Detection Of Survivin mRNA Research Articles

Engineering of catalytic hairpin-rigidified Y-shaped DNA-functionalized nanomachine to rapidly detect mRNA.

The catalytic hairpin-rigidified Y-shaped DNA through layer-by-layer assembly has beenfixed on the surface of copper sulfide nanoparticles for the detection of survivin mRNA. The distance between the CHA probes fixed on the Y-shaped DNA is significantly shortened. The results show that the fluorescence of this nanomachine reached the maximum value in 50 min (excitation wavelength at 488 nm and emission wavelength 526 nm), and its reaction rate is more than 5-fold faster than that of the free-CHA control system. In addition, the nanomachine showed high sensitivity (LOD of 3.5 pM) and high specificity for the survivin mRNA detection. Given its fast response time and excellent detection performance, we envision that the catalytic hairpin-rigidified Y-shaped DNA-functionalized nanomachine will offer potential applications in disease diagnostics and clinical applications.

A novel binding-induced DNAzyme motor triggered by survivin mRNA.

The accurate and sensitive detection of survivin mRNA is of great significance for cancer diagnosis and treatment. However, limited by the low-abundance mRNA in live cells, most strategies of survivin mRNA detection that were one-to-one signal-triggered model (one target triggered one signal) were inapplicable in practice. Here, we reported a binding-induced DNAzyme motor triggered by the survivin mRNA, which was a one-to-more signal-triggered model (one target triggered more signals), amplifying the detection signal and enhancing the sensitivity. The nanomotor is constructed by assembling several DNAzyme motor strands silenced by the blocker strands, and dozens of FAM-labeled substrate strands on a single gold nanoparticle (AuNP), forming three-dimensional DNA tracks. Through building the survivin mRNA bridge between the blocker and the DNAzyme motor strand, the binding-induced DNA nanomotor could be triggered by survivin mRNA. The operation of the DNAzyme motor was self-powered. And each walking step of the DNAzyme motor was fueled by DNAzyme-catalyzed substrate cleavage, along with the cleavage of the fluorescent molecule, resulting in autonomous and progressive walking along the AuNP-based tracks, and the fluorescence increase. The DNAzyme motor exhibited excellent sensitivity and remarkable specificity for survivin mRNA, providing the potential for cell image.

Diagnostic accuracy of urinary survivin mRNA expression detected by RT-PCR compared with urine cytology in the detection of bladder cancer: A meta-analysis of diagnostic test accuracy in head-to-head studies.

Survivin is a promising marker for the diagnosis of bladder cancer. The accuracy and clinical value of urinary survivin mRNA expression were compared with urine cytology, which is the standard diagnostic method for bladder cancer. Scientific databases, including PubMed, Web of Science, Cochrane Library and China National Knowledge Infrastructure, were searched in order to find studies that examined urinary survivin mRNA expression and urine cytology in the diagnosis of bladder cancer. Quality assessment was performed using the Quality Assessment of Diagnostic Accuracy Studies 2 tool in Revman 5.3 and data analysis was conducted using Stata/MP. The I2 statistic was used to evaluate heterogeneity and Deeks' funnel plot was generated to assess the possibility of publication bias. A total of 15 studies that evaluated a total of 1,624 patients were included in the present meta-analysis. The pooled sensitivity and specificity values for the detection of urinary survivin mRNA expression in the diagnosis of bladder cancer were 0.86 [95% confidence interval (CI), 0.81–0.90] and 0.95 (95% CI, 0.93–0.96), respectively. Regarding urine cytology, the pooled sensitivity and specificity values were 0.42 (95% CI, 0.36–0.48) and 1.00 (95% CI, 0.98–1.00), respectively. Furthermore, the differences in pooled sensitivity were statistically significant in the diagnosis of grade 1 and 2 bladder tumors. Summary receiver operating characteristic curve values for urinary survivin mRNA expression and urine cytology were 0.95 (95% CI, 0.93–0.97) and 0.86 (95% CI, 0.83–0.89), respectively. Urinary survivin mRNA expression was also more accurate compared with other diagnostic indicators, including positive likelihood ratios, negative likelihood ratios, diagnostic odds ratios and Youden's index. Compared with traditional urine cytology, urinary survivin mRNA detection using reverse transcription-PCR was identified to be more effective in the diagnosis of early bladder cancer.

Highly Integrated, Biostable, and Self-Powered DNA Motor Enabling Autonomous Operation in Living Bodies.

An ultimate goal of synthetic DNA motor studies is to mimic natural protein motors in biological systems. Here, we rationally designed a highly integrated and biostable DNA motor system with high potential for living body operation, through simple assembly of a Mn2+-dependent DNAzyme-powered DNA motor with a degradable MnO2 nanosheet. The motor system shows outstanding high integration and improved biostability. High integration confers the motor system with the ability to deliver all the core components to the target sites as a whole, thus, enabling precise control of the spatiotemporal distribution of these components and achieving high local concentrations. At the target sites, reduction of the MnO2 nanosheet by intracellular glutathione (GSH) not only releases the DNA motor, which can then be initiated by the intracellular target, but also produces Mn2+ in situ to power the autonomous and progressive operation of the DNA motor. Interestingly, the resultant consumption of GSH in turn protects the DNA motor from destruction by physiological GSH, thus, conferring our motor system with improved biostability, reduced false-positive outputs, and consequently, an increased potential to be applied in a living body. As a proof of concept, the highly integrated DNA motor system was demonstrated to work well for amplified imaging detection of survivin mRNA (mRNA), an important tumor biomarker, in both living cancer cells and living tumor-bearing mice. This work reveals concepts and strategies promoting synthetic DNA motor applications in biological systems.

Optical Biosensing System for the Detection of Survivin mRNA in Colorectal Cancer Cells Using a Graphene Oxide Carrier-Bound Oligonucleotide Molecular Beacon.

The anti-apoptotic protein survivin is one of the most promising cancer biomarkers owing to its high expression in human cancers and rare occurrence in normal adult tissues. In this work, we have investigated the role of supramolecular interactions between a graphene oxide (GO) nanosheet nanocarrier and a survivin molecular beacon (SurMB), functionalized by attaching fluorophore Joe and quencher Dabcyl (SurMB-Joe). Molecular dynamics simulations revealed hydrogen bonding of Joe moiety and Dabcyl to GO carriers that considerably increase the SurMB-GO bonding strength. This was confirmed in experimental work by the reduced fluorescence background in the OFF state, thereby increasing the useful analytical signal range for mRNA detection. A new mechanism of hairpin–hairpin interaction of GO@SurMB with target oligonucleotides has been proposed. A low limit of detection, LOD = 16 nM (S/N = 3), has been achieved for complementary tDNA using GO@SurMB-Joe nanocarriers. We have demonstrated an efficient internalization of SurMB-Joe-loaded GO nanocarriers in malignant SW480 cells. The proposed tunability of the bonding strength in the attached motifs for MBs immobilized on nanocarriers, via structural modifications, should be useful in gene delivery systems to enhance the efficacy of gene retention, cell transfection and genomic material survivability in the cellular environment.

The Relation between Survivin Gene Expression and Urinary Bladder Cancer Disease

Purpose: The aim of this study is to investigate the expression of survivin gene in patients suffering from urinary bladder cancer, thus contributing to further understanding of the molecular etiology of the disease by assessment of Survivin gene expression on both RNA and protein levels in urinary bladder tissue samples and Correlate between its expression and pathological prognostic parameters of patients with urinary bladder cancer risk. Methods: In the present study 26 cases of urinary bladder cancer and 26 cases of normal tissues far from tumor sites in the urinary bladder of the same patients (as control samples) were collected, also 10 paraffin-embedded tissue samples from consecutive of archival bladder specimens patients and 10 tissue samples that of healthy tissue from the same individual as control group were collected for immunohistochemical detection of survivin protein and real time PCR for survivin mRNA detection. Results: In normal tissues of control samples no expression for survivin gene has been noticed. However, in patients affected with bladder cancer the survivin expression was highly increased by increasing degree of tumor malignancy as indicated by real time PCR for survivin mRNA and immunohistochemistry for survivin protein. Conclusion: From this study, it is concluded that survivin protein was not expressed in normal bladder urothelium but was present in a high percentage in tissue samples of patients suffering from bladder cancer and its expression is associated with disease recurrence, tumor grade, progression and mortality.

Molecular beacon-decorated polymethylmethacrylate core-shell fluorescent nanoparticles for the detection of survivin mRNA in human cancer cells

One of the main goals of nanomedicine in cancer is the development of effective drug delivery systems, primarily nanoparticles.Survivin, an overexpressed anti-apoptotic protein in cancer, represents a pharmacological target for therapy and a Molecular Beacon (MB) specific for survivin mRNA is available. In this study, the ability of polymethylmethacrylate nanoparticles (PMMA-NPs) to promote survivin MB uptake in human A549 cells was investigated.Fluorescent and positively charged core PMMA-NPs of nearly 60nm, obtained through an emulsion co-polymerization reaction, and the MB alone were evaluated in solution, for their analytical characterization; then, the MB specificity and functionality were verified after adsorption onto the PMMA-NPs.The carrier ability of PMMA-NPs in A549 was examined by confocal microscopy. With the optimized protocol, a hardly detectable fluorescent signal was obtained after incubation of the cells with the MB alone (fluorescent spots per cell of 1.90±0.40 with a mean area of 1.04±0.20µm2), while bright fluorescent spots inside the cells were evident by using the MB loaded onto the PMMA-NPs. (27.50±2.30 fluorescent spots per cell with a mean area of 2.35±0.16µm2). These results demonstrate the ability of the PMMA-NPs to promote the survivin-MB internalization, suggesting that this complex might represent a promising strategy for intracellular sensing and for the reduction of cancer cell proliferation.

Plasmonic nanohalo optical probes for highly sensitive imaging of survivin mRNA in living cells.

A strategy is designed for sensitive detection of tumor biomarker survivin mRNA based on resonance Rayleigh scattering of a single AuNP nanohalo probe that couples large gold nanoparticles (L-AuNPs, 52 nm) with small AuNPs (S-AuNPs, 18 nm) through the affinity interaction between streptavidin and biotin. This core-satellite plasmon ruler is further applied to imaging survivin mRNA in living cells.

Integration of DNA bio-gates and duplex-specific nuclease signal amplification: towards electrochemiluminescence detection of survivin mRNA.

Using the DNA bio-gate and duplex-specific nuclease assisted target recycling, a facile electrochemiluminescence assay was developed for the sensitive detection of survivin mRNA.

On-chip selective capture of cancer cells and ultrasensitive fluorescence detection of survivin mRNA in a single living cell

The rapid recognition of cancer cells and detection of tumor biomarker survivin mRNA plays a critical role in the early diagnosis of many cancers. Based on the integration of specific cancer cell capture and intracellular survivin mRNA detection, this work presents a novel and sensitive on-chip approach for the bioanalysis of survivin mRNA in a single living cell. The microchannel surface was firstly modified with a prostate stem cell antigen (PSCA) monoclonal antibody as the recognition element for prostate cancer cells (PC-3). As a result of the antigen-antibody specific affinity interactions, PC-3 cells could be selectively captured on the microchannel surface. After cell capture, nano-sized graphene oxide-poly(ethylene glycol) bis(amine) (NGO-PEG) was employed as a quencher and carrier of a signal tag, fluorescein isothiocyanate (FITC)-labeled antisense oligonucleotide (F-S1), which is complementary to part of survivin mRNA (target survivin mRNA), to transfect into the captured PC-3 cells. Upon the selective binding of S1 to intracellular survivin mRNA, F-S1 will be released from the NGO-PEG, inducing the fluorescence recovery of FITC. This antibody-based microfluidic device enables simple and inexpensive monitoring of the amount of survivin mRNA in single captured cell without the need for sample pretreatment. The survivin mRNA content in each PC-3 cell was estimated to be (4.8 ± 1.8) × 10(6) copies. This strategy opens a different perspective for ultrasensitive survivin mRNA detection, which may facilitate the early screening for malignancy.

A Novel Molecular Beacon Bearing a Graphite Nanoparticle as a Nanoquencher for In situ mRNA Detection in Cancer Cells

Molecular beacons (MBs) have shown fascinating applications in many biological fields. However, exploration of cost-effective, sensitive, stable and efficient MB for in situ live cell- based assay has still room for improvement. In this regards, we have developed a novel MB which bears a spherical graphite nanoparticle (GN) as a fluorescent quencher. The GN resulted in the high quenching efficiency, and the presence of GN enhanced the biological stability and transfection of the MB into the cells, thereby enabling the real-time survivin mRNA detection and quantification in the MCF-7 breast cancer cells. These results demonstrated that the advancement of the proposed MB containing a GN nanoquencher can be used as a robust molecular probe for genetic analysis in the cells.

Expression of survivin in human oocytes and preimplantation embryos

To determine whether [1] survivin is expressed in human oocytes and embryos; [2] embryos grown in vitro secrete survivin protein; and [3] survivin levels are correlated with embryo cleavage rates. Experimental. University-affiliated IVF clinic. Couples undergoing IVF-ET cycles. Conventional reverse transcriptase-polymerase chain reaction (PCR), real-time PCR, immunohistochemistry, Western blot on oocytes, embryos and control choriocarcinoma JEG-3 cells, and ELISA analysis of conditioned culture media. Detection of survivin mRNA and protein in oocytes and preimplantation embryos and in JEG-3 cancer cells. Detection of survivin concentrations in embryo culture media. Survivin mRNA and protein were expressed during human oocyte maturation, from germinal vesicle to metaphase II stage, and throughout embryo development, from pronuclear stage to blastocyst stage. Survivin was localized predominantly in the cytoplasm of all cells examined and in the oocytes on the chromatin of metaphase chromosomes and midbodies. Western blot analysis of human oocyte and cancer cell extracts detected a full-length (primary) survivin band of 16.5 kDa. Survivin was also detected in conditioned media samples from embryo cultures and showed a positive correlation with embryo cleavage rates. Our data have demonstrated for the first time that human oocytes/embryos not only express but also secret survivin, suggesting that survivin may play an important role in human oogenesis and embryogenesis.

Switchable ‘on–off–on’ electrochemical technique for direct detection of survivin mRNA in living cells

Survivin is a new member of apoptosis inhibition protein in cells which can promote cell proliferation and inhibit apoptosis through karyokinesis. Survivin mRNA in cells has become an important biomarker for early and accurate cancer diagnostics. Here, a switchable electrochemical sensor for survivin mRNA in living cells has been designed, which is based on the changes of electron transfer efficiency between ferrocene at the end of DNA and the electrode surface after reaction with survivin mRNA. The method was then used to study populations of individual cells with different levels of mRNA. Based on the different current changes, it was possible to evaluate different cell populations and cell status on the basis of the amount of mRNA levels in cells. This strategy may provide a new direction in the fabrication of high-performance electrochemical biosensors for the detection of biomolecules in living cells.

The diagnostic value of urine-based survivin mRNA test using reverse transcription-polymerase chain reaction for bladder cancer: a systematic review

Survivin has gradually become an important target in diagnosis, prognosis prediction and treatment of tumor. There are many studies on urine-based survivin mRNA test using reverse transcription-polymerase chain reaction (RT-PCR) as a noninvasive examination for bladder cancer. However, its clinical value remains controversial. This study was to evaluate the diagnostic value of urine survivin mRNA detection with RT-PCR for bladder cancer by a systematic review of related studies. With the search terms such as bladder neoplasm, survivin, RT-PCR, sensitivity, specificity and diagnosis, we systematically searched through PubMed, EMBASE, SCI, Cochrane Library, Chinese Biomedical Literature Database (CBM), Chinese Scientific Journal Full-text Database (CSJD), China Journal Full-text Database (CJFD), Chinese Medical Association (CMA) digital periodicals and Google Scholar totally from January 1997 to April 2009 for diagnostic trials with RT-PCR detection of urine survivin mRNA for bladder cancer. The Quality Assessment of Diagnostic Accuracy Studies (QUADAS) items were used to evaluate the quality of the included studies. Meta-disc software was used to calculate outcome indicators. Twenty-six studies, totally 2 416 patients, were eligible. Meta-analysis showed that compared with pathologic examination, the summary values of sensitivity, specificity, positive likelihood ratio, negative likelihood ratio and summary receiver operating characteristic curve (SROC) of urine-based survivin mRNA test using RT-PCR for bladder cancer were 88%, 94%, 14.56, 0.13 and 0.9736, respectively. Nested RT-PCR got the highest sensitivity, specificity and SROC and the values were 91%, 95% and 0.9805, respectively. The sensitivity and specificity of general RT-PCR were the second highest, which were 87% and 94%, respectively. The sensitivity of quantitative RT-PCR was 80% and the specificity was 93%. Comparing with pathologic examination, the sensitivity and specificity of urine-based survivin mRNA test using RT-PCR are relatively high. It can be used as an important adjunct method for cystoscope in early screening and postoperative monitoring of bladder cancer.

Preoperative Survivin mRNA Detection in Peripheral Blood is an Independent Predictor of Outcome in Esophageal Carcinoma

Survivin (SVV) mRNA expression levels in peripheral blood of patients with gastrointestinal malignancies change significantly during the course of treatment. We wanted to scrutinize these findings in patients with esophageal carcinoma and furthermore evaluate whether the detection of mRNA and the change in detecting ability have an association with overall survival. Whole blood was drawn 1 day pre- and 10 days post-operatively from 62 patients with esophageal carcinoma. Tumor cells were enriched from whole blood by density-gradient centrifugation prior to extraction of total cellular RNA and subsequent direct quantitative reverse transcriptase-PCR assays. SVV was detectable in 48 out of 62 patients (77%). Stepwise multivariate Cox linear regression models demonstrated a significant and independent association of measured SVV with overall survival (6.6 exp[b]; 95% CI: 1.97-22.12; p = 0.002). Increased SVV levels after the operation were linked to shorter overall survival (p = 0.04). Preoperative SVV expression levels appear to be associated with overall survival in patients with esophageal cancers. Increasing levels could potentially indicate a higher risk for shorter overall survival and therefore demand adapted treatment modalities.

Detection of survivin mRNA in healthy oral mucosa, oral leucoplakia and oral cancer

Survivin is involved in modulation of cell death and cell division processes. Survivin expression in normal adult tissues has not been fully understood, although it is markedly lower than in cancer, where it is over-expressed. To investigate survivin expression in normal, potentially malignant and cancerous oral mucosa. We measured survivin mRNA levels by real-time RT-PCR in specimens of oral mucosa (15 from normal mucosa, 17 from potentially malignant lesions, 17 from neoplasms). Scores were compared using Kruskal-Wallis test and post hoc according to Conover. Chi-squared test was used for dichotomous data. The median relative levels of survivin mRNA resulted six for normal mucosa, eight for potentially malignant lesions, 13 for cancers: differences among these three groups were statistically significant, as between cancer and potentially malignant lesions. Expression in normal mucosa and potentially lesions group showed no significant difference. Low, but not marginal expression of survivin in normal mucosa is a new finding, and it could be explained with the higher sensibility of our methods. Survivin expression in oral potentially malignant lesions might indicate a progressive deregulation of expression paralleling oncogenesis, particularly during the first stages of process, suggesting a putative predictive role for survivin.

Detection of Survivin mRNA in nasopharyngeal carcinoma by real-time fluorescence quantitative RT-PCR

To establish the method of real time fluorescence quantitative RT-PCR for detecting the expression of Survivin mRNA in nasopharyngeal carcinoma (NPC) tissues. The total RNA was extracted from NPC cell line CNE-2 and tissues with Trizol and then been transcribed reversely to cDNA, a method of real time fluorescence quantitative RT-PCR for detecting the expression of Survivin mRNA in NPC tissues had been established, in which chronic nasopharyngitis patients’ nasopharynx tissues treated as control group. The expression of Survivin mRNA all could be detected either in CNE-2 cells, NPC tissues or in chronic nasopharyngitis patients’ nasopharynx tissues, and there was higher the expression level of Survivin mRNA in NPC tissues than which in chronic nasopharyngitis patients’ nasopharynx tissues, the difference was significant (P 0.05). There was no relationship between Survivin mRNA expression and age and sex of NPC patients (P > 0.05). Real time fluorescence quantitative RT-PCR is a rapid, effective and high sensitive method for detecting the expression of Survivin mRNA in NPC tissues. The overexpression of Survivin mRNA may play some roles in pathogenesis of NPC.

Detection of survivin mRNA in urine of patients with superficial urothelial cell carcinomas

The aim of the present study was to assess if the presence of survivin mRNA in exfoliated cells present in urine samples can be a reliable marker of the presence of bladder tumour and recurrence. Urine samples from 30 patients with superficial urothelial cell carcinomas (UCC) were collected prior to transurethral resection (TUR) of the tumour and in the first routine follow-up, three months after TUR. Detection of survivin mRNA was performed by reverse transcription-polymerase chain reaction (RT-PCR). No correlation was observed between survivin detection and the clinicopathological variables analysed, nevertheless, when patients were grouped into low-grade (G1) and high-grade (G2+G3) tumours, statistically significant differences were found between both groups (p=0.04). When we analysed the results of survivin detection and urinary cytology together, we observed that informative cases rose from 27.8% to 44.4%. Also, Kaplan-Meier curves for patients with negative cytology in the first followup, categorised according to survivin detection, revealed that survivin mRNA positive cases recurred earlier than negative ones. From our results we can conclude that detection of survivin expression can be a reliable tumour marker, but more studies are needed to clarify the potential of survivin to predict recurrences. These results showed that survivin detection in combination with conventional urinary cytology can be a useful tool to increase the sensitivity in detecting the presence of a recurrence after TUR.

Evaluation of Survivin Reverse Transcriptase-Polymerase Chain Reaction for Noninvasive Detection of Bladder Cancer

Survivin, a member of the inhibitor of apoptosis gene family, is expressed in most common cancers. We investigated the expression pattern of survivin in the tumors of patients with bladder cancer and assessed the diagnostic potential of RT-PCR detection of survivin mRNA in urine. RT-PCR was used to analyze mRNA expression of survivin in 161 cases of bladder cancer, including TCC in 97, SCC in 53 and adenocarcinoma in 11, and their matched nontumor tissues. Urine specimens (50 ml) were collected from 84 patients in whom bladder cancer was documented by transurethral resection or biopsy, 41 with nonbladder cancer urological diseases and 42 healthy volunteers. Total RNA was extracted from urine sediments and RT-PCR was performed for survivin. Survivin expression was detected in all bladder cancer tissues. In contrast, survivin was not detectable in normal urothelium specimens. Urinary survivin was detected in urine samples from 51 of 53 patients with TCC, 22 of 25 with SCC and 6 of 6 with adenocarcinoma. Overall sensitivity was 94%. Survivin mRNA was not detected in any healthy volunteers. Positive results were obtained in 2 patients with renal cell carcinoma, 1 with hematuria and 1 with a contracted bladder but in none with other urological diseases. Overall specificity was 95%. Survivin mRNA detection in urine sediment using RT-PCR shows high sensitivity and specificity for bladder cancer. It may prove useful for the routine screening and monitoring of patients.

Survivin Gene Expression Positively Correlates with Proliferative Activity of Cancer Cells in Esophageal Cancer

Objective: The correlation between survivin gene expression and the occurrence of spontaneous apoptosis, proliferative activity of cancer cells, tumor angiogenesis, and abnormal p53 nuclear accumulation was evaluated in esophageal cancer. Methods: A total of 57 patients, on whom surgical esophageal resection had been performed between 1993 and 2000, were enrolled in this study. Total RNA was extracted from tumors and noncancerous epithelia. Expression levels of survivin and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) messenger RNA (mRNA) were analyzed quantitatively by real-time reverse transcriptase polymerase chain reaction (quantitative PCR). The proliferative activity of cancer cells, apoptotic cancer cells, microvessel density, and abnormal p53 nuclear accumulation were analyzed in these tumors. Results: The expression level of tumor survivin mRNA described as survivin/GAPDH (s/G) ratio was higher than that of noncancerous tissue (p = 0.0003), but did not correlate with lymph node metastasis, with the depth of tumor invasion, with the occurrence of apoptosis or with tumor angiogenesis. However, the tumor s/G ratio positively correlated with the proliferative activity and p53 nuclear accumulation in cancer cells. The 5-year survival rate of 53 patients was 37% (we excluded 4 patients who died from operative complications from this survival study). The 5-year survival rate of 27 patients with a high tumor s/G ratio (28.6%) was lower than that of 26 patients with a low tumor s/G ratio (46.2%, p = 0.041). High tumor s/G ratio was detected as an important prognostic factor independent of tumor stage. Conclusion: Detection of survivin mRNA by quantitative PCR provided us with important prognostic and biological information regarding esophageal cancer patients.