ORIGINAL ARTICLE

Vertebral bone density by quantitative computed tomographymirrors bone structure histomorphometric parametersin hemodialysis patients

Aluizio Barbosa Carvalho • Ricardo Carneiro • Graziella M. Leme •

Carlos E. Rochitte • Raul D. Santos • Marcio H. Miname • Rosa M. Moyses •

Vanda Jorgetti • Maria Eugenia F. Canziani

Received: 4 June 2012 / Accepted: 14 February 2013

� The Japanese Society for Bone and Mineral Research and Springer Japan 2013

Abstract Diagnosing low bone mass is of clinical

importance for hemodialysis (HD) patients due to its

association with fractures and cardiovascular disease. We

investigated whether bone density obtained by quantitative

computed tomography (QCT) is associated with the his-

tologically determined bone volume and microarchitecture

parameters in HD patients. Twenty-six HD patients were

studied. Bone biopsy samples were obtained from the iliac

crest and trabecular bone volume, thickness, number and

separation were evaluated by histomorphometry. Vertebral

trabecular bone density (VTBD) was evaluated by QCT.

VTBD correlated positively with trabecular bone volume

(r = 0.69, p \ 0.001), trabecular thickness (r = 0.45,

p = 0.022) and trabecular number (r = 0.62, p \ 0.001),

and negatively with trabecular separation (r = -0.50,

p \ 0.01). In the multiple linear regression analysis

adjusting for age, gender and diabetes, VTBD remained

associated with bone volume by histomorphometry

(b = 0.06; 95 % CI 0.02–0.11; p = 0.006; R2 = 0.49).

VTBD measured by QCT mirrored bone volume and

microarchitecture parameters obtained by histomorphom-

etry in HD patients.

Keywords Bone density � Quantitative computed

tomography � Bone biopsy � Histomorphometry �Hemodialysis

Introduction

Renal osteodystrophy is highly prevalent among patients

with chronic kidney disease (CKD) on hemodialysis (HD),

and its deleterious effects on morbidity and mortality have

been well-recognized [1]. Bone biopsy followed by histo-

morphometry is the gold standard method to evaluate renal

osteodystrophy, especially by the determination of the state

of bone turnover and mineralization [2]. Histomorpho-

metric analysis also allows the measurement of bone

parameters related to bone volume and microarchitecture,

considered the mirror of bone mass and quality. Histo-

morphometry has proved to be a useful tool to identify

bone mineral disturbances in the general population [3] as

well as in CKD patients undergoing HD [4]. Nevertheless,

bone biopsy is an invasive, time-consuming and expensive

method, besides requiring a specialized physician for his-

tomorphometry analysis. Hence, noninvasive techniques

have been introduced for the routine diagnosis of low bone

density, which is of relevant clinical importance for HD

patients considering its association with fractures [5] and

cardiovascular disease [6].

Dual-energy X-ray absorptiometry (DEXA) has been the

most widely used method for assessing bone density. This

method has been used for assessing bone strength and

fracture risk. However, there are a number of potential

reasons that compromise the accuracy of this method in

patients with CKD. First, many bone abnormalities (altered

microarchitecture and turnover) and changes in trabecular

bone volume and cortical thickness that often occur in

A. B. Carvalho (&) � G. M. Leme � M. E. F. Canziani

Nephrology Division, Federal University of Sao Paulo, Rua

Borges Lagoa, 960, Sao Paulo, SP 04038-002, Brazil

e-mail: aluizio@uol.com.br

R. Carneiro � C. E. Rochitte � R. D. Santos � M. H. Miname

Heart Institute (InCor), University of Sao Paulo, Sao Paulo,

Brazil

R. M. Moyses � V. Jorgetti

Nephrology Division, University of Sao Paulo,

Medical School Hospital, Sao Paulo, Brazil

123

J Bone Miner Metab

DOI 10.1007/s00774-013-0442-0

CKD [7] are not detected by DEXA, whose technique is

based on the attenuation of radiation by bones or soft tis-

sues. Second, bone size may confound bone density mea-

surements by DEXA. And, finally, arthritic conditions and

the frequent presence of aortic calcification in the face of

CKD may overestimate bone density measurements by this

method [8].

The quantitative computed tomography (QCT) method

has emerged as a superior technique with the advantage of

measuring the bone in three dimensions. Thus, bone den-

sity measurement by QCT is less dependent on bone size.

Additional advantage of QCT is the ability of distin-

guishing between trabecular and cortical bone with a

minimal radiation exposure [9]. Indeed, vertebral QCT has

a high spatial resolution, which allows precise definition of

trabecular bone without contamination of cortical bone,

osteophytes and aortic calcification. Accordingly, vertebral

QCT has been used to evaluate bone mass in the general

[10] and CKD populations [3, 6]. However, the compari-

son of bone density by QCT with the gold-standard

method of bone biopsy has been poorly reported in the

literature, particularly in patients with CKD. Therefore,

herein we aimed to test whether vertebral trabecular bone

density (VTBD) assessed by QCT is associated not only

with bone volume but also with microarchitecture

parameters evaluated by bone biopsy in CKD patients

undergoing HD.

Materials and methods

This cross-sectional study evaluated 26 clinically stable

HD patients who participated in the multicenter Brazilian

Study on Bone Mineral Disturbances in Hemodialysis,

following the inclusion criteria previously described [11].

Patients who had undergone bone biopsy and computed

tomography evaluation in an interval \3 months were

included.

The study protocol was reviewed and approved by the

local institutional ethics committee and all patients signed

the informed consent.

Biochemical parameters

Laboratory data included serum measurements of ionized

calcium, phosphorus, alkaline phosphatase (reference ran-

ges \270 U/L for male, \240 U/L for female), intact

parathyroid hormone (PTH) (chemiluminescence, DPC,

Medlab, USA, reference ranges 10–65 pg/mL) and

25-hydroxyvitamin D [25(OH)D] (chemiluminescence,

DiaSorin, Minnesota, USA, reference range 18–62 ng/mL).

Bone biopsy and histomorphometry

Bone biopsies were carried out in either the right or left

iliac crest, using a 7 mm inner diameter electrical trephine

(Gauthier Medical, Rochester, MN, USA). Bone fragments

were submitted to the usual processing and histological

studies [12]. Sections were stained with toluidine blue

staining. Bone histomorphometry was conducted using the

semi-automatic method contained in the software Oste-

omesure (Osteometrics Inc., Atlanta, GA, USA). Histo-

morphometric structure parameters of trabecular bone

volume, thickness, separation and number were analyzed in

accordance with the standards of the American Society of

Bone and Mineral Research [13].

Histomorphometric results of static parameters were

compared to gender- and age-matched controls selected

from a large bone histomorphometry database compiled

from the analyses of iliac biopsies of healthy individuals

submitted to autopsy after an early demise, as follows:

bone volume (normal values = 24.0 ± 6.1 % for male and

21.8 ± 7.2 % for female), trabecular thickness (normal

values = 127.9 ± 29.7 lm for male and 126.0 ± 28.8 lm

for female), trabecular separation (420.6 ± 124.1 lm for

male and 498.3 ± 195.9 lm for female) and trabecular

number (1.89 ± 0.42 n/mm for male and 1.76 ± 0.52 n/

mm for female) [14].

Quantitative computed tomography

QCT analysis was performed in a thoracic vertebral image

obtained in a 16-detector-row scanner (Somatron Volume

Zoom Siemens AG, Erlhagen, Germany). VTBD was

measured within a region of interest placed at the mid-

vertebral body (Fig. 1). The vertebral body chosen was the

one located at the level of the largest heart area on the axial

images. VTBD was resulted by the attenuation of vertebral

bone expressed in Hounsfield Units (HU) [15]. Analyses

were performed in a Vitrea 2 workstation (Vital Images

Inc., Plymouth, MN). In order to evaluate the interobserver

error for VTBD measurement two observers blinded to the

clinical data performed QCT. The interobserver variability

of the QCT technique was evaluated in the present study.

Bland and Altman analysis showed a low error of

4.8 ± 14.7 HU (p = 0.11) and the limits of agreement

were -24.6 and 34.2 HU.

Statistical analysis

Data were shown as mean and standard deviation or pro-

portions, as appropriated. Data normality was tested by the

Shapiro–Wilk test. The relationship between VTBD and

J Bone Miner Metab

123

bone biopsy parameters was investigated by linear corre-

lation with Pearson coefficient test. Multiple linear

regression analysis was applied to evaluate potential

independent association between VTBD and bone volume.

The predictability of bone volume by VTBD was also

assessed by the receiver operating characteristic (ROC)

curve analysis, using the reference for bone volume [14] as

the cut-off point. A p value \0.05 was considered signifi-

cant. All statistical analyses were performed using SPSS

version 15 (SPSS Inc, Chicago, IL) and STATA 8.0

(StataCorp, College Station, TX).

Results

Table 1 depicts the demographic, clinical, bone biopsy and

QCT data of the 26 patients enrolled in the present study.

Patients were relatively young and 15 % had diabetes. The

length on HD therapy of the patients varied from 4 to

103 months. Hypertension was the main cause of CKD

(69 %). Thirty-eight percent of the patients were over-

weight or obese (BMI C25 kg/m2), and only one patient

had BMI lower than 18 kg/m2.

Trabecular bone volume, thickness and number were

decreased and trabecular separation was increased in our

HD patients when compared to the reference values

(Table 1). Abnormalities of bone volume were observed in

58 % of the patients, of trabecular thickness in 62 %, of

trabecular number in 77 % and of trabecular separation in

46 %. Strong correlations were found between VTBD by

QCT and the histomorphometric parameters. As can be

seen in Fig. 2, VTBD correlated positively with trabecular

bone volume (r = 0.69, p \ 0.001), trabecular thickness

(r = 0.45, p = 0.022) and trabecular number (r = 0.62,

p \ 0.001), and negatively with trabecular separation

(r = -0.50, p \ 0.010). In the multiple linear regression

analysis adjusting for age, gender and diabetes, QCT

derived VTBD was independently associated with bone

volume assessed by histomorphometry (b = 0.06; 95 % CI

0.02–0.11; p = 0.006; R2 = 0.49). In addition, the ROC

curve analysis revealed VTBD as predictor of bone volume

(area under the curve 0.76; Fig. 3).

Discussion

This study showed that VTBD by QCT reflects bone vol-

ume and microarchitecture parameters determined by bone

biopsy in CKD patients undergoing HD. Considering the

unacceptably high prevalence of histological and structural

bone abnormalities in CKD patients, finding a noninvasive,

Fig. 1 Quantitative computed tomography images of the thoracic vertebral body (a). The white circle depicts the tissue attenuation data

measured by the computer software in the region of interest (trabecular bone) chosen for analysis (b)

Table 1 Demographic, clinical and bone characteristics of the

hemodialysis patients

N = 26

Age (years) 45.9 ± 13.7

Male (%) 53

White (%) 46

Length on hemodialysis (months) 38.2 ± 30.2

Body mass index (kg/m2) 23.7 ± 4.3

Ionized calcium (mmol/L) 1.24 ± 0.1

Serum phosphorus (mg/dL) 6.5 ± 1.9

Alkaline phosphatase (U/L) 294.1 ± 226.2

Parathyroid hormone (pg/mL) 413.5 ± 353.7

25-hydroxyvitamin D (ng/dL) 33.1 ± 19.2

Bone biopsy

Trabecular bone volume (%) 20.3 ± 8.8

Trabecular thickness (lm) 120 ± 21.5

Trabecular number (n/mm) 1.63 ± 0.52

Trabecular separation (lm) 569.4 ± 317

Quantitative computed tomography

Vertebral trabecular bone density (HU) 190.1 ± 88.9

Mean ± standard deviation or proportions

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123

less time-consuming and cost-effective method able to

mirror bone volume and microarchitecture abnormalities

obtained by bone biopsy is of relevant importance for

clinical and research purposes in the management of CKD

patients.

Computed tomography technology has undergone

innovations since the advent of QCT in the 1980s. QCT

provides higher spatial resolution and superior image

quality compared with earlier computed tomography sys-

tems. However, at present, a limited number of studies

have used QCT method in CKD population. In the present

study, loss of trabecular bone volume and derangements of

microarchitecture were observed, which is in accordance

with previous bone biopsy findings in HD patients [4]. To

our knowledge, the current study is the first to demonstrate

the association of VTBD by QCT with histomorphometric

parameters in CKD.

We recognized that the findings of this study are limited

to a relatively young and asymptomatic cohort of HD

patients with low frequency of diabetes. However, the

present finding of a good conformity between VTBD

measured by QCT in the thoracic spine and bone volume

and structural parameters obtained by bone biopsy in HD

patients draws attention to the feasibility of measuring

VTBD in studies of coronary calcification. This might

allow simultaneous assessment of cardiovascular disease

and bone loss. In accordance, a previous study by Lenchik

et al. [10] has demonstrated that cardiac gated computed

tomography acquisition coupled with a QCT phantom is

useful to assess thoracic bone density. The authors found a

high correlation (r = 0.93) between trabecular thoracic

bone density and trabecular lumbar bone density by QCT,

considered this latter the place of choice for monitoring

Fig. 2 Correlations between vertebral trabecular bone density (VTBD) by quantitative computed tomography and histomorphometric structure

parameters by bone biopsy in hemodialysis patients

Fig. 3 ROC curve analysis of vertebral trabecular bone density

(VTBD) by quantitative computed tomography and bone volume by

bone biopsy in hemodialysis patients

J Bone Miner Metab

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age-, disease- and treatment-related bone density changes

by the International Society for Clinical Densitometry

Official Position [16]. Recently, Tamminen et al. [17]

comparing micro-CT and bone histomorphometry in bone

samples from healthy subjects and patients with osteopo-

rosis or renal osteodystrophy found that micro-CT was an

effective method to determine structural bone changes.

In conclusion, we demonstrated that VTBD assessed by

QCT mirrors the bone volume and structure histomorpho-

metric parameters in HD patients. Prospective studies are

warranted to investigate the predictive power of QCT to

detect overtime changes in histomorphometry as well as to

evaluate the impact of QCT derived bone density on clin-

ical outcomes such as fractures and cardiovascular

diseases.

Acknowledgments We are indebted to Maria Ayako Kamimura for

reviewing and improving this manuscript.

Conflict of interest All authors have no conflicts of interest.

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