Troubleshooting in platelet storage temperature and new perspectives through proteomics

Dear Sir,

With reference Dr. Badlou’s letter “Reaction to the review article Troubleshooting in platelet storage temperature and new perspectives through proteomics”¹, we would first of all like to recall the basic aims of our review, which were to describe cold- and agonist-induced platelet activation and their basic differences, cover the main platelet storage lesions occurring both at room and refrigerating temperatures and, finally, glance at the latest approaches which can be applied to dissect untoward effects of the storage of platelet concentrates. This is only to underline our intention: to provide a state-of-art review of storage-related troubleshooting in platelet preservation, and what proteomics can tell regarding this issue.

As already shown by many studies in this field, proteomics offers the great advance of revealing virtually all protein modifications present in a certain cellular system, highlighting time- and condition-dependent changes. The study of proteins rather than genes allows an analysis of the final molecular scenario, including post-translational modifications, protein degradation and so on. Proteomics can unravel previously unknown products, giving a snapshot of a certain and variable cellular status. It can, therefore, be used as a non-targeted strategy, which is particularly useful when a certain molecular mechanism has been only partially dissected. Furthermore, proteomics is a holistic approach, which has a resemblance only with microarray technology. Nevertheless, we had no wish to disdain other methods applied in this field; we simply wanted to have a look at this topic from our point of view, without forgetting to note the intrinsic limitations. In fact, while describing two-dimensional electrophoresis, we emphasized its unsuitability for the dissection of membrane signalling pathways, which are rich in hydrophobic protein species. However, we claimed that platelets are an ideal target for proteomic analysis, since they lack both a nucleus and huge transcriptional activity.

Moreover, many published works dealing with proteomics emphasise the need to complement proteomic techniques to get a complete view of a certain issue, being aware of their technical limitations; others tried to gather “omic” information, others underlined the need for proteomic findings to be translated from the laboratory to the clinic. A wealth of articles have been published in the last few years on platelet proteomics, from analysis of platelet activation to storage-related modifications to protein profiling and structural/functional characterisation (we cannot list all the references for editorial constraints, but just a quick search in Pubmed gives an idea). This undoubtedly reflects the collective interest of the scientific community, or, if preferable, of a part of it. Wrong global fascination?

We wonder why Dr. Badlou said we stated that other “omic” approaches apart from proteomics failed to improve platelet quality, since at the end of our review we wrote: “Undoubtedly, the rapid buildup of proteomic, transcriptomic and metabolomic information in medical sciences, due to the rapid technical progress in “omic” sciences, could be potentially translated into dramatic improvements of current clinical practice”.

Later on, Dr. Badlou disagreed with our sentence “In order to improve storage of platelet concentrates (PCs), lowering temperature has been attempted as well, although results were not as positive as expected, since platelet do not tolerate refrigeration”. This statement was written after reading several papers concerning increased clearance of refrigerated platelets (one for all, see the paper by Wandell et al.²). This phenomenon, and particularly the mechanism of clearance of long-term (=48 h) chilled platelets, was recently clarified³ at the molecular level. Although platelet refrigeration prevents lactate accumulation and pH variations better than storage at 22 °C, change in shape and exposure of P-selectin, both suggested as a triggering factor for platelet clearance, were also noticed in chilled platelets precisely by Dr. Badlou and his colleagues⁴. When looking at the final product ready for the recipient, in our opinion the main parameter to consider is clearance: in fact, the best product becomes the worst if not available for the recipient. Nonetheless, we know that platelet clearance does not correlate with platelet functionality.

Dr. Badlou said we forgot to mention their considerable work in this field; if so, we must clarify that this was not intentional. We made our description as accurate as we could, but it would be impossible to cover all aspects of this topic.

Perhaps, we should not focus on refrigeration, and not even on platelet treatment before cold storage, as Dr. Badlou did, but instead on platelet re-warming after refrigeration. In fact, cold-induced anomalies in platelet concentrates emerge at this time. We refer to 0/37 °C-triggered GPIba perturbations and loss-of-function, cold-induced production of thromboxane A2 after re-warming, metabolic alterations due to functional damages of proteins and apoptotic pathways. Even if galactosylation could be a feasible way to avoid platelet clearance of short-term chilled platelets, it has been demonstrated not to impair phagocytosis of long-term chilled products (long-term being a period longer than 48 hours). The reason for this behaviour has been partially clarified by the recent findings of Hoffmeinster and colleagues³, who showed that hepatocytes and not macrophages are responsible for the clearance of long-term chilled platelets. We referred to platelets stored for 48 hours at 4 °C as long-term stored platelets since their behaviour has been compared to short-term chilled counterparts (2 hours), but in general both could be considered short times in the context of blood bank needs. Thus, when Dr. Badlou claims the main question is not room temperature or cold storage but rather “how under cold conditions?”, let us whisper “perhaps”.

As rightly mentioned by Dr. Badlou et al., it is important to maintain a realistic approach when dealing with matters of clinical interest. Nonetheless, his first approach towards assessing the usefulness of metabolic suppression in avoiding untoward effects in cold-stored platelets were performed in closed tubes and with antimycin A⁵, which differ from current clinical practices and incompatible with recipient’s safety, respectively. We guess the same is true for the proteomic approach, which is a relatively new in this field: let us wait for its findings to evolve from broad, “metaphysical” speculation to transferable knowledge in the daily routine of assessing platelet quality.