The American Journal of PATHOLOGY MARCH 1975 * VOLUME 78, NUMBER 3
Metabolic Bone Disease in Chronic Renal Failure I. Dialyzed Uremics W. E. Huffer, MD, D. Kuzela, MD and M. M. Popovtzer, MD
Garner and Ball's point counting technic was used to compare metabolic bone disease in dialyzed and nondialyzed uremic patients. Histologic measurements of bone from dialyzed and nondialyzed uremic patients dying between 1966 and 1971 showed that dialyzed patients have quantitatively more severe bone resorp. tion, distortion of trabecular architecture and mineralization defects. Mineralization defects become more severe as the duration of dialysis increases but are not related to serum calcium and phosphorus levels. Bone volume in both groups is normal or increased and in dialysis patients increases in proportion to the elevation of serum phosphorus. Mean serum phosphorus and calcium levels, bone volume, and volume:surface ratios all decreased in dialysis patients between 1966 and 1971, while bone resorption and mineralization defects did not change. These results suggest that lowering of serum phosphorus without increasing serum calcium may aggravate the uremic bone disease by reducing bone volume without improvement of mineralization and resorption defects. (Am J Pathol 78:365-384, 1975)
MAINTENANCE HEMODIALYSIS has been successful in prolonging the life of patients suffering from chronic renal failure, but morbidity due to metabolic bone disease has not been eliminated.1'2 QuantiFrom the Departments of Pathology and Medicine, University of Colorado Medical Center, Denver, Colo. Presented in part at the Seventy-first Annual Meeting of the American Association of Pathologists and Bacteriologists, San Francisco, Calif, March 10, 1974. Supported in part by Training Grant GM-00977 from the US Public Health Service. Accepted for publication October 10, 1974. Address reprint requests to Dr. William E. Huffer, Department of Pathology, School of Medicine, University of Colorado Medical Center, 4200 East Ninth Ave, Denver, CO 80220. 365
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tative histopathologic studies of bone disease in nondialyzed-7 and dialyzed -22 patients have produced conflicting data about the occurrence of major types of bone abnormalities and hence different conclusions about the relationship between predialysis renal osteodystrophy and dialysis boine disease. The most prevalent patterns of bone response to chronic renal failure reported in different studies seem to be determined not only by common- pathogenetic responses to altered blood mineral levels, hormone secretion and vitamin utilization associated with renal failure, but also to variations in baseline mineral and bone homeostasis in the populations studied. These are influenced by the diet, climate and age of the individuals developing renal failure.23 In dialysis patients, differences in the specific therapeutic regimens may be expected to add to the many variables that determine the major patterns of bone response.24 The University of Colorado Medical Center has had an active maintenance hemodialysis program for several years. Presumably, the population treated at this center has not changed dramatically during this time. However, the details of the therapeutic regimen have changed, not only with respect to dialysis procedures, but also with respect to auxillary therapy such as the institution of oral phosphate-binding therapy in 1970. Since differences in the form of dialysis bone disease reported from different treatment centers might be related to differences in therapy, it seemed appropriate to ask if there were any changes in the form of bone disease seen in dialysis patients treated at this center during a time when therapy was in a state of evolution. This quantitative histopathologic study was designed to answer that question and to try to identify any specific changes in therapy which might explain changes in the form of bone disease. Quantitative indices of bone resorption, mineralization and bone volume in autopsy specimens of bone from dialyzed, nondialyzed and normal control patients were compared to establish the similarities and dissimilarities between conventional renal osteodystrophy and dialysis bone disease in patients treated at this medical center. The relationships between quantitative indices of bone morphology, parathyroid weight, antemortem serum calcium and phosphate levels, and duration of dialysis were investigated by statistical methods. Finally, the relationships of quantitative indices of bone morphology, antemortem serum mineral levels and duration of dialysis to time of institution of dialysis therapy or the time the patient died were investigated by statistical methods.
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Materials and Methods Sections of lumbar vertebrae were obtained from 44 dialysis patients, 15 conservatively treated uremics, and 20 normal control patients autopsied at Colorado General Hospital or the Denver Veterans Hospital between January 1966 and December 1971. The dialysis group was subdivided into a short-term group (9 patients, 0 to 6 months dialysis) and a long-term group (35 patients, >6 months dialysis). Conservatively treated patients had chronic renal failure not treated by maintenance hemodialysis or renal transplantation. Normal control patients were individuals without renal disease dying suddenly as a result of trauma. Twenty patients from a large pool of such individuals were matched by age and sex with 20 patients selected from 94 patients with renal disease using a table of random numbers. (Patients with renal failure included those in this study plus a group of patients treated by renal transplantation; lesions in the transplant patients are described in a subsequent paper 25). Specimens of lumbar vertebral bone were fixed for 24 hours in 10% neutral butfered formalin and decalcified in acid citrate. The major variable in identifying osteoid in decalcified hematoxylin- and eosin-stained sections is the decalcification time. Decalcified tissue was checked three times a day until a sharp pin could first be made to penetrate the cortical bone, which still felt hard to palpation. At this stage decalcification was sufficient to allow sectioning and still maintain the clearcut definition between basophilic (mineralized) and eosinophilic (nonmineralized) bone matrix in hematoxylin- and eosin-stained sections. After decalcification, specimens were processed routinely for paraffin embedding, sectioned at 7 to 10 ji and further processed for hematoxylin and eosin staining. Sections were stained with Harris hematoxylin (Harleco), pH 2.8, for 5 minutes and counterstained with eosin for 45 seconds. Sections of cancellous bone, 0.25 sq cm, were evaluated by a modification of Garner and Ball's point counting method.s Twenty-five nonoverlapping fields, encompassed by a square grid with 121 intersections, were counted resulting in a total count of 3,025 intersections/specimen. Parameters counted included calcified bone, osteoid, internal trabecular resorption spaces, and internal and extemal trabecular surfaces. The data were then used to calculate the following parameters: Parameters measuring resorptive activity and distortion of trabecular architecture Resorptive volume (RV) = internal resorption space counts/3025 Resorptive index (RI) = internal resorption space counts/calcified bone and osteoid counts Volume to surface ratio (V: S) = calcified bone and osteoid counts/surface counts
Parameters measuring changes in volume Total bone volume (BV) = calcified bone and osteoid counts/3025 Calcified bone volume (Ca++ BV) = calcified bone counts/3025 Parameter measuring mineralization defects Osteoid index (OI) = osteoid counts/calcified bone and osteoid counts Reproducibility of counts on a single specimen was approximately + 5%, similar to the results of Ellis and Peart.6 Mean values, standard deviations and standard errors for each of these parameters were calculated, and differences between groups were tested for by a standard t test applicable to two random
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of Pathology
groups of different sizes.268 The percentage of individuals in each renal disease group who fall within the normal control mean ± 2 SD or within specified limits above or below this range for several of the parameters was also calculated. Correlations between different parameters of bone histology or between a given parameter of bone histology and clinical or anatomic measurements such as duration of dialysis, parathyroid weight or serum phosphorus levels were tested by regression analysis using the University of Colorado Basic Library Computer Program, Stat 10. This program computes the slope and other statistics including the F ratio for a simple linear regression with one independent variable. Parathyroid weights were determined gravimetrically at the time of autopsy. Antemortem serum calcium, phosphate and Ca x P04 product levels were the mean of all predialysis determinations done during a patient's entire course of maintenance hemodialysis. For individual patients, the standard deviations of mean predialysis Ca and P04 values were approximately + 1 mg%, and there were no significant deviations above or below the means as a function of duration of dialysis. Parathyroid weights, serum mineral levels and quantitative indices of bone morphology were available in a group of 10 long-term dialysis patients whose indices of bone morphology are representative of the entire group of long-term dialysis patients. Mean antemortem serum mineral levels were available from 8 additional long-term dialysis patients (including 1 patient with quantitative bone indices) autopsied between 1966 and 1971. These 8 patients plus the 10 in the previous group were used to determine the relationship between serum mineral levels and the relative time of autopsy by regression analysis. The time of autopsy of the first dialysis patient was January 1966, and the time of autopsy of the last was December 1971. Regression analyses of bone indices and serum mineral levels as a function of relative time of autopsy have number of months after January, 1966, as the X axis. The earliest time of starting a maintenance dialysis program by a patient in this study was in 1965. The time axis for regression analysis of duration of dialysis as a function of relative time of starting maintenance dialysis was the number of months after January 1965, when the patient was started on the dialysis program. Since this was a retrospective study, the bone specimens used were all decalcified, and the von Kossa technic could not be used to determine osteoid index. As noted above, with careful decalcification and staining, osteoid could be identified in decalcified hematoxylin- and eosin-stained specimens by its relative eosinophilia, homogeneity and its relationship to either a calcification front or a cement line and adjacent basophilic bone matrix. In this study osteoid was consistently lamellar. To determine the accuracy of these criteria for identifying osteoid, each of five surgical bone biopsies from patients in another prospective study were divided into two portions. One portion was used to prepare frozen undecalcified sections which were stained by the von Kossa technic.27 The other was processed routinely after decalcification and stained with hematoxylin and eosin. The osteoid index was determined on all ten preparations in a double blind study. The results of the determinations on paired samples from the same biopsies are compared in Table 1. In addition to the quantitative parameters, a semiquantitative grading system was used to evaluate the degree of bone resorption. Grade 0 resorption was defined as trabeculae with smooth contours; Grades 1 to 4 are illustrated and defined in Figure 1.
Results Methodology
The osteoid index of five surgical biopsies as determined on vonKossa-stained sections was indistinguishable from osteoid index as
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Table 1-Osteoid Index on von-Kossa-Stained Undecalcified Sections and Hematoxylinand Eosin-Stained Decalcified Sections of the Same Biopsies von Kossa H&E Sample Difference 1 3.0 3.8 -0.8 2 1.2 0.8 +0.4 3 7.7 7.4 +0.3 4 0.0 0.0 0.0 5 0.6 0.5 +0.1 Mean 2.5 2.5 0.0
determined by histologic criteria applied to decalcified hematoxylinand eosin-stained sections of the same biopsies (Table 1). There was good correspondence between resorption grade as determined by established histologic criteria of bone resorption and the parameters used to quantitate bone resorption -(Text-figure 1). As the resorption grade increased, the mean V:S ratio decreased, and the resorption volume and resorptive index increased. Resorptive Activity and Distortion of Trabecular Architecture
Both dialyzed and nondialyzed patients had increased resorptive activity as judged by semiquantitative methods (Text-figure 2). None of the normal controls had significant resorption (100% Grade 0). All nondialyzed patients had significant resorption with an equal distribu20
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TEXT-FIG 1-Relationships among resorptive grade and V: S ratio (A), resorptive index x 10-2 (B) and RV x 3025 (C). The mean SE is shown for each resorptive grade. Figures include uremic groups from this study, a group of patients with renal transplants described in another study,25 plus the controls. The number of individuals in Grade 0 was 31; in Grade 1, 27; in Grade 2, 22; in Grade 3, 24; and in Grade 4, 10. Grade 4 lesions occur only in transplant patients.
HUFFER ET AlL
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tion among Grades 1 to 3. One-fifth to one-fourth of the dialyzed group had no increased resorption. The short-term dialysis patients had the lowest incidence of Grade 3 resorption. In contrast, the long-term dialysis group had a significant percentage of individuals with Grade 3 resorption. None of the patients included in this study had Grade 4 resorption. Differences in mean value of RV and RI between patient groups were similar (Table 2). The mean values of RV for all renal disease groups and of RI for the long-term dialysis group were significantly higher than the control. mean (P < .005). There were no significant differences in RV or RI between the nondialyzed patients and the combined dialysis group, but the means of these two resorptive parameters for long-term dialysis patients were significantly greater than those for short-term dialysis patients (P < .005). The short-term dialysis group had the greatest percentage of individuals within the normal range of RI and had no individuals whose
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in the short-term dialysis group (Text-figure 3). The incidence of severely reduced ratios (one-fifth to three-fifths normal) was also greater in the long-term group. Changes in Bone Volume
Mean bone volumes of all renal disease groups were slightly greater than the normal mean (Table 2). While the majority of patients within these renal disease groups had bone volumes within the normal range (Text-figure 3), all groups had individuals with increased bone volume (22 to 27% of each group). Nondialyzed and long-term dialysis groups had only small numbers of individuals with reduced bone volume. None of the short-term dialysis patients bad reduced bone volume. No significant relationships between bone volume and age (in normal controls or any renal disease group), duration of dialysis, osteoid index, or semiquantitative or quantitative resorption indices were found. Mineralization Defects
All renal disease groups had increased mean osteoid indices in comparison to the normal control group (Table 2). The elevation in mean osteoid index was statistically significant at P < .005 for long- and short-term dialysis groups and at P between .05 and .025 for the nondialyzed group. Elevations of osteoid index were more extreme in dialysis patients. The mean osteoid index for combined long- and short-term dialysis patients was significantly greater than the mean OI of the nondialyzed group (P < .005). Osteoid index in dialysis patients increased as a function of duration of dialysis (Text-figure 4) and as a function of age (slope 2.9% per year, P < .005). In contrast, there was no increase 40
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Discussion
The major abnormalities of bone reported in nondialyzed patients with chronic renal failure include excessive bone resorption, or osteitis fibrosa 3,4,6,7,9, 11,12,15,1720; mineralization defects, or osteomalacia 3,57,11, 12,15,17-19; and increased bone volume, or osteosclerosis.3 4'6'7"0""'2"8 Decreased bone volume or osteoporosis is rare in nondialyzed patients,19 20 but at least one contradictory report had appeared.7 One study from Australia found no significant mineralization defects in nondialyzed patients.4 Conflicting views on the relative prevalance and time of onset of osteomalacia and osteitis fibrosa have also been reported.'628 Many of these discrepancies may be reconciled by Parfitt's conclusion that the course of renal osteodystrophy will be influenced by the age of onset of renal failure and sociologic and geographic differences in vitamin D nutrition.23 The major forms and the course of dialysis bone disease are equally variable. Most studies of dialysis bone disease have found a significant degree of osteomalacia. 12'1419'22 and some have reported progression of mineralization defects during hemodialysis.""1,2"l4"15'22 In contrast, one study found no significant mineralization defects in dialysis patients,'3 and several studies reported no progression of osteomalacia with duration of dialysis.16'18"9 Increased bone resorption occurs frequently in dialysis patients.8'9"1 13,1519,21,22 Some studies report progression of bone resorption during the course of dialysis,9" 3"17'21 whereas others found no change or decreased bone resorption.11'12"15"18'19'22 Decreased 8"12"13"15'16'20'21 normal,8,10,12,17-19 and increased 8,11,12 total bone volume has been reported in dialysis patients. Many workers stress selective reduction in mineralized bone volume as a unique feature of dialysis bone disease.10 12'15 Conclusions about the nature of dialysis bone disease and its relationship to conventional renal osteodystrophy are as divergent as the data reported above. Dialysis bone disease, or at least some forms of dialysis bone disease, have been regarded as a) new patterns of renal osteodystrophy with progression of osteomalacia and a lessening of osteitis fibrosa;" b) disuse osteoporosis with mineralization defects not specific for dialysis therapy;21 c) a disease quantitatively different from renal osteodystrophy because of progressive loss of mineralized bone, increased osteoid and a change in character of osteitis fibrosa;'2 d) a dialysis osteopenia differing from osteitis fibrosa, osteoporosis, and osteomalacia;20 e) a progression of conventional renal osteodystrophy with (in some patients) a less familiar tendency to develop osteopo-
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rosis;'5 or f) a disease essentially identical to conventional renal osteodystrophy. 17-19
The literature thus indicates that the form and course of both conventional renal osteodystrophy and dialysis bone disease will vary from individual to individual, and from one patient population to another. The present study confirms the individual variability of bone pathology in both dialyzed and nondialyzed patients as shown by the large standard deviations of all quantitative parameters of bone morphology studied. It adds another set of quantitative data to the literature, illustrating the major patterns of bone disease in dialyzed and nondialyzed uremics at a single treatment center. The data show that at this center dialysis bone disease is qualitatively similar, but quantitatively more severe than conventional renal osteodystrophy. Both groups have increased bone resorption, mineralization defects, and a normal or increased bone volume. Dialysis patients, however, have significantly more severe mineralization defects which progress with increasing duration of dialysis. Bone resorption, as measured by resorptive index, resorptive volume, and volume: surface ratios is somewhat more severe in dialysis patients, and although there is no linear increase in any of these indices with duration of dialysis, the data on percentage distributions of resorptive defects show that bone resorption becomes more frequent and severe as dialysis is prolonged. The new data presented in this study relates patterns of bone disease, serum mineral levels and duration of dialysis to the relative time of treatment and death during the interval between 1965 and 1971. Several facts were established. First, patients dying near the end of this time interval did not survive dialysis for significantly longer periods than those dying near the beginning of the interval. This does not necessarily imply that more recent dialysis procedures did not increase longevity, since the study does not include patients still surviving. Second, total bone volume and the volume: surface ratio, both presumably measures of the structural integrity of the bone, decreased as mean serum phosphorus and/or Ca X P04 products decreased. Third, the serum calcium, phosphorus and Ca X PG4 products of dialysis patients decreased during the period from 1966 through 1971. Fourth, the bone volume and volume: surface ratio of dialysis patients also decreased during this period, but the osteoid index, resorptive volume and resorptive index did not change. Fifth, all of the long-term dialysis patients whose parathyroid weights were known had hyperplastic glands. These observations may have bearing on the relationship between
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conventional renal osteodystrophy and dialysis bone disease, and the variability in the form of bone disease seen at different dialysis centers. They imply that therapy aimed at reducing serum phosphate levels without precise control of serum calcium levels may change the form of bone disease, resulting in a decrease in bone volume without correcting resorption and mineralization defects. Bone resorption in renal osteodystrophy is generally acknowledged to result from excessive parathormone production by hyperplastic parathyroid glands responding to lowered serum calcium levels.6'23'24'29'30 Hyperparathyroidism as demonstrated in this study is frequent in longterm dialysis patients,3' and measurements of serum parathormone levels in dialysis patients have shown a transient decline at the onset of dialysis followed by a progressive increase in chronic dialysis patients.30 There is good reason to believe, therefore, that the increased bone resorption seen in dialysis patients in this study resulted from hyperparathyroidism which was not effectively controlled in the period between 1966 and 1971. Elevated phosphate levels have been shown to decrease parathormone-induced resorption in vitro.33 However, in normal rats, phosphate infusions do not cause decreased hydroxyproline excretion or mobilization of 90Sr in response to parathormone. They cause retention of calcium by bone and/or soft tissues and increase bone formation.34 These observations suggest at least two mechanisms to account for the association between decrease in bone volume and V:S ratio and decreased mean serum phosphate levels found in this study. One is a loss of stimulation to bone formation, and the other is a loss of suppression of parathormone-induced bone resorption. In either case, decreases in bone volume and V: S ratio would be predicted. Equally important in this connection is the observation that bone volume, V: S ratio and mean serum phosphate levels all decreased during the period between 1966 and 1971. Bone volume and V: S ratio changes might be explained by the same phosphate-regulated mechanisms. Since mean serum calcium levels also fell during this period, parathyroid mechanisms also may have contributed to decreasing bone volume. It is not possible to determine from this study what changes in therapy brought about decrease in mean serum phosphorus levels. It seems likely that the institution of oral phosphate-binding therapy in 1970 is implicated. This form of therapy has been used in conjunction with increased concentrations of dialysate calcium to decrease parathormone production and to reverse osteitis fibrosa in dialysis patients.34 It has also been used to reverse metastatic calcification in dialysis patients with
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osteitis fibrosa.36 If lowered serum phosphate levels are implicated in decreased bone volume and V: S ratios, it might be important to maintain higher calcium levels,34 a form of therapy shown to be efficacious by the Mayo Clinic studies.34 Lack of correlations between osteoid index, serum mineral levels or the time the patients died during a period when serum mineral levels were changing, suggest that osteomalacia in dialysis patients is related to some specific effect of dialysis. This suggestion is supported by the demonstration of a statistically significant increase in severity of osteomalacia with increased duration of dialysis in this study, and also by clinical studies showing that osteomalacia in dialysis patients does not respond favorably to appropriate vitamin D therapy that will produce beneficial results in nondialyzed patients.24 Among the factors suggested as possible causes of impaired mineralization in dialysis patients are fluorides 37 and/or hypermagnesemia.38 References 1. Pendras JP, Erickson RV: Hemodialysis: A successful therapy for chronic uremia. Ann Intern Med 64:293-311, 1966 2. Ritz E, Krempien B, Mehls 0, Malluche H, Stroble Z, Zimmerman H: Skeletal complications of renal insufficiency and maintenance dialysis. Nephron 10:195-207, 1973 3. Gamer A, Ball J: Quantitative observations on mineralized and unmineralized bone in chronic renal azotemia and intestinal malabsorption syndrome. J Pathol Bacteriol 91:545-561, 1966 4. Ireland AW, Cameron DA, Stewart JH, Posen S: Quantitative histology of bone in advanced renal failure. Calcif Tiss Res 4:282, 1969 5. Binswanger U, Fischer J, Schenk R, Merz W: Osteopathie bei chronischer Niereninsuffizienz. Dtsch Med Wochenschr 96:1914-1919, 1971 6. Ellis HA, Peart KM: Azotaemic renal osteodystrophy: A quantitative study of iliac bone. J Clin Pathol 26:83-101, 1973 7. Ingham JP, Stewart JH, Posen S: Quantitative skeletal histology in untreated end-stage renal failure. Br Med J 2:745-748, 1973 8. Kim D, Bell NH, Bundeson W, Putong P, Simm NM, Walker C, del Greco F: Renal osteodystrophy in the course of periodic dialysis for chronic uremia. Trans Am Soc Artif Intern Organs 14:367-371, 1968 9. Jowsey J, Massry SG, Cobum JW, Kleenan CR: Microradiographic studies of bone in renal osteodystrophy. Arch Intern Med 124:539-543, 1969 10. Kerr DNS, Walls J, Ellis H, Simpson W, Uldall PR, Ward MK: Bone disease in patients undergoing regular hemodialysis. J Bone Joint Surg 51B:578, 1969 11. de Verber GA, Oreopoulos DG, Rabinovich S, Lloyd GJ, Mesma HE, Beattis BC, Levy D, Hudsan H, Rappaport A: Changing patterns of renal osteodystrophy with chronic hemodialysis. Trans Am Soc Artif Intern Organs 16: 479-485, 1970
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12. Ellis HA, Peart KM: Renal osteodystrophy with particular reference to the effects of chronic intermittent hemodialysis. Nephron 8:402, 1971 13. Kuhlencordt F, Kruse HP, Lozano-Tonkin C: Histomorphometric studies of bone in patients on chronic hemodialysis. Isr J Med Sci 7:517, 1971 14. Ritz E, Krempien B, Kuhn H, Heuck F: Dialysis bone disease. Isr J Med Sci 7:520, 1971 15. Bishop MC, Woods CG, Oliver DO, Ledingham JGG, Smith R, Tibbut DA: Effects of hemodialysis on bone in chronic renal failure. Br Med J 3:664-667, 1972 16. Delling G: Quantitative analysis of bone changes during chronic hemodialysis. Verh Dtsch Gesamte Pathol 56:436-438, 1972 17. Duursma SA, Visser WJ, Nijo L: A quantitative histological study of bone in thirty patients with renal insufficiency. Calcif Tiss Res 9:216-225, 1972 18. Krempien B, Ritz E, Beck U, Keilbach H: Osteopathy in maintenance hemodialysis. Virchows Arch [Pathol Anat] 357:257-274, 1972 19. Krempien B, Ritz E, Hauck F: Osteopathy in maintenance hemodialysis. Verh Dtsch Gesamte Pathol 56:439-442, 1972 20. Parfitt AM, Massry SG, Winfield AC: Osteopenia and fractures occurring during maintenance hemodialysis: A new form of renal osteodystrophy. Clin Orthop 87:287-302, 1972 21. Woods CG, Bishop MC, Nicholson GD: Bone histologic changes occurring after hemodialysis treatment for chronic renal failure. J Pathol 107:137-143,
1972
22. Binswanger U, Sherrard D, Rich C, Curtis FK: Dialysis bone disease. Nephron 12:1-9, 1973 23. Parfitt AM: Renal osteodystrophy. Orthop Clin N Am 3:681-698, 1972 24. Rubini ME, Cobun JW, Massry SG, Shinaberger JH: Renal osteodystrophy: Some therapeutic consideration relative to long-term dialysis and transplantation. Arch Intern Med 124:663-669, 1969 25. Huffer WE, Kuzela D, Popovtzer MM, Starzl T: Metabolic bone disease in chronic renal failure. II. Renal transplant patients. Am J Pathol 78:385-400,
1975
26. Snedecor GW: Statistical Methods Applied to Experiments in Agriculture and Biology. Ames, Iowa, Iowa State College Press. Chapter 4, 1959 27. Pearse AGE: Histochemistry: Theoretical and Applied, Second edition. Boston, Little, Brown and Co, p 934, 1961 28. Stanbury SW: Bone disease in uremia. Am J Med 44:714-724, 1968 29. Stanbury SW, Lumb GA: Parathyroid function in chronic renal failure: A statistical survey of the plasma biochemistry in azotaemic renal osteodystrophy. Q J Med 35:1-23, 1966 30. Bricker NS, Slatopolsky E, Reiss E, Avioli LV: Calcium, phosphorus, and bone in renal disease and transplantation. Arch Intern Med 123:543-553, 1969 31. Vosik WM, Anderson CF, Steffee WP, Johnson WJ, Arnaud CD, Goldsmith RS: Successful management of osteitis fibrosa due to "tertiary" hyperparathyroidism. Mayo Clin Proc 47:110-113, 1972 32. Johnson JW, Hattner RS, Hampers CL, Bernstein DS, Merrill JP, Sherwood LM: Effect of hemodialysis on secondary hyperparathyroidism in patients with chronic renal failure. Metabolism 21:18-29, 1972 33. Raisz LG, Nieman I: Effect of phosphate calcium and magnesium on bone
382
34.
35. 36.
37. 38.
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resorption, and hormonal responses in tissue culture. Endocrinology 85:446452, 1969 Pechet MM, Bobadilla E, Carroll EL, Hesse RH: Regulation of bone resorption and formation: Influences of thyrocalcitonin, parathyroid hormone, neutral phosphate and vitamin D3. Am J Med 43:696-710, 1967 Goldsmith RS, Furszyfer J, Johnson WJ: Differences between acute and chronic suppressibility of parathyroid function during chronic hemodialysis. J Clin Invest 50:37A, 1971 Pingerra WF, Popovtzer MM: Uremic osteodystrophy: The therapeutic consequences of effective control of serum phosphorus. JAMA 222:1640-1642, 1972 Taves DR, Freeman RB, Kamm DE, Ramos CP, Scribner BH: Hemodialysis with fluoridated dialysate. Trans Am Soc Artif Intern Organs 14:412414, 1968 Feagin FF, Waller AA, Pigman W: Evaluation of the calcifying characteristics of biological fluids and inhibitors of calcification. Calcif Tiss Res 4:
231-244, 1969
- |
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Fig lA-Grade 1 resorption is limited to occasional formation of Howship's lacunae (arrow). B-Grade 2 resorption includes Howship's lacunae, hookshaped resorption areas (arrow) and occasional internal resorption spaces. C-Grade 3 resorption has increased surface resorption and many internal resorption spaces. DGrade 4 resorption is characterized by massive trabecular resorption with a significant loss of trabecular mass (Decalcified, paraffin-embedded, H&E sections, X 75).
.'
American Journal of Pathology
HUFFER ET AL BONE DISEASE OF HEMODIALYSIS
[End of Article]
Metabolic Bone Disease in Chronic Renal Failure I. Dialyzed Uremics W. E. Huffer, MD, D. Kuzela, MD and M. M. Popovtzer, MD
Garner and Ball's point counting technic was used to compare metabolic bone disease in dialyzed and nondialyzed uremic patients. Histologic measurements of bone from dialyzed and nondialyzed uremic patients dying between 1966 and 1971 showed that dialyzed patients have quantitatively more severe bone resorp. tion, distortion of trabecular architecture and mineralization defects. Mineralization defects become more severe as the duration of dialysis increases but are not related to serum calcium and phosphorus levels. Bone volume in both groups is normal or increased and in dialysis patients increases in proportion to the elevation of serum phosphorus. Mean serum phosphorus and calcium levels, bone volume, and volume:surface ratios all decreased in dialysis patients between 1966 and 1971, while bone resorption and mineralization defects did not change. These results suggest that lowering of serum phosphorus without increasing serum calcium may aggravate the uremic bone disease by reducing bone volume without improvement of mineralization and resorption defects. (Am J Pathol 78:365-384, 1975)
MAINTENANCE HEMODIALYSIS has been successful in prolonging the life of patients suffering from chronic renal failure, but morbidity due to metabolic bone disease has not been eliminated.1'2 QuantiFrom the Departments of Pathology and Medicine, University of Colorado Medical Center, Denver, Colo. Presented in part at the Seventy-first Annual Meeting of the American Association of Pathologists and Bacteriologists, San Francisco, Calif, March 10, 1974. Supported in part by Training Grant GM-00977 from the US Public Health Service. Accepted for publication October 10, 1974. Address reprint requests to Dr. William E. Huffer, Department of Pathology, School of Medicine, University of Colorado Medical Center, 4200 East Ninth Ave, Denver, CO 80220. 365
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tative histopathologic studies of bone disease in nondialyzed-7 and dialyzed -22 patients have produced conflicting data about the occurrence of major types of bone abnormalities and hence different conclusions about the relationship between predialysis renal osteodystrophy and dialysis boine disease. The most prevalent patterns of bone response to chronic renal failure reported in different studies seem to be determined not only by common- pathogenetic responses to altered blood mineral levels, hormone secretion and vitamin utilization associated with renal failure, but also to variations in baseline mineral and bone homeostasis in the populations studied. These are influenced by the diet, climate and age of the individuals developing renal failure.23 In dialysis patients, differences in the specific therapeutic regimens may be expected to add to the many variables that determine the major patterns of bone response.24 The University of Colorado Medical Center has had an active maintenance hemodialysis program for several years. Presumably, the population treated at this center has not changed dramatically during this time. However, the details of the therapeutic regimen have changed, not only with respect to dialysis procedures, but also with respect to auxillary therapy such as the institution of oral phosphate-binding therapy in 1970. Since differences in the form of dialysis bone disease reported from different treatment centers might be related to differences in therapy, it seemed appropriate to ask if there were any changes in the form of bone disease seen in dialysis patients treated at this center during a time when therapy was in a state of evolution. This quantitative histopathologic study was designed to answer that question and to try to identify any specific changes in therapy which might explain changes in the form of bone disease. Quantitative indices of bone resorption, mineralization and bone volume in autopsy specimens of bone from dialyzed, nondialyzed and normal control patients were compared to establish the similarities and dissimilarities between conventional renal osteodystrophy and dialysis bone disease in patients treated at this medical center. The relationships between quantitative indices of bone morphology, parathyroid weight, antemortem serum calcium and phosphate levels, and duration of dialysis were investigated by statistical methods. Finally, the relationships of quantitative indices of bone morphology, antemortem serum mineral levels and duration of dialysis to time of institution of dialysis therapy or the time the patient died were investigated by statistical methods.
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Materials and Methods Sections of lumbar vertebrae were obtained from 44 dialysis patients, 15 conservatively treated uremics, and 20 normal control patients autopsied at Colorado General Hospital or the Denver Veterans Hospital between January 1966 and December 1971. The dialysis group was subdivided into a short-term group (9 patients, 0 to 6 months dialysis) and a long-term group (35 patients, >6 months dialysis). Conservatively treated patients had chronic renal failure not treated by maintenance hemodialysis or renal transplantation. Normal control patients were individuals without renal disease dying suddenly as a result of trauma. Twenty patients from a large pool of such individuals were matched by age and sex with 20 patients selected from 94 patients with renal disease using a table of random numbers. (Patients with renal failure included those in this study plus a group of patients treated by renal transplantation; lesions in the transplant patients are described in a subsequent paper 25). Specimens of lumbar vertebral bone were fixed for 24 hours in 10% neutral butfered formalin and decalcified in acid citrate. The major variable in identifying osteoid in decalcified hematoxylin- and eosin-stained sections is the decalcification time. Decalcified tissue was checked three times a day until a sharp pin could first be made to penetrate the cortical bone, which still felt hard to palpation. At this stage decalcification was sufficient to allow sectioning and still maintain the clearcut definition between basophilic (mineralized) and eosinophilic (nonmineralized) bone matrix in hematoxylin- and eosin-stained sections. After decalcification, specimens were processed routinely for paraffin embedding, sectioned at 7 to 10 ji and further processed for hematoxylin and eosin staining. Sections were stained with Harris hematoxylin (Harleco), pH 2.8, for 5 minutes and counterstained with eosin for 45 seconds. Sections of cancellous bone, 0.25 sq cm, were evaluated by a modification of Garner and Ball's point counting method.s Twenty-five nonoverlapping fields, encompassed by a square grid with 121 intersections, were counted resulting in a total count of 3,025 intersections/specimen. Parameters counted included calcified bone, osteoid, internal trabecular resorption spaces, and internal and extemal trabecular surfaces. The data were then used to calculate the following parameters: Parameters measuring resorptive activity and distortion of trabecular architecture Resorptive volume (RV) = internal resorption space counts/3025 Resorptive index (RI) = internal resorption space counts/calcified bone and osteoid counts Volume to surface ratio (V: S) = calcified bone and osteoid counts/surface counts
Parameters measuring changes in volume Total bone volume (BV) = calcified bone and osteoid counts/3025 Calcified bone volume (Ca++ BV) = calcified bone counts/3025 Parameter measuring mineralization defects Osteoid index (OI) = osteoid counts/calcified bone and osteoid counts Reproducibility of counts on a single specimen was approximately + 5%, similar to the results of Ellis and Peart.6 Mean values, standard deviations and standard errors for each of these parameters were calculated, and differences between groups were tested for by a standard t test applicable to two random
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of Pathology
groups of different sizes.268 The percentage of individuals in each renal disease group who fall within the normal control mean ± 2 SD or within specified limits above or below this range for several of the parameters was also calculated. Correlations between different parameters of bone histology or between a given parameter of bone histology and clinical or anatomic measurements such as duration of dialysis, parathyroid weight or serum phosphorus levels were tested by regression analysis using the University of Colorado Basic Library Computer Program, Stat 10. This program computes the slope and other statistics including the F ratio for a simple linear regression with one independent variable. Parathyroid weights were determined gravimetrically at the time of autopsy. Antemortem serum calcium, phosphate and Ca x P04 product levels were the mean of all predialysis determinations done during a patient's entire course of maintenance hemodialysis. For individual patients, the standard deviations of mean predialysis Ca and P04 values were approximately + 1 mg%, and there were no significant deviations above or below the means as a function of duration of dialysis. Parathyroid weights, serum mineral levels and quantitative indices of bone morphology were available in a group of 10 long-term dialysis patients whose indices of bone morphology are representative of the entire group of long-term dialysis patients. Mean antemortem serum mineral levels were available from 8 additional long-term dialysis patients (including 1 patient with quantitative bone indices) autopsied between 1966 and 1971. These 8 patients plus the 10 in the previous group were used to determine the relationship between serum mineral levels and the relative time of autopsy by regression analysis. The time of autopsy of the first dialysis patient was January 1966, and the time of autopsy of the last was December 1971. Regression analyses of bone indices and serum mineral levels as a function of relative time of autopsy have number of months after January, 1966, as the X axis. The earliest time of starting a maintenance dialysis program by a patient in this study was in 1965. The time axis for regression analysis of duration of dialysis as a function of relative time of starting maintenance dialysis was the number of months after January 1965, when the patient was started on the dialysis program. Since this was a retrospective study, the bone specimens used were all decalcified, and the von Kossa technic could not be used to determine osteoid index. As noted above, with careful decalcification and staining, osteoid could be identified in decalcified hematoxylin- and eosin-stained specimens by its relative eosinophilia, homogeneity and its relationship to either a calcification front or a cement line and adjacent basophilic bone matrix. In this study osteoid was consistently lamellar. To determine the accuracy of these criteria for identifying osteoid, each of five surgical bone biopsies from patients in another prospective study were divided into two portions. One portion was used to prepare frozen undecalcified sections which were stained by the von Kossa technic.27 The other was processed routinely after decalcification and stained with hematoxylin and eosin. The osteoid index was determined on all ten preparations in a double blind study. The results of the determinations on paired samples from the same biopsies are compared in Table 1. In addition to the quantitative parameters, a semiquantitative grading system was used to evaluate the degree of bone resorption. Grade 0 resorption was defined as trabeculae with smooth contours; Grades 1 to 4 are illustrated and defined in Figure 1.
Results Methodology
The osteoid index of five surgical biopsies as determined on vonKossa-stained sections was indistinguishable from osteoid index as
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Table 1-Osteoid Index on von-Kossa-Stained Undecalcified Sections and Hematoxylinand Eosin-Stained Decalcified Sections of the Same Biopsies von Kossa H&E Sample Difference 1 3.0 3.8 -0.8 2 1.2 0.8 +0.4 3 7.7 7.4 +0.3 4 0.0 0.0 0.0 5 0.6 0.5 +0.1 Mean 2.5 2.5 0.0
determined by histologic criteria applied to decalcified hematoxylinand eosin-stained sections of the same biopsies (Table 1). There was good correspondence between resorption grade as determined by established histologic criteria of bone resorption and the parameters used to quantitate bone resorption -(Text-figure 1). As the resorption grade increased, the mean V:S ratio decreased, and the resorption volume and resorptive index increased. Resorptive Activity and Distortion of Trabecular Architecture
Both dialyzed and nondialyzed patients had increased resorptive activity as judged by semiquantitative methods (Text-figure 2). None of the normal controls had significant resorption (100% Grade 0). All nondialyzed patients had significant resorption with an equal distribu20
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HUFFER ET AlL
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in the short-term dialysis group (Text-figure 3). The incidence of severely reduced ratios (one-fifth to three-fifths normal) was also greater in the long-term group. Changes in Bone Volume
Mean bone volumes of all renal disease groups were slightly greater than the normal mean (Table 2). While the majority of patients within these renal disease groups had bone volumes within the normal range (Text-figure 3), all groups had individuals with increased bone volume (22 to 27% of each group). Nondialyzed and long-term dialysis groups had only small numbers of individuals with reduced bone volume. None of the short-term dialysis patients bad reduced bone volume. No significant relationships between bone volume and age (in normal controls or any renal disease group), duration of dialysis, osteoid index, or semiquantitative or quantitative resorption indices were found. Mineralization Defects
All renal disease groups had increased mean osteoid indices in comparison to the normal control group (Table 2). The elevation in mean osteoid index was statistically significant at P < .005 for long- and short-term dialysis groups and at P between .05 and .025 for the nondialyzed group. Elevations of osteoid index were more extreme in dialysis patients. The mean osteoid index for combined long- and short-term dialysis patients was significantly greater than the mean OI of the nondialyzed group (P < .005). Osteoid index in dialysis patients increased as a function of duration of dialysis (Text-figure 4) and as a function of age (slope 2.9% per year, P < .005). In contrast, there was no increase 40
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BONE DISEASE OF HEMODIALYSIS
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Discussion
The major abnormalities of bone reported in nondialyzed patients with chronic renal failure include excessive bone resorption, or osteitis fibrosa 3,4,6,7,9, 11,12,15,1720; mineralization defects, or osteomalacia 3,57,11, 12,15,17-19; and increased bone volume, or osteosclerosis.3 4'6'7"0""'2"8 Decreased bone volume or osteoporosis is rare in nondialyzed patients,19 20 but at least one contradictory report had appeared.7 One study from Australia found no significant mineralization defects in nondialyzed patients.4 Conflicting views on the relative prevalance and time of onset of osteomalacia and osteitis fibrosa have also been reported.'628 Many of these discrepancies may be reconciled by Parfitt's conclusion that the course of renal osteodystrophy will be influenced by the age of onset of renal failure and sociologic and geographic differences in vitamin D nutrition.23 The major forms and the course of dialysis bone disease are equally variable. Most studies of dialysis bone disease have found a significant degree of osteomalacia. 12'1419'22 and some have reported progression of mineralization defects during hemodialysis.""1,2"l4"15'22 In contrast, one study found no significant mineralization defects in dialysis patients,'3 and several studies reported no progression of osteomalacia with duration of dialysis.16'18"9 Increased bone resorption occurs frequently in dialysis patients.8'9"1 13,1519,21,22 Some studies report progression of bone resorption during the course of dialysis,9" 3"17'21 whereas others found no change or decreased bone resorption.11'12"15"18'19'22 Decreased 8"12"13"15'16'20'21 normal,8,10,12,17-19 and increased 8,11,12 total bone volume has been reported in dialysis patients. Many workers stress selective reduction in mineralized bone volume as a unique feature of dialysis bone disease.10 12'15 Conclusions about the nature of dialysis bone disease and its relationship to conventional renal osteodystrophy are as divergent as the data reported above. Dialysis bone disease, or at least some forms of dialysis bone disease, have been regarded as a) new patterns of renal osteodystrophy with progression of osteomalacia and a lessening of osteitis fibrosa;" b) disuse osteoporosis with mineralization defects not specific for dialysis therapy;21 c) a disease quantitatively different from renal osteodystrophy because of progressive loss of mineralized bone, increased osteoid and a change in character of osteitis fibrosa;'2 d) a dialysis osteopenia differing from osteitis fibrosa, osteoporosis, and osteomalacia;20 e) a progression of conventional renal osteodystrophy with (in some patients) a less familiar tendency to develop osteopo-
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rosis;'5 or f) a disease essentially identical to conventional renal osteodystrophy. 17-19
The literature thus indicates that the form and course of both conventional renal osteodystrophy and dialysis bone disease will vary from individual to individual, and from one patient population to another. The present study confirms the individual variability of bone pathology in both dialyzed and nondialyzed patients as shown by the large standard deviations of all quantitative parameters of bone morphology studied. It adds another set of quantitative data to the literature, illustrating the major patterns of bone disease in dialyzed and nondialyzed uremics at a single treatment center. The data show that at this center dialysis bone disease is qualitatively similar, but quantitatively more severe than conventional renal osteodystrophy. Both groups have increased bone resorption, mineralization defects, and a normal or increased bone volume. Dialysis patients, however, have significantly more severe mineralization defects which progress with increasing duration of dialysis. Bone resorption, as measured by resorptive index, resorptive volume, and volume: surface ratios is somewhat more severe in dialysis patients, and although there is no linear increase in any of these indices with duration of dialysis, the data on percentage distributions of resorptive defects show that bone resorption becomes more frequent and severe as dialysis is prolonged. The new data presented in this study relates patterns of bone disease, serum mineral levels and duration of dialysis to the relative time of treatment and death during the interval between 1965 and 1971. Several facts were established. First, patients dying near the end of this time interval did not survive dialysis for significantly longer periods than those dying near the beginning of the interval. This does not necessarily imply that more recent dialysis procedures did not increase longevity, since the study does not include patients still surviving. Second, total bone volume and the volume: surface ratio, both presumably measures of the structural integrity of the bone, decreased as mean serum phosphorus and/or Ca X P04 products decreased. Third, the serum calcium, phosphorus and Ca X PG4 products of dialysis patients decreased during the period from 1966 through 1971. Fourth, the bone volume and volume: surface ratio of dialysis patients also decreased during this period, but the osteoid index, resorptive volume and resorptive index did not change. Fifth, all of the long-term dialysis patients whose parathyroid weights were known had hyperplastic glands. These observations may have bearing on the relationship between
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conventional renal osteodystrophy and dialysis bone disease, and the variability in the form of bone disease seen at different dialysis centers. They imply that therapy aimed at reducing serum phosphate levels without precise control of serum calcium levels may change the form of bone disease, resulting in a decrease in bone volume without correcting resorption and mineralization defects. Bone resorption in renal osteodystrophy is generally acknowledged to result from excessive parathormone production by hyperplastic parathyroid glands responding to lowered serum calcium levels.6'23'24'29'30 Hyperparathyroidism as demonstrated in this study is frequent in longterm dialysis patients,3' and measurements of serum parathormone levels in dialysis patients have shown a transient decline at the onset of dialysis followed by a progressive increase in chronic dialysis patients.30 There is good reason to believe, therefore, that the increased bone resorption seen in dialysis patients in this study resulted from hyperparathyroidism which was not effectively controlled in the period between 1966 and 1971. Elevated phosphate levels have been shown to decrease parathormone-induced resorption in vitro.33 However, in normal rats, phosphate infusions do not cause decreased hydroxyproline excretion or mobilization of 90Sr in response to parathormone. They cause retention of calcium by bone and/or soft tissues and increase bone formation.34 These observations suggest at least two mechanisms to account for the association between decrease in bone volume and V:S ratio and decreased mean serum phosphate levels found in this study. One is a loss of stimulation to bone formation, and the other is a loss of suppression of parathormone-induced bone resorption. In either case, decreases in bone volume and V: S ratio would be predicted. Equally important in this connection is the observation that bone volume, V: S ratio and mean serum phosphate levels all decreased during the period between 1966 and 1971. Bone volume and V: S ratio changes might be explained by the same phosphate-regulated mechanisms. Since mean serum calcium levels also fell during this period, parathyroid mechanisms also may have contributed to decreasing bone volume. It is not possible to determine from this study what changes in therapy brought about decrease in mean serum phosphorus levels. It seems likely that the institution of oral phosphate-binding therapy in 1970 is implicated. This form of therapy has been used in conjunction with increased concentrations of dialysate calcium to decrease parathormone production and to reverse osteitis fibrosa in dialysis patients.34 It has also been used to reverse metastatic calcification in dialysis patients with
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osteitis fibrosa.36 If lowered serum phosphate levels are implicated in decreased bone volume and V: S ratios, it might be important to maintain higher calcium levels,34 a form of therapy shown to be efficacious by the Mayo Clinic studies.34 Lack of correlations between osteoid index, serum mineral levels or the time the patients died during a period when serum mineral levels were changing, suggest that osteomalacia in dialysis patients is related to some specific effect of dialysis. This suggestion is supported by the demonstration of a statistically significant increase in severity of osteomalacia with increased duration of dialysis in this study, and also by clinical studies showing that osteomalacia in dialysis patients does not respond favorably to appropriate vitamin D therapy that will produce beneficial results in nondialyzed patients.24 Among the factors suggested as possible causes of impaired mineralization in dialysis patients are fluorides 37 and/or hypermagnesemia.38 References 1. Pendras JP, Erickson RV: Hemodialysis: A successful therapy for chronic uremia. Ann Intern Med 64:293-311, 1966 2. Ritz E, Krempien B, Mehls 0, Malluche H, Stroble Z, Zimmerman H: Skeletal complications of renal insufficiency and maintenance dialysis. Nephron 10:195-207, 1973 3. Gamer A, Ball J: Quantitative observations on mineralized and unmineralized bone in chronic renal azotemia and intestinal malabsorption syndrome. J Pathol Bacteriol 91:545-561, 1966 4. Ireland AW, Cameron DA, Stewart JH, Posen S: Quantitative histology of bone in advanced renal failure. Calcif Tiss Res 4:282, 1969 5. Binswanger U, Fischer J, Schenk R, Merz W: Osteopathie bei chronischer Niereninsuffizienz. Dtsch Med Wochenschr 96:1914-1919, 1971 6. Ellis HA, Peart KM: Azotaemic renal osteodystrophy: A quantitative study of iliac bone. J Clin Pathol 26:83-101, 1973 7. Ingham JP, Stewart JH, Posen S: Quantitative skeletal histology in untreated end-stage renal failure. Br Med J 2:745-748, 1973 8. Kim D, Bell NH, Bundeson W, Putong P, Simm NM, Walker C, del Greco F: Renal osteodystrophy in the course of periodic dialysis for chronic uremia. Trans Am Soc Artif Intern Organs 14:367-371, 1968 9. Jowsey J, Massry SG, Cobum JW, Kleenan CR: Microradiographic studies of bone in renal osteodystrophy. Arch Intern Med 124:539-543, 1969 10. Kerr DNS, Walls J, Ellis H, Simpson W, Uldall PR, Ward MK: Bone disease in patients undergoing regular hemodialysis. J Bone Joint Surg 51B:578, 1969 11. de Verber GA, Oreopoulos DG, Rabinovich S, Lloyd GJ, Mesma HE, Beattis BC, Levy D, Hudsan H, Rappaport A: Changing patterns of renal osteodystrophy with chronic hemodialysis. Trans Am Soc Artif Intern Organs 16: 479-485, 1970
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12. Ellis HA, Peart KM: Renal osteodystrophy with particular reference to the effects of chronic intermittent hemodialysis. Nephron 8:402, 1971 13. Kuhlencordt F, Kruse HP, Lozano-Tonkin C: Histomorphometric studies of bone in patients on chronic hemodialysis. Isr J Med Sci 7:517, 1971 14. Ritz E, Krempien B, Kuhn H, Heuck F: Dialysis bone disease. Isr J Med Sci 7:520, 1971 15. Bishop MC, Woods CG, Oliver DO, Ledingham JGG, Smith R, Tibbut DA: Effects of hemodialysis on bone in chronic renal failure. Br Med J 3:664-667, 1972 16. Delling G: Quantitative analysis of bone changes during chronic hemodialysis. Verh Dtsch Gesamte Pathol 56:436-438, 1972 17. Duursma SA, Visser WJ, Nijo L: A quantitative histological study of bone in thirty patients with renal insufficiency. Calcif Tiss Res 9:216-225, 1972 18. Krempien B, Ritz E, Beck U, Keilbach H: Osteopathy in maintenance hemodialysis. Virchows Arch [Pathol Anat] 357:257-274, 1972 19. Krempien B, Ritz E, Hauck F: Osteopathy in maintenance hemodialysis. Verh Dtsch Gesamte Pathol 56:439-442, 1972 20. Parfitt AM, Massry SG, Winfield AC: Osteopenia and fractures occurring during maintenance hemodialysis: A new form of renal osteodystrophy. Clin Orthop 87:287-302, 1972 21. Woods CG, Bishop MC, Nicholson GD: Bone histologic changes occurring after hemodialysis treatment for chronic renal failure. J Pathol 107:137-143,
1972
22. Binswanger U, Sherrard D, Rich C, Curtis FK: Dialysis bone disease. Nephron 12:1-9, 1973 23. Parfitt AM: Renal osteodystrophy. Orthop Clin N Am 3:681-698, 1972 24. Rubini ME, Cobun JW, Massry SG, Shinaberger JH: Renal osteodystrophy: Some therapeutic consideration relative to long-term dialysis and transplantation. Arch Intern Med 124:663-669, 1969 25. Huffer WE, Kuzela D, Popovtzer MM, Starzl T: Metabolic bone disease in chronic renal failure. II. Renal transplant patients. Am J Pathol 78:385-400,
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26. Snedecor GW: Statistical Methods Applied to Experiments in Agriculture and Biology. Ames, Iowa, Iowa State College Press. Chapter 4, 1959 27. Pearse AGE: Histochemistry: Theoretical and Applied, Second edition. Boston, Little, Brown and Co, p 934, 1961 28. Stanbury SW: Bone disease in uremia. Am J Med 44:714-724, 1968 29. Stanbury SW, Lumb GA: Parathyroid function in chronic renal failure: A statistical survey of the plasma biochemistry in azotaemic renal osteodystrophy. Q J Med 35:1-23, 1966 30. Bricker NS, Slatopolsky E, Reiss E, Avioli LV: Calcium, phosphorus, and bone in renal disease and transplantation. Arch Intern Med 123:543-553, 1969 31. Vosik WM, Anderson CF, Steffee WP, Johnson WJ, Arnaud CD, Goldsmith RS: Successful management of osteitis fibrosa due to "tertiary" hyperparathyroidism. Mayo Clin Proc 47:110-113, 1972 32. Johnson JW, Hattner RS, Hampers CL, Bernstein DS, Merrill JP, Sherwood LM: Effect of hemodialysis on secondary hyperparathyroidism in patients with chronic renal failure. Metabolism 21:18-29, 1972 33. Raisz LG, Nieman I: Effect of phosphate calcium and magnesium on bone
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35. 36.
37. 38.
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resorption, and hormonal responses in tissue culture. Endocrinology 85:446452, 1969 Pechet MM, Bobadilla E, Carroll EL, Hesse RH: Regulation of bone resorption and formation: Influences of thyrocalcitonin, parathyroid hormone, neutral phosphate and vitamin D3. Am J Med 43:696-710, 1967 Goldsmith RS, Furszyfer J, Johnson WJ: Differences between acute and chronic suppressibility of parathyroid function during chronic hemodialysis. J Clin Invest 50:37A, 1971 Pingerra WF, Popovtzer MM: Uremic osteodystrophy: The therapeutic consequences of effective control of serum phosphorus. JAMA 222:1640-1642, 1972 Taves DR, Freeman RB, Kamm DE, Ramos CP, Scribner BH: Hemodialysis with fluoridated dialysate. Trans Am Soc Artif Intern Organs 14:412414, 1968 Feagin FF, Waller AA, Pigman W: Evaluation of the calcifying characteristics of biological fluids and inhibitors of calcification. Calcif Tiss Res 4:
231-244, 1969
- |
A i tt|i^
-* 4 tEw_
A%
44"
Fig lA-Grade 1 resorption is limited to occasional formation of Howship's lacunae (arrow). B-Grade 2 resorption includes Howship's lacunae, hookshaped resorption areas (arrow) and occasional internal resorption spaces. C-Grade 3 resorption has increased surface resorption and many internal resorption spaces. DGrade 4 resorption is characterized by massive trabecular resorption with a significant loss of trabecular mass (Decalcified, paraffin-embedded, H&E sections, X 75).
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HUFFER ET AL BONE DISEASE OF HEMODIALYSIS
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