Search Results (36)

Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive malignancy characterized by therapeutic resistance and poor prognosis, underscoring the need for new therapeutic strategies. Bufalin, a major bioactive constituent of Venenum bufonis, has shown antitumor activity in several cancer types; however, its mechanism of action in PDAC remains incompletely defined. In this study, we investigated the antitumor effects of bufalin in PDAC using in vitro assays, mouse tumor models, and integrative transcriptomic, proteomic, metabolomic, and bioinformatic analyses. Bufalin inhibited PDAC cell viability, clonogenic growth, migration, and tumor progression in vivo. Pharmacological rescue experiments indicated that ferroptosis contributes importantly to bufalin-induced cytotoxicity, although apoptosis- and pyroptosis-related pathways may also be involved. Multi-omics analyses revealed coordinated alterations in calcium homeostasis, endoplasmic reticulum (ER) stress/unfolded protein response (UPR) signaling, and ferroptosis-related metabolic pathways. Further experiments showed that bufalin was associated with disrupted intracellular Ca²⁺ homeostasis, IP₃R-linked ER Ca²⁺ release, activation of PERK/eIF2α/ATF4 signaling, increased ATF3 expression, reduced SLC7A11 and GPX4 expression, glutathione depletion, and enhanced lipid peroxidation. Molecular docking and surface plasmon resonance assays supported an in vitro physical interaction between bufalin and IP₃R1/IP₃R3, while inhibition of ER stress attenuated several bufalin-induced ferroptosis-related phenotypes. Bioinformatic analyses further showed that higher ER stress and ferroptosis signature scores were associated with improved overall survival in PDAC, and concurrent activation of both signatures was linked to the most favorable prognosis. Collectively, these findings support that bufalin suppresses PDAC progression through coordinated ER stress- and ferroptosis-related responses, highlighting ER stress-ferroptosis crosstalk as a potential therapeutic vulnerability in PDAC. Full article

Latrophilin 1 (LPHN1/ADGRL1), an adhesion G-protein-coupled receptor (GPCR), is the principal receptor for α-latrotoxin (αLTX), a toxin that triggers massive neurotransmitter release. However, its endogenous signaling mechanism remains elusive. Here, we dissect the LPHN1 signaling pathway at the vertebrate neuromuscular junction, using the pore-deficient αLTX mutant LTX^N4C as a selective agonist. Combining electrophysiological recordings from LPHN1 knockout mice with pharmacological inhibitors, calcium imaging, and biochemical assays, we delineate the cascade from receptor activation to spontaneous quantal acetylcholine release. We demonstrate that LPHN1 is specifically localized to the presynaptic membrane and mediates LTX^N4C-evoked release. Upon activation, LPHN1 engages the G_αq–phospholipase C pathway to generate inositol 1,4,5-trisphosphate (IP₃), triggering Ca²⁺ release from intracellular stores via IP₃ receptors. This store depletion activates store-operated Ca²⁺ entry (SOCE), providing sustained Ca²⁺ required for LTX^N4C-induced burst-like exocytosis. We uncover distinct roles for Ca_V2.1 and Ca_V1 channels in initiating and sustaining this response. These findings establish LPHN1 as a GPCR that harnesses intracellular stores and SOCE to drive spontaneous neurotransmission, revealing a novel signaling paradigm for adhesion GPCRs in presynaptic function. Full article

Background/Objectives: Primary dysmenorrhea is a common gynecological disorder characterized by painful uterine contractions. Danggui Buxue Decoction (DBD) is used to treat menstrual irregularities, but its mechanism in primary dysmenorrhea remains unclear. This study investigated the efficacy of DBD against dysmenorrhea and its calcium signaling-related mechanism. Methods: DBD components were analyzed by UPLC–Orbitrap MS. Isolated uterine muscle strips precontracted with oxytocin (OT, 50 ng/mL) or KCl (60 mM) were used to assess the effects of DBD and its active compounds (Quercetin, Formononetin, Ononin, Ferulic acid, Senkyunolide I, Calycosin, Ligustilide, Calycosin-7-O-β-D-glucoside). Ca²⁺-dependent experiments, intracellular calcium release assays, and inhibitor treatments (Nifedipine, 2-APB) were performed to evaluate the involvement of L-type calcium channels and the IP₃R pathway. A primary dysmenorrhea model induced by estradiol benzoate and oxytocin was used to assess the analgesic effects, histopathology, inflammatory factors, and IP₃/Ca²⁺-related proteins and genes following DBD and Quercetin treatment. Results: A total of 161 compounds were identified in DBD. DBD and its eight active constituents relaxed OT (50 ng/mL) or KCl (60 mM)-induced uterine contractions, with Quercetin, Calycosin, and Ligustilide showing particularly prominent relaxant activity. These three compounds suppressed extracellular calcium influx and intracellular calcium release through the blockade of L-type calcium channels and IP₃R. In vivo, DBD and Quercetin alleviated pain, reduced inflammation, and decreased uterine Ca²⁺ and IP₃ levels in dysmenorrhea mice. Conclusions: DBD and its active component Quercetin promote uterine relaxation by lowering Ca²⁺ levels, which is achieved through suppression of L-type calcium channels and the IP₃/Ca²⁺ pathway. This contributes to their therapeutic action against primary dysmenorrhea. Full article

Background/Objectives: Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive cancer with a five-year survival rate of approximately 13%, partly because of limited treatment options and resistance to therapies. Although the recently discovered KRAS G12D inhibitor MRTX1133 has shown promising efficacy in preclinical models, its clinical efficacy as a single agent is expected to be limited, as is the case with KRAS G12C inhibitors. Therefore, in this study, we evaluated potential combination strategies to enhance the therapeutic effect of MRTX1133. We combined MRTX1133 with the BCL-xL proteolysis-targeting chimera (PROTAC) DT2216 and the mTOR inhibitor everolimus. Methods: The sensitization of MRTX1133 by the combination of DT2216 + everolimus was tested in KRAS G12D-mutant PDAC cell lines using colony formation and apoptosis assays. The effects of MRTX1133 and/or DT2216 + everolimus on KRAS signaling and BCL-2 family proteins were assessed by immunoblotting and/or RT-PCR. The functional roles of BIM/NOXA were elucidated via immunoprecipitation (IP) and siRNA knockdown. Triple combination efficacy was evaluated in AsPC1 parental and MRTX1133-resistant xenografts, with pharmacodynamic effects confirmed by immunoblotting and immunohistochemistry. Results: The triple combination leads to significantly greater colony growth inhibition and apoptosis induction as compared with single agents or two-drug combinations in multiple KRAS G12D-mutant PDAC cell lines. Mechanistically, MRTX1133 treatment increased BIM and decreased NOXA levels, and the combination of DT2216/everolimus simultaneously enhanced BIM release and stabilized NOXA. In vivo, DT2216/everolimus combination significantly potentiated the anti-tumor activity of MRTX1133 in the AsPC1 PDAC xenograft model. Furthermore, the triple combination effectively overcame acquired MRTX1133 resistance in vitro and in the AsPC1 xenograft model. Conclusions: Collectively, our findings suggest that the combination of DT2216/everolimus potentiates the anti-tumor efficacy of MRTX1133 associated with enhanced apoptosis induction and inhibition of compensatory survival signaling. Full article

Latent tuberculosis infection (LTBI) represents a major global health concern as it constitutes the principal reservoir for future tuberculosis (TB) disease. Its identification is particularly important in Bacille Calmette–Guérin (BCG)-vaccinated populations, where cross-reactivity of purified protein derivative limits the specificity of the tuberculin skin test and hampers targeted preventive therapy. Early Mycobacterium tuberculosis antigens encoded within the RD1 region, especially ESAT-6, CFP-10 and TB7.7, have enabled the development of antigen-specific interferon-gamma release assays (IGRAs) and recombinant skin tests with improved BCG-independent specificity. This narrative review integrates and critically appraises current evidence on the immunobiological properties of early and latency-associated antigens, the cellular mechanisms underlying T-cell-dependent immune reactivity, and the diagnostic performance of IGRAs and ESAT-6/CFP-10-based skin tests, rather than merely summarizing individual studies. Although these platforms rely on different assay principles (in vitro cytokine release versus in vivo delayed-type hypersensitivity), both measure antigen-specific T-cell memory and do not define the biological stage of infection or reliably distinguish latent from incipient or active TB. Across most adult populations, IGRAs demonstrate high specificity and acceptable sensitivity, whereas reduced sensitivity and higher rates of indeterminate results are observed in young children and immunocompromised individuals. ESAT-6/CFP-10-based skin tests show diagnostic accuracy comparable to IGRAs and may offer operational advantages in resource-limited settings. Latency-associated antigens and host biomarkers such as IP-10, together with multi-analyte immune signatures, represent promising avenues for improving diagnostic sensitivity and prognostic stratification but currently lack sufficient validation for routine clinical use. Overall, RD1-encoded antigens remain central to LTBI immunodiagnosis, while future research should focus on developing stage-resolving and prognostic biomarkers, optimized antigen panels, and standardized interpretive frameworks. Full article

Objectives: This study aimed to evaluate the cytotoxic effects of Ti6Al4V particles and implantoplasty (IP)-treated surfaces on human fibroblasts and osteoblasts, and to investigate the influence of these particles on multispecies oral biofilm formation. Methods: Ti6Al4V particles generated during implantoplasty were collected. Human fibroblasts (HFF-1) and osteoblast-like cells (SaOs-2) were used to assess cytotoxicity through indirect lactate dehydrogenase (LDH) assays. Multispecies biofilms composed of Streptococcus oralis, Actinomyces viscosus, Veillonella parvula and Porphyromonas gingivalis were evaluated based on colony-forming units (CFUs) and metabolic activity. Fibroblasts and osteoblasts were co-cultured with biofilm-contaminated particles for 2, 4 and 6 h. Cell morphology and biofilm association were examined by phase-contrast microscopy, while metabolic activity was measured spectrophotometrically. Results: IP-treated surfaces showed no significant cytotoxicity (metabolic activity > 92%, LDH < 20%). Ti6Al4V particles selectively promoted A. viscosus and V. parvula growth (metabolic activity increases of ≈192% and ≈203%; CFU significantly higher versus controls, p < 0.05). Co-culture with biofilm-contaminated particles drastically reduced cell activity (fibroblasts < 25%, osteoblasts < 10%), whereas bacteria-free particles did not. Conclusions: Biofilm-contaminated particles released during implantoplasty markedly impair fibroblast and osteoblast cytocompatibility and selectively alter bacterial growth, whereas IP-treated surfaces per se are biocompatible. Minimizing particle dissemination and bacterial contamination during IP is therefore crucial. Full article

Background. Peru ranks among the countries with the highest burden of tuberculosis in Latin America. Interferon Gamma Release Assays (IGRAs), such as QuantiFERON-TB (QFT), cannot distinguish latent tuberculosis infection (LTBI) from active tuberculosis (ATB), but they provide a more specific and quantitative assessment of prior exposure to Mycobacterium tuberculosis. Methods. We enrolled 56 participants and measured 38 cytokines/chemokines from plasma obtained through QFT in patients with active TB (n = 11), LTBI (n = 17), and QFT-negative healthy controls (n = 28) using a Multiplex Bead Assay. Results. Levels of IL-1RA, IL-2, IL-5, IFN-γ, and IP-10 significantly differentiated both ATB and LTBI groups from healthy controls (p < 0.035). Furthermore, IL-5 and IP-10 specifically distinguished LTBI from healthy controls (p < 0.015), while GM-CSF differentiated ATB from controls (p < 0.05). Conclusions. Beyond IFN-γ as a differential proinflammatory cytokine in TB, cytokines such as IL-5, and IP-10 show potential as biomarkers to discriminate infection status in high-burden TB settings. Full article

This study aimed to evaluate cytokine profiling in a periapical lesion to provide a rationale for future treatment strategies for periapical lesions. Thirteen samples of exudative fluid were collected from such a lesion directly through the root canal. Cytokine profiling was performed using the Bio-Plex system. CXCL10 (C-X-C motif chemokine ligand 10, IP10) was found to be elevated in apical exudates of patients exhibiting favorable healing. To evaluate the role of CXCL10 in cell migration, a Transwell assay was conducted using bone marrow-derived mononuclear cells (BMMCs). Different types of cytokines were detected from the samples of periapical lesion at the initial visit. However, cytokine production varied across patient samples. Release of interleukin (IL)-1β, IL-2, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12, IL-13, IL-17, granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage colony-stimulating factor (GM-CSF), interferon gamma (IFN-γ), monocyte chemoattractant protein (MCP)-1, macrophage inflammatory protein (MIP)-1α, MIP-1β, and tumor necrosis factor (TNF)-α showed differential expression. Comparison of cytokine profiles indicated that cytokine production was variable before and after root canal treatment. In vitro, CXCL10 significantly improved BMMC migration in a dose-dependent manner, supporting clinical findings that elevated CXCL10 levels are associated with favorable healing in apical lesions. Although this study was limited by the small sample size and exploratory design, the cytokine profile of periapical lesions may be useful for assessing the condition of periapical lesions and modulating the immune response to bacterial infection. Full article

Breast cancer (BC) is a major cause of cancer-related mortality. Mortalin and Vimentin—two proteins implicated in BC progression and metastasis—have been identified as binding partners of the Secretion Modification Region (SMR) peptide from the HIV Nef protein. These interactions disrupt exosome release and offer novel therapeutic strategies. This study investigates the binding interactions between the SMR peptide, Mortalin, and Vimentin using surface plasmon resonance (SPR), co-immunoprecipitation (Co-IP), and Western blot assays. We also map the SMR binding sites on Mortalin through scanning peptide mapping and then identify a similar site on the Vimentin protein. Based on these data, we propose that the SMR peptide and its analogs interact with specific amino acid sequences in Mortalin and Vimentin, thereby disrupting cellular processes essential for Epithelial–Mesenchymal Transition (EMT) and tumor progression. SPR analysis revealed that the Nef protein exhibited the highest binding affinity to Vimentin (KD = 0.75 ± 1.1 nM) and Mortalin (KD = 3.16 ± 0.03 nM). The SMRwt peptide also demonstrated direct binding to both proteins with micromolar affinities (KD = 6.63 ± 0.74 µM for Vimentin; KD = 20.73 ± 2.33 µM for Mortalin), though the binding affinity was weaker than the full Nef protein. Co-IP experiments using MDA-MB-231, MCF-7, and BT474 BC cell lines confirmed that SMRwt, but not SMRmut, co-immunoprecipitated with Mortalin. Western blot analysis validated these interactions. Further, Mortalin peptide #56, derived from the substrate-binding domain, did not bind the SMR domain or inhibit Nef function. In contrast, peptides #61 and #62 from the C-terminal domain of Mortalin bound the SMR domain and effectively inhibited Nef activity. Notably, Mortalin peptide #61 inhibited SMRwt binding to both Mortalin and Vimentin, disrupting complex formation on the SPR sensor chip. These findings suggest that specific Mortalin-derived peptides can block SMR interactions, offering a potential therapeutic mechanism. Full article

Background: DEAD/H box 5 (DDX5) serves as a transcriptional coactivator for several transcription factors including E2F1, the primary target of the tumor suppressor pRB. E2F1 physiologically activated by growth stimulation activates growth-related genes and promotes cell proliferation. In contrast, upon loss of pRB function due to oncogenic changes, E2F1 is activated out of restraint by pRB (deregulated E2F1) and stimulates tumor suppressor genes such as ARF, which activates the tumor suppressor p53, to suppress tumorigenesis. We have recently reported that DDX5 augments deregulated E2F1 activity to induce tumor suppressor gene expression and apoptosis. During the analyses, we noted that over-expression of E2F1 increased DDX5 expression, suggesting a feed forward loop in E2F1 activation through DDX5. Objective: We thus examined whether the DDX5 gene is a target of deregulated E2F1. Method: For this purpose, we performed promoter analysis and ChIP assay. Result: The DDX5 promoter did not possess typical E2F binding consensus but contained several GC repeats observed in deregulated E2F1 targets. Insertion of point mutations in these GC repeats decreased responsiveness to deregulated E2F1 induced by over-expression of E2F1, but scarcely affected responsiveness to growth stimulation. ChIP assays showed that deregulated E2F1 induced by over-expression of E2F1 or expression of E1a, which binds pRB and releases E2F1, bound to the DDX5 gene, while physiological E2F1 induced by growth stimulation did not. Conclusions: These results suggest that the DDX5 gene is a target of deregulated E2F1, generating a feed forward loop mediating tumor suppressive E2F1 activity. Full article

Objectives: Ischemic stroke is a severe neurological disorder with high morbidity, mortality, and disability rates, posing a substantial burden on patients, families, and healthcare systems. Soy isoflavone (SI), a naturally occurring phytoestrogen, has demonstrated promising neuroprotective effects. This study aimed to evaluate the anti-stroke efficacy of SI and elucidate its underlying mechanisms through integrated phytochemical profiling, network pharmacology, and both in vitro and in vivo experimental validation. Methods: Active constituents of SI were extracted via reflux and identified using liquid chromatography–mass spectrometry (LC-MS). Network pharmacology was employed to predict therapeutic targets and signaling pathways. The neuroprotective effects of SI were first assessed in PC12 cells subjected to oxygen–glucose deprivation/reoxygenation (OGD/R) injury in vitro. For in vivo evaluation, transient cerebral ischemia–reperfusion injury was induced using the bilateral common carotid artery occlusion (BCCAO) model in adult male ICR rats (27.3 ± 1.8 g; 6–8 weeks old), obtained from the Shanghai Experimental Animal Center, Chinese Academy of Sciences. Forty-eight rats were randomly assigned into four groups (n = 12): sham, model (BCCAO), SI-treated (100 mg/kg, oral gavage for 5 days), and edaravone (EDA)-treated (10 mg/kg, i.p., positive control). All procedures were approved by the Institutional Animal Care and Use Committee of Changchun Normal University (Approval No. 2024003, 13 March 2024) and conducted in accordance with the NIH guidelines and ARRIVE 2.0 reporting standards. Results: In vitro, SI significantly enhanced PC12 cell viability from 57.23 ± 2.88% to 80.76 ± 4.43% following OGD/R. It also reduced intracellular Ca²⁺ by 58.42%, lactate dehydrogenase (LDH) release by 37.67%, caspase-3 activity by 55.05%, and reactive oxygen species (ROS) levels by 74.13% (p < 0.05). A flow cytometry analysis revealed that OGD/R increased the apoptosis rate from 5.34% (control) to 30.85% (model group), which was significantly attenuated by SI treatment, especially in the 560 µg/mL group (20.00%), followed by the 140 and 280 µg/mL groups. In vivo, SI improved neurological scores from 8.3 ± 1.09 to 6.8 ± 1.68, reduced cerebral infarction volume by 18.49%, and alleviated brain edema by 10.42% (p < 0.05). SI also decreased malondialdehyde (MDA) and LDH levels by 31.15% and 39.46%, respectively, while increasing the activity of antioxidant enzymes: superoxide dismutase (SOD) by 11.70%, catalase (CAT) by 26.09%, and glutathione peroxidase (GSH-px) by 27.55% (p < 0.01). Scratch assay results showed that SI restored the impaired migratory ability of the OGD/R-treated PC12 cells, further supporting its role in cellular repair. A Western blot analysis demonstrated the upregulation of nuclear factor erythroid 2–related factor 2 (Nrf2), heme oxygenase-1 (HO-1), and NAD(P)H:quinone oxidoreductase 1 (NQO1) and the downregulation of Kelch-like, ECH-associated protein 1 (Keap1) in the cerebral ischemia–reperfusion model. Conclusions: These findings indicate that soy isoflavone confers significant neuroprotective effects against cerebral ischemia–reperfusion injury by enhancing endogenous antioxidant defense mechanisms, reducing oxidative stress, inhibiting apoptosis, and promoting cell migration. The protective effects are likely mediated through the activation of the Nrf2/Keap1 signaling pathway, supporting the therapeutic potential of SI in ischemic stroke treatment. Full article

Mitochondria-ER contact sites (MERCS) are vital for mitochondrial dynamics, lipid exchange, Ca²⁺ homeostasis, and energy metabolism. We examined whether mitochondrial metabolism changes during the cell cycle depend on MERCS dynamics and are regulated by the outer mitochondrial protein mitochondrial rho GTPase 1 (MIRO1). Wound healing was assessed in mice with fibroblast-specific deletion of MIRO1. Wild-type and MIRO1^-/- fibroblasts and vascular smooth muscle cells were evaluated for proliferation, cell cycle progression, number of MERCS, distance, and protein composition throughout the cell cycle. Restoration of MIRO1 mutants was used to test the role of MIRO1 domains; Ca²⁺ transients and mitochondrial metabolism were evaluated using biochemical, immunodetection, and fluorescence techniques. MERCS increased in number during G1/S compared with during G0, which was accompanied by a notable rise in protein–protein interactions involving VDAC1 and IP3R as well as GRP75 and MIRO1 by proximity-ligation assays. Split-GFP ER/mitochondrial contacts of 40 nm also increased. Mitochondrial Ca²⁺ concentration ([Ca²⁺]), membrane potential, and ATP levels correlated with the formation of MERCS during the cell cycle. MIRO1 deficiency blocked G1/S progression and the cell-cycle-dependent formation of MERCS and altered ER Ca²⁺ release and mitochondrial Ca²⁺ uptake. MIRO1 mutants lacking the Ca²⁺-sensitive EF hands or the transmembrane domain did not rescue cell proliferation or the formation of MERCS. MIRO1 controls an increase in the number of MERCS during cell cycle progression and increases mitochondrial [Ca²⁺], driving metabolic activity and proliferation through its EF hands. Full article

Vitamin D (VD), a crucial micronutrient, regulates bone health and immune responses. Recent studies suggest that VD may confer protective effects against chronic inflammatory diseases. Additionally, plant-based peptides can show biological activities. Furthermore, the supplementation of protein hydrolysates with VD could potentially enhance the bioactivity of peptides, leading to synergistic effects. In this study, THP-1 cells were exposed to low concentrations of Lipopolysaccharide (LPS) to induce inflammation, followed by treatment with vitamin D at different concentrations (10, 25, or 50 nM) or a chickpea protein hydrolysate (“H30BIO”) supplemented with VD. The cytotoxicity of VD was evaluated using viability assay to confirm its safety. The cytokine secretion of TNF-α, IL-1β, and IL6 was assessed in the cell supernatant, and the gene expression of TNF-α, IL-1β, IL6, IL8, CASP-1, COX2, NRF2, NF-ĸB, NLRP3, CCL2, CCR2, IP10, IL10, and RANTES was quantified by qRT-PCR. Treatment with VD alone significantly decreased the expression of the pro-inflammatory genes TNF-α and IL6, as well as their corresponding cytokine levels in the supernatants. While IL-1β gene expression remained unchanged, a reduction in its cytokine release was observed upon VD treatment. No dose-dependent effects were observed. Interestingly, the combination of VD with H30BIO led to an increase in TNF-α expression and secretion in contrast with the LPS control, coupled with a decrease in IL-1β levels. Additionally, genes such as IP10, NF-κB, CCL2, COX2, NRF2, and CASP-1 exhibited notable modulation, suggesting that the combination treatment primarily downregulates NF-κB-related gene activity. This study demonstrates a synergistic interaction between VD and H30BIO, suggesting that this combination may enhance pathways involving TNF-α, potentially aiding in the resolution and modulation of inflammation through adaptive processes. These findings open new avenues for research into the therapeutic applications of enriched protein hydrolysates with VD to manage low-grade inflammatory-related conditions. Full article

The unprecedented research effort associated with the emergence of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) included several extensive proteomic studies that identified host proteins that interact with individual viral gene products. However, in most cases, the consequences of those virus–host interactions for virus replication were not experimentally pursued, which is a necessary step in determining whether the interactions represent pro- or anti-viral events. One putative interaction commonly identified in multiple studies was between the host adaptor protein complex 3 (AP-3) subunit B1 (AP3B1) and the SARS-CoV-2 envelope protein (E). AP3B1 is one subunit of AP-3 required for the biogenesis of lysosomal-related organelles (LROs), and its function impacts important disease processes including inflammation and vascular health. Thus, interactions between AP3B1 and SARS-CoV-2 might influence the clinical outcomes of infection. To determine if AP3B1 has a role in the SARS-CoV-2 replication cycle, we first confirmed the interaction in virus-infected cells using immunoprecipitation (IP) and immunofluorescence assays (IFA). AP3B1 is required by multiple viruses to aid in the replication cycle and therefore may be a therapeutic target. However, we found that the overexpression of AP3B1 suppressed SARS-CoV-2 replication, whereas the siRNA-mediated depletion of AP3B1 increased the release of infectious virus, suggesting an antiviral role for AP3B1. Together, our findings suggest that AP3B1 is an intrinsic barrier to SARS-CoV-2 replication through interactions with the viral E protein. Our work justifies further investigations of LRO trafficking in SARS-CoV-2 target cells and their role in viral pathogenesis. Full article

The development of targeted drug delivery systems has been a pivotal area in nanomedicine, addressing challenges like low drug loading capacity, uncontrolled release, and systemic toxicity. This study aims to develop and evaluate dual-functionalized mesoporous silica nanoparticles (MSN) for targeted delivery of celecoxib, enhancing drug loading, achieving controlled release, and reducing systemic toxicity through amine grafting and imidazolyl polyethyleneimine (PEI) gatekeepers. MSN were synthesized using the sol–gel method and functionalized with (3-aminopropyl) triethoxysilane (APTES) to create amine-grafted MSN (MSN-NH₂). Celecoxib was loaded into MSN-NH₂, followed by conjugation of imidazole-functionalized PEI (IP) gatekeepers synthesized via carbodiimide coupling. Characterization was conducted using Fourier-transform infrared spectroscopy (FTIR) and proton nuclear magnetic resonance (¹H-NMR). Drug loading capacity, entrapment efficiency, and in vitro drug release at pH 5.5 and 7.4 were evaluated. Cytotoxicity was assessed using the MTT assay on RAW 264.7 macrophages. The synthesized IP was confirmed by FTIR and ¹H-NMR. Amine-grafted MSN demonstrated a celecoxib loading capacity of 12.91 ± 2.02%, 2.1 times higher than non-functionalized MSN. In vitro release studies showed pH-responsive behavior with significantly higher celecoxib release from MSN-NH₂-celecoxib-IP at pH 5.5 compared to pH 7.4, achieving a 33% increase in release rate within 2 h. Cytotoxicity tests indicated significantly higher cell viability for IP-treated cells compared to PEI-treated cells, confirming reduced toxicity. The dual-functionalization of MSN with amine grafting and imidazolyl PEI gatekeepers enhances celecoxib loading and provides controlled pH-responsive drug release while reducing systemic toxicity. These findings highlight the potential of this advanced drug delivery system for targeted anti-inflammatory and anticancer therapies. Full article