β-actin Recombinant Mouse Monoclonal Antibody

Glaucoma is a leading cause of irreversible blindness, characterized by the progressive degeneration of retinal ganglion cells (RGCs) and optic nerve damage. Although current intraocular pressure (IOP)-lowering therapies exhibit varying degrees of efficacy in preventing RGC loss, neurodegeneration can persist even in cases where IOP remains within the normal range, underscoring the need for direct neuroprotective strategies. Emerging evidence suggests that neuroinflammation mediated by retinal microglia plays a pivotal role in the pathogenesis of glaucoma. Here, a nucleic acid-based nanotherapeutic platform is proposed that employs tetrahedral framework nucleic acids (tFNAs) conjugated with microRNA-124 (miR-124), referred to as Tmi, to achieve targeted immunomodulation. Compared with conventional delivery systems, this self-assembled nanostructure exhibits superior cellular penetration and nucleic acid protection capabilities. Mechanistic studies demonstrate that Tmi-mediated inhibition of the STAT3 pathway attenuates excessive microglial activation, reduces migration-associated inflammation, and suppresses pathological cell migration in an acute ocular hypertension model. This is an alternative treatment strategy beyond intraocular pressure control, offering greater possibilities for exploring therapies for acute high intraocular pressure glaucoma.

Pancreatic cancer is highly challenging, with most patients developing intrinsic or acquired resistance to first-line chemotherapy drug gemcitabine (GEM). Although Matrix Metalloproteinase 28 (MMP28) is upregulated in pancreatic cancer and predicts a poor prognosis, its role in GEM resistance and molecular mechanism remain unclear. Here, we aimed to investigate the role of MMP28 in GEM resistance and molecular mechanism. First, differentially expressed genes in pancreatic cancer were identified through bioinformatics and validated in clinical samples and cells. MMP28 was significantly overexpressed in pancreatic cancer tissues and Capan-1 and PANC-1 cells, correlating with poor prognosis. Then, MMP28 knockdown was performed in Capan-1 and PANC-1 cells, followed by GEM treatment. Furthermore, in vivo experiments evaluated GEM sensitivity after MMP28 knockdown. The results showed that MMP28 knockdown enhanced GEM sensitivity both in vitro , reducing cell proliferation and survival, and in vivo , where tumor growth was significantly suppressed. Additionally, glycolysis-related changes were assessed. We revealed that glycolysis was implicated as a key pathway in this process, with reduced glucose uptake and lactate production observed after MMP28 knockdown. Protein-protein interaction analysis identified Staphylococcal nuclease domain-containing protein 1 (SND1) as a key interactor, and SND1 expression was upregulated in pancreatic cancer tissues. Moreover, MMP28 interacted with SND1 to regulate SND1′s recruitment of HK2 mRNA to promote glycolysis. However, overexpression of SND1 reversed the effects of MMP28 knockdown, restoring glycolysis and GEM resistance. In conclusion, MMP28 promoted tumor growth and GEM resistance in pancreatic cancer by regulating glycolysis via interaction with SND1.

Background Ubiquitination regulates diverse cellular processes and is implicated in various pathophysiological conditions, including sepsis-induced acute lung injury (ALI). Ubiquitin-specific peptidase 48 (USP48), a deubiquitinating enzyme, has been shown to promote pyroptosis in tumor cells. However, its role in sepsis-induced ALI has not been elucidated. This study aimed to investigate the function and underlying mechanism of USP48 in sepsis-induced ALI. Methods A murine model of sepsis-induced ALI was established to assess USP48 expression in lung tissues. In parallel, USP48 levels were examined in lipopolysaccharide (LPS)-stimulated alveolar macrophages. USP48 was then knocked down both in vivo and in vitro to evaluate its functional role. Co-immunoprecipitation (Co-IP) was performed to examine the interaction between USP48 and NEK7. The effects of USP48 on NEK7 ubiquitination and stabilization were assessed in cell-based assays. To further delineate the mechanistic pathway, NEK7 was overexpressed following USP48 knockdown. Results USP48 expression was significantly upregulated in the lung tissues of septic mice and in LPS-induced macrophages. In vivo knockdown of USP48 alleviated sepsis-induced ALI and reduced macrophage pyroptosis. In vitro , USP48 enhanced LPS-induced NLRP3 inflammasome activation and macrophage pyroptosis. Co-IP analysis confirmed a direct interaction between USP48 and NEK7. Mechanistically, USP48 promoted NEK7 stabilization by facilitating its deubiquitination. Furthermore, USP48-induced pyroptosis was dependent on NEK7-mediated NLRP3 inflammasome activation. In vivo , USP48 aggravated sepsis-induced ALI through the NEK7/NLRP3/caspase-1/GSDMD signaling axis. Conclusions USP48 exacerbates sepsis-induced ALI by promoting pyroptosis of alveolar macrophages through NEK7-mediated activation of the NLRP3 inflammasome, which is facilitated by USP48-induced deubiquitination of NEK7.

Current therapeutic options remain insufficient for sepsis, driving the search for alternative treatment approaches. Accumulating evidence suggests that resistin (RETN) serves as a crucial factor in sepsis initiation and development. Nevertheless, the specific pathways through which RETN influences sepsis pathophysiology have yet to be elucidated. Single-cell sequencing analysis reveals RETN is primarily expressed in monocytes/macrophages. RETN in macrophages is markedly upregulated in septic patients, exhibiting a marked positive correlation with pro-inflammatory cytokines and disease severity. Bioinformatics analysis and in vitro experiments reveal that knockdown of RETN alleviated macrophage pyroptosis. RNA-Seq analysis and in vitro experiments revealed that the overexpression of RETN markedly upregulates the expression of GBP5 and NLRP3. Further in vivo experiments revealed that RETN knockdown markedly downregulates GBP5 expression, inhibits NLRP3 activation, and mitigates macrophage pyroptosis. This consequently reduces organ (lung, spleen, and heart) damage and improves survival in sepsis. Finally, knocking down GBP5 can reverse the promoting effect of overexpressed RETN on macrophage pyroptosis, organ damage and sepsis lethality. This investigation initially demonstrates that the RETN/GBP5/NLRP3 signaling axis regulates macrophage pyroptosis to aggravate sepsis, providing new potential targets and theoretical support for the research on the pathogenic mechanism of sepsis and clinical treatment.

Patients suffering from sepsis-induced acute lung injury (ALI) exhibit a high mortality rate, and their prognosis is closely associated with infiltration of neutrophils into the lungs. In this study, we found a significant elevation of CD64 + neutrophils, which highly expressed p75 neurotrophin receptor (p75 NTR ) in peripheral blood of mice and patients with sepsis-induced ALI. p75 NTR+ CD64 + neutrophils were also abundantly expressed in the lung of ALI mice induced by lipopolysaccharide. Conditional knock-out of the myeloid lineage's p75 NTR gene improved the survival rates, attenuated lung tissue inflammation, reduced neutrophil infiltration and enhanced the phagocytic functions of CD64 + neutrophils. In vitro, p75 NTR+ CD64 + neutrophils exhibited an upregulation and compromised phagocytic activity in blood samples of ALI patients. Blocking p75 NTR activity by soluble p75 NTR extracellular domain peptide (p75 ECD -Fc) boosted CD64 + neutrophil phagocytic activity and reduced inflammatory cytokine production via activation of the NF-κB pathway. The findings strongly indicate that p75 NTR+ CD64 + neutrophils are a novel pathogenic neutrophil subpopulation promoting sepsis-induced ALI.

The pathogenesis of preeclampsia (PE) involves endoplasmic reticulum stress (ERS) and the subsequent induction of mitophagy. Ariadne RBR E3 ubiquitin protein ligase 1 (ARIH1) is a key factor regulating mitophagy, but its role in PE has not been reported. In this study, we aimed to analyze the role of ARIH1 in the pathogenesis of PE. The role of ARIH1 in the pathogenesis of PE was investigated in a PE rat model and in an in vitro hypoxia/reoxygenation (H/R) model using HTR8 trophoblast cells. The study revealed that ARIH1 was upregulated while Mitochondrial fusion protein 2 (MFN2) was downregulated in PE rats and H/R-treated HTR8 cells. Inhibition of ARIH1 reversed the suppressed proliferation and invasion capacities of HTR8 cells under H/R conditions, reduced intracellular reactive oxygen species (ROS) and calcium ions (Ca 2+ ), and modulated the protein expression of LC3II/LC3I, p62, glucose-regulatory protein 78 (GRP78), and C/EBP homologous protein (CHOP). Additionally, mitochondrial membrane potential was improved. Interestingly, treatment with Tunicamycin or Thapsigargin could reverse the inhibitory effects of ARIH1 downregulation on trophoblastic cells by activating endoplasmic reticulum stress (ERS) and mitophagy. Notably, the study identified for the first time that ARIH1 mediates the ubiquitination degradation of MFN2. Inhibition of MFN2 abolished the regulatory effects of ARIH1 downegulation on ERS and mitophagy in trophoblast cells, as well as the associated damage in PE rats. Overall, the findings underscore the crucial role of ARIH1 in regulating mitophagy and ERS through MFN2, highlighting its significance in the pathogenesis of PE. Graphical The mechanism by which ubiquitin ligase ARIH1 promotes the occurrence of preeclampsia through MFN2. Our research found that ARIH1 is an important factor mediating the occurrence and development of PE, as well as mitochondrial autophagy and endoplasmic reticulum stress, and this effect is achieved through MFN2. Mechanistically, it was discovered for the first time that ARIH1 mediated the ubiquitination and degradation of MFN2.

Danggui Buxue Decoction (DBD) has shown potential antitumor effects in a variety of cancers. This study aims to delve into the effects and mechanisms of DBD on the metastasis of triple-negative breast cancer (TNBC). The mouse model of TNBC was constructed, and the function of DBD was investigated. The active components and core targets of DBD on TNBC were identified through liquid chromatography-mass spectrometry, network pharmacology, and molecular docking techniques. A set of in vitro experiments was conducted to examine the key components and core targets of DBD that affect the metastasis of TNBC. DBD inhibited the growth and metastasis of TNBC tumors. Formononetin is one of the key active ingredients of DBD, exerts antitumor effects by targeting Snail family transcriptional repressor 2 (SNAI2). The addition of DBD or Formononetin inhibited the malignant activity and reduced the expression of SNAI2. However, this ability of Formononetin was reversed due to the overexpression of the SNAI2 gene. Formononetin inhibits the metastasis of TNBC by downregulating SNAI2. The outcomes offer a basis for the future clinical utilization of DBD and Formononetin in TNBC therapy.

Glioblastoma stem cells (GSCs) have been implicated in the self-renewal and treatment resistance of glioblastoma (GBM). Our previous study found that 4,5-dimethoxycanthin-6-one has the potential to inhibit GBM cell proliferation. This current study aims to elucidate the molecular mechanism underlying the effects of 4,5-dimethoxycanthin-6-one in GBM development. The effect of 4,5-dimethoxycanthin-6-one on GSC formation and differentiation was explored in human GBM cell lines U251 and U87. Subsequently, 4,5-dimethoxycanthin-6-one binding to tetraspanin 1 (TSPAN1) / transmembrane 4 L six family member 1 (TM4SF1) was analyzed by molecular simulation docking. Co-immunoprecipitation (Co-IP) and immunofluorescence (IF) were used to assess the interactions between TSPAN1 and TM4SF1 in GSCs. Cell proliferation was detected by cell counting kit-8 (CCK-8) and colony formation assay. To evaluate cell migration, invasion and apoptosis, we employed wound healing assay, transwell and flow cytometry, respectively. Furthermore, subcutaneous xenograft tumor models in nude mice were constructed to evaluate the impact of 4,5-dimethoxycanthin-6-one on GSCs in vivo by examining tumor growth and histological characteristics. 4,5-Dimethoxycanthin-6-one inhibited GSC formation and promoted stem cell differentiation in a concentration-dependent manner. Molecular docking models of 4,5-dimethoxycanthin-6-one with TM4SF1 and TSPAN1 were constructed. Then, the interaction between TSPAN1 and TM4SF1 in GSC was clarified. Moreover, 4,5-dimethoxycanthin-6-one significantly inhibited the expressions of TM4SF1 and TSPAN1 in vitro and in vivo. Overexpression of TSPAN1 partially reversed the inhibitory effects of 4,5-dimethoxycanthin-6-one on GSC formation, proliferation, migration and invasion. 4,5-Dimethoxycanthin-6-one inhibited GBM progression by inhibiting TSPAN1/TM4SF1 axis. 4,5-Dimethoxycanthin-6-one might be a novel and effective drug for the treatment of GBM.

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.