INFLUENCE OF HEAD-DOWN TILT BEDREST ON DNA REPAIR CAPACITY IN PERIPHERAL BLOOD MONONUCLEAR CELLS

Abstract

INTRODUCTION Major limiting factors in human spaceflight are deleterious effects of reduced gravity and of space radiation exposure on health and performance. Radiation-induced cellular DNA damage, if not repaired or not correctly repaired, increases the risk of cancer and degenerative diseases. Likewise, reduced gravity may lead to musculoskeletal and cardiovascular deconditioning without appropriate countermeasures. We hypothesize that deconditioning could possibly hinder the recovery of cells from radiation damage. We aimed to develop a terrestrial ex vivo model to investigate cellular DNA repair while simulating microgravity effects using head-down-tilt (HDT) bedrest during the Spaceflight-Associated Neuro-Ocular Syndrome Countermeasures (SANS-CM) studies. METHODS The SANS CM campaigns comprised three experimental phases: 1) a 14-days baseline data collection (BDC) phase (BDC-14 through BDC- 1); 2) 30 days of 6° HDT bedrest phase (HDT1 through HDT30); and 3) a 14-days recovery (R+) phase (R+0 through R+13). Twelve participants enrolled in each campaign were healthy non-smoking men and women of 24-55 years with body height of 153-190 cm and body mass index of 19-30 kg/m². In the study, the countermeasure applied was either lower body negative pressure (LBNP) or upright sitting, both 2 x 3 h per day. Blood samples were obtained from the subjects 14 days before the bedrest (BDC-14), 10 and 28 days into head-down-tilt bedrest (HDT-10 and HDT-28) and after 10 days of recovery (R+10). Peripheral blood mononuclear cells (PBMC) were isolated by density gradient centrifugation using Greiner LeucosepTM tubes. We studied the ex vivo induction and repair of DNA double strand breaks, for which the cells were exposed to 1 and 4 Gy of X-rays and harvested after 0.5, 1, 2, 4 and 24 h after irradiation. DNA double strand breaks were detected via immunofluorescence staining of γH2AX that was quantified using flow cytometry. RESULTS γH2AX fluorescence intensity and the percentage of cells with unrepaired DNA reached a peak value after 2 h of X-rays irradiation and were greatly reduced after 24 h. For all blood collection time-points during the study, BDC-14, HDT-10, HDT-28 and R+10, the γH2AX fluorescence intensity did not differ significantly. Also, there was no significant difference between the two countermeasure groups, LBNP (n=12) and upright sitting (n=12). Furthermore, there was no significant interpersonal variance of DNA double strand break repair capacity. CONCLUSION DNA double strand break repair activity in PBMC remained unaffected by one month of HDT bedrest combined with the countermeasures LBNP or upright sitting, suggesting that the physical deconditioning does not modulate DNA repair capacity in case that daily countermeasures are applied. ACKNOWLEDGEMENT The study was funded by the National Aeronautics and Space Administration (NASA) and the German Aerospace Center, and performed at the :envihab research facility of the DLR Institute of Aerospace Medicine. The authors wish to gratefully acknowledge the bedrest participants who volunteered their time, without whom this project would not have been possible. We also thank the SANS CM study staff for their dedicated work and tireless effort.