苏木素伊红(HE)染色液

Background Hypertensive intracerebral hemorrhage (HICH) is a life-threatening disease and lacks effective treatments. Previous studies have confirmed that metabolic profiles altered after ischemic stroke, but how brain metabolism changes after HICH was unclear. This study aimed to explore the metabolic profiles after HICH and the therapeutic effects of soyasaponin I on HICH.Methods HICH model was established first. Hematoxylin and eosin staining was used to estimate the pathological changes after HICH. Western blot and Evans blue extravasation assay were applied to determine the integrity of the blood–brain barrier (BBB). Enzyme-linked immunosorbent assay was used to detect the activation of the renin–angiotensin–aldosterone system (RAAS). Next, liquid chromatography–mass spectrometry-untargeted metabolomics was utilized to analyze the metabolic profiles of brain tissues after HICH. Finally, soyasaponin I was administered to HICH rats, and the severity of HICH and activation of the RAAS were further assessed.Results We successfully constructed HICH model. HICH significantly impaired BBB integrity and activated RAAS. HICH increased PE(14:0/24:1(15Z)), arachidonoyl serinol, PS(18:0/22:6(4Z, 7Z, 10Z, 13Z, 16Z, and 19Z)), PS(20:1(11Z)/20:5(5Z, 8Z, 11Z, 14Z, and 17Z)), glucose 1-phosphate, etc., in the brain, whereas decreased creatine, tripamide, D-N-(carboxyacetyl)alanine, N-acetylaspartate, N-acetylaspartylglutamic acid, and so on in the hemorrhagic hemisphere. Cerebral soyasaponin I was found to be downregulated after HICH and supplementation of soyasaponin I inactivated the RAAS and alleviated HICH.Conclusion The metabolic profiles of the brains changed after HICH. Soyasaponin I alleviated HICH via inhibiting the RAAS and may serve as an effective drug for the treatment of HICH in the future.

Metabolic dysfunction-associated steatohepatitis (MASH) is a complex liver disease whose pathogenesis involving endoplasmic reticulum (ER) stress and ferroptosis. However, key regulatory genes remain poorly understood, hindering the development of effective therapeutic targets. This study aims to identify genes linked to ER stress and ferroptosis through bioinformatics and experimental validation, providing insights into MASH pathogenesis and potential therapeutic strategies. We first identified ER stress and ferroptosis as key processes in MASH through differential analysis and functional enrichment. This was subsequently validated in a high-fat diet (HFD)-induced MASH model in ApoE -/- mice, where ER stress and ferroptosis were confirmed to occur in the liver tissue of MASH mice. Additionally, daily intraperitoneal injection of the ferroptosis inhibitor ferrostatin-1 (Fer-1) alleviated MASH progression. In vitro, Fer-1 mitigated inflammation, lipid accumulation, and fibrosis in free fatty acid (FFA)-treated HepG2 cells. To identify key genes, we employed bioinformatics analysis and machine learning approaches, which led to the identification of cyclin dependent kinase inhibitor 1A ( CDKN1A) and early growth response 1 ( EGR1) as feature genes associated with MASH-related ER stress and ferroptosis. Increased expression of CDKN1A and decreased expression of EGR1 were observed in the liver tissue of MASH mice and FFA-treated HepG2 cells. Furthermore, in CDKN1A overexpression and EGR1 silencing cell models, treatment with the ER stress inhibitor 4-Phenylbutyric acid improved the ferroptosis. In summary, all results indicate that CDKN1A and EGR1 are key genes driving ER stress-induced ferroptosis in MASH. Our findings not only provide new evidence for the pathogenesis of MASH but also highlight novel therapeutic targets for intervention.

Objectives To unravel the heterogeneity and function of microenvironmental neutrophils during intervertebral disc degeneration (IDD). Methods Single-cell RNA sequencing (scRNA-seq) was utilized to dissect the cellular landscape of neutrophils in intervertebral disc (IVD) tissues and their crosstalk with nucleus pulposus cells (NPCs). The expression levels of macrophage migration inhibitory factor (MIF) and ACKR3 in IVD tissues were detected. The MIF/ACKR3 axis was identified and its effects on IDD were investigated in vitro and in vivo . Results We sequenced here 71520 single cells from 5 control and 9 degenerated IVD samples using scRNA-seq. We identified a unique cluster of neutrophils abundant in degenerated IVD tissues that highly expressed MIF and was functionally enriched in extracellular matrix organization (ECMO). Cell-to-cell communication analyses showed that this ECMO-neutrophil subpopulation was closely interacted with an effector NPCs subtype, which displayed high expression of ACKR3. Further analyses revealed that MIF was positively correlated with ACKR3 and functioned via directly binding to ACKR3 on effector NPCs. MIF inhibition attenuated degenerative changes of NPCs and extracellular matrix, which could be partially reversed by ACKR3 overexpression. Clinically, a significant correlation of high MIF/ACKR3 expression with advanced IDD grade was observed. Furthermore, we also found a positive association between MIF + ECMO-neutrophil counts and ACKR3 + effector NPCs density as well as higher expression of the MIF/ACKR3 signaling in areas where these two cell types were neighbors. Conclusions These data suggest that ECMO-neutrophil promotes IDD progression by their communication with NPCs via the MIF/ACKR3 axis, which may shed light on therapeutic strategies.

N6-methyladenosine (m6A) modification, a dynamically reversible epigenetic mechanism, is implicated in pulmonary fibrosis (PF) progression. The function and molecular mechanisms of m6A reader, insulin-like growth factor 2 mRNA binding protein 1 (IGF2BP1) in PF remain elusive. This study investigates the mechanistic contributions of IGF2BP1 to PF development. We found IGF2BP1 was overexpressed in macrophages of PF mice. IGF2BP1 knockdown markedly attenuated bleomycin (BLM)-induced lung pathology, as evidenced by reduced inflammatory cell infiltration, fibroblast accumulation, Ashcroft fibrosis scores, and hydroxyproline deposition. Furthermore, IGF2BP1 knockdown downregulated PF-associated markers in lung tissues and embryonic lung fibroblasts (ELFs), including TGF-β1, α-SMA, Collagen-I/III, Arg1, CCL18, Ym1, CD163, IL-6, IL-1β, and TIMP1, while decreasing the CD68 + /CD163 + macrophage proportion. Mechanistic studies revealed that IGF2BP1 bound to and stabilized thrombospondin-1 (THBS1) in an m6A-dependent manner. THBS1 overexpression rescued the suppression of macrophage M2 polarization caused by IGF2BP1 knockdown. Additionally, THBS1 overexpression counteracted IGF2BP1 knockdown-mediated inhibition of glycolysis, restoring HK2, LDHA, and PKM2 expression, lactate/glucose metabolism, and ATP production. Intriguingly, THBS1 physically interacted with toll-like receptor 4 (TLR4), and TLR4 overexpression reversed the inhibitory effect of THBS1 knockdown on macrophage M2 polarization and glycolytic reprogramming. Collectively, our findings demonstrate that IGF2BP1 drives PF progression by stabilizing THBS1 mRNA via m6A modification, thereby promoting TLR4-mediated macrophage M2 polarization and glycolytic activation. This study unveils a novel IGF2BP1/THBS1/TLR4 regulatory axis in PF pathogenesis, offering potential therapeutic targets.

Background Quinic acid (QA) and its derivatives have good lipid-lowering and hepatoprotective functions, but their role in atherosclerosis remains unknown. This study attempted to investigate the mechanism of QA on atherogenesis in Apoe−/− mice induced by HFD. Methods HE staining and oil red O staining were used to observe the pathology. The PCSK9, Mac-3 and SM22a expressions were detected by IHC. Cholesterol, HMGB1, TIMP-1 and CXCL13 levels were measured by biochemical and ELISA. Lipid metabolism and the HMGB1-SREBP2-SR-BI pathway were detected by PCR and WB. 16 S and metabolomics were used to detect gut microbiota and serum metabolites. Results QA or low-frequency ABX inhibited weight gain and aortic tissue atherogenesis in HFD-induced Apoe−/− mice. QA inhibited the increase of cholesterol, TMA, TMAO, CXCL13, TIMP-1 and HMGB1 levels in peripheral blood of Apoe−/− mice induced by HFD. Meanwhile, QA or low-frequency ABX treatment inhibited the expression of CAV-1, ABCA1, Mac-3 and SM22α, and promoted the expression of SREBP-1 and LXR in the vascular tissues of HFD-induced Apoe−/− mice. QA reduced Streptococcus_danieliae abundance, and promoted Lactobacillus_intestinalis and Ileibacterium_valens abundance in HFD-induced Apoe−/− mice. QA altered serum galactose metabolism, promoted SREBP-2 and LDLR, inhibited IDOL, FMO3 and PCSK9 expression in liver of HFD-induced Apoe−/− mice. The combined treatment of QA and low-frequency ABX regulated microbe-related Glycoursodeoxycholic acid and GLYCOCHENODEOXYCHOLATE metabolism in HFD-induced Apoe−/− mice. QA inhibited TMAO or LDL-induced HCAECs damage and HMGB1/SREBP2 axis dysfunction, which was reversed by HMGB1 overexpression. Conclusions QA regulated the gut-liver lipid metabolism and chronic vascular inflammation of TMA/TMAO through gut microbiota to inhibit the atherogenesis in Apoe−/− mice, and the mechanism may be related to the HMGB1/SREBP2 pathway.

Ulcerative colitis (UC) is a recurrent inflammatory disease without a specific cure or treatment for improvement. Here, we investigated the potential therapeutic effect and mechanism of ginsenoside Rg3 (Gin Rg3) on UC. We constructed an in vitro cellular inflammatory model and a dextran sulfate sodium (DSS)-induced UC mouse model. We also used Gin Rg3, MCC950 (NLRP3 inhibitor), MSU (NLRP3 activator), and fecal transplantation (FMT) to intervene the model. The results showed that Gin Rg3 inhibited NLRP3 inflammasome activation, pyroptosis, and apoptosis in vitro and in vivo. DSS-induced changes in the abundance of gut microbiota at the phylum or genus level were partially restored by Gin Rg3. Furthermore, gin Rg3 affected intestinal metabolism in mice by inhibiting the activation of NLRP3 inflammasome. The gut microbiota treated with Gin Rg3 was sufficient to alleviate DSS-induced UC. In summary, Gin Rg3 alleviated DSS-induced UC by inhibiting NLRP3 inflammasome activation and regulating gut microbiota homeostasis.

Background Pulmonary arterial hypertension (PAH) is characterized by lipid accumulation and mitochondrial dysfunction. This study was designed to investigate the effects of hypoxia-inducible factor-1α (HIF-1α) on fatty acid uptake and mitophagy in PAH. Methods Peripheral blood samples were obtained from PAH patients. Human pulmonary arterial smooth muscle cells and rat cardiac myoblasts H9c2 were subjected to hypoxia treatment. Male Sprague–Dawley rats were treated with monocrotaline (MCT). Right ventricular systolic pressure (RVSP), right ventricular hypertrophy index (RVHI), pulmonary artery remodeling, and lipid accumulation were measured. Cell proliferation and ROS accumulation were assessed. Mitochondrial damage and autophagosome formation were observed. Co-immunoprecipitation was performed to verify the interaction between HIF-1α and CD36/PI3K p85α. Results HIF-1α, CD36, Parkin, and PINK1 were upregulated in PAH samples. HIF-1α knockdown or PI3K p85α knockdown restricted the expression of HIF-1α, PI3K p85α, Parkin, PINK1, and CD36, inhibited hPASMC proliferation, promoted H9c2 cell proliferation, reduced ROS accumulation, and suppressed mitophagy. CD36 knockdown showed opposite effects to HIF-1α knockdown, which were reversed by palmitic acid. The HIF-1α activator dimethyloxalylglycine reversed the inhibitory effect of Parkin knockdown on mitophagy. In MCT-induced rats, the HIF-1α antagonist 2-methoxyestradiol (2ME) reduced RVSP, RVHI, pulmonary artery remodeling, lipid accumulation, and mitophagy. Recombinant CD36 abolished the therapeutic effect of 2ME but inhibited mitophagy. Activation of Parkin/PINK1 by salidroside (Sal) promoted mitophagy to ameliorate the pathological features of PAH-like rats, and 2ME further enhanced the therapeutic outcome of Sal. Conclusion PI3K p85α/HIF-1α induced CD36-mediated fatty acid uptake and Parkin/PINK1-dependent mitophagy to accelerate the progression of experimental PAH. Graphical Abstract

Background: While programmed death-ligand 1 (PD-L1)-targeted immunotherapy represents an advancement in non-small cell lung cancer (NSCLC), patient outcomes remain suboptimal. Aberrant activation of the cyclic adenosine monophosphate (cAMP) response element binding protein (CREB)-regulated transcription coactivator (CRTC) is linked to malignant proliferation and functionality in lung cancer cells. This study investigates the involvement of CRTC1 in tumor immunity.Methods: CRTC1 and Notch1 expression were regulated in A549 and NCI-H1299 NSCLC lines through plasmid-mediated overexpression/silencing to assess their effects on cell viability, apoptosis, migration, and invasion. CRTC1/Notch1-dysregulated Lewis lung carcinoma (LLC) cells were co-cultured with T cells to evaluate T cell activation and function. The efficacy of combined CRTC1 knockdown/overexpression and atezolizumab (anti-PD-L1) was tested in an LLC xenograft mouse model.Results: CRTC1 promoted cell viability, migration, and invasion while suppressing apoptosis across NSCLC models. In LLC cells, CRTC1 upregulated tumor cell PD-L1 expression, suppressed T cell-derived IFN-γ and IL-2 production, diminished endogenous CXCL10/11 secretion, and impaired T cell proliferation and cytotoxicity. Mechanistically, CRTC1 interacted with Notch1 to activate the Notch1/Akt pathway, stimulating PD-L1 upregulation, thereby facilitating tumor immunosuppression and growth. Notably, CRTC1 overexpression reversed the protective effects of atezolizumab on tumor growth. Combining CRTC1 knockdown with atezolizumab synergistically enhanced anti-tumor T cell immunity, achieving the most significant tumor regression in xenografts.Conclusion: These findings indicate that CRTC1 in tumor cells suppresses PD-L1-mediated anti-tumor immunity and promotes tumorigenesis via the Notch1/Akt signaling axis. Dual targeting of CRTC1 and PD-L1 demonstrates therapeutic synergy, suggesting CRTC1 pathway inhibition could optimize immunotherapy outcomes in NSCLC patients.

Background Intervertebral disc (IVD) degeneration (IDD) represents a predominant origin of low back pain and disability, yet current therapeutic interventions remain suboptimal. Emerging evidence highlights autophagy activation as a therapeutic strategy against IDD. This study investigates the mechanistic interplay between N6-methyladenosine (m6A) modifications and autophagy dysregulation in IDD pathogenesis. Methods Bioinformatics analysis identified ring finger protein 41 ( RNF41 ) as a key autophagy-IDD intersection gene. Functional validation utilized tert-butyl hydroperoxide (TBHP)-treated human nucleus pulposus (NP) cells to assess RNF41’s effects on senescence (CDKN2A), autophagy (LC3-II/p62), apoptosis (TUNEL), inflammation (IL-18/IL-1β), and extracellular matrix (ECM) homeostasis (aggrecan/MMP). Key m6A regulators modulating autophagy were screened via correlation analysis. In vivo validation employed adeno-associated virus (AAV)-mediated methyltransferase-like 3 (METTL3)/RNF41 delivery in puncture-induced IDD rat models. Results RNF41 expression was downregulated in human IVD tissues. Overexpression of RNF41 mitigated TBHP-induced senescence, apoptosis, activated AMPK/mTOR-mediated autophagy, suppressed inflammation, and restored ECM balance. The autophagy inhibitor chloroquine (CQ) abolished the protective effects of RNF41 overexpression on degenerative NP cells. Mechanistically, METTL3/YTHDC1 co-regulation in degenerative NP cells mediated m6A hypermethylation of RNF41 mRNA, shortening its half-life via YTHDC1-dependent decay. Intradiscal METTL3-silencing AAV attenuated puncture-induced disc loss and histopathological degeneration, whereas RNF41-silencing AVV exacerbated ECM disruption and annular disorganization. Conclusion METTL3/YTHDC1-mediated m6A modification drives IDD progression by silencing RNF41 , thereby impairing autophagy and ECM integrity. Targeting this axis offers a clinically actionable strategy to delay disc degeneration, particularly in patients with early-stage IDD. This evidence establishes RNF41’s role as a theragnostic biomarker and therapeutic targe, enabling precision-guided interventional approaches.

Background Docetaxel (DTX) resistance reduces therapeutic efficacy in prostate cancer (PCa). Accumulating reports support the role of phytochemicals in the reversal of DTX resistance. This study aimed to determine whether Epimedium brevicornu and Curcuma zedoaria extracts (ECe), specially icariin-curcumol, attenuates DTX resistance and explore their potential mechanisms.Methods Regulatory pathways were predicted between ECe active ingredients and PCa using network pharmacology. DTX-resistant cell LNCaP/R were established based on DTX-sensitive LNCaP, and xenograft models were further established. Active ingredients in ECe by HLPC-MS were identified. The binding of icariin and curcumol to the target was analyzed by molecular docking. Biochemical experiments were applied to determine the possible mechanisms by which Icariin-Curcumol regulates DTX sensitivity.Results Akt1 and the PI3K-Akt signaling pathway were predicted as the primary functional target between drug and PCa. ECe and DTX inhibited xenograft tumor growth, inflammation, cell viability and promoted apoptosis. Icariin and curcumol were detected in ECe, and icariin and curcumol docked with Akt1. ECe, Icariin-Curcumol and DTX downregulated AR, PSA, PI3K, Akt1, mTOR, and HIF-1ɑ. Moreover, ECe, Icariin-Curcumol and DTX increased glucose and PDH, decreased lactic acid, ATP and LDH, and downregulated c-Myc, hnRNPs, VEGF, PFK1, and PKM2. Notably, the anti-PCa effect of DTX was attenuated compared to ECe or Icariin-Curcumol in the LNCaP/R model. The combined effect of Icariin-Curcumol and DTX was superior to that of DTX.Conclusion Our data support that Icariin-Curcumol reverses DTX resistance by inhibiting the PI3K-Akt signaling and the Warburg effect, providing new ideas for improving therapeutic measures for PCa.