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Proteomic Profiling of Ectosomes Derived from Paired Urothelial Bladder Cancer and Normal Cells Reveals the Presence of Biologically-Relevant Molecules.

ABSTRACT: Protein content of extracellular vesicles (EVs) can modulate different processes during carcinogenesis. Novel proteomic strategies have been applied several times to profile proteins present in exosomes released by urothelial bladder cancer (UBC) cells. However, similar studies have not been conducted so far on another population of EVs, i.e., ectosomes. In the present study we used a shotgun nanoLC-MS/MS proteomic approach to investigate the protein content of ectosomes released in vitro by T-24 UBC cells and HCV-29 normal ureter epithelial cells. In addition, cancer-promoting effects exerted by UBC-derived ectosomes on non-invasive cells in terms of cell proliferation and migratory properties were assessed. In total, 1158 proteins were identified in T-24-derived ectosomes, while HCV-29-derived ectosomes contained a lower number of 259 identified proteins. Qualitative analysis revealed 938 proteins present uniquely in T-24-derived ectosomes, suggesting their potential applications in bladder cancer management as diagnostic and prognostic biomarkers. In addition, T-24-derived ectosomes increased proliferation and motility of recipient cells, likely due to the ectosomal transfer of the identified cancer-promoting molecules. The present study provided a focused identification of biologically relevant proteins in UBC-derived ectosomes, confirming their role in UBC development and progression, and their applicability for further biomarker-oriented studies in preclinical or clinical settings.

SUBMITTER: Surman M

PROVIDER: S-EPMC8268130 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Proteomic profiling of ectosomes derived from paired urothelial bladder cancer and normal cells reveals the presence of biologically-relevant molecules

Project description:Protein content of extracellular vesicles (EVs) can modulate different processes during carcinogenesis. Novel proteomic strategies have been applied several times to profile proteins present in exosomes released by urothelial bladder cancer (UBC) cells. However, similar studies have not been conducted so far on another population of EVs i.e. ectosomes. In the present study we used a shotgun nanoLC-MS/MS proteomic approach to investigate the protein content of ectosomes released in vitro by T-24 UBC cells and HCV-29 normal ureter epithelial cells. In addition, cancer-promoting effects exerted by UBC-derived ectosomes on non-invasive cells in terms of cell proliferation and migratory properties were assessed. A total of 1158 proteins was identified in T-24-derived ectosomes, while HCV-29-derived ectosomes contained a lower number of 259 identified proteins. Qualitative analysis revealed 938 proteins present uniquely in T-24-derived ectosomes, suggesting their potential applications in bladder cancer management as diagnostic and prognostic biomarkers. Also, T-24-derived ectosomes increased proliferation and motility of recipient cells, likely due to the ectosomal transfer of identified cancer-promoting molecules. The present study provided a focused identification of biologically relevant proteins in UBC-derived ectosomes, confirming their role in UBC development and progression, and their applicability for further biomarker-oriented studies in preclinical or clinical settings.

2021-05-06 | MSV000087370 | MassIVE

Mass Spectrometry-Based Proteomic Characterization of Cutaneous Melanoma Ectosomes Reveals the Presence of Cancer-Related Molecules.

Project description:Cutaneous melanoma (CM) is an aggressive type of skin cancer for which effective biomarkers are still needed. Recently, the protein content of extracellular vesicles (ectosomes and exosomes) became increasingly investigated in terms of its functional role in CM and as a source of novel biomarkers; however, the data concerning the proteome of CM-derived ectosomes is very limited. We used the shotgun nanoLC-MS/MS approach to the profile protein content of ectosomes from primary (WM115, WM793) and metastatic (WM266-4, WM1205Lu) CM cell lines. Additionally, the effect exerted by CM ectosomes on recipient cells was assessed in terms of cell proliferation (Alamar Blue assay) and migratory properties (wound healing assay). All cell lines secreted heterogeneous populations of ectosomes enriched in the common set of proteins. A total of 1507 unique proteins were identified, with many of them involved in cancer cell proliferation, migration, escape from apoptosis, epithelial-mesenchymal transition and angiogenesis. Isolated ectosomes increased proliferation and motility of recipient cells, likely due to the ectosomal transfer of different cancer-promoting molecules. Taken together, these results confirm the significant role of ectosomes in several biological processes leading to CM development and progression, and might be used as a starting point for further studies exploring their diagnostic and prognostic potential.

| S-EPMC7215915 | biostudies-literature

The clonal relation of primary upper urinary tract urothelial carcinoma and paired urothelial carcinoma of the bladder.

Project description:The risk of developing urothelial carcinoma of the bladder (UCB) in patients treated by radical nephroureterectomy (RNU) for an upper urinary tract urothelial carcinoma (UTUC) is 22% to 47% in the 2 years after surgery. Subject of debate remains whether UTUC and the subsequent UCB are clonally related or represent separate origins. To investigate the clonal relationship between both entities, we performed targeted DNA sequencing of a panel of 41 genes on matched normal and tumor tissue of 15 primary UTUC patients treated by RNU who later developed 19 UCBs. Based on the detected tumor-specific DNA aberrations, the paired UTUC and UCB(s) of 11 patients (73.3%) showed a clonal relation, whereas in four patients the molecular results did not indicate a clear clonal relationship. Our results support the hypothesis that UCBs following a primary surgically resected UTUC are predominantly clonally derived recurrences and not separate entities.

| S-EPMC7821318 | biostudies-literature

Water-structuring molecules and nanomaterials enhance radiofrequency heating in biologically relevant solutions.

Project description:For potential applications in nano-mediated radiofrequency cancer hyperthermia, the nanomaterial under investigation must increase the heating of any aqueous solution in which it is suspended when exposed to radiofrequency electric fields. This should also be true for a broad range of solution conductivities, especially those that artificially mimic the ionic environment of biological systems. Herein we demonstrate enhanced heating of biologically relevant aqueous solutions using kosmotropes and a hexamalonoserinolamide fullerene.

| S-EPMC5079531 | biostudies-literature

Messina: a novel analysis tool to identify biologically relevant molecules in disease.

Project description:BackgroundMorphologically similar cancers display heterogeneous patterns of molecular aberrations and follow substantially different clinical courses. This diversity has become the basis for the definition of molecular phenotypes, with significant implications for therapy. Microarray or proteomic expression profiling is conventionally employed to identify disease-associated genes, however, traditional approaches for the analysis of profiling experiments may miss molecular aberrations which define biologically relevant subtypes.Methodology/principal findingsHere we present Messina, a method that can identify those genes that only sometimes show aberrant expression in cancer. We demonstrate with simulated data that Messina is highly sensitive and specific when used to identify genes which are aberrantly expressed in only a proportion of cancers, and compare Messina to contemporary analysis techniques. We illustrate Messina by using it to detect the aberrant expression of a gene that may play an important role in pancreatic cancer.Conclusions/significanceMessina allows the detection of genes with profiles typical of markers of molecular subtype, and complements existing methods to assist the identification of such markers. Messina is applicable to any global expression profiling data, and to allow its easy application has been packaged into a freely-available stand-alone software package.

| S-EPMC2671167 | biostudies-literature

Quantitative Proteomic Analysis Reveals Caffeine-Perturbed Proteomic Profiles in Normal Bladder Epithelial Cells.

Project description:Lower urinary tract symptoms (LUTSs) are highly prevalent among the elderly and negatively impact quality of life. Since caffeinated beverages are enjoyed worldwide and the relationship between LUTS and caffeine is still not fully understood, it would be of particular interest to examine the underlying mechanisms that drive caffeine's influence on LUTS development and progression. The aim of this study is to characterize the effects of caffeine on hTert-immortalized normal bladder epithelial cells by investigating whether exposure to caffeine can cause potential changes in the bladder proteome and/or biological pathways. In labeled LC-MS/MS proteomic analysis, 57 proteins are found as being differentially expressed in caffeine-treated bladder epithelial cells, compared to controls; this included 32 upregulated and 25 downregulated proteins. Further functional gene enrichment analysis reveals that caffeine affects major biological pathways, including those for "muscle contraction" and "chromatin assembly." These findings provide new scientific insights that may be useful in future studies investigating the role of caffeine in bladder dysfunctions.

| S-EPMC6493332 | biostudies-literature

Structure and Properties of DNA Molecules Over The Full Range of Biologically Relevant Supercoiling States.

Project description:Topology affects physical and biological properties of DNA and impacts fundamental cellular processes, such as gene expression, genome replication, chromosome structure and segregation. In all organisms DNA topology is carefully modulated and the supercoiling degree of defined genome regions may change according to physiological and environmental conditions. Elucidation of structural properties of DNA molecules with different topology may thus help to better understand genome functions. Whereas a number of structural studies have been published on highly negatively supercoiled DNA molecules, only preliminary observations of highly positively supercoiled are available, and a description of DNA structural properties over the full range of supercoiling degree is lacking. Atomic Force Microscopy (AFM) is a powerful tool to study DNA structure at single molecule level. We here report a comprehensive analysis by AFM of DNA plasmid molecules with defined supercoiling degree, covering the full spectrum of biologically relevant topologies, under different observation conditions. Our data, supported by statistical and biochemical analyses, revealed striking differences in the behavior of positive and negative plasmid molecules.

| S-EPMC5906655 | biostudies-literature

Comparative Proteomic Profiling of Ectosomes Derived from Thyroid Carcinoma and Normal Thyroid Cells Uncovers Multiple Proteins with Functional Implications in Cancer.

Project description:Proteins carried by tumor-derived ectosomes play an important role in cancer progression, and are considered promising diagnostic markers. In the present study, a shotgun nanoLC-MS/MS proteomic approach was applied to profile and compare the protein content of ectosomes released in vitro by normal human thyroid follicular epithelial Nthy-ori 3-1 cells and human anaplastic thyroid carcinoma (TC) 8305C cells. Additionally, the pro-migratory and pro-proliferative effects of Nthy-ori 3-1- and 8305C-derived ectosomes exerted on the recipient cells were assessed in wound closure and Alamar Blue assays. A total of 919 proteins were identified in all replicates of 8305C-derived ectosomes, while Nthy-ori 3-1-derived ectosomes contained a significantly lower number of 420 identified proteins. Qualitative analysis revealed 568 proteins present uniquely in 8305C-derived ectosomes, suggesting their applicability in TC diagnosis and management. In addition, 8305C-derived ectosomes were able to increase the proliferation and motility rates of the recipient cells, likely due to the ectosomal transfer of the identified cancer-promoting molecules. Our description of ectosome protein content and its related functions provides the first insight into the role of ectosomes in TC development and progression. The results also indicate the applicability of some of these ectosomal proteins for further investigation regarding their potential as circulating TC biomarkers.

| S-EPMC8997476 | biostudies-literature

Proteomic Identification of the Galectin-1-Involved Molecular Pathways in Urinary Bladder Urothelial Carcinoma.

Project description:Among various heterogeneous types of bladder tumors, urothelial carcinoma is the most prevalent lesion. Some of the urinary bladder urothelial carcinomas (UBUCs) develop local recurrence and may cause distal invasion. Galectin-1 de-regulation significantly affects cell transformation, cell proliferation, angiogenesis, and cell invasiveness. In continuation of our previous investigation on the role of galectin-1 in UBUC tumorigenesis, in this study, proteomics strategies were implemented in order to find more galectin-1-associated signaling pathways. The results of this study showed that galectin-1 knockdown could induce 15 down-regulated proteins and two up-regulated proteins in T24 cells. These de-regulated proteins might participate in lipid/amino acid/energy metabolism, cytoskeleton, cell proliferation, cell-cell interaction, cell apoptosis, metastasis, and protein degradation. The aforementioned dys-regulated proteins were confirmed by western immunoblotting. Proteomics results were further translated to prognostic markers by analyses of biopsy samples. Results of cohort studies demonstrated that over-expressions of glutamine synthetase, alcohol dehydrogenase (NADP⁺), fatty acid binding protein 4, and toll interacting protein in clinical specimens were all significantly associated with galectin-1 up-regulation. Univariate analyses showed that de-regulations of glutamine synthetase and fatty acid binding protein 4 in clinical samples were respectively linked to disease-specific survival and metastasis-free survival.

| S-EPMC5979315 | biostudies-literature

A urinary microRNA signature can predict the presence of bladder urothelial carcinoma in patients undergoing surveillance.

Project description:BackgroundThe objective of this study was to determine whether microRNA (miRNA) profiling of urine could identify the presence of urothelial carcinoma of the bladder (UCB) and to compare its performance characteristics to that of cystoscopy.MethodsIn the discovery cohort we screened 81 patients, which included 21 benign controls, 30 non-recurrers and 30 patients with active cancer (recurrers), using a panel of 12 miRNAs. Data analysis was performed using a machine learning approach of a Support Vector Machine classifier with a Student's t-test feature selection procedure. This was trained using a three-fold cross validation approach and performance was measured using the area under the receiver operator characteristic curve (AUC). The miRNA signature was validated in an independent cohort of a further 50 patients.ResultsThe best predictor to distinguish patients with UCB from non-recurrers was achieved using a combination of six miRNAs (AUC=0.85). This validated in an independent cohort (AUC=0.74) and detected UCB with a high sensitivity (88%) and sufficient specificity (48%) with all significant cancers identified. The performance of the classifier was best in detecting clinically significant disease such as presence of T1 Stage disease (AUC=0.92) and high-volume disease (AUC=0.81). Cystoscopy rates in the validation cohort would have been reduced by 30%.ConclusionsUrinary profiling using this panel of miRNAs shows promise for detection of tumour recurrence in the surveillance of UCB. Such a panel may be useful in reducing the morbidity and costs associated with cystoscopic surveillance, and now merits prospective evaluation.

| S-EPMC4815774 | biostudies-literature

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