MiR-33a Overexpression Exacerbates Diabetic Nephropathy Through Sirt6-dependent Notch Signaling.

INTRODUCTION: Diabetic nephropathy (DN) belongs to the major cause of end-stage kidney disease. We probed the functions of a microRNA miR-33a in inducing podocytes injury during childhood  DN (CDN). METHODS: Kidney samples were collected from 20 children with DN. Matrix deposition and glomerular basement membranes thickness were examined by periodic acid-Schiff staining. Immunofluorescence staining was performed to assess kidney function-related proteins. MicroRNA (MiR)-33a mimic together with miR-33a inhibitor was transfected into podocytes for determining the roles of miR-33a. Glomerular podocyte apoptosis was determined by terminal deoxynucleotidyl transferase (TdT) dUTP Nick-End Labeling (TUNEL) staining along with flow cytometry. RESULTS: Down-regulation of Nephrin and Podocin and increased podocyte apoptosis rate were observed in the glomerulus of CDN as well as podocytes treated with high glucose. MiR-33a was up regulated in the glomeruli and glucose-treated podocytes. Injury in podocytes was aggravated with miR-33a elevation but alleviated with miR-33a inhibition. Moreover, the expression of Sirtuin 6 (Sirt6) was decreased while the levels of notch receptor 1 (Notch1) and notch receptor 4 (Notch4) were elevated in the glomerulus and glucose-treated podocytes. Decreased level of Sirt6 upon glucose treatment was abrogated by miR-33a inhibition, and the podocytes injury induced by glucose exposure was relieved by Sirt6 via Notch signaling. CONCLUSION: These findings indicated that miR-33a promoted podocyte injury via targeting Sirt6-dependent Notch signaling in CDN, which might provide a novel sight for CDN treatment. DOI: 10.52547/ijkd.7904.

Bioinformatics-based screening and analysis of the key genes involved in the influence of antiangiogenesis on myeloid-derived suppressor cells and their effects on the immune microenvironment.

This study aimed to screen differentially expressed genes (DEGs) involved in the influence of antiangiogenic therapy on myeloid-derived suppressor cell (MDSC) infiltration and investigate their mechanisms of action. Data on DEGs after the action of antiangiogenic drugs in a pan-cancer context were obtained from the Gene Expression Omnibus (GEO) database. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses were performed using the clusterProfiler package in R software. Single-sample gene set enrichment analysis was performed using the gene set variation analysis package to evaluate the levels of immune cells and the activity of immune-related pathways. The relationships of DEGs with the infiltration levels of MDSCs and specific immune cell subpopulations were investigated via gene module analysis. The top 10 key genes were subsequently obtained from PPI network analysis using the cytoHubba plugin of the Cytoscape platform. When the DEGs of the four datasets were intersected, a DEG in the intersection of three datasets and 12 DEGs in the intersection of two datasets were upregulated, and 28 DEGs in the intersection of two datasets were downregulated. GO and KEGG pathway enrichment analyses revealed that the DEGs were associated with multiple important signaling pathways closely related to tumor onset and development, including cell differentiation, cell proliferation, the cell cycle, and immune responses. Most downregulated genes in lung adenocarcinoma (LUAD) were positively correlated with MDSC expression. Only MGP was negatively correlated; the correlation between CACNG6 and MDSC expression was statistically insignificant. In lung squamous cell carcinoma (LUSC), the relationships of PMEPA1, PCDH7, NEURL1B, and CACNG6 with MDSC expression were statistically insignificant; MGP was negatively correlated with MDSC expression. The top 10 key genes with the highest degree scores obtained using the cytoHubba plugin of Cytoscape were AURKB, RRM2, BUB1, NUSAP1, PRC1, TOP2A, NCAPH, CENPA, KIF2C, and CCNA2. Most of these genes were upregulated in LUAD and associated with immune cell infiltration and prognosis in tumors. An analysis of the relationships between DEGs and infiltration by other specific immune cells revealed the presence of consistent patterns in the downregulated genes, which exhibited positive correlations with the levels of Th2 cells, γδ T cells, and CD56dim NK cells, and negative correlations with other infiltrating immune cells. Antiangiogenic therapy may regulate MDSC infiltration through multiple important signaling pathways closely associated with tumor onset and development, such as cell differentiation, cell proliferation, the cell cycle, and immune responses. Antiangiogenic drugs may exert effects by affecting various types of infiltrating cells associated with immune suppression.

Anlotinib reduces the suppressive capacity of monocytic myeloid-derived suppressor cells and potentiates the immune microenvironment normalization window in a mouse lung cancer model.

By exploring the effects of an antiangiogenic small molecule drug named anlotinib on the levels of myeloid-derived suppressor cells (MDSCs) in a mouse xenograft model of lung cancer, the role of anti-angiogenesis in remodeling the immune microenvironment was discussed. In addition, the impact of anlotinib on the normalization of the immune microenvironment and time window was examined, providing a theoretical basis for the optimization of clinical strategies applying anlotinib combined with PD-1 inhibitors. On the basis of the LLC mouse xenograft model, MDSCs and MDSCs + immune microenvironment were examined in tissues, respectively, according to different samples. The former observation included the control (group A) and anlotinib monotherapy (group B) groups; the latter also included the control (group C) and anlotinib monotherapy (group D) groups. The levels of MDSCs in peripheral blood at different time points were analyzed by flow cytometry, and the levels of MDSCs in tissue samples at different time points were evaluated by immunofluorescence and immunohistochemistry. The volumes of subcutaneous xenografts were significantly smaller in the anlotinib treatment group compared with the control group ( P < 0.005). Flow cytometry showed that compared with the control group, the intratumoral percentages of total MDSCs ( P < 0.01) and mononuclear-MDSCs ( P < 0.05) were significantly decreased on days 3 and 17 after anlotinib treatment in peripheral blood samples; however, there was no significant difference in granulocytic-MDSCs changes between the experimental and control groups. Immunofluorescence showed that the levels of MDSCs in both the experimental and control groups reached the lowest points 10 days after drug administration, and were significantly lower in the experimental group than in the control group ( P < 0.05). Anlotinib reduces the levels of MDSCs in the mouse xenograft model of lung cancer, with the characteristics of time window. This study provides a basis for further exploring strategies for anti-angiogenic treatment combined with immunotherapy in lung cancer based on time-window dosing.

Induction of ligustrazine apoptosis of A549 cells through activation of death receptor pathway.

Background: The aim of this study was to evaluate the effect of ligustrazine on the apoptosis of A549 cells and clarify the mechanism of ligustrazine-induced apoptosis. Methods: Ligustrazine was prepared with medium according to the gradient concentration. Based on a cytotoxicity test, 3 different concentrations of ligustrazine were selected to form low, medium, and high groups, with a 0 mg/mL dose used as the control. The apoptosis degree and Fas (Fas cell surface death receptor) and Fas-L (Fas Ligand) expression were detected by flow cytometry and quantitative polymerase chain reaction (qPCR), respectively; meanwhile, the activity of caspase 8 and caspase 3 was analyzed by enzyme-linked immunosorbent assay (ELISA) and qPCR, respectively. Results: After 24 hours of ligustrazine administration, the survival rate of A549 cells decreased with the increase of drug concentration, while the rate of apoptosis increased with the increase of drug concentration. Meanwhile, Fas and Fas-L expression was found to be significantly increased at both the gene and protein level, which was positively correlated with drug concentration. Furthermore, the expression of caspase 8 and caspase 3 was positively correlated with the concentration of ligustrazine, and there was significant difference compared with the control group. Conclusions: Ligustrazine can induce the apoptosis of A549 cells via the upregulation of Fas- and caspase-activating death receptor pathway expression.

LncRNA SNHG3 Promotes Proliferation and Metastasis of Non-Small-Cell Lung Cancer Cells Through miR-515-5p/SUMO2 Axis.

Lung cancer is a global disease and a major cause of cancer-related mortality worldwide. Accumulated studies have confirmed the essential role of long non-coding RNAs (lncRNAs) in the occurrence and development of cancers. Meanwhile, there have been reports concerning the role of Small Nucleolar RNA Host Gene 3 (SNHG3) in various cancers. However, there are so far few studies on the function and mechanism of SNHG3 in lung cancer. In the present study, SNHG3 was found to be highly expressed in lung cancer tissues and cells. Downregulation of SNHG3 could inhibit cell proliferation, migration, invasion and epithelial-mesenchymal transition (EMT) process. In addition, SNHG3 was found to have the ability to bind to miR-515-5p. Furthermore, Small Ubiquitin Like Modifier 2 (SUMO2) was identified to be the downstream target of miR-515-5p, which was negatively correlated with miR-515-5p expression. SNHG3 could positively regulate SUMO2 expression by sponging miR-515-5p. In addition, the rescue experiment showed that simultaneous transfection of miR-515-5p or SUMO2 siRNA could reverse the effect of SNHG3 expression on cell proliferation and metastasis. Collectively, our study demonstrates that SNHG3 can act on miR-515-5p in the form of competitive endogenous RNA (ceRNA) to regulate SUMO2 positively and thus affect the proliferation and metastasis of NSCLC cells. Findings in our study support that SNHG3/miR-515-5p/SUMO2 regulatory axis may become a potential therapeutic target for lung cancer.