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Extracellular vesicles (EVs) regulate vascular injury and homeostasis; however, the intracellular mechanisms for the EV effects are unknown. While EV components and functions are well studied, their cellular uptake and intracellular processing remain unclear. Following our previous work on EVs, we investigated EV uptake mechanisms and organelle localization. Pulmonary endothelial-derived EVs showed preferential interaction with pulmonary vessels, particularly injured vessels. Co-culture experiments confirmed endothelial cells as exclusive EV targets via energy-dependent, clathrin-mediated endocytosis. Intracellularly, EVs colocalized with lysosomes and trans-Golgi, suggesting degradation and protein recycling pathways. These findings provide insights into temporal EV uptake dynamics and organelle interactions, establishing a foundation for understanding EV processing and content distribution mechanisms.
The hypothalamus coordinates energy-balance regulation and is also a neurogenic/plastic region in the adult brain. Tanycytes, a specialized population of radial glial-like cells lining the third ventricle, reside at the critical interface between the blood, cerebrospinal fluid, and hypothalamic parenchyma. This unique positioning enables them to sense metabolic and nutrient-derived signals, and to shuttle molecules between periphery and brain. Tanycytes can respond to glucose and lipids, as demonstrated by a calcium transient down their long processes that extend into the hypothalamic nuclei. Tanycytes are also capable of self-renewal and differentiation after brain injury, supporting their classification as putative neural stem cells in the adult hypothalamus. Bone morphogenetic protein (BMP) signaling regulates neuroplasticity and contributes to metabolic regulation, including appetite and sympathetic drive to adipose. We previously demonstrated that central administration of BMP7 suppresses appetite, and BMP receptor 1A (BMPR1A) in anorectic hypothalamic POMC neurons impacts appetite regulation. BMPR1A is also tightly and highly co-expressed in hypothalamic tanycytes. Here, we attempted to genetically inactivate BMPR1A in adult tanycytes to explore its functional roles. Using the Rax-CreERT2xBMPR1Aflox mouse line, we tested multiple routes of tamoxifen administration, as well as its metabolite, without success. Cre recombinase activity was successfully induced via dietary tamoxifen (shown by recombination of the BMPR1A locus and fluorescent reporter induction), but efficient BMPR1A knockout in adult tanycytes was not achieved. Similarly, adeno-associated viral (AAV)-mediated BMPR1A knockdown via the Dio2 promoter and intracerebroventricular delivery yielded limited efficiency, despite confirmed Cre activity indicated by reporter expression. We also observed a compensatory increase in BMPR1A in cells not targeted by these knock-out/knockdown systems, as we observed previously with POMC-Cre knockout of BMPR1A, indicating a responsiveness of the hypothalamic niche to manipulation of BMPR1A levels. Together, our findings support that Cre-driven reporter activity doesn't guarantee gene depletion, and demonstrate that current strategies for loss of function of BMPR1A in adult hypothalamic tanycytes remain technically challenging and require careful validation before interpretation of phenotypes. More efficient and reliable methods are required to elucidate the molecular signaling and functional roles of molecules expressed in adult tanycytes.
Osteoarthritis (OA)-related meniscal degeneration involves complex interactions between oxidative stress and proteasomal dysfunction. However, the molecular drivers of regional meniscal vulnerability remain poorly defined. This study integrated multiple transcriptomic datasets from OA and control menisci to identify functional networks and hub genes by using weighted gene co-expression network analysis. Human meniscal tissues from medial and lateral compartments were harvested during total knee arthroplasty and subjected to western blot analysis. In vitro assays on the basis of human chondrocytes were exposed to lipopolysaccharide or the proteasome inhibitor MG132 (carbobenzoxy-l-leucyl-l-leucyl-l-leucinal) to evaluate the stimulus-specific regulation of identified network and hub genes. Weighted gene co-expression network analysis revealed microsomal glutathione S-transferase (MGST1) as the hub gene within a module enriched for ubiquitination and proteasome activity. Experimental validation in human meniscal tissues demonstrated pronounced upregulation of MGST1, ubiquitin-conjugating enzyme E2 N (UBE2N), and proteasome activator complex subunit alpha (PSMA) in mechanically overloaded medial compartments compared to lateral regions. In vitro studies demonstrated stimulus-specific modulation: lipopolysaccharide-induced inflammatory stress upregulated MGST1, whereas proteasome inhibition via MG132 led to its downregulation. These findings highlight a dynamic interplay between redox adaptation and proteostasis, where chronic mechanical stress drives MGST1-mediated antioxidant responses and compensatory ubiquitination. Together, these results suggest that joint tissues dynamically adapt to mechanical and inflammatory challenges by modulating oxidative stress defenses and protein quality control mechanisms, processes central to OA pathophysiology.
The oxygen equivalent of blood lactate accumulation (ΔLa) is commonly expressed relative to body mass (BM), yet BM includes adipose tissue which does not substantially contribute to glycolytic energy production nor lactate distribution space and may confound sex comparisons. We examined whether replacing BM with fat-free mass (FFM) (i) improves the correspondence between calculated glycolytic work and 15-s sprint work and (ii) attenuates sex differences in the work-lactate relationship. Seventy-one trained cyclists (48 men, 23 women) performed a 15-s all-out seated cycling sprint on a Cyclus2 ergometer. Blood lactate was sampled at rest and repeatedly for 8 min post-sprint; ΔLa was defined as peak minus pre-exercise lactate. Glycolytic work (WGly) was calculated using an oxygen-equivalent approach with ΔLa scaled to either BM or FFM. General linear models included glycolytic work, sex, and 15-s work. Men exhibited higher 15-s work and ΔLa than women, but the slope of the relationship between WGly and 15-s work did not differ by sex for either BM or FFM. Regression models using FFM explained slightly more variance in 15-s work than BM (R 2 = 0.79 vs. 0.75). Adding sex improved model fit for both formulations (R 2 = 0.85 and 0.85, respectively), indicating primarily an intercept effect rather than a slope difference. Replacing BM with FFM provides only a small improvement in explaining 15-s work and does not reveal sex-specific differences in the work-lactate slope. Thus, the lactate oxygen equivalent appears sex-invariant while FFM-based scaling may still be preferred for a more physiologically grounded estimate of WGly.
Dysfunction of the circadian clock has been implicated in the pathogenesis of various diseases, including metabolic disorders, inflammatory conditions, and cancer. While the significance of circadian rhythm in diabetic nephropathy is gaining attention, the specific alterations in circadian profiles in diabetic nephropathy remain unexplored. In the present study, we performed RNA sequencing on renal cortex samples collected every 4 h across the day from both control and diabetic mice. The rhythmicity of genes was identified using the JTK_CYCLE algorithm for each group. Genes that lost, acquired, or sustained rhythmicity in diabetic mice were denoted the circadian dysregulation gene set. Subsequent bioinformatic analyses focused on this gene set to investigate the circadian reprogramming in diabetic nephropathy. We observed significant circadian disruption in the kidney of diabetic mice, marked by both the gain and loss of rhythmicity, along with alterations in the phase and relative amplitude of genes that retained rhythmic expressions. Circadian disturbances, such as phase shifts and alterations in relative amplitude or mesor, were also noted in core clock genes. Furthermore, genes that lost rhythmicity in diabetic nephropathy were predominantly associated with protein homeostasis and glycolipid metabolism, whereas those that gained rhythmicity were mainly linked to gene regulation, fatty acid metabolism, and protein transport. The genes in the circadian dysregulation gene set that exhibit differential expression at least at one Zeitgeber time were most prominently enriched in the lipid metabolic process. WGCNA and correlation analysis revealed co-expression networks involving core clock genes and PPAR signaling pathway with renal triglyceride levels. Our study reveals substantial circadian disruption in diabetic nephropathy, with significant impacts on protein homeostasis and glycolipid metabolism. Furthermore, our findings highlight the potential influence of circadian system dysregulation on the disorder of fatty acid metabolism in diabetic nephropathy.
Gut microbes are key regulators of immune homeostasis. Their composition fluctuates over time and between individuals and is also influenced by disease. We and others have reported changes in gut bacterial composition following induction of experimental autoimmune encephalomyelitis (EAE), a well-established model for multiple sclerosis (MS). Specifically, we observed reductions in the abundance of bacteria capable of producing gamma-aminobutyric acid (GABA). Because GABA regulates immune cell function, we genetically engineered a Lactococcus lactis strain to overproduce GABA (P8s-GAD L. lactis) and hypothesized that this strain would have protective activity in EAE. To test this hypothesis, a suspension of P8s-GAD L. lactis was administered by gavage to C57BL/6 Envigo (Env) and Jackson Laboratories (Jax) mice at the time of EAE induction. Controls included mice treated with unmodified L. lactis (P-L. lactis) and mice treated with sterile bacterial medium. P8s-GAD L. lactis was clinically protective in Env mice but not in Jax mice. To understand the lack of protection in Jax mice, we examined the effects of treatments on intestinal micro- and mycobiota using 16S rRNA and IST sequencing, and samples were collected at disease induction, 14 days after, and at the end of the experiment (day 28). We also examined the impact of treatments on the brain, using whole-brain proteomics (day 28). Despite the lack of disease protection, P8s-GAD L. lactis significantly modified the gut microbiome by affecting broad taxonomic composition, as quantified by beta-diversity changes over time, and the CNS protein profile, including an increase in Gabra6 expression, the alpha-6 subunit of the GABA type A (GABARA) receptor. These changes, combined with reduced EAE severity observed in Env mice, suggest that GABA-producing bacteria could be considered for the treatment of neuroinflammatory conditions. The study also highlights the importance of controlling the mouse source in probiotic and microbiota research within experimental models of immune-mediated diseases.
[This corrects the article DOI: 10.1096/fba.2025-00226.].
Idiopathic pulmonary fibrosis (IPF) is a chronic lung disease characterized by the excessive accumulation of collagen-rich extracellular matrix (ECM), leading to the replacement of normal lung architecture. This pathological remodeling is primarily caused by the epithelial-to-mesenchymal transition (EMT), in which epithelial cells lose their polarity and adhesion and acquire mesenchymal characteristics that encourage the deposition of ECM and the growth of fibrotic tissue. The disruption of ECM homeostasis caused by dysregulated MMP-2 and MMP-9 activity in IPF paradoxically promotes abnormal tissue remodeling and fibrosis. IPF's unknown etiology, delayed diagnosis, and lack of effective treatments point to a crucial knowledge gap regarding ECM- and EMT-driven fibrosis. The microRNA-200 (miR-200) family has been found to be important EMT regulators in recent research, suggesting that they may be able to influence the course of fibrosis. Hereby the current study provides an insight on the role of miR-200a in regulating MMP-2 and MMP-9 in bleomycin (BLM)-induced lung fibrosis using both in vitro (A549 cells) and in vivo (C57BL/6 mice) models. A549 cells were transfected with a synthetic miR-200a mimic, and MMP expression was analyzed using RT-qPCR. In vivo, mice were intranasally administered a lentiviral vector expressing miR-200a prior to BLM induction, followed by tissue analysis at days 14 and 21 using histological stains and immunofluorescence. Gene and protein expression were quantified via RT-qPCR and western blotting. Our findings indicate that miR-200a mitigates fibrosis by downregulating MMPs and PAI-1 while upregulating uPA and uPAR, suggesting a protective role of miR-200a and its potential as a therapeutic target for pulmonary fibrosis.
Heat stroke (HS) is the most severe form of hyperthermia, with mortality exceeding 50% in severe cases. The liver is highly vulnerable to HS-induced injury, often triggering multi-organ failure. Although rapid cooling remains the primary treatment, the molecular mechanisms underlying hepatic damage remain elusive, highlighting an urgent need for mechanistic insights, especially given global extreme heat events. We established a HS model by gradually increasing the core temperature of mice from 40°C to 43°C. Mice were sacrificed at each target temperature to collect blood and liver tissues for hematological, biochemical, and histopathological analyses. Transcriptomic profiling was conducted on murine livers, and differentially expressed genes (DEGs) were identified and analyzed. The peroxisome proliferator-activated receptor (PPAR) signaling pathway was identified as a significantly enriched pathway and 12 key DEGs were validated by reverse transcription quantitative PCR (RT-qPCR) to assess temperature-dependent metabolic reprogramming. The expression of CD36, ACOX3, and PPARα was validated by immunohistochemistry at the protein level to investigate their response to heat stress. A graded murine HS model was established and histopathology analysis showed significant liver injury with core temperatures ≥ 42°C, manifesting as weight loss, elevated serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), neutrophilia, thrombocytopenia, hepatocyte necrosis, and sinusoidal congestion. Transcriptomic profiling revealed temperature-dependent DEGs from 41°C onward mainly involved in inflammatory/immune, lipid metabolism, apoptosis, and stress response pathways. DEGs consistently dysregulated across different temperatures were enriched in PPAR, insulin signaling, and endoplasmic reticulum (ER) stress-related pathways. RT-qPCR analysis revealed the altered expression of PPAR-related key genes, indicating functional disruption in lipid metabolism. Immunohistochemistry further confirmed these transcriptomic findings at the protein level, suggesting that heat stress induced reprogramming of the PPAR signaling pathway. Together, these findings suggest that HS-induced liver injury is closely associated with progressive metabolic reprogramming, with dysregulated lipid metabolism playing a central pathogenic role. By combining a murine stepwise HS model and transcriptomic analysis, we identified dysregulated PPAR signaling as a key temperature-dependent feature of liver injury, suggesting its potential role as a temperature-sensing node and therapeutic target. This work provides a framework with precise temporal windows and molecular candidates for the development of mechanism-directed intervention strategies for HS.
Breast cancer (BC) is one of the most common cancers in women around the world, and utilizing a combined approach is a crucial strategy. Induction of cuproptosis in tumor cells is a novel antitumor approach, though its standalone efficacy remains unclear. In this study, we prepared a novel liposome loaded with the photosensitizer indocyanine Green (ICG) and the cuproptosis inducer elesclomol-Cu (ES-Cu) to examine the synergistic effects of photodynamic-cuproptosis treatment on BC. The cuproptosis inducer ES-Cu and the photosensitizer ICG were encapsulated in nanoliposomes with a membrane hydration approach and then validated in vitro and in vivo. JC-1, MDA, GSH, and other cuproptosis-related indicators were used to confirm the ability of PDT to enhance ES-Cu-induced cuproptosis in MCF-7 breast cancer cells. For confirming the cytotoxic impact of PDT in conjunction with the cuproptosis inducer, tests for CCK-8 and cell death staining were performed. The drugs were administered to animals via tail vein injection to observe their tumor inhibition effects in vivo. Their safety was assessed by monitoring changes in body weight. The average particle size of liposomes loaded with ES-Cu and ICG was 208.3 ± 1.07 nm, exhibiting a consistent nanospherical morphology. ICG produced cytotoxic reactive oxygen species (ROS) that enhanced ES-Cu-induced cell cupping under NIR laser irradiation. The therapeutic effect of the synergistic treatment combining PDT and cuproptosis was validated in both in vitro and in vivo experiments. This investigation proved that PDT markedly augments the ES-Cu-induced cuproptosis in breast cancer cells, demonstrating a synergistic therapeutic effect. This synergistic effect presents a novel therapy approach for BC with substantial practical application potential.
The brain relies on interoceptive feedback signals to regulate bodily functions. Female mice with low serum IGF-1 levels (LID mice) exhibit reduced spontaneous running compared to control females, an effect not seen in males. Reduced activity normalized after sustained systemic IGF-1 treatment. This observation led us to hypothesize that circulating IGF-1-a key regulator of skeletal muscle and bone mass that crosses the blood-brain barrier during physical activity-may convey body vigor information to the brain. Since hypothalamic orexin neurons, which are involved in regulating physical activity, express IGF-1 receptors (IGF-1R) and are modulated by this growth factor, we hypothesized that these neurons might gauge circulating IGF-1 levels to modulate physical activity. Indeed, inactivation of IGF-1R in mouse orexin neurons (Firoc mice) was associated with less time spent in free running. These mice maintain physical fitness but display altered mood and are less sensitive to the rewarding actions of exercise. Further, in response to exercise, Firoc mice showed limited c-fos activation of hypothalamic orexin neurons and monoaminergic neurons of the ventro-tegmental area (VTA) in the brainstem. This area is involved in the rewarding component of exercise that seems to be modulated by IGF-1, as mice receiving systemic IGF-1 showed increased c-fos expression in VTA neurons, while mice with reduced IGF-1R expression in VTA neurons showed no improved mood after exercise. Collectively, these results suggest that circulating IGF-1 is gauged by orexin neurons to modulate physical activity, and that VTA neurons convey the rewarding properties of exercise through direct actions of IGF-1 on them. Hence, serum IGF-1 may constitute an interoceptive signal acting on orexin/VTA neurons to modulate physical activity according to physical vigor (muscle and bone mass).
Mitochondrial function is essential for skeletal muscle health, and its disruption leads to atrophy and functional decline. This study examines the impact of denervation on skeletal muscle mitochondria in polymerase gamma (PolG)(+/mut) mice, which accumulate mitochondrial DNA (mtDNA) mutations due to a partial deficiency in polymerase gamma proofreading. Using a 14-day denervation protocol, we assessed muscle mass, mtDNA copy number, oxidative stress and mitochondrial dynamics in wild-type (WT) and PolG(+/mut) mice. Our findings reveal that while denervation significantly reduced muscle wet weight and mitochondrial enzyme activity, no genotype-specific differences in muscle atrophy were observed. However, PolG(+/mut) mice displayed more disorganized mitochondrial cristae and elevated oxidative stress markers, indicating greater mitochondrial vulnerability. Despite these changes, the lack of significant differences in mitochondrial proteins and gene expression between genotypes may reflect an adaptive antioxidant response, including increased catalase expression, although the compensatory nature of this response cannot be conclusively determined. These results suggest that oxidative stress-related responses are involved in mitochondrial adaptations during denervation-induced muscle atrophy. The increased expression of antioxidant enzymes, such as catalase, in PolG(+/mut) mice suggests that antioxidant mechanisms are activated in response to increased oxidative stress. These findings underscore the importance of controlling oxidative stress for maintaining muscle health.
Aberrant anabolic activity is critical to tumor biology; however, much remains to be learned about the regulators of protein anabolism in cancer and how this regulation may affect cancer pathophysiology. MicroRNA (miRNA), a family of small nucleotide regulatory molecules, may serve as a potential source of proteostatic regulation. Here, we examined the ability of two co-transcribed miRNA species, miR15a and miR16 (jointly described as miR15a/16) to regulate protein handling and pathophysiology in non-small cell lung cancer (NSCLC). We found that miR15a/16 regulates genes in numerous metabolic and pathological pathways, including those related to protein metabolism. Transfection of cellular models of NSCLC with miR15a/16 mimetics caused reductions in both cell growth and protein synthesis rates. These findings indicate that miR15a/16 acts as regulators of protein anabolism in NSCLC, serving as novel metabolic regulators and potential clinical therapeutic targets for malignant lung cancer.
The National Institutes of Health (NIH) has launched a major initiative to expand human-based New Approach Methodologies (NAMs) in biomedical research and reduce reliance on animal models. While NAMs offer powerful complementary tools, animal-based research remains indispensable in musculoskeletal science for understanding complex cellular and systemic processes, disease onset and progression, and developing effective therapies. Foundational knowledge of embryonic development, disease mechanisms, tissue regeneration, gene function, and systemic pharmacology has emerged from animal models and will continue to do so. This review underscores the essential role of animal models in five key areas of musculoskeletal biology: osteoporosis, osteoarthritis, bone fracture repair and regeneration, bone cancer, and Inherited Skeletal Disorders (ISDs). We also examine NAMs including organoids, engineered scaffolds, organ-on-chip platforms, and Artificial Intelligence (AI)/computational modeling, highlighting their strengths in mechanistic and high-throughput studies but also their limitations in replicating in vivo structural, physiological, biomechanical, and systemic complexity. Animal models remain the gold standard for exploring disease mechanisms, testing preclinical therapeutic and diagnostic efficacy and safety, and translating discoveries into clinical practice. Rather than replacing animal research, NAMs should be integrated as complementary approaches to advance understanding and innovation. Curtailing animal research would jeopardize medical progress and hinder life-saving interventions for humans and animals alike. This review aims to inform the public and policymakers on the continued necessity of ethically conducted animal research as a cornerstone of musculoskeletal health.
Hypoxia is associated with a range of maladies, inflammation, and impaired immunity. The airway epithelial barrier contends with constant exposure to microbes but can be weakened with hypoxia and diseases, such as cystic fibrosis (CF). People with CF (pwCF) have defective cystic fibrosis transmembrane conductance regulator (CFTR) function leading to reduced immune function, excess mucous accumulation, and chronic infection. CFTR is a cAMP-dependent anion channel that is regulated in part by adenylyl cyclase 6 (AC6). G protein-coupled receptors (GPCRs) such as the chemosensory bitter taste receptors (T2Rs) have been shown to alter inhibitory G proteins and cAMP levels. T2Rs also mediate innate immunity responses and detect quorum sensing molecules (QSMs) through T2R14. The impact of hypoxia on these processes, in human airways, has not yet been characterized. We analyzed protein expression and functional endpoints at normal (21%), mild (10%), and severe (1%) oxygen levels to establish the effects of hypoxia on these processes in human bronchial epithelial cells. Our results show that severe hypoxia leads to decreased AC6 expression without altering Gαi/Gαs/T2R14 compared to wild-type controls. Hypoxia induced ligand and oxygen dependent effects on T2R14 functional responses to fungal QSMs, farnesol, and tyrosol. IL-5 release was increased in QSM treated CF cells at 1% oxygen. Severe hypoxia inhibited forskolin-induced currents due to CFTR and reduced cAMP. These results demonstrate expression level and functional alterations due to hypoxia in airway epithelia, including evidence that reduced AC6 expression and function in severe hypoxia is associated with CFTR dysfunction, establishing a potential link between these proteins and the functional outcome of airway epithelial response to hypoxia.
Gasping respiration enhances survival chances during cardiac arrest by activating the suprahyoid muscles (SHMs), which are crucial for airway dilation. We previously reported that the high concentration of sevoflurane (6.5%: 2.0 minimum alveolar concentration, MAC) leads to gasping-like respiration in mice. Here, to understand the molecular mechanisms of this phenomenon, we compared the hypothalamic transcriptome profiles among control, 2.3% sevoflurane (0.7 MAC; eupnea), and 2.0 MAC groups and identified the differentially expressed genes (DEGs), in which hypocretin (orexin) precursor (Hcrt) gene expression was significantly elevated in the 2.0 MAC group. Notably, the intracerebroventricular administration of orexin enhanced SHM activity at 0.7 MAC. Our findings suggest that the 2.0 MAC sevoflurane-induced increases in orexin enhance activation of SHMs resulting in the involvement of gasping respiration.
Pancreatic ductal adenocarcinoma (PDAC) is the fourth leading cause of cancer-related mortality, characterized by intrinsic resistance to conventional therapies and limited effective treatment options. In this study, we investigated the role of the CXCR2 axis in PDAC therapy resistance. CXCR2, a chemokine receptor, is actively involved in inflammation, tumor angiogenesis, and metastasis. Our working hypothesis is that CXCR2 contributes to PDAC chemotherapy resistance. To test this, we generated gemcitabine-resistant (GemR) lines using T3M4 and CD18/HPAF (CD18) cell lines. Baseline expression of CXCL1, CXCL5, and CXCL8 ligands was higher in GemR cells compared to parental cells. Upon gemcitabine treatment, parental cells exhibited a greater increase in CXCL1 and CXCL8 expression than GemR cells. Further analysis in T3M4 cells revealed a dose- and time-dependent increase in CXCL1 and CXCL8 expression following gemcitabine exposure. Next, we assessed whether targeting CXCR2 could enhance the therapeutic response. We treated parental and GemR cell lines with gemcitabine in combination with a CXCR2 antagonist, Navarixin. Notably, lower concentrations of gemcitabine combined with Navarixin were more effective than higher concentrations of gemcitabine alone in GemR cell lines. In both parental and GemR xenograft models, combination therapy with Navarixin and gemcitabine demonstrated superior antitumor and antimetastatic activity compared to either treatment alone. In conclusion, these findings highlight the critical role of the CXCR2 axis in PDAC therapy resistance. Targeting CXCR2 enhances gemcitabine efficacy, offering a potential therapeutic strategy to overcome resistance in PDAC.
The cellular and molecular complexity of acne pathogenesis has hindered progress toward effective targeted therapies. While keratinocytes are known to influence skin inflammation, their precise transcriptional programs and regulatory circuitry in acne remain unclear. We developed an integrative computational framework that combines single-cell RNA sequencing (scRNA-seq), gene co-expression network analysis (WGCNA), and two complementary machine learning algorithms (SVM-RFE, LASSO) to identify disease-relevant biomarkers. We mapped acne lesion cellular composition, reconstructed keratinocyte differentiation trajectories, and integrated miRNA-transcription factor-drug interaction networks to link molecular signatures to potential interventions. We uncovered marked keratinocyte heterogeneity and enriched late-stage pro-inflammatory states in acne lesions, accompanied by increased macrophage/monocyte and T cell infiltration. Six keratinocyte-associated biomarkers (PYGL, C10orf99, C12orf75, S100A2, PI3, CARD18) were identified, achieving high diagnostic accuracy (AUC > 0.85). Functional enrichment connected these genes to cytokine and chemokine signaling, while regulatory analysis revealed upstream modulators (hsa-let-7b-5p, FOXC1). Drug-gene network mapping suggested repurposing potential for cyclosporin A and valproic acid. In conclusion, our study delineates a keratinocyte-centered molecular signature that shapes acne pathogenesis and provides potential therapeutic biomarkers.
Emerging evidence highlights the pivotal role of the gut microbiota (GM) in regulating host metabolism and contributing to the development of insulin resistance (IR). Gut dysbiosis alters the production of critical metabolites, including short-chain fatty acids (SCFAs), bile acids, indole derivatives, and trimethylamine N-oxide (TMAO), which influence intestinal barrier integrity, inflammatory pathways, and glucose homeostasis. Recent clinical and translational studies indicate that SCFAs can improve fasting insulin and HOMA-IR, although the magnitude of benefit varies substantially across individuals, highlighting ongoing controversy surrounding their metabolic effects. Altered microbial regulation of bile-acid metabolism has also been implicated in impaired lipid and glucose signaling, reinforcing the relevance of FXR- and TGR5-mediated pathways in IR. Elevated TMAO levels have further been associated with adverse metabolic outcomes, though debate persists regarding its causal role versus its function as a diet-dependent biomarker. Microbiota-targeted strategies, including dietary fiber, probiotics, and fecal microbiota transplantation (FMT), show potential to modulate these metabolic pathways, yet clinical results remain inconsistent. This narrative review synthesizes recent mechanistic discoveries and clinical findings on microbiota-derived metabolites in IR, highlights key controversies, and outlines future priorities for translating microbiome science into effective and personalized interventions for metabolic disease prevention and management.