Introduction

GTP cyclohydrolase, GCH1 is the rate-limiting enzyme in the de novo biosynthesis of tetrahydrobiopterin (BH4), which is an enzyme cofactor essentially required for the production of monoamine neurotransmitters and nitric oxide (Werner et al., 2011), and the metabolism of ether-lipids via alkylglycerol monooxygenase (AGMO) (Watschinger et al., 2010). The synthesis is a 3-step enzymatic cascade starting with GCH1. The downstream enzymes are PTPS (pyruvoyltetrahydropterin synthase) and SPR (sepiapterin reductase). The expression of GCH1 is increased on demand to meet requirements of BH4, which is particularly high in inflammatory conditions owing to the upregulation of inducible nitric oxide synthase in myeloid derived inflammatory cells (McNeill et al., 2015).
Recently, Cronin et al. demonstrated that GCH1 is also upregulated in activated CD4 positive and CD8+ T-cells (Cronin et al., 2018). By using T-cell-specific GCH1 depletion and overexpression, or SPR inhibition, the authors show that BH4 acts as a regulator of T-cell receptor dependent T-cell proliferation in various models of allergy, autoimmunity and immune mediated cancer surveillance (Cronin et al., 2018). GCH1 deficiency attenuated T-cell proliferation, and overexpression had opposite effects. Intriguingly, nitric oxide (NO) was apparently not involved but BH4 acted as a regulator of iron homeostasis (Cronin et al., 2018), suggested by low iron levels and low conversion of ferri Fe3+ to ferro Fe2+. As a result, cytochrome c activity and ATG generation were reduced. Hence, the proliferation defect may originate from an energy deficit (Cronin et al., 2018).
Mostly, the authors used T-cell transfer models that skip the active immunization. T-cell transfer models do not require antigen recognition, presentation and initiation of antigen-specific T-cell proliferation. The consideration of the model is important in the light of the protective anti-oxidative and anti-inflammatory effects of BH4/sapropterin in models of cardiovascular diseases (Katusic et al., 2009; Li et al., 2011) or colitis (Zschiebsch et al., 2016), and inhibition of tumor growth upon GCH1 inhibition (Pickert et al., 2013). Sapropterin-hydrocholoride (KuvanĀ®) is a clinically available BH4 medication approved for treatment of genetic BH4 deficiency (Howells et al., 1986; Williams et al., 1979), and was also suggested as adjunctive treatment for cardiovascular disease (Jeong et al., 2019; Rodriguez-Miguelez et al., 2018), diabetes (Gangula et al., 2018), depression and schizophrenia (Clelland et al., 2020) and mycobacterial infection (McNeill et al., 2018).
The duality of results suggest that the net outcome of high or low BH4 concentrations in vivo depends on the cellular source and the complex functions in the disease-specific and site-specific (auto)-immune context. Because sapropterin is an approved drug, it is crucial to know if it boosts T-cell responses under certain conditions. In particular, autoimmune diseases of the peripheral and central nervous system such as multiple sclerosis differ from other sites because blood-to-brain and brain-to-CSF barriers normally hinder immune cells from invasion. BH4-dependent endothelial NOS is highly expressed in brain endothelial cells, and pro-oxidative metabolites that are generated via NOS in the absence of BH4, promote a disruption of the BBB (Wu et al., 2009). It is well known that MS pathophysiology has a strong oxidative contribution (Fischer et al., 2012; Licht-Mayer et al., 2015; Linker et al., 2011; Mossakowski et al., 2015). In addition to redox targets, BH4 alters bioactive lipids (Zschiebsch et al., 2016) presumably via AGMO, which are crucial for BBB integrity.
To assess the putative benefit or adversity of the currently available BH4 drug (sapropterin, KuvanĀ®), we used the experimental autoimmune encephalomyelitis (EAE) model, in which the MS like disease is evoked by active immunization against proteins of myelin sheaths. We analyzed the disease course, immune cell proliferation, and invasion and alterations of lipid signaling molecules. The mechanistic focus on bioactive lipids was motivated by the functions of AGMO in inflammation and resolution (Watschinger et al., 2015) and iron dependency of fatty acid metabolism.