MicroRNA-21 accelerates hepatocyte proliferation in vitro via PI3K/Akt signaling by targeting PTEN

MicroRNAs (miRNAs) are involved in controlling hepatocyte proliferation during liver regeneration. In this study, we established the miRNAs-expression patterns of primary hepatocytes in vitro under stimulation of epidermal growth factor (EGF), and found that microRNA-21 (miR-21) was appreciably up-regulated and peaked at 12 h. In addition, we further presented evidences indicating that miR-21 promotes primary hepatocyte proliferation through in vitro transfecting with miR-21 mimics or inhibitor. We further demonstrated that phosphatidylinositol 3′-OH kinase (PI3K)/Akt signaling was altered accordingly, it is, by targeting phosphatase and tensin homologue deleted on chromosome 10, PI3K/Akt signaling is activated by miR-21 to accelerate hepatocyte rapid S-phase entry and proliferation in vitro.     

Deoxycholyltaurine rescues human colon cancer cells from apoptosis by activating EGFR-dependent PI3K/Akt signaling

Recent studies indicate that secondary bile acids promote colon cancer cell proliferation but their role in maintaining cell survival has not been explored. We found that deoxycholyltaurine (DCT) markedly attenuated both unstimulated and TNF-alpha-stimulated programmed cell death in colon cancer cells by a phosphatidylinositol 3-kinase (PI3K)-dependent mechanism. To examine the role of bile acids and PI3K signaling in maintaining colon cancer cell survival, we explored the role of signaling downstream of bile acid-induced activation of the epidermal growth factor receptor (EGFR) in regulating both apoptosis and proliferation of HT-29 and H508 human colon cancer cells. DCT caused dose- and time-dependent Akt (Ser(473)) phosphorylation, a commonly used marker of activated PI3K/Akt signaling. Both EGFR kinase and PI3K inhibitors attenuated DCT-induced Akt phosphorylation and Akt activation, as demonstrated by reduced phosphorylation of a GSK-3-paramyosin substrate. Transfection of HT-29 cells with kinase-dead EGFR (K721M) reduced DCT-induced Akt phosphorylation. In HT-29 cells, EGFR and PI3K inhibitors as well as transfection with dominant negative AKT attenuated DCT-induced cell proliferation. DCT-induced PI3K/Akt activation resulted in downstream phosphorylation of GSK-3 (Ser(21/9)) and BAD (Ser(136)), and nuclear translocation (activation) of NF-kappaB, thereby confirming that DCT-induced activation of PI3K/Akt signaling regulates both proproliferative and prosurvival signals. Collectively, these results indicate that DCT-induced activation of post-EGFR PI3K/Akt signaling stimulates both colon cancer cell survival and proliferation.     

LEM domain containing 1 promotes proliferation via activating the PI3K/Akt signaling pathway in gastric cancer

Gastric cancer (GC) is one of the most common cancers worldwide and has especially high morbidity and mortality in China. LEM domain containing 1 (LEMD1), an important cancer-testis antigen, has been reported to be overexpressed in various cancers and promotes the progression of cancers. However, the biological characteristics of LEMD1 remain to be explored in GC. The connection between LEMD1 expression and GC progression was analyzed by using The Cancer Genome Atlas datasets and our human microarray datasets. A Kaplan-Meier plot was used to analyze the relationship between LEMD1 expression and prognosis. The expression of LEMD1 was analyzed by quantitative real-time polymerase chain reaction and Western blot, and the proliferation ability of GC cells was analyzed by cell proliferation and colony formation assays and 5-ethynyl-2′-deoxyuridine analysis. The cell cycle and apoptosis were analyzed by flow cytometry. Furthermore, subcutaneously implanted tumor models in nude mice were used to demonstrate the role of LEMD1 in promoting tumor proliferation in vivo. In this study, we demonstrated that the LEMD1 expression level was increased in GC tissues and cells compared with normal tissues and GES-1. The in vivo and in vitro assays showed that LEMD1 promoted GC cell proliferation by regulating the cell cycle and apoptosis. Moreover, we showed that LEMD1 regulated cell proliferation by activating the phosphatidylinositol 3 kinase (PI3K) / protein kinase B (AKT) signaling pathway. Overall, the results of our study suggest that LEMD1 contributes to GC proliferation by regulating the cell cycle and apoptosis via activation of the PI3K/AKT signaling pathway. LEMD1 may act as a potential target for GC treatment.     

Overexpression of fucosyltransferase IV promotes A431 cell proliferation through activating MAPK and PI3K/Akt signaling pathways

Lewis Y (LeY) is a carbohydrate tumor‐asssociated antigen. The majority of cancer cells derived from epithelial tissue express LeY type difucosylated oligosaccharide. Fucosyltransferase IV (FUT4) is an essential enzyme that catalyzes the synthesis of LeY oligosaccharide. Our previous studies have shown that FUT4 overexpression promotes A431 cell proliferation, but the mechanism is still largely unknown. Herein, we investigated the role of the mitogen‐activated protein kinases (MAPKs) and phosphoinositide‐3 kinase (PI3K)/Akt signaling pathways on FUT4‐induced cell proliferation. Results show that overexpression of FUT4 increases the phosphorylation of ERK1/2, p38 MAPK, and PI3K/Akt. Inhibitors of PI3K (LY294002 and Wortmannin) prevented the phosphorylation of ERK1/2, p38 MAPK, and Akt PI3K). Moreover, phosphorylation of Akt is abolished by inhibitors of ERK1/2 (PD98059) and p38 MAPK (SB203580). These data suggested that FUT4 not only activates MAPK and PI3K/Akt signals, but also promotes the crosstalk among these signaling pathways. In addition, FUT4‐induced stimulation of cell proliferation correlates with increased cell cycle progression by promoting cells into S‐phase. The mechanism involves in increased expression of cyclin D1, cyclin E, CDK 2, CDK 4, and pRb, and decreased level of cyclin‐dependent kinases inhibitors p21 and p27, which are blocked by the inhibitors of upstream signal molecules, MAPK and PI3K/Akt. In conclusion, these studies suggest that FUT4 regulates A431 cell growth through controlling cell cycle progression via MAPK and PI3K/Akt signaling pathways. J. Cell. Physiol. 225: 612–619, 2010. © 2010 Wiley‐Liss, Inc.     

PI3K/Akt信号传导通路与肿瘤

信号转导通路的异常激活是肿瘤细胞的发生、发展重要步骤,PI3K/Akt 信号通路在人类绝大多数恶性肿瘤中被异常激活,其在肿瘤的增殖、存活、细胞运动、抵抗凋亡、血管发生和转移以及对化疗耐药、放疗抗拒中发挥了重要作用.因此,通过对PI3K/Akt 通路的研究进一步了解肿瘤的发生、发展机制,并寻求抗肿瘤药物的新靶点,本文就 PI3K/Akt 信号转导通路的结构特点、与肿瘤发生、发展的关系及其时放化疗的影响作一综述.     

Erianin induces triple-negative breast cancer cells apoptosis by activating PI3K/Akt pathway

Background: Triple-negative breast cancer (TNBC) is a refractory subtype of breast cancer, 25–30% of which have dysregulation in the PI3K/AKT pathway. The present study investigated the anticancer effect of erianin on TNBC cell line and its underlying mechanism.Methods: After treatment with erianin, MTT assay was employed to determine the MDA-MB-231 and EFM-192A cell proliferation, the nucleus morphological changes were observed by DAPI staining. The cell cycle and apoptotic proportion were detected by flow cytometry. Western blot was performed to determine the cell cycle and apoptosis-related protein expression and PI3K pathways. Finally, the antiproliferative activity of erianin was further confirmed by adding or not adding PI3K agonists SC79.Results: Erianin inhibited the proliferation of MDA-MB-231 and EFM-192A cells in a dose-dependent manner, the IC₅₀ were 70.96 and 78.58 nM, respectively. Erianin could cause cell cycle arrest at the G₂/M phase, and the expressions of p21 and p27 were up-regulated, while the expressions of CDK1 and Cyclin B1 were down-regulated. Erianin also induced apoptosis via the mitochondrial pathway, with the up-regulation of the expression of Cyto C, PARP, Bax, active form of Caspase-3, and Caspase-9. Furthermore, p-PI3K and p-Akt expression were down-regulated by erianin. After co-incubation with SC79, the cell inhibition rate of erianin was decreased, which further confirmed that the attenuated PI3K/Akt pathway was relevant to the pro-apoptotic effect of erianin.Conclusions: Erianin can inhibit the proliferation of TNBC cells and induce cell cycle arrest and apoptosis, which may ascribe to the abolish the activation of the PI3K/Akt pathway.     

miR-188-5p regulates proliferation and invasion via PI3K/Akt/MMP-2/9 signaling in keloids

Keloids(KDs)and hypertrophic scars(HSs),two forms of pathological scars,seriously affect the physical and psychological health of patients.Despite many similarities with HSs,KDs are characterized by invasion and a high rate of recurrence after surgery,features they share in common with tumors.The underlying molecular mechanisms of this phenomenon have not been fully elucidated.In this study,we used microRNA(miRNA)array analysis to search for invasion-associated miRNAs in KDs.The expression of miR-188-5p in KDs,HSs,normal skin(NS)tissues,and cell lines was measured by quantitative real-time polymerase chain reaction.Furthermore,cell proliferation,migration,and invasion were detected in KD fibroblasts(KFs)and HS fibroblasts(HSFs),and interrelated proteins were ascertained by western blot analysis.It was found that miR-188-5p was significantly decreased in KD tissue compared with HS and NS tissues.Upregulated expression of miR-188-5p suppressed KF proliferation,migration,and invasion;and decreased expression of miR-188-5p also promoted HSF proliferation,migration,and invasion.The protein levels of MMP-2,MMP-9,PI3K,and p-Akt in miR-188-5p mimic-transfected KFs were repressed.In contrast,after transfection with miR-188-5p inhibitor,the protein levels of MMP-2,MMP-9,PI3K,and p-Akt were higher than the control in HSFs.Treatment with PI3K/Akt inhibitor LY294002 in KFs with miR-188-5p inhibitor did not further reduce their proliferation,migration,and invasion.The upregulation of MMP-2 and MMP-9 by miR-188-5p inhibitor could be abolished by LY294002.These findings together demonstrate a tumor-suppressive role of miR-188-5p in KD proliferation and invasion via PI3K/Akt/MMP-2/9 signaling,indicating that miR-188-5p may be a potential prognostic marker and therapeutic target for KDs.     

Follistatin could promote the proliferation of duck primary myoblasts by activating PI3K/Akt/mTOR signalling

FST (follistatin) is essential for skeletal muscle development, but the intracellular signalling networks that regulate FST-induced effects are not well defined. We sought to investigate whether FST promotes the proliferation of myoblasts through the PI3K (phosphoinositide 3-kinase)/Akt (protein kinase B)/mTOR (mammalian target of rapamycin) signalling. In the present study, we transfected the pEGFP-duFST plasmid and added PI3K and mTOR inhibitors to the medium of duck primary myoblasts. Then, we analysed the cellular phenotypic changes that occurred and analysed the expression of target genes. The results showed that FST promoted myoblast proliferation, induced the mRNA expression of PI3K, Akt, mTOR, 70-kDa ribosomal protein S6K (S6 kinase) and the protein expression of phospho-Akt (Thr³⁰⁸), mTOR, phospho-mTOR (serine 2448), phospho-S6K (Ser⁴¹⁷), inhibited the mRNA expression of FoxO1, MuRF1 (muscle RING finger-1) and the protein expression of phospho-FoxO1 (Ser²⁵⁶). Moreover, we found that the overexpression of FST could alleviate the inhibitory effect of myoblast proliferation caused by the addition of {"type":"entrez-nucleotide","attrs":{"text":"LY294002","term_id":"1257998346","term_text":"LY294002"}}LY294002, a PI3K inhibitor. Additionally, the overexpression of duck FST also relieved the inhibition of myoblast proliferation caused by the addition of rapamycin (an mTOR inhibitor) through PI3K/Akt/mTOR signalling. In light of the present results, we hypothesize that duck FST could promote myoblast proliferation, which is dependent on PI3K/Akt/mTOR signalling.     

PI3K/Akt信号通路与肺纤维化

肺纤维化(pulmonary fibrosis)是进行性、致命性的疾病。其致病机制不明,治疗效果差。PI3K/Akt信号通路主要与细胞的生长、增殖、分化、凋亡及血管形成等有关。近年来,随着对PI3K/Akt信号通路的深入研究,发现其活化后可激活下游中的一些因子参与肺纤维化,且与其他通路协同作用促进肺纤维化的形成。因此该通路有可能成为治疗肺纤维化的新靶点。将PI3K/Akt信号通路参与肺纤维化形成的研究进展作一综述。     

Downregulation of uPAR inhibits migration,invasion, proliferation,FAK/PI3K/Akt signaling and induces senescence in papillary thyroid carcinoma cells

Papillary thyroid carcinoma (PTC) is the most common endocrine and thyroid malignancy. The urokinase plasminogen activator receptor (uPAR) plays an important role in cancer pathogenesis, including breakdown of the extracellular matrix, invasion and metastasis. Additionally, there is increasing evidence that uPAR also promotes tumorigenesis via the modulation of multiple signaling pathways. BRAF^V600E, the most common initial genetic mutation in PTC, leads to ERK1/2 hyperphosphorylation, which has been shown in numerous cancers to induce uPAR. Treatment of the BRAF^V600E-positive PTC cell line, BCPAP, with the MEK/ERK inhibitor U0126 reduced uPAR RNA levels by 90%. siRNA-mediated downregulation of uPAR in BCPAP cells resulted in greatly decreased activity in the focal adhesion kinase (FAK)/phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway. This phenomenon was concurrent with drastically reduced proliferation rates and decreased clonigenic survival, as well as demonstrated senescence-associated nuclear morphology and induction of β-galactosidase activity. uPAR-knockdown BCPAP cells also displayed greatly reduced migration and invasion rates, as well as a complete loss of the cells'' ability to augment their invasiveness following plasminogen supplementation. Taken together, these data provide new evidence of a novel role for uPAR induction (as a consequence of constitutive ERK1/2 activation) as a central component in PTC pathogenesis, and highlight the potential of uPAR as a therapeutic target.Key words: urokinase plasminogen activator receptor (uPAR), papillary thyroid carcinoma, invasion, migration, proliferation, senescence, FAK, PI3K, Akt     

Gadolinium promoted proliferation in mouse embryo fibroblast NIH3T3 cells through Rac and PI3K/Akt signaling pathways

Nephrogenic systemic fibrosis (NSF) is a fibrosing disorder disease developed in patients with underlying renal insufficiency following exposure to gadolinium-based contrast agents (GBCAs). Previous studies have demonstrated that GdCl₃ can promote NIH3T3 fibroblast cell proliferation, which provide a new clue to the role of GBCAs in the development of NSF. In the present study, we further clarify the molecular mechanism of Gd-promoted proliferation. The results showed that intervention with the Rac inhibitor NSC23766 abrogated Gd-promoted proliferation. The levels of active Rac1 significantly increased in Gd-treated cells detected by pull-down assays. In addition, the phosphorylation of Akt was significantly elevated in the treatment group, which was blocked by NSC23766. NSC23766 also reduced the migration of NIH3T3 cells enhanced by Gd. Moreover, the F-actin cytoskeleton was strengthened and the mitotic cell numbers was significantly increased after exposure to Gd. These results suggest that Rac and PI3K/Akt signaling pathways, as well as integrin-mediated signal pathway may play important roles in Gd-induced cell proliferation. In addition, under serum-free condition, Gd could decrease ROS accumulation and increase NIH3T3 cell survival.     

Roxarsone induces angiogenesis via PI3K/Akt signaling

Background

3-Nitro-4-hydroxy phenyl arsenic acid, roxarsone, is widely used as an organic arsenic feed additive for livestock and poultry, which may increase the level of arsenic in the environment and the risk of exposure to arsenic in human. Little information is focused on the angiogenesis roxarsone-induced and its mechanism at present. This paper aims to study the role of PI3K/Akt signaling in roxarsone-induced angiogenesis in rat vascular endothelial cells and a mouse B16–F10 melanoma xenograft model.

Results

The results showed that treatment with 0.1–10.0 µmol/L roxarsone resulted in an increase in the OD rate in the MTT assay, the number of BrdU-positive cells in the proliferation assay, the migration distance in the scratch test and the number of meshes in tube formation assay. Further, treatment with 1.0 µmol/L roxarsone was associated with significantly higher phosphorylation of PI3K/Akt and expression of VEGF than the control treatment. The PI3K inhibitor was found to significantly combat the effects of 1.0 µmol/L roxarsone. Furthermore, roxarsone treatment was observed to increase the weight and volume of B16–F10 xenografts and VEGF expression and PI3K/Akt phosphorylation in a dose-dependent manner, with the 25 mg/kg dose having significant effects.

Conclusions

These results demonstrate that roxarsone has the ability to promote growth and tube formation in vascular endothelial cells and the growth of mouse B16–F10 xenografts. Further, the findings also indicate that PI3K/Akt signaling plays a regulatory role in roxarsone-induced angiogenesis in vivo and in vitro.

     

Low-intensity pulsed ultrasound promotes the proliferation of human bone mesenchymal stem cells by activating PI3K/AKt signaling pathways

Low-intensity pulsed ultrasound (LIPUS) is a promising therapy that is widely used in clinical applications and fundamental research. Previous research has shown that LIPUS exposure has a positive effect on stem cell proliferation. However, the impact of LIPUS exposure on human bone marrow mesenchymal stem cells (hBMSCs) remains unknown. In our study, the effect and mechanism of LIPUS exposure on the proliferation of hBMSCs were investigated, and the optimal parameters of LIPUS were determined. hBMSCs were obtained and identified by flow cytometry, and the proliferation of hBMSCs was measured using the Cell Counting Kit-8 assay to determine cell cycle and cell count. Expression levels of the phosphoinositide 3-kinase (PI3K)/protein kinase B (AKt) pathway proteins and cyclin D1 were determined by western blot analysis. Next, hBMSCs were successfully cultured and identified as multipotent mesenchymal stem cells. We found that LIPUS could promote the proliferation of hBMSCs when the exposure time was 5 or 10 minutes per day. Furthermore, 50 or 60 mW/cm² LIPUS had a more significant effect on cell proliferation, but if cells were irradiated by LIPUS for 20 minutes once a day, an intensity of at least 50 mW/cm² could markedly inhibit cell growth. Cell cycle analysis demonstrated that LIPUS treatment drives cells to enter S and G2/M phases from the G0/G1 phase. LIPUS exposure increased phosphorylation of PI3K/AKt and significantly upregulated expression of cyclin D1. However, these effects were inhibited when cells were treated with PI3K inhibitor (LY294002), which in turn reduced LIPUS-mediated proliferation of hBMSCs. These results suggest that LIPUS exposure may be involved in the proliferation of hBMSCs via activation of the PI3K/AKt signaling pathway and high expression of cyclin D1, and the intensity of 50 or 60 mW/cm² and exposure time of 5 minutes were determined to be the optimal parameters for LIPUS exposure.     

Knockdown of SRPX2 inhibits the proliferation,migration, and invasion of prostate cancer cells through the PI3K/Akt/mTOR signaling pathway

Sushi repeat‐containing protein X‐linked 2 (SRPX2), a novel chondroitin sulfate proteoglycan, is reported to play a critical role in tumorigenesis. However, the expression and functional role of SRPX2 in prostate cancer have not been defined. Thus, the aim of this study was to investigate the expression and functional role of SRPX2 in human prostate cancer. Our results showed that the expression of SRPX2 was obviously increased in human prostate cancer tissues and cell lines. In addition, knockdown of SRPX2 inhibited the proliferation, migration, and invasion of prostate cancer cells, as well as prevented the epithelial‐mesenchymal transition process in prostate cancer cells. Mechanically, knockdown of SRPX2 efficiently inhibited the activation of PI3K/Akt/mTOR pathway in prostate cancer cells. Taken together, these data demonstrated that knockdown of SRPX2 inhibits the proliferation and metastasis in human prostate cancer cells, partly through the PI3K/Akt/mTOR signaling pathway. Thus, SRPX2 may be a novel therapeutic target for the treatment of prostate cancer.     

miR-21 promotes proliferation and inhibits apoptosis of hepatic stellate cells through targeting PTEN/PI3K/AKT pathway

Abstract

To investigate the effect of microRNA 21 (miR-21) on hepatic stellate cells (HSCs) proliferation and apoptosis, and further to study its potential mechanisms. LX-2 cells were divided into miR-21 mimic group (Mimic), miR-21 mimic negative control group (NM), miR-21 inhibitor group (Inhibitor), miR-21 inhibitor negative control group (NC), and blank control group (Control). The cell proliferation was detected by CCK-8 assay and the cell migration and invasion were detected by scratch and transwell assay. Cell cycle and apoptosis were detected by flow cytometry. The levels of interleukin (IL)-6, tumor necrosis factor (TNF)-α, and transforming growth factor (TGF)-β1 were detected by enzyme-linked immunosorbent assay (ELISA). Proliferation, apoptosis, and phosphatase and tensin homolog (PTEN)/phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT) signaling pathway related genes and proteins were detected by quantitative real-time polymerase chain reaction (qRT-PCR) and western blot, respectively. The cells proliferation, migration, and invasion were promoted in Mimic group. The levels of IL-6, TNF-α, and TGF-β1 were increased after miR-21 administration. The expression of α-smooth muscle actin (SMA) and collagen 1 (Colla1) were increased, while Bax/B-cell lymphoma (Bcl)-2 ratio and programed cell death 4 (PDCD4) were reduced after miR?21 treatment. Meanwhile, the mRNA and protein expression of PTEN were reduced and PI3K/AKT pathway been promoted. Our study demonstrated that miR-21 could promote proliferation and inhibit apoptosis of HSCs, and its mechanism may be related to PTEN/PI3K/AKT pathway.     

TWEAK promotes hepatic stellate cell migration through activating EGFR/Src and PI3K/AKT pathways

Activated human hepatic stellate cells (HSCs) showed enhanced ability of migration compared with quiescent HSCs, which is pivotal in liver fibrogenesis. The aim of the present study was to investigate the effects of tumor necrosis factor‐like weak inducer of apoptosis (TWEAK) on the migration of activated HSCs and to explore the relevant potential mechanisms. Human HSCs LX‐2 cells were cultured with TWEAK. TNFRSF12A‐downexpressing lentiviruses were used to infect LX‐2 cells. The specific matrix metalloproteinases inhibitor BB94, the Src family kinase inhibitor, Dasatinib, and the specific inhibitor of phosphoinositide 3‐kinase (PI3K), LY294002 were used to treat LX‐2 cells combined with TWEAK. Cell migration and invasion was tested by the transwell assay. The expression of EGFR/Src, PI3K/AKT, and matrix metallopeptidase 9 (MMP9) was identified by real‐time polymerase chain reaction or western blotting. The result showed TWEAK promoted HSC migration and collagen production. BB94 significantly attenuated the migration of LX‐2 induced by TWEAK. Dasatinib inhibited the ability of cell migration stimulated by TWEAK. TWEAK upregulated the phosphorylation of epidermal growth factor receptor (EGFR) and Src. The phosphorylation of PI3K and AKT was significantly activated by TWEAK stimulation. Inhibition of PI3K/AKT reduced the expression of MMP9 induced by TWEAK. The present study, for the first time, demonstrated that TWEAK promoted HSC migration through the activation of EGFR/Src and PI3K/AKT pathways, and showed a novel potential mechanism of HSC migration regulated by TWEAK.