The well-established increased cardiovascular risk that is a hallmark of chronic kidney disease (CKD) has directed research to metabolic changes that are typical of CKD. Epidemiological data point to derangements of mineral metabolism to be involved in this risk profile. Subsequently, newly discovered humoral factors—such as fibroblast growth factor (FGF)-23—that are involved in mineral and vitamin D homeostasis turned out to be associated with clinical outcome, independently of the minerals they regulate. Additional proteins involved in FGF-23 signaling, such as Klotho, subsequently appeared to have FGF-23-independent effects as well. In this review, the discovery, mode of action, and clinical implications of these new factors are outlined.
Identification of fibroblast growth factor (FGF)-23 has in many ways revolutionized our current understanding of mineral metabolism. It was initially discovered through attempts to identify the predicted existence of ‘phosphatonins’, that is, phosphate-regulating hormones. Summarizing the accomplishments made by many different researchers, FGF-23 was found to be the primary cause of autosomal dominant hypophosphatemic rickets,
Structurally, FGF-23 was the twenty-third member of the FGF family to be discovered with approximately 25–30% homology to other FGFs. The first 24 amino acids of the amino terminus function as a signal peptide for its transport from the Golgi network to the extracellular space, and it is consequently a circulating factor. The carboxy terminus is distinct from other FGFs, providing unique characteristics in terms of glycosylation and receptor activation. Two arginines—located at residues 176 and 179, respectively—provide a consensus site for proteolytic cleavage by furin-like enzymes that inactivate and degrade the active FGF-23 protein
There is striking concordance between the phenotypic changes found in patients with primary or secondary FGF-23 excess and/or deficiency and data from the extensive animal and in vitro studies. Collectively, FGF-23 is a potent negative regulator of circulating phosphate and 1,25-dihydroxyvitamin D (calcitriol; 1,25D) levels.
Another role of FGF-23 in vitamin D metabolism is to enhance the degradation pathway of vitamin D through stimulation of the 24-hydroxylase. In the context of physiology, more recent studies have also convincingly shown that FGF-23, at least in the short term, directly decreases the transcript level and secretion of parathyroid hormone (PTH).
This further underscores the fact that bone, beyond its capacity to store minerals and provide mechanical support, is a highly active endocrine organ. Further, there is robust evidence for the presence of a previously unidentified bone–kidney axis. The interplay between bone and kidney is not farfetched given that the kidney is the main determinant of circulating phosphate levels and actively participates in maintaining calcium homeostasis, providing the skeleton with sufficient minerals to form hydroxyapatite crystals at the mineralization front.
Because FGF-23 holds promise as a biomarker for patient outcome (see below), especially in patients with CKD, it is important to understand its mode of regulation. The most rapid stimuli for FGF-23 expression both in vitro and in vivo is 1,25D, evoking a response in serum FGF-23 level within 3–4 h after intravenous administration.
This completes a feedback loop between vitamin D and FGF-23, and FGF-23 can in that sense be viewed as a counter-regulatory hormone for vitamin D. As a result, the decline in vitamin D level that occurs already in the initial phase of CKD can likely be attributed to a rise in FGF-23 rather than a reduced renal mass per se.
FGF-23 production is also promoted by high dietary phosphate intake,
as well as chronic hyperphosphatemia, although rapid changes in serum phosphate concentrations may not invoke acute increments in FGF-23. One hypothesis is that FGF-23 responds to the net phosphate balance rather than the serum phosphate level, but experimental data supporting this hypothesis is weak. Further, the complete chain of events from high dietary phosphate intake and hyperphosphatemia to increased FGF-23 synthesis in bone is currently unknown.
It also stands clear that vitamin D and phosphate regulate FGF-23 through independent pathways, because mice lacking the vitamin D receptor are still highly responsive to high dietary phosphate intake.
As a final remark, the response in FGF-23 elicited by dietary phosphate intake in humans is much weaker than that in rodents.
Despite the fact that FGF-23 belongs to the FGF family, in which all members signal through one or several of the known FGF receptors, it has been difficult to unveil the ‘true’ FGF-23 receptor both in vivo and in vitro. A major breakthrough came from studies by Urakawa et al.,
who demonstrated that type I membrane-bound α-Klotho (Klotho) directly binds to FGF receptor 1c, converting it into a specific FGF-23 receptor (Figure 2).
Accordingly, FGF-23 is dependent on Klotho to induce FGF-receptor signaling, at least in the kidneys and parathyroid glands. The importance of Klotho in FGF-23 signaling is evidenced by Klotho-null mice, which harbor nearly an identical biochemical phenotype compared with FGF-23-knockout mice, despite exceptionally high circulatory FGF-23 levels.
There are, however, still controversies and unresolved issues around FGF-23 receptor signaling. First, Klotho expression in the kidney is largely confined to the distal tubules, whereas renal phosphate reabsorption occurs in the proximal tubules. It is currently unclear how FGF-23 signaling in distal tubules modifies phosphate reabsorption in proximal tubules. Second, it is possible that high levels of FGF-23, as present in many patients with advanced CKD, could induce unspecific ‘off-target’ (that is, Klotho-independent) FGF-receptor signaling.
FGF-23 and Klotho are likely to have important roles in the pathophysiology of secondary hyperparathyroidism. Although FGF-23 in the short term suppresses PTH secretion, chronically high exposure of FGF-23 may override this effect by lowering the systemic levels of 1,25D and attenuate parathyroid vitamin D receptor signaling. Equally important, it was recently demonstrated that FGF-23 reduces the expression level of Klotho and that parathyroid Klotho expression in surgically removed human parathyroid adenomas declines in parallel with loss of renal function.
In summary, the discovery of FGF-23 and Klotho has led to significant advances in our understanding of mineral metabolism. This knowledge is now gradually being translated into the clinic with many potential implications, including the endorsement of FGF-23 as a predictive biomarker and the possibility of FGF-23/Klotho as a novel therapeutic target.
Klotho was discovered in 1997 in a mouse strain with a phenotype consistent with premature aging as the principle hallmark.
as shown in Figure 2. Klotho exists as a membrane-bound form and two circulating forms—one being the shed product of the membrane form and the other a truncated form derived from the same gene by alternative splicing.
Besides its above-described role as a cofactor in FGF-23 signaling, as shown in Figure 2, Klotho harbors at least two important additional functions.
The latter is of importance because Klotho retains this enzymatic activity even when it is shed from the plasma membrane. This is in contrast to the role of Klotho as a cofactor in FGF-23, where the membrane-bound form appears to be necessary.
The glucuronidase-like activity of Klotho modulates the sugar moieties, leading to sustained retention of these highly important calcium transporters in the kidney. In this way, Klotho promotes reabsorption of ultrafiltrated calcium and prevents calciuria.
This calcium-retaining effect of Klotho in the healthy kidney is yet another mechanism used to prevent overactivation of native vitamin D, besides its suppressive role on vitamin D metabolism mediated by FGF-23 signaling. Active vitamin D in turn upregulates Klotho expression, closing a feedback loop between vitamin D and Klotho.
Remarkable about the latter finding is the fact that this effect is located on the proximal tubule, although most investigators found the distal tubule to be the most prominent site of Klotho expression. The other remarkable fact is that the Klotho-induced phosphaturic effects were also observed in FGF-23-knockout mice, indicating a direct effect on NaPt2a.
A third recognized mode of action of Klotho is its protective effect against oxidative stress. This was demonstrated by Kuro-o et al.
Given its central role in regulating mineral metabolism, the obvious question arises: How important is FGF-23 clinically? In early CKD, FGF-23 appears to be beneficial, compensating for reduced phosphate excretory capacity by increasing fractional excretion of phosphate. Although phosphate retention is also a stimulus for PTH secretion, in early-stage CKD the rise in FGF-23 is more pronounced than that of PTH, possibly because of the inhibitory effects of FGF-23 on PTH.
A trade-off of the increase of FGF-23 may be a reduction in levels of 1,25D by the mechanism described above. As CKD progresses, the efficacy of FGF-23 to induce phosphaturia declines, due to at least two mechanisms. First, the loss of functioning nephrons reduces the amount of phosphate being ultrafiltrated; second, lowered renal Klotho expression dismantles the FGF-23 receptor, leading to higher phosphate reabsorption per nephron. For these reasons it is expected that in advanced CKD, FGF-23 could be an indicator for dismal outcome. Indeed, this has been shown for more advanced CKD, where FGF-23 independently predicted progression of disease.
The most convincing argument for the clinical meaning of FGF-23 comes from the analysis of a large cohort of dialysis patients, in which an independent association between FGF-23 and mortality was demonstrated, as shown in Figure 3.
Even after correcting for established predictors of mortality, the hazard ratios for the higher ranges of FGF-23 outranked the others.
The fact that correcting for phosphate level, PTH, and vitamin D use did not mitigate predictive value of FGF-23 for dismal outcome is remarkable, because these parameters were thought to be in the same causal pathway as FGF-23. This could mean that FGF-23 is a sensitive marker for phosphate burden, or induces harm itself. In support for the first hypothesis, FGF-23 has, in large observational studies of elderly subjects with normal or only mildly impaired renal function, been associated with vascular dysfunction,
If FGF-23 indeed turns out to be either a sensitive biomarker of phosphate load, or has some different pathological effect in advanced stages of CKD, the next clinical question would be: Is it modifiable? From a theoretical point of view, options to lower FGF-23 would be to reduce levels of active vitamin D, PTH, and phosphate burden. Indeed, parathyroidectomy has been shown to induce a significant decline in FGF-23,
but this approach is not likely to be sufficient in CKD. The use of phosphate binders has shown inconsistent results as FGF-23-lowering agents. Sevelamer declined FGF-23 in a dose-dependent manner in an experimental model of uremia,
This absence of effect of lanthanum carbonate on FGF-23 in this study, however, could have been caused by a too short study period of two weeks only, since more time may be required for FGF-23 to slow down. Indeed, in a very recent study, extended use of lanthanum carbonate in an otherwise comparable study population did significantly suppress FGF-23 by almost 25%.
Currently, there is no evidence supporting the fact that FGF-23 is a modifiable risk factor that leads to improvement of clinical outcomes such as mortality. Prospective trials are underway that study changes in vascular function after FGF-23-targeted interventions. A critical question is whether FGF-23 is purely a biomarker of phosphate exposure or if it has ‘off-target’ effects directly contributing to vascular toxicity. If FGF-23 levels are reduced by means of improving phosphate control, it will be difficult to analyze whether changes in outcomes are related to a more stringent phosphate control or a consequence of a lower FGF-23 level per se. Another option that could shed light on this question would be to intervene at the level of FGF-23-producing cells, either osteocytes in bone or ectopic production from bone cells in the vessel wall in the presence of calcified arterial lesions,
an enigma emerges: How is it possible that a hormone such as FGF-23 has such an impact on outcome, while its main target organ—the kidney—is non-functioning? Of course, the parathyroid is another target for FGF-23; however, in CKD, PTH suppression by FGF-23 is abolished by parathyroid resistance.
theoretically, other tissues expressing Klotho and FGF receptor 1c might be unidentified targets for FGF-23. Recently, both Klotho and FGF receptor-1 were shown to be present in human aortic smooth muscle cell,
but at present it is unclear whether this is membrane-bound or soluble Klotho, and whether actual signal transduction by FGF-23 can occur at this site. Nevertheless, the arterial wall and cardiomyocytes as target tissues for FGF-23 action is an attractive concept, given the central role of cardiovascular disease in CKD-associated morbidity. Alternatively, indirect effects of FGF-23 on the arterial wall are also plausible. These could be mediated by phosphate itself or by a reduction in Klotho level, given that FGF-23 suppresses Klotho,
In conclusion, data pointing to FGF-23 and Klotho being active factors in the burden of CKD are ever increasing. Both epidemiological data and pathophysiological mechanisms have set the stage for targeted intervention in the clinical setting. These new apparently important factors in CKD provide hope for improved future targeted therapy in CKD.
Autosomal dominant hypophosphataemic rickets is associated with mutations in FGF-23.
This issue was organized by the VDR Expert Centers and was funded by Abbott Laboratories with a collaborative effort to advance and support the science and the improvement of quality of life for renal patients. MGV has received consulting fees from or participated in paid advisory boards for Amgen, Abbott Laboratories, and Fresenius; received lecture fees for speaking at the invitation of Abbott Laboratories and Amgen; and received grant support from Genzyme and Shire. TEL has received consulting fees from or participated in paid advisory boards for Genzyme, Abbott Laboratories, and Swedish-Orphan, and has received lecture fees for speaking at the invitation of Amgen, AstraZeneca, Shire, Genzyme, and Abbott Laboratories.
TO CITE THIS ARTICLE: Vervloet MG, Larsson TE. Fibroblast growth factor-23 and Klotho in chronic kidney disease. Kidney inter., Suppl. 2011; 1: 130–135.