Pub Date : 2015-11-18eCollection Date: 2015-01-01DOI: 10.2147/COPD.S94211
Xuesong Chen, Kouying Liu, Zhiyue Wang, Yinsu Zhu, Yang Zhao, Hui Kong, Weiping Xie, Hong Wang
Computed tomography (CT) is widely used for evaluation of lung diseases. To evaluate the value of CT measurement of pulmonary artery for diagnosis of chronic obstructive pulmonary disease (COPD) and its comorbidity pulmonary hypertension (PH), we retrospectively reviewed the CT of 221 patients with COPD and 115 control patients without cardiovascular or lung disease. Patients with COPD were divided into PH (COPD-PH) and non-PH according to systolic pulmonary artery pressure. Main pulmonary artery (MPA), right pulmonary artery (RPA) and left pulmonary artery branches, and ascending aorta (AAo) and descending aorta (DAo) diameters were measured. Meanwhile, the ratios of MPA/AAo and MPA/DAo were calculated. MPA, RPA, and left pulmonary artery diameters were significantly larger in COPD than those in the controls, and this augment was more obvious in COPD-PH. AAo and DAo diameters did not vary obviously between groups, while MPA/AAo and MAP/DAo increased significantly in COPD and PH. MPA could be helpful for COPD diagnosis (MPA diameter ≥27.5 mm, sensitivity 54%, and specificity 80%), and RPA could be applied for COPD-PH diagnosis (RPA diameter ≥23.4 mm, sensitivity 67%, and specificity 76%). There was a marked correlation between MPA/DAo and systolic pulmonary artery pressure (r=0.594, P<0.001). Therefore, chest CT could be a simple and effective modality for diagnostic evaluation of COPD and its comorbidity, PH.
{"title":"Computed tomography measurement of pulmonary artery for diagnosis of COPD and its comorbidity pulmonary hypertension.","authors":"Xuesong Chen, Kouying Liu, Zhiyue Wang, Yinsu Zhu, Yang Zhao, Hui Kong, Weiping Xie, Hong Wang","doi":"10.2147/COPD.S94211","DOIUrl":"10.2147/COPD.S94211","url":null,"abstract":"<p><p>Computed tomography (CT) is widely used for evaluation of lung diseases. To evaluate the value of CT measurement of pulmonary artery for diagnosis of chronic obstructive pulmonary disease (COPD) and its comorbidity pulmonary hypertension (PH), we retrospectively reviewed the CT of 221 patients with COPD and 115 control patients without cardiovascular or lung disease. Patients with COPD were divided into PH (COPD-PH) and non-PH according to systolic pulmonary artery pressure. Main pulmonary artery (MPA), right pulmonary artery (RPA) and left pulmonary artery branches, and ascending aorta (AAo) and descending aorta (DAo) diameters were measured. Meanwhile, the ratios of MPA/AAo and MPA/DAo were calculated. MPA, RPA, and left pulmonary artery diameters were significantly larger in COPD than those in the controls, and this augment was more obvious in COPD-PH. AAo and DAo diameters did not vary obviously between groups, while MPA/AAo and MAP/DAo increased significantly in COPD and PH. MPA could be helpful for COPD diagnosis (MPA diameter ≥27.5 mm, sensitivity 54%, and specificity 80%), and RPA could be applied for COPD-PH diagnosis (RPA diameter ≥23.4 mm, sensitivity 67%, and specificity 76%). There was a marked correlation between MPA/DAo and systolic pulmonary artery pressure (r=0.594, P<0.001). Therefore, chest CT could be a simple and effective modality for diagnostic evaluation of COPD and its comorbidity, PH. </p>","PeriodicalId":11675,"journal":{"name":"Ejc Supplements","volume":"4 1","pages":"2525-33"},"PeriodicalIF":2.8,"publicationDate":"2015-11-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4655902/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"78389513","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2015-11-01DOI: 10.1016/j.ejcsup.2015.08.012
A. Bondar , A. Kurilshikov , E. Morozkin , M. Zaripov , M. Kabilov , V. Voytsitskiy , V. Vlassov , P. Laktionov
Cancer cells display altered methylation signatures distinguishing them from normal cells. Originating from all tissues and cells of the body cell-free DNA (cfDNA) including aberrantly methylated DNA reflect epigenetic aberrations occurs not only in tumor cells but also in tumor microenvironment. Actually, aberrantly methylated cfDNA has proved to be a promising biomarker for noninvasive detection of cancer with several clinically-certified tests (Epi-pro Colon®, Epi-pro Lung®, Cologuard®). However only one test (Epi-pro Colon®) uses blood plasma – the most convenient source of cfDNA. Input of tissue and age specific methylation along with unidentified reasons lead to the presence of molecules with every conceivable cytosine-methylation patterns which decrease probability of tumor DNA identification. Among numerous variants of cfDNA methylation patterns only few reflect cancer related changes. To identify those tumor-specific profiles single nucleotide resolution of methylated cytosine locations in the individual circulating DNA molecules is obviously required.
We performed target bisulfite sequencing of potential prostate cancer (PC) cfDNA markers (GSTP1; RNF219) isolated from blood plasma of 18 healthy donors (HD), 17 benign hyperplasia (BPH) and 20 PC patients using MiSeq platform (Illumina). RNF219 gene was shown to have high diagnostic potential in our previous comparative study of cfDNA from HD, PC and BHP patients using HCGI12k microarrays (Cortese, et al., 2012). GSTP1 is a common pathological DNA methylation event in PC and is most widely studied and promising methylation marker in the cfDNA of PC patients (Wu, et al., 2011).
Selected loci were amplified after bisulfite conversion (Zymo Research) of cfDNA with methyl-independent barcoded primers and sequenced with coverage ranging from 23509 to 143953. Identification of CpG methylation status in DNA fragments was performed with the BiQ Analyzer HT Software. All statistical analysis was performed using R Statistical Software (version 3.1.1) To reveal diagnostically significant differences, several approaches to data analysis were used. Conventional approach is a prediction of patient’s diagnosis based on differences in methylation level of CpG-sites. Another approach used in this study relies on discrimination of cancer-related correlation between methylation statuses of CpG-sites within individual molecules of cfDNA – Intramolecular Correlation of Methylation Statuses (ICoMS).
Study population was randomly subsampled into training and test cohorts. The logit regression model based on methylation level of CpG-sites achieved an area under the ROC curve (AUC) exceeding 0.94 in both cohorts for GSTP1 gene and 0.81 – for RNF219 gene. A novel approach to identify diagnostic significance of cfDNA methylation was based on comparison of correlation matrices (pairwise phi coefficient between methylation statuses of CpG sites) for HD, BHP and PC groups. Binomial regression
{"title":"P124","authors":"A. Bondar , A. Kurilshikov , E. Morozkin , M. Zaripov , M. Kabilov , V. Voytsitskiy , V. Vlassov , P. Laktionov","doi":"10.1016/j.ejcsup.2015.08.012","DOIUrl":"10.1016/j.ejcsup.2015.08.012","url":null,"abstract":"<div><p>Cancer cells display altered methylation signatures distinguishing them from normal cells. Originating from all tissues and cells of the body cell-free DNA (cfDNA) including aberrantly methylated DNA reflect epigenetic aberrations occurs not only in tumor cells but also in tumor microenvironment. Actually, aberrantly methylated cfDNA has proved to be a promising biomarker for noninvasive detection of cancer with several clinically-certified tests (Epi-pro Colon®, Epi-pro Lung®, Cologuard®). However only one test (Epi-pro Colon®) uses blood plasma – the most convenient source of cfDNA. Input of tissue and age specific methylation along with unidentified reasons lead to the presence of molecules with every conceivable cytosine-methylation patterns which decrease probability of tumor DNA identification. Among numerous variants of cfDNA methylation patterns only few reflect cancer related changes. To identify those tumor-specific profiles single nucleotide resolution of methylated cytosine locations in the individual circulating DNA molecules is obviously required.</p><p>We performed target bisulfite sequencing of potential prostate cancer (PC) cfDNA markers (GSTP1; RNF219) isolated from blood plasma of 18 healthy donors (HD), 17 benign hyperplasia (BPH) and 20 PC patients using MiSeq platform (Illumina). RNF219 gene was shown to have high diagnostic potential in our previous comparative study of cfDNA from HD, PC and BHP patients using HCGI12k microarrays (Cortese, et al., 2012). GSTP1 is a common pathological DNA methylation event in PC and is most widely studied and promising methylation marker in the cfDNA of PC patients (Wu, et al., 2011).</p><p>Selected loci were amplified after bisulfite conversion (Zymo Research) of cfDNA with methyl-independent barcoded primers and sequenced with coverage ranging from 23509 to 143953. Identification of CpG methylation status in DNA fragments was performed with the BiQ Analyzer HT Software. All statistical analysis was performed using R Statistical Software (version 3.1.1) To reveal diagnostically significant differences, several approaches to data analysis were used. Conventional approach is a prediction of patient’s diagnosis based on differences in methylation level of CpG-sites. Another approach used in this study relies on discrimination of cancer-related correlation between methylation statuses of CpG-sites within individual molecules of cfDNA – Intramolecular Correlation of Methylation Statuses (ICoMS).</p><p>Study population was randomly subsampled into training and test cohorts. The logit regression model based on methylation level of CpG-sites achieved an area under the ROC curve (AUC) exceeding 0.94 in both cohorts for GSTP1 gene and 0.81 – for RNF219 gene. A novel approach to identify diagnostic significance of cfDNA methylation was based on comparison of correlation matrices (pairwise phi coefficient between methylation statuses of CpG sites) for HD, BHP and PC groups. Binomial regression","PeriodicalId":11675,"journal":{"name":"Ejc Supplements","volume":"13 1","pages":"Page 7"},"PeriodicalIF":0.0,"publicationDate":"2015-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/j.ejcsup.2015.08.012","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"54308744","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2015-11-01DOI: 10.1016/j.ejcsup.2015.08.014
O. Bryzgunova , E. Lekhnov , T. Skvortsova , E. Morozkin , I. Zaporozhchenko , A. Grigorieva , M. Zaripov , E. Ryabchikova , V. Vlassov , P. Laktionov
Differently sized microparticles, including exosomes (30–100 nm), prostasomes (50–500 nm), oncosomes (50–500 nm) and other microparticles (100–1000 nm) were found in blood and urine. Exosomes from prostate cancer (PCa) patients can potentially contain cancer-specific nucleic acids, and thus can represent a valuable source of diagnostic material. In this study, we have investigated microvesicles and miRNA from urine of healthy donors and PCa patients. To isolate miRNAs from urine and microparticles, novel methods for miRNA isolation were elaborated (Rus. patent application No 2014137763, priority date 17.09.2014). The study population included 14 patients with PCa (63–82 years, T2-3NxMx1) and control group of 20 healthy volunteers with no previous history of prostate disease (48–73 years). Urine was clarified by two serial centrifugations at 400g, 20 °C, 20 min and at 17000g, 20 °C, 20 min. Microparticles were precipitated from the resulting supernatant by high-speed centrifugation at 100000g, 18 °C, 90 min, the pellet was resuspended and pelleted by centrifugation under the same conditions. To isolate exosomes, total microparticles were filtered through 0.1 μm pore filters and reprecipitated. The resulting pellets were resuspended and exosome samples were investigated by transmission electron microscopy (TEM). MiRNAs were isolated by one-step single-phase protocol and purified using “BioSilica” spin- columns (Zaporozhchenko et al., Anal. Biochem, upcoming, doi: 10.1016/j.ab.2015.03.028) and by recently developed method based on precipitation of excess biopolymers, allowing to isolate miRNAs with better efficiency than commercially available kits. The size and quantity assessment of extracellular RNA were performed using capillary electrophoresis system on Agilent 2100 Bioanalyzer. Concentrations of miRNAs (miR19b, miR25, miR205, miR125b, miR126) were measured by qRT-PCR and normalized to miR-16 using dCq method.
TEM demonstrated the presence of 20–300 nm microparticles in urine of healthy donors and PCa patients. Approximately 50–70% of all urine microparticles are represented by 30–100 nm exosomes and residual 30–50% by particles larger than 100 nm. (The pool of urine microvesicles consists of 50–70% exosomes and 30–50% particles larger than 100 nm.).
The major part of extracellular RNA found both in exosomes and total microparticles fraction of healthy donors and patients with PCa, is 25–200 nt long and can include tRNA (73–93 n.), 5.8 rRNA (∼150 n.), snoRNK (10–20 n.), snRNA (60–300 n.) piRNAs (29–30 n.), miRNAs (20–25 n.), siRNA (21–25 n.). Concentration of extracellular urine RNA in exosomes and total microparticles of healthy donors and patients with PCa amounts to100 pg/ml of urine on av
{"title":"P67","authors":"O. Bryzgunova , E. Lekhnov , T. Skvortsova , E. Morozkin , I. Zaporozhchenko , A. Grigorieva , M. Zaripov , E. Ryabchikova , V. Vlassov , P. Laktionov","doi":"10.1016/j.ejcsup.2015.08.014","DOIUrl":"10.1016/j.ejcsup.2015.08.014","url":null,"abstract":"<div><p>Differently sized microparticles, including exosomes (30–100<!--> <!-->nm), prostasomes (50–500<!--> <!-->nm), oncosomes (50–500<!--> <!-->nm) and other microparticles (100–1000<!--> <!-->nm) were found in blood and urine. Exosomes from prostate cancer (PCa) patients can potentially contain cancer-specific nucleic acids, and thus can represent a valuable source of diagnostic material. In this study, we have investigated microvesicles and miRNA from urine of healthy donors and PCa patients. To isolate miRNAs from urine and microparticles, novel methods for miRNA isolation were elaborated (Rus. patent application No 2014137763, priority date 17.09.2014). The study population included 14 patients with PCa (63–82<!--> <!-->years, T2-3NxMx1) and control group of 20 healthy volunteers with no previous history of prostate disease (48–73<!--> <!-->years). Urine was clarified by two serial centrifugations at 400<em>g</em>, 20<!--> <!-->°C, 20<!--> <!-->min and at 17000<em>g</em>, 20<!--> <!-->°C, 20<!--> <!-->min. Microparticles were precipitated from the resulting supernatant by high-speed centrifugation at 100000<em>g</em>, 18<!--> <!-->°C, 90<!--> <!-->min, the pellet was resuspended and pelleted by centrifugation under the same conditions. To isolate exosomes, total microparticles were filtered through 0.1<!--> <!-->μm pore filters and reprecipitated. The resulting pellets were resuspended and exosome samples were investigated by transmission electron microscopy (TEM). MiRNAs were isolated by one-step single-phase protocol and purified using “BioSilica” spin- columns (Zaporozhchenko et al., Anal. Biochem, upcoming, doi: 10.1016/j.ab.2015.03.028) and by recently developed method based on precipitation of excess biopolymers, allowing to isolate miRNAs with better efficiency than commercially available kits. The size and quantity assessment of extracellular RNA were performed using capillary electrophoresis system on Agilent 2100 Bioanalyzer. Concentrations of miRNAs (miR19b, miR25, miR205, miR125b, miR126) were measured by qRT-PCR and normalized to miR-16 using dCq method.</p><p>TEM demonstrated the presence of 20–300<!--> <!-->nm microparticles in urine of healthy donors and PCa patients. Approximately 50–70% of all urine microparticles are represented by 30–100<!--> <!-->nm exosomes and residual 30–50% by particles larger than 100<!--> <!-->nm. (The pool of urine microvesicles consists of 50–70% exosomes and 30–50% particles larger than 100<!--> <!-->nm.).</p><p>The major part of extracellular RNA found both in exosomes and total microparticles fraction of healthy donors and patients with PCa, is 25–200<!--> <!-->nt long and can include tRNA (73–93<!--> <!-->n.), 5.8 rRNA (∼150 n.), snoRNK (10–20 n.), snRNA (60–300<!--> <!-->n.) piRNAs (29–30 n.), miRNAs (20–25 n.), siRNA (21–25 n.). Concentration of extracellular urine RNA in exosomes and total microparticles of healthy donors and patients with PCa amounts to100<!--> <!-->pg/ml of urine on av","PeriodicalId":11675,"journal":{"name":"Ejc Supplements","volume":"13 1","pages":"Page 8"},"PeriodicalIF":0.0,"publicationDate":"2015-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/j.ejcsup.2015.08.014","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"54308765","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2015-11-01DOI: 10.1016/j.ejcsup.2015.08.019
M. Chernyavskaya , V. Chernych , A. Efremov , V. Atamanov
Purpose
To research the characteristics of the inflammation immune disorders and cell destruction at the local level in the lacrimal and intraocular liquids in patients with choroidal melanoma.
Materials and methods
Investigation of concentration of the cytokines interleukin (IL) IL-4, IL-6, IL-8, IL-10, autoantibodies to antigens of native DNA (AAB to Ag of nDNA) in the lacrimal and intraocular liquids was performed in 36 patients (72 eyes) aged 34–83 years. The mean age of the study group patients was 60.45 years (15 males and 21 females). As standard indicators, survey results of 20 “healthy” donor lacrimal liquid – volunteers were used. For statistical data processing, descriptive statistics and methods of inter-group comparisons were used. Quantitative characteristics are presented as the median (25, 75 percentile). For comparison of quantitative traits, regression analysis was used. P value of <0.05 considered significant (95% confidence interval). Statistical analysis was performed using the software R (Team RC, 2013).
Results
A significant positive relation of concentration of IL-4, IL-6, IL-8, IL-10, AAB to Ag of nDNA in lacrimal liquid of eye with choroidal melanoma and paired “healthy” eye was reveled (p < 0.05). Concentrations of IL-6, IL-8, IL-10 and AAB to Ag of nDNA in the lacrimal liquid of the eye with choroidal melanoma were significantly higher as compared with the levels of the control group (p < 0.05). The level concentration of IL-4 in lacrimal liquid in eye with choroidal melanoma was significantly low (p < 0.05). The level concentration of IL-6 was significantly higher in the early (T1–T2) than in the later stages (T3–T4) of choroidal melanoma development (p < 0.05). All stages of choroidal melanoma were characterized by higher level of IL-6, compared with the control group (p < 0.05). Given the role of IL-6 in the pathogenesis of malignant diseases, the activity may be indicative of tumor progression in the early stages of development of choroidal melanoma. A significant increase in IL-10 was shown at stages T1 and T4 compared with the control group (p < 0.05). Regression analysis revealed a significant positive relation of concentration of IL-4, IL-6, IL-8, IL-10, AAB to Ag of nDNA between lacrimal liquid and intraocular liquids of the eye with choroidal melanoma (p < 0.05).IL-4 r = 0,84 (0.58; 1.47), p < 0.05 IL-6 r = 0.98 (0.64; 1.32), p < 0.01 IL-8 r = 0.82 (0.12; 1.73), p = 0.05 IL-10 r = 0.84 (0.01; 1.81), p = 0.05AAB to Ag of nDNA r
目的探讨脉络膜黑色素瘤患者泪液及眼内液的炎症、免疫功能紊乱及局部细胞破坏的特点。材料与方法对36例(72眼)34 ~ 83岁的泪液及眼内液中白细胞介素(IL)、IL-4、IL-6、IL-8、IL-10及自体DNA抗原抗体(AAB对nDNA抗原的抗原)的浓度进行测定。研究组患者的平均年龄为60.45岁(男性15例，女性21例)。以20名“健康”供体泪液志愿者的调查结果作为标准指标。统计数据处理采用描述性统计和组间比较的方法。数量特征以中位数(25,75百分位数)表示。数量性状比较采用回归分析。P值为<0.05认为显著(95%置信区间)。使用R软件进行统计分析(Team RC, 2013)。结果脉络膜黑色素瘤眼和配对“健康”眼泪液中IL-4、IL-6、IL-8、IL-10、AAB浓度与nDNA Ag呈显著正相关(p <0.05)。脉络膜黑色素瘤患者眼泪液中IL-6、IL-8、IL-10及nDNA中AAB与Ag的比值明显高于对照组(p <0.05)。脉络膜黑色素瘤眼泪液中IL-4水平明显降低(p <0.05)。IL-6水平浓度在脉络膜黑色素瘤早期(T1-T2)明显高于晚期(T3-T4) (p <0.05)。与对照组相比，脉络膜黑色素瘤的所有阶段均以更高水平的IL-6为特征(p <0.05)。鉴于IL-6在恶性疾病发病机制中的作用，其活性可能在脉络膜黑色素瘤发展的早期阶段指示肿瘤进展。与对照组相比，T1和T4期IL-10显著升高(p <0.05)。回归分析显示，脉络膜黑色素瘤眼泪液和眼内液中IL-4、IL-6、IL-8、IL-10、AAB浓度与nDNA Ag呈显著正相关(p <0.05)。IL-4 r = 0,84 (0.58;1.47)， p <0.05 IL-6 r = 0.98 (0.64;1.32)， p <0.01 IL-8 r = 0.82 (0.12;1.73)， IL-10 r = 0.84 (0.01;1.81)， p = 0.05AAB对Ag的nDNA r = 0.79 (0.36;1.25)， p <0.05.Conclusions1。我们发现，在脉络膜黑色素瘤的发病过程中，局部炎症起着重要的作用，表现为IL-6、IL-8浓度明显升高，而IL-4水平较低，表达免疫功能紊乱，细胞破坏，表现为nDNA中AAB - Ag水平较对照组显著升高。泪液适用于脉络膜黑色素瘤患者局部炎症、免疫功能紊乱和细胞破坏的研究。在脉络膜黑色素瘤的所有阶段，泪液中IL-6水平均显著升高，但在早期(T1-T2)比晚期(T3-T4)更为明显。在脉络膜黑色素瘤的机制中，IL-10的显著升高可能提示肿瘤生长引起的局部免疫抑制的严重程度。在脉络膜黑色素瘤患者中，眼泪液中IL-4、IL-6、IL-8、IL-10、nDNA中AAB与Ag的浓度与配对的“健康”眼呈显著正相关。
{"title":"P132","authors":"M. Chernyavskaya , V. Chernych , A. Efremov , V. Atamanov","doi":"10.1016/j.ejcsup.2015.08.019","DOIUrl":"10.1016/j.ejcsup.2015.08.019","url":null,"abstract":"<div><h3>Purpose</h3><p>To research the characteristics of the inflammation immune disorders and cell destruction at the local level in the lacrimal and intraocular liquids in patients with choroidal melanoma.</p></div><div><h3>Materials and methods</h3><p>Investigation of concentration of the cytokines interleukin (IL) IL-4, IL-6, IL-8, IL-10, autoantibodies to antigens of native DNA (AAB to Ag of nDNA) in the lacrimal and intraocular liquids was performed in 36 patients (72 eyes) aged 34–83<!--> <!-->years. The mean age of the study group patients was 60.45<!--> <!-->years (15 males and 21 females). As standard indicators, survey results of 20 “healthy” donor lacrimal liquid – volunteers were used. For statistical data processing, descriptive statistics and methods of inter-group comparisons were used. Quantitative characteristics are presented as the median (25, 75 percentile). For comparison of quantitative traits, regression analysis was used. <em>P</em> value of <0.05 considered significant (95% confidence interval). Statistical analysis was performed using the software R (Team RC, 2013).</p></div><div><h3>Results</h3><p>A significant positive relation of concentration of IL-4, IL-6, IL-8, IL-10, AAB to Ag of nDNA in lacrimal liquid of eye with choroidal melanoma and paired “healthy” eye was reveled (<em>p</em> <!--><<!--> <!-->0.05). Concentrations of IL-6, IL-8, IL-10 and AAB to Ag of nDNA in the lacrimal liquid of the eye with choroidal melanoma were significantly higher as compared with the levels of the control group (<em>p</em> <!--><<!--> <!-->0.05). The level concentration of IL-4 in lacrimal liquid in eye with choroidal melanoma was significantly low (<em>p</em> <!--><<!--> <!-->0.05). The level concentration of IL-6 was significantly higher in the early (T1–T2) than in the later stages (T3–T4) of choroidal melanoma development (<em>p</em> <!--><<!--> <!-->0.05). All stages of choroidal melanoma were characterized by higher level of IL-6, compared with the control group (<em>p</em> <!--><<!--> <!-->0.05). Given the role of IL-6 in the pathogenesis of malignant diseases, the activity may be indicative of tumor progression in the early stages of development of choroidal melanoma. A significant increase in IL-10 was shown at stages T1 and T4 compared with the control group (<em>p</em> <!--><<!--> <!-->0.05). Regression analysis revealed a significant positive relation of concentration of IL-4, IL-6, IL-8, IL-10, AAB to Ag of nDNA between lacrimal liquid and intraocular liquids of the eye with choroidal melanoma (<em>p</em> <!--><<!--> <!-->0.05).IL-4 <em>r</em> <!-->=<!--> <!-->0,84 (0.58; 1.47), <em>p</em> <!--><<!--> <!-->0.05 IL-6 <em>r</em> <!-->=<!--> <!-->0.98 (0.64; 1.32), <em>p</em> <!--><<!--> <!-->0.01 IL-8 <em>r</em> <!-->=<!--> <!-->0.82 (0.12; 1.73), <em>p</em> <!-->=<!--> <!-->0.05 IL-10 <em>r</em> <!-->=<!--> <!-->0.84 (0.01; 1.81), <em>p</em> <!-->=<!--> <!-->0.05AAB to Ag of nDNA <em>r</e","PeriodicalId":11675,"journal":{"name":"Ejc Supplements","volume":"13 1","pages":"Page 11"},"PeriodicalIF":0.0,"publicationDate":"2015-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/j.ejcsup.2015.08.019","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"54308865","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2015-11-01DOI: 10.1016/j.ejcsup.2015.08.029
M. Freidin
Blood-derived biomarkers, such as circulating tumour cells (CTCs) and circulating tumour DNA (ctDNA), are a valuable source of molecular genetic data for tumours they spring from. In translational cancer research, the “liquid biopsy” concept has been put forward to denote the detection and molecular characterization of these biomarkers.
The idea of liquid biopsy is based on a hypothesis that profiles of somatic mutations in CTCs and ctDNA are identical to those in the original tumour. Therefore, the mutation status of the source tumour can be revealed through the molecular analysis of the CTCs and ctDNA obtained from the blood. The major advantages of liquid biopsy are an essentially decreased invasiveness (no need for tissue biopsy, surgery or bronchoscopy) and an ability to carry out the analysis at patient’s follow up (e.g. to monitor for residual disease).
However, both the CTCs and ctDNA are not abundant in the bloodstream and their capture is technically challenging. Also, due to tumour heterogeneity, the CTCs may not fully represent the entire tumour, while ctDNA is naturally fragmented and degraded, so its utility for genetic analysis may be limited. Finally, the DNA extracted from CTCs and, especially, ctDNA are “contaminated” by DNA from non-tumour cells from the bloodstream raising a challenge of detecting mutant DNA among significantly prevailing wild-type DNA.
Some of these issues can be overcome by using such advanced techniques as BEAMing, digital PCR or ultra-deep sequencing, but their use in standard clinical settings is limited by the need for special equipment and associated costs. Inexpensive and less sophisticated, but still highly sensitive and specific, approaches, such as COLD-PCR or wild-type blocking PCR, are also available to detect “druggable” mutations in CTCs and ctDNA.
Application of these approaches of liquid biopsy in clinical practice may be highly beneficial for personalized care of lung cancer patients.
{"title":"T33","authors":"M. Freidin","doi":"10.1016/j.ejcsup.2015.08.029","DOIUrl":"10.1016/j.ejcsup.2015.08.029","url":null,"abstract":"<div><p>Blood-derived biomarkers, such as circulating tumour cells (CTCs) and circulating tumour DNA (ctDNA), are a valuable source of molecular genetic data for tumours they spring from. In translational cancer research, the “liquid biopsy” concept has been put forward to denote the detection and molecular characterization of these biomarkers.</p><p>The idea of liquid biopsy is based on a hypothesis that profiles of somatic mutations in CTCs and ctDNA are identical to those in the original tumour. Therefore, the mutation status of the source tumour can be revealed through the molecular analysis of the CTCs and ctDNA obtained from the blood. The major advantages of liquid biopsy are an essentially decreased invasiveness (no need for tissue biopsy, surgery or bronchoscopy) and an ability to carry out the analysis at patient’s follow up (e.g. to monitor for residual disease).</p><p>However, both the CTCs and ctDNA are not abundant in the bloodstream and their capture is technically challenging. Also, due to tumour heterogeneity, the CTCs may not fully represent the entire tumour, while ctDNA is naturally fragmented and degraded, so its utility for genetic analysis may be limited. Finally, the DNA extracted from CTCs and, especially, ctDNA are “contaminated” by DNA from non-tumour cells from the bloodstream raising a challenge of detecting mutant DNA among significantly prevailing wild-type DNA.</p><p>Some of these issues can be overcome by using such advanced techniques as BEAMing, digital PCR or ultra-deep sequencing, but their use in standard clinical settings is limited by the need for special equipment and associated costs. Inexpensive and less sophisticated, but still highly sensitive and specific, approaches, such as COLD-PCR or wild-type blocking PCR, are also available to detect “druggable” mutations in CTCs and ctDNA.</p><p>Application of these approaches of liquid biopsy in clinical practice may be highly beneficial for personalized care of lung cancer patients.</p></div>","PeriodicalId":11675,"journal":{"name":"Ejc Supplements","volume":"13 1","pages":"Page 16"},"PeriodicalIF":0.0,"publicationDate":"2015-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/j.ejcsup.2015.08.029","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"54309046","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2015-11-01DOI: 10.1016/j.ejcsup.2015.08.031
T. Gening, T. Abakumova, D. Dolgova, S. Gening, I. Antoneeva
Tumor-associated neutrophils (TANs) are the cell population that differs in morphofunctional characteristics from peripheral blood cells (Gregory and Houghton, 2011). The first study to identify the presence of TANs as an independent poor prognostic factor and to include TANs into a prognostic risk model was published in 2006 (Donskov , 2006). Specific signals during cancer progression have been shown to induce the emergence of a pro-tumor phenotype of neutrophils (PMN). Frinlender (2013) points out an anti-tumor phenotype of TANs. Of particular interest is the study of interaction of neutrophils with T cells because the latter are considered to be cytotoxic cells mediating antitumor immunity. The aim of the study was to assess the lymphoid cell infiltration (LCI) and the functional status of TANs in ovarian cancer. LCI was assessed by immunohistochemistry and the levels of mieloproxydase (MPO) and cationic proteins (CP) were evaluated by cytochemical methods in ovarian carcinoma surgical resection specimens. The results were presented as a mean cytochemical coefficient (MCC). The intensity of nitroblue tetrazolium (NBT) test was expressed as a percentage. Obtained results were analyzed by nonparametric statistical methods. The Kruskal–Wallis test was used to evaluate the differences between groups.
We established that low intensity of infiltration in general and formation of marginal lymphoid cell ridge in some tumors are typical for malignant ovarian neoplasms. More intensive infiltration was found in the regions of the rapid growth of tumor cells, i.e. invasion zone. In some cases, a group of tumor cells was separated from the main part of the parenchyma and surrounded by lymphoid elements. Conditions are created under which active spread of cancer cells in the parenchyma is associated with intensive lymphoid cells infiltration in the stroma. Total count and density of lymphoid cells can significantly vary: from single lymphoid elements uniformly scattered in the tumor tissue to focal accumulations. In tumor tissue location of lymphoid cells is irregular, so most of the infiltration is in the stroma and among the cancer cells only solitary lymphocytes are found. Lymphoid cells which make up the infiltration include 12.3 ± 1.52% of macrophages, 22.1 ± 2.3% of plasma cells, 62.5 ± 1.1% of lymphocytes and 3.1 ± 0.9% of PMN. Total amount of cells was 224.1 ± 32.8 per 1 mm2 of tumor.
Lymphocytes in ovarian carcinomas do not form large clusters; they are disposed among other lymphoid cells in the stroma on the periphery of the tumor. Plasma cells have a classical morphological structure and often locate in the stroma and on the periphery of the tumor. Macrophages are an integral part of the tumor infiltrate. In the tumor they become stretched and angular. Macrophages usually have large dimensions, and they are
{"title":"A18","authors":"T. Gening, T. Abakumova, D. Dolgova, S. Gening, I. Antoneeva","doi":"10.1016/j.ejcsup.2015.08.031","DOIUrl":"10.1016/j.ejcsup.2015.08.031","url":null,"abstract":"<div><p>Tumor-associated neutrophils (TANs) are the cell population that differs in morphofunctional characteristics from peripheral blood cells (Gregory and Houghton, 2011). The first study to identify the presence of TANs as an independent poor prognostic factor and to include TANs into a prognostic risk model was published in 2006 (Donskov , 2006). Specific signals during cancer progression have been shown to induce the emergence of a pro-tumor phenotype of neutrophils (PMN). Frinlender (2013) points out an anti-tumor phenotype of TANs. Of particular interest is the study of interaction of neutrophils with T cells because the latter are considered to be cytotoxic cells mediating antitumor immunity. The aim of the study was to assess the lymphoid cell infiltration (LCI) and the functional status of TANs in ovarian cancer. LCI was assessed by immunohistochemistry and the levels of mieloproxydase (MPO) and cationic proteins (CP) were evaluated by cytochemical methods in ovarian carcinoma surgical resection specimens. The results were presented as a mean cytochemical coefficient (MCC). The intensity of nitroblue tetrazolium (NBT) test was expressed as a percentage. Obtained results were analyzed by nonparametric statistical methods. The Kruskal–Wallis test was used to evaluate the differences between groups.</p><p>We established that low intensity of infiltration in general and formation of marginal lymphoid cell ridge in some tumors are typical for malignant ovarian neoplasms. More intensive infiltration was found in the regions of the rapid growth of tumor cells, i.e. invasion zone. In some cases, a group of tumor cells was separated from the main part of the parenchyma and surrounded by lymphoid elements. Conditions are created under which active spread of cancer cells in the parenchyma is associated with intensive lymphoid cells infiltration in the stroma. Total count and density of lymphoid cells can significantly vary: from single lymphoid elements uniformly scattered in the tumor tissue to focal accumulations. In tumor tissue location of lymphoid cells is irregular, so most of the infiltration is in the stroma and among the cancer cells only solitary lymphocytes are found. Lymphoid cells which make up the infiltration include 12.3<!--> <!-->±<!--> <!-->1.52% of macrophages, 22.1<!--> <!-->±<!--> <!-->2.3% of plasma cells, 62.5<!--> <!-->±<!--> <!-->1.1% of lymphocytes and 3.1<!--> <!-->±<!--> <!-->0.9% of PMN. Total amount of cells was 224.1<!--> <!-->±<!--> <!-->32.8 per 1<!--> <!-->mm<sup>2</sup> of tumor.</p><p>Lymphocytes in ovarian carcinomas do not form large clusters; they are disposed among other lymphoid cells in the stroma on the periphery of the tumor. Plasma cells have a classical morphological structure and often locate in the stroma and on the periphery of the tumor. Macrophages are an integral part of the tumor infiltrate. In the tumor they become stretched and angular. Macrophages usually have large dimensions, and they are ","PeriodicalId":11675,"journal":{"name":"Ejc Supplements","volume":"13 1","pages":"Pages 17-18"},"PeriodicalIF":0.0,"publicationDate":"2015-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/j.ejcsup.2015.08.031","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"54309086","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2015-11-01DOI: 10.1016/j.ejcsup.2015.08.033
E. Grigorieva , M. Karbyshev , V. Volkomorov , E. Kremmer , A. Huber , I. Mitrofanova , M. Zavyalova , E. Kaigorodova , J. Kzhyshkowska , N. Cherdyntseva
Background
Transmembrane prostate androgen-induced protein 1 (TMEPAI) is a membrane protein that has attracted significant attention of many researchers its involvement in TGF-β signaling pathway which involved in malignant transformation and metastatic tumor progression. We investigated the TMEPAI expression level in gastric adenocarcinomas in comparison to non-tumor mucosa samples and determined its potential prognostic significance.
Materials and methods
Fresh and paraffin-embedded gastric adenocarcinoma samples and paired adjacent normal tissues were collected from gastric cancer patients. Evaluation of the PMEPA 1 gene expression was carried out using RT-PCR. For evaluation of TMEPAI protein expression, monoclonal antibodies (mAbs) were developed by using hybridoma techniques. Specificity of prepared monoclonal antibodies against recombinant TMEPAI and evaluation of its expression in the clinical samples using selected mAbs were performed using immunoblotting and immunohistochemistry.
Results
We have identified more than two-fold increase in gene expression of PMEPA1 in tumor tissue in 44% of patients. The monoclonal antibodies have shown the capacity to specifically recognize the recombinant TMEPAI in HEK293T cell lysates. We also evaluate the ability of the selected antibodies to recognize the target protein in fixed cells by immunocytochemistry. The evaluation of TMEPAI in adenocarcinoma samples collected from gastric cancer patients revealed decreased protein expression. We have observed pronounced expression of TMEPAI in normal gastric epithelial cells, while tumor cells from gastric adenomas and adenocarcinomas samples were mostly negative for target protein expression. We found that gastric epithelium cells lose the TMEPAI expression concurrent with severe dysplasia.
Conclusion
Apparently, the TMEPAI may be a potential biomarker of malignant transformation risk of the stomach epithelium.
The presented study was financially supported by Grants from the Russian Fund for Basic Research (14-04-31500) and Tomsk State University Competitiveness Improvement Program.
{"title":"P68","authors":"E. Grigorieva , M. Karbyshev , V. Volkomorov , E. Kremmer , A. Huber , I. Mitrofanova , M. Zavyalova , E. Kaigorodova , J. Kzhyshkowska , N. Cherdyntseva","doi":"10.1016/j.ejcsup.2015.08.033","DOIUrl":"10.1016/j.ejcsup.2015.08.033","url":null,"abstract":"<div><h3>Background</h3><p>Transmembrane prostate androgen-induced protein 1 (TMEPAI) is a membrane protein that has attracted significant attention of many researchers its involvement in TGF-<em>β</em> signaling pathway which involved in malignant transformation and metastatic tumor progression. We investigated the TMEPAI expression level in gastric adenocarcinomas in comparison to non-tumor mucosa samples and determined its potential prognostic significance.</p></div><div><h3>Materials and methods</h3><p>Fresh and paraffin-embedded gastric adenocarcinoma samples and paired adjacent normal tissues were collected from gastric cancer patients. Evaluation of the PMEPA 1 gene expression was carried out using RT-PCR. For evaluation of TMEPAI protein expression, monoclonal antibodies (mAbs) were developed by using hybridoma techniques. Specificity of prepared monoclonal antibodies against recombinant TMEPAI and evaluation of its expression in the clinical samples using selected mAbs were performed using immunoblotting and immunohistochemistry.</p></div><div><h3>Results</h3><p>We have identified more than two-fold increase in gene expression of PMEPA1 in tumor tissue in 44% of patients. The monoclonal antibodies have shown the capacity to specifically recognize the recombinant TMEPAI in HEK293T cell lysates. We also evaluate the ability of the selected antibodies to recognize the target protein in fixed cells by immunocytochemistry. The evaluation of TMEPAI in adenocarcinoma samples collected from gastric cancer patients revealed decreased protein expression. We have observed pronounced expression of TMEPAI in normal gastric epithelial cells, while tumor cells from gastric adenomas and adenocarcinomas samples were mostly negative for target protein expression. We found that gastric epithelium cells lose the TMEPAI expression concurrent with severe dysplasia.</p></div><div><h3>Conclusion</h3><p>Apparently, the TMEPAI may be a potential biomarker of malignant transformation risk of the stomach epithelium.</p><p>The presented study was financially supported by Grants from the <span>Russian Fund for Basic Research</span> (<span>14-04-31500</span>) and Tomsk State University Competitiveness Improvement Program.</p></div>","PeriodicalId":11675,"journal":{"name":"Ejc Supplements","volume":"13 1","pages":"Pages 18-19"},"PeriodicalIF":0.0,"publicationDate":"2015-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/j.ejcsup.2015.08.033","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"54309115","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2015-11-01DOI: 10.1016/j.ejcsup.2015.08.053
T. Kunts , K. Karpukhina , E. Mikhailova , N. Varaksin , A. Autenshlyus
Growing tumor and its microenvironment are capable to produce a number of cytokines that alter the nature of the antitumor surveillance by host immune system.
Objective
Comparative evaluation of cytokine-producing function of the invasive ductal carcinoma and fibroadenoma of the breast in vitro.
Materials and methods
Similar biopsies (V = 8 mm3) of the breast tumors were obtained using a special device, cultivated in DMEM F-12 at 37 °C for 72 h. Concentrations of the following cytokines: IL-1β, IL-1Ra, TNFα, IL-2, IL-6, IL-8, IL-10, IL-17, IL-18, VEGF and IFNγ in the supernatant of the tumor were measured with enzyme-linked immunosorbent assay (ELISA).
Results
The investigation of cytokines level in the supernatant of malignant and benign breast tumors revealed significant differences only in concentrations of IL-10, IL-17, IL-18 and IFNγ which had a contrary tendency. For example, the concentration of IL-10 was lower at invasive ductal carcinoma in comparison with fibroadenomas. The concentrations of IL-17, IL-18 and IFNγ at invasive ductal carcinoma were significantly higher than those of breast fibroadenomas. IL-17 and IL-18 are known to be pro-oncogenic cytokines, and the higher the level, the higher the severity of tumor progression. Reduction in the IL-10 concentration might be explained by the already formed neoplasm, which depends on angiogenesis. In this case, IL-10 no longer exerts antiangiogenic action, which contributes to tumor progression. Reduction in the IL-10 concentration, which inhibits the production of IFNγ leads to an increase in the IFNγ level. In early stages of tumor development, IFNγ provides an antitumor effect and at the same time facilitates the selection of a more malignant clones but its pro-tumoral action predominates at advanced stages of tumorigenesis. Moreover, higher concentration of IFNγ is supposed to be associated with biological effects of IL-18, which is its immediate inductor. In addition, malignant tumor cells are capable to produce their own IL-18, which stimulates tumor progression and facilitates the migration of endothelial cells involved in angiogenesis, which leads to intensified invasion and metastasis.
Conclusion
Cytokine production in supernatants of invasive ductal carcinoma compared with fibroadenoma of the breast is characterized by increase in IL-17, IL-18 and IFNγ concentrations and decrease in IL-10 concentration. Findings suggest the ability of malignant tumor and its microenvironment to secrete the pro-oncogenic cytokines. Fibroadenoma is also able to produce cytokines due to fibroblasts, fibrocytes and some leukocytes in its content.
{"title":"P64","authors":"T. Kunts , K. Karpukhina , E. Mikhailova , N. Varaksin , A. Autenshlyus","doi":"10.1016/j.ejcsup.2015.08.053","DOIUrl":"10.1016/j.ejcsup.2015.08.053","url":null,"abstract":"<div><p>Growing tumor and its microenvironment are capable to produce a number of cytokines that alter the nature of the antitumor surveillance by host immune system.</p></div><div><h3>Objective</h3><p>Comparative evaluation of cytokine-producing function of the invasive ductal carcinoma and fibroadenoma of the breast in vitro.</p></div><div><h3>Materials and methods</h3><p>Similar biopsies (<em>V</em> <!-->=<!--> <!-->8<!--> <!-->mm<sup>3</sup>) of the breast tumors were obtained using a special device, cultivated in DMEM F-12 at 37<!--> <!-->°C for 72<!--> <!-->h. Concentrations of the following cytokines: IL-1<em>β</em>, IL-1Ra, TNF<em>α</em>, IL-2, IL-6, IL-8, IL-10, IL-17, IL-18, VEGF and IFN<em>γ</em> in the supernatant of the tumor were measured with enzyme-linked immunosorbent assay (ELISA).</p></div><div><h3>Results</h3><p>The investigation of cytokines level in the supernatant of malignant and benign breast tumors revealed significant differences only in concentrations of IL-10, IL-17, IL-18 and IFN<em>γ</em> which had a contrary tendency. For example, the concentration of IL-10 was lower at invasive ductal carcinoma in comparison with fibroadenomas. The concentrations of IL-17, IL-18 and IFN<em>γ</em> at invasive ductal carcinoma were significantly higher than those of breast fibroadenomas. IL-17 and IL-18 are known to be pro-oncogenic cytokines, and the higher the level, the higher the severity of tumor progression. Reduction in the IL-10 concentration might be explained by the already formed neoplasm, which depends on angiogenesis. In this case, IL-10 no longer exerts antiangiogenic action, which contributes to tumor progression. Reduction in the IL-10 concentration, which inhibits the production of IFN<em>γ</em> leads to an increase in the IFN<em>γ</em> level. In early stages of tumor development, IFN<em>γ</em> provides an antitumor effect and at the same time facilitates the selection of a more malignant clones but its pro-tumoral action predominates at advanced stages of tumorigenesis. Moreover, higher concentration of IFN<em>γ</em> is supposed to be associated with biological effects of IL-18, which is its immediate inductor. In addition, malignant tumor cells are capable to produce their own IL-18, which stimulates tumor progression and facilitates the migration of endothelial cells involved in angiogenesis, which leads to intensified invasion and metastasis.</p></div><div><h3>Conclusion</h3><p>Cytokine production in supernatants of invasive ductal carcinoma compared with fibroadenoma of the breast is characterized by increase in IL-17, IL-18 and IFN<em>γ</em> concentrations and decrease in IL-10 concentration. Findings suggest the ability of malignant tumor and its microenvironment to secrete the pro-oncogenic cytokines. Fibroadenoma is also able to produce cytokines due to fibroblasts, fibrocytes and some leukocytes in its content.</p></div>","PeriodicalId":11675,"journal":{"name":"Ejc Supplements","volume":"13 1","pages":"Page 30"},"PeriodicalIF":0.0,"publicationDate":"2015-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/j.ejcsup.2015.08.053","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"54309560","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2015-11-01DOI: 10.1016/j.ejcsup.2015.08.065
D. Meshalkina, M. Shevtsov, B. Margulis, I. Guzhova
Glioblastoma is one of the most malignant cancer types. Its median survival is 15 months with combined radio- and chemotherapy and only 4 months without therapy. Molecular chaperones play a very multifaced role in tumor development. Depletion of Hdj1 in cancer cells accelerates tumor growth. Decrease of Hdj2 level correlates with the increase of tumor aggressiveness, but also attenuates tumor protection against radiotherapy. Hsp70 provides considerable survival advantages to the cancer cells, but at the same time can act as a “chaperokine”, activating antitumoral immunity. For assessment of chaperone’s role in glioma progression, invasiveness and metastasis formation we chose rat model of intracranial injection of 105 C6 cells. We developed three C6-based cell lines with protein knock-down by RNA-interference: C6 shHsp70 (on 83%), C6 shHdj1 (on 96%) and C6 shHdj2 (on 52%). The last differed in roundish and easily detachable morphology. Following intracranial injection of the modified tumor cells, the animals’ survival was estimated. As compared to the groups of control C6 (25.4 ± 3.9 days) and C6 shHdj1 (25.5 ± 3.8) we observed a nearly 1.5-fold decrease in survival in C6 shHdj2 (16.8 ± 3.5 days) (P < 0.05). On the contrary, in C6 shHsp70 group the survival increased up to 42.5 ± 12.0 days (the increase is completely explainable by the slower growth rate of the culture). Subsequent MR imaging and histological analysis of tumors demonstrated elevated invasiveness and metastatic activity in C6 shHdj2 group in comparison to C6 shHsp70, C6 shHdj1 and control C6. High migration activity and the ability of floating C6 shHdj2 cells to adhere and settle on the substrate was proved in wound-healing assay, spot-healing assay, colony forming assay and transwell migration assay. Adhesion assay showed decreased adhesion ability of C6 shHdj2 cells and increased – of C6 shHsp70 cells on all types of tested extracellular matrixes. Immunofluorescence analyses showed loss of membrane- expressed N-cadherin and loss of intercellular contacts mediated by N-cadherin in C6 shHdj2 cells in comparison to other considered cell lines (although its level in western blot was elevated). Actin staining with rhodamine-falloidin revealed highly abundant leading edges in C6 shHdj2 culture. Matrix metalloprotease zymography proved an increased activity in gelatinases (mmp2 and mmp9) as well as in caseinases (mmp1 and mmp8) in C6 shHdj2 culture supernatant. Assay of stemness marker CD133 expression showed its 11.8 times increase in C6 shHdj2.
Our experiments proved the high importance of Hdj2 level in glioma progression, invasion and metastasis.
{"title":"P56","authors":"D. Meshalkina, M. Shevtsov, B. Margulis, I. Guzhova","doi":"10.1016/j.ejcsup.2015.08.065","DOIUrl":"10.1016/j.ejcsup.2015.08.065","url":null,"abstract":"<div><p>Glioblastoma is one of the most malignant cancer types. Its median survival is 15<!--> <!-->months with combined radio- and chemotherapy and only 4<!--> <!-->months without therapy. Molecular chaperones play a very multifaced role in tumor development. Depletion of Hdj1 in cancer cells accelerates tumor growth. Decrease of Hdj2 level correlates with the increase of tumor aggressiveness, but also attenuates tumor protection against radiotherapy. Hsp70 provides considerable survival advantages to the cancer cells, but at the same time can act as a “chaperokine”, activating antitumoral immunity. For assessment of chaperone’s role in glioma progression, invasiveness and metastasis formation we chose rat model of intracranial injection of 10<sup>5</sup> C6 cells. We developed three C6-based cell lines with protein knock-down by RNA-interference: C6 shHsp70 (on 83%), C6 shHdj1 (on 96%) and C6 shHdj2 (on 52%). The last differed in roundish and easily detachable morphology. Following intracranial injection of the modified tumor cells, the animals’ survival was estimated. As compared to the groups of control C6 (25.4<!--> <!-->±<!--> <!-->3.9<!--> <!-->days) and C6 shHdj1 (25.5<!--> <!-->±<!--> <!-->3.8) we observed a nearly 1.5-fold decrease in survival in C6 shHdj2 (16.8<!--> <!-->±<!--> <!-->3.5<!--> <!-->days) (<em>P</em> <!--><<!--> <!-->0.05). On the contrary, in C6 shHsp70 group the survival increased up to 42.5<!--> <!-->±<!--> <!-->12.0<!--> <!-->days (the increase is completely explainable by the slower growth rate of the culture). Subsequent MR imaging and histological analysis of tumors demonstrated elevated invasiveness and metastatic activity in C6 shHdj2 group in comparison to C6 shHsp70, C6 shHdj1 and control C6. High migration activity and the ability of floating C6 shHdj2 cells to adhere and settle on the substrate was proved in wound-healing assay, spot-healing assay, colony forming assay and transwell migration assay. Adhesion assay showed decreased adhesion ability of C6 shHdj2 cells and increased – of C6 shHsp70 cells on all types of tested extracellular matrixes. Immunofluorescence analyses showed loss of membrane- expressed N-cadherin and loss of intercellular contacts mediated by N-cadherin in C6 shHdj2 cells in comparison to other considered cell lines (although its level in western blot was elevated). Actin staining with rhodamine-falloidin revealed highly abundant leading edges in C6 shHdj2 culture. Matrix metalloprotease zymography proved an increased activity in gelatinases (mmp2 and mmp9) as well as in caseinases (mmp1 and mmp8) in C6 shHdj2 culture supernatant. Assay of stemness marker CD133 expression showed its 11.8 times increase in C6 shHdj2.</p><p>Our experiments proved the high importance of Hdj2 level in glioma progression, invasion and metastasis.</p></div>","PeriodicalId":11675,"journal":{"name":"Ejc Supplements","volume":"13 1","pages":"Pages 36-37"},"PeriodicalIF":0.0,"publicationDate":"2015-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/j.ejcsup.2015.08.065","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"54309742","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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