Carbon Monoxide Atmosphere Allows Platelets To Retain Function and Hinder Pathogen Growth.

Platelets are the sole blood fraction which is currently preserved at room temperature and thus exposed to microorganisms, specifically bacterial, growth. Bacterial contamination in platelet concentrate (PC), in a range of up to 0.4%, is the major cause of morbidity and mortality in transfusion medicine. To extend the current limited shelf life of 5 days for platelets, fast screening methods have been developed. Alternatively, methods for inactivation of bacterial growth by DNA crosslinking are provided. The latter include treatment by psoralen or photodynamic inactivation such as riboflavin activated by irradiation. The drawback of these efficient methods which relates to large investment and unknown future side effects, calls for alternative preservation methods. In the current study we demonstrate an option to store PC under sealed anaerobic, carbon monoxide atmosphere, in which neutral pH is kept by bicarbonate (Cob). The procedure is simple and cost effective.

Freshly isolated platelets (PC) were prepared by the Israel Central Blood Bank and stored in plastic flasks either allowing air transfer (control) or in gas non-transparent, Cob, at 22°C. Growth of inoculated bacteria in Cob stored PC was arrested immediately or after one log. Integrity of Cob platelets was better than control as assessed by leak of lactic dehydrogenase (LDH). Leak of LDH was lower as compared to control cells. P-selectin exposure on long term preserved platelets (days 6–9) was lower in Cob preserved platelets as compared to identical control PC. Phosophatydil serine (PS) measurement using flow cytometry indicated a large exposure in Cob platelets stored longer than 4 days, concomitant with consumption of all glucose, the energy source for anaerobic metabolism, in plasma. However, introduction of air resulted in a reversible reduction of the exposed PS. To better mimic in vivo viability of stored platelets, additional experiments were carried out using reconstituted blood (R-blood) prepared as follows: Platelets were removed from freshly drawn blood and stored PC were added to yield a concentration of ~ 350,000/μl. Aggregability of control and Cob R-blood were compared using a “paleletworks” kit (Helena company). Aggregability of stored PC was reduced with time. Nevertheless, aggregability of Cob platelets stored in R-blood was always significantly higher than its twin, air stored, sample. Adhesion of the stored platelets was estimated using Cone and Plate Analyzer (Diamed). No differences were found, within error, between air and Cob stored R-blood. In vivo survival of the stored platelets was assessed using a previously described rabbit model. Survival time declined with prolonged preservation in both control and COb stored platelets. No differences within error were observed between control and COb preserved platelets. These results indicate the option to use COb method for storage of platelets in a viable, pathogen reduced manner.