To investigate the interaction of miR-1290 and LHX6 in gliomas, and their influence on the propagation and metastasis of glioma cells. The differential expression of miR-1290 in glioma cells was identified by chip screening. The expression level of miR-1290 and LHX6 were determined by qRT-PCR and Western blot. The influence of miR-1290 on propagation of glioma cells were analyzed by MTT assay, EdU incorporation, and colony formation, while the impact of miR-1290 on metastasis was assessed by transwell assay. The relationship between LHX6 and miR-1290 was testified by luciferase reporter assay. The gliomas orthotopic implantation model of nude mice was established to investigate the influence of miR-1290 and LHX6 on tumor growth. Tumor volumes were evaluated by photon density, and the expression of Ki67 protein was analyzed by immunohistochemistry. MiR-1290 presented a higher expression in glioma cells and tissues. MiR-1290 overexpression significantly promoted propagation and metastasis of glioma cells, while miR-1290 knockdown inhibited glioma development. MiR-1290 suppressed LHX6 expression, facilitating development of glioma cells. The orthotopic implantation model showed that miR-1290 overexpression promoted tumor growth while LHX6 overexpression inhibited it. MiR-1290 could promote glioma cell propagation and metastasis by inhibiting LHX6.
The purpose of our study is to clarify the effect of microRNA-129-5p in the progression of human gastric cancer cells by regulating SPOCK1. The expression of microRNA-129-5p and SPOCK1 was tested by quantitative real-time polymerase chain reaction in tissues and cell lines. We validated the targeted relationship between microRNA-129-5p and SPOCK1 by dual luciferase reporter gene assay. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, colony formation, flow cytometry, transwell, and wound scratch assays were used to analyze the effects of microRNA-129-5p on SGC-7901 cell viability, proliferation, cell cycle and apoptosis, invasiveness, and migration. MicroRNA-129-5p was downregulated while SPOCK1 was upregulated in gastric cancer tissues and cell lines. The result of luciferase reporter gene assay demonstrated that microRNA-129-5p can target SPOCK1 by binding to the 3'untranslated region. The overexpression of microRNA-129-5p or the inhibition of SPOCK1 inhibited SGC-7901 viability, proliferation, migration, and invasion while promoted cell cycle arrest in G0/G1 stage and cell apoptosis. Our results suggested that microRNA-129-5p could directly specifically suppress SPOCK1, which might be one of the potential mechanisms in inhibiting cell processes including viability, proliferation, cell mitosis, migration, and invasiveness of gastric cancer cells.
We investigated the how miR-572 regulates PPP2R2C, and studied the effects of miR-572 and PPP2R2C on proliferation and migration as well as invasion of nasopharyngeal carcinoma (NPC) cells. NPC tissues and normal tissues were collected, and the expressions of miR-572 and PPP2R2C were detected by real-time PCR. Western blot was applied to detect the expression of PPP2R2C protein. The target relationship between miR-572 and PPP2R2C was confirmed by dual luciferase reporter gene assay. MTT assay and flow cytometry were applied to investigate the viability and apoptosis levels of NPC cells. Transwell as well as wound healing assays were used, respectively, to detect the invasiveness and migration of NPC cells. MiR-572 was highly expressed in NPC tissues as well as NPC cells, and there was lower expression of PPP2R2C in NPC tissues compared with normal samples. MiR-572 could bind to the 3' UTR of PPP2R2C and decrease its expression. Over-expressed miR-572 and decreased PPP2R2C expression could both inhibit proliferation and invasion and induce apoptosis of NPC cells. Thus, miR-572 promotes the proliferation and invasion of NPC by directly down-regulating PPP2R2C.
Hepatocyte growth factor (HGF) is an effective anti-fibrotic factor because of its bioactivity in inhibiting fibrosis-related proteins in the development of hepatic fibrosis. However, high-level production of bioactive mature form HGF is difficult because of its complex structure. Here, we report a non-fusion protein expression system to obtain truncated variant of N-terminal hairpin and first kringle domains of HGF (tvNK1) in Escherichia coli to determine its anti-fibrotic effects on hepatic stellate cells (HSCs). Under the selected conditions of cultivation and isopropyl-β-D-1-thiogalactopyranoside induction, the expression level of tvNK1 accounted for approximately 65 % of the total cellular protein and 50 % of fusion protein in the supernatant of whole cell lysates. The recombinant protein could be purified in one step with Ni(2+)-affinity chromatograph. Finally, about 65 mg recombinant tvNK1 was obtained from 1 l fermentation culture with no <95 % purity. In vitro, the final purified tvNK1 was shown to inhibit the proliferation of HSCs and decrease the mRNA and protein expression levels of fibrosis-related COL1A1 and α-smooth muscle actin genes.
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