Filippo Ingoglia: “Monocyte Derived Macrophages as an Alternative Cell Model for the Functional Diagnosis of Creatine Transporter Deficiency”
ABSTRACT
Measurement of creatine transporter activity is essential for establishing the diagnosis of creatine transporter deficiency (CTD) and to determine whether there is residual activity that might affect therapeutic responses. Primary skin fibroblasts are currently considered the gold standard cell type for functional assays. However, the minimally invasive nature of skin biopsy and the prolonged time required for fibroblast isolation and expansion limit their practical accessibility. To overcome these limitations, we investigated peripheral blood–derived cells as an alternative cellular model for the functional diagnosis of CTD. Peripheral blood cell fractions were isolated from apheresis cones of healthy donors using density gradient centrifugation. Based on isolation simplicity, consistency, and adherence properties, monocytes were identified as the most suitable cell type for further evaluation. Creatine transport and expression of the creatine transporter CT1 (SLC6A8) were assessed in monocytes and monocyte derived macrophages (MDM). Monocytes exhibited minimal creatine uptake (≈29 pmol/mg protein/h), whereas MDM showed a marked increase in saturable creatine uptake (≈517 pmol/mg protein/h), comparable to that observed in primary fibroblasts (≈607 pmol/mg protein/h). Consistent with these findings, SLC6A8 mRNA expression was increased approximately nine fold in MDM compared with monocytes. Kinetic analysis demonstrated that creatine uptake in MDM was linear for up to two hours and exhibited a Km of 44.7 ± 8.2 µM, closely matching fibroblast kinetic properties. In conclusion, monocyte derived macrophages represent a promising, faster, and less invasive cell system as compared to skin fibroblasts for the functional diagnosis of CTD.