AZD8931 br Methods br Results br Discussion

Methods
Results
Discussion The results demonstrate that O2 partial pressure influences SPCs mobilization with repetitive treatments. Whether this is due to augmented NOS-3 activation requires additional study. SPCs mobilization in response to a variety of drugs is compromised by older age, prior radiotherapy and use of several types of chemotherapy (e.g. platinum compounds, alkylating agents, AZD8931 analog and lenalidomide) (Jantunen and Kvalheim, 2010). None of these agents were being administered to patients during our study. We have reported previously that SPCs mobilization in response to a single 2.0 ATA O2 exposure is the same between normal adults and those exposed to radiotherapy (Thom et al., 2006). Obviously, all patients in this study received radiotherapy but there was no significant difference in radiation dosage or patient age between the 2.0 and 2.5 ATA treatment groups (Table 1). There were no notable deviations in the pattern of SPCs mobilization among the patients despite taking a variety of medications listed in Table 1. Some of these medications are known to have positive effects on SPCs mobilization (e.g. short term statin use, paclitaxel, certain β-blockers such as nebivolol and carvedilol); while others have a negative impact on mobilization (e.g. bisphosphonates, long-term use of statins and trimethoprim/sulfamethoxazole) (Hristov et al., 2007; Sorrentino et al., 2011; Besler et al., 2008; Wu et al., 2010; Fuchs et al., 2000; Xu et al., 2013; Fernandez et al., 2008). None of the medications listed in Table 1 had been started in the time frame while patients were receiving HBO2 and all had been prescribed for over 2months prior to patient enrollment. One patient in the 2.0 ATA group had HIV and one in the 2.5 ATA group had renal failure and was undergoing dialysis. HIV does not impede the efficacy of chemotherapeutic agent-mediated SPCs mobilization and renal failure may modify mobilization by some drugs but does not completely abrogate responses (Badros et al., 2001; Hill et al., 2012; Re et al., 2013; Gabarre et al., 2000). Whether these disorders influence HBO2-mediated mobilization will require additional study. Clearly, there are differences in mobilization mechanisms between chemotherapeutic agents and HBO2. Contrary to many of the stem cell mobilization drugs HBO2 does not activate platelets or elevate leukocyte counts which can be thrombogenic (Ma et al., 2011; Powell et al., 2005; Thom, 2006, 2009). Intracellular regulatory protein contents were elevated in all post-HBO2 samples with no significant differences between protocols. Elevations are likely a characteristic of the bone marrow SPCs population primed for mobilization and a higher percentage is released with higher O2 dose. Lower protein levels in pre-HBO2 samples at the 10th and 20th treatments may reflect preferential perivascular sequestration of newly mobilized cells and/or protein degradation in cells remaining in the circulation for many hours. The difference in protein contents of newly mobilized SPCs has not been appreciated in mobilization studies involving chemotherapeutic agents. This is probably because responses to chemical agents proceed over a much longer time course. A weakness of this investigation is that perhaps alternative or additional surface markers should be used to better characterize the mobilized cells. With regard to neovascularization potential, this is difficult to determine given the ongoing debate over EPCs characterization (Pober, 2012). Elevated intracellular proteins of HBO2-mobilized cells suggest they may have improved propensity for growth/differentiation based on animal studies (Milovanova et al., 2008, 2009). HIF-3 and PARP were probed because they provide evidence that cells were not merely circulating endothelial cells or cells undergoing apoptosis. PARP levels would be expected to be quite low in apoptotic cells (Gajdusek et al., 2001). EPCs can be distinguished from mature CECs by determining ‘clonogenic’ proliferative capacity, but not by flow cytometric evaluation of surface markers (Pober, 2012). Our approach for assessing intracellular markers after membrane permeabilization precludes ex vivo growth analysis, which is why we probed for HIF-3. In animals we have found HBO2-mobilized SPCs that form new blood vessels and hence not CECs are well endowed with HIF-3, whereas HIF-3 normally is highly tissue restricted (to thymus, lung and a lesser extent in brain, heart and kidney) (Milovanova et al., 2009; Gu et al., 1998). Therefore, we conclude that the cells mobilized by hyperoxia are SPCs and that treatment pressure influences mobilization efficiency. Functional consequences of this response require further study.