Search Results (70)

Cell death and inflammation are key innate immune responses, but excessive activation can cause tissue damage. The NLRP3 inflammasome is a promising target for reducing inflammation and promoting recovery. Immunometabolism regulates NLRP3 responses in neurological and inflammatory diseases through cyclic nucleotide signaling. Targeting phosphodiesterases (PDEs), which hydrolyze cAMP and cGMP, offer a novel approach to mitigate inflammation. While 14 PDE inhibitors are FDA-approved, PDE10A’s role in NLRP3 inflammasome activation remains unclear. This study investigates the effects of PDE10A inhibition on inflammasome-driven inflammation using two PDE10A inhibitors, MP-10 and TP-10, in macrophage and animal models of sepsis and traumatic nerve injury. Our results show that PDE10A inhibition reduces inflammasome activation by preventing ASC speck formation and by lowering levels of cleaved caspase-1, gasdermin D, and IL-1β, which are key mediators of pyroptosis. In the sepsis model, MP-10 significantly reduced inflammation, decreased plasma IL-1β, alleviated thrombocytopenia, and improved organ damage markers. In the nerve injury model, PDE10A inhibition enhanced motor function recovery and reduced muscle atrophy-related gene expression. These findings suggest that PDE10A inhibition could be a promising therapeutic approach for inflammatory and neuromuscular injuries. Given MP-10’s established safety in human trials, Phase 2 clinical studies for sepsis and nerve injury are highly promising. Full article

As part of chemotherapy regimens, Paclitaxel improves the overall survival of many non-small cell lung cancer (NSCLC) patients. However, the development of drug resistance and adverse events limits its clinical usage, reinforcing the need for further advancements in NSCLC therapeutics. We recently recognized the adiponectin receptor agonist AdipoRon as a promising anticancer compound in NSCLC. Consequently, this study aimed to evaluate the therapeutic potential of combining AdipoRon with Paclitaxel (Combo) in NSCLC cells. With respect to individual treatments, Combo triggered a stronger inhibition of both cell growth and clonogenic potential, as well as a greater induction of cell death. The Combo-mediated cytotoxicity was also corroborated by cleavage of poly-ADP ribose polymerase (PARP) and caspase-3 apoptotic markers. Notably, AMP-activated protein kinase (AMPK) emerged as a critical sensor in Combo efficacy, as its inhibition by Compound-C unveiled a significant rescue in cell growth. Although Combo caused a gradual downregulation of extracellular signal-regulated kinase 1/2 (ERK1/2), the hindrance in the upstream cascade by PD98059 partially counteracted the Combo outcomes. In conclusion, our findings designate AdipoRon as an effective candidate in Paclitaxel-based therapy. Nevertheless, future studies aimed at exploring the Combo aptitude in overcoming the Paclitaxel-related restraints need to be investigated in NSCLC. Full article

Background: The cyclic GMP-AMP synthase (cGAS)–type I interferon (IFN-I) pathway detects cytoplasmic DNA and triggers immune responses. Cancer cells often suppress this pathway to evade immune surveillance; however, its therapeutic potential remains unclear. Methods: Mouse oral squamous cell carcinoma models, representing a prominent subtype of head and neck squamous cell carcinoma (HNSCC), were employed in this study. Flow cytometry, Western blot, ELISA, and PCR were used for analysis. Results: We found that immune-unresponsive MOC2 tumors exhibited a deficiency of antigen-presenting cells and cytotoxic T lymphocytes, along with a significant suppression of the cGAS-IFN-I pathway, compared to immune-responsive MOC1 tumors. An MOC2-conditioned medium impaired the differentiation of bone marrow-derived cells into dendritic cells (DCs), reducing the expression of DC markers as well as class I and II major histocompatibility complex (MHC) molecules. The activation of the cGAS-IFN-I pathway in MOC2 cells, either through exogenous DNA or direct IFN-I expression, enhanced class I MHC expression and antigen presentation on MOC2 cells. Furthermore, IFNB1 expression in MOC2 cells induced apoptosis and upregulated chemokines, such as CXCL9 and CXCL10, which recruit immune cells. In immunocompetent mice, IFNB1 expression suppressed MOC2 tumor growth by attracting DCs and T cells, an effect amplified by co-expressing the granulocyte–macrophage colony-stimulating factor. Conclusions: These findings highlight the potential of enhancing cancer cell-intrinsic cGAS-IFN-I signaling to improve tumor immune surveillance and control the progression of immune-cold HNSCC tumors. Full article

Gestational diabetes mellitus (GDM) is characterized by an inadequate pancreatic β-cell response to pregnancy-induced insulin resistance, resulting in hyperglycemia. The pathophysiology involves reduced incretin hormone secretion and signaling, specifically decreased glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP), impairing insulinotropic effects. Pro-inflammatory cytokines, including tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6), impair insulin receptor substrate-1 (IRS-1) phosphorylation, disrupting insulin-mediated glucose uptake. β-cell dysfunction in GDM is associated with decreased pancreatic duodenal homeobox 1 (PDX1) expression, increased endoplasmic reticulum stress markers (CHOP, GRP78), and mitochondrial dysfunction leading to impaired ATP production and reduced glucose-stimulated insulin secretion. Excessive gestational weight gain exacerbates insulin resistance through hyperleptinemia, which downregulates insulin receptor expression via JAK/STAT signaling. Additionally, hypoadiponectinemia decreases AMP-activated protein kinase (AMPK) activation in skeletal muscle, impairing GLUT4 translocation. Placental hormones such as human placental lactogen (hPL) induce lipolysis, increasing circulating free fatty acids which activate protein kinase C, inhibiting insulin signaling. Placental 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) overactivity elevates cortisol levels, which activate glucocorticoid receptors to further reduce insulin sensitivity. GDM diagnostic thresholds (≥92 mg/dL fasting, ≥153 mg/dL post-load) are lower than type 2 diabetes to prevent fetal hyperinsulinemia and macrosomia. Management strategies focus on lifestyle modifications, including dietary carbohydrate restriction and exercise. Pharmacological interventions, such as insulin or metformin, aim to restore AMPK signaling and reduce hepatic glucose output. Emerging therapies, such as glucagon-like peptide-1 receptor (GLP-1R) agonists, show potential in improving glycemic control and reducing inflammation. A mechanistic understanding of GDM pathophysiology is essential for developing targeted therapeutic strategies to prevent both adverse pregnancy outcomes and the progression to overt diabetes in affected women. Full article

Gestation length (GL) is a critical indicator of reproductive performance in sows and is closely associated with other reproductive traits, such as total number born (TNB) and number born alive (NBA). Despite its importance, the genetic mechanisms underlying GL and its impact on reproductive traits remain poorly understood. In this study, we investigated the relationship between GL and reproductive traits using 7013 farrowing records and conducted an imputed whole-genome sequence-based genome-wide association study (GWAS) for GL in first-parity sows, involving 3005 French Large White sows. Our findings revealed that the heritability of GL ranged from 0.22 to 0.26. Both excessively short and long GLs were associated with negative impacts on TNB, NBA, and other reproductive traits. A total of 64 SNPs exceeded the significance threshold, leading to the identification of two novel quantitative trait loci (QTLs) on chromosome 5 (QTL-1: 15.29–15.39 Mb and QTL-2: 12.86–12.94 Mb) and three promising candidate genes: TROAP, RFX4, and ADCY6. Gene ontology and KEGG pathway enrichment analyses revealed that these candidate genes are enriched in key biological processes, including ovarian steroidogenesis, the GnRH signaling pathway, and the regulation of cAMP biosynthesis, all of which are crucial for gestation and pregnancy maintenance. These findings improve our understanding of the genetic architecture of GL in sows and offer valuable genetic markers for enhancing reproductive efficiency in breeding programs. Full article

This study evaluated the neuroprotective potential of peptain-1 conjugated to a cell-penetrating peptide (CPP-P1) in an ocular hypertension model of glaucoma. Brown Norway (BN) rats were subjected to intraocular pressure (IOP) elevation via intracameral injection of silicone oil (SO), with concurrent intravitreal injections of either CPP-P1 or a vehicle. Retinal cross-sections were analyzed for markers of neuroprotection, including cAMP response element-binding protein (CREB), phosphorylated CREB (p-CREB), growth-associated protein-43 (GAP43), synapsin-1 (SYN1), and superoxide dismutase 2 (SOD2). Hematoxylin and eosin staining was used to assess retinal-layer thickness. SO-treated rats exhibited significant reductions in the thickness of the inner nuclear layer (INL, 41%, p = 0.016), inner plexiform layer (IPL, 52%, p = 0.0002), and ganglion cell layer (GCL, 57%, p = 0.001). CPP-P1 treatment mitigated these reductions, preserving INL thickness by 32% (p = 0.059), IPL by 19% (p = 0.119), and GCL by 31% (p = 0.057). Increased levels of CREB (p = 0.17) and p-CREB (p = 0.04) were observed in IOP-elevated, CPP-P1-treated retinas compared to IOP-elevated, vehicle-treated retinas. Although overall GAP43 levels were low, there was a modest increase in expression within the IPL and GCL in SO- and CPP-P1-treated retinas (p = 0.15 and p = 0.09, respectively) compared to SO- and vehicle-treated retinas. SO injection reduced SYN1 expression in both IPL and GCL (p = 0.01), whereas CPP-P1 treatment significantly increased SYN1 levels in the IPL (p = 0.03) and GCL (p = 0.002). While SOD2 expression in the GCL was minimal across all groups, a trend toward increased expression was observed in CPP-P1-treated animals (p = 0.16). The SO model was replicated with SO removal after 7 days and monitored for 21 days followed by retinal flat-mount preparation to assess retinal ganglion cell (RGC) survival. A 42% loss in RGCs (p = 0.009) was observed in SO-injected eyes, which were reduced by approximately 37% (p = 0.03) with CPP-P1 treatment. These findings suggest that CPP-P1 is a promising neuroprotective agent that promotes retinal ganglion cell survival and the preservation of other retinal neurons, potentially through enhanced CREB signaling in a rat model of SO-induced ocular hypertension. Full article

Background: Buffaloes are considered an indispensable genetic resource for dairy production. However, improvements in lactation performance have been relatively limited. Advances in sequencing technology, combined with genome-wide association studies, have facilitated the breeding of high-quality buffalo. Methods: We conducted an integrated analysis of genomic sequencing data from 120 water buffalo, the high-quality water buffalo genome assembly designated as UOA_WB_1, and milk production traits, including 305-day milk yield (MY), peak milk yield (PM), total protein yield (PY), protein percentage (PP), fat percentage (FP), and total milk fat yield (FY). Results: The results identified 56 significant SNPs, and based on these markers, 54 candidate genes were selected. These candidate genes were significantly enriched in lactation-related pathways, such as the cAMP signaling pathway (ABCC4), TGF-β signaling pathway (LEFTY2), Wnt signaling pathway (CAMK2D), and metabolic pathways (DGAT1). Conclusions: These candidate genes (e.g., ABCC4, LEFTY2, CAMK2D, DGAT1) provide a substantial theoretical foundation for molecular breeding to enhance milk production in buffaloes. Full article

Sarcopenia, a condition caused by an imbalance between muscle growth and loss, can severely affect the quality of life of elderly patients with metabolic, inflammatory, and cancer diseases. Vigeo, a nuruk-fermented extract of three plants (Eleutherococcus senticosus Maxim (ESM), Achyranthes japonica (Miq.) Nakai (AJN), and Atractylodes japonica Koidzumi (AJK)) has been reported to have anti-osteoporotic effects. However, evidence of the effects of Vigeo on muscle atrophy is not available. Here, in the in vivo model of dexamethasone (Dex)-induced muscle atrophy, Vigeo treatment significantly reversed Dex-induced decreases in calf muscle volume, gastrocnemius (GA) muscle weight, and histological cross-section area. In addition, in mRNA and protein analyses isolated from GA muscle, we observed that Vigeo significantly protected against Dex-induced mouse muscle atrophy by inhibiting protein degradation regulated by atrogin and MuRF-1. Moreover, we demonstrated that Vigeo significantly promoted C2C12 cell line differentiation, as evidenced by the increased width and length of myotubes, and the increased number of fused myotubes with three or more nuclei. Vigeo alleviated the formation of myotubes compared to the control group. Vigeo also significantly increased the mRNA and protein expression of myosin heavy chain (MyHC), MyoD, and myogenin compared to that in the control. Vigeo treatment significantly reduced the mRNA and protein expression of muscle degradation markers atrogin-1 and muscle RING Finger 1 (MuRF-1) in the C2C12 cell line in vitro. Vigeo also activated the AMP-activated protein kinase (AMPK)/silent information regulator 1 (Sirt-1)/peroxisome proliferator-activated receptor-γ co-activator-1α (PGC1α) mitochondrial biogenesis pathway and the Akt/mTOR protein synthesis signaling pathway in Dex-induced myotube atrophy. These findings suggest that Vigeo may have protective effects against Dex-induced muscle atrophy. Therefore, we propose Vigeo as a supplement or potential therapeutic agent to prevent or treat sarcopenia accompanied by muscle atrophy and degeneration. Full article

We have previously demonstrated that the vasopressin type 2 receptor (AVPR2) antagonist tolvaptan reduces cell proliferation and invasion and triggers apoptosis in different human cancer cell lines. To study this effect in vivo, a xenograft model of small cell lung cancer was developed in Fox1^nu/nu nude mice through the subcutaneous inoculation of H69 cells, which express AVPR2. One group of mice (n = 5) was treated with tolvaptan for 60 days, whereas one group (n = 5) served as the control. A reduced growth was observed in the tolvaptan group in which the mean tumor volume was significantly smaller on day 60 compared to the control group. In the latter group, a significantly lower survival was observed. The analysis of excised tumors revealed that tolvaptan effectively inhibited the cAMP/PKA and PI3K/AKT signaling pathways. The expression of the proliferative marker proliferating cell nuclear antigen (PCNA) was significantly lower in tumors excised from tolvaptan-treated mice, whereas the expression levels of the apoptotic marker caspase-3 were higher than those in control animals. Furthermore, tumor vascularization was significantly lower in the tolvaptan group. Overall, these findings suggest that tolvaptan counteracts tumor progression in vivo and, if confirmed, might indicate a possible role of this molecule as an adjuvant in anticancer strategies. Full article

Fever is one of the most common clinical conditions and is characterized by pyrogenic infection, malignancy, inflammation, and tissue damage, among others. Ellagic acid (EA) can inhibit the expression of related proteins on the pathway by blocking the nuclear factor kappa-B(NF-κB) signaling pathway, inhibit the levels of pro-inflammatory factors interleukin-1β(IL-1β), interleukin-6(IL-6), and tumor necrosis factor-α(TNF-α), increase the level of anti-inflammatory factor IL-10, and effectively alleviate inflammatory symptoms. In addition, EA can also reduce the levels of malondialdehyde(MDA) and nitric oxide(NO) in the body, increase the activities of superoxide dismutase (SOD), glutathione (GSH), and catalase(CAT), scavenge oxidative free radicals, inhibit lipid oxidation, and achieve antipyretic and anti-inflammatory effects. The purpose of this study was to establish the relationship between EA and various inflammatory markers, such as TNF-α, IL-6, IL-1β, prostaglandin E2(PGE₂), and cyclic adenosine monophosphate(cAMP), and clarify the mechanism of the cyclooxidase-2(COX-2)/NF-κB signaling pathway. Combined with the metabolomics analysis, our study revealed the effects of EA on multiple endogenous biomarkers, reflecting the characteristics of a multi-component, multi-target, and multi-pathway mechanism. Compared to lipopolysaccharide (LPS)- treated animals, subsequent administration of EA significantly lowered the LPS-induced rectal temperature increase (p < 0.05 or p < 0.01), significantly increased serum SOD and GSH levels (p < 0.05 or p < 0.01), and significantly decreased serum MDA, IL-1β, IL-6, and TNF-α levels (p < 0.05 or p < 0.01). In addition, compared to LPS-treated animals, subsequent administration of EA significantly decreased cerebrospinal fluid cAMP and PGE₂ levels (p < 0.05 or p < 0.01), significantly decreased cAMP, significantly increased 5-HT levels (p < 0.05 or p < 0.01), and significantly down-regulated p-NF-κB p65 and COX-2 protein levels in the hypothalamus. Subsequent gas chromatography mass spectrometry(GC-MS) metabolite analysis indicated that 12 differential metabolites were detected in serum isolated 4 h after LPS treatment, and 10 differential metabolites were detected in serum collected 7 h after LPS treatment. Next, Pearson correlation analysis was used to systematically characterize the relationship between the identified metabolites and TNF-α, IL-6, MDA, SOD, PGE₂, and cAMP. The levels of propionic acid, pyridine, and L-valine were up-regulated by EA, which inhibited the expression of MDA, IL-1β, and TNF-α and increased the activity of GSH. The levels of inositol, urea, and 2-monopalmitin were down-regulated by EA, which inhibited the expression of MDA, IL-1β, and TNF-α, increased the activity of SOD and GSH, reduced the inflammatory response, and alleviated the oxidative stress state. Combined with the results of the metabolic pathway analysis, we suggest that the pathways of the galactose metabolism, synthesis and degradation of ketone bodies, as well as ascorbic acid and aldehyde acid metabolism are closely related to the antipyretic and anti-inflammatory effects of EA. Our study established the relationship between EA and various inflammatory markers, such as TNF-α, IL-6, IL-1β, PGE₂, and cAMP, and clarified the mechanism of the COX-2/NF-κB signaling pathway. Combined with the metabolomics analysis, our study revealed the effects of EA on multiple endogenous biomarkers, reflecting the characteristics of a multi-component, multi-target, and multi-pathway mechanism. Full article

Both hypertension and aging are known to increase the vulnerability of the brain to neurovascular damage, resulting in cognitive impairment. The present study investigated the efficacy of the antihypertensive drug losartan on age- and hypertension-associated cognitive decline and the possible mechanism underlying its effect in spontaneously hypertensive rats (SHRs). Losartan was administered (10 mg/kg, i.p. for 19 days) to 3- and 14-month-old SHRs. Age-matched Wistar rats were used as controls. Working memory, short-term object recognition, and spatial memory were assessed using the Y-maze, object recognition test (ORT) and radial arm maze (RAM) test. The expression of markers associated with aging, oxidative stress, and memory-related signaling was assessed in the frontal cortex (FC) and hippocampus. Motor activity measured over 24 h was not different between groups. Middle-aged vehicle-treated SHRs showed poorer performance in spontaneous alternation behavior (SAB) and activity in the first Y-maze test than their younger counterparts, suggesting age-related reduced “decision making” and reactivity in a novel environment. Losartan improved the age- and hypertension-induced decline in short-term recognition and spatial memory measured in the ORT and the second Y-maze test, particularly in the middle-aged rats, but was ineffective in the young adult rats. Changes in memory and age-related markers such as cAMP response element-binding protein (CREB) and amyloid-β_1–42 (Aβ_1–42) and increased oxidative stress were observed in the hippocampus but not in the FC between young adult and middle-aged vehicle-treated SHRs. Losartan increased CREB expression while reducing Aβ_1–42 levels and concomitant oxidative stress in middle-aged SHRs compared with vehicle-treated SHRs. In conclusion, our study highlights the complex interplay between hypertension, aging, and cognitive impairment. It suggests that there is a critical time window for therapeutic intervention with angiotensin II type 1 receptor blockers. Full article

Moderate-to-severe psoriasis (Ps) treatment includes systemic drugs and biological agents. Apremilast, a small molecule primarily metabolized by cytochrome CYP3A4, modulates the immune system by specifically inhibiting phosphodiesterase type 4 (PDE4) isoforms and is currently used for the treatment of Ps and psoriatic arthritis (PsA). Clinical trials and real-world data showed variable efficacy in response among Ps patients underlying the need for personalized therapy. This study implements a candidate-gene and a network-based approach to identify genetic markers associated with apremilast response in forty-nine Greek Ps patients. Our data revealed an association of sixty-four SNPs within or near PDE4 and CYP3A4 genes, four SNPs in ncRNAs ANRIL, LINC00941 and miR4706, which influence the abundance or function of PDE4s, and thirty-three SNPs within fourteen genes whose protein products either interact directly with PDE4 proteins or constitute components of the cAMP signaling pathway which is modulated by PDE4s. Notably, fifty-six of the aforementioned SNPs constitute eQTLs for the respective genes in relevant to psoriasis tissues/cells implying that these variants could be causal. Our analysis provides a number of novel genetic variants that, upon validation in larger cohorts, could be utilized as predictive markers regarding the response of Ps patients to apremilast treatment. Full article

Heparan sulphate (HS) can act as a co-receptor on the cell surface and alterations in this process underpin many pathological conditions. We have previously described the usefulness of mimics of HS (glycomimetics) in protection against β-glycerophosphate-induced vascular calcification and in the restoration of the functional capacity of diabetic endothelial colony-forming cells in vitro. This study aims to investigate whether our novel glycomimetic compounds can attenuate glycated low-density lipoprotein (g-LDL)-induced calcification by inhibiting RAGE signalling within the context of critical limb ischemia (CLI). We used an established osteogenic in vitro vascular smooth muscle cell (VSMC) model. Osteoprotegerin (OPG), sclerostin and glycation levels were all significantly increased in CLI serum compared to healthy controls, while the vascular calcification marker osteocalcin (OCN) was down-regulated in CLI patients vs. controls. Incubation with both CLI serum and g-LDL (10 µg/mL) significantly increased VSMC calcification vs. controls after 21 days, with CLI serum-induced calcification apparent after only 10 days. Glycomimetics (C2 and C3) significantly inhibited g-LDL and CLI serum-induced mineralisation, as shown by a reduction in alizarin red (AR) staining and alkaline phosphatase (ALP) activity. Furthermore, secretion of the osteogenic marker OCN was significantly reduced in VSMCs incubated with CLI serum in the presence of glycomimetics. Phosphorylation of cyclic AMP response element-binding protein (CREB) was significantly increased in g-LDL-treated cells vs. untreated controls, which was attenuated with glycomimetics. Blocking CREB activation with a pharmacological inhibitor 666-15 replicated the protective effects of glycomimetics, evidenced by elevated AR staining. In silico molecular docking simulations revealed the binding affinity of the glycomimetics C2 and C3 with the V domain of RAGE. In conclusion, these findings demonstrate that novel glycomimetics, C2 and C3 have potent anti-calcification properties in vitro, inhibiting both g-LDL and CLI serum-induced VSMC mineralisation via the inhibition of LDLR, RAGE, CREB and subsequent expression of the downstream osteogenic markers, ALP and OCN. Full article

Male and female Chinese soft-shelled turtles (Pelodiscus sinensis) have sex-dimorphic growth patterns, and males have higher commercial value because of their larger size and thicker calipash. Thus, developing sex-specific markers is beneficial to studies on all-male breeding in P. sinensis. Here, we developed an accurate and efficient workflow for the screening of sex-specific sequences with ZW or XY sex determination systems. Based on this workflow, female and male P. sinensis reference genomes of 2.23 Gb and 2.26 Gb were obtained using de novo assembly. After aligning and filtering, 4.01 Mb female-specific sequences were finally identified. Subsequently, the seven developed sex-specific primer pairs were 100% accurate in preliminary, population, and embryonic validation. The presence and absence of bands for the primers of P44, P45, P66, P67, P68, and P69, as well as two and one bands for the PB1 primer, indicate that the embryos are genetically female and male, respectively. NR and functional annotations identified several sex-determining candidate genes and related pathways, including Ran, Eif4et, and Crkl genes, and the insulin signaling pathway and the cAMP signaling pathway, respectively. Collectively, our results reveal that a ZW-type sex-determination system is present in P. sinensis and provide novel insights for the screening of sex-specific markers, sex-control breeding, and the studies of the sex determination mechanism of P. sinensis. Full article

Cinnamyl alcohol (CA) is an aromatic compound found in several plant-based resources and has been shown to exert anti-inflammatory and anti-microbial activities. However, the anti-adipogenic mechanism of CA has not been sufficiently studied. The present study aimed to investigate the effect and mechanism of CA on the regulation of adipogenesis. As evidenced by Oil Red O staining, Western blotting, and real-time PCR (RT-PCR) analyses, CA treatment (6.25–25 μM) for 8 d significantly inhibited lipid accumulation in a concentration-dependent manner and downregulated adipogenesis-related markers (peroxisome proliferator-activated receptor γ (PPARγ), CCAAT/enhancer-binding protein α (C/EBPα), fatty acid binding protein 4 (FABP4), adiponectin, fatty acid synthase (FAS)) in 3-isobutyl-1-methylxanthine, dexamethasone, and insulin(MDI)-treated 3T3-L1 adipocytes. In particular, among the various differentiation stages, the early stage of adipogenesis was critical for the inhibitory effect of CA. Cell cycle analysis using flow cytometry and Western blotting showed that CA effectively inhibited MDI-induced initiation of mitotic clonal expansion (MCE) by arresting the cell cycle in the G₀/G₁ phase and downregulating the expression of C/EBPβ, C/EBPδ, and cell cycle markers (cyclin D1, cyclin-dependent kinase 6 (CDK6), cyclin E1, CDK2, and cyclin B1). Moreover, AMP-activated protein kinase α (AMPKα), acetyl-CoA carboxylase (ACC), and extracellular signal-regulated kinase 1/2 (ERK1/2), markers of upstream signaling pathways, were phosphorylated during MCE by CA. In conclusion, CA can act as an anti-adipogenic agent by inhibiting the AMPKα and ERK1/2 signaling pathways and the cell cycle and may also act as a potential therapeutic agent for obesity. Full article