Dataset Information

Analysis of extracellular matrix proteins from 3D cultures of primary prostate fibroblasts, DU145 cells and mixed cells

ABSTRACT: Composition and organization of extracellular matrix (ECM) in tumor stroma are important regulators of cancer cell behavior. Therefore, ECM produced by cancer associated fibroblasts is often used in in vitro experiments. Here, a proteome wide analysis of ECM proteins in 3D culture is analysed.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture, Prostate Cancer Cell Line, Fibroblast

DISEASE(S): Prostate Adenocarcinoma

SUBMITTER: Pekka Rappu

LAB HEAD: Jyrki Heino

PROVIDER: PXD006563 | Pride | 2018-03-13

REPOSITORIES: Pride

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Publications

The composition of prostate core matrisome in vivo and in vitro unveiled by mass spectrometric analysis.

Ojalill Marjaana M Rappu Pekka P Siljamäki Elina E Taimen Pekka P Boström Peter P Heino Jyrki J

The Prostate 20180309 8

<h4>Background</h4>The composition and organization of extracellular matrix (ECM) are important regulators of cell behavior. In particular in the prostate, this central role of the ECM is further stressed by the fact that several potential markers of prostate stem cells are matrix receptors.<h4>Methods</h4>We established 12 fibroblastic cell lines from cancerous and non-cancerous areas of six prostates and allowed the cells to produce ECM under cell culture conditions. We also performed a proteo ...[more]

PMID: 29520855

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Project description:Proteomics, the temporal study of proteins expressed by an organism, is a powerful technique that can reveal how organisms respond to biological perturbations, such as disease and environmental stress. Yet, the use of proteomics for addressing ecological questions has been limited, partly due to inadequate protocols for the sampling and preparation of animal tissues from the field. Although RNAlater is an ideal alternative to freezing for tissue preservation in transcriptomics studies, its suitability for the field could be more broadly examined. Moreover, existing protocols require samples to be preserved immediately to maintain protein integrity, yet the effects of delays in preservation on proteomic analyses have not been thoroughly tested. Hence, we optimised a proteomic workflow for wild-caught samples. First, we conducted a preliminary in-lab test using SDS-PAGE analysis on aquaria-reared Octopus berrima confirming that RNAlater can effectively preserve proteins up to 6 h after incubation, supporting its use in the field. Subsequently, we collected arm tips from wild-caught Octopus berrima and preserved them in homemade RNAlater immediately, 3 h, and 6 h after euthanasia. Processed tissue samples were analysed by liquid chromatography tandem mass spectrometry to ascertain protein differences between time delay in tissue preservation, as well as the influence of sex, tissue type, and tissue homogenisation methods. Over 3500 proteins were identified from all tissues, with bioinformatic analysis revealing protein abundances were largely consistent regardless of sample treatment. However, nearly 10 % additional proteins were detected from tissues homogenised with metal beads compared to liquid nitrogen methods, indicating the beads were more efficient at extracting proteins. Our optimised workflow demonstrates that sampling non-model organisms from remote field sites is achievable and can facilitate extensive proteomic coverage without compromising protein integrity.

Project description:Background: Lamins A/C (encoded by the LMNA gene) can lead to dilated cardiomyopathy (DCM). Objectives: This study sought to undertake proteomic analysis of myocardial tissue to explore the postgenomic phenotype of end-stage lamin heart disease. Methods: Consecutive patients with end-stage lamin heart disease (LMNA-group, n=7) and ischaemic DCM (ICM-group, n=7) undergoing heart transplantation were enrolled. Samples were obtained from left atrium(LA), left ventricle(LV), right atrium(RA), right ventricle(RV) and interventricular septum(IVS). Liquid chromatography combined with mass-spectrometry was used for protein quantification. We compared protein concentrations in cardiac samples between LMNA and ICM groups. Proteins were considered differentially abundant if the quantitative difference was 1.5-fold and corrected p-value <0.05 at a false discovery rate of 0.01. Gene ontology(GO) enrichment analysis explored the related biological processes. Results: 4,247 proteins were identified in LMNA and ICM samples, of which 633 were differentially abundant in LA, 39 in LV, 181 in RA, 52 in RV, and 85 in IVS. Abundance of lamin A/C was reduced but lamin B (LMNB) increased in LMNA LA/RA tissue compared to ICM, but not in LV/RV. Transthyretin was more abundant in the LV/RV of LMNA compared to ICM while sarcomeric proteins such as titin and cardiac myosin heavy chain were generally reduced in RA/LA of LMNA. Protein expression profiling and GO enrichment analysis revealed sarcopenia, extracellular matrix(ECM) remodeling, deficient myocardial energetics, redox imbalances, and abnormal calcium handling in LMNA samples. Conclusion: Lamin heart disease is a biventricular and biatrial disease, characterized by sarcopenia, aberrant metabolism, and ECM remodeling. LMNB and transthyretin were unexpectedly abundant in the atria and ventricles respectively of patients with end-stage lamin heart disease potentially hinting to the possibility of compensatory responses.

Dataset Information

Analysis of extracellular matrix proteins from 3D cultures of primary prostate fibroblasts, DU145 cells and mixed cells

Publications

The composition of prostate core matrisome in vivo and in vitro unveiled by mass spectrometric analysis.

Similar Datasets

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