Liquid Chromatography-Mass Spectrometry-Based Metabolomics of Nonhuman Primates after 4 Gy Total Body Radiation Exposure: Global Effects and Targeted Panels.

Rapid assessment of radiation signatures in noninvasive biofluids may aid in assigning proper medical treatments for acute radiation syndrome (ARS) and delegating limited resources after a nuclear disaster. Metabolomic platforms allow for rapid screening of biofluid signatures and show promise in differentiating radiation quality and time postexposure. Here, we use global metabolomics to differentiate temporal effects (1-60 d) found in nonhuman primate (NHP) urine and serum small molecule signatures after a 4 Gy total body irradiation. Random Forests analysis differentially classifies biofluid signatures according to days post 4 Gy exposure. Eight compounds involved in protein metabolism, fatty acid ß oxidation, DNA base deamination, and general energy metabolism were identified in each urine and serum sample and validated through tandem MS. The greatest perturbations were seen at 1 d in urine and 1-21 d in serum. Furthermore, we developed a targeted liquid chromatography tandem mass spectrometry (LC-MS/MS) with multiple reaction monitoring (MRM) method to quantify a six compound panel (hypoxanthine, carnitine, acetylcarnitine, proline, taurine, and citrulline) identified in a previous training cohort at 7 d after a 4 Gy exposure. The highest sensitivity and specificity for classifying exposure at 7 d after a 4 Gy exposure included carnitine and acetylcarnitine in urine and taurine, carnitine, and hypoxanthine in serum. Receiver operator characteristic (ROC) curve analysis using combined compounds show excellent sensitivity and specificity in urine (area under the curve [AUC] = 0.99) and serum (AUC = 0.95). These results highlight the utility of MS platforms to differentiate time postexposure and acquire reliable quantitative biomarker panels for classifying exposed individuals.

Identifying Urinary and Serum Exosome Biomarkers for Radiation Exposure Using a Data Dependent Acquisition and SWATH-MS Combined Workflow.

PURPOSE: Early and accurate assessment of radiation injury by radiation-responsive biomarkers is critical for triage and early intervention. Biofluids such as urine and serum are convenient for such analysis. Recent research has also suggested that exosomes are a reliable source of biomarkers in disease progression. In the present study, we analyzed total urine proteome and exosomes isolated from urine or serum for potential biomarkers of acute and persistent radiation injury in mice exposed to lethal whole body irradiation (WBI). METHODS AND MATERIALS: For feasibility studies, the mice were irradiated at 10.4 Gy WBI, and urine and serum samples were collected 24 and 72 hours after irradiation. Exosomes were isolated and analyzed using liquid chromatography mass spectrometry/mass spectrometry-based workflow for radiation exposure signatures. A data dependent acquisition and SWATH-MS combined workflow approach was used to identify significantly exosome biomarkers indicative of acute or persistent radiation-induced responses. For the validation studies, mice were exposed to 3, 6, 8, or 10 Gy WBI, and samples were analyzed for comparison. RESULTS: A comparison between total urine proteomics and urine exosome proteomics demonstrated that exosome proteomic analysis was superior in identifying radiation signatures. Feasibility studies identified 23 biomarkers from urine and 24 biomarkers from serum exosomes after WBI. Urinary exosome signatures identified different physiological parameters than the ones obtained in serum exosomes. Exosome signatures from urine indicated injury to the liver, gastrointestinal, and genitourinary tracts. In contrast, serum showed vascular injuries and acute inflammation in response to radiation. Selected urinary exosomal biomarkers also showed changes at lower radiation doses in validation studies. CONCLUSIONS: Exosome proteomics revealed radiation- and time-dependent protein signatures after WBI. A total of 47 differentially secreted proteins were identified in urinary and serum exosomes. Together, these data showed the feasibility of defining biomarkers that could elucidate tissue-associated and systemic response caused by high-dose ionizing radiation. This is the first report using an exosome proteomics approach to identify radiation signatures.

Prevalence of and risks for internal contamination among hospital staff caring for a patient contaminated with a fatal dose of polonium-210.

BACKGROUND: Alexander Litvinenko died on November 23, 2006, from acute radiation sickness syndrome caused by ingestion of polonium-210 (²¹°Po). OBJECTIVE: The objective was to assess the prevalence of and risk factors for internal contamination with ²¹°Po in healthcare workers (HCWs) caring for the contaminated patient. SETTING: Hospital. PARTICIPANTS: HCWs who had direct contact with the patient. METHODS: We interviewed 43 HCWs and enquired about their activities and use of personal protective equipment (PPE). Internal contamination was defined as urinary ²¹°Po excretion above 20 mBq within 24 hours. We obtained risk ratios (RRs) for internal contamination using Poisson regression. RESULTS: Thirty-seven HCWs (86%) responded, and 8 (22%) showed evidence of internal contamination, all at very low levels that were unlikely to cause adverse health outcomes. Daily care of the patient (washing and toileting the patient) was the main risk factor (RR, 3.6 [95% confidence interval (CI), 1.1-11.6]). In contrast, planned invasive procedures were not associated with a higher risk. There was some evidence of a higher risk associated with handling blood samples (RR, 3.5 [95% CI, 0.8-15.6]) and changing urine bags and/or collecting urine samples (RR, 2.7 [95% CI, 0.8-9.5]). There was also some evidence that those who reported not always using standard PPE were at higher risk than were others (RR, 2.5 [95% CI, 0.8-8.1]). CONCLUSIONS: The sensitive quantitative measurement enabled us to identify factors associated with contamination, which by analogy to other conditions with similar transmission mechanisms may help improve protection and preparedness in staff dealing with an ill patient who experiences an unknown illness.