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Abstract

Chronic kidney disease stage 5 (CKD5) marks the fifth stage of renal failure, frequently causing dysregulation of bone and mineral metabolism. Challenges exist in evaluating and managing chronic kidney disease–mineral bone disorder (CKD-MBD) with the standard panel of biomarkers. Our objective was to profile osteopontin (OPN) in patients with CKD5 on maintenance hemodialysis (CKD5-HD) and elucidate its relationship to phosphorus (P), calcium (Ca²⁺), alkaline phosphatase (AP), and intact parathyroid hormone (iPTH) to improve understanding of the present model of CKD-MBD. Elevation of plasma OPN was seen in the CKD5-HD cohort (n = 92; median: 240.25 ng/mL, interquartile range [IQR]: 169.85 ng/mL) compared to a normal group (n = 49; median: 63.30 ng/mL, IQR: 19.20 ng/mL; p < .0001). Spearman correlation tests revealed significant positive correlations of OPN with iPTH (p < .0001; r = 0.561, 95% confidence interval = 0.397-0.690) and OPN with AP (p < .0001; r = 0.444, 95% confidence interval = 0.245-0.590) in CKD5-HD patients. Ultimately, OPN may play an integral role in the MBD axis, suggesting that it may be important to actively monitor OPN when managing CKD5-HD.

Keywords

kidney disease hemodialysis biomarkers mineral and bone disorder osteopontin

Introduction

Chronic kidney disease stage 5 (CKD5) is the fifth stage of renal failure, with treatment necessitating dialysis or kidney transplant. The number of CKD5 prevalent cases rises by approximately 20,000 per year, and in 2016, the number of prevalent cases reached 726,331.¹ Chronic kidney disease leads to dysregulation of calcium (Ca²⁺), phosphorus (P), parathyroid hormone (PTH), and vitamin D metabolism, resulting in biochemical laboratory abnormalities, significant bone disease, and/or vascular calcification that define chronic kidney disease–mineral and bone disorder (CKD-MBD).^2
-4 Kidney Disease Improving Global Outcomes (KDIGO) recommends evaluation of CKD-MBD by monitoring serum Ca²⁺, phosphate, PTH, and alkaline phosphatase (AP) starting in stage G3a of CKD.^2
-4 However, even with careful monitoring of these markers, patients with CKD5 on maintenance hemodialysis (CKD5-HD) have poorer health outcomes related to MBD, such as increased risk of developing cardiovascular disease and increased fracture risk.^5
-7 While obtaining abdominal radiographs, computed tomography–based imaging, or dual-energy X-ray absorptiometry scans or performing a bone biopsy to help evaluate calcification and bone mineral density (BMD) status, the strength of recommendation of these practices is low.^3,4 Therefore, the utility in identifying additional biomarkers to better evaluate, diagnose, and treat patients with CKD5-HD with MBD is appealing.^8

-11

Osteopontin (OPN) is a glycol-phosphoprotein found in bone, acute and chronic inflammatory cells, smooth muscle, epithelial, and endothelial cells, neurons, and fetal renal tissue and is expressed in the thick ascending limb of the loop of Henle.^12,13 Some of its functions include increasing macrophage and T-cell counts, perpetuation of inflammation, wound healing, tumor development and progression, roles in diabetes, and possible roles in the regulation of nephrolithiasis and nephrogenesis.^12,14 OPN was also found to promote angiogenesis, encourage growth and invasion of renal cancer, impact the development of lupus nephritis in patients with systemic lupus erythematosus, and potentially be useful as a marker of acute allograft rejection in kidney transplants.^15

-20 Additionally, OPN is important in regulation of vascular calcification and bone mineralization. Local increases in OPN in vessel walls have been linked to atherosclerotic plaque formation, inflammation within arteries, and smooth muscle mineralization.^12,21,22 The function of OPN in bone is defined by its ability to anchor osteoclasts via the α_vβ₃ integrin.²³ By anchoring osteoclasts, OPN plays a significant role in bone resorption and turnover.

A number of biomarkers have been evaluated in CKD-MBD, but there is a clear need for better characterization of novel markers, especially within the context of CKD5-HD.^9,10,11 Serum monitoring of OPN may satisfy this need for improved diagnosis and management of CKD-MBD as it (1) has clear physiologic functions associated with bone and mineral regulation and (2) has been shown to have high sensitivity and specificity in diagnosing bone turnover disease and coronary calcification in certain populations.^24,25 Our study sought to evaluate the plasma levels of OPN in a CKD5-HD population and describe its relationship to the current markers used for evaluating MBD established by the KDIGO group.

Methods

Whole blood samples were collected in sodium citrate tubes from stable patients with CKD5-HD treated at Loyola University Hospital System’s Outpatient Dialysis Center (n = 92). Normal plasma was obtained from 49 nonsmoking, drug-free healthy volunteers (25 males and 24 females) from George King Bio-Medical, Inc (Overland Park, Kansas). Both CKD5-HD and normal plasma samples were frozen at −80°C for storage and later analysis. Plasma levels of OPN in both populations were measured using a commercially available OPN sandwich enzyme-linked immunosorbent assays (R&D Systems, Minneapolis, Minnesota). Epic electronic medical record charts of the 92 CKD5-HD patients were reviewed for levels of iPTH, AP, P, and Ca²⁺ at the time blood samples were collected. Demographics such as age, sex, race/ethnicity, body mass index, comorbidities, medications, and supplementation were obtained through the chart review and are represented in Table 1.

Table 1.

Demographics of Patient With CKD5-HD, Loyola University Hospital System Outpatient Dialysis Center.

Demographic	No. of Patients^a (% of Total)
Age	60.5 (20.2-94.8)^b
Sex
Male	44 (47.8%)
Female	48 (52.2%)
Race/ethnicity
White	24 (26.1%)
Black	49 (53.3%)
Asian/Pacific Islander	0 (0.0%)
Native American/Alaskan Native	0 (0.0%)
Hispanic/Latino/other	19 (20.7%)
BMI
<20 kg/m²	4 (4.3%)
20-24.9 kg/m²	25 (27.2%)
25-30 kg/m²	26 (28.3%)
>30 kg/m²	37 (40.2%)
Comorbidities
Diabetes	60 (65.2%)
Coronary artery disease	44 (47.9%)
Osteoporosis/osteopenia	23 (25.0%)
Comorbidities
Atrial fibrillation	14 (15.2%)
Stroke history	17 (18.5%)
Medications
Calcium acetate	29 (31.5%)
Lanthanum	5 (5.4%)
Cinacalcet	16 (17.4%)
Sevelamer carbonate	42 (45.7%)
ACE inhibitor	15 (16.3%)
ARB	5 (5.4%)
Supplementation
Vitamin D	82 (91.3%)
Calcium carbonate	10 (10.9%)

Abbreviations: ACE, angiotensin-converting enzyme; ARB, angiotensin II receptor blocker; BMI, body mass index; CKD5-HD, chronic kidney disease on maintenance hemodialysis.

^a n = 92.

^b Age reported as mean age (minimum age-maximum age).

Statistical Analyses

Comparison of OPN levels between the CKD5-HD and normal population was performed utilizing a Mann-Whitney t test, with a p < .05 demonstrating statistical significance. Results were expressed as median and interquartile range (IQR: IQR presented as one value, Q3 - Q1). Relationships of OPN levels with iPTH, AP, P, and Ca²⁺ levels were identified via Spearman correlation tests. Results were deemed statistically significant with p < .05. The r values were produced to determine the strength of correlation between 2 markers. The data was collected in Microsoft Excel and analyzed using Windows GraphPad Prism v7 software (GraphPad Software, La Jolla, California). Normality was determined via the IBM SPSS Statistics software (SPSS Inc., Chicago, IL).

Results

Primary analysis compared plasma OPN levels in the CKD5-HD cohort to the normal population. A statistically significant elevation of OPN plasma levels in the CKD5-HD cohort (median: 240.25 ng/mL, IQR: 169.85 ng/mL) was observed compared to the normal group (median: 63.30 ng/mL, IQR: 19.20 ng/mL) with a p value of <.0001 via the Mann-Whitney t test (Figure 1).

Figure 1.

Plasma level differences of osteopontin (OPN) in a normal control group and patients with chronic kidney disease stage 5 on hemodialysis (CKD5-HD). Normal control: n = 49; median = 63.3 ng/mL; interquartile range (IQR) = 19.20 ng/mL; CKD5-HD: n = 92; median = 240.25 ng/mL; IQR = 169.85 ng/mL. Plasma levels of OPN are elevated in CKD5-HD compared to normal control group (p < .0001 via Mann-Whitney t test).

Correlation analyses were conducted between OPN and iPTH, OPN, and AP, OPN and Ca²⁺, and OPN and P. Spearman correlation tests revealed a significant positive correlation of OPN with iPTH (p < .0001; r = 0.561, 95% confidence interval = 0.397-0.690) and OPN with AP (p < .0001; r = 0.444, 95% confidence interval = 0.245-0.590) in the CKD5-HD cohort (Figure 2). No other correlations were found between OPN and total Ca²⁺ (p = .36, r = 0.096; data not shown) or OPN and P (p = .269, r = 0.116; data not shown).

Figure 2.

Spearman correlational analysis comparing osteopontin (OPN) to intact parathyroid hormone (iPTH), and OPN to alkaline phosphatase (AP) in patients with chronic kidney disease stage 5 on hemodialysis (CKD5-HD). Results show a significantly positive correlation between OPN and iPTH (p < .0001; r = 0.561, 95% confidence interval = 0.397-0.690) and OPN and AP (p < .0001; r = 0.444, 95% confidence interval = 0.245-0.590).

Discussion

Our results show a significant increase in the plasma level of OPN in patients with CKD5-HD. These findings are consistent with previous studies in CKD showing elevated OPN.²⁶ Elevation of OPN in our patient samples may be due to its upregulation in chronic inflammatory states and/or due to MBD.^9,27 High levels of OPN are important to note as they are associated with all-cause mortality in patients with CKD5-HD.²⁸

Chronic kidney disease leads to Ca²⁺-P-vitamin D-PTH dysregulation.²⁶ Progression of CKD causes more severe MBD, which can lead to severe bone disease such as osteitis, osteomalacia, and possibly osteoporosis.^29,30

The relationship of OPN to the Ca²⁺-P-vitamin D-PTH axis is more complex. A study from Shen and Christakos demonstrated marked increase in OPN transcription with stimulation of the vitamin D receptor.³¹ Animal studies of vascular calcification have demonstrated an increase in OPN expression after treatment with vitamin D receptor agonists.³² OPN knockout mice have shown to have higher bone formation secondary to PTH-induced increase in osteoblast activity, and PTH infusion has been shown to increase OPN levels.^33,34 These studies exemplify how OPN expression can be significantly altered by changes in a number of variables in the Ca²⁺-P-vitamin D-PTH axis.

AP is a common marker of bone turnover and has shown to be elevated in patients with CKD with low BMD and is also reported to be associated with higher mortality in patients with or without MBD.^4,35
-37 Our results show a positive correlation between OPN and AP. OPN has been shown to play a role in the inhibition of vessel calcification.²¹ Lomashvili and colleagues demonstrated in rat models how AP can both dephosphorylate OPN and increase aortic calcification.^21,38 Both OPN and AP have increased expression in high bone turnover states.^23,36 However, variability of duration on dialysis, degree of inflammation, and status of calcification and bone turnover states in our CKD5-HD cohort make it difficult to predict the cause of the direct relationship between OPN and AP.^35,36,38

Limitations

Our study has several limitations. The sample size is relatively small, and samples were collected at varying time points during the day. Circadian rhythm and diet may have affected the levels of proteins and minerals that were measured.³⁹ The amount of time since starting dialysis varied for each patient, which could have affected levels of certain biomarkers.³⁵ The patients with CKD5-HD had a significant number of comorbidities, such as diabetes, hypertension, and coronary/peripheral artery disease which could have affected the regulation of Ca²⁺, P, OPN, and iPTH. However, our cohort represents the typical patients with CKD5-HD as most are found to have upward of 3 comorbidities at any given time.⁴⁰ Serum bicarbonate and bone-specific AP have been studied in CKD-MBD; however, we did not include these markers in our study due to lower strength of recommendation outlined by KDIGO as well as cost of add-on laboratory testing.⁴ Future studies may warrant multivariate modeling of OPN with these markers in addition to the ones studied.

Conclusion

This study demonstrates a significant elevation in OPN in patients with CKD5-HD compared to normal blood samples and identified a direct relationship between OPN and iPTH and OPN and AP in the same CKD5-HD cohort. These findings suggest OPN may play an integral role in ion homeostasis, vascular calcification, and bone turnover axes in CKD5-HD. This underscores that it may be important to include OPN in the group of markers used to evaluate patients with CKD5-HD for mineral and bone dysregulation.

Footnotes

Authors’ Note

Ethical approval to report this case was obtained from Loyola University Chicago Health Sciences Institutional Review Board (#LU107346). Verbal informed consent was obtained from the patients for their anonymized information to be published in this article.

Acknowledgments

The authors are grateful to the patients and staff involved with the collection of samples at the Loyola Outpatient Dialysis Center. The authors would like to thank the Hemostasis and Thrombosis Laboratories at Loyola University Chicago Health Sciences Campus for their mentorship and guidance.

Declaration of Conflicting Interests

The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Funding

The author(s) disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: Research reported in this manuscript was supported by the T35 training grant from the National Institute of Allergy and Infectious Diseases (NIAID) of the National Institutes of Health (NIH) under the award number T35 AI125220.

ORCID iDs

Aleksander Druck

Dimpi Patel

Jawed Fareed

References

United States Renal Data System. United States Renal Data System. 291–332. https://www.usrds.org/2016/view/Default.aspx.

Moe

Drueke

Cunningham

, et al. Definition, evaluation, and classification of renal osteodystrophy: a position statement from Kidney Disease: Improving Global Outcomes (KDIGO). Kidney Int. 2006;69(11):1945–1953. doi:S0085-2538(15)51415-4.

Kidney Disease: Improving Global Outcomes (KDIGO) CKD-MBD Work Group. KDIGO clinical practice guideline for the diagnosis, evaluation, prevention, and treatment of chronic kidney disease-mineral and bone disorder (CKD-MBD). Kidney Int Suppl. 2009;(113):S1–130. doi:10.1038/ki.2009.188.

KDIGO 2017 clinical practice guideline update for the diagnosis, evaluation, prevention, and treatment of chronic kidney Disease–Mineral and Bone Disorder (CKD-MBD). Kidney Int Suppl. 2017;7(1):1–59. https://www.sciencedirect.com/science/article/pii/S2157171617300011. doi:10.1016/j.kisu.2017.04.001.

Kalantar-Zadeh

Kuwae

Regidor

, et al. Survival predictability of time-varying indicators of bone disease in maintenance hemodialysis patients. Kidney Int. 2006;70(4):771–780. http://www.sciencedirect.com/science/article/pii/S0085253815519959. doi:10.1038/sj.ki.5001514.

Alem

Sherrard

Gillen

, et al. Increased risk of hip fracture among patients with end-stage renal disease. Kidney Int. 2000;58(1):396–399. doi:S0085-2538(15)47111-X.

Fried

Biggs

Shlipak

, et al. Association of kidney function with incident hip fracture in older adults. J Am Soc Nephrol. 2007;18(1):282–286. doi:ASN.2006050546.

Babayev

Nickolas

. Can one evaluate bone disease in chronic kidney disease without a biopsy? Curr Opin Nephrol Hypertens. 2014;23(4):431–437. doi:10.1097/01.mnh.0000447014.36475.58.

Alderson

Ritchie

Green

Chiu

Kalra

. Potential for biomarkers of chronic kidney disease-mineral bone disorder to improve patient care. Nephron Clin Pract. 2013;124(3-4):141–150. https://www.karger.com/DOI/10.1159/000356394. doi:10.1159/000356394.

10.

Tan

Cai

. Is there a role for newer biomarkers in chronic kidney disease‐mineral and bone disorder management? Nephrology. 2017;22(S2):14–18. https://onlinelibrary.wiley.com/doi/abs/10.1111/nep.13015. doi:10.1111/nep.13015.

11.

Ortiz

Massy

Fliser

, et al. Clinical usefulness of novel prognostic biomarkers in patients on hemodialysis. Nat Rev Nephrol. 2011;8(3):141–150. doi:10.1038/nrneph.2011.170.

12.

Icer

Gezmen-Karadag

. The multiple functions and mechanisms of osteopontin. Clin Biochem. 2018;59:17–24. http://www.sciencedirect.com/science/article/pii/S0009912018303837. doi:10.1016/j.clinbiochem.2018.07.003.

13.

Denhardt

Guo

. Osteopontin: a protein with diverse functions. FASEB J. 1993;7(15):1475–1482.

14.

Hudkins

Giachelli

Cui

Couser

Johnson

Alpers

. Osteopontin expression in fetal and mature human kidney. J Am Soc Nephrol. 1999;10(3):444. http://jasn.asnjournals.org/content/10/3/444.abstract.

15.

Kaleta

. The role of osteopontin in kidney diseases. Inflamm Res. 2019;68(2):93–102. doi:10.1007/s00011-018-1200-5.

16.

Liu

Chen

Guo

Yuan

. Influence of osteopontin short hairpin RNA on the proliferation and invasion of human renal cancer cells. J Huazhong Univ Sci Technolog Med Sci. 2010;30(1):61–68. doi:10.1007/s11596-011-0111-7.

17.

Wai

Kuo

. Osteopontin: regulation in tumor metastasis. Cancer Metastasis Rev. 2008;27(1):103–118. doi:10.1007/s10555-007-9104-9.

18.

Bai

, et al. Osteopontin gene polymorphism in association with systemic lupus erythematosus in Chinese patients. Chin Med J (Engl). 2007;120(23):2124–2128.

19.

Wong

Lit

Tam

Lam

. Elevation of plasma osteopontin concentration is correlated with disease activity in patients with systemic lupus erythematosus. Rheumatology (Oxford). 2005;44(5):602–606. doi:10.1093/rheumatology/keh558.

20.

Jin

Tian

Wang

, et al. Kidney injury molecule-1 and osteopontin: New markers for prediction of early kidney transplant rejection. Mol Immunol. 2013;54(3-4):457–464. doi:10.1016/j.molimm.2013.01.013.

21.

Scatena

Liaw

Giachelli

. Osteopontin: a multifunctional molecule regulating chronic inflammation and vascular disease. Arterioscler Thromb Vasc Biol. 2007;27(11):2302–2309. http://atvb.ahajournals.org/cgi/content/abstract/27/11/2302. doi:10.1161/ATVBAHA.107.144824.

22.

Abdalrhim

Marroush

Austin

, et al. Plasma osteopontin levels and adverse cardiovascular outcomes in the PEACE trial. PLoS One. 2016;11(6):e0156965. https://www.ncbi.nlm.nih.gov/pubmed/27284698. doi:10.1371/journal.pone.0156965.

23.

Chellaiah

Hruska

. The integrin {alpha}v{beta}3 and CD44 regulate the actions of osteopontin on osteoclast motility. Calcif Tissue Int. 2003;72(3):197–205. https://search.proquest.com/docview/213010437. doi:10.1007/s00223-002-1025-6.

24.

Wei

Huang

Tan

Chen

Deng

. Serum osteopontin levels in relation to bone mineral density and bone turnover markers in postmenopausal women. Scand J Clin Lab Invest. 2016;76(1):33–39. doi:10.3109/00365513.2015.1087045.

25.

Kardesoglu

Yiginer

, et al. The relationship between coronary calcification and the metabolic markers of osteopontin, fetuin-A, and visfatin. Turk Kardiyol Dern Ars. 2009;37(6):397–402.

26.

Barreto

Lenglet

Liabeuf

, et al. Prognostic implication of plasma osteopontin levels in patients with chronic kidney disease. Nephron Clin Pract. 2011;117(4):c363–372. https://www.karger.com/DOI/10.1159/000321520. doi:10.1159/000321520.

27.

Lund

Giachelli

Scatena

. The role of osteopontin in inflammatory processes. J Cell Commun Signal. 2009;3(3-4):311–322. doi:10.1007/s12079-009-0068-0.

28.

Scialla

Kao

WHL

Crainiceanu

, et al. Biomarkers of vascular calcification and mortality in patients with ESRD. Clin J Am Soc Nephrol. 2014;9(4):745–755. http://cjasn.asnjournals.org/content/9/4/745.abstract. doi:10.2215/CJN.05450513.

29.

Yuen

Ananthakrishnan

Campbell

. Hyperparathyroidism of renal disease. Perm J. 2016;20(3):15–127. doi:10.7812/TPP/15-127.

30.

Goodman

. Renal osteodystrophy: pathogenic mechanisms and therapeutic options. In: Bilezikian

Raisz

Martin

, eds. Principles of Bone Biology. 3rd ed. San Diego, CA: Academic Press; 2008:1479–1510. Chap 68. http://www.sciencedirect.com/science/article/pii/B978012373884400001X. doi:10.1016/B978-0-12-373884-4.00001-X.

31.

Shen

Christakos

. The vitamin D receptor, runx2, and the notch signaling pathway cooperate in the transcriptional regulation of osteopontin. J Biol Chem. 2005;280(49):40589–40598. doi:10.1074/jbc.M504166200.

32.

Lau

Leaf

, et al. Vitamin D receptor agonists increase klotho and osteopontin while decreasing aortic calcification in mice with chronic kidney disease fed a high phosphate diet. Kidney Int. 2012;82(12):1261–1270. doi:10.1038/ki.2012.322.

33.

Yuan

Sato

Densmore

, et al. Deletion of PTH rescues skeletal abnormalities and high osteopontin levels in klotho−/− mice. PLOS Genet. 2012;8(5):e1002726. doi:10.1371/journal.pgen.1002726.

34.

Ono

Nakashima

Rittling

, et al. Osteopontin negatively regulates parathyroid hormone receptor signaling in osteoblasts. J Biol Chem. 2008;283(28):19400–19409. doi:10.1074/jbc.M800005200.

35.

Blayney

Pisoni

Bragg-Gresham

, et al. High alkaline phosphatase levels in hemodialysis patients are associated with higher risk of hospitalization and death. Kidney Int. 2008;74(5):655–663. http://www.sciencedirect.com/science/article/pii/S0085253815533711. doi:10.1038/ki.2008.248.

36.

Park

Kovesdy

Duong

, et al. Association of serum alkaline phosphatase and bone mineral density in maintenance hemodialysis patients. Hemodial Int. 2010;14(2):182–192. doi:10.1111/j.1542-4758.2009.00430.x.

37.

Fan

Jin

Jiang

Fang

. Elevated serum alkaline phosphatase and cardiovascular or all-cause mortality risk in dialysis patients: a meta-analysis. Sci Rep. 2017;7(1):13224. doi:10.1038/s41598-017-13387-z.

38.

Lomashvili

Cobbs

Hennigar

Hardcastle

O’Neill

. Phosphate-induced vascular calcification: role of pyrophosphate and osteopontin. J Am Soc Nephrol. 2004;15(6):1392–1401. http://jasn.asnjournals.org/content/15/6/1392.abstract. doi:10.1097/01.ASN.0000128955.83129.9C.

39.

Portale

Halloran

Morris

Jr . Dietary intake of phosphorus modulates the circadian rhythm in serum concentration of phosphorus. Implications for the renal production of 1,25-dihydroxyvitamin D. J Clin Invest. 1987;80(4):1147–1154. doi:10.1172/JCI113172.

40.

Tonelli

Wiebe

Guthrie

, et al. Comorbidity as a driver of adverse outcomes in people with chronic kidney disease. Kidney Int. 2015;88(4):859–866. doi:10.1038/ki.2015.228.

Osteopontin Levels in Patients With Chronic Kidney Disease Stage 5 on Hemodialysis Directly Correlate With Intact Parathyroid Hormone and Alkaline Phosphatase

Abstract

Keywords

Introduction

Methods

Statistical Analyses

Results

Discussion

Limitations

Conclusion

Footnotes

Authors’ Note

Acknowledgments

Declaration of Conflicting Interests

Funding

ORCID iDs

References