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Background: Mitochondrial dysfunction is a key link in neuronal damage, and targeted mitochondrial repair may be an improvement mechanism for cerebral ischemia-reperfusion (CI/R) injury.Objective: To investigate the effect and mechanism of Guilou Guizhi decoction (GLGZD) on CI/R injury.Materials and Methods: A middle cerebral artery occlusion/reperfusion (MCAO/R) mouse model was used to mimic ischemic stroke. GLGZD was administered by gavage, and the PI3K pathway inhibitor LY294002 was injected intraperitoneally. Neurological deficits, infarct volume, brain edema, and neuronal survival were assessed using mNSS scoring, TTC, HE, Nissl, and TUNEL staining. Mitochondrial function was evaluated by measuring membrane potential, ATP, and ROS levels. Western blot detected Cytc release, apoptosis markers, and PI3K/AKT/mTOR pathway activation.Results: After GLGZD intervention, mNSS score, infarct area, and cerebral tissue water content were dramatically lowered in MCAO/R mice, histopathological damage and neuronal apoptosis in the cerebral cortex and CA1 region were significantly improved. Additionally, the mitochondrial membrane potential and ATP content were elevated, the level of ROS was lowered, the release of Cytc into the cytoplasm was reduced, and the level of neuronal apoptosis-related proteins. Furthermore, GLGZD therapy dramatically increased the phosphorylation status of PI3K/AKT/mTOR pathway molecules, and the pathway inhibitor LY294002 effectively reduced GLGZD's ameliorative effect on a variety of lesions in MCAO/R mice brain tissues.Conclusion: GLGZD restores mitochondrial function via the PI3K/AKT/mTOR pathway to alleviate neuronal apoptosis in MCAO/R mice, suggesting its potential as a therapeutic agent for ischemic stroke.
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Background: Turner Syndrome is a rare disorder resulting from abnormalities in the number or structure of a single X chromosome. It stands as the sole survivable monosomy observed in humans. Individuals affected by Turner Syndrome commonly face significant complications, including impaired gonadal development, short stature, and intellectual disabilities. Despite extensive research, the complete understanding of the pathogenesis behind Turner Syndrome remains elusive.Objectives: This study aimed to elucidate the molecular mechanisms of Turner Syndrome by identifying key X chromosome inactivation escape genes and exploring their potential contributions to genotype–phenotype correlations and therapeutic targets through integrative bioinformatics analyses.Methods: The study included three RNA-seq datasets, consisting of 72 patients with Turner syndrome and 58 individuals with normal cytogenetics. Differentially expressed gene analysis, protein-protein interaction network analysis, and functional enrichment analysis were conducted to ascertain hub genes. Furthermore, pivotal lncRNAs and transcription factors were identified through ceRNA network analysis and transcription factor enrichment analyses.Results: This study made a significant contribution by exploring the relationship between Turner Syndrome genotypes and various phenotypes at the transcriptomic level. We found hub genes associated with hormone regulation, including growth hormone. Moreover, we highlighted 10 crucial X chromosome inactivation escape genes (ASMTL, CD99, DHRSX, EIF1AX, JPX, KDM5C, KDM6A, SLC25A6, XIST, and ZFX), which may serve as core pathogenic factors in Turner Syndrome. Additionally, based on our findings, we proposed certain drugs as potential new therapeutic strategies for Turner Syndrome.Conclusions: Our study provides substantial evidence supporting the correlation between genotype and various phenotypes in Turner Syndrome. Furthermore, we shed light on the potential pathogenic significance of X chromosome inactivation escape genes. These findings open up new possibilities for therapeutic interventions.
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Background: Triple-negative breast cancer (TNBC) is one of the most aggressive breast cancer subtypes, with limited therapeutic options due to the lack of specific molecular targets and frequent drug resistance. Biotechnological advances in nanocarrier systems and ultrasound-targeted drug delivery provide new opportunities for precision cancer therapy.Objectives: This study aimed to develop paclitaxel (PTX)-loaded poly(lactide-co-glycolide) (PLGA) lipid microbubbles and evaluate whether ultrasound-targeted microbubble destruction (UTMD) could enhance the therapeutic efficacy against TNBC cells.Materials and Methods: PTX-loaded microbubbles were synthesized and characterized using dynamic light scattering (particle size, zeta potential), scanning electron microscopy (morphology), and UV spectrophotometry (drug loading efficiency). In vitro anticancer efficacy was assessed through MTT and Transwell assays (cell proliferation and migration), flow cytometry and Western blotting (apoptosis), and mitochondrial damage was analyzed using membrane potential assays and transmission electron microscopy.Results: The PTX-loaded PLGA microbubbles were spherical, well-dispersed, and stable, with an average diameter of 390.5 nm, zeta potential of –22.97 mV, and a drug loading efficiency of 4.34±0.08%. Ultrasound exposure significantly enhanced the inhibitory effects of PTX-loaded microbubbles on TNBC cell proliferation and invasion, promoted apoptosis through Bax/Caspase-3 activation and Bcl-2 downregulation, and exacerbated mitochondrial damage compared with microbubbles or PTX alone.Conclusion: This study demonstrates a nanobiotechnology-based strategy in which ultrasound-triggered PLGA microbubbles serve as an effective carrier for PTX delivery. The findings highlight UTMD as a promising targeted drug delivery platform in medical biotechnology, warranting further in vivo validation for clinical translation in TNBC therapy.
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Objective: Multisystem inflammatory syndrome in children (MIS-C) is a severe, post-infectious inflammatory condition linked to SARS-CoV-2. Its pathogenesis involves complex immune dysregulation, but the role of non-apoptotic cell death pathways, particularly ferroptosis, in immune cells remains unexplored. This study aimed to define the dynamics of ferroptosis-related gene expression within specific immune cell populations during the acute (T1) and recovery (T3) phases of MIS-C to uncover potential mechanistic contributors to disease pathology and identify novel targets for immunomodulation. Materials and Methods: We performed integrated bioinformatic analysis on single-cell RNA sequencing (scRNA-seq) data from peripheral blood mononuclear cells (PBMCs) of children with MIS-C at admission (T1, n=(Specify if available)) and convalescence (T3, n=(Specify if available)). Cell populations were identified and quantified. A curated panel of ferroptosis-related genes (including drivers like ACSL4, SLC7A11 and defenders like GPX4), alongside iron metabolism genes (FTH1, FTL), was analyzed for cell-type-specific expression patterns across the two time points. Results: Immune cell composition shifted significantly from T1 to T3, with B cell proportion increasing (12.1% to 21.4%) and NK cells decreasing (10.8% to 5.3%), reflecting immune response evolution. Critically, in monocytes and macrophages, we observed a concurrent upregulation of pro-ferroptotic genes (ACSL4, SLC7A11) and iron storage genes (FTH1, FTL), indicating a heightened ferroptotic susceptibility. Paradoxically and importantly, the expression of the key anti-ferroptotic enzyme GPX4 was significantly elevated in these same cells (1.8-fold in monocytes, 2.1-fold in macrophages at T3), suggesting an active compensatory cellular defense against lipid peroxidation. Conclusion: Our single-cell analysis unveils a cell-state-specific ferroptosis signature in MIS-C. Monocytes and macrophages exhibit a unique molecular tension, poised for ferroptosis yet simultaneously upregulating potent defensive mechanisms. This imbalance in iron metabolism and redox homeostasis likely contributes to inflammatory mediator release and multi-organ injury. The identified upregulation of GPX4 represents a critical endogenous protective response, highlighting it as a potential biomarker for disease resolution. These findings provide a novel theoretical framework for modulating inflammation in MIS-C through targeted intervention in the ferroptosis pathway, offering a promising avenue for applied therapeutic strategy development, and position GPX4 as a tractable therapeutic target and a robust biomarker for monitoring disease resolution in MIS-C.
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Background: Perioperative anesthetic strategies are increasingly recognized as important modulators of postoperative immune responses and intestinal microecological homeostasis following gastrointestinal surgery. Objective: To systematically explore the differential effects of inhalational anesthesia (sevoflurane) and intravenous anesthesia (propofol) on postoperative intestinal microbiota, and to investigate the potential association between anesthetic methods, NF-κB/HIF-1α signaling pathway, and macrophage polarization using integrated bioinformatics analyses. Methods: Publicly available datasets were retrieved from NCBI SRA, GEO, and EMBL-EBI to identify three independent cohorts with clear annotations of gastrointestinal surgery and anesthetic methods. A total of 90 samples were included (30 control, 30 inhalational anesthesia, and 30 intravenous anesthesia). Intestinal microbiota profiles derived from 16S rRNA sequencing were integrated with expression data of NF-κB/HIF-1α pathway–related genes. Microbial diversity, community composition, differential taxa, functional enrichment, and microbe–gene correlations were analyzed using QIIME2, limma, vegan, and related bioinformatics tools to construct the regulatory axis of “anesthetic method–intestinal microbiota–signaling pathway–macrophage polarization.” Results: The inhalational anesthesia group exhibited significantly reduced α-diversity (Chao1 and Shannon indices) compared with both the control and intravenous anesthesia groups (P<0.05). β-diversity analysis revealed distinct microbial community structures among the three groups (PERMANOVA R²=0.27, P=0.001). Genus-level analysis demonstrated a marked enrichment of Bacteroides in the inhalational anesthesia group, whereas Lactobacillus and Bifidobacterium were predominant in the control group. LEfSe analysis identified eight significantly differential genera. Expression of pro-inflammatory NF-κB/HIF-1α pathway genes (NFKB1, IL-6, TNF-α) was significantly up-regulated in the inhalational anesthesia group, accompanied by down-regulation of anti-inflammatory markers (ARG1, IL-10). These alterations were less pronounced in the intravenous anesthesia group. Correlation analysis identified Bacteroides–NFKB1 and Lactobacillus–IL-10 as key regulatory nodes. Functional enrichment revealed inflammation-related pathways enriched under inhalational anesthesia, whereas short-chain fatty acid synthesis pathways were enriched in controls. Conclusion: Inhalational anesthesia is associated with greater disruption of postoperative intestinal microbiota homeostasis than intravenous anesthesia, and is correlated with NF-κB/HIF-1α-related pro-inflammatory macrophage polarization. These findings provide bioinformatics evidence supporting propofol-based intravenous anesthesia and microbiota-targeted strategies to improve postoperative outcomes.
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Background: Severe acute pancreatitis (SAP) exacerbates inflammatory damage due to mitochondrial autophagy disorders, but the mechanism and diagnostic markers remain unclear. Objectives: This study aimed to identify mitophagy-related biomarkers in severe acute pancreatitis through integrated bioinformatics and machine-learning approaches, and to elucidate their potential regulatory mechanisms and diagnostic value. Methods: Differentially expressed genes (DEGs) related to SAP were screened based on the GSE45670 dataset, and candidate genes were obtained by taking the intersection with the mitophagy gene set. Key genes were screened by combining the least absolute shrinkage and selection operator (LASSO) regression and Support Vector Machine-Recursive Feature Elimination (SVM-RFE) algorithms; the diagnostic efficacy was evaluated using the ROC curve, and a nomogram model was constructed. Furthermore, gene functions and regulatory mechanisms were revealed through Gene Set Enrichment Analysis (GSEA), Gene Set Variation Analysis (GSVA), immune infiltration analysis, and ceRNA network, and potential targeted drugs were predicted using molecular docking. Results: A total of 16 DEGs related to mitophagy were identified. Functional enrichment analysis showed that they were significantly associated with metabolic and immune regulation pathways. Machine learning and expression level validation jointly screened PGD and LMNB1 genes as two key genes. The key genes showed significant expression differences in the training set and external validation set (upregulated in the SAP group), and had excellent diagnostic efficacy (area under the curve (AUC) > 0.85). Immune infiltration analysis showed that the infiltration of 13 types of immune cells increased in the SAP group, and PGD was highly positively correlated with immune cells such as activated dendritic cells (r = 0.82). Molecular docking indicated that estradiol and progesterone might target and regulate PGD and LMNB1. Conclusion: PGD and LMNB1 are key genes related to mitophagy in SAP and have excellent diagnostic value. This study provides a theoretical basis for the analysis of the molecular mechanism of SAP and the development of precise diagnosis and treatment strategies.
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Background: Cyclophosphamide (CP) induces asthenozoospermia, a common model of male infertility. L-Asparagine, derived from Asparagus cochinchinensis, is traditionally used for reproductive disorders. Objective: This study investigates its protective role via the PI3K/AKT/mTOR pathway and autophagy regulation. Methods: Male rats (n=8/group) received CP with or without L-asparagine (low, medium, high doses), rapamycin, or leucine. After four weeks, sperm parameters, hormone levels, oxidative stress markers, histology, and protein expression were evaluated. Results: CP administration significantly impaired testicular weight, hormone levels, sperm count and motility, and antioxidant capacity, while increasing oxidative stress and autophagy. These effects were accompanied by suppression of the PI3K/AKT/mTOR/p70S6K pathway. L-Asparagine treatment dose-dependently restored spermatogenic function (e.g., sperm motility increased by ~40% in the high-dose group), reduced oxidative stress (MDA decreased by 30%, SOD increased by 25%), and inhibited excessive autophagy. Rapamycin co-treatment reversed the benefits of L-asparagine, while leucine co-treatment potentiated them. Conclusion: L-Asparagine mitigates CP-induced spermatogenic dysfunction by activating the PI3K/AKT/mTOR pathway, reducing oxidative damage, and suppressing aberrant autophagy. These findings highlight its potential as a nutraceutical for managing male infertility, particularly post-chemotherapy.
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Background: Chronic kidney disease-associated pruritus (CKD-aP) is a common complication of chronic kidney disease with unclear pathogenesis and limited treatment options. Objectives: This study applied network pharmacology and molecular docking to investigate the interactions between gut microbiota metabolites and biological targets, aiming to identify potential microbiota-derived therapeutic targets for CKD-aP. Methods: The study first extracted gut microbial metabolites from the gutMGene database, and then searched in multiple public databases including DisGeNET, GeneCards and OMIM to find potential targets related to CKD-aP. To identify core targets, we analyzed the protein-protein interaction (PPI) network and conducted functional enrichment studies using the DAVID database, with the methods of gene ontology (GO) classifications and KEGG pathways. Additionally, we mapped out a microbial-substrate-metabolism-target (MSMT) network to better understand their interconnected relationships. Results: 44 common targets were found linking gut microbial metabolites to the disease, with IL6, AKT1, and PPARG emerging as key intervention points for CKD-aP. Our analysis revealed a total of 44 common targets shared between gut microbial metabolites and CKD-aP. Among these, IL6, AKT1, and PPARG were selected as central targets for potential therapeutic intervention. By developing integrated MSMT networks, we discovered that these pivotal targets are influenced by factors such as Lacticaseibacillus paracasei, tryptophan metabolic pathways, and linoleic acid metabolism in alleviating CKD-aP. Further KEGG pathway analysis pinpointed critical signaling, including IL-17 signaling pathway, the C-type lectin receptor signaling pathway and Toll-like receptor signaling pathway as key pathways. Furthermore, molecular docking analysis was performed on active metabolites and core targets, showing that 10-keto-12Z-octadecenoic acid-PPARG, Genipin-CASP3, and Urolithin A-CYP1A1 have good binding activity. Conclusion: Gut microbiota metabolites play a therapeutic role in CKD-aP mainly through microflora (Lacticaseibacillus paracasei), metabolites (tryptophan metabolism, linoleic acid, 10-keto-12Z-octadecenoic acid, Genipin, and Urolithin A), signaling pathways (IL-17 signaling pathway, the C-type lectin receptor signaling pathway and Toll-like receptor signaling pathway), and targets (L6, AKT1, PPARG, EGFR, CASP3, JUN, CXCL8, RELA, CYP1A1). The above findings are hoped to provide scientific basis for researching the potential mechanisms and possible treatment approaches for CKD-aP.
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Background: Hypervirulent Klebsiella pneumoniae (hvKp) is often characterized by hypervirulence and hypermucoviscosity, traits classically associated with the virulence regulators rmpA and rmpA2. However, some clinically virulent isolates lack these canonical regulators, suggesting the existence of non-canonical pathways driving hypervirulence. Objective: This study aimed to investigate the genetic basis of hypervirulence and hypermucoviscosity in clinical K. pneumoniae isolates that lack the rmpA and rmpA2. Methods: A total of 20 clinical isolates were collected. Virulence was assessed using the Galleria mellonella infection model. The hypermucoviscous phenotype was determined via the string test and mucoviscosity assay. Whole-genome sequencing (WGS) and comparative genomic analysis were performed to identify genetic variations, with a particular focus on an ST1-KL122 isolate (1744) that exhibited hypervirulence and hypermucoviscosity despite the absence of rmpA/rmpA2. Results: All 20 isolates induced at least 25% mortality in G. mellonella. Most isolates displayed a hypermucoviscous phenotype, except for isolates 2355, 2324, and 1951. WGS confirmed the absence of rmpA and rmpA2 in isolates 2355, 2324, 1951, and the hypermucoviscous isolate 1744. In the hypervirulent and hypermucoviscous ST1-KL122 isolate 1744, comparative genomic analysis identified two nonsynonymous mutations in the wzc gene, which encodes a tyrosine kinase involved in capsular polysaccharide (CPS) export: c.1397A>C (p.Asn466Thr) and c.1721A>T (p.His574Leu). Conclusions: Our findings provide evidence that mutations in the wzc represent a novel, non-canonical regulatory mechanism that drives CPS-mediated hypermucoviscosity and hypervirulence in rmpA/rmpA2-negative hvKp.