Search Results (7,168)

Background: Dietary crude protein (CP) acts as a key nutritional factor that affects the growth performance and liver metabolism of fattening Hu sheep, with metabolizable energy (ME) representing a major confounding factor in CP-related responses. To isolate the specific effects of CP on liver metabolism and minimize energy–protein interactions, we standardized dietary ME at 9.4 MJ/kg dry matter. Methods: We then established three isoenergetic CP concentrations: 11.07%, 13.07%, and 15.11%. A total of ninety 4-month-old male Hu sheep (with an initial body weight of 27.09 ± 1.83 kg) were allocated at random to three dietary treatment groups, each containing 30 animals distributed across three replicate pens, and fed pelleted total mixed rations (PTMRs) for 75 days under pen conditions in southern Xinjiang. Exploratory combined transcriptomic and metabolomic profiling of liver tissue was conducted to characterize how graded CP levels modulate growth traits and hepatic metabolic pathways, thereby identifying the appropriate dietary CP level for efficient and sustainable fattening of Hu sheep in this region. Result: Results indicated that animals fed the 15.11% CP diet showed a significantly higher average daily gain (ADG) and cumulative weight gain compared with those fed 11.07% or 13.07% CP (p < 0.05). Exploratory multi-omics enrichment analysis demonstrated significant overrepresentation (p < 0.05) of differentially expressed genes and metabolites in key biological pathways—including bile secretion, AMP-activated protein kinase (AMPK) signaling, steroid biosynthesis, peroxisome proliferator-activated receptor (PPAR) signaling, and oxidative stress-related and oxidative phosphorylation. Correlation analyses characterized two hub genes—ATP6AP1 and LOC101119853—that were significantly and negatively correlated with ADG (p < 0.05), whereas two metabolites—calcidiol and ADP—displayed significant positive relationships with ADG (p < 0.05). Pathway-level comparisons further demonstrated that both the 13.07% vs. 15.11% CP and the 11.07% vs. 15.11% CP contrasts yielded significant enrichment in AMPK signaling and steroid biosynthesis. Notably, calcidiol and ADP both declined numerically in the 13.07% vs. 15.11% CP comparison, whereas only ADP reached statistical significance in the 11.07% vs. 15.11% CP contrast. Conclusions: Collectively, under an ME level of 9.4 MJ/kg, a dietary CP concentration of 15.11% contributes to favorable growth of 4-month-old fattening Hu sheep housed in pens in southern Xinjiang. This level is associated with improved growth performance and coordinated regulation of central hepatic regulatory networks—particularly those involved in energy homeostasis and steroidogenesis—thereby supporting metabolic stability without compromising animal health or production efficiency. These findings provide a preliminary molecular basis for precision protein nutrition in Hu sheep feeding systems and offer translational insights for optimizing ruminant nutrition under arid and semi-arid environmental constraints. All correlations indicate potential associations, not causal relationships. Full article

Background: Yes-associated protein 1 (YAP1), a key effector of the Hippo signaling pathway and mechanosensitive transcriptional coactivator, plays a complex role in osteoarthritis (OA) and cartilage regeneration. While YAP1 is essential for tissue homeostasis, its dysregulation has been implicated in both inflammatory and degenerative joint pathologies. However, its precise function remains ambiguous. Methods: We silenced YAP1 with small interfering RNA (siYAP1) in two human-cell-based models relevant to OA pathogenesis and cartilage repair: (1) IL-1β (10 ng/mL)-stimulated articular chondrocytes in monolayer and pellet cultures, and (2) TGF-β1 (10 ng/mL)-induced chondrogenesis in MSC pellet cultures. Outcome measures comprised YAP1 nuclear localization; inflammatory/catabolic markers in chondrocytes (IL6, IL8, ADAMTS5, MMP13); and, in MSC pellets, chondrogenic or hypertrophic markers (COL2A1, ACAN, RUNX2, MMP13, COL10A1) together with glycosaminoglycan (GAG) deposition. Statistical significance was assessed using an ANOVA or Friedman test with post hoc correction (Tukey or Dunn’s test, respectively); p < 0.05 was considered significant. Results: In human chondrocytes, siYAP1 reduced IL-1β-induced nuclear YAP1 localization and suppressed pro-inflammatory mediators IL6 and IL8, indicating an anti-inflammatory effect. YAP1 silencing also downregulated ADAMTS5 expression in 2D monolayers but not in 3D pellet cultures, suggesting reduced regulatory influence in the three-dimensional environment. Notably, MMP13 expression was paradoxically increased following YAP1 knockdown, underscoring the complexity of YAP1’s role in catabolic regulation. In MSC chondrogenesis, siYAP1 enhanced TGF-β1-induced chondrogenesis by increasing COL2A1 and ACAN expression and promoting GAG deposition on day 21. Additionally, it reduced hypertrophic markers RUNX2 and MMP13 on day 7, though COL10A1 remained elevated compared to negative siRNA, indicating only partial suppression of hypertrophic differentiation. Nuclear YAP1 levels were increased by day 21 despite reduced mRNA, suggesting post-transcriptional regulation or enhanced nuclear translocation. Conclusions: These findings demonstrate that YAP1 knockdown exerts context-specific anti-inflammatory and pro-chondrogenic effects while partially mitigating hypertrophy. However, divergent outcomes, namely elevated MMP13 in chondrocytes and upregulated COL10A1 in MSCs, indicate that YAP1 silencing does not uniformly suppress inflammation or hypertrophy. YAP1 represents a potential therapeutic target for OA, but its modulation requires careful consideration of cellular context, siRNA delivery method, and timing to optimize outcomes for cartilage repair and joint preservation. Full article

Pulmonary arterial hypertension (PAH) is a progressive, fatal disease of the pulmonary vasculature characterized by obliterative remodeling of small pulmonary arteries, leading to sustained elevation of pulmonary vascular resistance, right ventricular failure, and premature death. The diagnostic gold standard remains right heart catheterization, requiring a mean pulmonary artery pressure greater than 20 mmHg at rest, a pulmonary arterial wedge pressure of 15 mmHg or below, and a pulmonary vascular resistance exceeding 2 Wood units. PAH is an autosomal dominant disorder with markedly incomplete penetrance of approximately 20–30%, indicating that germline mutations alone are insufficient to cause disease. Disease manifestation requires additional “second hits”, including chronic hypoxia, systemic inflammation, hemodynamic stress, hormonal influences, and common genetic modifiers such as single-nucleotide polymorphisms (SNPs). This genetic and environmental complexity underpins the broad clinical heterogeneity observed across PAH subtypes, which include idiopathic PAH, heritable PAH, and disease associated with connective tissue disorders, HIV infection, portal hypertension, congenital heart disease, schistosomiasis, and drug or toxin exposure. This review provides a comprehensive and critical appraisal of the molecular-genetic architecture of PAH. Thirty genes have now been implicated in disease pathogenesis, spanning seven functional categories: receptors of the TGF-β/BMP signaling family (BMPR2, ACVRL1, ENG, BMPR1B); circulating BMP ligands (GDF2, BMP10); transcription factors (TBX4, SOX17, KLF4, FOXF1, SMAD1, SMAD4, SMAD9); membrane and polyamine transporters (ATP13A3, AQP1); potassium channel regulators (KCNA5, KCNK3, ABCC8); metabolic and mitochondrial genes (EIF2AK4, NFU1, GGCX); signaling receptors and structural proteins (NOTCH3, KDR, CAV1, PLEKHH2); vasoactive and extracellular matrix regulators (KLK1, CBLN2, CD248); and epigenetic regulators (TET2, TOPBP1). Among these, BMPR2 is the dominant contributor, accounting for 53–86% of heritable PAH and 14–35% of idiopathic cases. The remaining genes each account for fewer than 5% of cases individually, collectively reflecting a broad landscape of rare and ultra-rare genetic contributions. For each gene, we critically evaluate the strength of genetic evidence, pathogenic mechanisms, degree of mechanistic resolution, and clinical relevance. We further discuss the contribution of emerging technologies, including whole-genome sequencing, single-cell and spatial transcriptomics, multi-omics integration, iPSC-derived vascular models, and artificial intelligence, to expanding the PAH genetic architecture beyond single-gene discovery. A key theme across this landscape is convergence: despite mechanistic diversity at the gene level, most PAH-associated variants ultimately impair endothelial quiescence, promote smooth muscle proliferation, and drive apoptosis resistance through disruption of BMP signaling amplitude, transcriptional stability, ion channel homeostasis, metabolic integrity, or epigenetic regulation. This convergence supports both a unified therapeutic rationale and a precision medicine framework for genotype-stratified intervention in PAH. Full article

The optimal timing and therapeutic role of dexamethasone for neuroprotection in neonatal hypoxic–ischemic (HI) brain injury remain unclear. We investigated whether dexamethasone-mediated neuroprotection is time-dependent and explored its underlying molecular mechanisms in a neonatal HI mouse model. The Rice–Vannucci model (unilateral carotid artery ligation followed by 8% O₂ for 90 min) was constructed utilizing postnatal day 7 mice who received vehicle (n = 5), dexamethasone pre-treatment (0.5 mg/kg, 6 h before HI; n = 6), or dexamethasone post-treatment (0.5 mg/kg, 6 h after HI; n = 6). Brain injury severity was evaluated by two blinded investigators 72 h after HI, who measured the whitish discoloration in the ipsilateral hemisphere. Transcriptomic analysis was performed using five representative brain samples from each group. Dexamethasone pre-treatment significantly reduced the area of whitish discoloration compared with the vehicle (p < 0.001); dexamethasone post-treatment exerted no significant protective effect. Transcriptomic profiling identified 962 (407 upregulated and 555 downregulated) differentially expressed genes. Genes with upregulated expressions were enriched in pathways related to central nervous system development, synaptic signaling, and calcium homeostasis; those with downregulated expressions were associated with cellular metabolic processes. Protein–protein interaction network analysis identified Dlg4, Calm1, and Grin1 as hub genes. qRT-PCR validation confirmed significant upregulation of Grin1 and Calm1, whereas Dlg4 showed a concordant but non-significant trend. These findings suggest that dexamethasone pre-treatment may be associated with time-dependent changes in synaptic- and calcium-related gene expression following neonatal HI injury, providing insight into the optimal therapeutic window for neonatal HI brain injury. Full article

Background: Pancreatic ductal adenocarcinoma (PDAC) is associated not only with tumor progression but also with profound metabolic and nutritional disturbances. Thyroid hormone homeostasis may reflect this systemic response; however, perioperative data in PDAC remain limited. We aimed to assess perioperative changes in thyroid-related parameters in patients with PDAC undergoing different types of surgical management and to explore their associations with nutritional, metabolic, and tumor-burden variables. Methods: We performed a retrospective single-center study including 101 patients with PDAC. Thyroid-related and metabolic laboratory parameters were assessed before surgery and again 4–6 weeks later. The analyzed variables included thyroid-stimulating hormone (TSH), free triiodothyronine (FT3), free thyroxine (FT4), the FT3/FT4 ratio, albumin, total protein, glucose, insulin, HbA1c, lipid parameters, and CA 19-9. Patients were analyzed according to resectional versus non-resectional treatment and according to four procedure types. The primary endpoint was perioperative change in the FT3/FT4 ratio. Results: At baseline, resectional patients had significantly higher FT3 and FT3/FT4 ratio values and lower FT4 and CA 19-9 levels than non-resectional patients. In the whole cohort, FT3 and the FT3/FT4 ratio decreased significantly after treatment, whereas TSH increased, and FT4 remained unchanged. These endocrine changes occurred in parallel with significant declines in albumin, total protein, glucose, insulin, HbA1c, and HDL cholesterol, together with an increase in triglyceride levels. Baseline FT3 and FT3/FT4 ratios correlated positively with albumin and total protein and negatively with CA 19-9. Although perioperative changes did not differ significantly between resectional and non-resectional groups except for triglycerides, significant procedure-dependent differences were observed across the four surgical categories for FT3, FT4, TSH, and the FT3/FT4 ratio; glucose; insulin; and triglycerides. The prevalence of low-T3 syndrome increased from 11.1% preoperatively to 38.7% postoperatively. Conclusions: In PDAC, perioperative changes in thyroid hormone indices are pronounced and strongly depend on the type of surgical management. FT3 and the FT3/FT4 ratio appear to reflect systemic metabolic and nutritional adaptation as well as disease burden rather than acting as tumor-specific markers. Full article

The rumen epithelium of Tibetan sheep plays a critical role in energy metabolism and immune defense; however, its post-transcriptional regulatory mechanisms under high-altitude hypoxia stress remain unclear. In this study, we employed integrated mRNA and miRNA transcriptome sequencing to analyze the adaptive strategies of the rumen epithelium in Tibetan sheep at different altitudes. A total of 2183 differentially expressed genes (DEGs) and 135 differentially expressed miRNAs (DEmiRNAs) were identified. Functional enrichment analysis revealed that DEGs and their target genes were significantly enriched in immune-related pathways such as the NF-κB signaling pathway and cytokine–cytokine receptor interaction, as well as metabolic pathways including oxidative phosphorylation and branched-chain amino acid degradation. Integrated network analysis highlighted key regulatory pairs, including oar-miR-370-3p targeting PCK2 and IL1R2, and novel-miR-781 regulating PIK3R5, suggesting coordinated modulation between mitochondrial homeostasis and immune responses. Specifically, the upregulation of immune genes (CCL19, MADCAM1) and heat shock proteins at TS4500m indicates enhanced mucosal immunity and stress tolerance, while altered expression of metabolic genes reflects a shift in energy substrate utilization. These findings elucidate a complex mRNA-miRNA regulatory network that enables Tibetan sheep to maintain rumen epithelial integrity and energy balance under extreme high-altitude conditions, providing novel insights into the molecular basis of hypoxia adaptation in ruminants. Full article

Sebacic acid (SA), a ten-carbon medium-chain dicarboxylic acid, has emerged as a multifunctional bioactive metabolite with potential applications in metabolic regulation and regenerative medicine. Evidence indicates that SA exerts anti-inflammatory effects by modulating nuclear factor kappa B (NF-kB), mitogen-activated protein kinase (MAPK), and signal transducer and activator of transcription (STAT) pathways, improves glucose homeostasis by enhancing mitochondrial function and suppressing hepatic gluconeogenesis, and contributes to lipid metabolism via peroxisomal and mitochondrial β-oxidation. Beyond metabolic regulation, SA promotes bone repair by stimulating osteoblast differentiation and inhibiting osteoclast activity, and supports muscle regeneration by enhancing energy supply, cell proliferation, and the microenvironment. SA also serves as a monomer for poly (glycerol sebacate) (PGS), enabling its use in biodegradable tissue engineering scaffolds. This review synthesizes current experimental and preclinical findings on the biological functions of SA, elucidates the underlying molecular mechanisms, and highlights its translational potential for the treatment of metabolic disorders and tissue regeneration. Full article

Perilipins (PLINs) are lipid droplet-associated proteins that regulate lipid storage, mobilization, and metabolism, yet their roles in invertebrates remain poorly characterized. This study aimed to investigate the evolutionary conservation and functional adaptation of PLIN2 in the sea cucumber Apostichopus japonicus. Phylogenetic analysis placed A. japonicus PLIN2 within the PLIN2 clade, forming an echinoderm-specific branch distinct from vertebrate PLIN2s. Structural prediction revealed an N-terminal PAT domain containing an amphipathic helix that was required for lipid droplet targeting, as deletion of this region abolished its localization to lipid droplet. Functionally, PLIN2 abundance positively correlated with lipid droplet formation, and its knockdown reduced triacylglycerol accumulation while upregulating lipolysis-related genes. Pull-down and co-immunoprecipitation assays identified interactions between PLIN2 and the endoplasmic reticulum protein ERP44, as well as the mitochondrial protein TRXR2, suggesting a role in lipid droplet–organelle coupling. Consistently, disruption of the PLIN2-TRXR2 module impaired fatty acid transfer from lipid droplets to mitochondria, leading to suppressed β-oxidation and decreased ATP production. In addition, PLIN2 mediates the protective role of lipid droplets against Vibrio splendidus-induced ER stress. Together, these findings establish A. japonicus PLIN2 as a multifunctional lipid droplet-associated protein that coordinates lipid droplet stability with organelle communication, energy metabolism, and ER homeostasis. Full article

Progranulin (PGRN) is a secreted protein composed of 7.5 granulin domains. The protein is implicated in various functions, including cell survival, inflammation, lysosomal homeostasis, tumorigenesis, and aging. Haploinsufficiency and complete loss of PGRN function cause the neurodegenerative disorders frontotemporal lobar degeneration and neuronal ceroid lipofuscinosis type 11, respectively. In the nervous system, administration of exogenous PGRN has been shown to promote the survival of various nerve cell types under different pathological conditions and to stimulate neurite outgrowth in vitro and axonal regeneration in vivo. In the retina, PGRN dysfunction results in photoreceptor and retinal ganglion cell (RGC) loss, whereas PGRN administration promotes photoreceptor cell survival. In the present study, we analyzed whether a sustained intravitreal administration of PGRN promotes the survival of axotomized RGCs and the regrowth of the lesioned axons. To this end, we generated a PGRN-overexpressing clonal neural stem cell line and injected the cells into the vitreous cavity of a mouse optic nerve crush model. The progression of the lesion-induced degeneration of RGCs was studied at different time points after the nerve crush. The regeneration of the injured RGC axons into the distal optic nerve stump was analyzed one month after nerve lesioning. We found that the intravitreally administered PGRN slowed the degeneration of the injured RGCs for up to four months, the latest post-lesion interval analyzed. Furthermore, PGRN stimulated the regeneration of some RGC axons over long distances into the distal optic nerve stumps. Taken together, our results identify PGRN as a novel neurotrophic factor for retinal ganglion cells. Full article

Neurodegenerative disorders, including Parkinson’s, Alzheimer’s, and multiple sclerosis, are significant global health issues characterized by escalating neuronal dysfunction and cognitive decline. Studies suggest that microbial toxins originating from fungi and bacteria may contribute to neurodegenerative processes by altering neuronal homeostasis in several ways. Toxins formerly associated with infectious diseases have now been associated with neuroinflammation, oxidative stress, and protein misfolding, all of which are common in neurodegenerative diseases. According to recent studies, microbial toxins generated by the gut microbiota may cross the blood–brain barrier and possibly contribute to neuroinflammatory cascades linked to the development of neurodegenerative diseases. The complex interplay of microbial metabolites, microbial responses, and mitochondrial dysfunction demonstrates the diverse character of neurodegenerative processes. This review delves into the current understanding of microbial toxins, which are produced by diverse bacteria and can have a direct or indirect impact on neuronal health via multiple signaling pathways. Understanding the signaling mechanisms of microbial and toxin-mediated neurodegenerative diseases could result in the development of effective alternative therapeutics for neurological disorders. Full article

Growth hormone-releasing hormone receptor 1a (GHS-R1a) is a key G protein-coupled receptor (GPCR) governing feeding and energy homeostasis. Accumulating evidence shows that GHS-R1a forms functional heterodimers with multiple metabolic-related GPCRs, including dopamine 2 receptor (D2R), melanocortin 3 receptor (MC3R), 5-hydroxytryptamine 2c receptor (5-HT2cR), orexin receptor 1 (OX1R) and cannabinoid receptor 1 (CB1R). These heterodimers undergo distinct signal transduction reprogramming, generating novel physiological effects that are not observed with individual receptors: for instance, GHS-R1a/D2R mediates an atypical calcium signaling pathway to regulate appetite, while GHS-R1a/5-HT2cR antagonizes ghrelin-induced orexigenic effects. Meanwhile, diverse detection techniques, including co-immunoprecipitation and fluorescence resonance energy transfer, have been developed to identify and validate GHS-R1a heterodimerization, laying a solid foundation for mechanistic research. This review systematically summarizes the molecular mechanisms of GHS-R1a heterodimer formation, the characteristic signal regulation patterns of different heterodimers, and their specific regulatory roles in feeding circuits. Furthermore, we discuss the existing research gaps in this field, such as the lack of in vivo detection methods for heterodimers and the unclear structural basis of dimerization. Finally, we highlight the potential of targeting specific GHS-R1a heterodimers as a novel therapeutic strategy for obesity and anorexia, providing new directions for future pharmaceutical development and clinical translation. Full article

The liver is a central metabolic organ that integrates nutrient sensing, lipid handling, and detoxification to maintain systemic homeostasis. In metabolic dysfunction–associated steatotic liver disease (MASLD), chronic metabolic overload accelerates hepatocyte senescence, impairing regenerative capacity and promoting progression toward fibrosis and hepatocellular carcinoma. While transcriptomic studies have provided important insights into stress-responsive pathways, they incompletely capture the proteome remodeling and proteoform-level alterations that govern hepatocyte function during aging and disease. Recent mass spectrometry–based proteomics studies have revealed that disruption of autophagy-dependent proteome homeostasis is a defining feature of senescent hepatocytes. Quantitative analyses demonstrate coordinated alterations in selective autophagy pathways—including lipophagy, mitophagy, ferritinophagy, ER-phagy, and pexophagy—accompanied by organelle-specific protein abundance signatures and remodeling of autophagy-related proteoforms. These findings position proteomics as an essential tool for resolving the spatial and functional reorganization of hepatocyte proteomes that cannot be inferred from transcript abundance alone. In this review, we synthesize proteomics-driven evidence defining selective autophagy dysfunction in aging and MASLD livers, critically evaluate methodological limitations, and propose a conceptual framework in which impaired selective autophagy acts as a proteome-level driver of hepatocyte senescence. We further outline future directions for proteoform-resolved and spatial proteomics approaches aimed at identifying actionable targets for therapeutic intervention in liver disease. Full article

Unexplained infertility affects up to 30% of couples and has been associated with heat shock proteins (HSP) and endometrial stress. HSPs and their co-chaperones are part of a complex network of proteins responsible for maintaining protein homeostasis and cell survival. This exploratory hypothesis-generating study investigated the possible relationship between HSPs and unexplained infertility. Twenty-five women were recruited from an IVF clinic. Eleven were confirmed for unexplained infertility (UI), while fourteen were age- and body mass index (BMI)-matched couples with confirmed male factor infertility (MFI), acting as controls. Blood samples were obtained at day 21 of the luteal phase, and plasma measurement of 19 HSPs and co-chaperones undertaken using the slow off-rate modified aptamer (SomaScan) platform. Welch’s t-test and a permutation test were used to compare group means, and Pearson’s correlations to examine relationships with HSPs. Of the 19 proteins measured, plasma HSP70 was decreased (permutation p = 0.002) in cases with unexplained infertility, while HSC70 and STIP1 were increased (permutation p = 0.017 and p = 0.001, respectively) when compared to MFI control. HSP70 was negatively correlated to both HSC70 and STIP 1 in UI (r = −0.77, permutation p = 0.017; −0.80, permutation p = 0.003, respectively), but not in MFI, whilst HSC70 and STIP1 were positively correlated in both UI and MFI (r = 0.93, permutation p = 0.001; r = 0.65, permutation p = 0.035, respectively). The HSP70-HSC70-STIP1 axis showed HSC70-STIP1 coupling with an inverse relationship with inducible HSP70, findings that may suggest dysregulation of constitutive and stress-inducible chaperone systems in UI. Full article

Introduction: This study sought to explore the impact of dietary Cu deficiency on colonic health, including assessments of histopathology, barrier function, inflammatory response, and gut microbiota composition. Methods: Weaned mice were fed a copper-deficient diet for four weeks, followed by one week of intraperitoneal copper sulfate administration as a proof-of-concept rescue intervention. Colonic pathology was assessed by H&E staining, goblet cell changes by AB-PAS staining, and intestinal barrier integrity by immunofluorescence. Inflammatory cytokine levels were measured by ELISA, while protein and mRNA expression of inflammatory markers were detected by Western blot and qRT-PCR. Gut microbiota composition, diversity, and signature genus abundance were analyzed by 16S sequencing. Results: Compared to the control group, CuD mice exhibited histopathological damage in the colon, including mucosal thinning and inflammatory cell infiltration. The number of goblet cells and the expression of mucin MUC2 were significantly reduced, and the expression of tight junction proteins (ZO-1, Occludin) was downregulated, indicating impairment of both the physical and chemical intestinal barriers. Concurrently, Cu deficiency markedly elevated systemic and colonic levels of pro-inflammatory cytokines (TNF-α, IL-1β, and IL-6), and enhanced NF-κB phosphorylation. To explore potential microbial contributions to these colonic alterations, we subsequently analyzed the gut microbiota composition by 16S rRNA sequencing. This analysis revealed that Cu deficiency significantly reduced the α-diversity and species richness of the gut microbiota. This dysbiosis was characterized by a decreased abundance of beneficial bacteria (e.g., Bacteroidota, Muribaculaceae) and an increased abundance of Desulfobacterota, a pro-inflammatory taxon, as well as Akkermansia, a mucin-degrading bacterium with context-dependent effects on gut health. Intraperitoneal administration of copper sulfate (CuD + CuSO₄) partially reversed the histopathological and inflammatory changes; its effect on the gut microbiota was not assessed. Conclusions: Dietary Cu deficiency is associated with colonic injury, and these alterations were accompanied by intestinal barrier disruption, an activated inflammatory response, and gut microbiota dysbiosis. These findings provide experimental evidence highlighting the importance of copper nutrition in maintaining colonic homeostasis, though further mechanistic studies are needed to establish causal relationships. Full article

Background/objectives: Epithelial ovarian cancer (EOC) is the deadliest gynaecologic malignancy, largely due to late-stage diagnosis and ineffective therapy. EOC commonly spreads through the peritoneal cavity as multicellular spheroids, which are metastatic structures that enhance survival under detachment stress, promote dissemination, and contribute to therapeutic resistance. We previously showed that ULK1, a serine/threonine kinase classically linked to macroautophagy initiation, supports EOC progression, suggesting non-canonical roles in spheroid biology and pathogenesis. Methods: CRISPR/Cas9 ULK1 knockout (ULK1KO) models were generated in OVCAR8, HEYA8, and ES2 cells. Mitochondrial degradation phenotypes were assessed in spheroids by immunoblotting and fluorescence microscopy. Label-free proteomics with bioinformatic pathway analysis identified ULK1-associated programs in EOC spheroids. Bioenergetic consequences were quantified using Seahorse ATP-Rate assays. Therapeutic interactions were evaluated using multi-dose combination matrices testing the ULK1 inhibitor DCC-3116 with metformin. Results: ULK1 modulated mitochondrial degradation in a cell-line-specific manner, either promoting or protecting against mitochondrial loss through mechanisms that were uncoupled from canonical autophagy machinery. Proteomic and bioinformatic analyses revealed significant alterations in mitochondria-related processes, aligning with emerging ULK1 functions in mitochondrial homeostasis. ULK1 loss broadly reduced OXPHOS complex proteins in EOC spheroids and consistently decreased hexokinase 2 (HK2), indicating coordinated metabolic remodeling. Seahorse profiling mirrored these shifts: OVCAR8 ULK1KO spheroids showed reduced OCR and ATP production, whereas HEYA8 and ES2 ULK1KO spheroids exhibited increased mitochondrial ATP production. Combination matrices showed potential synergy between DCC-3116 and metformin. Conclusions: These data show that ULK1 differentially regulates mitochondrial degradation across EOC spheroid models through potential mechanisms alternative to canonical autophagy machinery, while reshaping spheroid metabolism and revealing potential therapeutic vulnerabilities in advanced EOC. Full article