The incidence of metabolic bone disease (MBD) of prematurity is as high as 23% of very low-birthweight infants and as high as 55% of extremely low-birthweight infants, but a lack of consensus in diagnostic criteria makes the actual incidence difficult to assess. Understanding the pathogenesis and biomarkers to detect MBD may help clinicians identify patients at risk for MBD and provide appropriate nutritional management to these patients.After completing this article, readers should be able to: Explain the physiology of bone formation and mineral utilization of the fetus and newborn.Describe the risk factors for and diagnostic tools to detect metabolic bone disease.List the mineral requirements and explain the management options for infants with metabolic bone disease.Metabolic bone disease (MBD) of prematurity is defined as a reduction in bone mineralization below expected values at a specific gestational age and is characterized by specific biochemical and radiographic findings. The incidence of MBD is as high as 23% of very low-birthweight (VLBW) infants and as high as 55% of extremely low-birthweight (ELBW) infants. It is usually detected between 10 and 16 weeks’ postnatal age (1) but a lack of consensus in diagnostic criteria makes the actual incidence difficult to assess. While our understanding of MBD and nutrition of prematurity infants has improved significantly in recent decades, MBD continues to be a comorbidity in premature infants that is challenging to detect and treat, potentially leading to long-term poor growth and pathologic fractures.Skeletal morphogenesis or the process by which the skeletal framework develops in utero, begins in the sixth week of gestation. (2) The fetal skeleton develops in 3 distinct pathways. The craniofacial skeleton arises from cranial neural crest cells, the axial skeleton arises from paraxial mesoderm cells, and the limb skeleton arises from lateral plate mesoderm cells. The basic mechanism by which the skeleton forms is firstly through migration of these 3 cell types into their correct spatial location to form the anatomic framework for the fetal skeleton. This is then followed by proliferation of these cell types into areas of high cellular density, which serves to further delineate the fetal skeletal anatomy. Finally, the cells either differentiate into chondrocytes, which form a cartilaginous skeletal structure that eventually is replaced by bone (endochondral bone formation) or they differentiate into osteoblasts, which form the osteoid bone matrix directly (intramembranous bone formation). (3)Intramembranous bone formation occurs for most of the craniofacial region and the clavicles. Osteoblasts, which differentiate from cranial neural crest cells, begin to form and secrete osteoid, the unmineralized protein mixture that forms the organic bone matrix. Osteoclasts, which differentiate from a hematopoietic stem cell progenitor, also begin to invade the region and have the function of bone resorption. Calcium circulating in utero binds to osteoid, which hardens the matrix. As the secreted osteoid surrounds osteoblasts, the osteoblasts then may further differentiate into osteocytes, which remain present in the mature skeleton. These osteocytes have signaling and mechanoreceptor functions to regulate osteoblast and osteoclast activity, as well as bone mineral metabolism.Endochondral bone formation forms the remainder of the axial and limb skeleton. At 8 weeks’ gestational age, the cartilaginous framework for the fetal skeleton is established. (4) Osteoblasts begin to form primary ossification centers and secrete osteoid, whereas osteoclasts work to resorb the chondrocytes that the osteoblasts replace. Calcium binding by the osteoid framework continues throughout gestation, but the majority of skeletal mineralization occurs in the third trimester. The process of linear bone growth then continues for years after birth.Parathyroid hormone (PTH) is an 84-amino acid polypeptide hormone that is central to the metabolism of calcium and exerts its effects on 2 organ systems, the bone and kidney. Changes in serum calcium ions are sensed by the calcium-sensing receptor of the parathyroid gland, which regulates gene transcription within the parathyroid cells. Increases in serum calcium decrease the production and secretion of PTH by parathyroid chief cells. (5)PTH breaks down bone by binding to PTH type 1 receptors on osteoblasts. PTH binding causes the osteoblasts to upregulate macrophage colony-stimulating factor and receptor activator of nuclear factor κ B ligand (RANKL) expression. Activation of RANKL in turn by RANK binding increases osteoclast formation. (6) This ultimately leads to bone resorption and the release of calcium into the bloodstream. PTH also increases calcium reabsorption in the distal tubules through the upregulation of a calcium ion transporter, TRPV5. In addition, PTH decreases phosphate reabsorption and upregulates the production of α1-hydroxylase, which converts vitamin D into its biologically active metabolite, 1,25-dihydroxy-vitamin D.1,25-dihydroxyvitamin D, or calcitriol, is the biologically active form of vitamin D. It is created by the conversion of vitamin D2 or vitamin D3 by α1-hydroxylase in the kidneys. Vitamin D2 is obtained enterally and vitamin D3 is obtained from sunlight exposure to the skin. Calcitriol increases gastrointestinal absorption of calcium by increasing epithelial calcium transport proteins such as TRPV6 and calbindin. (7)PTH-related peptide is produced by many tissues in the body including placental tissue, parathyroid tissue, skeletal tissue, and smooth muscle cells. It has many effects on the developing fetus, including increasing calcium transfer across the placenta and is involved in proper chondrocyte development and endochondral bone formation. (8)Calcitonin is a 32-amino acid polypeptide hormone produced and secreted from the thyroid gland as well as the placenta and upregulated in response to increases in serum calcium. It upregulates calcitriol production in the kidney and decreases osteoclast activity through calcitonin receptor activation. (9)To support the growing skeletal structure in the fetus, concentrations of calcium, magnesium, and phosphorus are maintained higher than maternal values by 1.2 to 2 mg/dL (0.3–0.5 mmol/L), 0.12 mg/dL (0.05 mmol/L), and 1.5 mg/dL (0.5 mmol/L), respectively. (10) Eighty percent of calcium accretion by the fetus occurs in the third trimester as this is when the majority of skeletal mineralization occurs. (11) A fetus in the third trimester takes in an average of 100 to 120 mg/kg per day of calcium and 50 to 65 mg/kg per day of phosphorus. (12) The main source of calcium to the fetus is active transport across the placenta. In addition, a small amount of body calcium lost through filtering by the fetal kidneys is excreted as amniotic fluid and ingested by the fetus, returning it to the fetal circulation via intestinal intake. The relative hypercalcemia of the fetus is maintained via low serum concentrations of fetal PTH and calcitriol, and high concentrations of fetal PTH-related peptide. Low calcitriol levels are maintained due to the low PTH levels, as well as high concentrations of placental 24-hydroxylase, which prevents the conversion of inactive vitamin D to calcitriol. (13) Calcitonin produced by the fetal thyroid and placenta is also increased in fetal circulation and aids in bone mineralization. (10)Immediately after birth, the umbilical cord is cut, which removes the neonate’s placental supply of calcium. As a result, intestinal intake of calcium becomes the neonate’s source of calcium and this remains in balance with skeletal calcium stores to maintain normal serum calcium homeostasis. Intestinal uptake of calcium occurs via active and passive transport. It is suggested that the majority of calcium intake is initially passive and concentration dependent. (10) As the gastrointestinal system matures, active transport becomes more important and is mediated by calbindins, which are vitamin D–dependent calcium-binding receptors present on enterocytes.Mineral accretion occurs across the placenta, therefore, prenatal risk factors include conditions that might impair nutrient delivery to the fetus. Uteroplacental insufficiency, preeclampsia, fetal growth restriction, and chorioamnionitis are associated with an increased risk of MBD. (14) Other risk factors may include maternal vitamin D deficiency and male sex (Table 1). (15)Prematurity is a major risk factor for MBD, as the majority of calcium accretion occurs during the third trimester. Conditions that affect nutrient intake postnatally also increase the risk of MBD, including feeding intolerance, bronchopulmonary dysplasia, and prolonged parenteral nutrition (Table 1). All of these have the potential to cause deficiencies in calcium, phosphorus, or vitamin D. (16) Medications that increase bone mineral excretion, including loop diuretics, glucocorticoids, caffeine, and sodium bicarbonate, also increase the risk for MBD. Sepsis, acidosis, renal disease, liver disease, and gastrointestinal disease such as necrotizing enterocolitis or abdominal wall defects have also been shown to contribute to MBD. (14) A reduction in bone formation may also occur due to a lack of mechanical stimulation from movement, (17) as is the case in neonates who require sedation or paralysis.There are no universal screening or diagnostic criteria for MBD; however, many biochemical and radiographical markers can aid in the detection and diagnosis of MBD.Serum calcium levels normally reach a nadir in both preterm and term neonates 24 to 30 hours after birth and then begin to rise due to an increase in PTH levels. (18) In the kidney, PTH increases calcium reabsorption and decreases phosphate reabsorption. It increases intestinal absorption of calcium and phosphate via the synthesis of calcitriol, and increases bone resorption to release calcium into the serum. Serum calcium levels can be misleading in the diagnosis of MBD, as infants may maintain normal serum calcium levels by decreasing calcium mineralization in bone. Moreover, calcium levels less than 8.5 mg/dL (2.13 mmol/L) suggest inadequate calcium intake. (19)If nutritional calcium intake is chronically insufficient, the biochemical representation of MBD can include hypophosphatemia due to the effects of PTH on the kidney. A serum phosphate level below 3.6 mg/dL (1.16 mmol/L), in exclusively breastfed newborns, confers a greater risk for MBD. (20)Tubular reabsorption of phosphate (TRP) can also be measured as follows: urinary phosphate/serum phosphate × serum creatinine/urinary creatinine. If serum phosphate is less than 5.5 mg/dL (1.78 mmol/L) and TRP is greater than 95%, this may suggest insufficient phosphorus intake. (21)Alkaline phosphatase is an enzyme present in many body tissues, including skeletal, liver, and intestine; the majority comes from skeletal tissue and represents bone metabolism. (22) Serum levels in the newborn continue to rise until 4 to 6 weeks of age, when they peak. Alkaline phosphatase levels then decrease around 8 to 10 weeks of age. Different levels of alkaline phosphatase have been shown to be associated with MBD in various neonatal populations. A level greater than 900 IU/L (15 μkat/L) in infants of less than 33 weeks’ gestational age, along with serum phosphate lower than 5.6 mg/dL (1.8 mmol/L), has shown sensitivity of 70% and specificity of 100% for MBD. (23) In ELBW infants (<30 weeks’ gestational age), a level greater than 500 IU/L (8.3 μkat/L) is associated with the development of MBD. (16)Levels of vitamin D are measured as 25-hydroxyvitamin D, its inactive form. Values less than 20 ng/mL (50 nmol/L) indicate a deficiency; values less than 30 ng/mL (75 nmol/L) are also useful in identifying neonates at risk of developing MBD. (19)As PTH is released in response to hypocalcemia and is a marker of bone resorption, elevated levels suggest the development of MBD. Levels higher than 100 pg/mL (100 ng/L) may be useful to identify neonates who are at risk of developing MBD. (24)A reduction of bone mineralization by 20% to 40% is needed before it becomes detectable on radiography, so radiographs are not reliable in early MBD when bone demineralization is not usually significant. (25) Pathologic fractures due to MBD also do not occur frequently in early MBD. A grading scale describing radiographic abnormalities has been developed, reflecting the gradual chondrocyte expansion and its failure to mineralize (26): Normal: No abnormalities noted, with normal bone density and a normal dense white line at the metaphysis.Grade 1: Loss of a dense white line at the metaphysis and thinning of the cortex.Grade 2: Changes seen in grade 1, along with fraying of the metaphysis, with splaying (metaphyseal widening) and cupping (metaphyseal concavity), that is, rachitic changes.Grade 3: Changes seen in grade 2, along with fractures.This diagnostic test is the gold standard to assess bone mineral density in neonates, including in preterm infants. The scan measures bone mineral density in grams of hydroxyapatite per centimeter squared and is directed at the neonate’s lumbar spine, forearm, or calcaneus. Technical aspects of performing this examination limit its usefulness in neonates. One study showed that a bone mineral density greater than 0.068 g/cm2 confers a lower risk of developing MBD in VLBW infants of less than 31 weeks’ gestational age. (27)This test uses ultrasonography to measure the speed of sound along the tibia to indirectly quantify bone mineral density in infants. (28) In comparison with term infants, preterm infants born between 24 and 28 weeks’ gestational age were shown to have lower quantitative ultrasonography parameters at corrected term age, indicating a reduction in bone mineralization related to prematurity. (29)The initial step is to identify infants who are most at risk for developing MBD, which include premature infants (VLBW or ELBW infants) and those with gastrointestinal disorders that lead to syndromes of malabsorption. Routine biochemical evaluation of MBD is recommended to start at 4 weeks of age and at minimum should include measurement of serum alkaline phosphatase and phosphorus levels. Serum calcium and vitamin D can also be measured at this time. Monitoring of biochemical markers should continue at least every 2 weeks thereafter. Monitoring can be discontinued if the patient is at low risk of developing MBD, is receiving full enteral feedings, has a stable level of alkaline phosphatase less than 500 to 600 IU/L (8.3–10 μkat/L), and a stable phosphorus level greater than 4 mg/dL (1.3 mmol/L). (30) Monitoring should continue every 2 weeks if these criteria are not met and managed appropriately.If alkaline phosphatase levels are greater than 800 IU/L (13.3 μkat/L), or if there is a clinical concern for fractures, radiographic monitoring of rachitic changes should be initiated.The primary treatment strategy for MBD is to optimize the neonate’s intake of calcium, phosphorus, and vitamin D. Other considerations include discontinuation of medications that promote bone demineralization, including loop diuretics and glucocorticoids.Guidelines for calcium and phosphorus requirements for the premature infant were released by the American Academy of Pediatrics in 2013. (30) To support proper skeletal development in the preterm infant and prevent MBD, adequate amounts of calcium, phosphorus, and vitamin D must be supplied in a manner that mimics the in utero accretion. In a healthy preterm infant, enteral calcium absorption is around 50% to 60% of intake and phosphorus absorption is around 80% to 90% of intake. (31) Unfortified human milk contains 25 mg/dL of calcium and is therefore insufficient for the mineral needs of the premature infant. Preterm formulas and human milk fortifiers designed for preterm infants have increased concentrations of calcium to meet this need.Recommended enteral requirements by the 2013 American Academy of Pediatrics report for VLBW infants are 150 to 220 mg/kg per day of calcium, 75 to 140 mg/kg per day of phosphorus, and 200 to 400 IU/day of vitamin D. If the neonate requires parenteral nutrition, the recommended parenteral requirements are 75 to 100 mg/kg per day of calcium and 50 to 80 mg/kg per day of phosphorus. (32) In parenteral nutrition, the ratio of calcium and phosphorus is optimally 1.7:1 (mg/mg) (32); infants may develop hypercalcemia or hypophosphatemia, in which the calcium-to-phosphorus ratio is too high. The recommended mineral intake for prevention and treatment of MBD of prematurity is summarized in Table 2.If an infant is meeting these daily requirements, but continues to have evidence of MBD, further mineral supplementation can be given in the form of elemental calcium and phosphorus; these can be given via both enteral and parenteral routes. The starting dose of elemental calcium is 20 mg/kg per day and can be increased to a maximum of 70 to 80 mg/kg per day; the starting dose of elemental phosphorus is 10 to 20 mg/kg per day and can be increased to 40 to 50 mg/kg per day. (30)While there is not much research into long-term outcomes of neonates affected by MBD, most premature infants do well after discharge with respect to bone mineralization. (30) Continued mineral supplementation in the form of transitional formula may be appropriate for infants who continue to have elevated alkaline phosphatase levels after discharge. Vitamin D supplementation should also be continued for infants to receive at least 400 IU/L daily.