Monocyte Cytokine Responses to TLR Ligands in Children with ASD

Background:

Increasing evidence has suggested that dysregulation of the immune system occurring prenatally or in early childhood may contribute to the development of autism spectrum disorders (ASD). We hypothesize that innate immune mechanisms may be abnormal in ASD and lead to inappropriate response to specific bacterial and viral antigens. Furthermore, failure of peripheral monocytes to trigger immune responses appropriately could result in ongoing inflammation that could contribute to peripheral immune dysfunction and microglial inflammation in the brain.

Objectives:

We examined the response of isolated monocytes to specific viral and bacterial ligands in children diagnosed with ASD compared to typically developing control children who were enrolled in the CHARGE study at UC Davis.

Methods:

Peripheral blood from 17 children with ASD (average age 3.9 years) and 16 typically developing children (average age 3.3 years) was isolated and mononuclear cells obtained by density gradient centrifugation. CD14+ monocytes were isolated by magnetic bead separation and stimulated for 24 hours with specific toll-like receptor (TLR) ligands: lipoteichoic acid/LTA (TLR2), poly I:C (TLR3), lipopolysaccharide/LPS (TLR4), flagellin (TLR5), or CpG-B (TLR9). Supernatants were harvested from the cell cultures and the production of IL-1β, IL-6, TNFα, MCP-1, and GM-CSF were determined by Luminex analysis.

Results:

The production of the proinflammatory cytokine IL-1β was increased in monocyte cell cultures from ASD children compared to typically developing controls following stimulation with LTA (p=0.02) or LPS (p=0.04). Further, the proinflammatory cytokines TNFα and IL-6 were also significantly increased in monocyte cell cultures from ASD compared to controls following LTA stimulation. However, stimulation with flagellin and poly I:C did not produce differential production of these cytokines in monocyte cell cultures from ASD children compared to controls. Conversely, CpG stimulation produced decreased IL-β, IL-6, GM-CSF and TNFα in monocyte cell cultures from ASD children compared to controls (p<0.05). Similarly, production of the chemokine MCP-1 was decreased two-fold in monocyte cell cultures from ASD children following stimulation with poly (I:C) (p=0.006) or CpG (p=0.002).

Conclusions:

These results demonstrate a differential cytokine response upon stimulation of monocyte cell cultures with some TLR ligands between children with ASD compared to typically developing controls. Furthermore, these differences are associated with the specific nature of the antigen presented, suggesting that separate signaling pathways within monocytes may be affected. As monocytes are key regulators of the immune response, dysfunction in these cells could result in long-term immune alterations in children with ASD. Furthermore, as monocytes share lineage and functional similarity with microglial cells, these findings may parallel similar responses of microglial cells in the brains of children with ASD. Overall, these data suggest that monocyte cell cultures from children with ASD have changes in signaling via select TLRs when compared to age matched controls that may be indicative of an altered neuroimmune network.