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关键词： angiogenesis   human microvascular endothelial cells   human umbilical vein endothelial cells   interleukin‐6   pentraxin 3  

摘要： Interleukin-6 (IL-6) is a pleiotropic cytokine primarily involved in inflammation and vascular remodeling. While it is known to influence angiogenesis, the formation of new blood vessels, the underlying mechanisms and downstream targets in endothelial cells (ECs) remain poorly understood. This study aimed to explore the angiogenic effect of IL-6 in human umbilical vein endothelial cells (HUVECs) and human microvascular endothelial cells (HMEC-1 cells) and to gain insights into the mechanistic pathways involved. We found that IL-6 promoted EC migration in a wound healing assay and stimulated tube formation in both cell types, while also enhancing blood vessel formation in the chicken chorioallantoic membrane assay. Proteomic analysis of IL-6-treated HUVECs identified pentraxin 3 (PTX3) as an IL-6-responsive factor. Subsequent validations using ELISA, qRT-PCR, and western blot confirmed increased PTX3 expression in HUVECs but revealed a downregulation in HMEC-1 cells, indicating a cell-type-specific regulatory effect of IL-6. To further elucidate the cellular response to IL-6, we assessed the expression of the IL-6 receptor subunits, IL-6Rα and gp130, in both EC types following IL-6 stimulation. qRT-PCR revealed increased gp130 gene expression, but flow cytometry showed no change in cell surface expression, while IL-6Rα surface expression increased in both EC types, supporting sustained IL-6 signaling. Overall, these findings provide updated insights into the pro-angiogenic actions of IL-6 and how it regulates PTX3 expression in human ECs.

关键词： Cytokine   Inflammation   Neurotoxicity   Oxidative stress   Sodium/glucose cotransporter 2 inhibitor  

摘要： Background: Neuroinflammation drives neuronal degeneration and cognitive decline through various mechanisms. Interleukin (IL)-17A, secreted by Th17 cells, activates glycogen synthase kinase 3 beta (GSK3 beta), worsening neuroinflammation via the nuclear factor kappa B (NF-kappa B) pathway. Dapagliflozin (DPG), a sodium/ glucose cotransporter 2 inhibitor with neuroprotective effects, reduces oxidative stress, apoptosis, and inflammation. This study examines DPG's molecular impact on neuroinflammation in an LPS-induced rat model. Methods: The experiment was designed with four groups as control, lipopolysaccharide (LPS) (5 mg/kg, intraperitoneal), LPS + DPG (10 mg/kg via oral gavage), and DPG, with a total of thirty-two female Wistar Albino rats. After five days of treatment, the rats were euthanized. Brain and cerebellum tissues were gathered for biochemical analysis to examine oxidative stress parameters spectrophotometrically;histological and immunostaining analysis focusing on caspase-9 (Cas-9), NF-kappa B, and IL-10 immunoexpressions, and genetic analysis as IL-17A, GSK3 beta, IL-6, and cyclooxygenase 2 gene expressions by qRT-PCR. Results: Histopathological evaluation revealed hyperemia, edema, mild degeneration, neuronal death, and modest gliosis in the LPS group. Increment of Cas-9 and NF-kappa B immunoexpressions, oxidative stress parameters, and the mRNA expressions of all four genes, in addition to the decrement of IL-10 and total antioxidant status, have been observed. DPG treatment significantly reversed all these findings and protected the neuronal tissues against LPS-induced substantial neuronal damage. Conclusion: In conclusion, DPG may exert neuroprotective effects through the involvement of the IL-17A/GSK3 beta pathway and reduction of oxidative stress;however, further studies are needed to clarify causal mechanisms and broader anti-inflammatory actions.

关键词： Diamine oxidase   IFN-gamma   NK cells   Inflammation  

摘要： Aims: Diamine oxidase (DAO), a well-established biomarker for intestinal damage, histamine intolerance or tumorigenesis, has rarely been reported in immune regulation. This study aimed to identify DAO as a critical enhancer of abnormal inflammation by promoting interferon-gamma (IFN-gamma) production from natural killer (NK) cells. Main methods: Clinical bioinformatics analyzed aoc1 (DAO-coding gene) expression in PBMCs from patients with inflammatory diseases. Murine models (LPS-induced systemic inflammation, sepsis, DSS-induced colitis) using DAO-/- mice, alongside DAO-/- NK92 cells and DAO inhibitor DIZE, were employed for phenotypic validation. Cellular profiling, bone marrow chimeras, reciprocal transplantation, RNA-sequence, non-targeted metabolomics, and flow cytometry were utilized to dissect DAO's mechanisms in NK cells. Key findings: DAO deficiency protected mice from inflammatory pathology by suppressing IFN-gamma production. NK cells were identified as the primary target cells during the process, with DAO acting intracellularly to promote IFN-gamma via a reactive oxygen species (ROS)-autophagy axis. DAO-derived ROS, distinct from mitochondrial or NOX2 sources, enhanced autophagic flux during NK activation, enabling IFN-gamma biosynthesis. DAO did not affect NK homeostasis, including maturation, proliferation, or receptor expressions. Significance: DAO is a novel risk factor in inflammatory diseases, driving IFN-gamma production through ROSautophagy signaling in NK cells. Targeting DAO may offer therapeutic strategies for conditions involving dysregulated IFN-gamma responses, including sepsis, colitis, and autoimmune disorders.

关键词： JAK2/STAT3 signaling pathway   interferon‐gamma   jawbone marrow stromal cells   methyltransferase‐like 3   osteogenic differentiation   postmenopausal osteoporosis  

摘要： To investigate whether interferon-gamma (IFN-γ) alleviates postmenopausal osteoporosis (POP) by regulating METTL3 via the JAK2/STAT3 pathway to enhance osteogenic differentiation of jawbone marrow stromal cells (JBMSCs). Ovariectomized (OVX) rats received IFN-γ (5000 IU/dose, 3×/week for 24 weeks), with jawbone mass assessed via micro-CT and HE staining. JBMSCs were cultured, and osteogenic differentiation under IFN-γ (optimal concentration: 10 ng/mL) was evaluated using qRT-PCR, ALP/alizarin red staining, and CCK-8. METTL3's role was analyzed via lentiviral overexpression (OE-METTL3) or knockdown (sh-METTL3). JAK2/STAT3 pathway activity was tested using AG490 (JAK2 inhibitor) and Western blot for phosphorylated JAK2/STAT3. IFN-γ reduced alveolar bone loss in OVX rats, increased osteocytes, and decreased osteoclasts. In JBMSCs, 10 ng/mL IFN-γ upregulated osteogenic genes (Runx2, OPN, OPG) and mineralization. METTL3 expression was elevated by IFN-γ; sh-METTL3 suppressed Runx2, while OE-METTL3 reversed OVX-induced osteogenic inhibition. IFN-γ activated JAK2/STAT3 signaling, but AG490 reduced both osteogenic gene expression and METTL3 levels. METTL3 overexpression amplified JAK2/STAT3 phosphorylation, synergizing with IFN-γ to enhance osteogenesis. IFN-γ activates the JAK2/STAT3 pathway to drive METTL3 transcription, upregulating osteogenic genes and mitigating jawbone loss in POP. METTL3 overexpression counteracts estrogen deficiency-induced osteogenic suppression and forms a positive feedback loop with JAK2/STAT3. AG490's complete blockade of IFN-γ/METTL3 effects confirms the JAK2/STAT3-METTL3 axis as central to bone immune metabolism, highlighting therapeutic potential for targeting this pathway in POP.

关键词： Neurodegeneration   Noradrenergic   Axon   Acrylamide   Microglia   Oxidative stress   TNF-alpha  

摘要： Environmental toxicants such as acrylamide or 1-bromopropane induce cognitive dysfunction in humans. We previously reported specific noradrenergic neuronal degeneration induced by acrylamide or 1-bromopropane in rodents. In this study, we applied in vivo and in vitro models as well as bulk and single-cell transcriptomic analyses to uncover the underlying mechanisms. RNA-seq of brains of acrylamide-exposed mice revealed a transcriptomic profile involving genes related to multiple neurodegenerative diseases and oxidative stress pathways. Single-cell RNA-seq for microglia identified upregulation of immunoregulation-, inflammation-, and oxidative stress- related pathways, and identified the upregulation of Tnfaip2 (a TNF-alpha effector), in multiple microglial sub-clusters. Further results of our in vitro interaction model showed that compared to direct acrylamide exposure, exposure to conditioned medium (CM) of acrylamide-exposed BV2 microglia significantly decreased 1C11NE axon density, and RNA-seq for 1C11NE identified similar transcriptomic profiles to those of brains of acrylamide-exposed mice. RNA-seq for BV2 microglia showed upregulation of various oxidative stress related genes. Further inhibition experiments demonstrated that TNF-alpha inhibition or anti-oxidation alleviated acrylamide-induced axonal degeneration in 1C11NE neurons. Finally, in vivo TNF-alpha knockout alleviated acrylamide-induced neurotoxicity. Our study demonstrated that acrylamide-induced noradrenergic axon degeneration is promoted via a non-cell autonomous mechanism, involving microglial Tnfaip2/TNF-alpha and

关键词： 17H   17α-hydroxyprogesterone   ACTH   Adrenocorticotropic hormone   AMH   Anti-Müllerian hormone   AND   Androstenedione   APDC   Ammonium pyrrolidine dithiocarbamate   AUC   Area under the curve   BMI   Body mass index   BP   Biological Process   CC   Cellular Component   Cd   Cadmium   CI   Confidence intervals   CRH   Corticotropin-releasing hormone   CTD   Comparative Toxicogenomics Database   DHE   Dehydroepiandrosterone sulfate   DisGeNET   Disease Gene Network   DRC   Dynamic reaction cell   ES1   Estrone sulfate   ESO   Estrone   EST   Estradiol   FDR   False discovery rate   FSH   Follicle-stimulating hormone   FSHR   Follicle-stimulating hormone receptor   GnRH   Gonadotropin-releasing hormone   GO   Gene Ontology   Hg   Mercury   HPA   Hypothalamic-pituitary-adrenal   HPG   Hypothalamic-pituitary-gonadal   HPLC-MS/MS   High performance liquid chromatography-tandem mass spectrometry   ICP-MS   Inductively Coupled Plasma Mass Spectrometry   ID-LC-MS/MS   Isotope dilution liquid chromatography-tandem mass spectrometry   KEGG   Kyoto Encyclopedia of Genes and Genomes   LC-MS/MS   Liquid chromatography-tandem mass spectrometry   LLOD   Lower limit of detection   LUH   Luteinizing hormone   MF   Molecular Function   Mn   Manganese   NHANES   National Health and Nutrition Examination Survey   Pb   Lead   PG4   Progesterone   PIR   Poverty income ratio   PPI   Protein-protein interaction   RCS   Restricted cubic splines   ROC   Receiver operating characteristic   Se   Selenium   SHBG   Sex hormone-binding globulin   SRM   Selected reaction monitoring   SWAN   Study of Women's Health Across the Nation   TMAH   Tetramethylammonium hydroxide   TNF   Tumor necrosis factor   TST   Testosterone   Bioinformatic analysis   Effect modification   Folate   Metals   Sex hormones  

摘要： Background Hormonal disruption evidence of metal exposure is lacking and contradictory. Objectives To elucidate the association of multiple metals exposure with a comprehensive panel of sex hormones in women across the lifespan. Methods 5492 women aged 3–80 from a national survey were included. Multivariable linear regression, restricted cubic splines, and receiver operating characteristic curves were employed to assess individual metal exposure with sex hormones. Quantile-based g computation was used to explore mixture exposure association. Vitamin D and folate were examined as the modifiers. Integrative bioinformatics analysis was conducted leveraging a network of databases to reveal underlying mechanisms. Results Exposure to cadmium, lead (Pb), mercury (Hg), selenium, and manganese was associated with sex hormones. For example, in young adult (20–49 years), Pb was associated with androstenedione [3.89 %, 95 % CI: 0.81, 7.06], estrone sulfate [8.14 %, 95 % CI: 1.54, 15.18], and luteinizing hormone [12.29 %, 95 % CI: 5.01, 20.07]; Hg was associated with estrone sulfate [4.84 %, 95 % CI: 0.62, 9.22]. Metals mixture was mainly associated with young adults’ increased sex hormones with effect ranging from 5.37 % (95 % CI: 1.77, 9.10) to 17.83 % (95 % CI: 9.04, 27.34). Folate modified such associations with hazardous effect appeared in high folate level in young adults, while in low folate level in other age groups. Inflammatory signaling pathways such as TNF and IL-17 may mediate metal-induced sex hormones disturbance. Conclusions Our findings highlight the significant associations, folate modifications, and potential pathways of metal exposure on female sex hormones, which provides insights for prevention avenues and mechanisms.

关键词： 1,2-Napthoquinone   Dendritic cell   IFN-γ   MHC-I   CD8 T cell   APC   allophycocyanin   BQ   benzoquinone   CIITA   class II major histocompatibility complex transactivator   DEM   diethyl maleate   FITC   fluorescein isothiocyanate   GAPDH   glyceraldehyde-3-phosphate dehydrogenase   IFN   interferon   MHC   major histocompatibility complex   JAK   janus kinase   MD   menadione   NAC   N-acetylcysteine   NQ   napthoquinone   NLRC5   NOD-, LRR- and CARD-containing 5   OVA   ovalbumin   PE   phycoerythrin   RT-PCR   reverse transcription-polymerase chain reaction   ROS   reactive oxygen species   STAT   signal transducers and activator of transcription   TLR   toll like receptor  

摘要： Dendritic cells play a crucial role in immune responses by capturing pathogens and presenting antigens to T cells via major histocompatibility complex (MHC) molecules, thus triggering adaptive immune responses. 1,2-naphthoquinone (1,2-NQ), a quinone found in diesel exhaust and cigarette smoke, has various physiological functions. In this study, we investigated the effect of 1,2-NQ on the expression of antigen presentation-related molecules in the dendritic cell line DC2.4. The results revealed that 1,2-NQ enhanced the IFN-γ-induced upregulation of MHC-I expression at the transcriptional level. Moreover, it upregulated the expression of NLRC5, a transcriptional activator of MHC-I. 1,2-NQ is a reactive oxygen species (ROS) producing reagent. The 1,2-NQ-induced upregulation of MHC-I expression and downregulation of MHC-II expression were abolished by the ROS scavenger N-acetylcysteine. Similar effects on MHC expression were also observed with ROS-inducing reagents, such as paraquat and diethyl maleate. In addition, dendritic cells stimulated with 1,2-NQ exhibited enhanced efficacy in CD8 T cell activation, which was accompanied by increased IFN-γ production by T cells. These findings demonstrate that 1,2-NQ enhances the IFN-γ-induced activation of dendritic cells and promotes the activation of CD8 T cells.

关键词： class I   CRISPR   gene editing   genome editing   immunogenicity   immunotoxicity   MHC I  

摘要： In vivo genome editing with CRISPR-Cas9 systems is generating worldwide attention and enthusiasm for the possible treatment of genetic disorders. However, the consequences of potential immunogenicity of the bacterial Cas9 protein and the AAV capsid have been the subject of considerable debate. Here, we model the antigen presentation in cells after in vivo gene editing by in vitro transduction of a human cell line with an AAV2 vector that delivers the Staphylococcus aureus Cas9 transgene. Through HLA class I enrichment, peptide elution, and highly sensitive LC-MS interrogation, we identified a highly conserved saCas9-derived T cell epitope in the catalytic domain of the enzyme that is restricted to HLA-A*02:01 and induces CD8+ T cell activation and killing. We conclude that AAV delivery of Cas9 results in presentation of a T cell epitope that can activate CD8+ cells and induce killing of the transduced cell, with important ramifications for in vivo genome editing strategies.

关键词： IL-12 mRNA   Lipid nanoparticle   IFN gamma-IDO pathway   Cancer immunotherapy   Prodrug synthesis  

摘要： The administration of recombinant cytokines, particularly interleukin-12 (IL-12), holds promising clinical potential for treating various cancers. Sustained intratumoral delivery of IL-12 can restore tumor resident CD8+ effector T cells and induce the priming of antitumor CD8+ effector T cells. However, these CD8+ T cell-dependent anticancer efficacy is usually transient and accompanies the activation of immune suppressive CD4+Foxp3+ T regulatory cells. The underlying mechanism of T regulatory cell activation in IL-12 therapy is the upregulation of IFN gamma dependent indoleamine 2,3-dioxygenase (IDO) expression. Due to this negative feedback, the combinatorial use of drugs should be considered to enhance the efficacy of IL-12-mediated therapy. Herein, we designed a lipid nanoparticle (LNP) system which can successfully deliver an IDO inhibitor indoximod (IND) and IL-12 encoding mRNA. In order to increase the loading efficiency, the IND prodrug was synthesized by conjugating IND with cholesterol by ester linkage. Optimized IND prodrug encapsulating LNP successfully transfected tumor cells and macrophages, resulting in the secretion of IL-12 cytokine. With IL-12 transfection, macrophages upregulated T cell co-stimulation factor and released TNF alpha cytokine, indicating that the tumor microenvironment could be changed from cold tumor to hot tumor for immunotherapy. Also, by the elevated secretion of IL-12 cytokine, T cells release high levels of IFN gamma, which is a central role in IL-12-mediated immunotherapy. This co-delivery system presents a promising strategy to overcome the limitations of single IL-12-mediated therapy by simultaneously promoting antitumor immune responses and inhibiting immunosuppressive mechanisms, thereby enhancing the overall efficacy of cancer immunotherapy.