ABSTRACT
Objective To explore the role of miR-199a-3p in osteoblast proliferation induced by fluid shear stress (FSS) and the potential molecular mechanism. Methods Osteoblast MC3T3-E1 was treated with 1. 2 Pa FSS with time gradients of 0, 15, 30, 45, 60, 75 and 90 min, respectively. MC3T3-E1 cells were transfected with miR-199a-3p mimic or miR-199a-3p inhibitor. MC3T3-E1 cells were transfected with miR-199a-3p mimic and itsnegative control and then treated with 1. 2 Pa FSS for 45 min. The pc DNA NC, pc DNA-CABLES -1, si RNA NC and si RNA CABLES-1 were transfected into MC3T3-E1 cells. The pc DNA-CABLES-1 and mir-199a-3p mimic and SI NA-cables-1 and miR-199a-3p inhibitor were co-transfected, respectively. Cell activity was detected by CCK-8 assay. Real-time quantitative PCR (RT-qPCR) was used to detect expression levels of CABLES-1, miR-199a-3p, CDK 6, Cyclin D1 and PCNA. Luciferase reporting assay was used to detect targeting relationship between CABLES-1 and miR-199a-3p. Immunofluorescence was used to detect protein expression of CABLES-1.Western blot was used to detect protein expression of CABLES-1, CDK 6, PCNA and Cyclin D1. Results Mir- 199a-3p in MC3T3-E1 cells was significantly down-regulated by FSS. Over-expressed miR-199a-3p inhibitedosteoblast proliferation, and down-regulated miR-199a-3p expression promoted osteoblast proliferation. miR-199a- 3p could reverse the FSS-induced proliferation in osteoblasts. Dual luciferase assay showed that miR-199a-3p targeted to CABLES-1 and over-expressed miR-199a-3p inhibited expression of CBALES-1 protein. CABLES-1 could promote proliferation of osteoblasts. miR-199a-3p inhibited osteoblast proliferation induced by FSS through CABLES-1. Conclusions FSS-induced osteoblast proliferation can be realized by down-regulated miR-199a-3p expression via targeting CABLES-1. The findings in this study provide new direction for researches on mechanism of FSS-induced osteoblast proliferation, as well as new ideas for future research on clinical application of mechanical loading in the treatment of bone and joint diseases.
ABSTRACT
Objective@#To investigate the role of miR-199a-3p in children with severe mycoplasmal pneumonia (MP) and to study its effect on Th17 cells.@*Methods@#Sixty children with severe MP (severe group), 50 children with recovery MP (recovery group) and 40 healthy children (control group) were enrolled. Venous blood samples were collected from all subjects, and the expression of miR-199a-3p in the peripheral blood leukocytes was detected by RT-PCR. The levels of IL-17a, IL-13, GM-CSF and IL-6 in serum were determined by enzyme-linked immunosorbent assay (ELISA). The effect of miR-199a-3p on the differentiation of Th17 cells was investigated by inducing differentiation of Th17 cells and transfecting miR-199a-3p mimics or inhibitors in vitro.@*Results@#Compared with the control group, the expression of miR-199a-3p in peripheral blood leukocytes of children with severe MP was significantly decreased (P<0.05). The levels of IL-17a, IL-13, GM-CSF and IL-6 in serum were significantly higher than those in the control group (all P<0.05). High expression of miR-199a-3p inhibited the differentiation of Th17 cells in vitro.@*Conclusions@#The expression of miR-199a-3p is decreased in the pathogenesis of MP. Th17 cells are the target cells of miR-199a-3p, and their differentiation is regulated by miR-199a-3p. The high expression of miR-199a-3p significantly inhibites the differentiation of Th17 cells.
ABSTRACT
Objective To investigate the role of miR-199a-3p in children with severe mycoplasmal pneumonia (MP) and to study its effect on Th17 cells. Methods Sixty children with severe MP (severe group), 50 children with recovery MP (recovery group) and 40 healthy children (control group) were enrolled. Venous blood samples were collected from all subjects, and the expression of miR-199a-3p in the peripheral blood leukocytes was detected by RT-PCR. The levels of IL-17a, IL-13, GM-CSF and IL-6 in serum were determined by enzyme-linked immunosorbent assay (ELISA). The effect of miR-199a-3p on the differentiation of Th17 cells was investigated by inducing differentiation of Th17 cells and transfecting miR-199a-3p mimics or inhibitors in vitro. Results Compared with the control group, the expression of miR-199a-3p in peripheral blood leukocytes of children with severe MP was significantly decreased (P<0.05). The levels of IL-17a, IL-13, GM-CSF and IL-6 in serum were significantly higher than those in the control group (all P<0.05). High expression of miR-199a-3p inhibited the differentiation of Th17 cells in vitro. Conclusions The expression of miR-199a-3p is decreased in the pathogenesis of MP. Th17 cells are the target cells of miR-199a-3p, and their differentiation is regulated by miR-199a-3p. The high expression of miR-199a-3p significantly inhibites the differentiation of Th17 cells.
ABSTRACT
@#【Objective】To investigate the role and the potential target of miR-199a-3p in mouse cardiac hypertrophy.【Methods】Neonatal mouse ventricular cardiomyocytes（NMVC）were isolated from the hearts of 0- 3- day- old newborn C57BL/6 mice. MiR-199a-3p mimic and retinoblastoma transcriptional corepressor 1（Rb-1）siRNA were transfected in? to NMVC to elevate the level of miR-199a-3p and inhibit Rb-1 expression，respectively. NMVC were stained with FITC- phalloidin solution to determine the size of NMVC. Dual luciferase reporter assay was performed to identify the interaction between miR- 199a- 3p and the 3’UTR of Rb- 1. mRNA and protein expression of cardiac hypertrophy associated genes were determined by RT-qPCR and Western blotting assay，respectively.【Results】（1）Over-expression of miR-199a-3pcould significantly enhance the expression of cardiac hypertrophy-related genes in NMVC ；（2）Dual-luciferase reporter assay results verified that miR- 199a-3p can interact with the 3’UTR of Rb-1. MiR-199a-3p could suppress Rb-1 ex? pression at the post-transcriptional level；（3）Functionally，miR-199a-3p mimic，consistent with Rb-1 siRNA，could increase cell size and the expression of Nppa，Acta1 and Myh7 in NMVC，and promote the nuclear translocation of E2f2 in NMVC.【Conclusions】MiR-199a-3p promotes the entry of E2f2 into the nucleus through inhibiting the expression of Rb-1，contributing to cardiomyocyte hypertrophy.
ABSTRACT
Objective To investigate the effect of miRNA-199a-3p overexpression on the expression of MAP3K4 protein in gastric cancer. Methods 35 gastric cancers and the matched adjacent tissue specimens were collected. Expression of miRNA-199a-3p and MAP3K4 were detected by stem-loop real-time reverse transcription polymerase chain reaction and western blot. Cell transfection was employed to explore the regulation of miRNA-199a-3p on MAP3K4 gene. Luciferase reporter assay was performed to identify target MAP3K4 gene. Results Com-pared with the adjacent tissue specimens ,miRNA-199a-3p was upregulated in the gastric cancers ,and MAP3K4 protein was down-regulated in the gastric cancers. Cells transfected with miR-199a-3p mimics showed lower MAP3K4 protein. MAP3K4 was identified as target gene of miR-199a-3p. Conclusions miRNA-199a-3p acts as an oncogene in gastric cancer and functions by targeting MAP3K4.
ABSTRACT
Objective To investigate the effect of miRNA-199a-3p overexpression on the expression of MAP3K4 protein in gastric cancer. Methods 35 gastric cancers and the matched adjacent tissue specimens were collected. Expression of miRNA-199a-3p and MAP3K4 were detected by stem-loop real-time reverse transcription polymerase chain reaction and western blot. Cell transfection was employed to explore the regulation of miRNA-199a-3p on MAP3K4 gene. Luciferase reporter assay was performed to identify target MAP3K4 gene. Results Com-pared with the adjacent tissue specimens ,miRNA-199a-3p was upregulated in the gastric cancers ,and MAP3K4 protein was down-regulated in the gastric cancers. Cells transfected with miR-199a-3p mimics showed lower MAP3K4 protein. MAP3K4 was identified as target gene of miR-199a-3p. Conclusions miRNA-199a-3p acts as an oncogene in gastric cancer and functions by targeting MAP3K4.
ABSTRACT
Background and purpose:Multiple microRNAs (miRNAs) are abnormally expressed in breast cancer and play an important role in the regulation of breast cancer. miRNAs may be a new target for the treatment of breast cancer. This study aimed to investigate the expression of miR-199a-3p in breast cancer and the effect of miR-199a-3p on proliferation and apoptosis of breast cancer.Methods:Real-time PCR was used to test the expression of miR-199a-3p in breast cancer tissues, normal breast tissues, breast cancer cells and normal breast cells. Overexpression (or silencing the expression) of miR-199a-3p was conducted by transfecting MDA-MB-231 with miR-199a-3p mimics (or inhibitors). The proliferation of MDA-MB-231 was detected by MTT method. The apoptosis of MDA-MB-231 was investigated by Hoechst staining and caspase-3 activity assay kit.Results:Compared to corresponding non-tumor breast tissues (or normal breast cell HBL-100), lower levels of miR-199a-3p were expressed in breast cancer tissues or breast cancer cells. Overexpression of miR-199a-3p induced by miR-199a-3p mimic inhibited the proliferation and promoted the apoptosis of MDA-MB-231, while silencing the expression of miR-199a-3p induced by miR-199a-3p in-hibitor increased the proliferation and suppressed the apoptosis of MDA-MB-231.Conclusion:The expression of miR-199a-3p is lower in breast cancer, which shows its tumor suppression effect by regulating the proliferation and apoptosis of breast cancer cells.
ABSTRACT
OBJECTIVE: To explore the effect of Aitongxiao granule (traditional Chinese medicines) therapy on miRNAs of replant liver cancer rats. METHODS: The 60 replant liver cancer rats were randomly divided into high dose(A), middle dose(B), lower dose(C), 5-Fu(D), distilled water(E) groups. After 14 days of experimental treatment, the expressions of miR-199a-3p,miR-199a-5p, miR-18 in liver cancer tissues were detected with qRT-PCR. RESULTS: Before experimental treatment, in comparison with E group, the level of miR-18 was higher but level of miR-199a-3p, miR-199a-5p were lower in A,B,C,D groups(P < 0.01). After 14 days of experimental treatment, in comparison with E group, the level of miR-18 was lower but level of miR-199a-3p, miR-199a-5p were higher in A,B,C,D groups(P < 0.01). The more significant differences were found in B group than that in D group. CONCLUSION: The Aitongxiao granule has the regulative effects on miRNAs of replant liver cancer rats. Its therapeutical effects relate to the miRNAs expressions of liver cellular cancer development.