Serum exosomal miR-16-5p functions as a tumor inhibitor and a new biomarker for PD-L1 inhibitor-dependent immunotherapy in lung adenocarcinoma by regulating PD-L1 expression.

OBJECTIVES: We aimed at investigating whether serum exosomal miR-16-5p could be utilized as an immunotherapy biomarker in lung adenocarcinoma (LUAD) patients administered by programmed cell death ligand-1 (PD-L1) inhibitors, and to evaluate its functions in LUAD progression. METHODS: Sixty LUAD sufferers and 20 healthy controls (HCs) were covered in this work. We applied both IHC and WB to examine PD-L1 level in clinical tissue samples and utilized WB to quantify PD-L1 expression in LUAD cells and LUAD xenograft tissues, respectively. Transmission electron microscopy (TEM), WB, and nanoparticle tracking analysis (NTA) were executed to confirm the exosomes isolated from serum specimens and cell culture media. To quantify of exosomal miR-16-5p level from serum and culture medium of cultured cell, qRT-PCR experiment was utilized. The connection between tissue PD-L1 level and serum exosomal miR-16-5p expression in PD-L1-positive sufferers administered by PD-L1 inhibitors was verified using Spearman correlation coefficient analysis. In addition, the overall survival (OS) and progression-free survival (PFS) rates among PD-L1 inhibitor managed sufferers were acquired through a follow-up visit. Finally, we used a group of assays, including 5-bromo-2'-dexoyuridine (BrdU) and colony formation test, wound healing experiment, flow cytometry, and nude mice xenograft experiment, to explore the functions of circulating exosomal miR-16-5p on LUAD cell proliferation, apoptosis, and migration, as well as tumor development, respectively. RESULTS: PD-L1 expression was positively related to T stage (tumor size stage), and PD-L1 inhibitor treatment reduced the PD-L1 expression and mitigated T stage in PD-L1-positive LUAD sufferers. For PD-L1-positive LUAD sufferers, elevated PD-L1 expression or reduced serum exosomal miR-16-5p level were linked to longer PFS and OS upon PD-L1 inhibitor treatment. The number of exosomes in patient's serum was more than that in the serum of healthy individuals, and PD-L1 inhibitor treatment decreased the number of serum-derived exosomes in PD-L1-positive LUAD sufferers. Exosome-derived miR-16-5p was downregulated in patient's serum and cell culture medium, and this was negatively linked to tumor stage and PD-L1 expression. Meanwhile, PD-L1 inhibitor treatment could increase the serum exosomal miR-16-5p expression, and the expression change of serum exosomal miR-16-5p was diametrically related to PD-L1 after the treatment. Moreover, the overexpression of PD-L1 accelerated tumor growth and decreased the exosomal miR-16-5p content in cell culture media, while exosomal miR-16-5p overexpression in cell culture media inhibited tumor development by decreasing the PD-L1 expression. Exosomal miR-16-5p overexpression in cell culture media also depressed LUAD cell proliferation and migration, and stimulated cell apoptosis, especially in the cells which cultured in the mediums with PD-L1 inhibitor in vitro. CONCLUSIONS: Serum exosomal miR-16-5p may be a latent tumor inhibitor and a new biomarker for PD-L1 inhibitor-dependent immunotherapy in LUAD by regulating the PD-L1 expression.

Circulating Tumor Cells and Fibronectin 1 in the Prognosis of Nasopharyngeal Carcinoma.

OBJECTIVE: Nasopharyngeal carcinoma is highly endemic in Southeast China. Circulating tumor cell is an important biomarker in the prognosis of variety kinds of cancers. Overexpression of fibronectin 1 was observed in variety kinds of malignancies and may contribute to progress and metastasis of the cancers. The current study was aimed to investigate phenotypes of circulating tumor cell in nasopharyngeal carcinoma blood and fibronectin 1 expression in the circulating tumor cell, and their clinical application in predicting nasopharyngeal carcinoma prognosis. METHODS: Blood samples were obtained from nasopharyngeal carcinoma patients before and after treatment. CanPatrol circulating tumor cell enrichment and RNA in situ hybridization were applied to identify circulating tumor cell and its phenotypes. Fibronectin 1 messenger RNA expression in the cells of circulating tumors was examined by messenger RNA-in situ hybridization. RESULTS: Circulating tumor cell was not associated with tumor characteristics or lymph node metastasis. Patients with >9 circulating tumor cells or >5 mesenchymal phenotype circulating tumor cell per 5-mL blood had poorer progression-free survival (P < .05). Multivariable analysis demonstrated that 2 or more mesenchymal phenotype circulating tumor cells with high fibronectin 1 messenger RNA expression predicted shorter progression-free survival (P < .05). CONCLUSIONS: Circulating tumor cells with high-level fibronectin 1 expression was associated with poor survival in patients with nasopharyngeal carcinoma and could be an independent prognostic factor for nasopharyngeal carcinoma.

LEF1-AS1 is implicated in the malignant development of glioblastoma via sponging miR-543 to upregulate EN2.

Glioblastoma (GBM) has been regarded as the most aggressive disease in the nervous system. Accumulating literatures have illustrated the crucial role of competing endogenous RNAs (ceRNAs) network in the pathogenesis and progression of various tumors. The promoting effect of LEF1-AS1 on GBM development has been previously identified. This study attempted to explore the underlying mechanism of LEF1-AS1 in GBM. Data of clinical GBM patients was downloaded from TCGA and GEO databases. The proliferative ability, clonogenic vitality, invasive, and migratory capabilities of GBM cells were measured using Cell counting kit-8 (CCK-8), colony formation and transwell assays. Luciferase reporter gene analysis was performed to verify the correlations between LEF1-AS1/EN2 and miR-543. qRT-PCR and western blotting were implemented to evaluate the mRNA and protein levels, respectively. Our results consolidated that LEF1-AS1 was highly expressed in GBM tissue specimens and its up-regulation induced unfavorable prognosis. The loss/gain-of-function analyses verified that LEF1-AS1 promoted the GBM cell malignant behaviors. Mechanically, LEF1-AS1 acted as a ceRNA for miR-543 and positively regulated engrailed homeobox 2 (EN2) expression. Down-regulation of miR-543 elevated GBM cell malignant behaviors, which was reversed by LEF1-AS1 knockdown. Meanwhile, the LEF1-AS1 inhibition could arrest the promoting effect of high-regulated EN2 on GBM cell aggressiveness and vice versa. In conclusion, our findings identified LEF1-AS1 as a ceRNA for miR-543 and showed that EN2 was positively regulated by LEF1-AS1. The LEF1-AS1/miR-543/EN2, as a novel ceRNA network, was implicated in the progression of GBM, which provided a novel insight for GBM treatment.

A preliminary study on the potential of Mycoplasma pneumoniae to induce dyskaryotic change in respiratory epithelium in adult community-acquired pneumonia.

BACKGROUND: This study aimed to explore the cellular morphology of respiratory epithelium in Mycoplasma pneumonia (MpP) patients. MATERIALS AND METHODS: The cast-off cell morphological findings from bronchoscopic brushings in MpP and community-acquired pneumonia (CAP) caused by typical pathogens were reviewed. RESULTS: Compared with the CAP group, cellular dysplasia in respiratory tract epithelial brushings was significantly greater in MpP patients (P = 0.033). CONCLUSION: Unique biological characteristics and mechanisms of pathogenesis of Mycoplasma pneumoniae (Mp) may result in dyskaryotic changes in respiratory epithelium in adult MpP.

Rapid Identification of Bacterial Species Associated with Bronchiectasis via Metagenomic Approach.

Bronchiectasis is a chronic lung disorder and a number of bacterial pathogens are involved. However, 30%-40% of sputum and purulent samples in good quality failed to grow any pathogenic bacteria, making it difficult to confirm the pathogen. In this study, we collected bronchoalveolar lavage fluid from a bronchiectasis patient undergoing acute exacerbation, and sent for 16S rDNA pyrosequencing by a 454 GS Junior machine. Metagenomic analysis showed the composition of bacterial community in sample was complex. More than a half of reads (51.3%) were from Pseudomonas aeruginosa. This result was corresponding with the culture result but came out 2 d earlier, which is meaningful for early diagnosis and treatment. The detection with 16S rDNA pyrosequencing technology is more sensitive and rapid than routine culture, and can detect the co-infection or symbiosis in airway, giving us a novel and convenient approach to perform rapid diagnosis.