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Proteomics analysis of extracellular vesicles purified by heparin chromatography.

ABSTRACT: Extracellular vesicles (EVs) were isolated from U2OS cell (Human) conditioned medium (Input) was fractionated into two subpopulations named non-heparin binding (NHB) and heparin-binding (HB). Proteomics analysis was performed on Input, NHB and HB samples. The two main goals for this analysis are: first, do a proteomics profiling study to identify proteins present in each of the 3 samples; second, perform quantitative analysis of the level of identified proteins in the samples and investigate if there is a certain enrichment pattern differences in samples "NHB" and "HB". Three biological replicates are included.

INSTRUMENT(S): Thermo Scientific instrument model

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

DISEASE(S): Bone Osteosarcoma

SUBMITTER: Yijun Zhou

LAB HEAD: Dirk Dittmer

PROVIDER: PXD039845 | Pride | 2023-06-13

REPOSITORIES: pride

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			Action	DRS
	20220401_OE480_PC951-Dittmer_1.raw	Raw
	20220401_OE480_PC951-Dittmer_2.raw	Raw
	20220401_OE480_PC951-Dittmer_3.raw	Raw
	20220401_OE480_PC951-Dittmer_4.raw	Raw
	20220401_OE480_PC951-Dittmer_5.raw	Raw

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Project description:Background: Mesenchymal stromal cells (MSCs) are a promising cell source for tissue engineering and regenerative medicine. In our lab, we found that cell preparations from bone marrow of many donors had a limited capacity for in vitro-differentiation into the osteogenic and chondrogenic lineages although this is a capacity claimed to be inherent to this kind of cells. Therefore the current study was designed to test the hypothesis whether the amount of heparin used as anti-coagulant during bone marrow harvest has inhibitory influence on the in vitro-differentiation capacity of the isolated MSCs. Methods: Bone marrow was obtained from twelve donors with good physical condition during preparation of the femoral cavity for the implantation of the femoral stem for a total hip arthroplasty in the absence or presence of heparin. Isolated MSCs were characterized by plastic adhesion, morphology, population doubling times, expression of cell surface antigens and in vitro-differentiation into the adipogenic, osteogenic and chondrogenic lineages. In addition, transcriptome analyses were performed. Results: Bone marrow prepared in the absence of heparin showed a comparable processability as bone marrow in the presence of heparin. Notably, no coagulation was observed. The number of mononuclear cells retrieved from the density gradient was independent of heparin addition. Moreover, none of the phenotypic or functional parameters assessed correlated with the addition of heparin. Transcriptome and quantitative realtime PCR analyses demonstrated that some selected genes were up- or downregulated in the cells isolated with the addition of heparin. However, this statement held true for only some but not all of the donors investigated in the present study. Conclusions: No correlation with heparin addition was observed with respect to the parameters assessed in the present study. This implies the absence of consequences both for the results of in vitro-investigations and also during in vivo-application, thereby ruling out heparin as a potential source of disparate results that are described in the literature.

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Proteomics analysis of extracellular vesicles purified by heparin chromatography.

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