Reprogramming the Tumor Immune Microenvironment Through Activatable Photothermal Therapy and GSH depletion Using Liposomal Gold Nanocages to Potentiate Anti-Metastatic Immunotherapy.

Cancer immunotherapy offers significant clinical benefits for patients with advanced or metastatic tumors. However, immunotherapeutic efficacy is often hindered by the tumor microenvironment's high redox levels, leading to variable patient outcomes. Herein, a therapeutic liposomal gold nanocage (MGL) is innovatively developed based on photo-triggered hyperthermia and a releasable strategy by combining a glutathione (GSH) depletion to remodel the tumor immune microenvironment, fostering a more robust anti-tumor immune response. MGL comprises a thermosensitive liposome shell and a gold nanocage core loaded with maleimide. The flexible shell promotes efficient uptake by cancer cells, enabling targeted destruction through photothermal therapy while triggering immunogenic cell death and the maturation of antigen-presenting cells. The photoactivated release of maleimide depletes intracellular GSH, increasing tumor cell sensitivity to oxidative stress and thermal damage. Conversely, GSH reduction also diminishes immunosuppressive cell activity, enhances antigen presentation, and activates T cells. Moreover, photothermal immunotherapy decreases elevated levels of heat shock proteins in tumor cells, further increasing their sensitivity to hyperthermia. In summary, MGL elicited a robust systemic antitumor immune response through GSH depletion, facilitating an effective photothermal immunotherapeutic strategy that reprograms the tumor microenvironment and significantly inhibits primary and metastatic tumors. This approach demonstrates considerable translational potential and clinical applicability.

DENV NS1 and MMP-9 cooperate to induce vascular leakage by altering endothelial cell adhesion and tight junction.

Dengue virus (DENV) is a mosquito-borne pathogen that causes a spectrum of diseases including life-threatening dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS). Vascular leakage is a common clinical crisis in DHF/DSS patients and highly associated with increased endothelial permeability. The presence of vascular leakage causes hypotension, circulatory failure, and disseminated intravascular coagulation as the disease progresses of DHF/DSS patients, which can lead to the death of patients. However, the mechanisms by which DENV infection caused the vascular leakage are not fully understood. This study reveals a distinct mechanism by which DENV induces endothelial permeability and vascular leakage in human endothelial cells and mice tissues. We initially show that DENV2 promotes the matrix metalloproteinase-9 (MMP-9) expression and secretion in DHF patients' sera, peripheral blood mononuclear cells (PBMCs), and macrophages. This study further reveals that DENV non-structural protein 1 (NS1) induces MMP-9 expression through activating the nuclear factor κB (NF-κB) signaling pathway. Additionally, NS1 facilitates the MMP-9 enzymatic activity, which alters the adhesion and tight junction and vascular leakage in human endothelial cells and mouse tissues. Moreover, NS1 recruits MMP-9 to interact with ß-catenin and Zona occludens protein-1/2 (ZO-1 and ZO-2) and to degrade the important adhesion and tight junction proteins, thereby inducing endothelial hyperpermeability and vascular leakage in human endothelial cells and mouse tissues. Thus, we reveal that DENV NS1 and MMP-9 cooperatively induce vascular leakage by impairing endothelial cell adhesion and tight junction, and suggest that MMP-9 may serve as a potential target for the treatment of hypovolemia in DSS/DHF patients.

Tumor Necrosis Factor-α Promotes the Tumorigenesis, Lymphangiogenesis, and Lymphatic Metastasis in Cervical Cancer via Activating VEGFC-Mediated AKT and ERK Pathways.

Background: Lymphatic metastasis is a common phenomenon of cervical cancer. Tumor necrosis factor-α (TNF-α) was found to be closely associated with lymphatic cancer metastasis. However, the mechanism through which TNF-α regulates lymphatic metastasis in cervical cancer remains unclear. Methods: In this study, cervical cancer cells were cultured in Dulbecco's modified Eagle's medium (DMEM) with or without TNF-α for 48 h, and then the corresponding conditional medium (CM-TNF-α or CM) was collected. The level of vascular endothelial growth factor (VEGFC) in the corresponding CM was then detected using an enzyme-linked immunosorbent assay (ELISA). Next, human lymphatic endothelial cells (HLECs) were cultured in CM-TNF-α or CM for 48 h. Cell viability was measured using the cell counting kit-8 (CCK-8) assay, and angiogenesis was detected using a tube formation assay. Subsequently, the expressions of AKT, p-AKT, ERK, and p-ERK in HLECs were detected using western blotting. In addition, to further investigate the effect of TNF-α on the progression of cervical cancer, a C33A subcutaneous xenograft model was established in vivo. Results: We found that TNF-α significantly stimulated cervical cancer cells to secrete VEGFC. Additionally, the CM collected from the TNF-α-treated cervical cancer cells notably promoted the proliferation, migration, and angiogenesis of HLECs; however, these changes were reversed by MAZ51, a VEGFR3 inhibitor. Moreover, TNF-α obviously elevated D2-40 and VEGFC protein expressions in tumor tissues, promoting lymphangiogenesis and lymphatic metastasis in vivo. Meanwhile, TNF-α markedly upregulated p-AKT and p-ERK expressions in tumor tissues, whereas these changes were reversed by MAZ51. Conclusion: Collectively, TNF-α could promote tumorigenesis, lymphangiogenesis, and lymphatic metastasis in vitro and in vivo in cervical cancer via activating VEGFC-mediated AKT and ERK pathways. These results may provide new directions for the treatment of cervical cancer.

Prognostic values of regulatory T cells (Tregs) and Treg-related genes in gastric cancer.

Introduction: This study attempted to investigate the potential of a risk model constructed for regulatory T cells (Tregs) and their related genes in predicting gastric cancer (GC) prognosis. Material and methods: We used flow cytometry to detect the content of CD4+CD25+ Tregs. After detecting expression of five Treg-related genes by quantitative real-time polymerase chain reaction (qRT-PCR), Pearson analysis was employed to analyze the correlation between Tregs and related gene expression. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), colony formation and transwell assays were used to detect the effects of a disintegrin and metalloproteinase with thrombospondin motifs 12 (ADAMTS12) on cell functions. A prognostic risk model was built after Cox regression analysis. The Kaplan-Meier method was employed to assess how Tregs, 5-gene risk scores and expression of 5 genes were correlated with the survival time. Results: A significantly increased content of Tregs was found in GC tissues (p < 0.05). 5 Treg- related genes were significantly up-regulated in GC with a positive correlation with the content of Tregs (p < 0.05). Overexpression of ADAMTS12 significantly enhanced the viability, proliferation, migration and invasion of tumor cells. Kaplan-Meier analysis demonstrated poor overall survival and disease-free survival in the high-risk group. The results of survival analysis of Treg content and related gene expression were consistent with those of Cox analysis. Conclusions: The risk model constructed based on five Treg-related genes can enable effective prediction in the prognosis of GC patients.

Interferon alpha treatment leads to a high rate of hepatitis B surface antigen seroconversion in Chinese children with chronic hepatitis B.

Chronic hepatitis B virus (HBV) infection (CHB) in children remains a public health challenge despite significant success in programme is established to prevent mother-to-child transmission. In particular, CHB in Chinese children are mostly acquired through vertical transmission, which differs from the common infection route reported in other countries and regions. This situation has resulted in a high endemic prevalence of CHB in Chinese adults. Thus, successful treatment of children with CHB will prevent the development of advanced liver diseases in late adulthood. However, there is still no consensus on the clinical guideline to treat paediatric CHB. In this study, we evaluated the potential of interferon alpha (IFNa) treatment for Chinese children with CHB. A total of 41 patients with CHB aged 3-17 years were enrolled in this retrospective study: 21 patients were treated with pegylated (PEG)-IFNa and 20 patients without treatment served as the control group. The rates of HBV DNA suppression, hepatitis B e antigen (HBeAg) clearance and hepatitis B surface antigen (HBsAg) clearance were significantly higher in the PEG-IFNa treatment group than in the control group (P < 0.05 at 48 weeks). Unexpectedly, PEG-IFNa treatment achieved a high rate of HBsAb production, far exceeding the clinical outcome in documented PEG-IFNa-treated CHB adults. Further analysis revealed that younger children (3-6 years old) were more responsive to PEG-IFNa treatment with respect to achieving a protective level of HBsAb in a short treatment cycle than adolescents (10-17 years old). Overall, these results indicate that the immune system of children might have a preserved PEG-IFNa-mediated mechanism to completely control HBV, which can help to design new strategies to treat CHB patients.

Preclinical efficacy of a RAF inhibitor that evades paradoxical MAPK pathway activation in protein kinase BRAF-mutant lung cancer.

Oncogenic activation of protein kinase BRAF drives tumor growth by promoting mitogen-activated protein kinase (MAPK) pathway signaling. Because oncogenic mutations in BRAF occur in ∼2-7% of lung adenocarcinoma (LA), BRAF-mutant LA is the most frequent cause of BRAF-mutant cancer mortality worldwide. Whereas most tumor types harbor predominantly the BRAFV600E-mutant allele, the spectrum of BRAF mutations in LA includes BRAFV600E (∼60% of cases) and non-V600E mutant alleles (∼40% of cases) such as BRAFG469A and BRAFG466V The presence of BRAFV600E in LA has prompted clinical trials testing selective BRAF inhibitors such as vemurafenib in BRAFV600E-mutant patients. Despite promising clinical efficacy, both innate and acquired resistance often result from reactivation of MAPK pathway signaling, thus limiting durable responses to the current BRAF inhibitors. Further, the optimal therapeutic strategy to block non-V600E BRAF-mutant LA remains unclear. Here, we report the efficacy of the Raf proto-oncogene serine/threonine protein kinase (RAF) inhibitor, PLX8394, that evades MAPK pathway reactivation in BRAF-mutant LA models. We show that PLX8394 treatment is effective in both BRAFV600E and certain non-V600 LA models, in vitro and in vivo. PLX8394 was effective against treatment-naive BRAF-mutant LAs and those with acquired vemurafenib resistance caused by an alternatively spliced, truncated BRAFV600E that promotes vemurafenib-insensitive MAPK pathway signaling. We further show that acquired PLX8394 resistance occurs via EGFR-mediated RAS-mTOR signaling and is prevented by upfront combination therapy with PLX8394 and either an EGFR or mTOR inhibitor. Our study provides a biological rationale and potential polytherapy strategy to aid the deployment of PLX8394 in lung cancer patients.

Mapping the molecular determinants of BRAF oncogene dependence in human lung cancer.

Oncogenic mutations in the BRAF kinase occur in 6-8% of nonsmall cell lung cancers (NSCLCs), accounting for more than 90,000 deaths annually worldwide. The biological and clinical relevance of these BRAF mutations in NSCLC is incompletely understood. Here we demonstrate that human NSCLC cells with BRAF(V600E), but not other BRAF mutations, initially are sensitive to BRAF-inhibitor treatment. However, these BRAF(V600E) NSCLC cells rapidly acquire resistance to BRAF inhibition through at least one of two discrete molecular mechanisms: (i) loss of full-length BRAF(V600E) coupled with expression of an aberrant form of BRAF(V600E) that retains RAF pathway dependence or (ii) constitutive autocrine EGF receptor (EGFR) signaling driven by c-Jun-mediated EGFR ligand expression. BRAF(V600E) cells with EGFR-driven resistance are characterized by hyperphosphorylated protein kinase AKT, a biomarker we validated in BRAF inhibitor-resistant NSCLC clinical specimens. These data reveal the multifaceted molecular mechanisms by which NSCLCs establish and regulate BRAF oncogene dependence, provide insights into BRAF-EGFR signaling crosstalk, and uncover mechanism-based strategies to optimize clinical responses to BRAF oncogene inhibition.

Identification of potential biomarkers for diabetic cardiomyopathy using LC-MS-based metabolomics.

Diabetic cardiomyopathy (DCM) is a serious complication of type 2 diabetes mellitus (T2DM) that contributes to cardiovascular morbidity and mortality. However, the metabolic alterations and specific biomarkers associated with DCM in T2DM remain unclear. In this study, we conducted a comprehensive metabolomic analysis using liquid chromatography-mass spectrometry (LC-MS) to investigate the plasma metabolite profiles of T2DM patients with and without DCM. We identified significant differences in metabolite levels between the groups, highlighting the dysregulation of various metabolic pathways, including starch and sucrose metabolism, steroid hormone biosynthesis, tryptophan metabolism, purine metabolism, and pyrimidine metabolism. Although several metabolites showed altered abundance in DCM, they also shared characteristics of DCM and T2DM rather than specific to DCM. Additionally, through biomarker analyses, we identified potential biomarkers for DCM, such as cytidine triphosphate, 11-ketoetiocholanolone, saccharopine, nervonic acid, and erucic acid. These biomarkers demonstrated distinct patterns and associations with metabolic pathways related to DCM. Our findings provide insights into the metabolic changes associated with DCM in T2DM patients and highlight potential biomarkers for further validation and clinical application. Further research is needed to elucidate the underlying mechanisms and validate the diagnostic and prognostic value of these biomarkers in larger cohorts.

Role of aryl hydrocarbon receptors in infection and inflammation.

The aryl hydrocarbon receptor (AhR) is a transcription factor that is activated by various ligands, including pollutants, microorganisms, and metabolic substances. It is expressed extensively in pulmonary and intestinal epithelial cells, where it contributes to barrier defense. The expression of AhR is pivotal in regulating the inflammatory response to microorganisms. However, dysregulated AhR expression can result in endocrine disorders, leading to immunotoxicity and potentially promoting the development of carcinoma. This review focuses on the crucial role of the AhR in facilitating and limiting the proliferation of pathogens, specifically in relation to the host cell type and the species of etiological agents involved in microbial pathogen infections. The activation of AhR is enhanced through the IDO1-AhR-IDO1 positive feedback loop, which is manipulated by viruses. AhR primarily promotes the infection of SARS-CoV-2 by inducing the expression of angiotensin-converting enzyme 2 (ACE2) and the secretion of pro-inflammatory cytokines. AhR also plays a significant role in regulating various types of T-cells, including CD4+ T cells and CD8+ T cells, in the context of pulmonary infections. The AhR pathway plays a crucial role in regulating immune responses within the respiratory and intestinal barriers when they are invaded by viruses, bacteria, parasites, and fungi. Additionally, we propose that targeting the agonist and antagonist of AhR signaling pathways could serve as a promising therapeutic approach for combating pathogen infections, especially in light of the growing prevalence of drug resistance to multiple antibiotics.

Mechanical strength affecting the penetration in microneedles and PLGA nanoparticle-assisted drug delivery: Importance of preparation and formulation.

Microneedles (MNs) prepared from polymeric materials are painless and minimally invasive, safe and efficient, but they hindered by low mechanical strength and single diverse drug release pattern. Due to the distinctive mechanical strength and dimensions of poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs), the integration of nano-technology with microneedles can effectively improve penetration and delivery efficiency through the stratum corneum. We herein designed a simple paroxetine (PAX)-loaded PLGA nanoparticles-integrated dissolving microneedles system (PAX-NPs-DMNs), aiming to improve the bioavailability of PAX through the synergistic permeation-enhancing effect of dissolving microneedles (DMNs) and NPs. PAX-NPs-DMNs had a complete tips molding rate (Neff) of (94.06 ± 2.16) %, a 15×15 quadrangular-conical microneedle array and an overall fracture force of 301.10 N, which were improved nearly 0.50 times compared with the blank microneedles (HA-DMNs) and PAX microneedles (PAX-DMNs). PAX-NPs-DMNs could extend the release duration of PAX from 1 h to 24 h and the cumulative permeability per unit area (Qn) was 47.66 times and 7.37 times higher than the PAX and the PAX-DMNs groups. PAX-NPs-DMNs could be rapidly dissolved within 10 min without hindering skin healing or causing adverse reactions. This study confirmed that PAX-NPs-DMNs can effectively improve the bioavailability of PAX and the mechanical strength of DMNs, which can easily penetrate the skin to provide sustained and painless delivery without causing adverse effects, thus offering a more convenient and effective method for central nervous diseases.