Background

In successful renal transplantation, the degree of renal function recovery is usually incomplete and information is scarce about the abnormalities of mineral metabolism in long-term adult renal recipients with normal renal function. This study was designed to investigate bone mineral metabolism in patients with a long-term normal functioning kidney.

Methods

Twenty-nine adult asymptomatic renal transplant (RT) recipients with stable graft function for more than 10 years and serum creatinine <2 mg/dL were studied. They were classified into two groups according to glomerular filtration rate: Group A (N = 12; nine men, three women)>70 mL/min (x: 126 55 mL/min) and Group B (N = 17; nine men, eight women) <70 mL/min (x: 56 11 mL/min). Circulating biochemical markers of bone remodelling, bone histomorphometry, and densitometry (lumbar spine and hip) were obtained to investigate bone disease in these patients.

Results

Serum PTH was slightly elevated in 10 patients (83%) in group A. Serum PTH levels were positively related to serum calcium, osteocalcin, BAP, telopeptide, OH-proline, and creatinine. There was no histologic data to support overactivity on bone in this group of patients, with only one showing high bone turnover. Mineralization was prolonged in 34% of patients. Twenty-two patients (75%) exhibited normal bone turnover. In the group with GFR>70 mL/min the prevalence of mineralization defect in the presence of normal serum levels of calcitriol suggested vitamin D resistance. Lumbar and femoral neck osteoporosis was present in 25% and 33% of patients in group A, and 23% and 53% in group B, respectively. T-score at lumbar spine was negatively correlated with months since transplantation. Patients under treatment with cyclosporine (CsA) showed increased concentrations of osteocalcin and D-pyr and higher lumbar bone mineral density (BMD), but bone histomorphometry was not influenced by CsA.

Conclusion

Patients with long-term renal transplantation with normal renal function frequently present with slight increases in PTH, but without an effect on bone histology. CsA did not induce changes in bone histology and delayed mineralization was frequently observed.

METHODS

Subjects

Twenty-nine renal transplant recipients with stable graft function for more than 10 years and serum creatinine <2 mg/dL were studied. They were classified into two groups according to glomerular filtration rate. Diabetic patients were excluded from the study. None of the patients was treated with vitamin D metabolites or calcium supplements or had undergone previous parathyroidectomy. All patients provided written informed consent for participation and bone biopsy. Clinical characteristics are shown in Table 1.

Table 1 - Clinical characteristics.

Full table

Bone biopsies

Transiliac bone biopsies were collected from the anterior iliac crest after double tetracycline labeling and processed as described previously⁷. Bone turnover was defined according to activation frequency⁷.

Laboratory tests

All samples were obtained the day of bone biopsy. Immunoradiometric assays were used to determine intact PTH and osteocalcin. Bone alkaline phosphatase (BAP) and propeptide of type I procollagen-PICP were measured by immunoassay. Carboxyterminal telopeptide of type I collagen-ICTP and 25-hydroxyvitamin D₃ were measured by radioimmunoassay (RIA). Total urinary OH-proline was determinated with a colorimetric method and urinary deoxypyridinoline-Dpyr by enzymoimmunoassay. 1,25 dihydroxyvitamin D₃ was determinated by RIA.

Bone densitometry

Bone mineral density (BMD) was evaluated in the femoral neck and lumbar spine (L1 to L4) by dual energy radiographic absorptiometry using a DPX scanner (Hologic QDR-4500; Hologic Corp., Waltham, MA, USA). Osteopenia and osteoporosis prevalences were assessed according to World Health Organization (WHO) recommendations⁸.

Statistical analysis

Results are expressed as mean SD. Unpaired Student t test was used to compare intergroup diferences. Pearson's linear correlation was used to detect relationships between variables. Computations were performed using SPSS statistical software (SPSS, Inc., Chicago, IL, USA).

RESULTS

The biochemical data for Groups A and B (discussed below) are shown in Table 2.

Table 2 - Biochemical parameters of bone metabolism.

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Group A. In spite of normal renal function, mean serum PTH was slightly higher than reference normal values. It was elevated in 10 patients. One patient had hypercalcemia and seven had hypophosphatemia. Biochemical parameters of bone formation (osteocalcin, bone alkaline phosphatase, and PICP) and resorption (ICTP, D-pyridinoline, and OH-proline) were within normal limits. All patients showed normal levels of 1,25(OH)₂D₃, but two patients had low levels of 25(OH)D. Osteopenia was observed both at lumbar and more severe at femoral neck. Twenty-five percent of patients showed lumbar osteoporosis and 33% showed femoral neck osteoporosis. Bone histomosphometry studies Table 3 revealed high bone turnover in three patients, normal bone turnover in eight patients, and only one patient exhibited low bone turnover. Delayed mineralization was observed in three patients and mild aluminum deposits in one.

Table 3 - Histomorphometric parameters.

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Group B. Mean serum PTH was elevated in 11 patients; only in six patients did it remain in normal range. Three patients were hypercalcemic and five showed hypophosphatemia. Bone remodeling parameters were within normal range. 1,25(OH)₂D₃ was below the normal range in six patients. All patients exhibited normal levels of 25(OH)D. Patients exhibited osteopenia both at lumbar spine and at femoral neck. Neck osteoporosis was present in 53% of patients and lumbar osteoporosis in 23% of patients. In histomorphometric studies Table 3, two patients had low bone turnover, 14 had normal bone turnover, and one had high bone turnover. Seven patients showed delayed mineralization. Mild aluminum deposits were found in one patient and peritrabecular fibrosis was found in one patient.

Bone turnover marker correlations

Serum PTH levels were positively related to serum calcium (r = 0.44, P < 0.05), osteocalcin (r = 0.40, P < 0.05), bone alkaline phosphatase (r = 0.42, P < 0.05), ICTP (r = 0.42, P < 0.05), OH-proline (r = 0.42, P < 0.05), and serum creatinine (r = 0.49, P < 0.01). There were no significant correlations between serum PTH and histomorphometric parameters.

Osteocalcin correlated significantly with all the other markers. Correlations between osteocalcin and BAP and propeptide as markers of bone formation reached statistical significance within the group as a whole (r = 0.40, P < 0.05; r = 0.50, P < 0.01, respectively). OH-proline and D-Pyr as markers of bone resorption also correlated significantly (r = 0.61, P < 0.001). In contrast, calcitriol levels did not correlate significantly with PTH or other biochemical markers.

Negative relationships were also observed between bone volume and telopeptide (r = -0.47, P < 0.05), OH-proline (r = -0.39, P < 0.05), and D-Pyr (r = -0.40, P < 0.05). Osteoclast surface was positively correlated with osteocalcin (r = 0.37, P < 0.05), telopeptide (r = 0.42, P < 0.05), and D-pyr (r = 0.40, P < 0.05). Calcitriol levels correlated with bone formation rate, activation fraction, and mineralizing surface (P < 0.05). No significant correlations were found between PTH, BAP, and propeptide with histomorphometric parameters.

No significant correlations were found between BMD and biochemical or histologic parameters of bone activity, with the exception of a negative relationship between T-score at the femoral neck and PTH (r = -0.43, P < 0.05). T-score at the lumbar spine was also correlated negatively with months since transplantation (r = -0.40, P < 0.05).

Immunosuppressive therapy

The 29 renal transplant patients were classified according to the immunosuppressive therapy received: CsA group (19 patients) and non-CsA group (10 patients) Table 4. Histomorphometric data were not different between the two groups.

Table 4 - Results according to immunosuppressive therapy.

Full table

DISCUSSION

This study was designed employing circulating biochemical markers of bone remodeling, bone histomorphometry, and densitometry to investigate bone disease in long-term adult renal recipients with normal renal function.

Persistent hyperparathyroidism was detected in 43% of long-term renal transplant patients with a serum creatinine <1.5 mg/dL⁹. In this study, according to activation frequency, only three patients out of 12 with normal renal function exhibited high bone turnover. Therefore, although past studies have reported a high frequency of parathyroid overactivity in renal transplant patients based on PTH levels, there is no histologic data to support overactivity on bone. Corticosteroids most likely counterbalance the bone effects of PTH.

Studies in renal transplant recipients measuring vitamin D metabolites have given conflicting results¹⁰. In the present study, in the group with GFR>70 mL/min, all patients had levels of 1,25(OH)D within the normal range, indicating the success of transplantation in restoring 1-hydroxylase activity. On the other hand, in the group with GFR <70 mL/min, six patients had levels of 1,25(OH)D below the normal range, with normal levels of 25(OH)D. Reduction in 1-hydroxylase activity reported in early renal insufficiency explains this observation¹¹.

The main histologic characteristic in the whole group of patients was a normal bone turnover. Our results are in agreement with the study of Sanchez et al¹² of a pediatric transplanted population, but differ from other studies in which the predominant lesion is a decrease in bone turnover^13,14,15. One important finding is the prevalence of mineralization defect in 34% of patients. The presence of normal serum levels of calcidiol and calcitriol in the absence of significant aluminum deposits suggests vitamin D resistance. This alteration in mineralization has been previously described^13,16. Although hypophosphatemia, present in 13 patients, could lead to a possible mineralization defect, we did not find differences in mineralization status between patients with normal and hypophosphatemia. Because the mineralization defect occurs in the presence of normal circulating vitamin D metabolites, it is conceivable that the apparent resistance of bone cells to vitamin D is due to abnormal response of its receptor (VDR) or postreceptor defect¹³.

Corticosteroid cumulative dose is considered the most important factor involved in bone mass reduction after transplantation^17,18,19. Corticosteroid bone loss affects predominantly trabecular bone³. Our study, performed at least 10 years after transplant, displays a mild reduction in BMD in cancellous bone (lumbar spine), and is more severe in cortical bone (femoral neck), where up to 43% of patients showed osteoporosis. Because most of our patients have some degree of elevation of PTH, this may represent the mechanism that underlies the preferential cortical bone loss

It has been suggested that CsA increases bone turnover by inhibiting antiresorptive cytokines²⁰. However, the clinical data in renal transplant (RT) do not support an adverse effect of CsA on bone mass because CsA monotherapy does not reduce BMD 12 to 18 months after grafting²¹. Data in humans suggest that cyclosporine may decrease the incidence of osteopenia in RT recipients because of the use of lower doses of prednisone²². Our CsA patients exhibited an increase in bone formation and resorption markers, suggesting higher bone turnover. However, this was not confirmed by histopathologic studies.

Residual hyperparathyroidism related to abnormal renal function and steroid-induced osteoporosis is a metabolic finding in renal transplant patients. Delayed mineralization, the histologic predominant observation, is probably more prevalent in the absence of hyperparathyroidism.

References

1. Hruska, KA, Teitelbaum, SL: Renal osteodystrophy. N Engl J Med 1995 333: 166–174,  | Article | PubMed | ISI | ChemPort |
2. Messa, P, Sindici, C, Canella, G, et al: Persistent secondary hyperparathyroidism after renal transplantation. Kidney Int 1998 54: 1704–1713,  | Article | PubMed | ISI | ChemPort |
3. Canalis, E: Mechanisms of glucocorticoid action in bone: Implications to glucocorticoid-induced osteoporosis. J Clin Endocrinol Metab 1969 81: 3441–3447,
4. Movsowitz, C, Epstein, S, Fallon, M, et al: Cyclosporin-A in vivo produces severe osteopenia in the rat: Effect of dose and duration of administration. Endocrinology 1988 123: 2571–2577,  | PubMed | ISI | ChemPort |
5. Westeel, FP, Mazouz, H, Ezaituni, F, et al: Cyclosporine bone remodeling effect prevents steroid osteopenia after kidney transplantation. Kidney Int 2000 58: 1788–1796,  | Article | PubMed | ISI | ChemPort |
6. Aroldi, A, Tarantino, A, Montagnino, G, et al: Effects of three immunosuppressive regimens on vertebral bone density in renal transplant recipients. Transplantation 1997 63: 380–386,  | PubMed | ISI | ChemPort |
7. Serrano, S, Mariñoso, ML, Torres, A, et al: Osteoblastic proliferation in bone biopsies from patients with end stage chronic renal failure. J Bone Miner Res 1997 12: 191–199,  | PubMed | ISI | ChemPort |
8. World Health Organization: Assessment of fracture risk and its application screening for postmenopausal osteoporosis. Technical report series. 1994, Geneva, WHO, p 843
9. Torres, A, Zarraga, S, Rodriguez, A, et al: Optimum PTH levels before renal transplantation to prevent persistent hyperparathyroidism [abstract]. J Am Soc Nephrol 1998 9: 572A, .
10. Riancho, JA, de Francisco, ALM, del Arco, C, et al: Serum levels of 1,25-dihydroxyvitamin D after renal transplantation. Miner Electrolyte Metab 1988 14: 332–337,  | PubMed | ISI | ChemPort |
11. Portale, AA, Booth, BE, Tsai, HC, Morris, RC: Reduced plasma concentration of 1.25 dihydroxyvitamin D in children with moderate renal insufficiency. Kidney Int 1982 21: 627–632,  | PubMed | ISI | ChemPort |
12. Sanchez, CP, Salusky, IB, Kuizon, BD, et al: Bone disease in children and adolescents undergoing successful renal transplantation. Kidney Int 1998 53: 1358–1364,  | Article | PubMed | ISI | ChemPort |
13. Monier-Faugere, MC, Mawad, H, Qi, Q, et al: High prevalence of low bone turnover and occurrence of osteomalacia after kidney transplantation. J Am Soc Nephrol 2000 11: 1093–1099,  | PubMed | ISI | ChemPort |
14. Velasquez-Forero, F, Mondragón, A, Herrero, B, Peña, JC: Adynamic bone lesion in renal transplant recipients with normal renal function. Nephrol Dial Transplant 1996 11: 58–64,  | PubMed |
15. Cueto-Manzano, AM, Konel, S, Hutchison, AJ, et al: Bone loss in long-term renal transplantation: Histopathology and densitometry analysis. Kidney Int 1999 55: 2021–2029,  | Article | PubMed | ISI | ChemPort |
16. Julian, BA, Laskow, DA, Dubovsky, J, et al: Rapid loss of vertebral mineral density after renal transplantation. N Engl J Med 1991 325: 544–550,  | PubMed | ISI | ChemPort |
17. Pichette, V, Bonnardeaux, A, Prudhomme, L, et al: Long term bone loss in kidney transplant recipients: A cross-sectional and longitudinal study. Am J Kidney Dis 1996 28: 105–114,  | PubMed | ISI | ChemPort |
18. Grotz, WH, Mundinger, FA, Rasenack, J, et al: Bone loss after kidney transplantation: A longitudinal study in 115 graft recipients. Nephrol Dial Transplant 1995 10: 2096–2100,  | PubMed | ISI | ChemPort |
19. Wolpaw, T, Deal, CL, Fleming-Brooks, S, et al: Factors influencing vertebral bone density after renal transplantation. Transplantation 1994 58: 1186–1189,  | PubMed | ISI | ChemPort |
20. Sprague, SM: Mechanism of transplantation-associated bone loss. Pediatr Nephrol 2000 14: 650–653,  | Article | PubMed | ISI | ChemPort |
21. Torregrosa, JV, Campistol, JM, Montesinos, M, et al: Evolution of bone mineral density after renal transplantation. Nephrol Dial Transplant 1995 10(Suppl 6):111–113,  | PubMed | ISI |
22. Westeel, FP, Mazouz, H, Ezaituni, F, et al: Cyclosporine bone remodeling effect prevents steroid osteopenia after kidney transplantation. Kidney Int 2000 58: 1788–1796,  | Article | PubMed | ISI | ChemPort |