Thrombin Inhibition Reduces the Expression of Brain Inflammation Markers upon Systemic LPS Treatment

Systemic inflammation and brain pathologies are known to be linked. In the periphery, the inflammation and coagulation systems are simultaneously activated upon diseases and infections. Whether this well-established interrelation also counts for neuroinflammation and coagulation factor expression in the brain is still an open question. Our aim was to study whether the interrelationship between coagulation and inflammation factors may occur in the brain in the setting of systemic inflammation. The results indicate that systemic injections of lipopolysaccharide (LPS) upregulate the expression of both inflammatory and coagulation factors in the brain. The activity of the central coagulation factor thrombin was tested by a fluorescent method and found to be significantly elevated in the hippocampus following systemic LPS injection (0.5 ± 0.15 mU/mg versus 0.2 ± 0.03 mU/mg in the control). A panel of coagulation factors and effectors (such as thrombin, FX, PAR1, EPCR, and PC) was tested in the hippocampus, isolated microglia, and N9 microglia cell by Western blot and real-time PCR and found to be modulated by LPS. One central finding is a significant increase in FX expression level following LPS induction both in vivo in the hippocampus and in vitro in N9 microglia cell line (5.5 ± 0.6- and 2.3 ± 0.1-fold of increase, resp.). Surprisingly, inhibition of thrombin activity (by a specific inhibitor NAPAP) immediately after LPS injection results in a reduction of both the inflammatory (TNFα, CXL9, and CCL1; p<0.006) and coagulation responses (FX and PAR1; p<0.004) in the brain. We believe that these results may have a profound clinical impact as they might indicate that reducing coagulation activity in the setting of neurological diseases involving neuroinflammation may improve disease outcome and survival

Recovery from trauma induced amnesia correlates with normalization of thrombin activity in the mouse hippocampus.

Transient amnesia is a common consequence of minimal traumatic brain injury (mTBI). However, while recent findings have addressed the mechanisms involved in its onset, the processes contributing to its recovery have not yet been addressed. Recently, we have found that thrombin is detected at high concentrations in the brain of mice after exposure to mTBI and that in such settings amnesia is rescued by either inhibiting thrombin activity or by blockade of PAR1. Here, we report that mice spontaneously recover from amnesia after two weeks from mTBI exposure. At this time point, long term potentiation was equally evoked in injured vs. control animals with thrombin concentration in the brain being normalized at this stage. These findings, which refer to the specific aspect of memory retrieval upon mTBI, together with our previous work, hint to a strong correlation between cognitive defects in the context of mTBI and thrombin concentrations in the brain. This may suggest that a possible scavenging of thrombin in the brain at early phases following mTBI may improve memory function

At two weeks upon injury, brain RNA levels of prothrombin, PAR1 and factor X were comparable between mTBI exposed animals and controls.

<p>At two weeks upon injury, brain RNA levels of prothrombin, PAR1 and factor X were comparable between mTBI exposed animals and controls.</p

Long Term Potentiation (LTP) is equally evoked in mTBI and control animals at two weeks following injury.

<p>(a) While twenty fours upon mTBI, animals exhibited a lower LTP compared to controls., (b) no difference in LTP could be detected in mTBI exposed animals vs. control at two weeks upon injury. Sample illustrations are at the indicated time points, the arrow indicates the time of high frequency stimulation delivery.</p

At two weeks upon injury, brain thrombin activity and concentration are similar between mTBI exposed animals and controls.

<p>(a) Thrombin activity as well as its protein levels (b) assessed as described in the methods section were comparable between mTBI and control brain slices. (c) PAR1 and (d) factor X were slightly elevated in mTBI animals compared to control. Refer to text for statistics.</p