STOREDB:STUDY1155 Radiation exposure of peripheral mononuclear blood cells alters the composition and function of released extracellular vesicles [DOI:10.20348/STOREDB/1155]

Study meta-data


STUDYIDSTOREDB:STUDY1155
CREATEDON2019-12-13 13:17:31
MODIFIEDON2019-12-13 13:17:31
UPLOADEROmid Azimzadeh
DOIDOI:10.20348/STOREDB/1155

Study details


STUDY NAME
Radiation exposure of peripheral mononuclear blood cells alters the composition and function of released extracellular vesicles
STUDY STATUS
Published: Open access to everyone
DATA SHARING POLICY
CC-Attribution Non-Commercial
COUNTRY
Germany
PRINCIPAL INVESTIGATOR
Dr.Simone Moertl
SPECIES
Homo sapiens
SIZE OF COHORT
0-999
OUTCOME
Cancer
MELODI RESEARCH PRIORITY
Identification, development and validation of biomarkers for radiation-induced health (cancer and non-cancer) effects through sound epidemiological studies
EXPOSURE CONTEXT
Medical
INTERNAL OR EXTERNAL EXPOSURE
External
TYPE OF EXTERNAL EXPOSURE
X-ray
TYPE OF INTERNAL EXPOSURE
X-ray
RADIONUCLIDE
caesium
AGE AT EXPOSURE
Adult
EXPOSURE PATTERN
Acute
DOSE RATE
High
BIOLOGICAL SAMPLE AVAILABLE
Yes
ECOLOGICAL DATA
No
STUDY DESCRIPTION
Normal tissue toxicity is a dose-limiting factor in radiation therapy. One component of normal tissue that is continuously exposed during therapeutic irradiation is the circulating population of blood mononuclear cells (PBMC). PBMCs are highly sensitive to IR, however little is known how ionizing radiation (IR) affects the PBMC response on a systemic level. Therefore it was the aim of this study to investigate whether IR was capable to induce changes in the composition and function of extracellular vesicles secreted from PBMCs after radiation exposure to different doses.
Whole blood samples from healthy donors were irradiated with 0 Gy, 0,1 Gy, 2 Gy or 6 Gy X-rays and PBMC-secreted EVs were isolated 72 h later. Proteome analysis of EVs by label-free proteomics identified 606 proteins, of which 283 significantly changed their abundance after irradiation. microRNA expression analysis by small RNA sequencing revealed 379 microRNAs, of which 34 were changed after irradiation. For both, proteome and microRNA data, principal component analysis showed a dose-dependent separation of control and exposed groups. An IPA network analysis of the radiation-regulated EV proteins and microRNAs consistently predicted an association of deregulated components with apoptosis, cell death and survival. Functional studies identified endothelial cells as efficient EV recipient system, whereby irradiation of recipient cells further increased the uptake. Furthermore we detected an apoptosis suppressive effect of EVs from irradiated PBMCs in endothelial recipient cells.
In summary, our study demonstrates that IR triggers the communication between PBMCs and endothelial cells. We identified EVs from irradiated PBMC donors as transmitters of protective signals to irradiated endothelial cells. Thus our data may lead to the discovery of biomarker candidates for radiation dosimetry and even more importantly, they suggest EVs a novel systemic communication pathway between irradiated and non-irradiated normal, non-cancer tissues.
PUBMED ID
1
MEAN DURATION OF FOLLOW-UP (WEEKS)
1

STOREDB:DATASET1220 Proteomic analysis on the extracellular vesicles of irradiated peripheral mononuclear blood cells [DOI:10.20348/STOREDB/1155/1220]


Created on:2019-12-19 10:18:30 Modified On:2019-12-19 11:26:18
DATASET NAME
Proteomic analysis on the extracellular vesicles of irradiated peripheral mononuclear blood cells
DOIDOI:10.20348/STOREDB/1155/1220
DATASET DESCRIPTION
Filter aided sample preparation (FASP) digest
Each 10 g were digested using a modified FASP procedure (Wiśniewski et al., 2009). After reduction and alkylation using DTT and IAA, the proteins were centrifuged on a 30 kDa cutoff filter device (Sartorius), washed thrice with UA buffer (8 M urea in 0.1 M Tris/HCl pH 8.5) and twice with 50 mM ammoniumbicarbonate. The proteins were digested for 2 hours at room temperature using 1 g Lys-C (Wako Chemicals, Neuss, Germany) and for 16 hours at 37C using 2 g trypsin (Promega, Mannheim, Germany). After centrifugation (10 min at 14 000 g) the eluted peptides were acidified with 0.5% TFA and stored at -20C.

LC-MSMS measurements
LC-MS/MS analysis was performed on a Q-Exactive HF mass spectrometer (Thermo Scientific) online coupled to an Ultimate 3000 nano-RSLC (Dionex). Tryptic peptides were automatically loaded on a C18 trap column (300 m inner diameter (ID) 5 mm, Acclaim PepMap100 C18, 5 m, 100 , LC Packings) at 30l/min flow rate prior to C18 reversed phase chromatography on the analytical column (nanoEase MZ HSS T3 Column, 100, 1.8 m, 75 m x 250 mm, Waters) at 250nl/min flow rate in a 95 minutes non-linear acetonitrile gradient from 3 to 40% in 0.1% formic acid. Profile precursor spectra from 300 to 1500 m/z were recorded at 60000 resolution with an automatic gain control (AGC) target of 3e6 and a maximum injection time of 50 ms. TOP10 fragment spectra of charges 2 to 7 were recorded at 15000 resolution with an AGC target of 1e5, a maximum injection time of 50 ms, an isolation window of 1.6 m/z, a normalized collision energy of 28 and a dynamic exclusion of 30 seconds.

Quantitative data analysis using Progenesis QI for proteomics
Generated raw files were analyzed using Progenesis QI for proteomics (version 3.0, Nonlinear Dynamics, part of Waters) for label-free quantification as described previously (Hauck et al., 2010; Merl et al., 2012). Features of charges 2-7 were used and all MSMS spectra were exported as mgf file. Peptide search was performed using Mascot search engine (version 2.5.1) against the Ensembl human protein database (83462 sequences, 31286148 residues). Search settings were: 10 ppm precursor tolerance, 0.02 Da fragment tolerance, one missed cleavage allowed. Carbamidomethyl on cysteine was set as fixed modification, deamidation of glutamine and asparagine allowed as variable modification, as well as oxidation of methionine. Applying the percolator algorithm (Brosch et al., 2009) resulted in a peptide false discovery rate (FDR) of 0.54%. Search results were reimported in the Progenesis QI software. Proteins were quantified by summing up the abundances of all unique peptides per protein. Resulting normalized protein abundances were used for calculation of fold-changes and statistical values were exported from the Progenesis QI software. For final quantifications, proteins were identified with at least 2 unique peptides and with ratios greater than 1.30-fold or less than 0.77-fold (p< 0.05) were defined as being significantly differentially expressed.
STOREDB:FILE11549

AZO9526A16MA3 016-Lym C 6Gy AZO9526A16MA3-40-349078

Uploaded on: 2019-12-19 12:13:29
Type: image/x-panasonic-raw
File size: 1 gigabytes
STOREDB:FILE11548

AZO9526A10MA3 010-Lym A 2Gy AZO9526A10MA3-40-349042

Uploaded on: 2019-12-19 12:06:07
Type: image/x-panasonic-raw
File size: 1 gigabytes
STOREDB:FILE11547

AZO9526A11MA3 011-Lym B 2Gy AZO9526A11MA3-40-349049

Uploaded on: 2019-12-19 12:05:45
Type: image/x-panasonic-raw
File size: 1 gigabytes
STOREDB:FILE11546

AZO9526A3MA3 003-Lym B 0Gy AZO9526A3MA3-40-349003

Uploaded on: 2019-12-19 11:58:54
Type: image/x-panasonic-raw
File size: 1 gigabytes
STOREDB:FILE11545

AZO9526A2MA3 002-Lym A 0Gy AZO9526A2MA3-40-348899

Uploaded on: 2019-12-19 11:58:28
Type: image/x-panasonic-raw
File size: 1 gigabytes
STOREDB:FILE11544

AZO9526A7MA3 007-Lym B 01Gy AZO9526A7MA3-40-343895

Uploaded on: 2019-12-19 11:51:42
Type: image/x-panasonic-raw
File size: 1 gigabytes
STOREDB:FILE11543

AZO9526A12MA3 012-Lym C 2Gy AZO9526A12MA3-40-349055

Uploaded on: 2019-12-19 11:51:11
Type: image/x-panasonic-raw
File size: 1 gigabytes
STOREDB:FILE11542

AZO9526A14MA3 014-Lym A 6Gy AZO9526A14MA3-40-349064

Uploaded on: 2019-12-19 11:44:16
Type: image/x-panasonic-raw
File size: 1 gigabytes
STOREDB:FILE11541

AZO9526A7MA4 007-Lym B 01Gy AZO9526A7MA4-40-349028

Uploaded on: 2019-12-19 11:43:41
Type: image/x-panasonic-raw
File size: 1 gigabytes
STOREDB:FILE11540

AZO9526A4MA3 004-Lym C 0Gy AZO9526A4MA3-40-349009

Uploaded on: 2019-12-19 11:36:31
Type: image/x-panasonic-raw
File size: 1 gigabytes
STOREDB:FILE11539

AZO9526A6MA3 006-Lym A 01Gy AZO9526A6MA3-40-349021

Uploaded on: 2019-12-19 11:36:17
Type: image/x-panasonic-raw
File size: 1 gigabytes