Development and validation a prognostic model based on natural killer T cells marker genes for predicting prognosis and characterizing immune status in glioblastoma through integrated analysis of single-cell and bulk RNA sequencing.

BACKGROUND: Glioblastoma (GBM) is an aggressive and unstoppable malignancy. Natural killer T (NKT) cells, characterized by specific markers, play pivotal roles in many tumor-associated pathophysiological processes. Therefore, investigating the functions and complex interactions of NKT cells is great interest for exploring GBM. METHODS: We acquired a single-cell RNA-sequencing (scRNA-seq) dataset of GBM from Gene Expression Omnibus (GEO) database. The weighted correlation network analysis (WGCNA) was employed to further screen genes subpopulations. Subsequently, we integrated the GBM cohorts from The Cancer Genome Atlas (TCGA) and Chinese Glioma Genome Atlas (CGGA) databases to describe different subtypes by consensus clustering and developed a prognostic model by least absolute selection and shrinkage operator (LASSO) and multivariate Cox regression analysis. We further investigated differences in survival rates and clinical characteristics among different risk groups. Furthermore, a nomogram was developed by combining riskscore with the clinical characteristics. We investigated the abundance of immune cells in the tumor microenvironment (TME) by CIBERSORT and single sample gene set enrichment analysis (ssGSEA) algorithms. Immunotherapy efficacy assessment was done with the assistance of Tumor Immune Dysfunction and Exclusion (TIDE) and The Cancer Immunome Atlas (TCIA) databases. Real-time quantitative polymerase chain reaction (RT-qPCR) experiments and immunohistochemical profiles of tissues were utilized to validate model genes. RESULTS: We identified 945 NKT cells marker genes from scRNA-seq data. Through further screening, 107 genes were accurately identified, of which 15 were significantly correlated with prognosis. We distinguished GBM samples into two distinct subtypes and successfully developed a robust prognostic prediction model. Survival analysis indicated that high expression of NKT cell marker genes was significantly associated with poor prognosis in GBM patients. Riskscore can be used as an independent prognostic factor. The nomogram was demonstrated remarkable utility in aiding clinical decision making. Tumor immune microenvironment analysis revealed significant differences of immune infiltration characteristics between different risk groups. In addition, the expression levels of immune checkpoint-associated genes were consistently elevated in the high-risk group, suggesting more prominent immune escape but also a stronger response to immune checkpoint inhibitors. CONCLUSIONS: By integrating scRNA-seq and bulk RNA-seq data analysis, we successfully developed a prognostic prediction model that incorporates two pivotal NKT cells marker genes, namely, CD44 and TNFSF14. This model has exhibited outstanding performance in assessing the prognosis of GBM patients. Furthermore, we conducted a preliminary investigation into the immune microenvironment across various risk groups that contributes to uncover promising immunotherapeutic targets specific to GBM.

Comprehensive analysis identifies cuproptosis-related gene DLAT as a potential prognostic and immunological biomarker in pancreatic adenocarcinoma.

BACKGROUND: Cuproptosis is a regulated cell death form associated with tumor progression, clinical outcomes, and immune response. However, the role of cuproptosis in pancreatic adenocarcinoma (PAAD) remains unclear. This study aims to investigate the implications of cuproptosis-related genes (CRGs) in PAAD by integrated bioinformatic methods and clinical validation. METHODS: Gene expression data and clinical information were downloaded from UCSC Xena platform. We analyzed the expression, mutation, methylation, and correlations of CRGs in PAAD. Then, based on the expression profiles of CRGs, patients were divided into 3 groups by consensus clustering algorithm. Dihydrolipoamide acetyltransferase (DLAT) was chosen for further exploration, including prognostic analysis, co-expression analysis, functional enrichment analysis, and immune landscape analysis. The DLAT-based risk model was established by Cox and LASSO regression analysis in the training cohort, and then verified in the validation cohort. Quantitative reverse transcriptase polymerase chain reaction (RT-qPCR) and immunohistochemistry (IHC) assays were performed to examine the expression levels of DLAT in vitro and in vivo, respectively. RESULTS: Most CRGs were highly expressed in PAAD. Among these genes, increased DLAT could serve as an independent risk factor for survival. Co-expression network and functional enrichment analysis indicated that DLAT was engaged in multiple tumor-related pathways. Moreover, DLAT expression was positively correlated with diverse immunological characteristics, such as immune cell infiltration, cancer-immunity cycle, immunotherapy-predicted pathways, and inhibitory immune checkpoints. Submap analysis demonstrated that DLAT-high patients were more responsive to immunotherapeutic agents. Notably, the DLAT-based risk score model possessed high accuracy in predicting prognosis. Finally, the upregulated expression of DLAT was verified by RT-qPCR and IHC assays. CONCLUSIONS: We developed a DLAT-based model to predict patients' clinical outcomes and demonstrated that DLAT was a promising prognostic and immunological biomarker in PAAD, thereby providing a new possibility for tumor therapy.

19-ring-air-core fiber supporting thousands of OAM modes for spatial division multiplexing.

We propose and design a 19-ring-air-core fiber that can support about 3000 orbital angular momentum (OAM) modes (156 modes in each ring) with <-80 dB inter-ring cross talk across the entire C and L bands after 100-km fiber propagation. Moreover, the eigenmodes are all separated from their adjacent modes by effective index differences >2.67 × 10-4 and mode groups by > 1.90 × 10-2, which can guarantee the stable transmission of OAM modes. This designed fiber is a potential candidate for applications in spatial division multiplexing (SDM) of optical channels to improve the capacity of next-generation high-speed optical communication systems, especially in short-distance applications. In this Letter, we also show the relationship between supported OAM mode numbers, total cross talk, and effective refractive index of intra-ring modes during the optimization of fiber through numerical simulations. It can provide a related reference for the future design of multi-ring-core fibers.

GPC3-targeted and curcumin-loaded phospholipid microbubbles for sono-photodynamic therapy in liver cancer cells.

More effective strategies are needed to improve the treatment of liver cancer. Sono-photodynamic therapy (SPDT) has a more obvious antitumor effect than sonodynamic therapy (SDT) or photodynamic therapy (PDT). We aimed to investigate Glypican-3-targeted, curcumin-loaded microbubbles (GPC3-CUR-MBs)-mediated SPDT in liver cancer cells in vitro and in vivo. GPC3-CUR-MBs were prepared by streptavidin-biotin interactions and the immune ligation method. The characterization and toxicity of GPC3-CUR-MBs and the anti-liver cancer effects of GPC3-CUR-MB-mediated SPDT in vitro and in vivo were studied. We synthetized GPC3-CUR-MBs and found that GPC3-CUR-MBs had no significant toxicity to HepG2 liver cancer cells. In terms of the anti-liver cancer effects in vitro and in vivo, when we used CUR, CUR-MBs or GPC3-CUR-MBs as the sono/photosensitizers, the outcome of SPDT was superior to that of SDT or PDT alone. The outcomes with GPC3-CUR-MBs were better than those with CUR or CUR-MBs in the SDT, PDT or SPDT groups. During the treatment period, the weight of the HepG2 tumor-bearing mice did not decrease significantly, and no significant evidence of lung, heart, liver, spleen and kidney damage was found with H&E staining. Our results indicated that the anti-liver tumor effect of SPDT was better than that of SDT and PDT and that GPC3-CUR-MBs were promising sono/photosensitizers.

5-Aminolevulinic acid hydrochloride loaded microbubbles-mediated sonodynamic therapy in pancreatic cancer cells.

5-Aminolevulinic acid hydrochloride (ALA)-mediated sonodynamic therapy (SDT) had anti-tumour effect on pancreatic cancer cells. Hence, ALA loaded lipid/poly(lactic-co-glycolic acid) (PLGA) microbubbles (MBs)-mediated SDT for pancreatic cancer has great potential. The average size of ALA-lipid MBs and ALA-PLGA MBs was about 3.0 µm. The two kinds of MBs had good biocompatibility to normal HPDE6-C7 cells and were not toxic to pancreatic cancer cells. Compared with ALA-induced SDT, a statistically significant decrease in cell viability was observed in ALA lipid/PLGA MBs combined with ultrasound groups in AsPC-1 and BxPC-3 cells (p < .05). Obvious effect on the apoptotic rate, apoptosis and pyroptosis morphology, enhanced reactive oxygen species was found in ALA-lipid/PLGA MBs mediated SDT in vitro. Through in vivo study, we found ALA-lipid/PLGA MBs-mediated SDT was a promise treatment for pancreatic cancer.

Curcumin-Loaded Poly(L-lactide-co-glycolide) Microbubble-Mediated Sono-photodynamic Therapy in Liver Cancer Cells.

Sono-photodynamic therapy (SPDT) activates the same photo-/sonosensitizer and exerts more marked antitumor effects than sonodynamic therapy or photodynamic therapy. We aimed to explore the utilization of curcumin (CUR)-loaded poly(L-lactide-co-glycolide) microbubble (MB)-mediated SPDT (CUR-PLGA-MB-SPDT) in HepG2 liver cancer cells. The cytotoxicity and intracellular accumulation of CUR were determined. We used 40 µM CUR as the photo-/sonosensitizer for 3 h. In a comparison of CUR-SDT or CUR-PDT, HepG2 cell viability decreased and apoptotic rate increased in CUR-SPDT. The CUR-PLGA MBs had round spheres with smooth surfaces and an average size of 3.7 µm. In CUR-PLGA MBs, drug entrapment efficiency and drug-loading capacity were 74.29 ± 2.60% and 17.14 ± 0.60%, respectively. CUR-loaded PLGA MBs (CUR-PLGA MBs) had good biocompatibility with normal L02 cells and were almost non-cytotoxic to HepG2 cells. Among CUR-SDT, CUR-PDT, CUR-SPDT or CUR-PLGA-MB-SDT, the cell CUR-PLGA-MB-SPDT had the lowest viability. Transmission electron microscopy revealed pyroptosis and apoptosis in the CUR-PLGA-MB-SPDT group; the potential mechanism was related to the mitochondrial membrane potential loss and increased production of intracellular reactive oxygen species. These findings suggested that CUR-PLGA-MB-SPDT may be a promising treatment for liver cancer.

ANXA2 Silencing Inhibits Proliferation, Invasion, and Migration in Gastric Cancer Cells.

Annexin A2 (ANXA2) has been well known to associate with the progress of malignant tumor. However, the biological behavior of ANXA2 in gastric cancer (GC) remains unclear. We made a hypothesis in transcriptome level from TCGA datasets. Then, we used immunohistochemical staining to quantify the expression level of ANXA2 protein in GC tissues compared with adjacent tissues. Quantitative real-time PCR and western blot were used for analyzing ANXA2 expression in human GC (SGC-7901, MKN-45, BGC-823, and AGS) cell lines. We investigated the effect of a lentivirus-mediated knock-down of ANXA2 on the proliferation, invasion and migration of gastric cancer AGS cells. Cell proliferation was examined by MTT and colony formation tests. Cell apoptosis and cycle were measured by flow cytometry. Migration and invasion were detected by transwell assay. We found that high expression of ANXA2 can increase the mobility of cancer cells from TCGA datasets. ANXA2 was upregulated in GC tissues compared with adjacent tissues. AGS cell line displayed significantly higher expression of ANXA2 among the four GC cell lines. In addition, ANXA2 silencing led to a weakened ability of proliferation, invasion, and migration in GC cells; targeting of ANXA2 may be a potential therapeutic strategy for GC patients.

808 nm Near-Infrared Light-Excited UCNPs@mSiO2-Ce6-GPC3 Nanocomposites For Photodynamic Therapy In Liver Cancer.

BACKGROUND: It is important to explore effective treatment for liver cancer. Photodynamic therapy (PDT) is a novel technique to treat liver cancer, but its clinical application is obstructed by limited depth of visible light penetration into tissue. The near-infrared (NIR) photosensitizer is a potential solution to the limitations of PDT for deep tumor tissue treatment. PURPOSE: We aimed to investigate 808 nm NIR light-excited UCNPs@mSiO2-Ce6-GPC3 nanocomposites for PDT in liver cancer. METHODS: In our study, 808 nm NIR light-excited upconversion nanoparticles (UCNPs) were simultaneously loaded with the photosensitizer chlorin e6 (Ce6) and the antibody glypican-3 (GPC3), which is overexpressed in hepatocellular carcinoma cells. The multitasking UCNPs@mSiO2-Ce6-GPC3 nanoparticles under 808 nm laser irradiation with enhanced depth of penetration would enable the effective targeting of PDT. RESULTS: We found that the UCNPs@mSiO2-Ce6-GPC3 nanoparticles had good biocompatibility, low toxicity, excellent cell imaging in HepG2 cancer cells and high anti-tumor effect in vitro and in vivo. CONCLUSION: We believe that the utilization of 808 nm NIR excited UCNPs@mSiO2-Ce6-GPC3 nanoparticles for PDT is a safe and potential therapeutic option for liver cancer.