Assessment of bone turnover and bone quality in type 2 diabetic bone disease:current concepts and future directions

Mishaela R Rubinand Janina M Patsch

Assessment of bone turnover and bone quality in type 2 diabetic bone disease:current concepts and future directions

Mishaela R Rubin1and Janina M Patsch2

Substantial evidence exists that in addition to the well-known complications of diabetes,increased fracture risk is an important morbidity.This risk is probably due to altered bone properties in diabetes.Circulating biochemical markers of bone turnover have been found to be decreased in type 2 diabetes(T2D)and may be predictive of fractures independently of bone mineral density(BMD).Serum sclerostin levels have been found to be increased in T2D and appear to be predictive of fracture risk independent of BMD.Bone imaging technologies,including trabecular bone score(TBS)and quantitative CT testing have revealed differences in diabetic bone as compared to non-diabetic individuals.Speci f i cally,high resolution peripheral quantitative CT(HRpQCT)imaging has demonstrated increased cortical porosity in diabetic postmenopausal women. Other factors such as bone marrow fat saturation and advanced glycation endproduct(AGE)accumulation might also relate to bone cell function and fracture risk in diabetes.These data have increased our understanding of how T2D adversely impacts both bone metabolism and fracture risk.

INTRODUCTION

Type 2 diabetes mellitus(T2D)is an exceedingly common chronic metabolic disorder that has an enormous impact on public health.Currently,diabetes affects over 387 million adults worldwide and is projected to reach 592 million by 2035.1Until recently,the list of target organs affected by T2D did not include the skeleton.Yet it is now well-established that T2D is an independent risk factor for fractures,which is not attributable to increased body mass index(BMI)nor other classical osteoporosis risk factors.2New data from epidemiologic and pathophysiologic reports,as well as from studies employing state-of theart investigational tools,have recently increased our understanding of how T2D adversely impacts both bone metabolism and fracture risk.

Epidemiological data indicate that older adults with T2D have a higher risk of fractures,with a 50%–80%increased extremity fracture risk.3–4A meta-analysis of 12 studies reported a relative risk of 1.7(95%con f i dence interval: 1.3–2.2)for hip fracture in both men and women with T2D.5The risk of all clinical fractures was also increased,with a summary relative risk of 1.2(95%con f i dence interval:1.0–1.5).5Other studies have reported similar results,6with a direct association between the duration of diabetes and increased fracture risk.7Given this increased fracture risk,it is perhaps surprising that bone mineral density(BMD)is generally higher in those with T2D compared with those without.8In a meta-analysis,Vestergaard et al8reported an increased Z-score of+0.41 at the spine and+0.27 at the hip associated with T2D.In addition,a large prospective study has shown that patients with T2D have a higher fracture risk for a given femoral neck BMD T-score.9Although dual-energy x-ray absorptiometry(DXA)is the gold standard method for the quantitative assessment of BMD,providing areal BMD of the hip,spine,radius,and total body(including body composition),it has limitations in patients with complex metabolic bone diseases such as chronic kidney disease or diabetic bone disease.It hastherefore been suggested that fragility fractures in T2D may result from diabetes-related alterations in skeletal properties not captured by DXA.2Similarly,patients with T2D have a higher fracture risk for a given FRAX probability(https:// www.shef.ac.uk/FRAX/).9The FRAX algorithm provides the 10-year probability of major osteoporotic fractures per individual tested.A recent study has demonstrated that diabetes mellitus does not modify the effects of risk factors incorporated into FRAX.10Nevertheless,the study has shown that diabetes has stronger effects on hip fracture risk in younger than older individuals which warrants special consideration for diabetic fracture prevention in clinical practise.

The paradox of higher BMD in association with increased fractures might be attributed to more frequent trauma,as diabetes is associated with an increased frequency of falls.11However,in studies of diabetes and fracture that controlled for fall frequency,diabetes still remained independently associated with increased fracture risk.3–4Thiazolidinediones use might also be considered as an explanation,since it has been proposed that these agents divert mesenchymal stem cells from the osteogenic to the adipocytic lineage and are associated with bone loss and increased fracture risk,particularly in women.12However,thiazolidinedione use cannot fully account for the increased risk of fracture observed with diabetes,since most studies included substantial observation time prior to the widespread use of these medications.Rather,it appears that other bone properties,which are undetectable by DXA,are probably contributing to fracture risk in diabetes.

PTH AND BIOCHEMICAL MARKERS OF BONE TURNOVER IN T2D

Decreased bone remodeling in T2D has been demonstrated by a number of lines of evidence.Levels of parathyroid hormone(PTH)tend to be 20%–50%lower in T2D subjects than in controls,even in the setting of reduced eGFR,suggesting a state of reduced PTH secretion in T2D.13–15Circulating biochemical markers of bone formation,including P1NP,osteocalcin(OCN)14–15and bone speci f i c alkaline phosphatase16have been found to be decreased in T2D.These decreases in formation measures are associated with reductions in the bone resorption marker serum CTx.13–16The decrease in bone remodeling in T2D appears to be predictive of fracture risk regardless of BMD.In a study of 255 T2D women and 240 controls,T2D women with the combination of the lowest PTH and OCN levels had nearly a f i vefold increased risk of vertebral fractures independent of lumbar spine BMD.15

DYNAMIC HISTOMORPHOMETRY IN T2D

Lower bone formation in T2D on biopsy was reported in one study,but the numbers were very small(n=6 T2D patients; 2 female),and the results were confounded by selecting for low BMD and a problematical control group.17In a more recent pilot study,low-bone formation was observed in six T2D postmenopausal women and six postmenopausal age-matched non-diabetic controls,where tetracycline double-labeled iliac crest bone biopsies showed virtually no uptake of label in diabetic subjects(Figure 1),with reduced mineralizing surface,osteoid surface,and osteoblast surface.18Interestingly,corresponding reductions in bone resorption indices were not present,perhaps suggesting a disproportionate reduction in bone formation in diabetes as compared with bone resorption.

OTHER BONE MARKERS IN T2D

Insulin-like growth factor(IGF)-1,an anabolic factor which stimulates osteoblast proliferation,has been inversely associated with the risk and number of vertebral fractures in diabetic women independent of BMD.14–19Another marker which might re f l ect bone formation is that of circulating osteogenic precursor cells,20which have been reported to be decreased in patients with T2D(Figure 2).Circulating osteogenic precursor cells can be detected in the peripheral blood by f l ow cytometry using antibodies speci f i c for the osteoblast matrix protein OCN.21Peripheral blood mononuclear cells that were positive for OCN were lower in postmenopausal women with T2D as compared with non-diabetic controls.18Moreover,within the decreased pool of overall OCN+cells,the T2D subjects had an increased subpopulation of immature OCN+cells, that is,cells that also had early markers CD146 and CD34, subpopulations which diminish when osteoblasts mature.20An additional novel bone marker in T2D may be sphingosine 1-phosphate(S1P),a lipid mediator which increases osteoclastogenesis by increasing RANKL.22S1P was found to be increased in T2D women(n=482)as compared with controls and was associated with vertebral fractures.Interestingly,this marker suggests an elevation in bone resorption in T2D,in contrast to the reports of decreased s-CTx levels.13–16It is possible that s-CTx underestimates the level of bone resorption in diabetes because enzymatic cross-linking of bone collagen by lysyl oxidase is reduced in diabetes,23–24such that less cross-linked telopeptides might be released in diabetes during bone resorption.

SCLEROSTIN IN T2D

Sclerostin,an osteocyte product,is a negative regulator of bone formation which competes with the anabolic Wnt β-catenin pathway by binding to LRP5 or 6.25In healthy adults,sclerostin levels are increased by factors including age,BMI,inactivity,bone mineral content,and possibly fractures.25It was f i rst reported in 2012 that sclerostin levels were higher in 74 T2D women and men versus 50 nondiabetic controls and that higher levels correlated with age,male gender and BMD.26This observation was corroborated by another report in which sclerostin levels were found to be twofold higher in T2D than in controls or T1D,after adjusting for age and BMI.27A correlation between Wnt disruption and decreased osteoblast activity was further observed in 40 T2D postmenopausal women who,as compared with controls,had decreased β-catenin levels which correlated with lower BAP.16In the largest diabetes sclerostin study,higher sclerostin levels in 321 men and women with T2D were associated with an increased risk of vertebral fractures independent of lumbar spine BMD.28Interestingly,diabetic postmenopausal women with fragility fractures were shown to demonstrate signi f icantly higher serum sclerostin levels than diabetic postmenopausal women without fragility fractures.29While BMI, renal function,glycemic control,and diabetes medication (including insulin)were comparable between women with and without fragility fractures,diabetes duration was signi f i cantly longer in those that had sustained fractures.It could be posited from these data that the higher sclerostin levels in T2D re f l ect the presence of more deeply embedded osteocytes in older bone that has accumulated more microscopic damage.Thus prolonged low-bone turnover in diabetes,as a result of Wnt inhibition, may lead to defective microdamage repair and increased bone microcrack accumulation in a manner reminiscent of high-dose bisphosphonate therapy,30thus contributing to greater bone fragility.Stressing the interplay between bone health and vascular health and considering diabetic fractures to be true diabetic complications,it is also noteworthy that in diabetics higher serum sclerostin levels appear to be associated with higher amounts of vascular calci f i cations.31

AGES AND BONE REMODELING

Decreased bone formation might occur in part because of increased advanced glycation endproducts(AGEs)in bone collagen.AGEs are a diverse group of compounds that are generated through the non-enzymatic glycation or glycoxidation of proteins,lipids,and nucleic acids32with the best-studied being carboxymethyl-lysine and pentosidine.33–36These compounds are markedly increased in patients with diabetes,35forming non-enzymatic cross-links within and across collagen f i bers.37–38AGEs interfere with normal osteoblast function39and attachment to the collagen matrix,40as well as impair osteoblast development.41–42AGEs also decrease bone resorption by altering the structural integrity of bone matrix proteins and inhibiting the osteoclastic differentiation process.43This might have long-lasting skeletal effects that are similar to the“hyperglycemic metabolic memory”that has been described with AGE accumulation in other tissues.44In the Diabetes Control and Complications Trial(DCCT), accumulation of AGEs in skin collagen of type 1 diabetes patients predicted complications decades later,regardless of subsequent improvements in glycemic control.45In the bone matrix,accumulation of AGEs leads to more biomechanically brittle bone that has lost its toughness and is less able to deform before fracturing.30Urinary pentosidine,the best-studied AGE,was associated with a 42% increase in clinical fracture incidence in T2D.38Although the relationship between in vivo bone and circulating levels of AGEs has not been fully elucidated,AGEs are likely related to both low-bone formation and increased bone fragility in T2D.

ASSESSMENT OF FRACTURE RISK IN T2D:FROM CURRENT CLINICAL PRACTICE TO NOVEL IMAGING BIOMARKERS

Irrespective of diabetes history,in clinical practice fracture risk is routinely assessed by measuring BMD and bydetermining the presence of clinical risk factors including age,sex,BMI,smoking and drinking habits,history of fragility fractures,parental fractures,rheumatoid arthritis, and known secondary osteoporosis.These factors are part of the aforementioned,questionnaire-based fracture risk assessment tool(FRAX).46

BMD can be measured by DXA and quantitative computed tomography(QCT)at central and peripheral sites.47Although DXA can predict fracture risk in diabetic patients to a certain extent,experts have suggested the introduction of a“diabetic correction factor”for T-scores because given T-scores were shown to be associated with a higher risk of fracture in older adults with T2D than in those without DM.48Similar concepts have been suggested for FRAX scores in elderly subjects with T2D.48The use of trabecular bone score(TBS)has been also suggested to improve the diagnostic performance of lumbar spine DXA in patients with T2D:from an image processing perspective,TBS is based on two-dimensional texture-analysis of DXA images.Applying methodologies originally used in pattern recognition(for example,in geostatistics characterizing landscapes from aerial views), textural variations are quanti f i ed between neighboring gray-scale pixels.49In clinical practise,operators and clinicians are only presented with the end-result of these calculations:A single(TBS)index which is higher in images with fewer inter-pixel variations(that is,indirect measure of better spinal bone microarchitecture)and lower in images with higher inter-pixel variations(that is,indirect measure of worse spinal bone microarchitecture).50While BMD is typically normal to higher in subjects with diabetes,TBS appears to be lower in diabetics than in non-diabetic subjects.51In the same publication,Leslie et al.51also showed that lumbar spine TBS can predict osteoporotic fractures irrespective of the presence or absence of diabetes.In addition,TBS has been shown to be positively associated with good glycemic control.52–53While these results are of major clinical interest and relevance,it remains to be stressed that—from an imaging standpoint—TBS only provides an indirect measure of bone microarchitecture at low resolution and reduced image quality.

While DXA is a projectional technique,central QCT provides volumetric data on BMD and bone geometry of the spine and the proximal femur.For QCT,a clinical multidetector-CT scanner,a calibration phantom mat, and dedicated software are needed.54Alterations in volumetric hip BMD—as determined by QCT in diabetic versus non-diabetic subjects—have been con f i rmed by two independent studies.29,53–55One of these studies also concluded that patients with T2D appear to have little bene f i t from elevated BMD due to unchanged load-tostrength ratios.55

HIGH-RESOLUTION PERIPHERAL QCT

Within the last two decades,dedicated peripheral QCT scanners have been developed to capture volumetric BMD of peripheral sites(that is,the extremities)at varying resolution.In addition to BMD and geometric measures, high-resolution peripheral QCT(HR-pQCT)provides quantitative access to bone microarchitecture,an important surrogate of bone quality and bone strength.56–57Many micro-architectural parameters quanti f i able by HR-pQCT refer to those initially coined by static histomorphometry (for example,trabecular number,trabecular thickness).58In the f i rst generation HR-pQCT scanner,which has a spatial resolution of～80μm,only two microstructural parameters—trabecular number and trabecular heterogeneity—are directly measured by three-dimensional distance transformations.59Trabecular separation and trabecular thickness are derived from trabecular BMD and trabecular number,which are determined by threshold-based segmentation and a ridge-counting technique.60Recently,a second generation scanner with shorter scan time and higher spatial resolution has been introduced.Using this second generation scanner,all micro-architectural indices can be calculated directly.61It is noteworthy that so far,all published HR-pQCT data on bone microarchitecture in T2D have been acquired with scanners of the f i rst generation.62–65

First evidence of associations between impaired cortical strength and T2D originally came from a pQCT study in a cohort of elderly men.66The f i rst HR-pQCT study investigating bone microarchitecture in T2D then raised the topic of high cortical porosity as a potential predominant structural phenotype:62while trabecular BMD and trabecular thickness were greater in postmenopausal women with T2D than in those without,high cortical porosity was observed (Figure 2).Based on the unstrati f i ed enrollment of diabetic women with and without fractures,this study lead to the formation of a key hypothesis for subsequent research: could cortical porosity serve as a discriminative feature between diabetic women with and without fragility fractures?In line with this hypothesis,Shu et al,63also reported no differences in peripheral bone microarchitecture between postmenopausal women with and without T2D(without history of fragility fractures).Speci f i cally enrolling postmenopausal diabetic women with and without fragility fractures and comparing them with nondiabetic controls with and without fractures,the importance of high cortical porosity with relatively maintained trabecular properties could be con f i rmed.64Interestingly, diabetic women with and without fractures were comparable with respect to clinical characteristics:age,BMI, glycemic control,25(OH)-vitamin D,kidney function,and PTH levels were similar.The only signi f i cant clinical difference was the duration of diabetes:diabetic women withfractures had been suffering from diabetes signi f i cantly longer than diabetic women without fractures(13 years versus 8 years).Using f i nite element analyses,de f i cits in stiffness,failure load,and cortical load fraction could be detected and attributed to cortical porosity.64Considering the high prevalence of insulin resistance and T2D in the African-American population,a forth HR-pQCT study speci f i cally focused on the assessment of bone microarchitecture in postmenopausal African-American women:In line with above mentionned studies of mixed racial recruitment,Yu et al.65found T2D and unfavorable cortical bone microarchitecture to be similarly associated in African-American women.

MAGNETIC RESONANCE IMAGING

Magnetic resonance imaging(MRI)provides an alternative method of depicting and quantifying bone geometry and bone microarchitecture.47High-resolution MRI has been used to study bone microarchitecture in postmenopausal and age-related osteoporosis,67–69secondary osteoporosis,70chronic kidney disease,71and with respect to treatment effects of anti-osteoporotic drugs.72–73Regarding diabetic bone disease,there is only a limited number of publications using MRI.Speci f i cally,a Canadian researcher reported poorer bone microarchitecture expressed as greater trabecular heterogeneity in subjects with diabetes mellitus than in healthy controls.74Although compared with HR-pQCT,MRI has certain disadvantages in studying bone microarchitecture(for example,lower resolution,higher technical complexity in acquisition and postprocessing of image data,no option of measuring BMD,and fewer reference data),the method does hold signi f i cant potential for bone research.Speci f i cally,MRI offers the option of imaging the skeleton beyond its geometry and structure.Using techniques such as magnetic resonance-perfusion and MR-spectrocopy,novel imaging surrogates of bone strength(for example,skeletal blood f l ow and regional biochemical composition of bone marrow)have been introduced.75–76Recent MR-imaging data have demonstrated that altered bone marrow fat composition is linked with T2D and fragility fractures in postmenopausal women.77Even after adjustment for age, race,and local(that is,spinal)BMD,lower unsaturated bone marrow fat and higher saturated bone marrow fat as determined by MR-spectroscopy were signi f i cantly associated with T2D.Of interest,postmenopausal diabetic women with a history of fragility fractures displayed the lowest unsaturation and the highest saturation levels.The relevance of these f i ndings is supported by epidemiologic data linking low dietary intake of n-3 poly-unsaturated fatty acids(PUFA)with low BMD78–80and the risk of hip fractures.81Preclinical data support a potential osteoprotective role of PUFA.82–85Vice versa,saturated dietary fatty acids appear to be linked with osteoporotic fractures.86With poor glycemic control also leading to accelerated aging of adipocytes and thus adipocyte dysfunction,87–90changes in bone marrow fat composition of subjects with T2D are likely to result from a multitude of interacting etiologic factors.It is conceivable that bonemarrow adiposity might re f l ect a shift in stem cell lineage away from osteoblastogenesis toward adipogenesis.91In the AGEs–Reykjavik cohort(115 men and 134 women; mean age 79;7%with diabetes),sclerostin levels were positively associated with marrow fat by MRI in men independent of BMD.91This positive relationship could be explained by a shift in precursor stem cell lineage away from osteoblastogenesis,as re f l ected by higher sclerostin levels,toward adipogenesis,as re f l ected by higher bone marrow fat.91

OBESITY AS A POTENTIAL CONFOUNDER

Confounding greater understanding of the mechanisms responsible for increased skeletal fragility in T2D is the frequent concurrence of obesity,which has also recently been shown to be a risk factor for fractures.92–94Mechanisms proposed to be responsible for increased skeletal fragility in both obesity and T2D include structural,metabolic,material,dynamic,and imaging abnormalities. However,the relative contribution of each condition separately and their combined effects on skeletal fragility remain unclear.The concurrence of obesity and T2D in most individuals has undermined attempts to gain pathogenetic clarity.For example,obese women have been reported to have trabecular and cortical microarchitectural abnormalities,but their diabetic status was not considered.95Moreover,obese diabetics have lower biochemical and quantitative histomorphometric indices of bone formation,17–96elevated circulating levels of the osteocyte product sclerostin,27increased adipocyte markers,97and abnormal bone marrow fat composition,77however,these abnormalities are also found in obese individuals without T2D.98–101Whether T2D has negative effects on speci f i c skeletal parameters that are distinct from the adverse skeletal effects of obesity remains to be clari f i ed.

A CONTINUING QUEST FOR NOVEL NON-INVASIVE BIOMARKERS REFLECTING HIGH FRACTURE RISK IN PATIENTS WITH T2D

Minimal invasive testing of bone strength(“micro-indentation”)is nowadays possible in the setting of clinical research.After local anesthesia at the mid-shaft point of the anterior tibia,a handheld device is inserted through soft tissue and periosteum,to touch the bone surface. Using triggered,pre-de f i ned impacts small bony indentations(that is,local super f i cial microfractures)are created (＜350 μm).The deeper the mean indentation distance, the weaker bone material properties are at the testing site of an individual subject.102–103Using the most recent generation in vivo testing device,Farr et al.104have shown that postmenopausal diabetic women had signi f i cantly lower bone material strength than non-diabetic controls. Poor bone material strength correlated with poor long-term glycemic control over the past 10 years.104In vivo micro-indentation has been used to quantify reduced bone strength in patients with osteoporotic hip fractures and atypical femoral fractures.105–106To the best of our knowledge,in vivo micro-indentation data are still lacking for patients with T1D,men,and patients with fragility fractures.

Given signi f i cant advances in knowledge within the last decade,dedicated research in diabetic bone disease with and without fractures and development of novel patient-oriented biomarkers continue to be warranted.

S UMMARY AND CONCLUSION

There is growing evidence that the skeleton is an important target organ for complications of T2D.Many large prospective studies have established that T2D is associated with an increased risk of fractures.Yet it remains unknown how to identify which patients with T2D are at increased risk because it is uncertain why bone fragility is increased when aBMD is normal.Available evidence suggests that compromised bone quality,not de f i cits in aBMD,is the underlying basis for fragility fractures in patients with T2D.Clues supporting this hypothesis include alterations in bone remodeling,increased cortical porosity,and decreased bone material properties.More information is needed about the pathogenesis of these abnormalities and their relationship to the increased fracture risk observed in T2D. Whether a circulating marker,such as an AGE or sclerostin, would be predictive of alterations in bone material properties or cortical porosity remains to be determined. Further investigation of skeletal parameters would shed light on the issue of greater bone fragility in T2D and potentially offset serious challenges in this population as they age.

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©The Author(s)2016

Research(2016)4,16001;

10.1038/boneres.2016.1;Published online:22 March 2016

1Metabolic Bone Disease Unit,Department of Medicine,Columbia University College of P&S,New York,USA and2Division of General Radiology and Pediatric Radiology,Department of Biomedical Imaging and Image-Guided Therapy,Medical University of Vienna,Vienna,Austria

Correspondence:Janina M Patsch(janina.patsch@meduniwien.ac.at)

Received:27 November 2015;Accepted:21 December 2015