Abstract
Fetuin-A is a hepatic secretory protein that promotes bone mineralization in vitro. Whether fetuin-A levels are associated with BMD in humans is unknown. The Health Aging and Body Composition study enrolled 3075 well-functioning black and white persons 70–79 yr of age and measured BMD. This cross-sectional study measured serum fetuin-A using ELISA among a random sample of 508 participants within sex and race strata. Multivariate linear regression analysis evaluated the associations of fetuin-A with BMD. Among women (n = 257), higher fetuin-A levels were significantly associated with higher total hip (p = 0.02), lumbar spine (p = 0.03), and whole body BMD (p = 0.01) in models adjusted for age, race, diabetes, alcohol and tobacco use, physical activity, body mass index, C-reactive protein levels, calcium supplement, and estrogen use. For example, each SD (0.38 g/liter) higher level of fetuin-A was associated with 0.016 g/cm2 higher total hip areal BMD. The association was of similar magnitude and direction for femoral neck BMD but did not reach statistical significance (p = 0.11). In contrast, among men (n = 251), fetuin-A had no significant associations with total hip (p = 0.79), lumbar spine (p = 0.35), whole body (p = 0.46), or femoral neck BMD (p = 0.54) in multivariable models. We conclude that higher fetuin-A levels are independently associated with higher BMD among well-functioning community-dwelling older women but not older men. Future studies should evaluate whether fetuin-A may refine fracture risk assessment in older women.
Key words: fetuin-A, BMD, elderly
INTRODUCTION
Osteoporosis is exceedingly common in older persons,(1) with an estimated 9 million osteoporotic fractures worldwide in 2000.(2) Hip fracture is strongly associated with loss of physical function, poor health-related quality of life,(3) and mortality.(4) Whereas advanced age and female sex are strong risk factors, biological pathways leading to osteoporosis remain incompletely understood. The discovery of novel pathways for osteoporosis could lead to new preventive strategies or therapeutic targets.
Fetuin-A (also called α-2 Heremans Schmid glycoprotein [AHSG]) is a 64-kDa serum protein that regulates calcium mineralization.(5) It is a hepatic secretory protein that is found in high levels in human serum (∼1 g/liter).(6) Fetuin-A complexes with calcium and phosphorus(7) and prevents the precipitation of these minerals from serum.(8) Several observations suggest that fetuin-A may play a role in the regulation of bone mineralization. First, it is among the most abundant noncollagenous proteins found in bone.(9,10) Because it is not produced by bone-resident cells and because it is selectively concentrated from serum into hydroxyapatite in vitro, bone fetuin-A is likely derived from hepatic synthesis and delivered to bone through serum.(11,12) In vivo, fetuin-A knockout mice develop osteomalacia by 3 mo of age.(13) Moreover, blood levels of fetuin-A differ in patients with Paget's disease of bone(14) and osteogenesis imperfecta(15) compared with healthy controls. However, whether and how fetuin-A might regulate bone mineralization remained unknown until recently. Building on observations that serum proteins could induce bone mineralization(16) and the size exclusion characteristics of type 1 collagen fibrils,(17) Toroian et al.(18) hypothesized that fetuin-A may simultaneously inhibit calcium precipitation in serum and promote calcification within bone. The investigators recently showed that fetuin-A directly promoted calcification within bone in an in vitro system. This important observation provides novel insights to the role of fetuin-A in bone homeostasis. However, the experiments were carried out using an acellular in vitro system and newborn rat bone.
Whether serum fetuin-A levels are associated with BMD in humans is uncertain. To that end, we evaluated the cross-sectional association of fetuin-A with BMD among 508 well-functioning older men and women who participated in the Health, Aging, and Body Composition (Health ABC) study. We hypothesized that higher fetuin-A levels would be associated with higher BMD, independent of previously recognized correlates of BMD in older persons.
MATERIALS AND METHODS
Participants
At the baseline study visit (April 1997 to June 1998), Health ABC enrolled 3075 well-functioning men and women 70–79 yr of age, recruited from a sample of Medicare beneficiaries at two clinical sites (Pittsburgh, PA, and Memphis, TN). To be eligible, participants reported no difficulty in walking a quarter mile, climbing 10 steps, or performing basic activities of daily living. Venous blood samples were obtained after overnight (8-h) fasts and stored at −70°C. Participants underwent a day-long evaluation that included medical history and physical activity assessment, medication use, physical examination, and BMD measurement. All participants provided written informed consent, and the study was approved by the institutional review boards at the University of Pittsburgh and the University of Tennessee Health Sciences Center, Memphis.
A stratified random sample of 508 participants was chosen from the Health ABC study to participate in this study. Because BMD varies substantially between men and women and blacks and whites, subjects were randomly selected from within sex and race strata (127 black men, 126 black women, 130 white men, and 125 white women) to ensure approximately equal numbers within each stratum.
Measurements
Fetuin-A
Fetuin-A levels were measured using a human fetuin-A ELISA kit (Epitope Diagnostics, San Diego, CA, USA). The assay uses a two-site “sandwich” technique with two polyclonal antibodies that bind to different epitopes of human fetuin-A. Measurements were performed at the Laboratory for Clinical Biochemistry Research at the University of Vermont. Fetuin-A was measured twice among each participant, and results were averaged. The intra-assay and interassay CVs were <5%. Among a 5% blind duplicate assessment at the LCBR, the intraindividual CVs were 13.5% and 20%, respectively.
BMD
Areal BMD was measured at the proximal femur and the whole body using DXA (Hologic 4500A, version 9.03; Hologic, Bedford, MA, USA) with identical protocols at both study sites. Lumbar spine BMD was determined from the lumbar spine subregion of the whole body scan. This method is highly correlated with site-specific lumbar spine BMD measurements and has predicted fractures in prior studies.(19) DXA quality assurance measurements, including use of daily and cross-calibration phantoms, were performed at both sites. The precision for whole body BMD was evaluated by performing duplicate scans on five volunteers for each scanner. The CV was 1.0% in Memphis and 0.4% in Pittsburgh for whole body BMD and 2.5% and 3.1%, respectively, for lumbar spine BMD. The precision for hip BMD was not available among these five volunteers but has been reported previously for the QDR 4500A as 0.9%(20) and 1.2%.(21)
Other measurements
Age, sex, and race were determined by self-report. Diabetes was defined by history of physician's diagnosis, use of hypoglycemic medications, fasting glucose level ≥126 mg/dl, or 2-h postchallenge plasma glucose level ≥200 mg/dl. Smoking and alcohol use and frequency were determined by questionnaire. Physical activity was assessed using self-report of walking and exercise and assigning kilocalories per week to activities. Height and weight were measured wearing light clothes and no shoes, and body mass index (BMI) was calculated (kg/m2). Creatinine was measured by colorimetry (Johnson & Johnson, New Brunswick, NJ, USA), and estimated glomerular filtration rate (eGFR) was calculated by the four-variable Modification of Diet in Renal Disease (MDRD) equation.(22) High-sensitivity C-reactive protein (CRP) was measured by ELISA (Calbiochem, San Diego, CA, USA) and was standardized to the World Health Organization's First International Reference Standard. Participants brought all medications that they had taken during the previous 2 wk including over-the-counter medications and supplements to the study visit. Medications, frequency, and dose were recorded by study personnel and matched to a dictionary of prescription and nonprescription drugs using the Iowa Drug Information System,(23) which identified the ingredients of each drug and allowed identification of the individual pharmaceutical compounds.
Statistical analyses
Because of marked sex differences in BMD shown in prior research, we stratified the study sample by sex, a priori. Results were qualitatively substantially different among men and women. p values for interaction for the association of fetuin-A with BMD by sex ranged from 0.11 to 0.37 across the measurement sites. To maximize power, we treated fetuin-A as a continuous variable (per SD) as our primary predictor variable. However, to evaluate differences in fetuin-A levels across demographic and clinical variables, we categorized subjects into tertiles groups of fetuin-A. This allowed us to maximize the number of subjects within each category while providing a cursory evaluation of the linearity of relation between of fetuin-A and each variable. Demographics and clinical characteristics were compared across tertiles by ANOVA or the Kruskal-Wallis test for continuous variables and the χ2 test or Fisher's exact test for categorical variables.
To explore the functional form of fetuin-A with BMD at each anatomic site, we used general additive models with a smoother to fit cubic B-spline functions to the data. The extreme 2.5% of the distribution of fetuin-A was excluded to avoid implausible extrapolations on both tails of the spline functions.(24) For the association of fetuin-A with whole body and lumbar spine BMD in women, the spline functions suggested deviation from linearity, particularly among fetuin-A levels in the highest third of the study sample. We evaluated quadratic terms (fetuin-A2), which were statistically significant, so all subsequent analyses for these outcomes included the quadratic functions to improve model fit. For these outcomes, data are presented graphically, as well as across fetuin-A tertiles. The associations of fetuin-A with BMD at all other sites in men and women were approximately linear. Quadratic terms at the other sites were evaluated in both sexes and were nonsignificant, so these models are presented without inclusion of the quadratic variable.
We evaluated the association of fetuin-A with BMD at the total hip, femoral neck, lumbar spine, and whole body using multivariate linear regression. Initial models were sex stratified and age and race adjusted. Final models were adjusted for age, race, physical activity, BMI, diabetes, smoking, alcohol use, CRP, estrogen use, and calcium use. Most of these variables were selected because prior publications showed that they are important correlates of BMD in older persons. In addition, any variable associated with fetuin-A with p ≤ 0.20 in univariate analysis in either men or women was included in the final model because relatively weak associations may still contribute substantial confounding effects.(25) This resulted in retention of CRP, estrogen use, and calcium use. Last, we created multiplicative interaction terms (fetuin × race), which were individually added to the final multivariable models to explore possible racial differences in the association of fetuin-A with BMD at each site. Two-tailed p < 0.05 was considered statistically significant. All analyses were conducted using SAS version 9.1.3 (SAS Institute, Cary, NC, USA) and SPlus V 6.1 (Insightful, Seattle, WA, USA).
RESULTS
The mean age of the 508 study participants was 74 yr. Fifty percent were women and black, reflecting the stratified sampling design. Distributions of all other variables evaluated in Table 1 were similar to the Health ABC parent study (data not shown). The mean BMI was 27 ± 5 kg/m2, 21% had diabetes, and 10% were current smokers. Mean BMD was lower among women at all measurement sites. For example, the mean total hip BMD was 0.81 ± 0.14 g/cm2 in women and 0.98 ± 0.15 g/cm2 in men (p < 0.0001). The mean serum fetuin-A level was 0.94 ± 0.42 g/liter and was similar in women and men (p = 0.53). Among women, demographics, medical history, body composition, and medication use were similar across fetuin-A tertiles; however, estrogen use was somewhat more common among women in the highest fetuin-A tertile. Men with higher fetuin-A levels were more frequently white and had lower CRP levels. Diabetes was more prevalent in the extremes of fetuin-A compared with men with intermediate levels (Table 1).
Table 1.
Demographic and Clinical Characteristics by Fetuin-A Tertile in Women and Men
In women, higher fetuin-A levels were associated with higher BMD at each measurement site (Fig. 1). Each SD (0.38 g/liter) increase in fetuin-A was associated with significantly higher total hip BMD in age- and race-adjusted models—an association that was essentially unaltered in fully adjusted models (Table 2). The association was in a similar direction but was of somewhat lower magnitude for femoral neck BMD, and the association did not reach statistical significance in the adjusted model (p = 0.11). Higher fetuin-A levels were also significantly associated with higher lumbar spine and whole body BMD; however, at these sites, the relationships were not linear. Fetuin-A was directly correlated with BMD throughout most of its range, but the association leveled at the highest third of the cohort (Fig. 1). A quadratic term (fetuin-A2) was included to account for this nonlinearity and showed that higher fetuin-A was significantly, albeit nonlinearly, associated with higher lumbar spine and whole body BMD in women. Table 3 provides adjusted mean BMD measurements by tertiles of fetuin-A among women at these measurement sites. We observed no effect modification by race for the association of fetuin-A with BMD at any site (all p values for interaction > 0.15).
FIG. 1.
Association of fetuin-A with BMD in older women. Natural cubic spline function showing β coefficients (solid lines) and 95% CIs (dotted lines) of fetuin-A with (A) total hip BMD; (B) femoral neck BMD; (C) lumbar spine BMD; and (D) whole body BMD. Spline functions were adjusted for age, race, diabetes, physical activity, BMI, alcohol use, tobacco use, CRP levels, and calcium and estrogen use.
Table 2.
Association of Fetuin-A and BMD, Stratified by Sex
Table 3.
Adjusted* Mean Areal BMD by Tertiles of Fetuin-A among Women (N = 257)
For the BMD measurement sites where the association was linear (total hip and femoral neck BMD), we compared the strength of the association with other continuous predictor variables; BMI, physical activity, and CRP levels. Skewed variables (physical activity and CRP) were log-transformed, and all variables were evaluated per SD change to facilitate comparisons. This analysis showed that the association of fetuin-A with BMD was approximately one fifth the strength of the association of BMI with BMD, approximately two thirds that of the association of physical activity with BMD, and approximately twice as strong as the association of CRP with BMD among women (Table 4).
Table 4.
Comparison of Strength of Associations of Continuous Predictors and BMD Among Women (N = 257)
In contrast, in men, we observed no significant association of fetuin-A with BMD at any site. Spline functions showed no evidence for a direct association between serum fetuin-A levels and BMD across the spectrum (Fig. 2). This was confirmed in multivariable-adjusted models, where the point estimates were substantially smaller than among women and were not statistically significant at any measurement site (Table 2). As with women, we observed no effect modification by race at any measurement site in men (all p values for interaction > 0.50).
FIG. 2.
Association of fetuin-A with BMD in older men. Natural cubic spline function showing β coefficients (solid lines) and 95% CIs (dotted lines) of fetuin-A with (A) total hip BMD; (B) femoral neck BMD; (C) lumbar spine BMD; and (D) whole body BMD. Spline functions were adjusted for age, race, diabetes, physical activity, BMI, alcohol use, tobacco use, CRP levels, and calcium use.
DISCUSSION
Recent studies have shown that fetuin-A is a serum protein that promotes bone mineralization in vitro.(18) Whether fetuin-A regulates bone mineralization in vivo is unknown. This study is novel in showing that higher serum fetuin-A levels are associated with higher BMD in humans, independent of previously recognized osteoporotic risk factors. The associations were of modest strength and consistently observed across BMD measurement sites but were limited to women. In the context of prior in vitro experiments, these observations are consistent with the hypothesis that fetuin-A may regulate bone mineralization in vivo in older women.
In 2004, Hamlin and Price(26) observed that demineralized bone would remineralize when bathed in serum but would not do so when bathed in buffered solution with physiologic levels of calcium and phosphorus. Serum protein separation columns showed that the responsible “serum calcification factor” was between 55 and 150 kDa in size.(16) More recently, Toroian et al.(18) showed that the aqueous space within type 1 collagen fibrils—the primary site of bone mineralization—excluded proteins >40 kDa. Therefore, the serum calcification factor was too large to penetrate type 1 collagen fibrils, suggesting that it might regulate bone mineralization without being physically present at the site of mineralization. Because fetuin-A was known to potently inhibit calcium and phosphorus mineralization in serum,(8,27,28) and had a molecular size of 64 kDa,(28) these investigators hypothesized that fetuin-A may promote calcification within the fibril by selectively inhibiting the growth of apatite mineral crystals outside the fibril. Because fetuin-A is too large to penetrate the fibril,(17) any mineral crystal within the fibril could grow unencumbered. The investigators recently showed that demineralized bone would not remineralize when exposed to serum depleted of fetuin-A but would remineralize when serum was reconstituted with fetuin-A.(18) Thus, in vitro experiments showed that fetuin-A directly promotes bone mineralization. The observations presented here are novel in showing a direct association of fetuin-A and BMD in older women in vivo.
Future studies are needed to fully elucidate the mechanisms by which fetuin-A may regulate bone structure and function. Whereas observations presented here and in prior studies showed that fetuin-A may promote bone mineralization, the significance of this relationship in regard to bone quality, and ultimately to fractures, is unknown. Not only the BMC but also its heterogeneity and spatial distribution are important determinants of bone quality.(29,30) In addition, the in vitro experiments described above were carried out using an acellular experimental system. Thus, fetuin-A may have additional regulatory effects on bone homeostasis conferred through osteoblasts or other bone-resident cells. In support of this hypothesis, vascular smooth muscle cells that are stimulated to calcify strongly resemble osteoblasts. They upregulate key transcription factors of the osteoblast cell lineage, produce calcium binding proteins, and secrete matrix vesicles into the extracellular space, which subsequently calcify.(31,32) Recently, Reynolds et al.(33) showed that fetuin-A could be internalized by vascular smooth muscle cells, were concentrated within matrix vesicles, and were resecreted into the extracellular space within these vesicles. The presence of fetuin-A within the matrix vesicles abrogated their ability to function as a nidus for calcification. Because osteoblasts also secrete matrix vesicles in the course of bone calcification(34) and have similar phenotype to vascular smooth muscle cells, it is plausible that fetuin-A may have cell-mediated local regulatory effects on bone mineralization above and beyond its effects in serum. This important physiologic question requires evaluation in future studies.
To our knowledge, only one prior study has evaluated the association of fetuin-A and BMD. Fiore et al.(35) evaluated 90 subjects with atherosclerosis and 80 healthy volunteers in a case-control study aimed to determine the associations of fetuin-A with subclinical cardiovascular disease. The participants also underwent lumbar spine and femoral neck BMD measurements, and the investigators reported no statistically significant association of serum fetuin-A levels and BMD. Differences in study design likely account for the discrepant findings. This study was larger, included older participants, and was stratified by sex. Because the associations observed in our study were limited to women, they may have been missed in the study by Fiore et al. In addition, subjects in the Fiori study were recruited on the basis of cardiovascular disease status. Because fetuin-A levels are lower in subjects with cardiovascular disease,(35,36) it is possible that its presence may have directly or indirectly influenced fetuin-A levels.
Fetuin-A was associated with higher BMD only among women. The reasons underlying this sex difference is unknown. We hypothesize that this may be explained by differences in the spectrum of disease. Men had significantly higher BMD at all measurement sites in our study and are at risk for hip fracture ∼10 yr after their female counterparts.(37) Alternatively, differences in sex hormones may modify the association of fetuin-A with BMD. In addition, the association of fetuin-A with whole body and lumbar spine BMD flattened in approximately the highest third of fetuin-A levels. The reasons underlying this deviation from linearity are also unknown. Fetuin-A has other putative effects including anti-inflammatory properties(38) and inhibition of insulin signaling.(39) It is possible that factors involved in the regulation of these processes affected fetuin-A levels at extreme values. These important questions require future study.
Strengths of this study include its community-based setting, balanced distribution of sex and race, available measurements of important confounding variables, and BMD measurement at multiple anatomic sites. However, the study has limitations. First, the study participation was limited to ages 70–79, and either black or white race, so the findings may not generalize to other age groups or race/ethnicities. The study is cross-sectional and can not evaluate the direction of observed associations. Although in vitro data suggest that fetuin-A influences bone mineralization, we can not exclude the possibility that BMD, or unmeasured factors associated with BMD, may have altered fetuin-A levels. Interassay and intra-assay CVs of fetuin-A were higher in this study than in previous studies.(40,41) This was mitigated to some degree by measuring fetuin-A twice in each subject and using averaged values. Any misclassification caused by measurement error should also have biased our results toward the null hypothesis. The lumbar spine BMD measurement was derived from the whole body BMD DXA scans. Other assessment modalities may yield different results. Last, fetuin-A was measured at only one point in time. Whether longitudinal trajectories of fetuin-A provide additional, or more specific, information regarding BMD is an important topic for future research.
In conclusion, higher fetuin-A levels are independently associated with higher BMD among well-functioning community-dwelling older women. In conjunction with previous research, these data are consistent with the hypothesis that fetuin-A may directly promote bone mineralization in older women. Future research is needed to evaluate whether cell-mediated processes may contribute to the role of fetuin-A in regulation of bone mineralization and to determine whether lower fetuin-A levels are associated with incident fractures in older women.
ACKNOWLEDGMENTS
The authors thank the other investigators, the staff, and the participants of the Health ABC study for valuable contributions and Dr Elsa Strotmeyer for her critical review of the manuscript. This study was supported by an American Diabetes Association (ADA)–ASP Young Investigator Innovation Award in Geriatric Endocrinology sponsored by the Atlantic Philanthropies, ADA, the John A. Hartford Foundation, and ASP, an American Heart Association Fellow to Faculty Transition Award (JHI), and Contracts N01-AG-6-2101, N01-AG-6-2103, and N01-AG-6-2106 and an Intramural Research Program from the National Institutes on Aging (NIA). The funding sources played no role in the design and conduct of the study; collection, management, analysis, or interpretation of the data, nor in the preparation of the manuscript. The NIA reviewed and approved this manuscript before submission.
REFERENCES
- 1.National Osteoporosis Foundation. Washington, DC, USA: National Osteoporosis Foundation; 2002. America's Bone Health: The State of Osteoporosis and Low Bone Mass in Our Nation. [Google Scholar]
- 2.Johnell O, Kanis JA. An estimate of the worldwide prevalence and disability associated with osteoporotic fractures. Osteoporos Int. 2006;17:1726–1733. doi: 10.1007/s00198-006-0172-4. [DOI] [PubMed] [Google Scholar]
- 3.Randell AG, Nguyen TV, Bhalerao N, Silverman SL, Sambrook PN, Eisman JA. Deterioration in quality of life following hip fracture: A prospective study. Osteoporos Int. 2000;11:460–466. doi: 10.1007/s001980070115. [DOI] [PubMed] [Google Scholar]
- 4.Gordon PC. The probability of death following a fracture of the hip. Can Med Assoc J. 1971;105:47–51, passim. [PMC free article] [PubMed] [Google Scholar]
- 5.Ketteler M, Vermeer C, Wanner C, Westenfeld R, Jahnen-Dechent W, Floege J. Novel insights into uremic vascular calcification: Role of matrix Gla protein and alpha-2-Heremans Schmid glycoprotein/fetuin. Blood Purif. 2002;20:473–476. doi: 10.1159/000063554. [DOI] [PubMed] [Google Scholar]
- 6.Ix JH, Chertow GM, Shlipak MG, Brandenburg VM, Ketteler M, Whooley MA. Fetuin-A and kidney function in persons with coronary artery disease–data from the Heart and Soul Study. Nephrol Dial Transplant. 2006;21:2144–2151. doi: 10.1093/ndt/gfl204. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 7.Price PA, Thomas GR, Pardini AW, Figueira WF, Caputo JM, Williamson MK. Discovery of a high molecular weight complex of calcium, phosphate, fetuin, and matrix gamma-carboxyglutamic acid protein in the serum of etidronate-treated rats. J Biol Chem. 2002;277:3926–3934. doi: 10.1074/jbc.M106366200. [DOI] [PubMed] [Google Scholar]
- 8.Heiss A, DuChesne A, Denecke B, Grotzinger J, Yamamoto K, Renne T, Jahnen-Dechent W. Structural basis of calcification inhibition by alpha 2-HS glycoprotein/fetuin-A. Formation of colloidal calciprotein particles. J Biol Chem. 2003;278:13333–13341. doi: 10.1074/jbc.M210868200. [DOI] [PubMed] [Google Scholar]
- 9.Quelch KJ, Cole WG, Melick RA. Noncollagenous proteins in normal and pathological human bone. Calcif Tissue Int. 1984;36:545–549. doi: 10.1007/BF02405363. [DOI] [PubMed] [Google Scholar]
- 10.Dickson IR, Poole AR, Veis A. Localisation of plasma alpha2HS glycoprotein in mineralising human bone. Nature. 1975;256:430–432. doi: 10.1038/256430a0. [DOI] [PubMed] [Google Scholar]
- 11.Wendel M, Heinegard D, Franzen A. A major non-collagenous 62 kDa protein from rat bone mineralized matrix is identical to pp63 a phosphorylated glycoprotein from liver. Matrix. 1993;13:331–339. doi: 10.1016/s0934-8832(11)80029-6. [DOI] [PubMed] [Google Scholar]
- 12.Mizuno M, Farach-Carson MC, Pinero GJ, Fujisawa R, Brunn JC, Seyer JM, Bousfield GR, Mark MP, Butler WT. Identification of the rat bone 60K acidic glycoprotein as alpha 2HS-glycoprotein. Bone Miner. 1991;13:1–21. doi: 10.1016/0169-6009(91)90046-3. [DOI] [PubMed] [Google Scholar]
- 13.Brandenburg V, Kruger T, Westenfeld R, Schafer C, Jahnen-Dechent W, Floege J, Martin D, Shalhoub V, Ketteler M. Administration of calcimimmetic AMG641 improves osteomalacia in fetuin-A−/− mice with mild renal insufficiency. Am Soc Nephrol [abstract] 2007 [Google Scholar]
- 14.Ashton BA, Smith R. Plasma alpha 2HS-glycoprotein concentration in Paget's disease of bone: Its possible significance. Clin Sci (Lond) 1980;58:435–438. doi: 10.1042/cs0580435. [DOI] [PubMed] [Google Scholar]
- 15.Dickson IR, Bagga M, Paterson CR. Variations in the serum concentration and urine excretion of alpha 2HS-glycoprotein, a bone-related protein, in normal individuals and in patients with osteogenesis imperfecta. Calcif Tissue Int. 1983;35:16–20. doi: 10.1007/BF02405000. [DOI] [PubMed] [Google Scholar]
- 16.Price PA, June HH, Hamlin NJ, Williamson MK. Evidence for a serum factor that initiates the re-calcification of demineralized bone. J Biol Chem. 2004;279:19169–19180. doi: 10.1074/jbc.M307880200. [DOI] [PubMed] [Google Scholar]
- 17.Toroian D, Lim JE, Price PA. The size exclusion characteristics of type I collagen: Implications for the role of noncollagenous bone constituents in mineralization. J Biol Chem. 2007;282:22437–22447. doi: 10.1074/jbc.M700591200. [DOI] [PubMed] [Google Scholar]
- 18.Toroian D, Price PA. The essential role of fetuin in the serum-induced calcification of collagen. Calcif Tissue Int. 2008;82:116–126. doi: 10.1007/s00223-007-9085-2. [DOI] [PubMed] [Google Scholar]
- 19.Melton LJ, III, Looker AC, Shepherd JA, O'Connor MK, Achenbach SJ, Riggs BL, Khosla S. Osteoporosis assessment by whole body region vs. site-specific DXA. Osteoporos Int. 2005;16:1558–1564. doi: 10.1007/s00198-005-1871-y. [DOI] [PubMed] [Google Scholar]
- 20.Baran D, Leahey D, von Stetten E. Clinical evaluation of a fourth generation fanbeam DXA system. J Bone Miner Res. 1994;10:S374. [Google Scholar]
- 21.Fuerst T, Gluer CC, Genant H. Performance evaluation of a new bone densitometer: Hologic QDR 4500. J Am Stat Assoc. 1995;10:S370. [Google Scholar]
- 22.Levey AS, Bosch JP, Lewis JB, Greene T, Rogers N, Roth D. A more accurate method to estimate glomerular filtration rate from serum creatinine: A new prediction equation. Modification of Diet in Renal Disease Study Group. Ann Intern Med. 1999;130:461–470. doi: 10.7326/0003-4819-130-6-199903160-00002. [DOI] [PubMed] [Google Scholar]
- 23.Pahor M, Chrischilles EA, Guralnik JM, Brown SL, Wallace RB, Carbonin P. Drug data coding and analysis in epidemiologic studies. Eur J Epidemiol. 1994;10:405–411. doi: 10.1007/BF01719664. [DOI] [PubMed] [Google Scholar]
- 24.Sarnak MJ, Katz R, Stehman-Breen CO, Fried LF, Jenny NS, Psaty BM, Newman AB, Siscovick D, Shlipak MG. Cystatin C concentration as a risk factor for heart failure in older adults. Ann Intern Med. 2005;142:497–505. doi: 10.7326/0003-4819-142-7-200504050-00008. [DOI] [PubMed] [Google Scholar]
- 25.Vittinghoff E, Glidden DV, Shiboski SC, McCulloch CE. New York, NY, USA: Springer Science + Business Media; 2005. Regression Methods in Biostatistics. [Google Scholar]
- 26.Hamlin NJ, Price PA. Mineralization of decalcified bone occurs under cell culture conditions and requires bovine serum but not cells. Calcif Tissue Int. 2004;75:231–242. doi: 10.1007/s00223-004-0190-1. [DOI] [PubMed] [Google Scholar]
- 27.Price PA, Lim JE. The inhibition of calcium phosphate precipitation by fetuin is accompanied by the formation of a fetuin-mineral complex. J Biol Chem. 2003;278:22144–22152. doi: 10.1074/jbc.M300744200. [DOI] [PubMed] [Google Scholar]
- 28.Ketteler M, Bongartz P, Westenfeld R, Wildberger JE, Mahnken AH, Bohm R, Metzger T, Wanner C, Jahnen-Dechent W, Floege J. Association of low fetuin-A (AHSG) concentrations in serum with cardiovascular mortality in patients on dialysis: A cross-sectional study. Lancet. 2003;361:827–833. doi: 10.1016/S0140-6736(03)12710-9. [DOI] [PubMed] [Google Scholar]
- 29.Ruffoni D, Fratzl P, Roschger P, Klaushofer K, Weinkamer R. The bone mineralization density distribution as a fingerprint of the mineralization process. Bone. 2007;40:1308–1319. doi: 10.1016/j.bone.2007.01.012. [DOI] [PubMed] [Google Scholar]
- 30.van der Linden JC, Birkenhager-Frenkel DH, Verhaar JA, Weinans H. Trabecular bone's mechanical properties are affected by its non-uniform mineral distribution. J Biomech. 2001;34:1573–1580. doi: 10.1016/s0021-9290(01)00146-4. [DOI] [PubMed] [Google Scholar]
- 31.Johnson RC, Leopold JA, Loscalzo J. Vascular calcification: Pathobiological mechanisms and clinical implications. Circ Res. 2006;99:1044–1059. doi: 10.1161/01.RES.0000249379.55535.21. [DOI] [PubMed] [Google Scholar]
- 32.Giachelli CM, Jono S, Shioi A, Nishizawa Y, Mori K, Morii H. Vascular calcification and inorganic phosphate. Am J Kidney Dis. 2001;38:S34–S37. doi: 10.1053/ajkd.2001.27394. [DOI] [PubMed] [Google Scholar]
- 33.Reynolds JL, Skepper JN, McNair R, Kasama T, Gupta K, Weissberg PL, Jahnen-Dechent W, Shanahan CM. Multifunctional roles for serum protein fetuin-a in inhibition of human vascular smooth muscle cell calcification. J Am Soc Nephrol. 2005;16:2920–2930. doi: 10.1681/ASN.2004100895. [DOI] [PubMed] [Google Scholar]
- 34.Anderson HC. Matrix vesicles and calcification. Curr Rheumatol Rep. 2003;5:222–226. doi: 10.1007/s11926-003-0071-z. [DOI] [PubMed] [Google Scholar]
- 35.Fiore CE, Celotta G, Politi GG, Di Pino L, Castelli Z, Mangiafico RA, Signorelli SS, Pennisi P. Association of high alpha2-Heremans-Schmid glycoprotein/fetuin concentration in serum and intima-media thickness in patients with atherosclerotic vascular disease and low bone mass. Atherosclerosis. 2007;195:110–115. doi: 10.1016/j.atherosclerosis.2006.08.052. [DOI] [PubMed] [Google Scholar]
- 36.Lim P, Collet JP, Moutereau S, Guigui N, Mitchell-Heggs L, Loric S, Bernard M, Benhamed S, Montalescot G, Rande JL, Gueret P. Fetuin-A is an independent predictor of death after ST-elevation myocardial infarction. Clin Chem. 2007;53:1835–1840. doi: 10.1373/clinchem.2006.084947. [DOI] [PubMed] [Google Scholar]
- 37.Donaldson LJ, Cook A, Thomson RG. Incidence of fractures in a geographically defined population. J Epidemiol Community Health. 1990;44:241–245. doi: 10.1136/jech.44.3.241. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 38.Demetriou M, Binkert C, Sukhu B, Tenenbaum HC, Dennis JW. Fetuin/alpha2-HS glycoprotein is a transforming growth factor-beta type II receptor mimic and cytokine antagonist. J Biol Chem. 1996;271:12755–12761. doi: 10.1074/jbc.271.22.12755. [DOI] [PubMed] [Google Scholar]
- 39.Mathews ST, Singh GP, Ranalletta M, Cintron VJ, Qiang X, Goustin AS, Jen KL, Charron MJ, Jahnen-Dechent W, Grunberger G. Improved insulin sensitivity and resistance to weight gain in mice null for the Ahsg gene. Diabetes. 2002;51:2450–2458. doi: 10.2337/diabetes.51.8.2450. [DOI] [PubMed] [Google Scholar]
- 40.Ix JH, Shlipak MG, Sarnak MJ, Beck GJ, Greene T, Wang X, Kusek JW, Collins AJ, Levey AS, Menon V. Fetuin-A is not associated with mortality in chronic kidney disease. Kidney Int. 2007;72:1394–1399. doi: 10.1038/sj.ki.5002549. [DOI] [PubMed] [Google Scholar]
- 41.Ix JH, Chertow GM, Shlipak MG, Brandenburg VM, Ketteler M, Whooley MA. Association of fetuin-A with mitral annular calcification and aortic stenosis among persons with coronary heart disease: Data from the Heart and Soul Study. Circulation. 2007;115:2533–2539. doi: 10.1161/CIRCULATIONAHA.106.682450. [DOI] [PMC free article] [PubMed] [Google Scholar]