Emerging role of microRNAs in lipid metabolism

microRNAs (miRNAs or miRs) are small non-coding RNAs that are involved in post-transcriptional regulation of their target genes in a sequence-specific manner. Emerging evidence demonstrates that miRNAs are critical regulators of lipid synthesis, fatty acid oxidation and lipoprotein formation and secretion. Dysregulation of miRNAs disrupts gene regulatory network, leading to metabolic syndrome and its related diseases. In this review, we introduced epigenetic and transcriptional regulation of miRNAs expression. We emphasized on several representative miRNAs that are functionally involved into lipid metabolism, including miR-33/33(⁎), miR122, miR27a/b, miR378/378(⁎), miR-34a and miR-21. Understanding the function of miRNAs in lipid homeostasis may provide potential therapeutic strategies for fatty liver disease.

The Expression of AGO2 and DGCR8 in Idiopathic Sudden Sensorineural Hearing Loss

OBJECTIVES: The microRNAs have been implicated in the development and function of the inner ear, especially in contribution to hearing. However, the impact of idiopathic sudden sensorineural hearing loss (SSNHL) on expression of miRNA biogenesis-related components has not been established. To investigate the regulations of microRNA (miRNA) biogenesis-related components, argonaute 2 (AGO2) and DiGeorge syndrome critical region gene 8 (DGCR8) mRNA expression in SSNHL and to evaluate the value of clinical parameters on their expression. METHODS: Thirty-seven patients diagnosed with SSNHL and fifty-one healthy volunteers were included in this study. We measured mRNA expression levels of AGO2 and DGCR8 in whole blood cells but erythrocytes of patients with SSNHL and controls, using reverse transcription and real-time polymerase chain reaction analysis. RESULTS: The mRNA expression level of AGO2 is upregulated in SSNHL. The expression level of AGO2 was significantly correlated with that of DGCR8 in both patients with SSNHL and controls. Expression level of AGO2 in SSNHL was correlated with white blood cell counts. CONCLUSION: This study demonstrated for the first time that the AGO2 mRNA expression level was upregulated in SSNHL, suggesting its important role in pathobiology of SSNHL development.

Argonaute-2-null embryonic stem cells are retarded in self-renewal and differentiation.

RNA interference (RNAi) pathways regulate self-renewal and differentiation of embryonic stem (ES) cells. Argonaute 2 (Ago2) is a vital component of RNA-induced silencing complex (RISC) and the only Ago protein with slicer activity. We generated Ago2-deficient ES cells by conditional gene targeting. Ago2-deficient ES cells are defective in the small-RNA-mediated gene silencing and are significantly compromised in biogenesis of mature microRNA. The self-renewal rate of Ago2-deficient ES cells is affected due to failure of silencing of Cdkn1a by EScell- specific microRNAs (miRNA) in the absence of Ago2. Interestingly, unlike Dicer- and Dgcr8-deficient ES cells, they differentiate to all three germ layers both in vivo and in vitro. However, early differentiation of Ago2-deficient ES cells is delayed by 2–4 days as indicated by persistence of higher levels of self-renewal/ pluripotency markers during differentiation. Further, appearance of morphological and differentiation markers is also delayed during the differentiation. In this study we show that Ago2 is essential for normal self-renewal and differentiation. Also, our data suggest that self-renewal and differentiation of ES cells are regulated by both siRNA and miRNA pathways.

Generation and preliminary application of polyclonal antibodies against human Argonaute2 / 中国免疫学杂志

Objective:To generate rabbit polyclonal antibody against human Argonaute2 (Ago2) protein and to identify its functional characterization for determination of differential expression and cellular localization of Ago2 protein in various cell lines.Methods:DNAstar software was applied for searching the high antigenicity region of Ago2 gene sequence termed k-Ago2.Prokaryotic expressing plasmid was constructed and transformed to E.coli BL21 (DE3) to induce expression by IPTG.The fusion protein was injected into rabbits subcutaneously to produce polyclonal antibodies after purification by gel regaining.ELISA was operated to detect antibody titer.Western blot was used to identify the specificity and sensitivity of the antibodies and detect the differential expression of Ago2 protein in various cell lines.Meanwhile,immunofluorescence experiments were arranged to show cellular localization of Ago2 protein.Results:The prokaryotic expressing plasmid was constructed correctly.K-Ago2 protein was expressed and purified,and then rabbit polyclonal antibodies against Ago2 were generated after immunization with k-Ago2 protein.The titer detected by ELISA was 1∶19 000.Western blot results demonstrated the high specificity of the antibodies.Finally,we successfully observed the differential expression and cellular localization of Ago2 protein in various cell lines.Conclusion:The polyclonal antibody against Ago2 protein has been achieved successfully.It will be propitious for the intensive study of the RNAi mechanism and even profound clinical application.