LncRNA MEG3靶向miR-181a-5p调控宫颈癌细胞放射敏感性

目的 研究LncRNA MEG3对宫颈癌细胞放射敏感性的影响,并探讨其作用机制。方法 运用qRT-PCR法检测放射抗性和放射敏感性宫颈癌细胞中LncRNA MEG3的表达;将过表达对照组(转染pcDNA 3.1)、过表达LncRNA MEG3组(转染pcDNA 3.1-LncRNA MEG3)、抑制miR-NC组(转染anti-miR-NC)、抑制miR-181a-5p组(转染anti-miR-181a-5p)、过表达LncRNA MEG3+过表达miR-NC组(共转染pcDNA 3.1-LncRNA MEG3和anti-miR-NC)、过表达LncRNA MEG3+过表达miR-181a-5p组(共转染pcDNA 3.1-LncRNA MEG3和anti-miR-181a-5p),均用脂质体法转染至SiHa细胞;克隆形成实验检测细胞的存活分数;流式细胞术检测细胞的凋亡率;双荧光素酶报告基因检测实验检测细胞的荧光活性;Western blot检测细胞中PTEN、p-Akt、Akt的蛋白表达。结果 与放射敏感组相比,放射抗性宫颈癌组织中LncRNA MEG3的表达明显降低(P<0.05),其表达量与宫颈癌细胞的放射敏感性呈正相关;过表达LncRNA MEG3、抑制miR-181a-5p均可显著增强宫颈癌细胞SiHa放射敏感性,促进凋亡(P<0.05);野生型LncRNA MEG3细胞的荧光活性受miR-181a-5p的抑制。过表达miR-181a-5p逆转了LncRNA MEG3对宫颈癌细胞放射增敏和促凋亡作用及对PTEN/Akt信号通路的调控。结论 长链非编码RNA LncRNA MEG3可增强宫颈癌细胞放射敏感性,其机制可能与靶向miR-181a-5p调控PTEN/Akt 信号通路有关,可为提高宫颈癌的预后提供新方向。     

MicroRNA-181c provides neuroprotection in an intracerebral hemorrhage model

Apoptosis is an important factor during the early stage of intracerebral hemorrhage.MiR-181 c plays a key regulatory role in apoptosis.However,whether miR-181 c is involved in apoptosis of prophase cells after intracerebral hemorrhage remains unclear.Therefore,in vitro and in vivo experiments were conducted to test this hypothesis.In vivo experiments:collagenase type VII was injected into the basal ganglia of adult Sprague-Dawley rats to establish an intracerebral hemorrhage model.MiR-181 c mimic or inhibitor was injected in situ 4 hours after intracerebral hemorrhage.Neurological functional defects(neurological severity scores)were assessed 1,7,and 14 days after model establishment.Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling and western blot assay were conducted 14 days after model establishment.In vitro experiments:PC12 cells were cultured under oxygen-glucose deprivation,and hemins were added to simulate intracerebral hemorrhage in vitro.MiR-181 c mimic or inhibitor was added to regulate miR-181 c expression.3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay,luciferase reporter system,and western blot assay were performed.Experimental results revealed differences in miR-181 c expression in brain tissues of both patients and rats with cerebral hemorrhage.In addition,in vitro experiments found that miR-181 c overexpression could upregulate the Bcl-2/Bax ratio to inhibit apoptosis,while inhibition of miR-181 c expression could reduce the Bcl-2/Bax ratio and aggravate apoptosis of cells.Regulation of apoptosis occurred through the phosphoinositide 3 kinase(PI3 K)/Akt pathway by targeting of phosphatase and tensin homolog deleted on chromosome ten(PTEN).Higher miR-181 c overexpression correlated with lower neurological severity scores,indicating better recovery of neurological function.In conclusion,miR-181 c affects the prognosis of intracerebral hemorrhage by regulating apoptosis,and these effects might be directly mediated and regulated by targeting of the PTEN\PI3 K/Akt pathway and Bcl-2/Bax ratio.Furthermore,these results indicated that miR-181 c played a neuroprotective role in intracerebral hemorrhage by regulating apoptosis of nerve cells,thus providing a potential target for the prevention and treatment of intracerebral hemorrhage.Testing of human serum was authorized by the Ethics Committee of China Medical University(No.2012-38-1)on February 20,2012.The protocol was registered with the Chinese Clinical Trial Registry(Registration No.ChiCTR-COC-17013559).The animal study was approved by the Institutional Animal Care and Use Committee of China Medical University(approval No.2017008)on March 8,2017.     

Notch1 contributes to TNF-α-induced proliferation and migration of airway smooth muscle cells through regulation of the Hes1/PTEN axis

Notch1 has been implicated in asthma pathogenesis. However, the function of Notch1 in regulating airway smooth muscle (ASM) cell proliferation and migration during airway remodeling of asthma remains unknown. Using an in vitro model induced by tumor necrosis factor (TNF)-α, we reported in this study that Notch1 participated in TNF-α-induced proliferation and migration of ASM cells. Our results demonstrated that Notch1 expression was significantly upregulated in ASM cells exposed to TNF-α. Notch1 inhibition significantly repressed TNF-α-induced ASM cell proliferation and migration, while Notch1 overexpression promoted the opposite effect. Moreover, Notch1 inhibition downregulated the expression of Notch-1 intracellular domain (NICD) and Hes1, while upregulated PTEN expression in TNF-α-exposed cells. Notably, Hes1 overexpression partially reversed the Notch1-inhibition-mediated inhibitory effect on TNF-α-induced ASM cell proliferation and migration. In addition, the promoting effect of Notch1 inhibition on PTEN expression was markedly abrogated by Hes1 overexpression. Overall, these findings demonstrated that Notch1 inhibition repressed TNF-α-induced ASM cell proliferation and migration by modulating the Hes1/PTEN signaling axis, a finding that highlights the involvement of Notch1/Hes1/PTEN in regulating airway remodeling of asthma.     

microRNA-21 mediates epithelial-mesenchymal transition of human hepatocytes via PTEN/Akt pathway

miR-21 has been shown to play fundamental role in diverse biological and pathological processes, including fibrotic diseases. In the present study, we investigated whether miR-21 regulated the fibrogenic epithelial-mesenchymal transition (EMT) in human hepatocytes QSG-7701 and explored underlying mechanisms. The results showed that treatment of QSG-7701 cells with pro-fibrogenic factor TGF-β₁ resulted in increased expression of miR-21 and promoted fibrogenic EMT in hepatocytes. Downregulation of miR-21 expression by transfection of anti-miR-21 into QSG-7701 cells inhibited fibrogenic EMT induced by TGF-β₁. Furthermore, overexpression of miR-21 alone also resulted in EMT-like transformation in QSG-7701 cells. TGF-β₁ treatment resulted in decreased PTEN and increased Akt phosphorylation and anti-miR-21 abolished this effect. Overexpression of miR-21 in QSG-7701 cells also downregulated PTEN and upregulated Akt phosphralation. Inhibition of Akt signaling by specific inhibitor Akt inhibitor IV blocked TGF-β₁ and miR-21-induced fibrogenic EMT. In summary, our results identify miR-21 as a key regulator of fibrogenic EMT in hepatocytes via PTEN/Akt pathway. Targeting miR-21 may provide a new therapeutic strategy against hepatic fibrosis.     

A Phase I Trial of IGF-1R Inhibitor Cixutumumab and mTOR Inhibitor Temsirolimus in Metastatic Castration-resistant Prostate Cancer

BackgroundDespite frequent PTEN (phosphatase and tensin homologue) loss and Akt/mammalian target of rapamycin (mTOR) signaling in prostate cancer, the disease is insensitive to single-agent mTOR inhibition. Insulin-like growth factor-1 receptor inhibition might mitigate the feedback inhibition by Torc1 inhibitors, suppressing downstream Akt activation and, thus, potentiating the antitumor activity of mTOR inhibition.Patients and MethodsIn the present phase I study, patients with metastatic castration-resistant prostate cancer received 6 mg/kg cixutumumab and 25 mg temsirolimus intravenously each week. The primary objective was safety and tolerability. Temsirolimus was decreased if ≥ 2 dose-limiting toxicities (DLTs) were observed in 6 patients. The correlative analyses included measurement of circulating tumor cells, [18F]-fluoro-2-deoxyglucose positron emission tomography, 16β-[18F]-fluoro-α-dihydrotestosterone positron emission tomography, and tumor biopsy.ResultsA total of 16 patients were enrolled across 3 cohorts (1, −1, −2). Two DLTs (grade 3 oral mucositis) were observed in cohort 1 (temsirolimus, 25 mg), and 1 DLT (grade 3 lipase) in cohort −1 (temsirolimus, 20 mg). The most common adverse events included hyperglycemia (100%; 31% grade 3), oral mucositis (63%; 19% grade 3), and diarrhea (44%; 0 grade 3). Low-grade pneumonitis occurred in 7 of 11 patients (44%; 0 grade 3), prompting the opening of a 3-weekly cohort (temsirolimus, 20 mg/kg), without pneumonitis events. No patient had a >50% decline in prostate-specific antigen from baseline. The best radiographic response was stable disease, with median study duration of 22 weeks (range, 7-63 weeks).ConclusionsDespite a strong scientific rationale for the combination, temsirolimus plus cixutumumab demonstrated limited antitumor activity and a greater than expected incidence of toxicity, including low-grade pneumonitis and hyperglycemia. Hence, the trial was stopped in favor of alternative androgen receptor/phosphatidylinositol 3-kinase–directed combinatorial therapies.     

抑癌基因PTEN与头颈肿瘤研究进展

PTEN是新近发现的具有磷酸酶活性的一种抑癌基因,它的异常表达见于各种散发性肿瘤。本文就其基因定位、结构特点、生物学活性、抑癌作用的信号传导机制及其在头颈恶性肿瘤中的表达及预后作一综述。     

PTEN、p27在口腔粘膜白斑和口腔癌组织中的表达及意义

目的　探讨抑癌基因PTEN、p2 7在口腔粘膜白斑 (oralleukoplakia ,OLK)和口腔癌 (oralsquamouscellcancer,OSCC)组织中的蛋白表达及意义。方法　应用免疫组化S -P法检测 10例正常口腔粘膜、2 5例口腔粘膜白斑 (其中白斑伴上皮异常增生 15例 ,白斑不伴上皮异常增生 10例 )及 32例口腔癌组织中抑癌基因PTEN和p2 7的蛋白表达情况。结果　正常口腔粘膜和白斑不伴上皮异常增生组织中PTEN和 p2 7蛋白全部阳性表达 ;白斑伴上皮异常增生组织中PTEN和 p2 7蛋白阳性表达率分别为93.3%、73.3%。OSCC组织中PTEN蛋白阳性表达率为71.9% ,其中高、中、低分化OSCC组织中PTEN蛋白阳性表达率分别为 85 .7%、75 %和 33.3% ,统计学分析表明PTEN在OSCC组织中的蛋白表达与组织分化程度明显相关 (P <0 .0 5 )。OSCC组织中 p2 7蛋白的阳性表达率为4 0 .6 % ,其中高、中、低分化OSCC组织中p2 7蛋白的阳性表达率分别为 4 2 .9%、4 1.7%和 33.3% ,统计学分析表明p2 7在OSCC组织中的蛋白表达与组织分化程度无明显相关 (P >0 .0 5 )。结论　OSCC组织中PTEN和 p2 7蛋白的阴性表达率较高 ,说明PTEN和 p2 7基因突变或缺失在OSCC的发生发展过程中起重要作用。     

三子颗粒下调miR-205-5p抑制小鼠脾虚型肠道腺瘤生长研究

目的 探讨三子颗粒通过降低微小核糖核酸-205-5p（miR-205-5p）水平抑制小鼠脾虚型肠道腺瘤生长的作用。方法 取70只4周龄的雄性C57BL/6J小鼠,采用对氧化偶氮甲烷（AOM）/葡聚糖硫酸钠（DSS）诱导小鼠结直肠腺瘤模型,将建模小鼠随机分为模型组、阿司匹林（200 mg·kg^-1）组、miR inhibitor-NC （2 mg·kg^-1）组、miR-205-5p inhibitor （2 mg·kg^-1）组和三子颗粒低、中、高剂量（1.7、3.4、6.8 g·kg^-1）组,造模期间ig给药,每天1次。比较各组小鼠肠道腺瘤的数量并测量腺瘤体积;苏木素-伊红（HE）染色观察小鼠肠道腺瘤的病理情况;CCK-8法检测各组小鼠肠道腺瘤细胞增殖活力;原位末端标记（TUNEL）法检测小鼠肠道腺瘤细胞凋亡;实时荧光定量PCR（qRT-PCR）法检测肠道腺瘤miR-205-5p表达量及磷酸酯酶与张力蛋白同源物（PTEN）、B淋巴细胞瘤-2（Bcl-2）、Bcl-2相关X蛋白（Bax）、Ki67 mRNA表达量;Western blotting法检测PTEN、Bcl-2、Bax、Ki67蛋白表达水平;荧光素酶活性实验验证miR-205-5p和PTEN的靶向关系。结果 与模型组比较,阿司匹林组和三子颗粒低、中、高剂量组小鼠肠道腺瘤数量、体积及细胞增殖活性均显著降低,凋亡率显著升高（P＜0.05）,miR-205-5p表达量、Bcl-2、Ki67 mRNA及蛋白表达量显著降低,PTEN、Bax mRNA及蛋白表达量显著升高（P＜0.05）,其中三子颗粒作用呈剂量相关性;与miR inhibitor-NC组比较,miR-205-5p inhibitor组小鼠肠道腺瘤数量、体积及细胞增殖活性均显著降低,凋亡率显著升高（P＜0.05）,miR-205-5p表达量、Bcl-2、Ki67 mRNA及蛋白表达量显著降低,PTEN、Bax mRNA及蛋白表达量显著升高（P＜0.05）;荧光素酶活性实验证实miR-205-5p可靶向调控PTEN。结论 三子颗粒可抑制小鼠脾虚型肠道腺瘤生长,可能是通过下调miR-205-5p,上调PTEN、Bax表达,下调Bcl-2、Ki67表达发挥作用的。