Thrombin-Induced Oxidative Phosphorylation in Platelets Is Repressed By Dengue Virus through Inhibition of the PI3K/Akt Signaling Pathway

Background:

There are many very important events that condition the progression to severe dengue, these include cytokine storm and a profound thrombocytopenia. Still, the molecular mechanisms underlying these events are not fully understood. Increasing evidence supports the fact that platelet dysfunction, in patients with dengue, could be due to direct interaction between platelets and dengue virus. This event induces changes in platelet function and contributes to pathogenesis. Recent results from our laboratory have shown that thrombin induces oxygen consumption through activation of glycoprotein Ib and subsequent phosphorylation of PI3K and Akt enzymes. Here we want to evaluate changes in these signalling pathways as well as their impact on mitochondrial function.

Aims: To evaluate DENV2 effects on platelet function, PI3K/Akt signaling, oxidative phosphorylation and GPIb expression.

Methods:

Blood samples from healthy volunteers were collected and platelet fraction was obtained. Purified DENV2 particles were incubated with platelets for 2 h at 37ºC. Then, morphological changes, activation and aggregation, mitochondrial function and surface markers expression were tested in platelets.

Results:

We found that DENV2 induces conspicuous morphological changes on platelets not induced by other arboviruses; increases CD41 (p<0.05), CD62P (8979 ± 1189 vs

1723 ± 160 MFI; p<0.05) and reduces CD42b (p<0.5) expressions on platelet surface; reduces agonist-induced platelet aggregation (30 ± 10 vs 71 ± 9%; p<0.05) through increases intracellular NO production (0.4 ± 0.2 vs 0.07 ± 0.01; p<0.05); inhibits thrombin-induced oxidative phosphorylation (45 ± 6.69 vs 348 ± 26.9 ngatomO/min/10 ⁹ platelets; p<0.0001) through PI3K/Akt signaling pathways inhibition (p<0.001). Surprisingly, rDC-SIGN treatment reverts DENV2 effects on platelet activation (9693 ± 1033 vs 2379 ± 390 MFI; p<0.01).

Conclusions:

DENV2 inhibits thrombin-induced oxidative phosphorylation through PI3K/Akt signaling pathway inhibition. Dengue virus inhibits thrombin-induced phosphorylation at different levels in the platelet. First, by reducing the expression of the GPIb molecule on the platelet surface. This molecule is important for thrombin-induced intracellular signalling. Second, dengue virus inhibits the PI3K/Akat signalling pathway, which is important for the induction of oxidative phosphorylation in mitochondria. And third, dengue virus increases intracellular nitric oxide levels and promotes its excretion into the extracellular milieu, this mechanism could be involved in the inhibition of platelet aggregation processes.These changes induce by short-term interaction of platelets with DENV2 has a direct effect over platelet function and may be correlated with the clinical manifestations observed during severe dengue stage, such as thrombocytopenia and hemorrhages. The overall impact of these changes in signaling pathways related to immune activity remains to be determinate, as does the potential to develop new pharmacological treatments.