Long-term Storage of Urine and Urinary EVs: A Systematic Evaluation of microRNA Stability
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE310686 | GEO | 2026/05/15
REPOSITORIES: GEO
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Project description:Progress and advancement in assisted reproductive technologies (ART) and its outcomes are limited by the importance of research of endometrial receptivity being overlooked. Due to endometrial biopsy being invasive and in vitro studies lacking reproducibility in vivo, urine is an appealing alternative biofluid source for biomarker research as it can be collected in large quantities non-invasively. The discovery of extracellular vesicles (EVs) in urine (uEVs), has also opened a new avenue in this biomarker research, with these EVs harbouring thousands of proteins that hold promise for biomarker development. In this study urine was collected from human female volunteers and samples representing the different phases of the menstrual cycle were subjected to EV isolation via differential centrifugation and size exclusion chromatography. The resulting uEVs were analysed via Nanoparticle tracking analysis (NTA) to examine the different concentration and size of particles and proteomic analysis performed using shotgun label-free mass spectrometry on the uEV samples and neat urine samples. Our results showed that uEVs were found in numbers depending on the menstrual cycle phase but uEV size was not statistically altered during different stages of the menstrual cycle. Proteomics showed 50% of proteins detected in the neat urine were also present in the uEV samples with 813 proteins were unique in the uEV samples. Proteomics analysis also showed that the menstrual cycle phase affect the uEVs proteomic profile, with some proteins shown to be significantly upregulated and downregulated during the window of implantation phase of the cycle compared to the other non-receptive periods. This data highlights that uEVs characteristics are altered depending on the menstrual cycle phase suggesting the potential of uEVs being used as biomarkers for improving fertility.
2023-12-04 | PXD047127 | Pride
Project description:Progress and advancement in assisted reproductive technologies (ART) and its outcomes are limited by the importance of research of endometrial receptivity being overlooked. Due to endometrial biopsy being invasive and in vitro studies lacking reproducibility in vivo, urine is an appealing alternative biofluid source for biomarker research as it can be collected in large quantities non-invasively. The discovery of extracellular vesicles (EVs) in urine (uEVs), has also opened a new avenue in this biomarker research, with these EVs harbouring thousands of proteins that hold promise for biomarker development. In this study urine was collected from human female volunteers and samples representing the different phases of the menstrual cycle were subjected to EV isolation via differential centrifugation and size exclusion chromatography. The resulting uEVs were analysed via Nanoparticle tracking analysis (NTA) to examine the different concentration and size of particles and proteomic analysis performed using shotgun label-free mass spectrometry on the uEV samples and neat urine samples. Our results showed that uEVs were found in numbers depending on the menstrual cycle phase but uEV size was not statistically altered during different stages of the menstrual cycle. Proteomics showed 50% of proteins detected in the neat urine were also present in the uEV samples with 813 proteins were unique in the uEV samples. Proteomics analysis also showed that the menstrual cycle phase affect the uEVs proteomic profile, with some proteins shown to be significantly upregulated and downregulated during the window of implantation phase of the cycle compared to the other non-receptive periods. This data highlights that uEVs characteristics are altered depending on the menstrual cycle phase suggesting the potential of uEVs being used as biomarkers for improving fertility.In this dataset we have the neat urine results.
2023-12-04 | PXD047128 | Pride
Project description:One of the challenges of current research in prostate cancer is to improve the differential non-invasive diagnosis of prostate cancer (PCa) and benign prostate hyperplasia (BPH). Extracellular vesicles (EV) are emerging structures with promising properties for intercellular communication. In addition, the characterization of EV in biofluids is an attractive source of non-invasive diagnostic, prognostic and predictive biomarkers. Here we show that urinary EV (uEV) from prostate cancer patients exhibit genuine and differential physical and biological properties. Importantly, transcriptomics characterization of uEVs led us to define the decreased abundance of Cadherin 3, type 1 (CDH3) transcript in uEV from PCa patients. Tissue and cell line analysis strongly suggested that the status of CDH3 in uEVs is a distal reflection of changes in the expression of this cadherin in the prostate tumor. Our results reveal that uEVs could represent a non-invasive tool to inform about the molecular alterations in prostate cancer. RNA extraction was performed in 10 ultracentrifuge-isolated-uEVs samples (4 BPH, 6 Pca)
2016-01-13 | E-GEOD-75441 | biostudies-arrayexpress
Project description:Bladder cancer exhibits molecular heterogeneity that complicates early diagnosis and prognosis, and drives confounding clinical outcomes. Non–muscle invasive and muscle-invasive subtypes, especially for intermediate to high grade, carry a 25 – 50% progression-free survival rate, underscoring the need for high precision prognostic strategy. Urinary extracellular vesicles (uEVs) are promising carriers of tumor-derived RNAs and proteins. However, significant challenges in studying uEVs arise from the diverse cellular origin of uEVs associated with the dynamic composition of urine, which presents roadblocks for developing the clinical utility of uEVs. We introduced an AI-driven EV liquid biopsy pipeline that integrates (1) standardized EV isolation via NanoPom magnetic beads, (2) transcriptomic profiling for molecular subtyping, and (3) prognostic scoring algorithm. In a discovery cohort of 16 bladder cancer patients including both MIBC and NMIBC, we compared NanoPom isolated uEVs with ExoEasy and Fujifilm MagCapture isolated uEVs, for identifying bladder cancer subtype-specific gene signatures, and externally validated them using UCSC Xena. The result outperformed currently reported bladder cancer diagnostic biomarkers from assays including Galeas, CxBladder, and Xpert. In a validation cohort of matched 7 patient plasma samples, we confirmed with plasma derived EVs for correlating with urinary EV biomarkers from NGS sequencing. The prognostic score stratified patients into low-, intermediate-, and high-grade risk groups based on Xena's bladder cancer survival outcomes. Our AI-driven uEV liquid biopsy pipeline proves the concept for high precision bladder cancer subtyping and prognosis, which could potentially facilitate treatment decision and lead to advanced profiling of bladder tumor biology using uEV liquid biopsy.
2025-09-28 | GSE308996 | GEO
Project description:Saccharomyces cerevisiae is exposed to freeze-thaw stress in commercial processes including frozen dough baking. The cell viability and fermentation activity after freeze-thaw were dramatically decreased due to freeze-thaw injury. Because freeze-thaw injury involves complex phenomena, the mechanisms of it are not fully understood. We attempted to analyze the mechanisms of freeze-thaw injury by indirect gene expression analysis during post-thaw incubation after freeze-thaw treatment using DNA microarray profiling. The results showed that a high frequency of the genes involved in the homeostasis of metal ions were up-regulated depending on the freezing period. The phenotype of the deletion mutants of the up-regulated genes extracted by indirect gene expression analysis was assessed. The deletion strains of the MAC1 and CTR1 genes involved in copper ion homeostasis exhibited freeze-thaw sensitivity, suggesting that copper ion homeostasis is required for freeze-thaw tolerance. Supplementation with copper ions during post-thaw incubation increased intracellular superoxide dismutase activity. Inverse correlated with intracellular superoxide dismutase activity, intracellular levels of reactive oxygen species were decreased. Moreover, cell viability increased by supplementation with copper ions under specific assessment conditions. This study suggested that insufficiency of copper ion homeostasis may be one of the causes of freeze-thaw injury.
2009-08-31 | GSE15461 | GEO
Project description:Saccharomyces cerevisiae is exposed to freeze-thaw stress in commercial processes including frozen dough baking. The cell viability and fermentation activity after freeze-thaw were dramatically decreased due to freeze-thaw injury. Because freeze-thaw injury involves complex phenomena, the mechanisms of it are not fully understood. We attempted to analyze the mechanisms of freeze-thaw injury by indirect gene expression analysis during post-thaw incubation after freeze-thaw treatment using DNA microarray profiling. The results showed that a high frequency of the genes involved in the homeostasis of metal ions were up-regulated depending on the freezing period. The phenotype of the deletion mutants of the up-regulated genes extracted by indirect gene expression analysis was assessed. The deletion strains of the MAC1 and CTR1 genes involved in copper ion homeostasis exhibited freeze-thaw sensitivity, suggesting that copper ion homeostasis is required for freeze-thaw tolerance. Supplementation with copper ions during post-thaw incubation increased intracellular superoxide dismutase activity. Inverse correlated with intracellular superoxide dismutase activity, intracellular levels of reactive oxygen species were decreased. Moreover, cell viability increased by supplementation with copper ions under specific assessment conditions. This study suggested that insufficiency of copper ion homeostasis may be one of the causes of freeze-thaw injury. Total RNA was extracted from the stress-treated yeast cells by using a hot phenol method. Poly(A)+ RNA was enriched from total RNA by using an Oligotex dT30 (Super) mRNA purification kit (Takara Bio, Ohtsu, Japan). cDNA synthesis, cRNA synthesis, and labeling were performed according to the Affymetrix user’s manual (Affymetrix, Santa Clara, USA). Biotinyated cRNA was fragmented and then used as a probe.Affimetrix Yeast Genome 2.0 arrays (Affymetrix) were used as DNA microarrays. All experiments were done in triplicate independently.
2009-08-31 | E-GEOD-15461 | biostudies-arrayexpress
Project description:Background: In autosomal dominant polycystic kidney disease (ADPKD) there is an unmet need for early markers of rapid disease progression to facilitate counseling and selection for kidney-protective therapy. Our aim was to identify markers for rapid disease progression in urinary extracellular vesicles (uEVs). Methods: uEVs were isolated at baseline in two independent cohorts including patients with rapid disease progression and stable disease; patients were matched by age, sex, total kidney volume and mutation. uEV biomarker candidates were identified by mass spectrometry and further analyzed in a third and fourth cohort using immunoblotting and an enzyme-linked immunosorbent assay (ELISA). Single-nucleotide RNA sequencing of healthy and ADPKD tissue was used to confirm findings and identify the cellular origin of the uEV biomarker. Results: In the first two cohorts matrix metalloproteinase-7 (MMP-7) was significantly higher in uEVs of patients with rapid disease progression compared to stable disease. In the third cohort, immunoblotting confirmed a >2-fold increase in uEV-MMP-7 in patients with rapid disease progression compared to stable disease. In the fourth cohort, whole urine rather than uEVs were analyzed with MMP-7 ELISA demonstrating a significant but weak correlation with kidney function decline. Single-nucleotide RNA sequencing showed higher MMP-7 expression in ADPKD kidney tissue than in healthy kidney tissue and localized MMP-7 to the proximal tubule and thick ascending limb. Conclusion: uEV-associated MMP-7 is elevated in ADPKD patients with rapid disease progression and should be further investigated as predictive biomarker. Our findings also suggests that MMP-7 performs better when analyzed in uEVs than in whole urine.
2023-12-18 | PXD042533 | Pride
Project description:In frozen dough baking technology, baker’s yeast Saccharomyces cerevisiae encounter freeze-thaw injury. After thawing, dramatically decrease in cell viability and fermentation activity is caused by freeze-thaw injury. The freezing period is critical factor in freeze-thaw injury, thus we focused and investigated time-dependent gene expression profiles in recovery process from freeze injury. First, changes in gene expression profiles in S. cerevisiae in recovery process from freeze-thaw injury were analyzed using a DNA microarray. The results showed the genes which were involved in homeostasis of metal ions were time-dependent up-regulated 2-fold or more in a series. Then we examined whether these genes were related to tolerance in freeze-thaw injury by using deletion strain. The results showed that deletion of MAC1, CTR1, and PCA1 genes which involved in copper ion transport exhibited freeze-thaw sensitivity in compared with wild type. These genes are involved in copper ion uptake to a cell under a copper deficiency condition or in copper ion homeostasis, suggesting that it may be related between freeze-thaw injury and copper ion transport. To determine the effect of supplementation of copper ion on cells after freeze-thaw treatment, cell viability, intracellular superoxide dismutase (SOD) activity, and intracellular levels of reactive oxygen species (ROS) were examined by various copper ion condition medium. The results showed that intracellular SOD activity was increased and intracellular levels of ROS were decreased by supplementation of copper ion, but there was no significant difference in cell viability. These results of the present study may suggest that copper ion concentration in yeast cell after freeze-thaw treatment is important to recovery from freeze-thaw injury due to redox control of intracellular levels of ROS, but copper ion did not directly affect cell viability.
2009-02-28 | GSE14942 | GEO
Project description:RNA-Seq is ubiquitous, but depending on the study, sub-optimal sample handling may be required, resulting in repeated freeze-thaw cycles. However, little is known about how each cycle impacts downstream analyses, due to a lack of study and known limitations in common RNA quality metrics, e.g., RIN, at quantifying RNA degradation following repeated freeze-thaws. Here we quantify the impact of repeated freeze-thaw on the reliability of downstream RNA-Seq analysis. To do so, we developed a method to estimate the relative noise between technical replicates independently of RIN. Using this approach we inferred the effect of both RIN and the number of freeze-thaw cycles on sample noise. We find that RIN is unable to fully account for the change in sample noise due to freeze-thaw cycles. Additionally, freeze-thaw is detrimental to sample quality and differential expression (DE) reproducibility, approaching zero after three cycles for poly(A)-enriched samples, wherein the inherent 3’ bias in read coverage is more exacerbated by freeze-thaw cycles, while ribosome-depleted samples are less affected by freeze-thaws. The use of poly(A)-enrichment for RNA sequencing is pervasive in library preparation of frozen tissue, and thus, it is important during experimental design and data analysis to consider the impact of repeated freeze-thaw cycles on reproducibility.
2020-05-09 | GSE150097 | GEO
Project description:In frozen dough baking technology, bakerâs yeast Saccharomyces cerevisiae encounter freeze-thaw injury. After thawing, dramatically decrease in cell viability and fermentation activity is caused by freeze-thaw injury. The freezing period is critical factor in freeze-thaw injury, thus we focused and investigated time-dependent gene expression profiles in recovery process from freeze injury. First, changes in gene expression profiles in S. cerevisiae in recovery process from freeze-thaw injury were analyzed using a DNA microarray. The results showed the genes which were involved in homeostasis of metal ions were time-dependent up-regulated 2-fold or more in a series. Then we examined whether these genes were related to tolerance in freeze-thaw injury by using deletion strain. The results showed that deletion of MAC1, CTR1, and PCA1 genes which involved in copper ion transport exhibited freeze-thaw sensitivity in compared with wild type. These genes are involved in copper ion uptake to a cell under a copper deficiency condition or in copper ion homeostasis, suggesting that it may be related between freeze-thaw injury and copper ion transport. To determine the effect of supplementation of copper ion on cells after freeze-thaw treatment, cell viability, intracellular superoxide dismutase (SOD) activity, and intracellular levels of reactive oxygen species (ROS) were examined by various copper ion condition medium. The results showed that intracellular SOD activity was increased and intracellular levels of ROS were decreased by supplementation of copper ion, but there was no significant difference in cell viability. These results of the present study may suggest that copper ion concentration in yeast cell after freeze-thaw treatment is important to recovery from freeze-thaw injury due to redox control of intracellular levels of ROS, but copper ion did not directly affect cell viability. Experiment Overall Design: Total RNA was extracted from the stress-treated yeast cells by using a hot phenol method. Poly(A)+ RNA was enriched from total RNA by using an Oligotex dT30 (Super) mRNA purification kit (Takara Bio, Ohtsu, Japan). cDNA synthesis, cRNA synthesis, and labeling were performed according to the Affymetrix userâs manual (Affymetrix, Santa Clara, USA). Biotinyated cRNA was fragmented and then used as a probe.Affimetrix Yeast Genome 2.0 arrays (Affymetrix) were used as DNA microarrays. All experiments were done in duplicate independently
2009-03-07 | E-GEOD-14942 | biostudies-arrayexpress