Macrophage-secreted exosomes inhibit breast cancer cell migration via the miR-101-3p/DLG5 axis

Objective:To investigate the role of macrophages in regulating breast cancer cell migration and its related mechanisms.Methods:Human leukemia monocytic cell line THP-1-secreted exosomes were isolated using multi-step ultracentrifugation and verified using nanoparticle tracking analysis.Differentially expressed miRNAs were identified using RNA sequencing.Overexpression of inhibitors of hsa-miR-101-3p in breast cancer MDA-MB-231 cells was performed by infecting their lentiviral constructs.The luciferase reporter assay was used to evaluate the interaction of DLG5 and miR-101.DGL5 expression was detected using qRT-PCR and Western blot analyses.Results:The migration of breast cancer cells was significantly inhibited after addition of exosomes.RNA sequencing results showed that miR-101-3p expression was significantly upregulated.Targetscan analysis predicted that miR-101-3p could target DLG5,and this prediction was verified using the luciferase assay.The addition of the miR-101-3p precursor significantly increased the expression of miR-101-3p,and the mRNA and protein levels of DLG5 were suppressed.In contrast,inhibiting the expression of miR-101-3p increased the mRNA and protein levels of DLG5.Furthermore,the scratch assay showed that inhibiting miR-101-3p could promote the migration of MDA-MB-231 cells.Conclusions:Macrophage exosomes can inhibit the migration of breast cancer cells,and increasing the expression of miR-101-3p to inhibit DLG5 expression may play an important role in this process,which needs further investigation.

下调METTL5通过Wnt/β-catenin信号通路抑制三阴乳腺癌细胞增殖、迁移与侵袭

目的探讨甲基转移酶5(methyltransferase-like 5,METTL5)在三阴乳腺癌(triple-negative breast cancer,TNBC)中的作用和潜在机制。方法采用免疫组织化学方法和Western blot检测TNBC肿瘤组织和细胞系中METTL5的表达情况。用靶向METTL5的shRNA(shRNA-METTL5)转染TNBC细胞后,用CCK-8、集落形成、伤口愈合以及Transwell实验分别检测细胞增殖活性、迁移与侵袭,Western blot检测Wnt/β-catenin信号关键蛋白的表达。构建异种移植瘤模型,验证敲降METTL5对TNBC细胞在体内生长以及Wnt/β-catenin信号活性的影响。结果METTL5在TNBC肿瘤组织和细胞系中表达上调(P<0.01)。敲降METTL5可抑制TNBC细胞的增殖、迁移和侵袭并降低了Wnt/β-catenin信号分子β-catenin、细胞周期蛋白(Cyclin)D1、基质金属蛋白酶(MMP)-2和MMP-7的表达(均P<0.01)。体内实验显示,敲降METTL5减缓了移植瘤生长和Wnt/β-catenin信号活性。结论敲降METTL5能抑制TNBC细胞的增殖、迁移与侵袭,其作用可能与抑制Wnt/β-catenin信号通路有关。

乳腺癌组织lncRNA CASC2、miR-532-3p表达水平与患者术后5年内生存的相关性

目的探究乳腺癌(BC)组织长链非编码RNA癌易感性候选基因2(lncRNA CASC2)、微小RNA-532-3p(miR-532-3p)表达与患者术后5年内生存的相关性。方法选择2015年1月—2018年6月大同市第五人民医院普通外科收治BC患者127例,术中收集BC组织及癌旁正常组织,荧光定量PCR法检测BC组织和癌旁正常组织中lncRNA CASC2、miR-532-3p表达;对BC患者术后进行为期5年的随访,记录患者5年内生存和死亡情况。比较癌旁正常组织和BC组织lncRNA CASC2及miR-532-3p表达,BC组织中lncRNA CASC2和miR-532-3p表达在不同临床病理特征中的差异,生存组和死亡组临床病理特征和BC组织中lncRNA CASC2及miR-532-3p表达的差异。分析BC组织lncRNA CASC2、miR-532-3p表达的相关性;BC组织中lncRNA CASC2和miR-532-3p表达与术后5年内生存的关系;影响BC患者术后5年内生存的因素;lncRNA CASC2、miR-532-3p对BC患者术后5年内生存的预测价值。结果BC组织中lncRNA CASC2表达水平低于癌旁正常组织,miR-532-3p表达水平高于癌旁正常组织(t/P=38.239/<0.001,49.406/<0.001);肿瘤直径≥2 cm、TNM分期Ⅲ期、肿瘤低分化、淋巴结转移者比例lncRNA CASC2低表达组高于高表达组,而miR-532-3p低表达组低于高表达组(lncRNA CASC2:χ^(2)/P=17.361/<0.001、17.052/<0.001、14.694/<0.001、13.173/<0.001;miR-532-3p:χ^(2)/P=10.733/0.001、9.813/0.002、10.134/0.001、7.444/0.006);127例BC患者术后随访5年,生存99例(生存组),死亡28例(死亡组),肿瘤直径≥2 cm、TNM分期Ⅲ期、肿瘤低分化、淋巴结转移者比例及miR-532-3p表达水平死亡组高于生存组,而lncRNA CASC2表达水平死亡组低于生存组[χ^(2)(t)/P=5.211/0.022、27.149/<0.001、27.990/<0.001、4.590/0.032、19.155/<0.001、10.818/<0.001];BC组织中lncRNA CASC2与miR-532-3p表达呈负相关(r/P=-0.561/<0.001);lncRNA CASC2高表达组BC患者术后5年内总生存率为89.23%(58/65),高于lncRNA CASC2低表达组66.13%(41/62)(χ^(2)/P=9.854/0.002);miR-532-3p高表达组BC患者术后5年内总生存率为65.57%(40/61),低于miR-532-3p低表达组89.39%(59/66)(χ^(2)/P=10.466/0.001);肿瘤直径≥2 cm、TNM分期Ⅲ期、肿瘤低分化、有淋巴结转移、lncRNA CASC2低表达、miR-532-3p高表达均是影响BC患者术后5年内生存的独立危险因素[HR(95%CI)=2.255(1.192~4.263)、2.143(1.252~3.666)、3.089(1.386~6.887)、2.219(1.223~4.026)、2.606(1.174~5.788)、2.855(1.592~5.120)];lncRNA CASC2、miR-532-3p及二者联合预测BC患者术后5年内生存的AUC分别为0.840、0.852、0.908,二者联合预测的AUC大于lncRNA CASC2、miR-532-3p各自单独预测的AUC(Z/P=2.246/0.025、2.033/0.042)。结论BC组织中lncRNA CASC2表达下调,miR-532-3p表达上调,且术后5年内死亡的BC患者较存活患者变化更显著,二者表达与临床病理特征相关,对预测BC患者术后5年内生存情况价值较高。

开心散治疗乳腺癌失眠症患者的临床疗效及对血清5-羟色胺水平的影响

目的观察开心散治疗乳腺癌失眠症患者的临床疗效及对血清5-羟色胺(5-hydroxytryptamine,5-HT)水平的影响。方法将63例乳腺癌失眠症患者随机分为对照组33例和治疗组30例。对照组患者采用艾司唑仑治疗,治疗组患者采用开心散联合艾司唑仑治疗。比较两组治疗前后匹兹堡睡眠质量指数量表(Pittsburgh sleep quality index,PSQI)、抑郁自评量表(self-rating depression scale,SDS)、焦虑自评量表(self-rating anxiety scale,SAS)、欧洲癌症研究和治疗协作组编制的癌症患者生活质量评分(European organization for research and treatment of cancer quality of life questionnaire-core 30,EORTC QLQ-C30)以及血清5-HT水平;在连续治疗4周后,比较两组临床疗效及不良反应发生率。结果治疗4周后,两组PSQI、SDS、SAS评分均较治疗前显著降低(P<0.05),且治疗组PSQI评分显著低于对照组(P<0.05);治疗组EORTC QLQ-C30躯体功能、情绪功能、认知功能以及社会功能评分均显著高于对照组(P<0.05);两组患者治疗后血清5-HT水平较治疗前显著升高(P<0.05),且治疗组血清5-HT水平高于对照组(P<0.05);治疗组疗效优于对照组(P<0.05);治疗组总不良反应发生率低于对照组,差异有统计学意义(P<0.05)。结论相较于单纯艾司唑仑治疗,开心散联合艾司唑仑能更好地改善乳腺癌患者睡眠,升高血清5-HT水平,提高乳腺癌患者生活质量,降低不良反应发生率。

PREOPERATIVE CHEMOTHERAPY OF CONTINUOUS INFUSION OF 5-FLUOROURACIL, EPIRUBICIN OR PIRARUBICIN AND CYCLO- PHOSPHAMINE IN OPERABLE PRIMARY BREAST CANCER

Objective: To evaluate the feasibility and activity of continuous-infusion of fluorouracil in association with epirubicin or pirarubicin and cyclophosphamine as neoadjuvant regimen in patients with primary breast cancer. Methods: A total of 111 (including 114 breasts) were entered into the study. Chemotherapy consisted of two to six cycles of epirubicin 50 mg/(m2穌) and cyclophosphamide 500 mg/(m2穌) on day 1 and 8, and continuous intravenous administration of 5-fluorouracil 200 mg/(m2穌) from day 1 to 28 with a microinfusional elastomer (CEFci) or pirarubicin 35 mg/(m2穌) on day 1 and 8 instead of epirubicin (CTFci). Results: The overall response rate was 87.7%. Forty-five patients (39.5%) attained a complete clinical response and 27 (23.7%) attained a pathological complete response. CTFci regimen was superior to CEFci regimen in response rate,the pathological complete response rate (pCR) of former regimen was significantly higher than that of latter regimen (34.8% vs. 16.2%) (P=0.022). The pCR rate in ER/PgR negative tumor was significantly higher than that of ER/PgR positive tumor, achieving 33.3% and 7.5% respectively (P=0.001, x2=11.043). There was no relationship between HER-2 expression and tumor response. The toxicity of two regimens was well tolerated. Alopecia was mild in CTFci regimen comparing with CEFci regimen but neutropenia in CTFci regimen was higher than CEFci regimen. Conclusion: Continuous-infusion of fluorouracil in association with epirubicin or pirarubicin and cyclophosphamine is effective regimens as neoadjuvant chemotherapy for primary breast cancer and the toxicity is well tolerated. Pirarubicin regimen was superior to epirubicin regimen in response rate.

乳腺癌化疗患者血清多巴胺、5-羟色胺与焦虑抑郁状态的关系分析

目的探讨乳腺癌化疗患者血清多巴胺(dopamine,DA)、5-羟色胺(5-hydroxytryptamine,5-HT)与焦虑抑郁状态的关系。方法选取2018年9月至2020年9月于成都市第四人民医院行乳腺癌化疗患者40例作为化疗组,同期选取体检中心进行健康体检人员40例作为对照组,采用酶联免疫吸附法检测血清DA、5-HT,采用汉密尔顿焦虑量表(Hamilton Anxiety Scale,HAMA)、汉密尔顿抑郁量表(Hamilton Depression Scale,HAMD)评估焦虑抑郁状态,分析血清DA、5-HT与焦虑抑郁状态的关系。结果化疗组血清DA、5-HT低于对照组,化疗组HAMA、HAMD评分高于对照组,差异有统计学意义(P<0.05);化疗组中,有焦虑情绪者血清DA、5-HT低于无焦虑情绪者,有抑郁情绪者血清DA、5-HT低于无抑郁情绪者,差异有统计学意义(P<0.05);Pearson相关性分析显示,血清DA、5-HT均与HAMA、HAMD评分呈负相关(P<0.05)。结论乳腺癌化疗患者血清DA、5-HT与焦虑抑郁状态有关,其中DA、5-HT减少可能是患者焦虑抑郁情绪出现的原因。

组蛋白去乙酰化酶HDAC5调控乳腺癌的研究进展

乳腺癌是世界范围内最常见的恶性肿瘤之一,在女性群体中发病率较高。作为IIa组蛋白去乙酰化酶(HDACs),组蛋白去乙酰化酶5(HDAC5)在乳腺癌患者和健康人群中的表达存在巨大的差异,从而成为在乳腺癌乃至其他癌症中具有潜在价值的生物标志物之一,被认为是抗癌药物的可靠分子治疗靶点。本文将对HDAC5的结构表征和其在乳腺癌发生发展中的作用,以及HDAC5抑制剂的应用作一简要总结,并为早期乳腺癌HDACs的检测、HDACs抑制剂(HDACi)的设计以及与HDACi联用的相关药物作用靶点等方面提供可行性建议,以期为乳腺癌肿瘤治疗提供理论策略参考。

Breast Cancer Resistance Protein Expression and 5-Fluorouracil Resistance

Objective To filtrate breast cancer resistance protein （BCRP）-mediated resistant agents and to investigate clinical relationship between BCRP expression and drug resistance. Methods MTT assay was performed to filtrate BCRP-mediated resistant agents with BCRP expression cell model and to detect chemosensitivity of breast cancer tissue specimens to these agents. A high performance liquid chromatography （HPLC） assay was established, and was used to measure the relative dose of intracellular retention resistant agents. RT-PCR and immunohistochemistry （IHC） were employed to investigate the BCRP expression in breast cancer tissue specimens. Results MTT assay showed that the expression of BCRP increased with the increasing resistance of 5-fluorouracil （5-Fu） （P〈0.05, n=3） in the cell model, while HPLC assay indicated that the intracellular retention dose of 5-Fu was significantly correlated with the expression of BCRP （t=-0.897, P〈0.05, n=3）. A total of 140 breast cancer tissue specimens were collected. BCRP-positive expression was detected in forty-seven specimens by both RT-PCR and IHC. As shown by MTT assay subsequently, the resistance index （RI） of 47 BCRP-positive breast cancer tissue specimens to 5-Fu was 7-12 times as high as that of adjacent normal tissue samples. BCRP expression was related to 5-Fu resistance （R2=0.8124, P〈0.01）. Conclusion Resistance to 5-Fu can be mediated by BCRR Clinical chemotherapy for breast cancer patients can be optimized based on BCRP-positive expression.

Expression of Rab25 correlates with the invasion and metastasis of gastric cancer

The objective of this study was to determine the expression of the important vesicle trafficking- regulating factor Rab25 in human gastric cancer tissues, to analyze the correlation between Rab25 protein expression with gastric cancer occurrence and development, and to discuss the correlation of Rab25 protein expression with gastric cancer cell metastasis. The overall aim was to provide experimental evidence that can be used to design future biological treatments of human gastric cancer. Human gastric cancer tissue and the adjacent normal gastric tissue were surgically removed, and immunohistochemistry and Western blotting were used to detect Rab25 protein expression. The correlation between Rab25 protein expression with the development and pathological characteristics of gastric cancer was analyzed. Using RNAi, Rab25 expression was reduced in the gastric cancer cell line MGC80-3, and the changes in MGC80-3 cell invasiveness were then monitored. Immunohistochemistry showed that the Rab25 protein expression rates were 78.21% and 23.08% in gastric carcinoma and the adjacent normal gastric tissue, respectively. Immunohistochemistry and Western blot results showed that Rab25 protein expression in gastric cancer was significantly higher than in adjacent normal gastric tissues （t）〈0.01）. Less differentiated gastric cancer cells had higher expression of Rab25 protein （P〈0.01）. Gastric carcinomas from patients with a late pathological stage （Ⅲ-Ⅳ） had significantly higher Rab25 protein expression than early stage （Ⅰ-Ⅱ） patients （P〈0.01）. Gastric carcinomas from patients with lymph node metastasis had significantly higher Rab25 protein expression than lymph node metastasis- free patients （P〈0.01）. Gastric carcinomas from patients with distant metastases had significantly higher Rab25 protein expression than the distant metastasis-negative patients （P〈0.01）. Rab25 protein expression in gastric cancer was not affected by the patients＇ sex, age, or tumor size （P〉0.05）. MGCS0-3 cells transfected with Rab25 siRNA had significantly lower Rab25 protein expression （P〈0.01） and a significantly lower number of cells that passed through a Transwell chamber compared with non-transfected controls and the transfected control group （P〈0.01）. Rab25 protein expression is associated with the development of gastric cancer, siRNA knockdown of Rab25 protein expression in MGC80-3 gastric cancer cells reduced MGC80-3 cell invasiveness and provided experimental evidence for potential future biological treatment strategies of human gastric cancer.

Adipocyte-derived SFRP5 inhibits breast cancer cells migration and invasion through Wnt and epithelial-mesenchymal transition signaling pathways

Objective:Obesity is closely associated with metastasis in breast cancer patients.Secreted frizzled-related protein 5(SFRP5),one of the novel adipokines with anti-inflammatory properties,is associated with obesity.This study aims to study the role of SFRP5 in the crosstalk between obesity and breast cancer metastasis and identify the underlying mechanism.Methods:3T3-L1 pre-adipocytes were differentiated to mature adipocytes and a hypertrophic adipocyte model was induced with palmitic acid(PA).Cell motility was measured in MDA-MB-231 and MCF-7 breast cancer cells co-cultured with adipocytes conditioned medium(CM)with or without SFRP5 protein.Wnt and epithelialmesenchymal transition(EMT)signal pathways were investigated by western blot.Circulating SFRP5 level in 218 breast cancer patients and the association with clinicopathologic characteristics of breast cancer were further determined.Online databases ENCORI and PREDICT Plus were used to exam the link between SFRP5 and prognosis.Results:Reduced SFRP5 level was detected in the hypertrophic adipocyte model.Recombinant SFRP5 protein inhibited MDA-MB-231 and MCF-7 cells invasion and migration induced by PA-treated adipocyte CM,and SFRP5 inhibition by specific antibody reversed the effect of SFRP5.Furthermore,SFRP5 significantly inhibited Wnt and downstream EMT in breast cancer cells.Low circulating SFRP5 level correlated with body mass index(BMI),lymph node(LN)metastasis,TNM stage and high Ki67 expression in breast cancer patients.Increased SFRP5 level was associated with favorable predicted survival.Kaplan-Meier curves showed high SFRP5 level in tumor tissue was associated with better outcome of breast cancer patients.Conclusions:Our findings demonstrated SFRP5 is a vital adipokine that mediates the crosslink between obesity and the metastatic potential of breast cancer.Promotion of SFRP5 expression in the adipose microenvironment may represent a novel approach for preventing breast cancer metastasis.

WWOX suppresses KLF5 expression and breast cancer cell growth

The WW domain-containing oxidoreductase（WWOX） is a tumor suppressor in a variety of cancers, including breast cancer. Reduced WWOX expression is associated with the basal-like subtype and a relatively poor disease-free survival rate among breast cancer patients. Though several WWOX partners have been identified, the functional mechanisms of WWOX＇s role in cancers have not been fully addressed to date. In the current study, we found WWOX suppresses expression of KLF5—an oncogenic transcription factor—at protein level, and suppresses cancer cell proliferation in both bladder and breast cancer cell lines. Furthermore, we demonstrated that WWOX physically interacts with KLF5 via the former＇s WW domains and the latter＇s PY motifs. Interestingly, we found the expression of WWOX negatively correlates with KLF5 expression in a panel of breast cancer cell lines. Taken together, we conjecture that WWOX may suppress cancer cell proliferation partially by reducing the expression of KLF5.

Effects of 5-Aza-CdR on Cell Proliferation of Breast Cancer Cell Line MDA-MB-435S and Expression of maspin Gene

The effects of 5-Aza-2＇-deoxycytidine （5-Aza-CdR） on the proliferation of MDA-MB-435S cells and the expression of tumor suppressor gene maspin were investigated. Human breast cancer cell line MDA-MB-435S was treated with 5 μmol/L 5-Aza-CdR, a specific demethylating agent for 0 to 8 days. The growth of MDA-MB-435S cells was observed by MTT assay before and after 5-Aza-CdR treatment, respectively. The expression of maspin mRNA was detected by reverse transcfiption-polymerase chain reaction （RT-PCR）. The cell cycle of MDA-MB-435S cells was analyzed by flow cytometry. The results showed that the growth of MDA-MB-435S cells treated with 5-Aza-CdR for 8 days was significantly suppressed as compared with the control groups, and the inhibition rate increased sharply from 5 day to 8 day （35.42% to 71.29%）. Flow cytometry showed that 5 μmol/L 5-Aza-CdR could induce G2/M cell cycle arrest and decrease the percentage of mitosis cell number in this cell line. Maspin mRNA was expressed in MDA-MB-435S cells after 5-Aza-CdR treatment, but it was weakly detectable before the treatment. It was concluded that Maspin gene might be transcriptional silencing by hypermethylation and the re-expression of maspin gene by 5-Aza-CdR can inhibit the proliferation and induce the G2/M arrest of MDA-MB-435S breast cancer cells.

Predictive model for 5.year mortality after breast cancer surgery in Taiwan residents

Background:Few studies of breast cancer surgery outcomes have used longitudinal data for more than 2 years.This study aimed to validate the use of the artificial neural network(ANN)model to predict the 5?year mortality of breast cancer patients after surgery and compare predictive accuracy between the ANN model,multiple logistic regression(MLR)model,and Cox regression model.Methods:This study compared the MLR,Cox,and ANN models based on clinical data of 3632 breast cancer patients who underwent surgery between 1996 and 2010.An estimation dataset was used to train the model,and a validation dataset was used to evaluate model performance.The sensitivity analysis was also used to assess the relative signifi?cance of input variables in the prediction model.Results:The ANN model significantly outperformed the MLR and Cox models in predicting 5?year mortality,with higher overall performance indices.The results indicated that the 5?year postoperative mortality of breast cancer patients was significantly associated with age,Charlson comorbidity index(CCI),chemotherapy,radiotherapy,hormone therapy,and breast cancer surgery volumes of hospital and surgeon(all P<0.05).Breast cancer surgery volume of surgeon was the most influential(sensitive)variable affecting 5?year mortality,followed by breast cancer surgery volume of hospital,age,and CCI.Conclusions:Compared with the conventional MLR and Cox models,the ANN model was more accurate in predict?ing 5?year mortality of breast cancer patients who underwent surgery.The mortality predictors identified in this study can also be used to educate candidates for breast cancer surgery with respect to the course of recovery and health outcomes.

Direct interaction between Rab5a and Rab4a enhanced epidermal growth factor-stimulated proliferation of gastric cancer cells

BACKGROUND Gastric cancer(GC)is one of the leading causes of cancer-related death worldwide.Although targeted therapies such as antibodies against human epidermal growth factor receptor 2 or vascular endothelial growth factor receptor 2 have been widely used in the treatment of metastatic cancer,the overall outcomes are poor.Therefore,elucidation of the mechanism underlying cancer progression is important to improve prognosis.Overexpression of the Rab5a gene has been confirmed to correlate with tumorigenesis of many cancers,but the mechanism underling,especially of GC,is still unclear.AIM To investigate the effects of Rab5a overexpression on the tumorigenesis of GC.METHODS First,the expression levels of Rab5a and Rab4a in primary tumorous tissues of GC patients diagnosed between 2015 and 2018 were analyzed.Then we constructed HGC-27 cell lines overexpressing green fluorescent protein-Rab5a or red fluorescent protein-Rab4a and investigated the interaction between Rab5a or Rab4a using Western blotting,co-immunoprecipitation,confocal microscopy,and colocalization analysis.Finally,epidermal growth factor-stimulated proliferation of these cell lines was analyzed using cell counting kit-8 cell viability assay.RESULTS Compared with normal gastric tissues,the expression levels of Rab5a and Rab4a increased progressively both in paracancerous tissues and in advanced cancerous tissues.Epidermal growth factor could promote the proliferation of HGC-27 cells,especially Rab5a-overexpressing HGC-27 cells.Notably,Rab5a and Rab4a cooverexpression promoted the proliferation of HGC-27 cells to the greatest extent.Further analysis identified a direct interaction between Rab5a and Rab4a in HGC-27 cells.CONCLUSION Co-overexpression of Rab5a and Rab4a in GC may promote the endosomal recycling of epidermal growth factor receptor,which in turn contributes to poor prognosis and tumor progression in GC patients.Inhibition of Rab5a or Rab4a expression might be a promising therapy for refractory GC.

Triple Effect of Doxorubicin, 5-Fluorouracil, Propranolol on Cell Survival on MCF-7 Breast Cancer Cell Line

Purpose: Investigating the triple effect of doxorubicin, 5-fluorouracil, propranolol on MCF-7 (ER+, WTp53) breast cancer cell line with MTT test and survival analysis. Materials/Methods: In order to determine effective dosages of a combination of doxorubicin, 5-fluorouracil, propranolol on the MCF-7 cell line by using MTT and survival analysis technique. Result: IC50 values acquired by MTT tests are 0.01 mg/ml for doxorubicin, 6 mg/ml for 5-fluorouracil, 30 mg/ml for propranolol and 0.2/1/30 mg/ml (with previous respect) if all three agents are combined. It is found that the use of doxorubicin, 5-fluorouracil, and propranolol in combination is much effective than their single application. Discussion: Moderate concentrations of doxorubicin, 5-fluorouracil, and propranolol, if they are applied individually, showed high toxicity. When we used these drugs in combination;toxic effects lessened with respect to monotherapy. In the MCF-7 cell line, doxorubicin (IC50: 0.01 μM) increases cell death rates significantly and propranolol (IC50: 3 μM) has minimum effects in monotherapy in contrast to others. Propranolol is only superior to itself in combination therapy (IC50: 4 μM). However 5-fluorouracil (IC50: 30 μM) showed antagonistic effects with respect to other drugs. Additionally, having applied the three drugs in combination on the MCF-7 cell line for the first time in literature, it is highly possible to assess the application of doxorubicin, 5-fluorouracil and propranolol combination as a novel therapy option.

Embryoprotein TWIST distribution in breast cancer cells MDA-MB-231 treated with 5-Fluorouracil and malnutrition

TWIST is a transcription factor that belongs to the family of helix-loop-helix proteins involved in metastasis with essential role regulating cell movement during early development, as well as in the tumor progression and metastasis of many cancers including breast cancer. It will be interesting to study the relation among cancer chemotherapy, malnutrition and the transcription factors like TWIST in order to explore the risk to metastasis. We used breast cancer line MDA-MB-231. Cell cultures were treated with 5-Fluorouracil (5-Fu), as well as changes in serum and nonessentials amino acid (NEAA), to explore the cell viability and the cellular distribution of TWIST by immunocytochemistry. Our results indicate that cell viability decreased significantly with 5-Fu treatment whereas no changes were observed in malnutrition treatment. On the other hand, TWIST protein significantly increased its distribution in cytoplasm of treated groups with malnutrition as well as in those treated with 5-Fu compared with the control. These results suggest that TWIST translocation was modified by the treatments and further studies are necessary to suggest that TWIST could be a tag protein to avoid metastasis.

Uncoupling tumor necrosis factor-αand interleukin-10 at tumor immune microenvironment of breast cancer through miR-17-5p/MALAT-1/H19 circuit

Triple Negative Breast Cancer(TNBC)immunotherapy has recently shown promising approach.However,some TNBC patients presented with resistance.One of the reasons was attributed to the excessive release of cytokines at the tumor microenvironment(TME)such as Tumor necrosis factor alpha(TNF-α)and Interleukin-10(IL-10).Fine regulation of these cytokines’levels via non-coding RNAs(ncRNAs)might alleviate the immune quiescent nature of TME at TNBC tumors.However,the extrapolation of ncRNAs as therapeutic tools is highly challenging.Therefore,disentanglement the nature for the isolation of natural compounds that could modulate the ncRNAs and their respective targets is an applicable translational therapeutic approach.Hence,this study aimed to targeting the chief immune suppressive cytokines at the TME(TNF-αand IL-10)via ncRNAs and to examine the effects of Rosemary aerial parts extract on the expression levels of these ncRNAs in TNBC.Results revealed miR-17-5p as a dual regulator of TNF-αand IL-10.Moreover,an intricate interaction has been shown between miR-17-5p and the oncogenic lncRNAs:MALAT1 and H19.Knocking down of MALAT1 and/or H19 caused an induction in miR-17-5p and reduction in TNF-αand IL-10 expression levels.miR-17-5p was found to be down-regulated,while TNF-α,IL-10,MALAT1 and H19 were up-regulated in BC patients.Forced expression of miR-17-5p in MDA-MB-231 cells reduced TNF-α,IL-10,MALAT1 and H19 expression levels,as well as several BC hallmarks.In a translational approach,ursolic acid(UA)isolated from rosemary induced the expression of miR-17-5p,MALAT1 and decreased H19 expression levels.In conclusion,this study suggests miR-17-5p as a tumor suppressor and an immune-activator miRNA in BC through tuning up the immunological targets TNF-α,IL-10 at the TME and the oncological mediators MALAT1 and H19 lncRNAs.

前列腺癌组织肝激酶B1,Rab鸟嘌呤核苷酸交换因子-5 mRNA表达水平与临床病理特征和预后的相关性研究

目的研究前列腺癌患者癌组织中肝激酶B1(liver kinase B1,LKB1)及Rab鸟嘌呤核苷酸交换因子-5(Rab guanine nucleotide exchage factor-5,Rab EX-5)mRNA表达及临床意义。方法应用实时荧光定量PCR(qRT-PCR)检测92例前列腺癌组织和80例良性前列腺增生组织中LKB1及RabEX-5 mRNA的表达,比较LKB1,RabEX-5 mRNA表达组间差异及与临床病理特征的关系。Pearson线性相关分析LKB1与RabEX-5 mRNA表达的相关性。Kaplan-Meier生存分析LKB1,RabEX-5 mRNA表达与前列腺癌患者生存预后的关系。多因素COX回归分析影响前列腺癌患者生存预后的危险因素。结果与前列腺增生组织相比,前列腺癌组织中RabEX-5 mRNA表达明显升高(1.311±0.374 vs 0.457±0.083),差异有统计学意义(t=20.758,P=0.000),而LKB1 mRNA表达显著降低(0.373±0.052 vs 1.234±0.268),差异有统计学意义(t=30.181,P=0.000)。LKB1,RabEX-5 mRNA表达与Gleason评分、骨转移、TNM分期有关(t=2.419~9.965,均P<0.05),与年龄、前列腺特异抗原(PSA)水平无关(t=0.379~1.453,均P>0.05)。前列腺癌组织中LKB1与RabEX-5 mRNA的表达呈显著负相关(r=-0.402,P=0.000)。低LKB1表达组3年总体生存率低于高LKB1表达组(χ^(2)=55.605,P=0.000),高RabEX-5表达组3年总体生存率低于低RabEX-5表达组(χ^(2)=29.435,P=0.000)。多因素COX回归分析结果表明,Gleason评分≥7分(OR=2.671,P=0.018)、骨转移(OR=1.528,P=0.031)、TNM分期为Ⅲ期(OR=1.634,P=0.037),LKB1 mRNA低表达(OR=1.764,P=0.008)及RabEX-5 mRNA高表达(OR=2.587,P=0.000)是影响前列腺癌患者生存预后的危险因素(P<0.05)。结论前列腺癌组织中RabEX-5 mRNA表达升高,而LKB1 mRNA表达降低,二者共同参与前列腺癌的发生发展,有望成为评估前列腺癌患者预后的组织肿瘤标志物。

CASC5通过BUB1促进乳腺癌发生发展的分子机制研究

目的探究易感性癌症候选基因5(CASC5)对乳腺癌发生发展的作用及其相关机制。方法收集50例乳腺癌患者的肿瘤组织及其癌旁组织,采用qRT-PCR检测CASC5的表达水平,并分析其与患者临床病理特征的关系。通过脂质体转染法将si-CASC5、si-BUB1和si-NC转染至MDA-MB-231和MCF-7细胞,采用qRT-PCR和Western blot检测细胞CASC5、BUB1表达水平,CCK-8检测细胞增殖能力,Transwell检测细胞侵袭能力。利用String数据库分析CASC5相关蛋白,并在MCF7细胞中验证。通过免疫共沉淀检测CASC5与BUB1的结合情况,qRT-PCR验证BUB1在肿瘤组织和癌旁组织中的表达水平,及其与CASC5表达水平的相关性。结果与癌旁组织相比,肿瘤组织中CASC5表达水平升高(P<0.05),CASC5表达水平与患者TNM分期、淋巴结转移相关(P<0.05)。与si-NC组相比,si-CASC5组MDA-MB-231和MCF-7细胞中CASC5表达水平降低(P<0.05),细胞增殖能力及侵袭能力减弱(P<0.05)。生物信息学预测CASC5可能与BUB1、NSL1、NDC80和MIS12等相互作用。与si-NC组相比,si-CASC5组MDA-MB-231细胞中BUB1表达水平降低(P<0.05),NSL1、NDC80和MIS12表达水平无统计学差异(P>0.05)。CASC5免疫沉淀产物中可检测到BUB1蛋白,BUB1免疫沉淀产物中可检测到CASC5蛋白。与si-NC组相比,si-BUB1组MDA-MB-231细胞中BUB1表达水平降低(P<0.05),细胞增殖能力及侵袭能力减弱(P<0.05)。与癌旁组织相比,肿瘤组织中BUB1表达水平升高(P<0.05),CASC5表达水平与BUB1表达水平呈正相关(r=0.4815)。结论CASC5可通过与BUB1相互作用,调控BUB1蛋白表达从而促进乳腺癌发生发展。

不同临床病理特征乳腺癌患者CK5/6、EGFR、AR、Ki67表达情况及临床指导意义

目的探讨不同临床病理特征乳腺癌患者细胞角蛋白5/6(CK5/6)、表皮生长因子受体(EGFR)、雄激素受体(AR)、增殖细胞核抗原(Ki67)表达情况及临床指导意义。方法回顾性收集239例乳腺癌患者临床资料,均经病理学检查确诊,统计乳腺癌组织中不同临床病理特征CK5/6、EGFR、AR、Ki67阳性表达情况,采用Pearson相关分析CK5/6、EGFR、AR、Ki67表达情况与乳腺癌患者临床病理特征的相关性。结果乳腺癌组织中CK5/6、EGFR、AR、Ki67阳性表达率分别为15.90%、20.92%、90.38%、31.80%。Pearson相关分析显示,CK5/6与肿瘤直径呈正相关(r=0.64,P<0.05),EGFR、Ki67与肿瘤直径、组织学分级呈正相关(r分别=0.58、0.68、0.50、0.52,P均<0.05),EGFR与临床分期呈正相关(r=0.68,P<0.05),AR与组织学分级呈正相关(r=0.30,P<0.05),Ki67与淋巴结转移、临床分期呈正相关(r分别=0.48、0.49,P均<0.05)。结论随着乳腺癌患者病情进展可引起CK5/6、EGFR、AR、Ki67阳性表达升高,乳腺癌患者CK5/6、EGFR、AR、Ki67阳性表达情况与患者临床病理特征具有显著相关性,临床可通过检测乳腺癌患者CK5/6、EGFR、AR、Ki67阳性表达情况对患者病情进展程度进行评估。