Increased bone mineral content in young adults with familial hypophosphatemic vitamin D refactory rickets

Increased Bone Mineral Content in Young Adults With Familial H ypophosphatemic Vitamin D Refractory Rickets J. E. Harrison,

W.

A. Cumming,

V. Fornasier,

D. Fraser,

Seven adults with familial hypophosphatemia have been investigated by histologic and radiographic examination of bone, and estimates of bone mineral status by in vivo neutron activation analysis (IVNAA). Histological examination showed severe osteomalacia and osteosclerosis in all cases. Radiography showed skeletal deformities and other sequelae of severe rickets of childhood in five of the seven cases, with, in addition, thickened well-mineralized bones; the other two showed normal radiographs. IVNAA measurements showed that the first five had

S. W.

Kooh,

and

K. G. McNeil1

greater than normal bone calcium and that the other two had normal values. Thus, in all cases then is a greater than normal total bone tissue (osteoid and mineralized bone together). The quantitative body calcium measurements show clearly that osteosclerosis occurs in familial hypophosphatemia, confirming the opinions based on histological and radiological data. Although there has been occasional reference to this sclerosis in the literature, up to the present it has received little attention.

H

YPOPHOSPHATEMIC VITAMIN D refractory rickets is characterized by rickets (or osteomalacia) and hypophosphatemia in individuals receiving physiological amounts of vitamin D. The serum calcium concentration is always normal. In the familial form of the condition hypophosphatemia is transmitted as an x-linked dominant trait but sporadic cases have been reported. In contrast to the diminished bone density which might be expected, skeletal radiographs of young adults known to have had hypophosphatemic vitamin D refractory rickets frequently demonstrate unusually thick bones which appear well mineralized. Similar findings have been reported’-’ but osteosclerosis in connection with this disease has received little attention. The technique of in vivo neutron activation analysis (IVNAA) allows a quantitative estimate to be made of the total (or a major part of the total) body calcium.4 Accordingly, such measurements have been made in a group of patients who were known to have had hypophosphatemia since infancy, and the results compared to radiological and histological data. MATERIALS Patienfs. were included

AND

METHODS

Seven young, fully grown individuals with vitamin D refractory hypophosphatemia in this study. Relevant clinical features are summarized in Table I. The criteria

for

From the Departments of Medicine, Radiology. Pathology, and Paediatrics. University of Toronto, and the Toronto General Hospital, the Princess Margaret Hospital. and the Hospital for Sick Children, Toronto, Canada. Receivedfor publication March 26. 1975. Supported in part by the Garfield Weston Charitable Foundation and Ontario Health Research Grant 399. Reprint requests should be addressed to Joan E. Harrison, M.D., Room 7326, Medical Science Building, University of Toronto, Toronto, Canada, MSS lA8. o 1976 by Grune & Stratton, Inc. Metabolism,

Vol. 25, No. 1 (January),

1976

33

1

6 7

30 23

20 28 24

17 29

Age

F

F

M M

F

F

F

Sex

1 had genu varum

861650 523188

3 4 5

‘Patient

038439 290834 261683 198426

2

range 380835

NO.

NO.

Normal

H.S.C.

CO*

and radiographic

<3 <<3

3 <<3 <3

10

25

Height percentile of normal

evidence

+ +

+ + +

0

0*

Skeletal deformities

of rickets

9.8 9.6

10.1

9.4 10.2

10.0

9.0- -10.5 9.6

CO WI lOOmI

for the first

1.5

1.8

2.3 2.7

2.2 2.6

1 .a

3.0-5.2

P me/ lOOmI

!s?rum

108 106 70 35

84 113

136

<90

Data

55 55 40

57 50 30

Marrow

~

Patient

Alk. PhoS’CW W/I

1.

5 yr of life.

Table

25 35 40

26 35 55

Mineral

20

10

20

17 15 15

Orteoid

Biopsy Volumes

44 22 33

O-7% 39 30 21

Osteoid Index

17 12 6

13 14

Y’

~ Duration

0 150,000

60,000

lO,OOo-175,000 25,OOC100,000 0 150.000-300.000

25,ooO-

25,000-

Range

150.000

60.000 0

current

Ofor5yr

0

Ofor4yr

Ofor3yr

Dose I.U. per day

Vitamin D therapy

BONE

MINERAL

35

CONTENT

diagnosis have been described previously.S*6 Patients 3, 4, and 5 were siblings and patient 1 was a first cousin of these. Except in the case of patient 7, instances of hypophosphatemic vitamin D refractory rickets were documented in relatives, and in each family the inheritance was consistent with an x-linked dominant pattern. Serum inorganic phosphate levels were more than 21 standard deviations below the age-related mean concentration for the normal population.’ Renal function, as measured by 24 hour creatinine clearance, was normal in all subjects. Five patients had received large doses of vitamin D,s during childhood, and two of these were still receiving vitamin D therapy at the time of study (Table I). One patient (patient 6) with dwarfism and moderately severe deformities, and one (patient 2) with normal stature and no deformities, grew up elsewhere and did not receive vitamin D therapy at any time. None of the patients were treated with oral phosphate supplements. Investigative procedures. Partial body calcium measurements by IVNAA and conventional skeletal radiography were carried out on all patients. Bone biopsies from the anterior iliac crest were obtained from all except patient 4. Fine-detail radiographs were made of the fresh tissue cores. Decalcified and undecalcified bone sections were studied by standard histological techniques. Volumes of marrow, osteoid, and mineralised bone were measured using the Chalkley grid. A measure of the unmineralised bone tissue was obtained according to the method of Sissons et al.,9 expressing as the osteoid index the relation of the osteoid to the total of osteoid and unmineralised bone. The technique of partial body IVNAA has been described in detail elsewhere.4 The skeletal calcium is measured in an area 60 x 30 cm which incorporates the trunk and upper thighs. Following exposure to neutrons, a count of gamma rays from radioactive 49Ca is obtained which is quantitatively indicative of the amount of calcium in the part of the body measured. Correction is made for variation in overall efficiency with subjects of different thickness.” To be clinically useful, the 49Ca counts or the absolute amount of calcium in the body must be related to the normal value for subjects of the same skeletal frame size. One would predict that the skeletal frame size is a function of (height)’ and therefore that in normal subjects the calcium in the body, or the 49Ca counts, would be a function of the (height).3 In previous experiments” we have shown that such a relationship does hold and a Ca bone index (CaBI) has been derived based on this relationship. In the previous work, IVNAA measurements were carried out on 18 volunteers, both men and women;‘between the ages of 34 and 53 yr. Their range of height was 1.51-1.89 metres, and their range of efficiency-corrected Ca count was 653-1295. The results demonstrated a linear correlation between the efficiency-corrected calcium counts and the (height)3 of the subjects. The regression line was expressed by the equation Calcium Count = 17 + 207 x (Ht)3, (with height in metres) and the correlation coefficient r = 0.91. From this regression line, a calcium bone index (CaBI) was obtained as follows: CaBI = (Ca count - 17)/(207 x (Ht)3). For the 18 volunteers the CaBI had a range of 0.9- 1.2 with a mean value of 1 .O. Thus, the CaBI provides an estimate of the mineral status of a subject as a fraction of the mineral status for normal subjects of the same skeletal frame size, but the validity of the CaBI depends on the assumption that Ht’ is a reliable estimate of skeletal frame size. This assumption

Table

2.

Calcium

gone

Index

(CaBI)

Bard

on Height CaBI'

Patient #

Calcium count

Height Ill

ArmSpan m

Basedon Height

Basedon ArmSpan

and

Arm

Span

Expected Heightonthe ofObservedCaCount m

1

892

1.59

1.59

1.05

1.05

1.52-1.67

2 3

862 1029

1.57 1.51

1.63 1.53

1.06 1.42

0.95 1.38

1.50-1.65 1.60-1.76

4 5 6

1450 1561 1160

1.56 1.60 1.45

1.69 1.70 1.47

1.82 1.83 1.81

1.43 1.52 1.74

1.79-1.97 1.84-2.02 1.66-1.85

756

1.25

1.27

1.83

1.74

1.44-1.58

7 *Normal

range

CaBl0.90-1.20.

Basis

36

may be invalid for subjects with rachitic used instead of height for the determination

HARRISON

deformities: of CaBI

as an alternative, in the present studies.

arm

span

has

ET Al.

also

been

RESULTS

The results of the calcium measurements are shown in Table 2 together with the CaBI values based on both the observed height and the arm span. Although in most subjects the CaBI value based on arm span is lower than the value based on height, the conclusions are the same. In all five patients with rachitic deformities (patients 3,4, 5, 6, and 7) the CaBI values were significantly above the normal range (p < 0.002). In contrast, the two subjects without rachitic deformities had CaBI within the normal range. To illustrate the significance of the CaBI, we have included in Table 2 the range of height that normal subjects would be expected to have for the same

Fig. 1. (A) A mdiogmph of the femur of diameter of the femur and the thickness of of the same height. The bone apwn well two. The femur appears normal. Them is no well minemlised.

patient six. The femur is wide and bowed. The the cortex an greater than these for normal minemlised. (B) A mdiogmph of the femur of bowing, no cortical thickening, and the bone

ovemll subjects patient appoors

BONE

MINERAL

CONTENT

37

efficiency-corrected calcium counts. The observed Ca counts would give normal CaBI values if the five subjects with deformities were each at least 9 cm taller. The radiographs of five patients (patients 3-7) showed the skeletal sequelae of severe rickets in childhood. These include bowing of long bones, thickening of cortices, and coarse trabecular pattern. Although the thickened cortices are in part due to buttressing of the concave periosteal and endosteal surfaces consequent on bowing of the bones, thickened cortices were seen in other areas as well. Similar generalized thickening of long bones has been noted by others.3 Ossification of ligaments and tendon attachments was not seen in any patient. A radiograph of the femur of patient 6 is shown in Fig. 1A. The femur is wide and bowed. The overall diameter of the femur and the thickness of the cortices are greater than those of normal subjects of the same height. The bone appears well mineralized. However, the trabecular pattern is abnormally coarse. Two subjects (patients 1 and 2) had normal skeletal radiographs. The femur of patient 1 is shown in Fig. 1B. There is no bowing, no cortical thickening, the bone is well mineralized and the trabeculae appear normal. Fine-detail radiographs of the cores of bone biopsies obtained prior to any processing showed increased bone density with widening and coarsening of trabecular outlines, indicating that there was an overall increase in the amount of mineralized bone. The increased bone density was observed in all six biopsies, including those from the two patients who had no skeletal deformities (patients 1 and 2). Fig. 2A shows increased bone density in a radiograph of the bone biopsy core from patient 6 while Fig. 2B shows a radiograph of normal bone (see also Table 1). All bone biopsies showed excessive amounts of unmineralized bone matrix, as shown by the high osteoid index (Table 1). In all the samples more than 70%

Fig. 2. (A) This is a fine-detail mdiogmph of a 3 x 5 mm core of tmbacular bone from the iliac crest biopsy in patient six. The coarsening and irregular outline of the tmbeculae is evident. The overall increase in bone results in decrease in marrow lucency with overlap of the widened irregular tmbeculae (x 10). (6) Fine-detail radiograph of normal tmbacular bone for comparison with Fig. 3. The iliac crest biopsy was taken at autopsy from a young adult with no known abnormality in calcium metabolism. The smooth contours and the regular pattern and distribution of the trabecular bone are evident ( x IO).

38

HARRISON

ET At.

Fig. 3. Undecolcifled section of bone biopsy from patient three, stained with Masson Trichrome and photogmphed under polarized light. Birefringent osteoid seams are present along the bony surfacesand appear light in the photograph. The irregular pattern of the tmbecular bone with irregular outlines and widening of the tmbeculae is evident. This demonstrates not only the excess of osteoid present, but also the ovemll increase in the amount of bone matrix that is present and minemlized ( x 100).

of the bone surfaces were covered by widened osteoid seams in which frequently more than 6 lamelae could be identified as shown by Fig. 3, (normal osteoid seams are no more than 3 lamelae in width”). In all biopsies, there was very little osteoblastic or osteoclastic activity, no fibrous tissue and no evidence of abnormal fibre bone. DISCUSSION

Although these seven adults with hypophosphatemia had extensive osteomalacia, the studies reported here show no evidence of bone mineral deficit. The CaBI values indicate normal bone mineral mass in two subjects (patients I and 2) and bone mineral mass above normal in the remaining five (patients 3-7). These IVNAA data emphasize the assessment based on conventional radiographs. In all patients the radiography indicates that the bones are well mineralized. In addition, because of the overall increase in the diameter of their long bones coupled with increased cortical thickness, the five patients with overt skeletal deformities would be expected to have more bone than normal, presumably due to excessive subperiosteal bone formation. These radiological findings are in agreement with reported data.’ Fine detail radiographic examination of all six bone biopsies revealed an increase in the width of individual trabeculae. On the assumption that the biopsies give true indications of the status of trabeculae throughout the body, there then

BONE

MINERAL

39

CONTENT

is a consequent increase in total trabecular bone mineral mass. It should be noted that trabecular bone is only about 20% of the total skeleton,13 and even a considerable increase in trabecular mineral mass might not be sufficient to increase the total mineral mass (i.e., CaBI) above the normal range; as noted, patients 1 and 2 have a normal CaBI. Nevertheless, the evidence of the high osteoid index taken with normal CaBI values suggests that the total bone tissue (mineralized bone plus osteoid) must be significantly greater than normal in patients 1 and 2 as it is in the other five patients. It is of considerable interest to find osteomalacia associated with evidence of increased total bone tissue (both trabecular and cortical bone tissue). Osteosclerosis may be associated with osteomalacia in other types of metabolic bone disease, for example renal osteodystrophy,‘c’6 and occasionally primary hyperparathyroidism, ‘7~‘8but the osteosclerosis in the present patients shows some features peculiar to hypophosphatemic rickets. The osteosclerosis of primary hyperparathyroidism and of renal osteodystrophy is usually patchy, and involves predominantly the spine, pelvis, and skull. In contrast, the patients with hypophosphatemia have generalized increased thickness of long bones. In addition, the osteosclerosis seen in primary or secondary hyperparathyroidism, is associated with extensive osteitis fibrosa cystica whereas the bone biopsies from our patients with hypophosphatemia showed no histological evidence of increased parathyroid hormone (PTH) activity. One can only speculate as to the cause of this osteosclerosis. It is difficult to attribute the osteosclerosis to PTH activity as had been done in the case of parathyroid bone disease. 19**0In the present small group of patients there is no evidence that vitamin D therapy had an effect in the bone pathology since one patient from each group (with deformities and without deformities) had never received any therapy and one from each group was maintained on massive doses of vitamin D from early childhood. Possibly, through mechanical stress and the piezoelectric effect *’ the structural weakness resulting from osteomalacia may induce formation of new albeit poorly mineralized bone tissue which, in long standing cases, may ultimately mineralize sufficiently to provide normal or possibly increased bone mineral mass. There is some evidence that the osteosclerosis is a late sequela of this disease since Steinbach and Noetzli’ observed that osteosclerosis developed in three children only after prolonged observation. Clearly, the pathogenesis of osteosclerosis in familial hypophosphatemia is unknown but its investigation could lead to new understanding of bone physiology. ACKNOWLEDGMENT We wish to express calcium measurements.

our

appreciation

to Mr.

John

Watts

for technical

assistance

in the total

body

REFERENCES 1. Steinbach HL, Noetzli M: Roentgen pearance of the skeleton in osteomalacia rickets. Am J Roentgen01 Radium Ther Med 91:955-972, 1964

apand Nucl

2. Stanbury SW: Osteomalacia, in MacIntyre I (ed): Calcium Metabolism and Bone Disease, Clinics in Endocrinology and Metabolism, Vol 1. Philadelphia, Saunders, 1972, p 239

40

3. Pa&t AM: Hypophosphatemic vitamin D refractory rickets and osteomalacia. Orthop Clin North Am 3:653-680, 1972 4. McNeil] KG, Thomas BJ, Sturtridge WC, Harrison JE: In vivo neutron activation analysis for calcium in man. J Nucl Med 14:502-506, 1973 5. Fraser D, Salter RB: The diagnosis and management of the various types of rickets. Pediatr Clin 5:417-441, 1958 6. Fraser D: Rickets, in Conn HF, Conn RB (eds): Current Diagnosis. Philadelphia, Saunders, 1968, p 521-524 7. Greenberg BG, Winters RW, Graham JB: The normal range of serum inorganic phosphorus and its utility as a discriminant in the diagnosis of congenital hypophosphatemia. J Clin Endocrinol20:364-379, 1960 8. Paunier L, Kooh SW, Conen PE, Gibson AAM, Fraser D: Renal function and histology after long-term vitamin D therapy of vitamin D refractory rickets. J Pediatr 73:833-844, 1968 9. Sissons HA, Halley KJ, Heighway J: Normal bone structure in relation to osteomalacia, in Masson, Cie (eds): L’Osteomalacie. Symposium organist par le Centre du Metabolisme Phospho-calcique, publie par D. J. Hioco. 1967, p 19-37 10. McNeil1 KG, Kostalas HA, Harrison JE: Effects of body thickness on in vivo neutron activation analysis. Int J Appl Radiat Isotopes 25:347-353, 1974 Il. Harrison JE, Sturtridge WC, Williams C, Watts J, McNeil1 KG: A bone calcium index based on partial body calcium measurements by in vivo activation analysis. J Nucl Med 16:116-122, 1975 12. Woods CC, Morgan DB, Paterson CR,

HARRISON

ET Al.

Grossman HH: Measurement of osteoid in bone biopsy. J Pathol Bact 95:441-449, 1968 13. Reference Man, published for The International Commission on Radiological Protection by Pergamon Press (in press) 14. Garner A, Ball J: Quantitative observations on mineralized and unmineralized bone in chronic renal azotaemia and intestinal malabsorption syndrome. J Path01 Bact 91:545560, 1966 15. Meema HE, Rabinovich S, Meema S, Lloyd GJ, Oreopoulos DC: Improved radiological diagnosis of azotemic osteodystrophy. Radiology 102:lllO, 1972 16. Meema HE, Oreopoulos DC, Rabinovich S, Husdan H, Rapoport A: Periosteal new bone formation (periosteal neostosis) in renal osteodystrophy. Radiology 110:513-522, 1974 17. Aitken RE, Kerr JL, Lloyd HM: Primary hyperparathyroidism with osteosclerosis and calcification in articular cartilage. Am J Med 37:813-820, 1964 18. Templeton AW, Jaconette JR, Ormond RS: Localized osteosclerosis in hyperparathyroidism. Radiology 78:955-957, 1962 19. Rasmussen H, Bordier P: The Physiological and Cellular Basis of Metabolic Bone Disease. Baltimore, Williams & Wilkins, 1974, p 155-175 20. Stanbury SW: Azotaemic renal osteodystrophy, in Maclntyre (ed): Calcium Metabolism and Bone Disease, Clinics in Endocrinology and Metabolism, Vol. 1.) Philadelphia, Saunders, 1972, p 267 21. Bassett CAL: Biophysical principles affecting bone structure, in Bourne GH (ed): The Biochemistry and Physiology of Bone, vol III. New York, Academic Press, 1971, p l-76