Activation of TLR4 promotes oxidative inflammation via NADPH oxidase signaling in platelets of subjects with autism spectrum disorder

Autism spectrum disorder (ASD) is a complex neurobehavioral disorder where both immune system and neurological dysregulations are known to play crucial roles. Platelets are known to play key roles in systemic oxidative inflammation through production of oxidants. However, there is still scarcity of data regarding the role of blood platelets in ASD subjects. Platelets are known to express innate receptors such as toll-like receptor 4 (TLR4), transcription factors such as NFkB, and oxidative enzymes such as NADPH oxidase (NOX2), however the molecular mechanisms in generation of oxidative inflammation in platelets of ASD subjects have not been investigated until now. Therefore, this study was aimed at investigating expression of TLR4, NFkB, NOX2, reactive oxygen species (ROS) and lipid peroxides (a marker of oxidative damage on lipid membranes) in platelets of ASD children (n = 27) and typically developing children (TDC; n = 22) through molecular and biochemical analyses. Further, correlation analysis was carried out with severity of the disease. Our data showed an upregulated expression of TLR4 and p-NFkB in ASD platelets as compared to TDC platelets. Further, elevated TLR4/ NFkB signaling was linked with elevated NOX2-derived ROS and oxidative stress. It was confirmed by the utilization of NFkB and NOX2 inhibitors which caused reduction in LPS-induced ROS and oxidative stress in ASD platelets. Further, there was positive correlation between platelet oxidative stress and severity of the disease. Our data suggests that elevated TLR4/ NFkB signaling in platelets of ASD subjects might predispose them to increased systemic oxidative inflammation through NOX-2 derived ROS generation and oxidative stress.