During disease, the complex forest of glycoproteins and glycolipids shielding the exterior of endothelial cells—called the glycocalyx— changes drastically. Should the glycocalyx degrade through enzyme activity, cell adhesion molecules and receptors key in regulating cell activation become exposed. Patients lose this protective wall of molecules, worsening disease.  A well-studied component of this glycocalyx is heparin sulfate; both its loss as a protective barrier and its resulting circulating fragments are associated with worsened diseases. The enzyme that degrades heparin sulfate is known as heparanase (HSPE), and in mouse models, overexpression of HSPE led to platelet hyperactivation and increased clotting. Therefore, we sought to examine the role of platelet HSPE in the context of sepsis, a disease that is known to have increase blood clotting and heparinase activity.

Using RNA-sequencing, we analyzed the change in mRNA levels of heparanase (HSPE) in platelets across multiple diseases: malaria, dengue, H1N1, and sepsis. This allowed us to measure which genes were being expressed more during which diseases. In every disease, HSPE mRNA was significantly increased but we observed the greatest change from healthy to diseased platelets in sepsis. Thus, we continued to focus on platelet HSPE in the context of sepsis.

When we measured the levels of HSPE mRNA in our septic platelets, we found that the production and expression of the mRNA drastically reduced by day 90 of sepsis (compared to the first day the patients were hospitalized for sepsis). This rise in HSPE mRNA production follows platelet activity, as platelets are hyperactive during active sepsis.

But are these mRNA being translated into HSPE proteins? To answer that, we first measured how many ribosomes were bound to the mRNA (from previously published data). We found a significant increase in bound ribosomes on the HSPE mRNA, signaling an increase in protein production. We then turned to our own platelet samples and measured protein levels via western blot. This also correlated with the increase in the initial inactive form of HSPE (pro-heparanase), further demonstrating that septic platelets begin to heavily produce HSPE during active disease. Interestingly, only the levels of active HSPE protein levels were higher in the platelets of deceased septic patients compared to those in surviving patients. Finally, we measured platelet HSPE activity and found that septic platelets have nearly 3-4 times more HSPE activity. Therefore, the increase in HSPE proteins translates into increase in heparin sulfate cleaving during sepsis.

This research sheds light on platelet’s role in degrading a component of the protective glycocalyx , which in turn drastically alters the inflammatory response during sepsis.