The role and molecular mechanism of NOP16 in the pathogenesis of nasopharyngeal carcinoma

We aimed to explore the effects of NOP16 on the pathogenesis of nasopharyngeal carcinoma (NPC) and the related mechanism. In this study, the expression level of NOP16 in NPC tissues and adjacent tissues was measured by qRT-polymerase chain reaction (PCR) and immunohistochemistry (IHC) tests. In the in vitro study, the expression levels of NOP16 and RhoA/phosphatidylinositol 3-kinase (PI3K)/Akt/c-Myc and IKK/IKB/NF-κB signalling pathway-related proteins in NPC cells were measured by qRT-PCR and Western blot (WB). CCK8 assays and colony formation assays were used to detect cell proliferation. Transwell assays were used to detect the migration and invasion ability of NPC cells. Flow cytometry and WB were used to measure the level of apoptosis. For the in vivo study, NPC xenograft models were established in nude mice, and tumour weight and volume were recorded. The expression levels of NOP16 and RhoA/PI3K/Akt/c-Myc signalling pathway-related proteins and mRNAs were measured by immunofluorescence, qRT-PCR and WB experiments. In clinical samples, the results of qRT-PCR and IHC experiments showed that the expression level of NOP16 was significantly increased in NPC tissues. In the in vitro study, the results of qRT-PCR and WB experiments showed that NOP16 was significantly increased in NPC cells. The CCK8 assay, colony formation assay, transwell assay and flow cytometry results showed that knocking out NOP16 inhibited the proliferation, migration and invasion of NPC cells and increased apoptosis. WB results showed that knocking out NOP16 inhibited the RhoA/PI3K/Akt/c-Myc and IKK/IKB/NF-κB signalling pathways. These effects were reversed by 740Y-P (PI3K activator). In the in vivo study, knockdown of NOP16 reduced tumour volume and weight and inhibited the RhoA/PI3K/Akt/c-Myc signalling pathway. In conclusion, knockdown of NOP16 inhibited the proliferation, migration and invasion of NPC cells and induced apoptosis by inhibiting the RhoA/PI3K/Akt/c-Myc and IKK/IKB/NF-κB pathways, leading to the malignant phenotype of NPC.     

miR-143 inhibits proliferation and metastasis of nasopharyngeal carcinoma cells via targeting FMNL1 based on clinical and radiologic findings

Mounting evidence has reported that microRNA-143 (miR-143) is involved in the development of multiple cancers. To investigate the underlying mechanisms of miR-143 regulating proliferation and metastasis in nasopharyngeal carcinoma (NPC) cells, we evaluated the levels of miR-143 and formin-like protein 1 (FMNL1) in NPC tissues. The results of qRT-PCR and Western blot analysis showed that the expression of miR-143 was decreased, while FMNL1 was increased in NPC tissues. The expression of miR-143 was significantly elevated in NPC cells compared with that of human nasopharyngeal epithelial cells. The results of MiRcode prediction, dual-luciferase reporter, and Western blot analysis assays indicated that miR-143 negatively regulated the expression of FMNL1 (r² = 0.4365P = 0.0001). Overexperssion of miR-143 or FMNL1 knockdown inhibited cell proliferation, migration, and invasion in NPC cells (P < 0.05). Ectopic expression of FMNL1 undermined the inhibition effect of miR-143 on proliferation, migration, and invasion in NPC cells. The findings of this study revealed that miR-143 functioned as a tumor suppressor and inhibited the NPC progression by targeting FMNL1.     

LncRNA CCAT1 modulates the sensitivity of paclitaxel in nasopharynx cancers cells via miR-181a/CPEB2 axis

Recent studies reported that long non-coding RNA (lncRNA) might play critical roles in regulating chemo-resistant of multiple types of cancer. This study aimed to investigate whether long non-coding RNA CCAT1 was involved in Paclitaxel resistance in nasopharyngeal carcinoma (NPC). qRT-PCR was used for testing the expression of CCAT1, miR-181a and CPEB2 in tumor tissues and NPC cancers. NPC cells were transfected with siRNAs to suppress the mRNA level of CCAT1 in NPC cells. MTT assays and flow cytometry analysis were used to assess the sensitivity of paclitaxel in NPC cells. Luciferase reporter assays were used to examine the interaction of CCAT1 or CPEB2 to miR-181a. Our findings revealed that the upregulated CCAT1 results in significantly enhancing paclitaxel resistance in nasopharyngeal cancer cells. Bioinformatics analysis and luciferase reporter assay indicated that the upregulated CCAT1 sponges miR-181a in NPC cells. Furthermore, RNA immuno-precipitation assays showed that miR-181a could directly bind to CCAT1 mRNA in NPC cells. We restored miR-181a in NPC cells, and found restoration of miR-181a re-sensitized the NPC cells to paclitaxel in vitro. In addition, our results also showed that miR-181a was a modulator of paclitaxel sensitivity due to its regulative effect on cell apoptosis via targeting CPEB2 in NPC cells. Taken together, lncRNA CCAT1 regulates the sensitivity of paclitaxel in NPC cells via miR-181a/CPEB2 axis.     

AKR1C2 acts as a targetable oncogene in esophageal squamous cell carcinoma via activating PI3K/AKT signaling pathway

The aldo‐keto reductases family 1 member C2 (AKR1C2) has critical roles in the tumorigenesis and progression of malignant tumours. However, it was also discovered to have ambiguous functions in multiple cancers and till present, its clinical significance and molecular mechanism in oesophageal squamous cell carcinoma (ESCC) has been unclear. The aim of this study was to explore the role of AKR1C2 in the tumorigenesis of ESCC. Here, we showed that AKR1C2 expression was found to be up‐regulated in ESCC tissues and was significantly associated with pathological stage, lymph node metastasis and worse outcomes. Functional assays demonstrated that an ectopic expression of AKR1C2 in ESCC cells resulted in increased proliferation, migration and cisplatin resistance, while knockdown led to inversing effects. Bioinformation analyses and mechanistic studies demonstrated that AKR1C2 activated the PI3K/AKT signalling pathway, furthermore, the inhibitor of PI3K or the selective inhibitor of AKR1C2 enzyme activity could reverse the aggressiveness and showed synergistic antitumour effect when combined with cisplatin, both in vitro and in vivo. In conclusion, Our findings revealed that AKR1C2 could function as an oncogene by activating the PI3K/AKT pathway, as a novel prognostic biomarker and/or as a potential therapeutic target to ESCC.     

Copper transporter Ctr1 contributes to enhancement of the sensitivity of cisplatin in esophageal squamous cell carcinoma

Increasing evidence has demonstrated that Ctr1 plays a crucial role in the regulation of cisplatin uptake in a variety of tumors. The purpose of this study was to investigate its role in mediating cisplatin sensitivity in ESCC cells. Immunohistochemistry (IHC), In situ hybridization (ISH) and semi-quantitative RT-PCR were used to detect Ctr1 expressions in ESCC tissues. qRT-PCR and Western blot was performed to investigate the levels of Ctr1 mRNA and protein in ESCC cells. CCK-8, Flow cytometry and Transwell chamber assay were carried out to examine cell proliferation, apoptosis, migration and invasion abilities in ESCC cells. We found that ESCC tissues and cells had higher Ctr1 level than normal tissues and Het-1A cell. Ctr1 expression was correlated with histological grade, invasion depth, TNM staging and lymph node metastasis in ESCC patients. Ctr1 depletion reduced the suppressive role of proliferation, migration and invasion as well as the inductive role of cell apoptosis and Caspase-3 activity evoked by cisplatin, whereas Ctr1 upregulation combined with cisplatin exerted the synergistic role in regulation of proliferation, apoptosis, Caspase-3 activity, migration and invasion in ESCC. In conclusion, Ctr1 is implicated in ESCC development and progression and its expression may be a novel predictor for assessment of cisplatin sensitivity in ESCC.     

Lentivirus-mediated RNAi knockdown of LMP2A inhibits the growth of nasopharyngeal carcinoma cell line C666-1 in vitro

Latent membrane protein 2A (LMP2A) is found to play a key role in the development of nasopharyngeal carcinoma (NPC). However, the role of LMP2A silencing in the inhibition of cell growth of NPC has not been clarified. In this study, we inhibited LMP2A gene expression by lentivirus-mediated RNAi, to explore the effects of LMP2A silencing on the growth of NPC cell line in vitro. A lentivirus-mediated RNAi technology was employed to specifically knock down the LMP2A gene in NPC cell line C666-1. Quantitative real-time polymerase chain reaction, Western blot, flow cytometry and colony formation assays were performed to evaluate the expression of LMP2A and biological behavior of cell line C666-1 in vitro. We successfully construct a highly efficient and stable lentivirus vector, which efficiently downregulate the expression of LMP2A gene in infected cell line C666-1. Down-regulation of the expression of LMP2A significantly inhibits the proliferation and colony formation of C666-1 cells. In addition, the specific down-regulation of LMP2A arrests cells in G0/G1 phase of cell cycle and increases apoptosis rate. Our findings suggest that lentivirus-mediated RNAi knockdown of LMP2A inhibits the growth of NPC cell line C666-1 in vitro, and LMP2A may be a potential target for gene therapy in treatment of NPC.     

circPVT1通过miR-24-3p/let-7a-5p/FSCN1轴促进鼻咽癌细胞侵袭迁移

目的 鼻咽癌是一种来源于鼻咽上皮的恶性肿瘤,其临床特征之一是易发生淋巴转移,但是目前鼻咽癌转移的分子机制尚未阐明。circPVT1是由PVT1基因2号外显子反向拼接形成的环状RNA (circRNA),在多种肿瘤中表达上调,本文探讨了circPVT1在鼻咽癌侵袭迁移中的作用和分子机制。方法 通过RT-qPCR检测circPVT1及其下游miRNA和FSCN1在鼻咽癌细胞的表达情况,Transwell和划痕愈合实验检测circPVT1对鼻咽癌细胞侵袭迁移的影响,RNA pull-down实验检测circPVT1结合的miRNA,双荧光素酶报告实验检测miR-24-3p和let-7a-5p靶向抑制FSCN1 mRNA表达。结果 在鼻咽癌细胞中过表达circPVT1可以促进鼻咽癌细胞侵袭迁移,而敲低circPVT1则可以抑制鼻咽癌细胞的侵袭迁移。进一步研究发现,circPVT1可以通过竞争性吸附miR-24-3p和let-7a-5p,上调FSCN1的表达,从而促进鼻咽癌细胞的侵袭迁移。结论 circPVT1通过miR-24-3p/let-7a-5p/FSCN1轴促进鼻咽癌细胞侵袭迁移,证实c...     

LTF基因在鼻咽癌细胞系中生物学功能的初步研究

目的:初步探讨野生型LTF基因在鼻咽癌细胞系中的生物学功能。方法:野生型LTF导入鼻咽癌细胞系,G418筛选,RT-PCR和Western-blotting分别在mRNA和蛋白质水平进行验证,得到稳定表达LTF基因的鼻咽癌细胞系。流式细胞术、平板克隆形成实验和MTT法分别检测细胞周期、细胞的克隆形成能力和细胞生长曲线。结果:成功导入LTF并稳定表达的鼻咽癌细胞系,G0-G1期细胞百分比例明显增加(72.01%vs 62.31%),G2-M期细胞百分比例减少(6.26%vs 10.81%);克隆形成能力降低(39.5%vs 59.7%),体外瘤细胞增殖能力降低(P0.05)。结论:LTF基因可阻滞细胞周期、抑制鼻咽癌细胞系的增殖能力和克隆形成率,同时为进一步的体内试验研究奠定基础。     

AKR1B10 confers resistance to radiotherapy via FFA/TLR4/NF-κB axis in nasopharyngeal carcinoma

Nasopharyngeal carcinoma (NPC) is one kind of human head and neck cancers with high incidence in Southern China, Southeast Asia and North Africa. In spite of great innovations in radiation and chemotherapy treatments, the 5-year survival rate is not satisfactory. One of the main reasons is resistance to radiotherapy which leads to therapy failure and recurrence of NPC. The mechanism underlying remains to be fully elucidated. Aldo-keto reductase B10 (AKR1B10) plays a role in the formation and development of carcinomas. However, its role in resistance to radiotherapy of NPC is not clear. In this research, the relationships between AKR1B10 expression and the treatment effect of NPC patients, NPC cell survival, cell apoptosis, and DNA damage repair, as well as the effect and mechanism of AKR1B10 expression on NPC radioresistance were explored. A total of 58 paraffin tissues of NPC patients received radiotherapy were collected including 30 patients with radiosensitivity and 28 patients with radioresistance. The relationships between AKR1B10 expression and the treatment effect as well as clinical characteristics were analyzed by immuno-histochemical experiments, and the roles of AKR1B10 in cell survival, apoptosis and DNA damage repair were detected using the AKR1B10 overexpressed cell models. Furthermore the mechanism of AKR1B10 in NPC radioresistance was explored. Finally, the radioresistance effect of AKR1B10 expression was evaluated by the tumor xenograft model of nude mice and the method of radiotherapy. The results showed AKR1B10 expression level was correlated with radiotherapy resistance, and AKR1B10 overexpression promoted proliferation of NPC cells, reduced apoptosis and decreased cellular DNA damage after radiotherapy. The probable molecular mechanism is that AKR1B10 expression activated FFA/TLR4/NF-κB axis in NPC cells. This was validated by using the TLR4 inhibitor TAK242 to treat NPC cells with AKR1B10 expression, which reduced the phosphorylation of NF-κB. This study suggests that AKR1B10 can induce radiotherapy resistance and promote cell survival via FFA/TLR4/NF-κB axis in NPC, which may provide a novel target to fight against radiotherapy resistance of NPC.     

LncRNA EPB41L4A-AS2 represses Nasopharyngeal Carcinoma Metastasis by binding to YBX1 in the Nucleus and Sponging MiR-107 in the Cytoplasm

Nasopharyngeal carcinoma (NPC) is known for its potential to progress to the lymph nodes and distant metastases at an early stage. As an important regulator in tumorigenesis biological processes, the functions of lncRNA in NPC tumor development remain largely unclear. In this research, the expression of EPB41L4A-AS2 in NPC tissues and cells was analyzed via real-time quantitative polymerase chain reaction (qRT-PCR). CCK8, colony formation, and EDU experiments were used to determine the viability of NPC cells. Transwell and wound healing assays were performed to test NPC cell migration and invasion. RNA pull-down and mass spectrometry analysis were used to identify potential binding proteins. Then, a popliteal lymph node metastasis model was established to test NPC metastasis. EPB41L4A-AS2 is repressed by transforming growth factor-beta, which is downregulated in NPC cells and tissue. It is associated with the presence of distant metastasis and adverse outcomes. The univariate and multivariate survival assays confirmed that EPB41L4A-AS2 expression was an independent predictor of progression-free survival (PFS) in patients with NPC. Biological analyses showed that overexpression of EPB41L4A-AS2 reduced the metastasis and invasion of NPC in vitro and in vivo, but had no significant effect on cell proliferation. Mechanistically, in the nucleus we identified that EPB41L4A-AS2 relies on binding to YBX1 to reduce the stability of Snail mRNA to enhance the expression of E-cadherin and reverse the progression of epithelial-to-mesenchymal transition (EMT). In the cytoplasm, we found that EPB41L4A-AS2 blocked the invasion and migration of NPC cells by promoting LATS2 expression via sponging miR-107. In a whole, the findings of this study help to further understand the metastasis mechanism of NPC and could help in the prevention and treatment of NPC metastasis.     

Silencing of LMP1 Induces Cell Cycle Arrest and Enhances Chemosensitivity Through Inhibition of AKT Signaling Pathway in EBV-Positive Nasopharyngeal Carcinoma Cells

The latent membrane protein 1 (LMP1) of Epstein-Barr virus (EBV) is closely associated with nasopharyngeal carcinoma (NPC). In this study, we investigated that the effect of silencing LMP1 on cell cycle distribution and chemosensitivity in EBV-positive nasopharyngeal carcinoma C666-1 cells. Silencing of LMP1 by specific siRNA induced G1 arrest in C666-1 cells. The protein expression of CDK4 and cyclin D1 decreased and P27 was upregulated following LMP1 knockdown. Phosphorylation of AKT and its downstream targets IКB, FKHR was inhibited by LMP1 siRNA. The chemosensitivity of C666-1 cells to bleomycin and cisplatin was enhanced by siRNA targeting LMP1. The cells treated with LMP1 siRNA showed enhanced cleavage of the effector caspase3 and PARP, and Bax had the tendency to exhibit higher expression. Also, co-transfection of constitutive active AKT plasmid with LMP-1 siRNA plasmid abrogates sensitivity of C666-1 to bleomycin and cisplatin. It is reported for the first time that AKT signaling pathway was directly involved in the effects induced by siRNA targeting LMP1. Our findings confirm LMP1 as a rational therapeutic target in NPC.     

Proteomic features of potential tumor suppressor NESG1 in nasopharyngeal carcinoma

We previously defined the recently revised NESG1 gene as a potential tumor suppressor in nasopharyngeal carcinoma (NPC). Here, we further used proteomics technology to globally examine NESG1‐controlled proteins in NPC cells. Twenty‐six proteins were found to be deregulated by NESG1 using proteomics analysis while enolase 1 (alpha) (ENO1), heat shock protein 90 kDa beta (Grp94), member 1 (HSP90B1), and cathepsin D (CTSD) proteins were differentially expressed by Western blot. Interestingly, a‐enolase (ENO1), an overexpressed gene in NPC, was confirmed as a NESG1‐regulated protein in NPC cells. Overexpressed ENO1 not only restored cell proliferation and cell‐cycle progression, but also antagonized the regulation of NESG1 to cell‐cycle regulators p21 and CCNA1 expression as well as induced the expression of C‐Myc, pRB, and E2F1 in NESG1‐ovexpressed NPC cells. Real‐time PCR and immunohistochemistry analysis showed that NESG1 expression is negatively correlated with ENO1 expression in NPC tissues. Our observations suggest that ENO1 downregulation plays an important role in NESG1‐induced growth inhibition of NPC cancer cells.     

沉默DEPDC1抑制鼻咽癌细胞HNE-1和CNE-1侵袭迁移

DEPDC1(DEP domain containing 1)是一个新的肿瘤相关基因,在多种恶性肿瘤的发生发展进程中起着重要作用。我们前期工作中在鼻咽癌细胞内沉默了DEPDC1的表达,发现抑制细胞增殖并诱发细胞凋亡。本研究旨在探讨沉默DEPDC1表达后,对鼻咽癌细胞HNE-1和CNE-1侵袭迁移能力的影响及其分子机制。结果显示,siRNA介导DEPDC1表达沉默后,细胞侧向运动能力、侵袭及迁移能力显著降低。qRT-PCR及Western印迹检测发现DEPDC1沉默导致EMT上游关键转录因子Twist1及间质细胞标志分子Vimentin表达显著下调。这些研究表明,鼻咽癌细胞中DEPDC1通过调节Twist1等EMT关键分子的表达在细胞侵袭转移过程中起关键作用。推测DEPDC1在鼻咽癌中高表达可能对于促进其侵袭转移具有重要作用,进而促进肿瘤发生发展,但具体分子机制仍有待更深入研究。     

褪黑素联合MPA抑制子宫内膜异常增生的作用及机制研究

摘要 目的：探索褪黑素联合MPA（醋酸甲羟孕酮）对子宫内膜异常增生细胞增殖活性的抑制作用及其机制。方法：取分化良好的子宫内膜增生细胞株Ishikawa和内膜癌细胞株ECC1于适宜条件培养,加入褪黑素、MPA单独或者联合处理48 h后,检测子宫内膜细胞株的增殖活性。收集褪黑素、MPA单独或者联合处理48 h后的子宫内膜增生细胞株Ishikawa细胞,提取细胞内的蛋白,检测人20α-羟基类固醇脱氢酶（AKR1C1）的表达情况。结果：褪黑素和MPA联合使用后对子宫内膜异常增生细胞的抑制作用明显高于褪黑素或MPA单独使用。褪黑素和MPA可抑制AKR1C1的表达,二者联合使用对AKR1C1的抑制高于两者单独使用。结论：褪黑素可提高子宫内膜异常增生细胞对MPA的敏感性,降低MPA的使用剂量,同时抑制AKR1C1的表达,使孕酮的代谢速率降低。褪黑素与MPA联合使用给子宫内膜增生和内膜癌的治疗策略带来新的思路。     

ALDH1A1诱导肺腺癌细胞发生上皮-间质转化及化疗耐药

目的:探索醛脱氢酶1A1(aldehyde dehydrogenase 1A1,ALDH1A1)在肺腺癌细胞(lung adenocarcinoma cell,LAC)化疗耐药中的作用及机制,为肺癌临床治疗和新型药物的研发提供实验依据。方法:采用慢病毒载体构建ALDH1A1高表达肺腺癌细胞模型,并通过流式细胞术和western blot技术对该细胞模型进行验证。通过CCK8法检测ALDH1A1高表达肺腺癌细胞对肺癌治疗药物顺铂(cisplatin,DDP)、紫杉醇(paclitaxcel)、厄洛替尼(erlotinib)和吉非替尼(gefitinib)的耐药性。通过检测肿瘤干细胞(cancer stem cell,CSC)分子标志物、上皮-间质转化(Epithelial-Mesenchymal Transition,EMT)分子标志物及细胞迁移能力探讨ALDH1A1高表达对肺腺癌细胞的干性和EMT特征的影响。双硫仑(disulfiram,DSF)是ALDH的抑制剂,我们通过CCK8法和transwell细胞迁移实验探究DSF对肺腺癌细胞体外生长和迁移能力的影响,体内实验探究DSF和厄洛替尼联合用药对HCC827-ALDH1A1细胞皮下异种移植瘤生长的影响。结果:ALDH1A1高表达诱导肺腺癌细胞对厄洛替尼、吉非替尼、紫杉醇和顺铂产生不同程度的耐药,干细胞标志物CD44、CD133蛋白表达上调,EMT间充质标志物vimentin蛋白表达上调,transwell实验结果显示ALDH1A1高表达肺腺癌细胞的迁移能力增强,使用ALDH靶向抑制剂DSF能选择性抑制ALDH1A1高表达肺腺癌细胞所增高的迁移能力并克服HCC827-ALDH1A1细胞皮下异种移植瘤的生长,延缓体内耐药。结论:ALDH1A1能诱导肺腺癌细胞对多种抗肺癌药物产生耐药并发生干细胞样转化,靶向抑制ALDH酶活性可克服由ALDH1A1高表达所产生的耐药,为肺癌的临床治疗提供新的思路。     

Placenta specific 8 gene induces epithelial-mesenchymal transition of nasopharyngeal carcinoma cells via the TGF-β/Smad pathway

The present study aimed to investigate the effects and mechanisms of PLAC8 on the epithelial-mesenchymal transition (EMT) of Nasopharyngeal carcinoma (NPC). The expression of PLAC8 in NPC and nasopharyngitis (NPG) tissues from 150 patients was determined using immunohistochemistry. The levels of PLAC8 in five NPC cell lines and nasopharyngeal permanent epithelial cell line were measured using western blotting. We then knocked out or overexpressed PLAC8 in CNE2 cells. Cell proliferation, wound healing, migration, and invasion assays were used to analyze the effects of PLAC8 on the proliferation, migration, and invasion in vivo and vitro. The results showed that the expression of PLAC8 was much higher in NPC tissues than in NPG tissues. The expression of PLAC8 was higher in all the cell lines than in the nasopharyngeal permanent epithelial cells. PLAC8 knockout resulted in significant decreases in cell proliferation, migration, and invasion; associated with lower protein levels of N-cadherin; and increased levels of E-cadherin. Overexpression of PLAC8 had the opposite effect. Furthermore, knockout of PLAC8 inactivated TGF-β/SMAD signaling pathway and suppressed the growth of NPC xenografts. PLAC8 may promote the carcinogenesis and EMT of NPC via the TGF-β/Smad pathway, which suggests that PLAC8 may be a potential biomarker for NPC.     

Knockdown of DIXDC1 Inhibits the Proliferation and Migration of Human Glioma Cells

DIX domain containing 1 (DIXDC1), the human homolog of coiled-coil-DIX1 (Ccd1), is a positive regulator of Wnt signaling pathway. Recently, it was found to act as a candidate oncogene in colon cancer, non-small-cell lung cancer, and gastric cancer. In this study, we aimed to investigate the clinical significance of DIXDC1 expression in human glioma and its biological function in glioma cells. Western blot and immunohistochemistry analysis showed that DIXDC1 was overexpressed in glioma tissues and glioma cell lines. The expression level of DIXDC1 was evidently linked to glioma pathological grade and Ki-67 expression. Kaplan–Meier curve showed that high expression of DIXDC1 may lead to poor outcome of glioma patients. Serum starvation and refeeding assay indicated that the expression of DIXDC1 was associated with cell cycle. To determine whether DIXDC1 could regulate the proliferation and migration of glioma cells, we transfected glioma cells with interfering RNA-targeting DIXDC1; investigated cell proliferation with Cell Counting Kit (CCK)-8, flow cytometry assays, and colony formation analyses; and investigated cell migration with wound healing assays and transwell assays. According to our data, knockdown of DIXDC1 significantly inhibited proliferation and migration of glioma cells. These data implied that DIXDC1 might participate in the development of glioma, suggesting that DIXDC1 can become a potential therapeutic strategy for glioma.     

Knockdown of NFBD1/MDC1 enhances chemosensitivity to cisplatin or 5-fluorouracil in nasopharyngeal carcinoma CNE1 cells

Nasopharyngeal carcinoma (NPC) is a rare but highly invasive cancer that is prevalent among people of southern Chinese ancestry in southern China and Southeast Asia. Radiotherapy and cisplatin (CDDP)-based chemotherapy are the main treatment options. Unfortunately, disease response to concurrent chemoradiotherapy varies among patients with NPC, and many cases are resistant to CDDP and radiotherapy. NFBD1 functions in cell cycle checkpoint activation and DNA repair following DNA damage. In this study, we identified the NFBD1 as a tractable molecular target to chemosensitize NPC cells. NFBD1 expression in NPC CNE1 cell lines was depleted using lentivirus-mediated short hairpin RNA, and the elevated sensitivity of these NFBD1-inhibited NPC cells to therapeutic reagent CDDP and 5-fluorouracil (5-FU) was evaluated using MTS assays. Flow cytometry analysis also showed that NFBD1 knockdown led to an obvious induction of apoptosis in CDDP- or 5-FU-treated CNE1 cells. Furthermore, we implicated the involvement of NFBD1 in Rad51 and DNA-PKcs foci formation following CDDP or 5-FU chemotherapy. In conclusion, NFBD1 knockdown improves the chemosensitivity of NPC cells by inhibiting cell growth and promoting apoptosis through the impairment of DNA damage repair, suggesting NFBD1 as a novel therapeutic target for NPC.

     

Effect of TET1 regulating MGMT on chemotherapy resistance of oral squamous cell carcinoma stem cells

The study was to evaluate the effect of ten‐eleven translocation 1 (TET1) regulating o6‐methylguanine‐DNA methyltransferase (MGMT) in chemotherapy resistance of oral squamous cell carcinoma (OSCC) stem cells. OSCC stem cells were divided into the blank, negative control (NC), TET1‐siRNA, TET1‐siRNA + MGMT‐OE, and MGMT‐OE groups. Methylation‐specific polymerase chain reaction (MSP), qRT‐PCR and Western blotting were conducted to detect the methylation status of MGMT, expressions of TET1, MGMT, ABCG2, and Oct‐4. Cell proliferation, cisplatin chemosensitivity, and cell cycle and apoptosis, were detected using CCK8 and flow cytometry. A chromatin immunoprecipitation (ChIP) assay was employed for detecting the link between TET1 and MGMT gene promoters. In comparison to the NC group, the TET1‐siRNA group exhibited increased levels of MGMT methylation, the number of apoptotic cells and cisplatin chemosensitivity consisting of varying concentrations, however, decreased levels of mRNA and protein expressions of TET1 as well as MGMT, cell viability, the number of cells in the S phase, and protein expressions of ABCG2 and Oct‐4 were all have diminished amounts. The TET1‐siRNA + MGMT‐OE and MGMT‐OE groups had higher MGMT mRNA and protein expression, as well as increased protein expressions of ABCG2 and Oct‐4, greater cell activity, higher number of cells in the S phase, decreased apoptotic rates in cells and decreased cisplatin chemosensitivity with different concentrations. Our study provided evidence that low‐expression of TET1 in OSCC stem cells may stimulate MGMT promoter methylation, while inhibiting MGMT mRNA expression, this ultimately strengthens the sensitivity of OSCC stem cells in regards to chemotherapeutics.