MicroRNAs regulate target gene expression and are involved in cell proliferation, apoptosis, differentiation, tumor invasion, and cancer stem cell regulation, among other processes. MicroRNA-26b (miR-26b) is closely related to tumor occurrence and development. In this study, we analyzed miR-26b expression in osteosarcoma tissue, its effect on Saos-2 osteosarcoma cell proliferation and invasion, and its relationship with 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3) expression. Osteosarcoma tissue was obtained from surgical patients and normal tissue adjacent to the tumor was used as a control. Real-time polymerase chain reaction was applied to detect miR-26b expression in cancer tissue and normal tissue. A vector expressing miR-26b was constructed and transfected into Saos-2. An MTT assay, cell invasion assay, and scratch experiment were used to analyze the effect of miR-26b on Saos-2 cell proliferation, invasion, and migration abilities. Western blotting analysis was performed to investigate the role of miR-26b on PFKFB3 expression. miR-26b expression in normal tissue was 7.55-fold higher than in osteosarcoma tissue (t = 10.20, P = 0.006). Compared with control tissue, miR-26b significantly inhibited osteosarcoma proliferation, migration, and invasion (P < 0.05). Western blotting results revealed that PFKFB3 protein expression decreased in Saos-2 cells after transfection with miR-26b. miR-26b was down-regulated in osteosarcoma tissue. miR- 26b may inhibit osteosarcoma cell proliferation, migration, and invasion by regulating PFKFB3 protein expression. miR-26b may have a tumor suppressor role in tumor occurrence and development.

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MicroRNA-26b inhibits osteosarcoma cell migration and invasion by down-regulating PFKFB3 expression.

Alternative splicing (AS) is a ubiquitous post-transcriptional regulatory mechanism in eukaryotes that generates multiple mRNA isoforms from a single gene, increasing diversity of mRNAs and proteins that are essential for eukaryotic developmental processes and responses to environmental stress. Results showed that a total of 37,463 AS events were identified in rainbow trout hepatocytes. In addition, a total of 364 differential alternative splicing (DAS) events were identified in hepatocytes under selenium nanoparticles (SeNPs) and 3632 DAS events were identified under a combination of SeNPs and heat stress (24 °C). Gene Ontology (GO) enrichment showed that some subcategories “immune effector processes”, “response to stimuli” and “antioxidant activity” were associated with immunity, abiotic stimuli and antioxidants. Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment showed that differentially expressed genes (DEGs) were significantly enriched in spliceosomes by adding SeNPs in heat-stressed hepatocytes. Splicing factor family (SRSF3, SRSF7, SRSF9, U2AF1 and U2AF2) and pre-RNA splicing factors (ACIN1 and PPRF18) were significantly upregulated and promoted AS. Furthermore, addition of SeNPs activated the phosphatidylinositol signaling system and upregulated the related genes PI4KA, DGKH, ITPK1 and Ocrl, and thus attenuated the inflammatory response to heat stress and enhanced resistance to heat stress by activating the adherent plaque kinase-PI3K-Akt signaling pathway and calcium channels. Those findings suggested that AS could be an essential regulatory mechanism in adaptation of rainbow trout to heat-stressed environments. 

Effect of selenium nanoparticles on alternative splicing in heat-stressed rainbow trout primary hepatocytes.

Bifunctional enzyme 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-2 (PFKFB-2) is represented by several alternative splice variants and plays a significant role in the glycolysis regulation in the brain, lung, testis and heart cells. The expression of PFKFB-2 mRNA and its alternative splice variants in these rat vital organs after single intratracheal injection of silver nanoparticles was studied. It was shown that the expression of PFKFB-2 mRNA is significantly changed in different rat tissues under silver nanoparticles action. The effect of silver nanoparticles on the expression of PFKFB-2 mRNA was observed one day after its injection to animals. In 3 and 14 days the effect of silver nanoparticles was increased (in testes) or kept on the approximately same level (in other investigated tissues). The expression of PFKFB-2 mRNA in most tissues is returned to its control levels one year after the injection of silver nanoparticles to the rats. It was also shown that the expression of alternative splice variants of PFKFB-2 mRNA without functional activity of 6-phosphofructo-2-kinase is significantly increased in different tissues 1, 3 and 14 days after single injection of silver nanoparticles. The results of this investigation demonstrate clearly that silver nanoparticles significantly affect the expression of PFKFB-2 mRNA on the alternative splicing level in different vital organs and show their effect on the important mechanisms of metabolism regulation in the cells on the level of key enzyme gene expression.

[Effect of silver nanoparticles on the expression of 6-phosphofructo-2-kinase/ fructose-2,6-bisphosphatase-2 mRNA and its alternative splice variants in different rat organs].

T O Zinchenko

Papers

[Effect of silver nanoparticles on the expression of 6-phosphofructo-2-kinase/ fructose-2,6-bisphosphatase-2 mRNA and its alternative splice variants in different rat organs].