Western及IP细胞裂解液(含抑制剂)

Pseudomonas aeruginosa pneumonia poses a significant therapeutic challenge. Nanoparticles serve as an effective tool for nucleic acid delivery to efficiently alleviate pneumonia. This study develops a hyaluronic acid (HA)-coated peptide nanoparticle system for targeted delivery of small interfering RNA (siRNA) against Tudor domain-containing protein 9 ( TDRD9 ), identified via RNA sequencing of bronchoalveolar lavage fluid-derived neutrophils from 21 recruited patients (11 males/10 females). Adoptive transfer of TDRD9-silenced polymorphonuclear neutrophils into neutrophil-depleted male mice attenuates lung inflammation and edema. Mechanistically, TDRD9 suppresses neutrophil cuproptosis by upregulating programmed death ligand 1 (PD-L1) through interaction with CD80 to activate p38 mitogen-activated protein kinase (MAPK) signaling. HA-si-TDRD9 nanoparticles enhance neutrophil cuproptosis, reduce pulmonary neutrophil accumulation, and ameliorate lung injury via PD-L1/CD80/MAPK. Importantly, HA-si-TDRD9 nanoparticles reduce bacterial growth, apoptosis, and inflammation in human lung organoids. This work demonstrates that targeting TDRD9 with siRNA nanoparticle platform presents a promising therapeutic strategy for treating bacterial lung injury.

Bevacizumab (Bev) resistance limits therapeutic efficacy in ovarian cancer (OC) patients. We identified ESM1 as a key gene in Bev-resistant OC. ESM1 secreted by OC-resistant cell lines activates the ITGB1/FAK axis to induce neovascularization and Bev resistance. Additionally, ESM1 overexpression promoted the growth and Bev resistance of OC, lung, intestinal, and hepatocellular carcinoma tumors. Then, we identified TRIM28 as an upstream regulator that stabilizes ESM1 by promoting SUMOylation, inhibiting its proteasomal degradation. In OC mice, TRIM28 overexpression promotes angiogenesis and Bev resistance via ESM1-mediated ITGB1/FAK activation. This work unveils a new molecular pathway underlying Bev resistance in OC and proposes TRIM28 and ESM1 as potential therapeutic targets.

Summary Hypoxic pulmonary hypertension (HPH) lacks effective pharmacologic treatments. Microarray-based gene expression indicates the crucial role of Cullin 5 (Cul 5) in HPH. This study showed that Cul 5 was upregulated in HPH patients and a murine model of HPH. In vitro , Cul 5 promoted the angiogenesis and adhesion capacity of human pulmonary artery endothelial cells (PAECs), which could be mitigated by Cul 5 inactivation mediated by pevonedistat or NEDD8 silence. In vivo , silencing of Cul 5 in the endothelium and Cul 5 inactivation by pevonedistat could also alleviate hypoxic vascular remodeling. Mechanistic research showed that Cul 5 participated in HPH pathogenesis via the TRAF6/NF-κB/HIF-1α/VEGF pathway. Inhibition of the TRAF6/NF-κB/HIF-1α/VEGF pathway could reverse Cul 5-induced human PAEC dysfunction. These findings demonstrate that Cul 5 is an important mediator of HPH via the TRAF6/NF-κB/HIF-1α/VEGF pathway firstly, and could be considered as a potential therapeutic target in the clinical treatment of HPH.

Despite the documented involvement of neutrophil extracellular traps (NETs) in various cancer types, their specific role in meningioma development remains understudied. Here, we report a mechanism by which the JAM3/Mac-1 interaction promotes meningioma development via NETs. We found that JAM3 knockdown in meningioma cells suppressed AKT phosphorylation, thereby reducing neutrophil migration and NET formation in co-culture; these effects were rescued by the AKT activator SC79. Co-IP confirmed the JAM3/Mac-1 interaction. In a xenograft model, JAM3 knockdown inhibited tumor growth and intratumoral NETs. Furthermore, NETs enhanced meningioma cell growth in vitro and in vivo , an effect abolished by DNase I. Conversely, Mac-1 knockdown in neutrophils impaired their function, which was also restored by SC79. Clinically, JAM3 expression correlated with Mac-1 and NET markers in meningioma samples. In conclusion, JAM3 promotes meningioma development by regulating the formation of NET through the Mac-1/AKT axis. These findings offer novel insights for treating these common primary tumors of the central nervous system.

Background To investigate the effects and mechanism of action of the main component, Stigmasterol (STG), of Liuwei Dihuang Pill (LP) on postmenopausal osteoporosis (PMOP) in ovariectomized (OVX) rats. Methods Cell viability was assessed using the cell counting kit 8 (CCK-8) assay. Osteoblast differentiation was evaluated through alkaline phosphatase (ALP) activity assay, ALP staining, and Alizarin Red S staining. Expression levels were assessed by real-time quantitative PCR (RT-qPCR), Western blot, and immunohistochemistry. Immunofluorescence was used to observe protein colocalization. Protein–protein interactions were detected using co-immunoprecipitation (Co-IP) and Molecular docking and drug affinity responsive target stability (DARTS) assays. Additionally, an OVX rat model was established through ovariectomy surgery. Utilizing micro-computed tomography (CT), we scanned and analyzed the fine structure of the rat femoral trabeculae across various groups, thereby gathering pertinent data. Enzyme-linked immunosorbent assay (ELISA) was performed to determine the levels of E2, key bone immune regulatory markers, bone turnover markers, and pro-inflammatory factor in serum. Results The main components of LP, including Quercetin, Kaempferol, β-sitosterol, and STG, significantly promote osteogenic differentiation of osteoblasts, with STG having the most significant effect. Moreover, STG also notably enhances the upregulation of the Akt/mTOR/ p70S6 kinase (p70S6K) signaling pathway. Bioinformatics analysis predicted the potential ubiquitin ligase synoviolin 1 (SYVN1) for Akt, which was experimentally validated. Molecular docking between STG and SYVN1 verified their binding activity, and STG downregulated the expression of SYVN1. Overexpression of SYVN1 reversed the promoting effect of STG on osteoblast differentiation and activation of the Akt/mTOR/p70S6K pathway. Furthermore, STG upregulated uterine index and serum levels of E2, OPG, BALP, PINP, OCN, and CTX-1, and downregulated levels of RANKL, IL-6, and TNF-α in OVX rats. Meanwhile, STG promoted the expression of OCN and inhibited the expression of NFATc1 and cathepsin-K in the femurs of OVX rats. Additionally, STG activated the Akt/mTOR/p70S6K pathway in the femurs of OVX rats. However, overexpressing SYVN1 reversed the effects of STG on OVX rats. Conclusion STG upregulated the Akt/mTOR/p70S6K signaling via SYVN1 to alleviate osteoporosis in OVX rats. Therefore, this study will provide new insights into discovering novel approaches for treating PMOP.

While circRNAs have been demonstrated to play critical roles in various neurological disorders, their functional mechanisms in post-intraventricular hemorrhage (IVH) communicating hydrocephalus remain poorly understood. Here we report a circular RNA, circular RNA homeodomain interacting protein kinase 3 (circHIPK3), significantly upregulated in meningeal tissues of IVH rat models. Knockdown of circHIPK3 markedly attenuated meningeal fibrosis, reduced ventricular dilation, and improved neurological function in IVH rats. Using both IVH rat models and thrombin-induced astrocyte cell lines, we validated the functional role of circHIPK3 in the pathogenesis of IVH-induced communicating hydrocephalus. Mechanistically, circHIPK3 functions as an endogenous sponge of miR-30a-3p to decrease its activity, resulting in upregulation of ACT1(TRAF3IP2) expression, which in turn triggers TGF-β1/Smad3 signaling pathway activation, ultimately driving fibrotic progression after IVH. Collectively, our findings suggest that circHIPK3 and its coupling mechanism are implicated in IVH, providing compelling evidence that circHIPK3 could be a key therapeutic target against post-IVH communicating hydrocephalus.