ABSTRACT
Objective To study the regulation of microRNA - 22(miR - 22)on glycometabolism of hemato-poietic stem cell TF - 1 and its molecular mechanism. Methods TF - 1 cells were cultured for 2 h under hypoxic con-ditions. The expression levels of Glut4 and miR - 22 was detected by quantitative real-time PCR(qRT - PCR). The sgRNA vector of the miR - 22 gene was constructed and miR - 22 gene of TK - 1 cells was knocked out by the CRISPR/ Cas9 technique. Overexpression vectors were constructed,and miR - 22 knocked - out cells were introduced to overexpress miR - 22,the expression of miR - 22 was detected by qRT - PCR and the expression levels of Glut4 and PPAR - γ were detected by qRT - PCR and Western blot. Results Compared with the control group,the expression of miR - 22 in TF - 1 cells decreased (0. 015 ± 0. 000 vs. 0. 056 ± 0. 001)and the expression of Glut4 (0. 351 ± 0. 038 vs. 0. 152 ± 0. 005)and PPAR - γ (0. 421 ± 0. 017 vs. 0. 248 ± 0. 008)increased,when TF - 1 cells were cultured for 2 h under hypoxic conditions,and the differences were statistically significant (all P < 0. 05). Compared with the control group,the expression levels of Glut4 (0. 019 ± 0. 00 vs. 0. 008 ± 0. 000)and PPAR - γ (0. 038 ± 0. 001 vs. 0. 019 ± 0. 000)were significantly increased after miR - 22 gene silencing,and they were significantly decreased (Glut4:0. 005 ± 0. 000 vs. 0. 008 ± 0. 000;PPAR - γ:0. 137 ± 0. 000 vs. 0. 019 ± 0. 000)after overexpression of miR - 22,and the differences were statistically significant (all P < 0. 05). Conclusions It suggests that miR - 22 ex-erts a negative regulation on glycometabolism of hematopoietic stem cell TF - 1 by downregulating the expression of PPAR - γ. A new regulatory factor of TF - 1 glycometabolism and the mechanisms are identified,which has provided new ideas for the targeted medication of diseases induced by hematopoietic stem cell dysfunction.
ABSTRACT
MicroRNA (miRNA) are a class of endogenous single -stranded non-coding RNA, which regulate gene expression post-transcriptionally by combining with the target mRNA and play a vital role in biological and pathological processes including the growth of organism , metabolic regulation, disease prediction and intervention.Thus miRNA detection is of considerable significance in disease diagnosis and the research of miRNA function.Because of the restriction factors about miRNA itself such as short sequence, low abundance and highly homologous , traditional methods for miRNA detection cannot meet the current demands due to the limitations like unsatisfactory sensitivity and complicated operation .This review summarizes the newly development about signal amplification -based methods for miRNA, including the advantages and limitations of all kinds of novel methods , and highlights the future trends as well.