Part Two Fibroblast Growth Factor 23 And Osteoporosis: Evidence From Bench To Bedside

Role of FGF23 in CKD-MBD Pathogenesis

Human mineral homeostasis largely depends on the kidney. Hyperphosphatemia, hypocalcemia, and reduced calcitriol are caused by the loss of renal functions in CKD and ESRD, which leads to secondary hyperparathyroidism and CKD-MBD [32]. The emergence of FGF23 has reformed the understanding of the mechanisms underlying the development of secondary hyperparathyroidism [33]. The loss of FGF23's ability to regulate phosphate levels through its phosphaturic effect and inhibit PTH secretion was shown in ESRD patients, resulting in hyperphosphatemia and increasing FGF23 levels. Although FGF23 can directly block PTH synthesis and secretion from the parathyroid gland primary cells via both the Klotho/FGFRs pathway [7] and the Klotho-independent pathway [16], it can also indirectly induce hyperparathyroidism. In CKD and ESRD patients, increased FGF23 induced by hyperphosphatemia inhibits vitamin D activation and subsequently decreases calcium absorption in the intestine [34,35]. As a result, low calcitriol levels and hypocalcemia promote PTH synthesis and release via VDR and CaSR in parathyroid, overriding the inhibitory effect of FGF23 [18]. The progressive kidney dysfunction will subsequently cause hyporesponsiveness of VDR [36] on the parathyroid gland with more excessive PTH synthesis and reduced expression of CaSR [37] on the parathyroid gland leading to parathyroid gland hyperplasia and will become autonomous [38]. This excess PTH leads to further calcium resorption from the bone, resulting in abnormalities of bone architecture. Consequently, the auto-/paracrine effects of FGF23 on bone cells, mediated by high levels of circulating FGF23, is another possible pathogenesis of CKD-MBD, in addition to secondary hyperparathyroidism. The recent preclinical study in the CKD model indicates that excessive FGF23 secretion driven by renal failure significantly inhibits bone mineralization via TNAP suppression and PPi accumulation [39]. FGF23 neutralization’s effects on bone quality had been explored in vivo for its therapeutic potential in CKD-MBD [40,41]. Anti-FGF23 treatment was found to significantly improve the bone quality in CKD mice by correcting the secondary hyperparathyroidism, and increased calcitriol levels, indicating that FGF23 is a key factor of CKD-related bone diseases. Nonetheless, other studies in CKD animal models indicated that FGF23 neutralization exacerbates hyperphosphatemia and elevated serum calcitriol, resulting in increased arterial calcification. Thus, the systemic mineral disturbances caused by FGF23 neutralization limit its benefit on bone quality in CKD-MBD, which certainly contributes to the increased risk of cardiovascular events and death.

Click here to know what is Cistanche

Potential Clinical Application of FGF23 in Osteoporosis and CKD-MBD

Although current clinical studies are insufficient to support those findings and the hypothesis that FGF23 is independently associated with bone mineralization decline in the elderly and CKD-MBD, several clinical studies show that FGF23 has been a potential predictor for fragility fractures. High circulating FGF23 levels were discovered to be an independent risk factor for overall fragility fracture in elderly men [29]. Furthermore, the three aforementioned studies discovered an independent association between elevated FGF23 and the incidence of fragility fractures in both moderate CKD and ESRD patients [28,34,35]. Even though FGF23 levels had an independent negative relationship with BMD in postmenopausal women [1,4], it was not an effective discriminator between osteopenia/osteoporosis and normal bone mass [4]. In addition, utilizing the FGF23 level for osteoporosis prediction in hemodialysis patients resulted in poor discrimination [44]. Considering that various fragility fracture prediction models (e.g., the FRAX score, Q Fracture) are based on well-established clinical predictors (e.g., gender, BMI, CKD, alcohol, smoking, and corticosteroid use), there are currently no models that incorporate clinical predictors describing both systemic and local bone mineral homeostasis. The growing evidence indicates that FGF23 can represent the status of systemic bone mineral balance, renal function, and probably bone remodeling, thus the value-added of FGF23 on fragility fracture prediction awaits upcoming studies to explore its clinical potential to improve the prognostication of osteoporosis and CKD-MBD patients.

Cistanche extract

FGF23 Measurement in Routine Clinical Practices

Since none of the commercially available FGF23 assays have been validated for clinical use, FGF23 is not presently applicable for routine clinical practices. For the measurement of FGF23, four immunoassays are commercially available: Immutopics (1st and 2nd generation, San Clemente, CA, USA), Kainos (Tokyo, Japan), Millipore (Billerica, MA, USA), and DiaSorin (Saluggia, Italy). The majority of assays detect the intact 251 amino acid protein (iFGF23) by simultaneously recognizing epitopes on the N- and C-terminal domains located near the proteolytic cleavage site. Additionally, Immutopics provides an assay that quantifies both iFGF23 and the C-terminal fragment (cFGF23) using two antibodies against two C-terminal epitopes. iFGF23 is measured in picograms per milliliter (pg./mL), with a normal reference range of 11.7–48.6 pg./mL in a healthy individual, whereas cFGF23 is reported in relative units (RU) per milliliter, with a normal reference range of 21.6–91.0 RU/mL [47]. Due to the possibility that iFGF23 may be degraded by protease enzyme or changed after venipuncture, two iFGF23 stability studies discovered decreasing FGF23 levels following an 8-h delay in centrifugation, but no evidence of deterioration after storing processed samples at −80 ◦C [48]. Biological variability studies in healthy individuals revealed that iFGF23 levels have a diurnal variation that peaks in the early morning and gradually declines during the day [47]. In comparison, the concentrations of cFGF23 could be slightly increased throughout the day [49] and see no significant change after dietary or phosphate intake [50]. Despite the stability and biological variability advantages of cFGF23, cFGF23 assays may be more applicable than iFGF23 assays, particularly for diagnostic and prognostic studies. In contrast, iFGF23 may outperform in representing the biological effects of FGF23 in diagnostic and therapeutic research because the c-terminal fragments might have counter-regulatory effects on the physiologically active FGF23 [51]. The schematic summary of FGF23 production and its measurement are illustrated in Figure 4.

Figure 4. FGF23 production and immunoassay measurements. (a) After completed transcription and translation, FGF23 can be transferred to two post-translation modification pathways, includingO-glycosylation with GALN13 on Thr178, or phosphorylation by the extracellular serine /threonine protein kinase FAM20C at Ser180. O-glycosylation modification by GALANT3, stabilized formcan prevents intact FGF23 from cleavage. In contrast, phosphorylated FGF23 by FAM20C can be cleaved into N-terminal and C-terminal fragments within the osteocyte/osteoblast. These peptides including full-length (intact) FGF23, N-terminal fragments, and C-terminal fragments, can be detected in the circulation. (b) For C-terminal assays, detecting antibodies bind to C-terminus epitopes to detect both fulllength FGF23 and its C-terminal fragments, whereas assays for intact FGF23 use antibodies to detect epitopes surrounding the FGF23 cleavage site for the detection of only full-lengthFGF23. This figure was generated with publication licensed by BioRender, Toronto, ON, Canada(Agreement number: DV237ONHF, 19 November 2021). Abbreviations: GALNT3, polypeptideN-acetyl galactosaminyltransferase 3; FAM20C, the extracellular protein kinase FAM20C; Ser, Serine;Thr, Threonine.

Cistanche capsules

Conclusions

A pivotal role of FGF23 was found in local and systemic bone remodeling with supraphysiological levels causing abnormal bone formation, although any direct effect on osteoblasts remains unclear as well as a controversial link between FGF23 with osteoclastogenesis and bone resorption. Current evidence from clinical studies indicates that FGF23 could be a risk factor for bone fragility in CKD-MBD, but not a major contributor to age-related osteoporosis. An increased FGF23 level may represent an abnormal state of bone mineral homeostasis but is not a direct indicator of decreased BMD. Since clinical studies, both in healthy elderly and in patients with impaired renal function, showed that elevated FGF23 levels were an independent risk factor of fragility fracture, a future predictive model for fragility fracture may incorporate FGF23 as a factor to represent bone mineral homeostasis status. FGF23 is a putative factor in the fragility of CKD-MBD, less so in age-related bone loss; future elucidation of pathogenesis requires remodeling biomarkers.

Cistanche supplement

The effects of Cistanche on the kidneys

Cistanche is an herbal medicine that has been widely used in traditional Chinese medicine for centuries. It is derived from the Cistanche plant, which is known for its potent medicinal properties. Among its many purported benefits, Cistanche is purported to have positive effects on kidney health.

One of the primary effects of Cistanche on the kidneys is its ability to promote renal regeneration. Studies have shown that Cistanche extracts contain active compounds that can stimulate the growth and proliferation of renal cells. This can be particularly beneficial for individuals suffering from kidney damage or disease, as it may help to restore the normal function of the kidneys.

Additionally, Cistanche has been found to possess diuretic properties, meaning that it can increase urine production and promote the elimination of toxins from the body. This diuretic effect helps to enhance the overall renal function and can be beneficial for individuals experiencing urinary tract infections or other kidney-related issues.

Furthermore, Cistanche is believed to possess anti-inflammatory properties, which can be particularly beneficial for individuals with kidney inflammation or infections. By reducing inflammation, Cistanche helps to alleviate pain and swelling in the kidneys, while also promoting the healing process.

However, it is important to note that while Cistanche may have positive effects on kidney health, it should not be used as a sole treatment for kidney diseases or other serious conditions. It is always advisable to consult with a healthcare professional before introducing any new herbal medicine into your routine, especially if you have pre-existing kidney issues or are taking other medications.

In conclusion, Cistanche has shown promising effects on kidney health, including renal regeneration, diuretic properties, and anti-inflammatory effects. However, further research is still needed to fully understand its mechanisms of action and establish its efficacy and safety.

Cistanche powder

References

1. Shen, J.; Fu, S.; Song, Y. Relationship of Fibroblast Growth Factor 23 (FGF-23) Serum Levels With Low Bone Mass in Postmenopausal Women. J. Cell. Biochem. 2017, 118, 4454–4459. [CrossRef] [PubMed]

2. Marsell, R.; Mirza, M.A.; Mallmin, H.; Karlsson, M.; Mellström, D.; Orwoll, E.; Ohlsson, C.; Jonsson, K.B.; Ljunggren, O.; Larsson, T.E. Relation between fibroblast growth factor-23, body weight and bone mineral density in elderly men. Osteoporos. Int. 2009, 20, 1167–1173. [CrossRef] [PubMed]

3. Rupp, T.; Butscheidt, S.; Vettorazzi, E.; Oheim, R.; Barvencik, F.; Amling, M.; Rolvien, T. High FGF23 levels are associated with impaired trabecular bone microarchitecture in patients with osteoporosis. Osteoporos. Int. 2019, 30, 1655–1662. [CrossRef]

4. Bilha, S.C.; Bilha, A.; Ungureanu, M.C.; Matei, A.; Florescu, A.; Preda, C.; Covic, A.; Branisteanu, D. FGF23 Beyond the Kidney: A New Bone Mass Regulator in the General Population. Horm. Metab. Res. 2020, 52, 298–304. [CrossRef] [PubMed]

5. Shimada, T.; Kakitani, M.; Yamazaki, Y.; Hasegawa, H.; Takeuchi, Y.; Fujita, T.; Fukumoto, S.; Tomizuka, K.; Yamashita, T. Targeted ablation of Fgf23 demonstrates an essential physiological role of FGF23 in phosphate and vitamin D metabolism. J. Clin. Investig. 2004, 113, 561–568. [CrossRef] [PubMed]

6. Takeshita, A.; Kawakami, K.; Furushima, K.; Miyajima, M.; Sakaguchi, K. Central role of the proximal tubular αKlotho/FGF receptor complex in FGF23-regulated phosphate and vitamin D metabolism. Sci. Rep. 2018, 8, 6917. [CrossRef]

7. Ben-Dov, I.Z.; Galitzer, H.; Lavi-Moshayoff, V.; Goetz, R.; Kuro-o, M.; Mohammadi, M.; Sirkis, R.; Naveh-Many, T.; Silver, J. The parathyroid is a target organ for FGF23 in rats. J. Clin. Investig. 2007, 117, 4003–4008. [CrossRef]

8. Kawaguchi, H.; Manabe, N.; Miyaura, C.; Chikuda, H.; Nakamura, K.; Kuro-o, M. Independent impairment of osteoblast and osteoclast differentiation in klotho mouse exhibiting low-turnover osteopenia. J. Clin. Investig. 1999, 104, 229–237. [CrossRef]

9. Masuyama, R.; Stockmans, I.; Torrekens, S.; Van Looveren, R.; Maes, C.; Carmeliet, P.; Bouillon, R.; Carmeliet, G. Vitamin D receptor in chondrocytes promotes osteoclastogenesis and regulates FGF23 production in osteoblasts. J. Clin. Investig. 2006, 116, 3150–3159. [CrossRef]

10. Yoshiko, Y.; Wang, H.; Minamizaki, T.; Ijuin, C.; Yamamoto, R.; Suemune, S.; Kozai, K.; Tanne, K.; Aubin, J.E.; Maeda, N. Mineralized tissue cells are a principal source of FGF23. Bone 2007, 40, 1565–1573. [CrossRef]

11. Mirams, M.; Robinson, B.G.; Mason, R.S.; Nelson, A.E. Bone as a source of FGF23: Regulation by phosphate? Bone 2004, 35, 1192–1199. [CrossRef] [PubMed]

12. Li, Y.; He, X.; Olauson, H.; Larsson, T.E.; Lindgren, U. FGF23 affects the lineage fate determination of mesenchymal stem cells. Calcif. Tissue Int. 2013, 93, 556–564. [CrossRef] [PubMed]

13. Bon, N.; Frangi, G.; Sourice, S.; Guicheux, J.; Beck-Cormier, S.; Beck, L. Phosphate-dependent FGF23 secretion is modulated by PiT2/Slc20a2. Mol. Metab. 2018, 11, 197–204. [CrossRef]

14. Takashi, Y.; Kosako, H.; Sawatsubashi, S.; Kinoshita, Y.; Ito, N.; Tsoumpra, M.K.; Nangaku, M.; Abe, M.; Matsuhisa, M.; Kato, S.; et al. Activation of unliganded FGF receptors by extracellular phosphate potentiates proteolytic protection of FGF23 by its O-glycosylation. Proc. Natl. Acad. Sci. USA 2019, 116, 11418–11427. [CrossRef] [PubMed]

15. Krajisnik, T.; Björklund, P.; Marsell, R.; Ljunggren, O.; Akerström, G.; Jonsson, K.B.; Westin, G.; Larsson, T.E. Fibroblast growth factor-23 regulates parathyroid hormone and 1alpha-hydroxylase expression in cultured bovine parathyroid cells. J. Endocrinol. 2007, 195, 125–131. [CrossRef] [PubMed]

16. Olauson, H.; Lindberg, K.; Amin, R.; Sato, T.; Jia, T.; Goetz, R.; Mohammadi, M.; Andersson, G.; Lanske, B.; Larsson, T.E. Parathyroid-Specific Deletion of Klotho Unravels a Novel Calcineurin-Dependent FGF23 Signaling Pathway That Regulates PTH Secretion. PLoS Genet. 2013, 9, e1003975. [CrossRef] [PubMed]

17. Kantham, L.; Quinn, S.J.; Egbuna, O.I.; Baxi, K.; Butters, R.; Pang, J.L.; Pollak, M.R.; Goltzman, D.; Brown, E.M. The calcium-sensing receptor (CaSR) defends against hypercalcemia independently of its regulation of parathyroid hormone secretion. Am. J. Physiol. Endocrinol. Metab. 2009, 297, E915–E923. [CrossRef]

18. Ferrè, S.; Hoenderop, J.G.J.; Bindels, R.J.M. Sensing mechanisms involved in Ca2+ and Mg2+ homeostasis. Kidney Int. 2012, 82, 1157–1166. [CrossRef]

19. Murali, S.K.; Roschger, P.; Zeitz, U.; Klaushofer, K.; Andrukhova, O.; Erben, R.G. FGF23 Regulates Bone Mineralization in a 1,25(OH)2D3 and Klotho-Independent Manner. J. Bone Mineral. Res. 2016, 31, 129–142. [CrossRef]

20. Mackenzie, N.C.W.; Zhu, D.; Milne, E.M.; can’t Hof, R.; Martin, A.; Quarles, D.L.; Millán, J.L.; Farquharson, C.; MacRae, V.E. Altered Bone Development and an Increase in FGF-23 Expression in Enpp1−/− Mice. PLoS ONE 2012, 7, e32177. [CrossRef]

21. Ho, A.M.; Johnson, M.D.; Kingsley, D.M. Role of the mouse ank gene in control of tissue calcification and arthritis. Science 2000, 289, 265–270. [CrossRef] [PubMed]

22. Russell, R.G.; Bisaz, S.; Donath, A.; Morgan, D.B.; Fleisch, H. Inorganic pyrophosphate in plasma in normal persons and patients with hypophosphatasia, osteogenesis imperfecta, and other disorders of bone. J. Clin. Investig. 1971, 50, 961–969. [CrossRef] [PubMed]

23. Lieben, L.; Masuyama, R.; Torrekens, S.; Van Looveren, R.; Schrooten, J.; Baatsen, P.; Lafage-Proust, M.H.; Dresselaers, T.; Feng, J.Q.; Bonewald, L.F.; et al. Normocalcemia is maintained in mice under conditions of calcium malabsorption by vitamin D-induced inhibition of bone mineralization. J. Clin. Investig. 2012, 122, 1803–1815. [CrossRef] [PubMed]

24. Yuan, Q.; Jiang, Y.; Zhao, X.; Sato, T.; Densmore, M.; Schüler, C.; Erben, R.G.; McKee, M.D.; Lanske, B. Increased osteopontin contributes to inhibition of bone mineralization in FGF23-deficient mice. J. Bone Miner. Res. Off. J. Am. Soc. Bone Mineral. Res. 2014, 29, 693–704. [CrossRef] [PubMed]

25. Murshed, M.; Harmey, D.; Millán, J.L.; McKee, M.D.; Karsenty, G. Unique coexpression in osteoblasts of broadly expressed genes accounts for the spatial restriction of ECM mineralization to the bone. Genes Dev. 2005, 19, 1093–1104. [CrossRef] [PubMed]

26. Addison, W.N.; Azari, F.; Sørensen, E.S.; Kaartinen, M.T.; McKee, M.D. Pyrophosphate inhibits the mineralization of osteoblast cultures by binding to minerals, up-regulating osteopontin, and inhibiting alkaline phosphatase activity. J. Biol. Chem. 2007, 282, 15872–15883. [CrossRef]

27. Allard, L.; Demoncheaux, N.; Machuca-Gayet, I.; Georgess, D.; Coury-Lucas, F.; Jurdic, P.; Bacchetta, J. Biphasic Effects of Vitamin D and FGF23 on Human Osteoclast Biology. Calcif. Tissue Int. 2015, 97, 69–79. [CrossRef]

28. Lane, N.E.; Parimi, N.; Corr, M.; Yao, W.; Cauley, J.A.; Nielson, C.M.; Ix, J.H.; Kado, D.; Orwoll, E. Association of serum fibroblast growth factor 23 (FGF23) and incident fractures in older men: The Osteoporotic Fractures in Men (MrOS) study. J. Bone Miner. Res. 2013, 28, 2325–2332. [CrossRef]

29. Mirza, M.A.; Karlsson, M.K.; Mellström, D.; Orwoll, E.; Ohlsson, C.; Ljunggren, O.; Larsson, T.E. Serum fibroblast growth factor-23 (FGF-23) and fracture risk in elderly men. J. Bone Miner. Res. 2011, 26, 857–864. [CrossRef]

30. Lewerin, C.; Ljunggren, Ö.; Nilsson-Ehle, H.; Karlsson, M.K.; Herlitz, H.; Lorentzon, M.; Ohlsson, C.; Mellström, D. Low serum iron is associated with high serum intact FGF23 in elderly men: The Swedish MrOS study. Bone 2017, 98, 1–8. [CrossRef]

31. Yamamoto, S.; Koyama, D.; Igarashi, R.; Maki, T.; Mizuno, H.; Furukawa, Y.; Kuro, O.M. Serum Endocrine Fibroblast Growth Factors as Potential Biomarkers for Chronic Kidney Disease and Various Metabolic Dysfunctions in Aged Patients. Intern. Med. 2020, 59, 345–355. [CrossRef]

32. Moe, S.M.; Drüeke, T.; Lameire, N.; Eknoyan, G. Chronic kidney disease-mineral-bone disorder: A new paradigm. Adv. Chronic Kidney Dis. 2007, 14, 3–12. [CrossRef] [PubMed]

33. Gutiérrez, O.M. Fibroblast Growth Factor 23 and Disordered Vitamin D Metabolism in Chronic Kidney Disease: Updating the “Trade-off” Hypothesis. Clin. J. Am. Soc. Nephrol. 2010, 5, 1710–1716. [CrossRef] [PubMed]

34. Kanda, E.; Yoshida, M.; Sasaki, S. Applicability of fibroblast growth factor 23 for evaluation of the risk of vertebral fracture and chronic kidney disease-mineral bone disease in elderly chronic kidney disease patients. BMC Nephrol. 2012, 13, 122. [CrossRef] [PubMed]

35. Desbiens, L.C.; Sidibé, A.; Ung, R.V.; Fortier, C.; Munger, M.; Wang, Y.P.; Bisson, S.K.; Marquis, K.; Agharazii, M.; Mac-Way, F. FGF23-klotho axis, bone fractures, and arterial stiffness in dialysis: A case-control study. Osteoporos. Int. 2018, 29, 2345–2353. [CrossRef] [PubMed]

36. Fukuda, N.; Tanaka, H.; Tominaga, Y.; Fukagawa, M.; Kurokawa, K.; Seino, Y. Decreased 1,25-dihydroxy vitamin D3 receptor density is associated with a more severe form of parathyroid hyperplasia in chronic uremic patients. J. Clin. Investig. 1993, 92, 1436–1443. [CrossRef] [PubMed]

37. Yano, S.; Sugimoto, T.; Tsukamoto, T.; Chihara, K.; Kobayashi, A.; Kitazawa, S.; Maeda, S.; Kitazawa, R. Association of decreased calcium-sensing receptor expression with proliferation of parathyroid cells in secondary hyperparathyroidism. Kidney Int. 2000, 58, 1980–1986. [CrossRef]

38. Tokumoto, M.; Tsuruya, K.; Fukuda, K.; Kanai, H.; Kuroki, S.; Hirakata, H. Reduced p21, p27, and vitamin D receptor in the nodular hyperplasia in patients with advanced secondary hyperparathyroidism. Kidney Int. 2002, 62, 1196–1207. [CrossRef]

39. Andrukhova, O.; Schüler, C.; Bergow, C.; Petric, A.; Erben, R.G. Augmented Fibroblast Growth Factor-23 Secretion in Bone Locally Contributes to Impaired Bone Mineralization in Chronic Kidney Disease in Mice. Front. Endocrinol. 2018, 9, 311. [CrossRef]

40. Sun, N.; Guo, Y.; Liu, W.; Densmore, M.; Shalhoub, V.; Erben, R.G.; Ye, L.; Lanske, B.; Yuan, Q. FGF23 neutralization improves bone quality and osseointegration of titanium implants in chronic kidney disease mice. Sci. Rep. 2015, 5, 8304. [CrossRef]

41. Shalhoub, V.; Shatzen, E.M.; Ward, S.C.; Davis, J.; Stevens, J.; Bi, V.; Renshaw, L.; Hawkins, N.; Wang, W.; Chen, C.; et al. FGF23 neutralization improves chronic kidney disease-associated hyperparathyroidism yet increases mortality. J. Clin. Investig. 2012, 122, 2543–2553. [CrossRef] [PubMed]

42. Coskun, Y.; Paydas, S.; Balal, M.; Soyupak, S.; Kara, E. Bone Disease and Serum Fibroblast Growth Factor-23 Levels in Renal Transplant Recipients. Transplant. Proc. 2016, 48, 2040–2045. [CrossRef] [PubMed]

43. Yavropoulou, M.P.; Vaios, V.; Pikilidou, M.; Chryssogonidis, I.; Sachinidou, M.; Tournis, S.; Makris, K.; Kotsa, K.; Daniilidis, M.; Haritanti, A.; et al. Bone Quality Assessment as Measured by Trabecular Bone Score in Patients with End-Stage Renal Disease on Dialysis. J. Clin. Densitom. 2017, 20, 490–497. [CrossRef] [PubMed]

44. Wu, Q.; Xiao, D.M.; Fan, W.F.; Ye, X.W.; Niu, J.Y.; Gu, Y. Effect of serum fibroblast growth factor-23, matrix Gla protein and Fetuin-A in predicting osteoporosis in maintenance hemodialysis patients. Ther. Apher Dial 2014, 18, 427–433. [CrossRef] [PubMed]

45. Slouma, M.; Sahli, H.; Bahlous, A.; Laadhar, L.; Smaoui, W.; Rekik, S.; Gharsallah, I.; Sallami, M.; Moussa, F.B.; Elleuch, M.; et al. Mineral bone disorder and osteoporosis in hemodialysis patients. Adv. Rheumatol. 2020, 60, 15. [CrossRef]

46. Desjardins, L.; Liabeuf, S.; Renard, C.; Lenglet, A.; Lemke, H.D.; Choukroun, G.; Drueke, T.B.; Massy, Z.A. FGF23 is independently associated with vascular calcification but not bone mineral density in patients at various CKD stages. Osteoporos. Int. 2012, 23, 2017–2025. [CrossRef]

47. Smith, E.R.; Cai, M.M.; McMahon, L.P.; Holt, S.G. Biological Variability of Plasma Intact and C-Terminal FGF23 Measurements. J. Clin. Endocrinol. Metab. 2012, 97, 3357–3365. [CrossRef]

48. Dirks, N.F.; Smith, E.R.; van Schoor, N.M.; Vervloet, M.G.; Ackermans, M.T.; de Jonge, R.; Heijboer, A.C. Pre-analytical stability of FGF23 with the contemporary immunoassays. Clin. Chim. Acta 2019, 493, 104–106. [CrossRef]

49. Vervloet, M.G.; van Ittersum, F.J.; Buttler, R.M.; Heijboer, A.C.; Blankenstein, M.A.; ter Wee, P.M. Effects of Dietary Phosphate and Calcium Intake on Fibroblast Growth Factor-23. Clin. J. Am. Soc. Nephrol. 2011, 6, 383–389. [CrossRef]

50. Burnett, S.M.; Gunawardene, S.C.; Bringhurst, F.R.; Jüppner, H.; Lee, H.; Finkelstein, J.S. Regulation of C-terminal and intact FGF-23 by dietary phosphate in men and women. J. Bone Min. Res. 2006, 21, 1187–1196. [CrossRef]

51. Goetz, R.; Nakada, Y.; Hu, M.C.; Kurosu, H.; Wang, L.; Nakatani, T.; Shi, M.; Eliseenkova, A.V.; Razzaque, M.S.; Moe, O.W.; et al. The isolated C-terminal tail of FGF23 alleviates hypophosphatemia by inhibiting FGF23-FGFR-Klotho complex formation. Proc. Natl. Acad. Sci. USA 2010, 107, 407–412. [CrossRef] [PubMed]

Wachiranun Sirikul 1 , Natthaphat Siri-Angkul 2,3,4, Nipon Chattipakorn 2,3,4 and Siriporn C. Chattipakorn 2,4,5,

1 Department of Community Medicine, Faculty of Medicine, Chiang Mai University, Chiang Mai 50200, Thailand; wachiranun.sir@cmu.ac.th

2 Neurophysiology Unit, Cardiac Electrophysiology Research and Training Center, Faculty of Medicine, Chiang Mai University, Chiang Mai 50200, Thailand; natthaphat.s@cmu.ac.th (N.S.-A.); nipon.chat@cmu.ac.th (N.C.)

3 Cardiac Electrophysiology Unit, Department of Physiology, Faculty of Medicine, Chiang Mai University, Chiang Mai 50200, Thailand

4 Center of Excellence in Cardiac Electrophysiology Research, Chiang Mai University, Chiang Mai 50200, Thailand

5 Department of Oral Biology and Diagnostic Sciences, Faculty of Dentistry, Chiang Mai University, Chiang Mai 50200, Thailand