Folate transport and binding by cultured human proximal tubule cells

Abstract

Because mechanisms for the renal regulation of folate excretion are poorly understood, a cell culture system representative of the human proximal tubule (HPT) was used for studies of renal folate transport. After confluent cultures of HPT cells were incubated with 3H-labeled folic acid (PteGlu), binding to the apical membrane was determined by an acid removal procedure, and transport was subsequently measured in solubilized cells. Although PteGlu binding was almost all specific (suppressed by excess unlabeled PteGlu), HPT cells transported PteGlu by specific and nonspecific processes. Specific PteGlu binding and transport were both saturable processes, reaching maxima of 0.5 and 0.1 pmol/mg protein, respectively, with half-maximal constants of 12 and 50 nM, respectively. The PteGlu analogues methotrexate and 5-methyltetrahydrofolic acid (5-CH3-H4PteGlu) inhibited both the binding and transport of PteGlu, with 5-CH3-H4PteGlu being more potent (lower half-maximal inhibitory concentration). In contrast, 5-formyltetrahydrofolic acid significantly reduced PteGlu transport at concentrations (100-250 nM) that had no effect on binding. These data suggest that the HPT cells will serve as a good model for studies of renal folate reabsorption. Initial characterization of the transport of folate by HPT cells suggests two distinct processes, binding to a high-affinity membrane folate-binding protein followed by a structurally specific transfer into the cell.

Renal Tubular Reabsorption of Folate Mediated by Folate Binding Protein 1

“…Injection of 3 H-MTHF also confirms the important role of folbp1 for renal tubular reabsorption, showing increased urinary excretion and decreased renal accumulation in folbp1Ϫ/Ϫ. These findings, establishing for the first time in an in vivo model a direct role for folbp1 in epithelia folate uptake, confirms the hypotheses raised by the identification of FR in the kidney proximal tubule and by observations on folate transport in the kidney and in cultured kidney cells (14,20,32,33). In addition, folbp1 is heavily expressed in other absorptive epithelia, including the rodent yolk sac (34) and choroid plexus (15).…”

Folate Receptor–Targeted Antioxidant Therapy Ameliorates Renal Ischemia–Reperfusion Injury

“…28 The binding of folate to the folate receptor occurs at a relatively high affinity with half maximal binding as high as 12 nM in human proximal tubule cells, making it ideal for pharmacological targeting. 25,26,29 In this study, we synthesized a novel folate-antioxidant conjugate to selectively target the kidney, to enhance localized superoxide scavenging, and to prevent the development of ARF while avoiding the associated side effects of systemic antioxidant therapy. [16][17][18]30,31 We hypothesize that the tempol-folate conjugate selectively targets the renal proximal tubule and protects from ischemic injury by way of scavenging reactive oxygen species, therefore preventing the cascade of events resulting in tubular dysfunction and ARF ( Figure 1A).…”

A New 18F-Labeled Folic Acid Derivative with Improved Properties for the PET Imaging of Folate Receptor–Positive Tumors

“…In general, folate-based radiopharmaceuticals show a high radiotracer accumulation in the kidneys due to a physiologic folate receptor expression in proximal tubule cells (16). Recently, Müller et al reported a favorable effect of the antifolate pemetrexed on kidney and tumor uptake (17,18); thus, the high radioactivity accumulation and the potential risk of damage to the kidneys may no longer be an issue.…”