle holds ecological, medicinal, and ornamental value yet faces conservation challenges due to limited natural populations and insufficient propagation techniques. Therefore, the present investigation evaluates the seed germination, in vitro propagation, acclimatization, and biochemical characteristics of in vitro grown plants of M. aculeata. Optimal seed germination was accomplished at 25°C on MS basal medium and with a maximum germination percentage (approximately 70
Pineapple (Ananas comosus L. Merr., cultivar MD2) is a key tropical crop but highly sensitive to salinity and drought, making temporary immersion bioreactors (TIBs) valuable for micropropagation and stress studies. Apical buds were cultured in vitro, transferred to TIBs with paclobutrazol, and subjected to nine treatments (control, NaCl, mannitol at 50.0 to 200.0 mM) for 30 d. Growth and biochemical traits were measured, data normalized (0 to 1), and analyzed by hierarchical cluster analysis (HCA) using average linkage and squared Euclidean distance. The dendrogram revealed clear treatment similarities: controls remained distinct until higher rescaled distances, low concentrations clustered together, and high concentrations formed separate groups, confirming dose-dependent responses. NaCl and mannitol did not cluster together initially, indicating distinct physiological profiles, with NaCl imposing additional ionic toxicity beyond osmotic effects. These findings complement earlier evaluations showing significant reductions in shoot multiplication and biomass above 50.0 mM, with critical concentrations for 50
The southern green stink bug, Nezara viridula L., is a highly polyphagous global pest that causes substantial economic losses on a broad range of crops. Current management strategies rely heavily on chemical insecticides, which can promote resistance development and negatively affect non-target beneficial organisms. The lack of a tractable in vitro cell culture system has limited the development of more targeted and sustainable control approaches for this species. In this study, we identified the optimal egg age for culture initiation and evaluated seven insect cell culture media or medium combinations, as well as several supplements, for their ability to support embryo-derived N. viridula primary cultures. A total of 200 primary cultures were initiated using 6,000 dissected N. viridula eggs over 16 months. Of the media tested, GS medium, a combination of CLG#2 and Shields and Sang medium, supplemented with 9
Coronary artery disease (CAD) involves intricate immune-related pathways; however, the contribution of epigenetic mechanisms remains inadequately defined. The fat mass and obesity-associated protein (FTO), which functions as an N⁶-methyladenosine (m⁶A) demethylase, has been implicated in the progression of atherosclerotic conditions. This study sought to elucidate how FTO downregulates proteinase 3 (PRTN3) through m⁶A demethylation and inhibits neutrophil activation via the C-X-C motif chemokine ligand 9/C-X-C motif chemokine receptor 3 (CXCL9/CXCR3) signaling pathway in the context of CAD. RNA sequencing was carried out on peripheral blood mononuclear cells (PBMCs) obtained from CAD patients and healthy individuals to detect gene expression differences. Functional enrichment analyses, including Gene Ontology and Kyoto Encyclopedia of Genes and Genomes, were performed along with experimental validation in endothelial progenitor cells (EPCs). The regulatory relationship between FTO and PRTN3 was examined via luciferase reporter assays, RNA immunoprecipitation, and methylated RNA immunoprecipitation quantitative polymerase chain reaction. Neutrophil activation was evaluated by measuring CXCL9/CXCR3 expression, tracking cell migration, and assessing reactive oxygen species (ROS) generation in HL-60 cells. For in vivo validation, apolipoprotein E-deficient (ApoE⁻/⁻) mice were maintained on a high-fat diet and treated with the AAV9 vector carrying FTO via tail vein injection to evaluate effects on atherosclerotic development. Findings indicated that PRTN3 is significantly upregulated in CAD patients, which was corroborated in EPCs. FTO was shown to directly bind to PRTN3 and decrease its expression by reducing m⁶A methylation. Overexpression of FTO enhanced proliferation and migration of EPCs and reduced apoptosis, whereas FTO silencing produced opposing outcomes. Furthermore, PRTN3 was found to stimulate the CXCL9/CXCR3 axis, leading to increased neutrophil migration and ROS production. In vivo, FTO inhibits the activation of neutrophils by down-regulating the expression of PRTN3, reduces the inflammatory response, and protects the occurrence and development of atherosclerosis in mice. These results uncover a novel regulatory pathway involving FTO, m⁶A, PRTN3, and CXCL9/CXCR3 in CAD pathogenesis, highlighting FTO as a promising target for therapeutic intervention.
Hepatocellular carcinoma (HCC) has a poor prognosis and limited treatment options. This study investigated the role and mechanism of Jiedu Xiaozheng Yin (JXY) in HCC using network pharmacology and in vivo/in vitro experiments. Active components of JXY and their potential therapeutic targets for HCC were identified via network pharmacology. A protein–protein interaction (PPI) network was constructed to identify core targets, and bioinformatics analysis confirmed their association with HCC. LC–MS/MS was used to isolate major components, which were then subjected to molecular docking with core targets. The effects of JXY on HCC were further validated through in vivo and in vitro experiments. Analysis of 171 core target genes was performed using PPI network screening and enrichment analysis. Quercetin was identified as the primary active component of JXY. Molecular docking confirmed its interaction with CYP1A2, which is overexpressed in HCC tissues. In vivo, JXY and quercetin significantly increased CYP1A2 and PTEN expression, while decreasing p-PI3K and p-Akt levels in HCC-bearing mice, thereby suppressing tumor growth and lung metastasis and promoting apoptosis. In vitro, both JXY and quercetin inhibited HCC cell proliferation and migration. CYP1A2 knockdown partially reversed, while its overexpression enhanced, these inhibitory effects. Quercetin, the main active component of JXY, upregulates CYP1A2 to promote PTEN expression, thereby inhibiting the PI3K/Akt signaling pathway and suppressing HCC progression.
Intramuscular fat deposition is a key determinant of meat quality in livestock species; however, yak (Bos grunniens) stromal vascular cells (SVCs) exhibit relatively low adipogenic differentiation efficiency under conventional in vitro conditions. In this study, we established a three-dimensional (3D) spheroid culture system to improve adipogenic differentiation of yak SVCs and compared it with traditional two-dimensional (2D) monolayer culture. Yak SVCs were isolated from longissimus dorsi muscle and characterized prior to differentiation. Cells were subjected to adipogenic induction under both 2D and 3D conditions. Adipogenic differentiation was evaluated by morphological observation, Oil Red O staining and quantification, and quantitative real-time PCR (RT-qPCR) analysis of adipogenic marker genes. Additionally, Western blot analysis of key adipogenic proteins was performed specifically on yak SVCs. To validate the induction system, murine C3H10T1/2 cells were included as a parallel control group and evaluated using the aforementioned non-protein assays. The results showed that 3D culture significantly enhanced lipid accumulation and increased the expression of adipogenic marker genes compared with 2D culture in yak SVCs. Consistently, protein levels of PPAR γ were significantly elevated in the 3D group of yak SVCs, while FABP4 showed an upward trend. Furthermore, the C3H10T1/2 control group exhibited successful adipogenic differentiation with similarly enhanced lipid accumulation and gene expression under 3D conditions, confirming the reliability of our system. In conclusion, 3D spheroid culture improves adipogenic differentiation of yak SVCs and provides a useful in vitro model for studying adipose development in yak. This system may facilitate future studies on the regulation of intramuscular fat deposition in livestock species.
Utilizing single-cell RNA sequencing and network pharmacology, this study explored the mechanism by which Bletilla striata polysaccharide (BSP) mitigates peri-implantitis-associated osteolysis. Integrated analyses revealed a significant expansion of pro-inflammatory macrophages and osteoclasts in diseased tissues, with differentially expressed genes enriched in glycolytic pathways. Mechanistic exploration indicated that BSP suppresses osteoclastogenesis primarily by targeting matrix metalloproteinase-9 (MMP9), thereby inhibiting proteolytic activity and cytoskeletal remodeling. Preliminary rescue experiments supported this notion, showing that while BSP effectively downregulated LPS-induced MMP9 upregulation and suppressed macrophage glycolytic metabolism, forced overexpression of MMP9 partially restored MMP9 levels and reversed the inhibitory effect of BSP on osteoclast formation. These findings suggest a potential “metabolism-protease” interplay, where BSP concurrently modulates immune metabolism and protease activity to alleviate inflammatory bone loss, providing a foundational basis for developing plant-derived polysaccharide therapeutics.
The aim of this study was to isolate human periapical cyst-derived mesenchymal stem cells (PC-MSCs) and compare their immunophenotypic profiles, clonogenic capacity, metabolic activity, migration capacity, and differentiation characteristics with those of dental pulp mesenchymal stem cells (DP-MSCs). PC-MSCs and DP-MSCs were isolated from six periapical cysts and six impacted third molars, respectively. The cells were characterized by flow cytometry and trilineage differentiation assays (osteogenic, adipogenic, chondrogenic). Proliferation was assessed using the MTT assay and colony-forming unit-fibroblast (CFU-F) efficiency. Migration potential was evaluated using an in vitro scratch assay. The secretion of odontogenic markers, dentin sialophosphoprotein (DSPP) and dentin matrix acidic phosphoprotein (DMP1), was quantified using enzyme-linked immunosorbent assay (ELISA). Both cell types expressed characteristic mesenchymal stem cell markers. PC-MSCs exhibited significantly higher colony-forming efficiency (CFU-F, p < 0.05) and superior migration capacity compared to DP-MSCs (p < 0.05). Both MSC populations demonstrated differentiation capacity. DP-MSCs showed more extensive Alizarin Red S staining, while PC-MSCs showed relatively stronger Oil Red O staining. ELISA analysis revealed no statistically significant differences in DSPP and DMP1 secretion levels between the two groups (p > 0.05). Although DP-MSCs exhibited superior mineralizing ability, PC-MSCs displayed higher clonogenic and migratory potential, while no statistically significant differences were observed in DSPP and DMP1 secretion levels between the groups. These findings suggest that periapical cysts, often regarded as pathological waste, may represent a promising and readily accessible source of MSC-like cells with potential relevance for regenerative endodontic research.
Mild thermal stimulation enhances skeletal muscle differentiation; however, its underlying metabolic basis remains unclear. Here, we demonstrate that thermal stimulation promotes myogenic differentiation through enhanced glucose uptake and transient lipid droplet (LD) accumulation in C2C12 myoblasts. Thermal stimulation at 39 °C induced a transient increase in LD formation during early differentiation. Early transient LD accumulation, particularly on days 1 and 2, was positively correlated with the day 5 fusion index, suggesting that early LD formation is associated with subsequent myogenic differentiation. Suppression of LD formation by Plin2 knockdown impaired myotube formation, indicating that LD formation is functionally required for myogenic differentiation. Thermal stimulation enhanced glucose uptake, accompanied by increased Slc2a4 and Ppargc1a expression and increased GLUT4 fluorescence intensity. Moreover, higher glucose availability further supported LD formation and myogenic differentiation even under serum-free conditions. Notably, mitochondrial mass, ATP content, and oxidative capacity remained largely unchanged, suggesting that enhanced glucose uptake is associated with LD accumulation without a corresponding increase in mitochondrial oxidative capacity. These findings identify transient LD formation as a key metabolic event driving myogenic differentiation under thermal stimulation.
Artemisia vulgaris L. has attracted considerable interest for its pharmaceutical, agricultural, and cosmetic applications due to its rich bioactive metabolites. However, low metabolite yields and challenges in large-scale cultivation constrain its commercial potential. Therefore, this study aimed to optimize a callus regeneration system, develop a comprehensive metabolite profile of the callus culture, and assess the effects of elicitor treatments on metabolite biosynthesis pathways to identify optimal concentrations for enhanced metabolite production. In vitro callus cultures were initiated on Murashige and Skoog (MS) medium with varying levels of benzylaminopurine (BAP), α-naphthaleneacetic acid (NAA), and 2,4-dichlorophenoxyacetic acid (2,4-D) phytohormones. Callus cultures were evaluated for colour, texture, moisture content, and biomass, and metabolite profiling was conducted using nuclear magnetic resonance (NMR) platform. Elicitor treatments of the callus cultures with different concentrations of methyl jasmonate (MJ) and salicylic acid (SA) were analysed via NMR-based metabolomics. MS medium containing 2.0 mg L⁻1 BAP and 0.2 mg L⁻1 NAA was optimal for callus culture establishment and maintenance. A total of eighteen compounds were identified, including dihydroartemisinic acid, caffeic acid, and gallic acid. MJ and SA functioned as stress elicitors, modulating the tricarboxylic acid cycle, phenylpropanoid, and artemisinin biosynthesis pathways, although they had limited direct effects on artemisinin concentrations. Overall, this study optimized callus culture production, generated the first detailed metabolite profile, and highlighted the potential of elicitation strategies to enhance bioactive metabolite yields, providing a foundation for future metabolic engineering and industrial applications.
An efficient micropropagation method based on axillary shoot bud proliferation was developed for Barleria prionitis, an important Indian woody medicinal plant. Highest (100
Colorectal cancer (CRC) is a malignant tumor with a high global incidence. Tianma granule (TMG) is a traditional Chinese medicine compound. However, its pharmacodynamic material basis and mechanism of action against CRC remained unclear. This study systematically analyzed the anti-CRC mechanism of TMG by integrating metabolomics, network pharmacology, and experimental validation. HPLC-MS identified the main active components of TMG. Core targets and pathways were predicted by network pharmacology. The binding affinity between components and targets was evaluated by molecular docking. In vitro experiments utilized the HCT116 and HCT-8 cells model, where the effects of TMG and its key component Apigenin on cell activity. Apigenin was identified as a key flavonoid component in TMG by metabolomics. Network pharmacology analysis indicated that TMG intervened in CRC through multi-target actions, with the Wnt/β-catenin pathway being the core regulatory pathway. In vitro experiments demonstrated that both TMG and Apigenin inhibited HCT116 and HCT-8 cell proliferation, promoted apoptosis, and weakened invasion and migration capabilities. Apigenin promotes β-catenin degradation and suppresses its nuclear translocation by upregulating GSK3β and APC expression. The ablation or mutation of GSK3β or APC abolishes this effect, whereas β-catenin knockdown alone recapitulates the anti-cancer effects of TMG and Apigenin, demonstrating that their actions are strictly dependent on β-catenin. Apigenin, as the core active ingredient of TMG, was demonstrated to target GSK3β/APC to promote β-catenin degradation and inhibit the activation of the Wnt/β-catenin pathway, thereby exerting anti-CRC effects.
The present study examined the impact of conjugated linoleic acid (CLA) on the differentiation and proliferation of bovine intramuscular preadipocyte (BIP) cells, specifically exploring the role of G protein-coupled receptor 41 (GPCR41). Cells were exposed to CLA at various concentrations ranging from 50 to 150 μmol/L. Flow cytometry revealed that treatment with 100 μmol/L CLA led to a significant increase in the proliferation rate. During differentiation, 100 and 150 μmol/L CLA markedly enhanced lipid droplet accumulation. Maximal cytoplasmic lipid content was observed when CLA was combined with insulin and dexamethasone. CLA treatment significantly enhanced the transcript abundance of the adipogenic transcription factors peroxisome proliferator-activated receptor gamma (PPARγ) and CCAAT-enhancer binding protein alpha (C/EBPα). Knockdown of GPCR41 via RNA interference in CLA-treated cells led to a significant suppression of GPCR41 expression, accompanied by decreased PPARγ mRNA levels and diminished cytoplasmic lipid droplet formation, indicating impaired differentiation. Taken together, these results show that CLA enhances both differentiation and proliferation of bovine intramuscular preadipocytes, possibly via activation of C/EBPα and PPARγ, with GPCR41 playing a critical role in mediating this adipogenic effect.
