Suppressive effect of platycodin D on bladder cancer through microRNA-129-5p-mediated PABPC1/PI3K/AKT axis inactivation

Platycodin D (PD) is a major constituent of Platycodon grandiflorum and has multiple functions in disease control. This study focused on the function of PD in bladder cancer cell behaviors and the molecules involved. First, we administered PD to the bladder cancer cell lines T24 and 5637 and the human uroepithelial cell line SV-HUC-1. Cell viability and growth were evaluated using MTT, EdU, and colony formation assays, and cell apoptosis was determined using Hoechst 33342 staining and flow cytometry. The microRNAs (miRNAs) showing differential expression in cells before and after PD treatment were screened. Moreover, we altered the expression of miR-129-5p and PABPC1 to identify their functions in bladder cancer progression. We found that PD specifically inhibited the proliferation and promoted the apoptosis of bladder cancer cells; miR-129-5p was found to be partially responsible for the cancer-inhibiting properties of PD. PABPC1, a direct target of miR-129-5p, was abundantly expressed in T24 and 5637 cell lines and promoted cell proliferation and suppressed cell apoptosis. In addition, PABPC1 promoted the phosphorylation of PI3K and AKT in bladder cancer cells. Altogether, PD had a concentration-dependent suppressive effect on bladder cancer cell growth and was involved in the upregulation of miR-129-5p and the subsequent inhibition of PABPC1 and inactivation of PI3K/AKT signaling.     

MircoRNA-129-5p suppresses the development of glioma by targeting HOXC10

BackgroundmiR-129-5p has been reported to be abnormally expressed and plays an important role in the progression of various malignancies. However, its role in gliomas and its exact molecular mechanism need further research.Methods and materialsRT-qPCR was performed to evaluate miR-129-5p and HOXC10 mRNA expression levels in tissues and cell lines. Cell proliferation was detected via Cell Counting Kit-8 (CCK-8), 5-ethynyl-2′-deoxyuridine (EdU) and clone formation assays. Luciferase assays were used to validate the binding of seeds between miR-129-5p and HOXC10. A tumor xenograft model was developed to study the effect of miR‐129-5p on glioma growth in vivo.ResultsmiR-129-5p was expressed at low levels in glioma tissues and cell lines. miR-129-5p overexpression inhibited glioma proliferation, migration and invasion. miR-129-5p negatively and directly targeted HOXC10. At the same time, HOXC10 was upregulated in glioma cancer, and HOXC10 knockdown inhibited cell proliferation, migration and invasion.ConclusionmiR-129-5p inhibits glioma development by altering HOXC10 expression and may therefore serve as a new diagnostic marker and therapeutic target for glioma in the future.     

miR-296-5p 靶向 PLK1 调控 PI3K/ AKT 通路诱导骨肉瘤细胞中的自噬并抑制上皮-间质转化

目的 探讨 miR-296-5p 靶向 PLK1 对骨肉瘤 (osteosarcoma, OS) 细胞自噬及抑制上皮-间质转化 (EMT) 的作用机制。 方法 qRT-PCR 检测 miR-296-5p 在 OS 细胞中的表达。 采用生物信息学分析预测 miR-296-5p 的靶基因, 验证 miR-296-5p 对靶基因 PLK1 的直接靶向调控; 细胞转染构建 miR-296-5p 过表达 和干扰细胞, CCK-8、 克隆形成、 Transwell 小室、 流式、 蛋白免疫印迹实验检测 miR-296-5p 的不同表达对 U2OS 细胞中 PTBP1 表达水平及细胞增殖、 侵袭、 凋亡、 自噬及 EMT 的影响。 结果 与对照组比较, miR-296-5p 在 OS 中表达降低, 而 PLK1 则升高 (P< 0. 05); 与 miR-NC 组比较, mimic 组的克隆形成率、 侵袭 细胞数目及 PTBP1、 p62、 N-cadherin、 Vimentin、 p-PI3K/ PI3K、 p-AKT/ AKT 水平降低, 细胞凋亡率、 Beclin-1、 LC3-Ⅱ/ Ⅰ、 E-cadherin 水平升高 (P< 0. 05); 与 PLK1 组比较, PLK1 + mimic 组的克隆形成率、 侵袭 细胞数目及 PTBP1、 p62、 N-cadherin、 Vimentin、 p-PI3K/ PI3K、 p-AKT/ AKT 水平降低, 细胞凋亡率、 Beclin-1、 LC3-Ⅱ/ Ⅰ、 E-cadherin 水平升高 (P< 0. 05)。 结论 miR-296-5p 可能能够靶向 PLK1 调控 PI3K/ AKT 通路诱导 OS 细胞中的自噬并抑制 EMT。     

Mutational and Immunohistochemical Study of the PI3K/Akt Pathway in Papillary Thyroid Carcinoma in Greece

PI3K/Akt signaling pathway plays critical role in many cell processes. There is indication that enhanced activation of PI3K/Akt cascade is implicated in thyroid tumors. Aim of this study was to evaluate the mutational status and expression of PI3K/Akt pathway mediators in papillary thyroid carcinoma in Greece. We evaluated the presence of mutations in PIK3CA (exons 9 and 20), AKT1 (exons 6–11), AKT2 (exons 6–11), AKT3 (exons 5–10), PTEN (exons 3–8), and PDPK1 (exons 4–10) genes in 83 papillary thyroid carcinomas by DNA sequencing. The expression levels of phospho-Akt and insulin-like growth factor I receptor (IGF-IR) were evaluated by immunohistochemistry. PIK3CA mutations were found in three samples. The analysis of AKT1 revealed one silent mutation in exon 9 (G726A) in 16 samples. One specimen carried an AKT3 mutation. One missense mutation was found in one sample in PTEN. No mutations were found in AKT2 and PDPK1. Increased levels of phosphorylated total Akt and IGF-IR were identified in some papillary cancers. Our findings indicate that PI3K/Akt signaling pathway is activated in some papillary tumors. However, mutations in genes coding most mediators of the pathway have not been proven to be the major modus of enhanced activation. These data suggest a potential role for PI3K/Akt-mediated signaling in papillary thyroid tumors.     

miR-218 modulate hepatocellular carcinoma cell proliferation through PTEN/AKT/PI3K pathway and HoxA10

Purpose: To investigate the regulatory mechanism of miR-218 in human hepatocellular carcinoma (HCC). Methods: qPCR was used to compare the expression levels miR-218 among six hepatocellular carcinoma cell lines and normal liver tissues. After transfecting MHCC97L cells with either miR-218 mimics or miR-218 inhibitor, western blotting was used to examine the expressing patterns of cyclinD1, p21, and PTEN/AKT/PI3K signaling pathway-related proteins. MTT and colony forming assay was used to assess the capability of cell proliferation. Bioinformatic method was applied to predict the binding of miR-218 on HoxA10, and western blotting was used to examine the modulatory effect of miR-218 AND HoxA10 on PTEN/AKT/PI3K pathway in HCC. Results: The expression levels of miR-218 were frequently lower in HCC cell lines than in normal liver tissues. Over-expression of miR-218 in HCC cells significantly decreased cell proliferation whereas inhibiting miR-218 promoted cancer cell proliferation. Western blotting analysis demonstrated that tumorigenesis related protein cyclin D1 and p21, as well as PTEN/AKT/PI3K signaling pathways were actively modulated by miR-218 in HCC cells. The expression of endogenous HoxA10 was also down-regulated by miR-218 over-expression, and silencing HoxA10 directly activated PTEN in HCC cells. Conclusion: Modulation of miR-218 actively affected HCC cancer cell development. The regulatory mechanism of miR-218 in HCC cells was acting through PTEN/AKT/PI3K pathway and possibly associated with HoxA10.     

LncRNA TMPO-AS1 up-regulates the expression of HIF-1α and promotes the malignant phenotypes of retinoblastoma cells via sponging miR-199a-5p

BackgroundLong non-coding RNA (lncRNA) TMPO antisense RNA 1 (TMPO-AS1) is reported to be oncogenic in prostate cancer and lung cancer. This study aims to investigate the expression and biological function of it in retinoblastoma (RB), and explore its regulatory role for miR-199a-5p and hypoxia-inducible factor-1α (HIF-1α).MethodsPaired RB samples were collected, and the expression levels of TMPO-AS1, miR-199a-5p and HIF-1α were examined by quantitative real-time polymerase chain reaction (qRT-PCR); TMPO-AS1 overexpressing plasmids and TMPO-AS1 shRNA were transfected into HXO-RB44 and SO-Rb50 cell lines respectively, and then proliferation, migration and invasion of RB cells were detected by CCK-8 assay and Transwell method. qRT-PCR and western blot were used to analyze the regulatory function of TMPO-AS1 on miR-199a-5p and HIF-1α; luciferase reporter gene assay was used to determine the regulatory relationship between miR-199a-5p and TMPO-AS1.ResultsTMPO-AS1 was significantly up-regulated in cancerous tissues of RB samples (relatively expression: 2.97 vs 3.93, p < 0.001), negatively correlated with miR-199a-5p (r=-0.4813, p < 0.01). There was one binding site on TMPO-AS1 for miR-199a-5p. After transfection of TMPO-AS1 shRNAs into RB cells, the proliferation, migration and invasion of cancer cells was significantly inhibited, while TMPO-AS1 had opposite effects; TMPO-AS1 was also demonstrated to regulate the expression of HIF-1α on both mRNA and protein levels via negatively regulating miR-199a-5p.ConclusionTMPO-AS1 is abnormally up-regulated in RB tissues, and it can modulate the proliferation and migration of RB cells. It has the potential to be the “ceRNA” to regulate HIF-1α expression by sponging miR-199a-5p.     

下调miR-21 抑制PI3K / AKT 信号通路对白血病细胞增殖凋亡的影响

目的:探讨miR-21 对白血病K562 细胞增殖凋亡的影响及对PI3K/ AKT 信号通路的调控作用。方法:将K562 细胞分为对照组、miR-21 NC 组和miR-21 干扰组,对照组不做处理,后两组采用阳离子脂质体LipofectamineTM2000 转染miR-21 inhibitor 和miR-21 negative control。转染48 h 后,利用实时荧光定量(qRT-PCR)检测各组细胞中miR-21 mRNA 的表达情况,采用MTT 比色法检测miR-21 对细胞增殖率的影响,流式细胞仪检测miR-21 对细胞周期和凋亡率的影响,蛋白质免疫印迹法(Western blot)检测miR-21 对各组细胞中PI3K/ AKT 信号通路相关蛋白PI3K、AKT 和p-AKT 表达的影响。结果:miR-21 干扰组中细胞的miR-21 mRNA 表达水平和细胞的存活率较对照组和miR-21 NC 组均明显降低;与对照组和miR-21 NC 组比较,流式细胞仪检测miR-3 干扰组中G0/ G1 期所占细胞比例明显升高,S 期细胞所占比例明显下降,细胞的凋亡率明显升高。Western blot 检测p-AKT 的表达水平较对照组和miR-21 NC 组明显下降,但PI3K 和AKT 蛋白的表达水平变化不大。结论:下调miR-21 能够抑制白血病K562 细胞的增殖,促进其凋亡,其作用机制可以与抑制PI3K/ AKT 信号通路有关。     

miR-141-3p promotes proliferation and metastasis of nasopharyngeal carcinoma by targeting NME1

PurposeThis study aimed to investigate the expression and biological function of miR-141-3p in nasopharyngeal carcinoma (NPC) via targeting neoplasm metastasis 1 (NME1).Materials and methodsThe expression of miR-141-3p and NME1 in 5-8F, C666-1, CNE-1, CNE-2, 6-10B and NP69 nasopharyngeal epithelial cells were detected using real-time Polymerase Chain Reaction (real-time PCR) and western blot, respectively. Cell proliferation was detected using Cell Counting Kit-8 (CCK-8), and the metastasis was detected using Transwell. The binding of miR-141-3p to NME1 was detected by dual luciferase reporter gene detection system. The effects of miR-141-3p on tumor growth were also determined in vivo.ResultsThe results showed that the expression of miR-141-3p significantly increased in various tumor cell lines and the expression of NME1 was higher in NP69 cells and lower in 5-8F cells, which had significant negative correlation. Furthermore, the expression of NME1 was significantly reduced after transfection of miR-141-3p and miR-141-3p promoted cell proliferation and metastasis. The double luciferase reporter gene detection system confirmed that NME1 was the target gene of miR-141-3p. Knockout of NME1 promoted the proliferation and metastasis of NP69 or 6-10B cells and the activation of p-Akt, which were abrogated by miR-141-3p. In vivo, the tumor volumes and weights in the miR-141-3p group significantly increased followed by down-regulation of NME1 and activation of p-Akt.ConclusionsWe confirmed that miR-141-3p promotes the proliferation and metastasis of NPC by targeting NME1.     

LncRNA DSCAM-AS1通过靶向miR-627-3p调控皮肤鳞状细胞癌增殖、侵袭和迁移

目的 探讨长链非编码RNA(LncRNA)唐氏综合征细胞粘附分子反义1(DSCAM-AS1)靶向miR-627-3p对人皮肤鳞状细胞癌增殖和侵袭能力的影响和分子机制.方法 采用实时荧光定量PCR(RT-qPCR)检测DSCAM-AS1和miR-627-3p在人永生化表皮细胞HaCaT和3种皮肤鳞状细胞癌细胞(SCC13...     

5,7,2,5-tetrahydroxy-8,6-dimethoxyflavone up-regulates miR-145 expression and inhibits proliferation of gastric cancer cells

IntroductionGastric cancer is a frequently detected malignancy, and its incidence has increased over the past decades in East Asia. The present study investigated the effect of 5,7,2, 5-tetrahydroxy-8,6-dimethoxyflavone (THDMF) on gastric cancer cells and explored the underlying mechanism. The study analysed cell viability changes, apoptotic features, and metastasis potential of treatment with THDMF.Material and methodsMTT colorimetric assay was used for measurement of MKN28, MKN45, and GES-1 cell proliferation and flow cytometry for the detection of apoptosis. Transwell and wound healing assays were used to observe the invasion and migration abilities of MKN28 cells. The expression of p21, MMP2/-9, PI3K, and c-Myc proteins was detected by western blotting.ResultsThe THDMF treatment significantly (p < 0.05) reduced MKN28 and MKN45 cell proliferation without changing GES-1 cell viability. A significant increase in apoptotic cell population on treatment with THDMF was observed. Treatment of MKN28 cells with THDMF increased the percentage of cells in the G1 phase. Exposure of MKN28 cells to THDMF caused a marked decrease in invasion and migration potential in comparison to control cells. The expression of miR-145 was markedly increased in MKN28 cells on treatment with THDMF. In MKN28 cells expression of c-Myc, PI3K, p-AKT, MMP-2, and MMP-9 was suppressed markedly on exposure to THDMF. The expression of p21 protein in MKN28 cells was markedly promoted on exposure to THDMF.ConclusionsTHDMF exhibits anti-cancer effect on gastric cancer cells in vitro by activation of cell apoptosis and arrest of cell cycle. In addition, THDMF promoted miR-145 expression and down-regulation of PI3K/AKT signalling pathway in MKN28 cells. Therefore, THDMF may be utilised as a potential novel therapeutic agent for the treatment of gastric cancer.     

DICER activates autophagy and promotes cisplatin resistance in non-small cell lung cancer by binding with let-7i-5p

ObjectiveDrug resistance is the main obstacle in the treatment of non-small cell lung cancer (NSCLC). This study aimed to explore the mechanism of DICER in NSCLC resistance and its downstream signaling pathways.MethodsThe A549 cisplatin (DDP)-resistant strain A549/DDP was established. A549/DDP cells were transfected with DICER- and let-7i-5p-related vectors, and treated with autophagy activator rapamycin. The cell viability and apoptosis were tested by CCK-8 assay and flow cytometry, respectively. The formation of autophagosomes was observed with a transmission electron microscopy. RT-qPCR and Western blot assay were conducted to detect expression levels of DICER, let-7i-5p, autophagy-related proteins, and the PI3K/AKT/mTOR pathway-related proteins. The dual luciferase reporter gene assay was implemented to confirm the targeted binding of DICER and let-7i-5p.ResultsDICER was highly expressed in DDP-resistant NSCLC tissues and cells, and DICER could target and negatively regulate the expression of let-7i-5p. DDP treatment could inhibit the viability and promote cell apoptosis of A549/DDP cells. Downregulation of DICER in A549/DDP cells exhibited a decrease of cell viability, a decreased ratio of LC3-II/LC3-I and autophagosomes, together with an elevation of cell apoptosis rate and the phosphorylation levels of PI3K/AKT/mTOR. Treatment of rapamycin and let-7i-5p inhibitor reversed the effects of downregulated DICER in cell viability, ratio of LC3-II/LC3-I, autophagosomes, cell apoptosis rate and the phosphorylation levels of PI3K/AKT/mTOR in A549/DDP cells.ConclusionOur research suggests that DICER promotes autophagy and DDP resistance in NSCLC through downregulating let-7i-5p, and inhibits the activation of PI3K/AKT/mTOR pathway.     

MicroRNA-145通过丝裂原活化蛋白激酶和磷脂酰肌醇3激酶/蛋白质丝氨酸苏氨酸激酶通路调控肺癌A549细胞系转移及侵袭

目的 探讨microRNA-145(miR-145)对非小细胞肺癌A549细胞转移、侵袭及对丝裂原活化蛋白激酶(MAPK)和磷脂酰肌醇3激酶/蛋白质丝氨酸苏氨酸激酶（PI3K/AKT）通路的作用。 方法 将非小细胞肺癌A549细胞分成miR-145模拟物（mimics）组和negative-mimics组（miR-NC）以及antago miR-145组（抑制剂组）和antago miR control 组（antago-NC）,采用Transwell迁移实验及基质胶侵袭实验等检测miR-145对人非小细胞肺癌A549迁移、侵袭能力的影响;Western blotting方法分析miR-145对MAPK和PI3K/AKT通路的影响。此外,采用细胞外调节蛋白激酶（ERK）及AKT的通路抑制剂分别作用于A549细胞系,检测A549细胞迁移、侵袭能力的改变。 结果 miR-145 mimics组穿过细胞数（90.67±10.33）明显少于miR-NC组（175.33±23.67）,miR-145 mimics组穿过基质胶的细胞数（153.33±22.33）少于miR-NC组 (77.33±13.67）,P<0.05;antago-NC组通过小室的细胞数量以及穿过基质胶的细胞数量明显少于antago miR-145组（P<0.05）,结果说明,miR-145具有抑制非小细胞肺癌A549细胞迁移、侵袭的能力;miR-145 mimics转染可分别抑制A549细胞中90%、78%以及73%的ERK1/2、AKT的ser-473位点和thr-308位点的磷酸化,antago miR-145转染可促进A549细胞中ERK1/2、AKT的ser-473位点和thr-308位点的磷酸化,增加115%、125%以及129%,而当抑制MAPK通路及PI3K/AKT通路的激活后,A549细胞的转移及侵袭能力下降。 结论 miR-145通过MAPK和PI3K/AKT通路调控肺癌A549细胞转移及侵袭。     

Long non-coding RNA LINC01296 promotes progression of oral squamous cell carcinoma through activating the MAPK/ERK signaling pathway via the miR-485-5p/PAK4 axis

IntroductionLong intergenic non-protein-coding RNA 1296 (LINC01296), a newly identified lncRNA, can function as an oncogenic driver to promote the development of multiple carcinomas. However, the effect of LINC01296 on oral squamous cell carcinoma (OSCC) is still unclear.Material and methodsWe determined the expression and role of LINC01296 in OSCC tissues and cell lines. The cell viability, migration and invasion were determined by MTT, wound healing assay and transwell assay, respectively. Flow cytometry was used for detecting cell cycle and apoptosis. The interaction and association between LINC01296, microRNA-485-5p (miR-485-5p) and p21 (RAC1) activated kinase 4 (PAK4) were analyzed by RNA immunoprecipitation (RIP) and luciferase reporter assays. The xenograft mouse model was established to detect the effect of LINC01296 on OSCC tumor growth.ResultsOur study showed that LINC01296 was over-expressed in OSCC tissues and cell lines. The level of LINC01296 was positively correlated with the patient’s tumor node metastasis (TNM) stage and nodal invasion. Knockdown of LINC01296 effectively inhibits cell viability, migration and invasion but promotes cell apoptosis in vitro. The in vivo experiment showed that LINC01296 knockdown inhibited OSCC tumor growth. The following analysis indicated that LINC01296 acted as a ceRNA for miR-485-5p, and PAK4 was identified as a direct target of miR-485-5p. Furthermore, we found that the effects of LINC01296 on OSCC progression were through regulating the expression of PAK4/p-MEK/p-ERK via sponging miR-485-5p.ConclusionsLINC01296 promote the cell cycle, proliferation, migration and invasion, and inhibit apoptosis of OSCC cells through activating the MAPK/ERK signaling pathway via sponging miR-485-5p to regulate PAK4 expression. These results suggested that the LINC01296/miR-485-5p/PAK4 axis was closely associated with OSCC progression. Our study provides a new insight into the molecular pathogenesis of OSCC, and may supply novel biomarkers for diagnosis and therapy of OSCC.     

lncRNAPVT1 targets miR-152 to enhance chemoresistance of osteosarcoma to gemcitabine through activating c-MET/PI3K/AKT pathway

Background: LncRNA PVT1 has been reported to be involved in a variety of biological processes, including cell proliferation, cell differentiation and cancer progression. However, the mechanism by which LncRNA PVT1 contributes to chemoresistance of osteosarcoma cell, has not been fully elucidated.Methods: We first generatedLncRNA PVT1-overexpressed MG63 cells and LncRNA PVT1 knockdown MG63/DOX cells. Then, we examined the effect of LncRNA PVT1 on cell viability and colony formation ability by MTT assay and soft agar assay, respectively. In addition, we performed flow cytometry analysis to detect apoptosis induced by GEM. Dual luciferase reporter assay and RIP were used to confirmed the interaction between LncRNA PVT1 and miR-152. Finally, we determined protein level of c-MET, p-PI3K, and p-AKT by westernblot.Results: LncRNA PVT1 overexpression promoted cell proliferation and exhibited the anti-apoptotic property in LncRNA PVT1-overexpressing MG63 cells treated with gemcitabine. While, LncRNA PVT1-depleted MG63/DOX cells treated with gemcitabine exhibited significant lower survival rate and high percentage of apoptosis. Next, we found that LncRNA PVT1 could target and downregulated the level of miR-152. Interestingly, miR-152 greatly rescued the biological outcomes of LncRNA PVT1 not only in MG63 but also in MG63/DOX cells. We observed that LncRNA PVT1 markedly induced PI3K/AKT pathway activation, which was abolished by miR-152 mimics overexpression. Finally, c-MET inhibitor was used to confirm the essential role of c-MET in LncRNA PVT1 and miR-152-regulated PI3K/AKT signaling.Conclusion: We showed thatlncRNA PVT1 played a contributory role in chemoresistance of osteosarcoma cells through c-MET/PI3K/AKT pathway activation, which was largely dependent on miR-152. Our findings advance our understanding of how lncRNA PVT1 promotes chemoresistance of osteosarcoma cells and facilitate development of novel strategies for treating osteosarcoma.     

Tumor necrosis factor-α promotes Staphylococcus aureus-induced osteomyelitis through downregulating endothelial nitric oxide synthase

BackgroundInfections of Staphylococcus aureus (S. aureus) often result in osteomyelitis, which is the acute or chronic infections of the bone marrow or bones. TNF-α is long recognized as a key factor contributing to the pathogenesis of osteomyelitis, but little is known about the underlying molecular mechanism.MethodsExpression levels of TNF-α, and several candidate genes, including endothelial nitric oxide synthase (eNOS), known to be downregulated by TNF-α were analysed in MC3T3-E1 cells with S. aureus infection and osteomyelitis patient blood. MicroRNA(miR)-129-5p was predicted and experimentally verified to target eNOS. Alizarin red sulfate (ARS) and alkaline phosphatase (ALP) staining assays were conducted on MC3T3-E1 cells with S. aureus infection to assess the role of TNF-α/miR-129-5p/eNOS on mineralization defect.ResultsTNF-α and miR-129-5p were upregulated while eNOS was downregulated in MC3T3-E1 cells with S. aureus infection and osteomyelitis patients, showing inversely correlated expression profiles. MiR-129-5p directly binds to the 3′-UTR of eNOS mRNA to suppress eNOS expression in MC3T3-E1 cells. TNF-α blocker inhibited miR-129-5p and elevated eNOS expression, likely contributing to rescued mineralization defect in S. aureus-infected MC3T3-E1 cells. During S. aureus infection, upregulated TNF-α increases endogenous miR-129-5p expression, which in turn inhibits eNOS, contributing to osteomyelitis.ConclusionOur study thereby proposes a novel signalling cascade involving TNF-α/miR-129-5p/eNOS in the pathogenesis of osteomyelitis, which may also serve as therapeutic targets.     

Ginsenoside Rh2 inhibits thyroid cancer cell migration and proliferation via activation of miR-524-5p

IntroductionThyroid cancer is an important disease that threatens the health of humans. Ginsenoside Rh2 is known as an anticancer molecule; however, its function in thyroid cancer cells has not been reported. In the present study, we identified that Rh2 treatment of the thyroid cancer cell line K1 inhibited cell migration and proliferation.Material and methodsWe determined the Rh2 function in thyroid cancer cell lines. By RT-PCR, expression of miR-524-5p and related genes were determined. The cell phenotype including cell migration and proliferation were detected after serials treatment. The relevant protein level were checked by Western blot.ResultsInterestingly, we observed that miR-524-5p, a type of miRNA, had lower expression in the thyroid cancer cell lines TPC-1, K1, and NPA than in the normal thyroid cell line Nthyri3-1. Additionally, Rh2 treatment induced miR-524-5p expression. Further examination using overexpression of miR-524-5p identified that the miR-524-5p mimic inhibited cell migration and proliferation of the K1 line. Similar to Rh2-treated cells, the miR-524-5p mimic-expressing cells had increased E-cadherin and reduced vimentin levels compared to the control cells. Next, we examined the relationship between Rh2 and miR-524-5p with respect to thyroid cell migration and proliferation. Treatment with Rh2 and miR-524-5p inhibitor suppressed Rh2 action on K1 thyroid cell migration and proliferation, and the rates were similar to those in control cells, suggesting that Rh2 might induce miR-524-5p expression to inhibit thyroid cancer cell migration and proliferation.ConclusionsOur analyses identified Rh2 and miR-524-5p action on thyroid cancer cell migration and proliferation as well as the linkage between Rh2 and miR-524-5p in thyroid cancer cell development.     

MicroRNA-146b-5p suppresses cholangiocarcinoma cells by targeting TRAF6 and modulating p53 translocation

BackgroundIn view of the poor prognosis and high mortality of cholangiocarcinoma, there is a need for new therapeutic strategies. This study aims to reveal the biological function of miR-146b-5p in cholangiocarcinoma cell and its possible mechanism.MethodsThe expression level and prognostic information on miR-146b-5p in cholangiocarcinoma were obtained in TCGA database. The biological function of miR-146b-5p on proliferation and vitality of cholangiocarcinoma cell HUCCT-1 was examined by EdU and MTT assay, and the apoptosis of HUCCT-1 cells transfected with miR-146b-5p mimic, mimic control, inhibitor, inhibitor control was detected by flow cytometry analysis. The western blot was done to evaluate the effect of miR-146b-5p targeting substrate and the expression of p53 in whole-cell protein and mitochondria fractions.ResultsOur finding revealed that miR-146b-5p expression in patients with CHOL was lower than the normal group(p＜0.001). MiR-146b-5p expression was down-regulated in human cholangiocarcinoma HUCCT-1 and RBE cells compared to normal control HIBEC and other cancer cells. The miR-146b-5p mimic could inhibit HUCCT-1 cell proliferation (p＜0.05) and promote HUCCT-1 cell apoptosis significantly (p＜0.05). The results of western blot showed that miR-146b-5p mimic could directly target TRAF6 3′UTR region and up-regulate the expression of p53 in mitochondria and miR-146b-5p inhibitor could down-regulated the level of p53 in mitochondria.ConclusionMiR-146b-5p is a cholangiocarcinoma suppressor by inhibiting cell proliferation and promoting cell apoptosis with targeting TRAF6, possibly via modulating p53 translocation to mitochondria.     

CAPG facilitates diffuse large B-cell lymphoma cell progression through PI3K/AKT signaling pathway

ObjectiveDiffuse large B-cell lymphoma (DLBCL) is a genetically heterogeneous tumor. Currently, macrophage-capping protein (CAPG) has been discovered to play a crucial part as a regulator in various cancers. However, it still remains unclear regarding its regulatory mechanism in DLBCL. Therefore, this study focused on revealing the mechanism underlying CAPG in DLBCL.MethodsThe bioinformatics analysis was conducted to predict the expression of CAPG in DLBCL and its downstream target genes. qRT-PCR was utilized to detect mRNA expression levels of CAPG and CEBPA. Western blot was performed to examine the expression levels of related proteins. Then we employed flow cytometry for the analysis of cell cycle and apoptosis. We also used MTT assay to measure cell survival and IHC assay to detect CAPG expression in DLBLC tissues. Then, ChIP was used to determine the binding of CEBPA and CAPG. For in vivo experiments, xenograft models were employed to investigate the effect of CAPG on DLBCL.ResultsHigh-level of CAPG expression was found in DLBCL cells and tissues. CAPG could strengthen the proliferative and invasive abilities of DLBCL cells, inhibit cell apoptosis, and activate PI3K/AKT signaling pathway. CAPG overexpression accelerated malignant progression of DLBCL cells. In addition, CAPG expression was regulated by CEBPA.ConclusionCAPG enhances the proliferation and invasion of DLBCL cells by promoting PI3K/AKT signaling pathway, which is expected to be a promising target for DLBCL.