MiRNA Stability in Frozen Plasma Samples.
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ABSTRACT:
SUBMITTER: Balzano F
PROVIDER: S-EPMC6331950 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Ovarian cancer is one of the most common cancer types in women characterized by a high mortality rate due to lack of early diagnosis. Circulating miRNAs besides being important regulators of cancer development could be potential biomarkers to aid diagnosis. We performed the circulating miRNA expression analysis in plasma samples obtained from ovarian cancer patients stratified into FIGO I, FIGO III, and FIGO IV stages and from healthy females using the NanoString quantitative assay. Forty-five miRNAs were differentially expressed, out of these 17 miRNAs showed significantly different expression between controls and patients, 28 were expressed only in patients, among them 19 were expressed only in FIGO I patients. Differentially expressed miRNAs were ranked by the network-based analysis to assess their importance. Target genes of the differentially expressed miRNAs were identified then functional annotation of the target genes by the GO and KEGG-based enrichment analysis was carried out. A general and an ovary-specific protein-protein interaction network was constructed from target genes. Results of our network and the functional enrichment analysis suggest that besides HSP90AA1, MYC, SP1, BRCA1, RB1, CFTR, STAT3, E2F1, ERBB2, EZH2, and MET genes, additional genes which are enriched in cell cycle regulation, FOXO, TP53, PI-3AKT, AMPK, TGFβ, ERBB signaling pathways and in the regulation of gene expression, proliferation, cellular response to hypoxia, and negative regulation of the apoptotic process, the GO terms have central importance in ovarian cancer development. The aberrantly expressed miRNAs might be considered as potential biomarkers for the diagnosis of ovarian cancer after validation of these results in a larger cohort of ovarian cancer patients.
| S-EPMC6769773 | biostudies-literature
Project description:The aim of the present study was to perform an additional global miRNA microarray analysis of tubular and tubulovillous adenoma biopsy specimen completed with colorectal adenocarcinomas using fresh frozen tisse
2016-10-19 | GSE83924 | GEO
Project description:Circulating microRNAs that are associated with specific diseases have garnered much attention for use in diagnostic assays. However, detection of disease-associated miRNA can be affected by several factors such as release of contaminating cellular miRNA during sample collection, variations due to amplification of transcript for detection, or controls used for normalization for accurate quantitation. We analyzed circulating miRNA in serum and plasma samples obtained concurrently from 28 patients, using a Nanostring quantitative assay platform. Total RNA concentration ranged from 32-125 ?g/ml from serum and 30-220 ?g/ml from plasma. Of 798 miRNAs, 371 miRNAs were not detected in either serum or plasma samples. 427 were detected in either serum or plasma but not both, whereas 151 miRNA were detected in both serum and plasma samples. The diversity of miRNA detected was greater in plasma than in serum samples. In serum samples, the number of detected miRNA ranged from 3 to 82 with a median of 17, whereas in plasma samples, the number of miRNA detected ranged from 25 to 221 with a median of 91. Several miRNA such as miR451a, miR 16-5p, miR-223-3p, and mir25-3p were highly abundant and differentially expressed between serum and plasma. The detection of endogenous and exogenous control miRNAs varied in serum and plasma, with higher levels observed in plasma. Gene expression stability identified candidate invariant microRNA that were highly stable across all samples, and could be used for normalization. In conclusion, there are significant differences in both the number of miRNA detected and the amount of miRNA detected between serum and plasma. Normalization using miRNA with constant expression is essential to minimize the impact of technical variations. Given the challenges involved, ideal candidates for blood based biomarkers would be those that are indifferent to type of body fluid, are detectable and can be reliably quantitated.
| S-EPMC5718466 | biostudies-literature
Project description:(1) Background: Glioblastoma multiforme (GBM) is among the most aggressive cancers with a poor prognosis. Treatment options are limited, clinicians lack efficient prognostic and predictive markers. Circulating miRNAs-besides being important regulators of cancer development-may have potential as diagnostic biomarkers of GBM. (2) Methods: In this study, profiling of 798 human miRNAs was performed on blood plasma samples from 6 healthy individuals and 6 patients with GBM, using a NanoString nCounter Analysis System. To validate our results, five miRNAs (hsa-miR-433-3p, hsa-miR-362-3p, hsa-miR-195-5p, hsa-miR-133a-3p, and hsa-miR-29a-3p) were randomly chosen for RT-qPCR detection. (3) Results: In all, 53 miRNAs were significantly differentially expressed in plasma samples of GBM patients when data were filtered for FC 1 and FDR 0.1. Target genes of the top 39 differentially expressed miRNAs were identified, and we carried out functional annotation and pathway enrichment analysis of target genes via GO and KEGG-based tools. General and cortex-specific protein-protein interaction networks were constructed from the target genes of top miRNAs to assess their functional connections. (4) Conclusions: We demonstrated that plasma microRNA profiles are promising diagnostic and prognostic molecular biomarkers that may find an actual application in the clinical practice of GBM, although more studies are needed to validate our results.
| S-EPMC8151942 | biostudies-literature
Project description:Background: As degradation of formalin-fixed paraffin-embedded (FFPE) samples limits ability to expression profile, we explored factors predicting success for FFPE profiling and investigated an approach overcoming this limitation. Methods: Bladder (n=141, stored 3-8 years) and cervix (n=160, stored 8-23 years) carcinoma FFPE samples were hybridised to Affymetrix Exon1.0ST arrays. Percent detection above background (%DABG) measured technical success. Biological signal was assessed by distinguishing cervix squamous cell carcinoma (SCC) and adenocarcinoma (AC) using a gene signature. Precursor mir-205 was measured by Exon array and mature miR-205 by qRT-PCR. Eight-old and -young cervix samples were compared using Affymetrix miRNA 2.0 arrays. For comparison, the 'cervix_tumour_cs1_113' (previsously submitted GSM677307) was included and re-analyzed with the samples from the current study (total 161 Cervix samples). Results: RNA quality controls (e.g. RNA integrity number) failed to predict profiling success, but sample age correlated with %DABG in bladder (R2=-0.30, p<0.01) and cervix (R2=-0.69, p<0.01). Biological signal was lost in older samples and neither a signature nor precursor mir-205 separated samples by histology. miR-205 qRT-PCR discriminated SCC from AC, validated by miRNA profiling (26-fold higher in SCC; p=1.10x10-5). Median miRNA probeset expression of eight-old and eight-young cervix samples correlated well (R2=0.95) overcoming the age-related bias of mRNA probesets, suggesting miR-205 stability generalises across microRNA. Conclusions: microRNA profiling overcomes the limitation of degraded FFPE samples
2012-07-04 | GSE39015 | GEO
Project description:PurposeMicrogravity has severe effects on cellular and molecular structures as well as on metabolic interactions. The aim of this study is to investigate the effects of microgravity (μg) exposure on human frozen sperm samples.MethodsSibling samples from 15 normozoospermic healthy donors were frozen using glycerol as cryoprotectant and analyzed under microgravity and ground conditions. Microgravity was obtained by parabolic flights using a CAP10B plane. The plane executed 20 parabolic maneuvers with a mean of 8.5 s of microgravity for each parabola.ResultsFrozen sperm samples preserved in cryostraws and stored in a secure and specific nitrogen vapor cryoshipper do not suffer significant alterations after μg exposure. Comparing the study group (μg) and the control group (1 g), similar results were obtained in the main parameters studied: sperm motility (M/ml) 13.72 ± 12.57 vs 13.03 ± 12.13 (- 0.69 95% CI [- 2.9; 1.52]), progressive a + b sperm motility (%) 21.83 ± 11.69 vs 22.54 ± 12.83 (0.03 95% CI [- 0.08; 0.15]), sperm vitality (%) 46.42 ± 10.81 vs 44.62 ± 9.34 (- 0.04 95% CI [- 0.13; 0.05]), morphologically normal spermatozoa (%) 7.03 ± 2.61 vs 8.09 ± 3.61 (0.12 95% CI [0.01; 0.24]), DNA sperm fragmentation by SCD (%) 13.33 ± 5.12 vs 13.88 ± 6.14 (0.03 95% CI [- 0.09; 0.16]), and apoptotic spermatozoa by MACS (%) 15.47 ± 15.04 vs 23.80 ± 23.63 (- 0.20 95% CI [- 0.66; 1.05]).ConclusionThe lack of differences obtained between frozen samples exposed to μg and those maintained in ground conditions provides the possibility of considering the safe transport of human male gametes to space. Nevertheless, further research is needed to validate the results and to consider the possibility of creating a human sperm bank outside the Earth.Trial registration numberClinicalTrials.gov: NCT03760783.
| S-EPMC7492354 | biostudies-literature
Project description:Background: As degradation of formalin-fixed paraffin-embedded (FFPE) samples limits ability to expression profile, we explored factors predicting success for FFPE profiling and investigated an approach overcoming this limitation. Methods: Bladder (n=141, stored 3-8 years) and cervix (n=160, stored 8-23 years) carcinoma FFPE samples were hybridised to Affymetrix Exon1.0ST arrays. Percent detection above background (%DABG) measured technical success. Biological signal was assessed by distinguishing cervix squamous cell carcinoma (SCC) and adenocarcinoma (AC) using a gene signature. Precursor mir-205 was measured by Exon array and mature miR-205 by qRT-PCR. Eight-old and -young cervix samples were compared using Affymetrix miRNA 2.0 arrays. For comparison, the 'cervix_tumour_cs1_113' (previsously submitted GSM677307) was included and re-analyzed with the samples from the current study (total 161 Cervix samples). Results: RNA quality controls (e.g. RNA integrity number) failed to predict profiling success, but sample age correlated with %DABG in bladder (R2=-0.30, p<0.01) and cervix (R2=-0.69, p<0.01). Biological signal was lost in older samples and neither a signature nor precursor mir-205 separated samples by histology. miR-205 qRT-PCR discriminated SCC from AC, validated by miRNA profiling (26-fold higher in SCC; p=1.10x10-5). Median miRNA probeset expression of eight-old and eight-young cervix samples correlated well (R2=0.95) overcoming the age-related bias of mRNA probesets, suggesting miR-205 stability generalises across microRNA. Conclusions: microRNA profiling overcomes the limitation of degraded FFPE samples 18 samples hybridised to miRNA microarray V2 arrays, 2 cervix cell lines and 16 clinical cervical tumour samples. There are no technical replicates.
2012-07-03 | E-GEOD-39015 | biostudies-arrayexpress
Project description:Using a microarray-based method we report on: i) adequate intra- and interarray reproducibility of miRNA profiling; ii) feasibility of using archival plasma samples stored for an extended period of time and available in limited amounts; and iii) good correlation between different batches.
2013-04-23 | GSE42654 | GEO
Project description:Background: As degradation of formalin-fixed paraffin-embedded (FFPE) samples limits ability to expression profile, we explored factors predicting success for FFPE profiling and investigated an approach overcoming this limitation. Methods: Bladder (n=141, stored 3-8 years) and cervix (n=160, stored 8-23 years) carcinoma FFPE samples were hybridised to Affymetrix Exon1.0ST arrays. Percent detection above background (%DABG) measured technical success. Biological signal was assessed by distinguishing cervix squamous cell carcinoma (SCC) and adenocarcinoma (AC) using a gene signature. Precursor mir-205 was measured by Exon array and mature miR-205 by qRT-PCR. Eight-old and -young cervix samples were compared using Affymetrix miRNA 2.0 arrays. For comparison, the 'cervix_tumour_cs1_113' (previsously submitted GSM677307) was included and re-analyzed with the samples from the current study (total 161 Cervix samples). Results: RNA quality controls (e.g. RNA integrity number) failed to predict profiling success, but sample age correlated with %DABG in bladder (R2=-0.30, p<0.01) and cervix (R2=-0.69, p<0.01). Biological signal was lost in older samples and neither a signature nor precursor mir-205 separated samples by histology. miR-205 qRT-PCR discriminated SCC from AC, validated by miRNA profiling (26-fold higher in SCC; p=1.10x10-5). Median miRNA probeset expression of eight-old and eight-young cervix samples correlated well (R2=0.95) overcoming the age-related bias of mRNA probesets, suggesting miR-205 stability generalises across microRNA. Conclusions: microRNA profiling overcomes the limitation of degraded FFPE samples
2012-07-04 | GSE39016 | GEO
Project description:Many emerging technologies are reliant on circulating cell-free DNA (cfDNA) and cell-free RNA (cfRNA) applications in the clinic. However, the impact of diurnal cycles or daily meals on circulating analytes are poorly understood and may be confounding factors when developing diagnostic platforms. To begin addressing this knowledge gap, we obtained plasma from four healthy donors serially sampled five times during 12 h in a single day. For all samples, we measured concentrations of cfDNA and cfRNA using both bulk measurements and gene-specific digital droplet PCR. We found no significant variation attributed to blood draw number for the cfDNA or cfRNA. This indicated that natural diurnal cycles and meal consumption do not appear to significantly affect abundance of total cfDNA, total cfRNA, or our two selected cfRNA transcripts. Conversely, we observed significant variation between individual donors for cfDNA and one of the cfRNA transcripts. The results of this work suggest that it will be important to consider patient-specific baselines when designing reliable circulating cfDNA or cfRNA clinical assays.
| S-EPMC7536441 | biostudies-literature