Laboratory Evaluation of Osteoporosis in Women

Case: A 65 y/o woman steps off of a curb, falls and severe pain develops in her left hip. After a 911 call, EMS brings the woman to the local ED. What is the most likely underlying case of this clinical scenario?

William E. Winter, MD University of Florida Departments of Pathology and Pediatrics 2-15-2015 Learning objectives: At the conclusion of this presentation, the laboratorian will be able to: 1. Describe the circumstances when the diagnosis of osteoporosis should be considered. 2. List the causes of pathological fractures. 3. Define osteoporosis. 4. Discuss the causes of osteoporosis. 5. Interpret the results of laboratory testing that is conducted during the evaluation of pt’s w/ possible osteoporosis. 6. Interpret bone marker testing results. 7. Understand the controversy that relates to the value of bone marker testing in the evaluation of osteoporosis and its therapy. Under what clinical circumstances is the diagnosis of osteoporosis considered in women? Women w/: (1) Loss in height, kyphosis, other acquired skeletal deformities. (2) Low BMD on DEXA scan. (3) Fractures.

Fracture

Pathologic Traumatic

(“Routine” Force > bone strength) (“Exceptional” Force > bone strength)

Stepping off a curb Falling off of a tall ladder ‐ > hip fracture ‐ > hip fracture

BMD = bone mineral density What are the causes of pathologic fractures?

Focal bone disease Neoplasms ‐ Malignant ‐ Benign

Non‐neoplastic disease (e.g., bone cyst, osteomyelitis; dysostoses, osteonecrosis)

Generalized bone disease Congenital disorders (e.g., dysplasias)

Acquired disorders

See: http://www.wheelessonline.com/ortho/pathologic_fracture What are examples of neoplasms that can cause pathologic fractures? Malignant ‐ Bone cancers ‐ Malignant fibrous histiocytoma (a.k.a. ‐ fibrous histiocytoma pleomorphic sarcoma) ‐ Osteosarcoma ‐ Malignant giant cell tumor of bone

‐ Non‐bone cancers (e.g., multiple myeloma)

‐ Metastasis (e.g., breast, lung, kidney, prostate, thyroid)

Benign ‐ Benign fibrous histiocytoma ‐ Nonossifying fibroma ‐ Benign giant cell tumor of bone What are the causes of pathologic fractures?

Focal bone disease Neoplasms ‐ Malignant ‐ Benign

Non‐neoplastic disease (e.g., bone cyst, osteomyelitis; dysostoses)

Generalized bone disease Congenital disorders (e.g., dysplasias)

Acquired disorders

See: http://www.wheelessonline.com/ortho/pathologic_fracture What are examples of dysplasias that can cause pathologic fractures? Dysplasia: congenital, global disorders of abnormal tissue development affecting cartilage and bone.

Examples Etiologies Achondroplasia GOF mutation in FGFR3

Thanatophoric dysplasia “ “ “

Osteopetrosis LOF in osteoclasts

Osteogenesis imperfecta Mutations in alpha 1 or alpha 2 chains that constitute type 1 collagen In DD of osteoporosis

McCune‐Albright syn. LOF in GS ATPase What are the causes of pathologic fractures?

Focal bone disease Neoplasms ‐ Malignant ‐ Benign

Non‐neoplastic disease (e.g., bone cyst, osteomyelitis; dysostoses)

Generalized bone disease Congenital disorders (e.g., dysplasias)

Acquired disorders

See: http://www.wheelessonline.com/ortho/pathologic_fracture Besides osteoporosis, what are examples of generalized, “acquired” disorders that can cause pathologic fractures? Examples Etiologies Rickets / Osteomalacia Decr. vit D activity; hypophosphatemia

10 hyperparathyroidism Excess PTH activity In DD of osteoporosis Renal osteodystrophy Vit D def., hyperphosphatemia, 20 hyperparathyroidism Paget disease of bone Excess bone turnover

Humoral hypercalcemia of Malignancy: PTHrP

Scurvy Dietary vitamin C def.

Gaucher disease Inborn error of sphingomyelin metab. What is osteoporosis?

NIH Consensus Statement, Vol 17 (1), March 27-29, 2000

“. . . . (A) skeletal disorder characterized by compromised bone strength predisposing to an increased risk of fracture.”

Eponym: “Porous bone disease”

Consequences Loss in height, collapsed of reduced bone vertebra, lumbar lordosis; strength: kyphosis; scoliosis; factures & associated mortality

Most common fractures: Hip, spine, and wrist 11 What determines bone strength?

Bone density - gm/area or volume of bone - Bone mineral density = ~70% of bone strength Bone quality - Architecture Vertebral bodies - Bone turnover - Damage accumulation - Mineralization

Young Old

Note: Can not measure bone strength directly Can measure bone density (DEXA scan) 12 What is the biology of osteoporosis in adults?

Following attainment of maximum adult BMD, over time:

Osteoclast activity > Osteoblast activity Resorb bone Build bone Outcome: - Reduced bone density (= osteopenia) - Disordered microarchitecture

Normal Osteoporosis

Scanning electron microscopy of 3rd lumbar vertebra. What cells build bone and which cells resorb bone?

Function Origin

Osteoblasts Build bone Mesoderm

Osteoclasts Resorb bone BM stem cell What is the cycle of normal bone turnover?

B OSTEOCLAST (resorption) OSTEOBLAST (formation)

“Basic multicellular unit”* A C

* includes osteocytes

E D

15 What regulates osteoclasts?

Stromal (osteoprogenitor) cell Osteoblast Intermittent PTH A mesenchymal cell

HSC

M‐CSF

OPG Monocyte RANKL Resting osteoclast Precursor RANK fusion

Macrophage Osteoclast precursor 16 M-CSF = macrophage colony stimulating factor; OPG = osteoprotegerin; RANK = receptor activator of nuclear factor NF kappaB NOTE: Normal pulses of PTH (“intermittent” secretion) from the parathyroid gland stimulate the development of osteoblasts from stromal cells.

Stimulated osteoblasts secrete macrophage colony‐stimulating factor (M‐CSF). This stimulates the replication of macrophages into osteoclast precursors.

Osteoblasts also express on their surface osteoclast differentiating factor (ODF) which stimulate osteoclast precursors differentiation into osteoclasts. ODF is also known as receptor activator of NF kappaB ligand (RANKL) and osteoprotegerin‐ligand (OPGL).

Osteoprotegerin (OPG), secreted by osteoblasts, antagonizes the effects of RANKL. OPG serves as a soluble decoy receptor for RANKL antagonizing RANKL’s ability to bind to its receptor which is RANK on osteoclast precursors.

17 Therefore: Osteoblasts regulate osteoclasts

Inhibition Osteoblast of osteoclast activation RANKL (incr. expression w/ IL‐1, IL‐6, IL‐11, TNF) Osteoprotegeren

Stimulation

RANK of osteoclast activation; Competition precursor Osteoclast fusion Resting precursor osteoclast

18 See fig 26‐3, Robbins OSTEOBLAST REGULATION OF OSTEOCLASTS

Inhibition of osteoclast activation

RANKL = receptor activator of NF‐kappaB ligand Osteoblast ‐ member of TNF superfamily of ligands & receptors ‐ essential for: osteoclast differentiation, activation, survival Osteoprotegeren ‐ also expressed on: marrow stromal cells & activated T cells

RANK = receptor activator of NF‐kappaB

NF‐kappaB

Stimulation of osteoclast activation

19 How do osteoclasts resorb bone?

Attachment to

bone via integrins (alphavbeta3) on foot‐like Resting podosomes that contain actin osteoclast

Bone Integrins Active bone resorption associate w/: ‐ osteopontin ‐ vitronectin (produce tight seal) Basolateral membrane Resorptive surface Osteoclast w/ ruffled boarder Mineral content of bone: hydroxyapatite . (Ca3(PO4)2)3 Ca(OH)2) 20 Released products activate osteoblasts HCO3‐

‐ Bicarbonate‐chloride Cl exchanger

Integrins (alphavbeta3) OSTEOCLAST

+ H20 + CO2 ‐CAII‐> H + HCO3‐

Howship lacuna or subosteoclastic compartment (a.k.a. ‐ resorption pit)

Chloride H+‐ATPase pump Cl‐ H+ channel Cathepsin K & MMP‐9 pH = 4.5 MINERALIZED BONE 21 BONE RESORPTION

‐ HCl ‐‐> solubilization of mineral content ‐ cathepsin K & matrix metalloproteinase 9 (MMP‐9) ‐‐> degrade connective tissue matrix Transcytosis

Bone Cl‐ H+ Cathepsin K & MMP‐9 pH = 4.5 22 cathepsin K: cysteine protease Definitions:

Transcytosis: A mechanism for transcellular transport in which a cell encloses extracellular material in an invagination of the cell membrane to form a vesicle (endocytosis), then moves the vesicle across the cell to eject the material through the opposite cell membrane by the reverse process (exocytosis).

ALSO: The transport mechanism by which most proteins reach the Golgi apparatus or the plasma membrane; ‐ the vesicles targeted toward lysosomes and secretory storage granules appear to be coated with clathrin.

Syn: cytopempsis, vesicular transport.

23 Where do osteocytes come from?

Osteoblast

Osteocyte

Osteoblasts form new bone where bone was eroded by osteoclasts; osteoblasts become entrapped in bone to

become osteocytes 24 What maintains bone mass?

Nutrition: dietary Ca++ intake and absorption (Vit D)*

Exercise & Gravity : load bearing (fosters bone remodeling)

Sex steroids E2 (women and men) Testosterone (men)

Growth hormone

Thyroid hormone

* Need: Functional intestine & 1,25(OH)2D; 1,25(OH)2D is a function of 25(OH)D, PTH, 25 ‐‐‐ PO4 , renal tubular cell What causes osteoporosis? Complex: multifactorial:

Environment (+) Genetics (+) Aging

Environment: Genetics:Aging: Vitamin D & Ca++ Femaleintake genderEstrogen (risk deficiency ♀ > ♂) in females General nutritionRace (risk:Androgen W > H > deficiencyAA) in males Exposure to sunlightFHx of osteoporosisChronic dis.: or osteoclast fracture activation Body weight Gene polymorphisms,Malnutrition / etc. malabsorption Skeletal trauma Current smoking What environmental factors contribute to osteoporosis? Environment (+) Genetics (+) Aging

Environment: Incr. risk w/:

Vitamin D & Ca++ intake Low intake General nutrition Poor nutrition Exposure to sunlight Lack of exposure Body weight Low body weight Skeletal trauma History of previous fracture Current smoking Chronic inflammation Drugs Glucocorticoids (>3 mo), What genetic factors contribute to osteoporosis? Environment (+) Genetics (+) Aging

Genetics: Comment:

Female gender Risk ♀ > ♂ Race W > H > AA FHx of osteoporosis In 10 relative(s) FHx of fractures In 10 relative(s) Gene polymorphisms Vitamin D receptor LRP5/6 (+) ~=>80 other genes Genome Med. 2013 May 29;5(5):44. doi: 10.1186/gm448. eCollection 2013. Genome-wide approaches for identifying genetic risk factors for osteoporosis. Wu S1, Liu Y2, Zhang L3, Han Y1, Lin Y1, Deng HW3. Author information Abstract

Osteoporosis, the most common type of bone disease worldwide, is clinically characterized by low bone mineral density (BMD) and increased susceptibility to fracture. Multiple genetic and environmental factors and gene- environment interactions have been implicated in its pathogenesis. Osteoporosis has strong genetic determination, with the heritability of BMD estimated to be as high as 60%. More than 80 genes or genetic variants have been implicated in risk of osteoporosis by hypothesis-free genome-wide studies. However, these genes or genetic variants can only explain a small portion of BMD variation, suggesting that many other genes or genetic variants underlying osteoporosis risk await discovery. Here, we review recent progress in genome-wide studies of osteoporosis and discuss their implications for medicine and the major challenges in the field. How does aging contribute to osteoporosis? Environment (+) Genetics (+) Aging

Aging: Comment:

Estrogen deficiency Menopause Androgen deficiency Andropause Chronic disease Cytokine-induced osteoclast activation Malnutrition Impaired appetite w\ aging Malabsorption Impaired absorption w\ aging What causes osteoporosis? Complex: multifactorial:

Environment (+) Genetics (+) Aging

Environment: Aging: Vitamin D & Ca++ intake Estrogen deficiency in females General nutrition Androgen deficiency in males Exposure to sunlight Chronic dis.: osteoclast activation Body weight Malnutrition / malabsorption Skeletal trauma Gravity (loading) Genetics: Female gender (risk ♀ > ♂) Race (risk: W > H > AA) vitamin D receptor polymorphisms LRP5/6 polymorphisms, etc. What are the laboratory findings in osteoporosis?

Result Ca++ Within reference interval

--- PO4 Within reference interval

ALP Within reference interval

Conclusion: There is NO diagnostic lab “signature” of osteoporosis! Why should laboratory testing be performed in cases of suspected osteoporosis?

Primary osteoporosis is a disease of exclusion!

Must: r/o other causes of orthopedic dis./osteopenia/fx’s

Differential diagnosis of ~common and/or important disorders that can present +/- similar to osteoporosis or may co-exist w/ osteoporosis:

Hyperparathyroidism Osteomalacia Lab: Major Dx role Renal osteodystrophy

Paget disease of bone Lab: Minor Dx role Osteogenesis imperfecta* X: gene testing Scurvy

How can hyperparathyroidism be diagnosed in women with orthopedic disease, osteopenia and/or fractures? Result Ca++ Increased

--- PO4 Decreased (or low normal)

ALP Increased (PTH - > osteoblast - > ALP)

Next diagnostic steps: --- Always: r/o renal disease (w/ untx’ed CRF: +/- incr. PO4 )

Confirm hypercalcemia: iCa++ (or alb): r/o dehydration

Measure: PTH (WNL or incr. = 10 hyperparathyroidism) How can osteomalacia be diagnosed in women with orthopedic disease, osteopenia and/or fractures? Result Ca++ Usually: lower range of reference interval

--- PO4 Decreased (Incr. PTH - > hyperphosphaturia - > hypophosphatemia)

ALP Increased (PTH - > osteoblast - > ALP)

Next diagnostic steps: (r/o renal disease) Measure: PTH (incr. = 20 hyperparathyroidism)

25-OHD (decr. w/ vit D def.) - Many rare forms: hepatic rickets, VDDR, VDRR, etc. How can renal osteodystrophy be diagnosed in women with orthopedic disease, osteopenia and/or fractures? Result Ca++ Within reference interval

------PO4 Increased (PO4 retention)

ALP Increased (PTH - > osteoblast - > ALP)

Cr Increased (decr. eGFR, CrCl)

Next diagnostic steps:

Measure: PTH (incr. = 20 hyperparathyroidism) How can Paget disease of bone (osteitis deformans) be diagnosed in women with orthopedic disease and/or fractures? Result Ca++ Within reference interval

--- PO4 Within reference interval

ALP Increased (cytokine? - > osteoblast - > ALP)

Paget disease of bone: Not a lab diagnosis!

Physical examination: deformities; hearing, CN problems

Radiologic findings

Remember: r/o renal disease How can osteogenesis imperfecta be diagnosed in women with orthopedic disease, osteopenia and/or fractures? Result Ca++ Within reference interval

--- PO4 Within reference interval

ALP Increased w/ fracture

Osteogenesis imperfecta: Not a chem. lab diagnosis!

Brittle bones that fracture +/- Hearing loss; blue sclera; dental problems Many disorders w/ variable severity

Radiologic findings: Extremely helpful; Gene testing avail. How can scurvy be diagnosed in women with orthopedic disease and/or fractures? Result Ca++ Within reference interval

--- PO4 Within reference interval

ALP Increased w/ fracture

Scurvy: ~Not a chem. lab diagnosis (unless vit C is measured)! (Dietary vit C def.)

4 H’s: Hemorrhage, hyperkeratosis, hypochondriasis*, and hematologic abnormalities (anemia from blood loss).

*an overwhelming fear that you have a serious disease, even though health care providers can find no evidence of illness. Plain film

This is the preferred initial examination.

head, neck and spine basilar invagination wormian bones kyphoscoliosis vertebral compression fractures codfish vertebrae platyspondyly chest pectus excavatum or carinatum pelvis acetabular protrusio general severe osteoporosis deformed, gracile (over-tubulated) bones cortical thinning hyperplastic callus formation popcorn calcification: the metaphyses and epiphyses exhibit numerous scalloped radiolucent areas with sclerotic margins 1 zebra stripe sign: cyclic bisphosphonate treatment produces sclerotic growth recovery lines in the long bones exuberant callus formation formation of pseudarthrosis at sites of healing fractures

Source: http://radiopaedia.org/articles/osteogenesis-imperfecta-1 How can bone formation be evaluated through laboratory testing? Markers of bone formation (incr. = bone accretion) Marker Comment

Bone alkaline Produced by osteoblasts phosphatase (BAP)

Osteocalcin Produced by osteoblasts Non-collagenous bone matrix protein

Procollagen Formed by conversion of procollagen N-terminal -- > collagen propeptide (s-PINP) How is procollagen N-terminal propeptide (s-PINP) formed? Procollagen

Procollagen amino proteinase Procollagen carboxy proteinase

Procollagen N-terminal propeptide (s-PINP) How can bone turnover be evaluated through laboratory testing? Markers of bone degradation Marker Comment N-telopeptides Bone collagen degradation product

Urine NTx 30-40% decr. from NTx baseline post-3 mos tx = typical response to anti-resorptive therapy.*

Serum NTx Target value for tx’ed post- menopausal women = premeno- pausal reference interval.**

Urine C-telo- Bone collagen degradation product Peptides (CTx) * http://ltd.aruplab.com/Tests/Pub/0070062; ** http://ltd.aruplab.com/Tests/Pub/0070500 How are NTx and CTx formed? N-Telopeptide: released with collagen degradation

One NTx assay: ex: MoAb 1H11 to this octapeptide

NTx from N-terminus of α2 collagen protein C-Telopeptide: released with collagen degradation

Type 1 collagen

CTx, CrossLapsTM assay: MoAb to this octapeptide How can bone turnover be evaluated through laboratory testing? Markers of bone degradation Marker Comment

Urine deoxy- Bone collagen degradation product pyridinoline Target value for tx’ed post- (DPD) menopausal women = premeno- pausal reference interval.*

Urine pyridino- Bone collagen degradation product line

* http://ltd.aruplab.com/Tests/Pub/0070212 How are deoxypyridinoline and pyridinoline formed?

Pyridinoline Deoxypyridinoline (more bone-specific)

NH2 NH2

CH‐COOH CH‐COOH

CH2 CH2

HO CH2‐CH2‐CH‐COOH HO CH2‐CH2‐CH‐COOH

NH2 NH2

N N

CH2‐CH‐CH2‐CH2‐CH‐COOH CH2‐CH2‐CH2‐CH2‐CH‐COOH

OH NH2 NH2 Pyridinium crosslinks: either: - pyridinoline or deoxypyridinoline

Released w/ collagen breakdown How can bone turnover be evaluated through laboratory testing? Markers of bone degradation Marker Comment

Tartrate-resistant Produced by osteoclasts acid phosphatase - reflect bone resorption (TRACP5b) Ir J Med Sci. 2014 Mar;183(1):47-52. doi: 10.1007/s11845-013-0970-6. Epub 2013 Jun 5. Limited utility of tartrate-resistant acid phosphatase isoform 5b in assessing response to therapy in osteoporosis. Brady JJ1, Crowley RK, Murray BF, Kilbane MT, O'Keane M, McKenna MJ. Author information Abstract BACKGROUND:

Tartrate-resistant acid phosphatase isoform 5b (TRACP5b) is a serum bone resorption marker. Our aim was to investigate its utility in monitoring metabolic bone disease. METHODS:

Serum TRACP5b, C-terminal cross-linking telopeptide of type I collagen, urine N-terminal cross-linking telopeptide of type I collagen and free deoxypyridinoline were measured pre- and post-treatment with a parathyroid hormone analogue [PTH (1-34)] (n = 14) or a bisphosphonate (N-BP) (n = 8). TRACP5b, bone alkaline phosphatase (bone ALP), 25-hydroxyvitamin D (25OHD) and parathyroid hormone (PTH) were measured in 100 osteoporosis patients on prolonged bisphosphonate therapy. RESULTS:

Changes in TRACP5b were smaller in magnitude but mimicked those of other bone resorption markers. Absolute changes in TRACP5b and the other resorption markers correlated significantly (p < 0.001). In patients on long- term bisphosphonates, TRACP5b and bone ALP levels were not suppressed. Vitamin D status was consistent with the level of supplementation. CONCLUSION:

TRACP5b has limited utility as a single marker of metabolic bone disease treatment. Clin Lab. 2006;52(9-10):499-509. Tartrate-resistant acid phosphatase 5b (TRACP 5b) as a marker of bone resorption. Halleen JM1, Tiitinen SL, Ylipahkala H, Fagerlund KM, Väänänen HK. Author information Abstract

Tartrate-resistant acid phosphatase (TRACP) is an enzyme that is expressed in high amounts by bone resorbing osteoclasts, inflammatory macrophages and dendritic cells. Two forms of TRACP circulate in human blood, TRACP 5a derived from macrophages and dendritic cells, and TRACP 5b derived from osteoclasts. Recent data have demonstrated the utility of TRACP 5b as a marker of osteoclast number and bone resorption, and serum TRACP 5a as a marker of inflammatory conditions. This review summarizes the scientific knowledge on the role of TRACP in osteoclastic bone resorption, the mechanism of TRACP 5b generation in osteoclasts and its secretion into the blood circulation, the methodology of measuring TRACP 5b, diagnostic evidence for the use of TRACP 5b as a resorption marker, and characteristics of TRACP 5b compared to other commonly used bone turnover markers. What are other novel bone markers?

Marker Comment

Sclerostin Produced by osteocytes - negative regulator of bone mass

Inter alpha-trypsin- Serum-borne reflection of the inhibitor heavy increased osteoclast activity chain H4 precursor (ITIH4) What are other novel bone markers?

Marker Comment

Undercarboxylated Used to assess the vitamin K osteocalcin (ucOC) status in bone metabolism

Serum N-terminal Stable fragment of osteocalcin midfragment Osteocalcin

Bone sialoprotein Produced by osteoblasts - Fn (?): protein nucleator of hydroxyapatite crystal formation What are other novel bone markers?

Marker Comment

Osteoprotegerin Produced by osteoblasts (OPG) - Decoy receptor for RANKL

N-terminal propeptide Serum concentrations correlate of the C-type with bone cortical area and natriuretic peptide endosteal circumference (NT-proCNP) J Bone Joint Surg Am. 2014 Oct 1;96(19):1659-68. doi: 10.2106/JBJS.M.01096. A review of osteocyte function and the emerging importance of sclerostin. Compton JT1, Lee FY1. Author information Abstract

➤ Osteocytes, derived from osteoblasts, reside within bone and communicate extensively with other bone cell populations to regulate bone metabolism. The mature osteocyte expresses the protein sclerostin, a negative regulator of bone mass.➤ In normal physiologic states, the protein sclerostin acts on osteoblasts at the surface of bone and is differentially expressed in response to mechanical loading, inflammatory molecules such as prostaglandin E2, and hormones such as parathyroid hormone and estrogen.➤ Pathologically, sclerostin dysregulation has been observed in osteoporosis-related fractures, failure of implant osseous integration, metastatic bone disease, and select genetic diseases of bone mass.➤ An antibody that targets sclerostin, decreasing endogenous levels of sclerostin while increasing bone mineral density, is currently in phase-III clinical trials.➤ The osteocyte has emerged as a versatile, indispensable bone cell. Its location within bone, extensive dendritic network, and close communication with systemic circulation and other bone cells produce many opportunities to treat a variety of orthopaedic conditions. Arch Biochem Biophys. 2014 Nov 1;561:74-87. doi: 10.1016/j.abb.2014.07.034. Epub 2014 Aug 23. Bone tissue remodeling and development: focus on matrix metalloproteinase functions. Paiva KB1, Granjeiro JM2. Author information Abstract

Bone-forming cells originate from distinct embryological layers, mesoderm (axial and appendicular bones) and ectoderm (precursor of neural crest cells, which mainly form facial bones). These cells will develop bones by two principal mechanisms: intramembranous and endochondral ossification. In both cases, condensation of multipotent mesenchymal cells occurs, at the site of the future bone, which differentiate into bone and cartilage- forming cells. During long bone development, an initial cartilaginous template is formed and replaced by bone in a coordinated and refined program involving chondrocyte proliferation and maturation, vascular invasion, recruitment of adult stem cells and intense remodeling of cartilage and bone matrix. Matrix metalloproteinases (MMPs) are the most important enzymes for cleaving structural components of the extracellular matrix (ECM), as well as other non-ECM molecules in the ECM space, pericellular perimeter and intracellularly. Thus, the bioactive molecules generated act on several biological events, such as development, tissue remodeling and homeostasis. Since the discovery of collagenase in bone cells, more than half of the MMP members have been detected in bone tissues under both physiological and pathological conditions. Pivotal functions of MMPs during development and bone regeneration have been revealed by knockout mouse models, such as chondrocyte proliferation and differentiation, osteoclast recruitment and function, bone modeling, coupling of bone resorption and formation (bone remodeling), osteoblast recruitment and survival, angiogenesis, osteocyte viability and function (biomechanical properties); as such alterations in MMP function may alter bone quality. In this review, we look at the principal properties of MMPs and their inhibitors (TIMPs and RECK), provide an up-date on their known functions in bone development and remodeling and discuss their potential application to Bone Bioengineering. J Bone Miner Res. 2008 Jul;23(7):1106-17. doi: 10.1359/jbmr.080235. Serum biomarker profile associated with high bone turnover and BMD in postmenopausal women. Bhattacharyya S1, Siegel ER, Achenbach SJ, Khosla S, Suva LJ. Author information Abstract

Early diagnosis of the onset of osteoporosis is key to the delivery of effective therapy. Biochemical markers of bone turnover provide a means of evaluating skeletal dynamics that complements static measurements of BMD by DXA. Conventional clinical measurements of bone turnover, primarily the estimation of collagen and its breakdown products in the blood or urine, lack both sensitivity and specificity as a reliable diagnostic tool. As a result, improved tests are needed to augment the use of BMD measurements as the principle diagnostic modality. In this study, the serum proteome of 58 postmenopausal women with high or low/normal bone turnover (training set) was analyzed by surface enhanced laser-desorption/ionization time-of-flight mass spectrometry, and a diagnostic fingerprint was identified using a variety of statistical and machine learning tools. The diagnostic fingerprint was validated in a separate distinct test set, consisting of serum samples from an additional 59 postmenopausal women obtained from the same Mayo cohort, with a gap of 2 yr. Specific protein peaks that discriminate between postmenopausal patients with high or low/normal bone turnover were identified and validated. Multiple supervised learning approaches were able to classify the level of bone turnover in the training set with 80% sensitivity and 100% specificity. In addition, the individual protein peaks were also significantly correlated with BMD measurements in these patients. Four of the major discriminatory peaks in the diagnostic profile were identified as fragments of interalpha-trypsin-inhibitor heavy chain H4 precursor (ITIH4), a plasma kallikrein-sensitive glycoprotein that is a component of the host response system. These data suggest that these serum protein fragments are the serum-borne reflection of the increased osteoclast activity, leading to the increased bone turnover that is associated with decreasing BMD and presumably an increased risk of fracture. In conjunction with the identification of the individual proteins, this protein fingerprint may provide a novel approach to evaluate high bone turnover states. Clin Calcium. 2009 Dec;19(12):1815-21. doi: CliCa091218151821. [Clinical implications of undercarboxylated osteocalcin]. [Article in Japanese] Hosoi T1. Author information Abstract

Importance of vitamin K has been suggested to maintain and improve bone strength. The serum concentration of undercarboxylated osteocalcin (ucOC) is utilized to assess the vitamin K status in bone metabolism. The clinical threshold for ucOC has been determined to discriminate the people at risk for fragility fractures from the view points of vitamin K metabolism. The measurement of ucOC would be useful to select the patients who require vitamin K(2) regimen and to assess the effectiveness of vitamin K(2) therapy.

Cytokine Growth Factor Rev. 2013 Oct;24(5):401-9. doi: 10.1016/j.cytogfr.2013.06.001. Epub 2013 Jul 1. Osteoprotegerin: multiple partners for multiple functions. Baud'huin M1, Duplomb L, Teletchea S, Lamoureux F, Ruiz-Velasco C, Maillasson M, Redini F, Heymann MF, Heymann D.

Osteoprotegerin (OPG) is an essential secreted protein in bone turnover due to its role as a decoy receptor for the Receptor Activator of Nuclear Factor-kB ligand (RANKL) in the osteoclasts, thus inhibiting their differentiation. However, there are additional ligands of OPG that confer various biological functions. OPG can promote cell survival, cell proliferation and facilitates migration by binding TNF-related apoptosis inducing ligand (TRAIL), glycosaminoglycans or proteoglycans. A large number of in vitro, pre-clinical and clinical studies provide evidences of OPG involvement in vascular, bone, immune and tumor biology. This review describes an overview of the different OPG ligands regulating its biological functions. Horm Res Paediatr. 2010;74(3):201-6. doi: 10.1159/000282111. Epub 2010 Jun 10. N-terminal C-type natriuretic propeptide is associated with the endosteal apposition of bone in females with a persistent eating disorder. Fricke O1, Beccard R, Semler O, Stabrey A, Tutlewski B, Lehmkuhl G, Schoenau E. Author information Abstract BACKGROUND/AIM:

Females with anorexia nervosa (AN) are often affected by osteoporosis. The study intends to investigate the association between serum levels of the N-terminal propeptide of the C-type natriuretic peptide (NT-proCNP) and bone development in anorexic females. SUBJECTS AND METHODS:

In a catamnestic visit, 21 females, formerly treated for AN, were assessed for the presence of eating disorders and analyzed for bone parameters of the distal radius (4% site) with peripheral quantitative computed tomography (pQCT), for maximal isometric grip force (MIGF) and for NT-proCNP serum levels. RESULTS:

The 9 females with a persistent eating disorder had lower height and weight than the recovered girls. NT-proCNP was correlated with the cortical area (r = 0.521), the endosteal circumference (CE, r = -0.468) and the ratio of MIGF to cross-sectional bone area (r = 0.434). CE explained 40% of the variance of NT-proCNP in females with persistent eating disorders, but was not associated with NT-proCNP in recovered girls (p = 0.691). The association between CE and NT-proCNP was not existent when the correlation was controlled for the duration of amenorrhea and the supplemented cumulative dose of ethinylestradiol (p = 0.275). CONCLUSION:

NT-proCNP reflects metaphyseal inwaisting which is modified by estrogens and the pressure on the growth plate. What is the value of bone marker testing in the evaluation of osteoporosis and its therapy? “BTMs (bone turnover markers) are widely used in bone research including therapeutic trials of new medications for osteoporosis and other bone disease. Whilst further data are awaited to confirm their utility in the clinical management of osteoporosis, they are currently used in specialist clinical practices, especially in monitoring treatment.”

-Vasikaran SD, Chubb SA, Schneider HG Towards optimising the provision of laboratory services for bone turnover markers. Pathology. 2014 Jun;46(4):267-73. Summary

Osteoporosis is NOT a biochemical diagnosis

Laboratory testing to: r/o:

Hyperparathyroidism Osteomalacia Renal osteodystrophy

Clinical/radiologic evaluation for:

Paget disease of bone Osteogenesis imperfecta

Bone turnover markers: clinical value? THE END What is hypophosphatasia?

Hypophosphatasia is an inherited disorder that affects the development of bones and teeth. This condition disrupts a process called mineralization, in which minerals such as calcium and phosphorus are deposited in developing bones and teeth. Mineralization is critical for the formation of bones that are strong and rigid and teeth that can withstand chewing and grinding.

The signs and symptoms of hypophosphatasia vary widely and can appear anywhere from before birth to adulthood. The most severe forms of the disorder tend to occur before birth and in early infancy. Hypophosphatasia weakens and softens the bones, causing skeletal abnormalities similar to another childhood bone disorder called rickets. Affected infants are born with short limbs, an abnormally shaped chest, and soft skull bones. Additional complications in infancy include poor feeding and a failure to gain weight, respiratory problems, and high levels of calcium in the blood (hypercalcemia), which can lead to recurrent vomiting and kidney problems. These complications are life‐threatening in some cases.

The forms of hypophosphatasia that appear in childhood or adulthood are typically less severe than those that appear in infancy. Early loss of primary (baby) teeth is one of the first signs of the condition in children. Affected children may have short stature with bowed legs or knock knees, enlarged wrist and ankle joints, and an abnormal skull shape. Adult forms of hypophosphatasia are characterized by a softening of the bones known as osteomalacia. In adults, recurrent fractures in the foot and thigh bones can lead to chronic pain. Affected adults may lose their secondary (adult) teeth prematurely and are at increased risk for joint pain and inflammation.

The mildest form of this condition, called odontohypophosphatasia, only affects the teeth. People with this disorder typically experience abnormal tooth development and premature tooth loss, but do not have the skeletal abnormalities seen in other forms of hypophosphatasia. How common is hypophosphatasia?

Severe forms of hypophosphatasia affect an estimated 1 in 100,000 newborns. Milder cases, such as those that appear in childhood or adulthood, probably occur more frequently.

Hypophosphatasia has been reported worldwide in people of various ethnic backgrounds. This condition appears to be most common in Caucasian (white) populations. It is particularly frequent in a Mennonite population in Manitoba, Canada, where about 1 in 2,500 infants is born with severe features of the condition. What genes are related to hypophosphatasia?

Mutations in the ALPL gene cause hypophosphatasia.

The ALPL gene provides instructions for making an enzyme called alkaline phosphatase. This enzyme plays an essential role in mineralization of the skeleton and teeth. Mutations in the ALPL gene lead to the production of an abnormal version of alkaline phosphatase that cannot participate effectively in the mineralization process. A shortage of alkaline phosphatase allows several other substances, which are normally processed by the enzyme, to build up abnormally in the body. Researchers believe that a buildup of one of these compounds, inorganic pyrophosphate (PPi), underlies the defective mineralization of bones and teeth in people with hypophosphatasia.

ALPL mutations that almost completely eliminate the activity of alkaline phosphatase usually result in the more severe forms of hypophosphatasia. Other mutations, which reduce but do not eliminate the activity of the enzyme, are often responsible for milder forms of the condition.

Read more about the ALPL gene. How do people inherit hypophosphatasia?

The severe forms of hypophosphatasia that appear early in life are inherited in an autosomal recessive pattern. Autosomal recessive inheritance means that two copies of the gene in each cell are altered. Most often, the parents of an individual with an autosomal recessive disorder each carry one copy of the altered gene but do not show signs and symptoms of the disorder.

Milder forms of hypophosphatasia can have either an autosomal recessive or an autosomal dominant pattern of inheritance. Autosomal dominant inheritance means that one copy of the altered gene in each cell is sufficient to cause the disorder.

Source: http://ghr.nlm.nih.gov/condition/hypophosphatasia