Mutations in SCN5A From Patients With IBS Result in Abnormal Na V1.5 Function

Abstract

Background: Central and peripheral serotonergic systems have been implicated in symptom pathogenesis in Irritable Bowel Syndrome (IBS) on the basis of serotonin's involvement in regulating gastrointestinal tract (GIT) homeostasis, in addition to its central role in the regulation of mood; accumulating evidence also points to the presence of low level immune activation in the condition. We previously showed that the activity of indoleamine-2,3dioxygenase (IDO), the immunoresponsive enzyme which is responsible for the degradation of tryptophan along the kynurenine pathway, is enhanced in IBS. The mechanism underpinning this phenomenon is not well understood. Recently, altered function in specific members of the toll-like receptor (TLR) family (TLR1-10) has also been demonstrated in IBS. However, the relationship between TLR activation and IDO activity in IBS is unknown. Aim: To investigate whether specific TLR activation elicits exaggerated kynurenine production in IBS patients compared to controls. Methods: 25 IBS patients and 37 healthy controls (HC) were recruited and venous blood was donated. Whole blood was cultured with a panel of nine different TLR agonists for 24 hours. Cell culture supernatants were then analysed for both trytophan and kynurenine by HPLC in a single isocratic run using sequential fluorescent and UV detection. Results: Kynurenine concentrations were elevated to similar levels in both IBS (240.9 ± 11.58 vs 322.4 ± 19.73ng/ml, p<0.001) and HC samples (224.2 ± 6.330 vs 316.5 ± 13.60ng/ml, p<0.001) following treatment with the TLR4 agonist lipopolysaccheride (LPS). A corresponding elevation in the kynurenine:tryptophan ratio, an indicator of IDO activity, was observed in both IBS patients (0.01774 ± 0.00089 vs 0.02367 ± 0.00137, p<0.001) and HCs (0.01681 ± 0.0006 vs 0.02354 ± 0.001117, p<0.001). In contrast, this ratio was elevated in IBS patients alone (0.01774 ± 0.00089 vs 0.02166 ± 0.00162, p<0.05) following treatment with the TLR8 ligand ssRNA40. Conclusion: These results demonstrate that TLR activation can drive elevations in IDO activity in a manner that corresponds to previously described functional TLR-agonist mediated responses in IBS. Moreover, it implicates TLR8 in the differential IDO activity observed in IBS patients compared to controls. This provides novel evidence for a putative TLR-dependant mechanism underpinning IDO activation in IBS and may point to new therapeutic avenues in this debilitating condition.