gms | German Medical Science

Objective: Previous research hypothesizes that eRNA is released upon tissue injury and subsequently activates inflammatory signaling. Previous data from our laboratory showed that eRNA is released into the subarchnoid space in an animal model of experimental SAH, and is associated with an activation of the innate immune system. This study aims to quantify eRNA release and investigate RNAse activity in patient samples.

Methods: CSF and EDTA samples from patients with aneurysmal SAH (n=20) were collected within the scope of a prospective study after acquiring approval by the ethics committee board. Control samples (n=5) were acquired from pseudotumor cerebri patients and healthy individuals. Sampling was conducted at four time points after bleeding onset (day 1,4,7,14). CSF and blood samples were spun down twice at 500g for 5min at 4C° to remove debris and blood. For eRNA quantification, 80U/ml RNAse Inhibitor was added. Isolation of RNA was conducted with the Master Pure RNA Purification Kit; quantification was done with the QubitTM RNA Assay Kit (Invitrogen) and measured with the fluorometer. For RNAse activity, supernatant was added to poly(c)solution, RNAse buffer, and BSA solution and incubated at 37C° for 15min. Substrate degradation was determined by measuring the absorbance of the supernatant at 280nm.

Results: In the acute stage of SAH, we found a significant increase of eRNA in the CSF on day 1 (Ctrl vs. SAH: 2ng/ml vs. 351.4ng/ml;p=0.033). In concomitance, RNAse activity was significantly decreasedt in the CSF of SAH patients (Ctrl. vs. SAH: 136.9U/ml vs. 75.23 U/ml; p=0.045). In peripheral blood samples, an eRNA peak was observed at a later time point on day 14 (Ctrl. vs. SAH: 361ng/ml vs. 736.5ng/ml;p=0.039). This was paralleled by a significant increase of RNAse activity in EDTA blood (Ctrl. vs. SAH: 76.44U/ml vs. 143.1U/ml; p=0.004).

Conclusion: Aneurysmal SAH is associated with a significant increase in eRNA in the CSF, paralleled by a decrease in RNAse activity, underpinning previously described mechanistic data on our mouse model. Hence, inhibition of eRNA may represent a potential strategy in modulating neuroinflammatory signaling but further ex vivo studies are needed to understand the underlying pathway mechanism.

Figure 1 [Fig. 1]