Parenteral aluminum administration in the dog: I. Plasma kinetics, tissue levels, calcium metabolism, and parathyroid hormone

Kidney International, Vol. 25 (1984), pp. 362—369

LABORATORY INVESTIGATION

Parenteral aluminum administration in the dog: I. Plasma kinetics, tissue levels, calcium metabolism, and parathyroid hormone DAN A. HENRY, WILLIAM G. GOODMAN, RONALD K. NUDELMAN, NICHOLAS C. DID0MENIcO, ALLEN C. ALFREY, EDUARDO SLATOPOLSKY, THOMAS M. STANLEY, and JACK W. COBURN The Medical, Laboratory and Research Services, Veterans Administration Wadsworth Medical Center and Sepulveda Veterans Administration Medical Center, The University of California Los Angeles-San Fernando Valley Program and the Department of Medicine, University of California Los Angeles School of Medicine, Los Angeles, California; The Medical and Research Services, Veterans Administration Medical Center, Denver, Colorado; The Department of Medicine, University of Colorado School of Medicine; and The Renal Division, Washington University School of Medicine, St. Louis, Missouri

Parenteral aluminum administration in the dog: I. Plasma kinetics, tissue levels, calcium metabolism, and parathyroid hormone. Aluminum

(Al) may cause both osteomalacia and encephalopathy in dialysis patients. Little is known about the biology of Al. This study examined the initial distribution kinetics of Al and its biological effects after injections of I mg/kg/day into dogs for 3 to 5 weeks. Following one intravenous dose, the plasma half-life (x SE) was 276 51.8 mm, with an apparent volume of distribution of 1.30 0.17 liters or 5.90 0.30% body wt; 10 to 21% of administered Al was excreted in the urine over 150 mm, and the renal contribution to plasma clearance of Al correlated with GFR (r 0.77, P < 0.05). The total plasma clearance of Al (4.43 2.83 mI/mm) exceeded the renal contribution to plasma clearance (1.94

0.30% do poids corporel; 10 a 21% de l'Al administré était excrété dans les urines en 150 mm, et Ia contribution rénale a l'epuration plasmatique de l'Al était corrélée avec le debit de filtration glomerulaire (r = 0,77, P < 0,05). La clearance plasmatique totale de l'Al (4,43 2,83 mI/mm) dépassait Ia contribution rénale ala clearance plasmatique (1,94 0,36

mi/mm) chez chaque chien, et seulement deux fois Ia contribution rénale a atteint 50% de Ia clearance plasmatique totale. La calcémie s'est élevée de 9,4 0,3 mg/dI et l'hormone parathyroi0,2 a 11,1 dienne immunoréactive (iPTH) a chute de 27 4% apres une injection

4% following one Al injection. With

d'Al. Lors d'injections répétées d'Al, Ia calcCmie s'est élevée du niveau debase de 10,2 0,07 mgldl a 11,1 0,22 et 11,3 0,46 mg/dl après I et 2 semaines, respectivement. La fonction rénale a diminué chez tous les chiens, et Ia créatinine sérique a dépassé 3,5 mg/dl chez quatre; au cours des 5 semaines de l'étude, Ia calcémie Ctait corrélée avec Ia créatininémie (r = 0,91, P <0.001). Le foie, le rein, et Ia rate avaient le plus grand contenu tissulaire en Al, et il y avait une fixation substan-

repeated Al injections, serum calcium increased from baseline levels of 0.46 rng/dl after 1 and 2 0.22 and 11.3 0.07 mg/dl to 11.1 10.2

tielle dans l'os; le contenu parathyroidien en Al était modeste. Les niveaux d'iPTH sérique étaient normaux ou élevés, et Ia rCponse de

weeks, respectively. Renal function declined in all dogs, and serum creatinine exceeded 3.5 mg/dl in four; over the 5 weeks of study, serum calcium correlated with serum creatinine (r = 0.91, P < 0.001). Liver, kidney, and spleen showed the highest tissue content of Al, and there was substantial uptake by bone; the parathyroid content of Al was

l'iPTH a une hypocalcémie induite par 1'EDTA n'était pas réduite par le traitement a I'Al. Ces données indiquent une retention tissulaire sub-

0.36 mI/mm) in each dog, and in only two instances did the renal contribution reach 50% of total plasma clearance. Serum calcium rose 0.3 mg/dl and imniunoreactive parathyroid 0.2 to 11.1 from 9.4

hormone (iPTH) fell by 27

modest. Serum iPTH levels were normal or elevated, and the response of serum iPTH to EDTA-induced hypocalcemia was not reduced by Al

stantielle d'Al aprés injections répétées d'Al, L'excrétiori rénale d'Al était limitée, probablement a cause d'une liaison importante aux protéines plasmatiques et aux tissus. L'Al parentéral a entrainé des perturbations marquees du métabolisme calcique et de Ia fonction rénale, mais Ia réponse de Ia PTH a l'hypocalcémie n'Ctait pas altérée.

treatment. These data show substantial tissue retention of Al with repeated Al injections. The renal excretion of Al was limited, probably

due to extensive plasma protein and tissue binding. Parenteral Al caused marked disturbances in calcium metabolism and renal function, but the PTH response to hypocalcemia was not impaired.

Administration parentérale d'aluminium chez le chien: I. Cinetique plasmatique, niveaux tissulaires, le métabolisme calcique, et l'hormone parathyroidienne. L'aluminium (Al) peut entrainer une ostéomalacie et une encephalopathie chez des hemodialysés. On salt peu de choses sur

Ia biologie de l'Al. Cette étude a examine Ia cinétique de distribution

initiale de l'Al et ses effets biologiques après des injections de I mg/kg/jour a des chiens pendant 3 a 5 semaines.AprCs une dose SE) était 276 51,8 mm, intraveineuse, Ia demi-vie plasmatique (x avec un volume de distribution apparent de 1.30 0.17 litres ou 5.90

Received for publication January 14, 1983 and in revised form June 8, 1983

© 1984 by the International Society of Nephrology

Aluminum has received considerable attention as a pathogenic factor in both dialysis dementia and osteomalacia [1—12]. The data in support of the association between aluminum accumula-

tion and osteomalacia are particularly convincing. Thus, clini-

cal studies of renal osteodystrophy indicate an association between bone aluminum content and both the frequency and severity of vitamin D-resistant osteomalacia in patients undergoing hemodialysis [8, 10, 11]. Ellis, McCarthy, and Herrington [13] showed that injections of aluminum to rats impaired the mineralization of the tibial metaphysis. In addition, the inadvertent administration of aluminum to patients treated with longterm total parenteral nutrition has been associated with the development of focal osteomalacia [14—161. Despite the potential toxic manifestations of aluminum accu-

mulation, little is known regarding the kinetics of aluminum 362

Kinetics of aluminum in the dog

distribution after its parenteral administration or its biological effects in humans or experimental animals. Aifrey, Hegg, and Craswell [17] demonstrated substantial accumulation of aluminum in tissues, particularly bone and liver, from uremic patients, whether treated with dialysis or not. Other investigators [18] found accumulation of aluminum in the parathyroid glands; moreover, the addition of aluminum to suspensions of parathyroid cells, in vitro, inhibits the secretion of PTH [19]. These latter observations may pertain to the syndrome of dialysis osteomalacia, which is characterized by altered calcium-PTH relationships [17]. Thus, these patients often show a propensity to hypercalcemia, and serum iPTH levels are often lower than predicted in the face of renal insufficiency [12, 20—221. Most observations have suggested that plasma aluminum is largely protein-bound and not readily removed by hemodialysis techniques [23, 24]. Thus, since the renal excretion of aluminum represents the predominant route for its elimination [23], aluminum accumulation is particularly likely to occur in the presence of renal insufficiency [17].

The present studies were conducted to investigate the kinetics of aluminum following its injection into dogs. Also, the biological consequences of both single and repeated injections of aluminum were studied, as was its tissue distribution. The

response of serum immunoreactive parathyroid hormone (iPTH) to acute hypocalcemia induced by sodium-EDTA was also evaluated before and after aluminum loading. Methods Studies were carried out in six normal, female mongrel dogs, weighing 18 to 34 kg. Prior to study, the animals had undergone

episiotomy for ease of bladder catheterization. The animals were studied during a 14-day control period prior to aluminum

363

specimens. An indwelling balloon catheter was inserted in the bladder for urine collections. A water diuresis was initiated by the administration of 40 ml/kg of water by a nasogastric tube followed by the infusion of 2.5% dextrose in water at 4 mI/mm.

Exogenous creatinine clearances were carried out after an initial loading dose of 8% creatinine (60 mg/kg) followed by a constant infusion of 1% creatinine at a rate of 1 mI/mm. After a stable urine flow rate was established, that is, above 50 mi/iS mm, the aluminum kinetic studies were carried out. Individual urine collections, of 15 mm duration, were obtained for 30 mm before and 150 mm after the aluminum injection; blood samples

were collected precisely at the midpoint of each collection period. In one animal, ultrafiltrates of plasma were prepared both in vivo during a hemodialysis procedure by isolated ultrafiltration using a cuprophan membrane and also in vitro by the centrifugation of plasma through a membrane (Centriflow, Amicon, Lexington, Massachusetts) at 2000 rpm at 21°C for 30 mm; the fraction of aluminum that was ultrafilterable in vivo was 3.9%; 2.5% was diffusable during centrifugation. Ultrafil-

terabie levels of aluminum were not measured during the present clearance experiments, but diffusable aluminum levels were determined in 15 samples from five other dogs given a comparable aluminum load; the percentage diffusability was 0.5 1%. During the present experiments, the renal 2.82 contribution to plasma clearance was calculated based on total plasma aluminum. Also, the renal clearance of aluminum, as calculated, does not have the same meaning regarding renal

handling as the usual renal clearance because of our lack of information regarding the quantity of aluminum that was actually filtered. In all dogs, the response of the parathyroid glands to hypocalcemia induced by the infusion of sodium EDTA was assessed 3

injections, and for 3 to 5 weeks while they received daily days before initiation of aluminum injections and again after aluminum injections. Fasting blood samples were obtained week 2 of aluminum injections; in two dogs, the sodium EDTA twice weekly for the measurement of calcium, phosphorus, infusion was repeated at the end of 4 weeks. For the studies creatinine, and aluminum concentrations. Renal function was evaluated initially by exogenous creatinine clearances and subsequently from serial serum creatinine levels. Basal serum iPTH levels were measured biweekly, and

done after initiation of aluminum administration, the sodium EDTA infusions were given 24 hr after the previous daily injection of aluminum. In all studies, sodium-EDTA (75 mg/kg

the secretory response to acute hypocalcemia was studied

samples were obtained for the determination of ionized calcium

during the control period and after 10 and 20 days of aluminum injections. The kinetics of aluminum disappearance from plasma were examined after its initial bolus injection and again in two dogs after 3 weeks of aluminum administration. Continued aluminum loading was produced by giving daily bolus injections of aluminum chloride, each providing elemental aluminum, 1 mg/kg, on

5 days each week for 3 to 5 weeks. Aluminum chloride was

and iPTH before the infusion of sodium EDTA, at 30-mm intervals during the EDTA infusion, and for 2 hr thereafter. The experiments were terminated after 3 to 5 weeks of aluminum loading. In two dogs, the studies were terminated after 3 weeks because of the rapid appearance of renal insufficiency (see Results). When the studies were terminated, the animals were killed with pentobarbitol, 40 mg/kg. Samples of bone, liver, kidney, thyroid, spleen, parathyroid, muscle, and

dissolved in 0.85% saline and infused intravenously in a volume of 8 to 15 ml over 45 to 60 sec on 5 mornings each week. This

brain were obtained for the measurement of aluminum content. Renal histology was evaluated in all animals except dogs 1 and

solution had a relatively low pH of 2,6, but the total acid load provided to each dog was only 0.10 mEq/kg/day. This small quantity of acid represents only a small fraction of the usual endogenous acid production of 3 mEq/kg/day in the dog [25]. The plasma clearance studies were conducted with the dogs

2. Tissue samples were fixed in neutral formalin and then

in the unanesthetized, fasting state while supported in the

body wt) was infused at a constant rate over 2 hr. Blood

embedded in paraffin. Sections were stained with hematoxylin

and eosin, and periodic acid-Schiff; the von Kossa stain for calcium and the Gram stain (Brown and Prenn technique) were also done. Total calcium concentrations were determined in serum and

urine by atomic absorption spectroscopy, as reported elsewhere [22], ionized calcium in serum by the flow-through superficial limb veins, one of which was used for the adminis- electrode (Orion Biomedical, Cambridge, Massachusetts), tration of parenteral fluids and the other for obtaining blood phosphorus and creatinine using an autoanalyzer (Technicon standing position by a sling-harness. The animals were prepared

for these clearance studies by cannulation of two separate

Henry et al

364

aluminum load excreted in the urine over 150 mm. The total

20,000 -

Vd = 1.32 liters

clearance of aluminum from plasma was 4.43 0.83 mI/mm. On the average, the kidney accounted for 44% of plasma aluminum removal and in only two dogs did this value exceed 50%. The renal contribution to plasma clearance of aluminum, measured after the initial injection, was related directly to GFR (r = 0.77,

PC Al = 4.63 mI/mm

P < 0.05).

19,000.. S

S

18,000-

S

t1/2 = 198 mm

17,000 E

= 0.96

E 16,000 E

15,000

I 14,000

A prompt and sustained increase in serum calcium concentration followed the initial injection of aluminum chloride in each dog (Fig. 2) which was accompanied by small but significant increments in urinary calcium excretion. There was a modest increase in serum phosphorus concentration which was significant only at 30 mm following the aluminum injection (4.5 0.6 to 5.6 0.7 mg/dl). In each dog, the serum iPTH level fell after the aluminum injection (Fig. 3), with an average decrease of 27 4%.

-c 0

50

100

Time after injection. rn/n

Fig. 1. Example of the disappearance curve of aluminum after its

injection (1 mg/kg elemental Al) in dog no. 1. The initial blood sample was obtained 7.5 mm after the injection with subsequent samples at 15mm intervals. PC Al represents the plasma clearance of aluminum.

Instruments Corporation, Tarrytown, New York). Aluminum was determined in serum, urine, and tissue samples by flameless atomic absorption spectroscopy [26]. Serum IPTH was determined by radioimmunoassay utilizing an antibody (Ch 9) which is directed against the mid-region of the carboxyl terminus of the PTH molecule [27]. The levels of iPTH, measured in dogs with moderate reductions in renal function, have been shown to correlate with histologic evidence of secondary hyperparathyroidism [28]. All values are expressed as the mean SE; statistical analysis of the data utilized Student's t test for paired samples and linear regression analysis. Results

Single dose aluminum administration Following the first intravenous dose of aluminum (1 mg/kg), the mean plasma aluminum concentration reached 17,835 1,294 and 16,069 1,295 g/1iter at 7.5 and 22.5 mm, respectively. The plasma levels then decreased slowly and followed a

Effects of repeated aluminum injections The continued injection of aluminum for 5 days each week for

3 to 5 weeks led to disturbances of calcium homeostasis and renal function. By the end of week 1, total serum calcium concentrations measured before the daily aluminum injection increased in five of the six dogs; the mean values increased from a baseline of 10.16 0.07 mg/dlto 11.05 0.22 and 11.33 0.46 mg/dl at the end of weeks 1 and 2, respectively (Fig. 4). Near the time for the studies to end, the total serum calcium concentration increased markedly in three of the dogs. Renal function, evaluated from serial serum creatinine levels and exogenous creatinine clearances, decreased in all dogs (Fig. 5). In four of six animals, serum creatinine levels exceeded 3.5 mg/dl at the completion of these studies; however, the marked increments in serum creatinine occurred only at the termination of the study in three of the animals. The rise in serum creatinine

closely paralleled the increments in serum calcium, with a significant correlation between serum calcium and serum creati-

nine concentrations over the course of the experiments (r = 0.91, P < 0.001). Despite these changes in serum calcium and creatinine, the body weight of the dogs remained within 0.5 kg, with the exception of one dog that had a weight loss of 2 kg. At the end of weeks 1 and 4 of aluminum injections, the basal morning plasma aluminum levels, which were measured 24 hr following the previous injection were related inversely to both the renal and total clearances of aluminum from plasma (Fig. 6),

pattern over 150 mm which was consistent with first-order which had been measured 4 days earlier. The basal plasma kinetics for the disappearance of aluminum from plasma; an aluminum levels, obtained prior to the daily injection of alumiexample is shown in Figure 1. The values for apparent volume of distribution (Vd), plasma half-life, and both the total plasma clearance and the renal contribution to the plasma clearance of aluminum for individual dogs are shown in Table 1. The mean plasma half-life, derived from the elimination curves for each of the six dogs was 276 52.8 mm. The Vd was 1.30 0.17 liters or 5.90 0.30% of body wt, representing values nearly twice the estimated plasma volume in these individual animals. From the subsequent tissue distribution of aluminum (described below), it is apparent that the volume of distribution was considerably larger; however, the entry of aluminum into the tissues may have occurred so slowly that it was not apparent over the 150 mm of this study. The mean renal contribution to the clearance of aluminum from plasma was 1.94 0.36 mI/mm, with 10 to 21% of the total

num and shown in Figure 7, remained relatively stable although there was substantial variation from dog to dog. Liver, kidney, and spleen tissues had the greatest accumulation of aluminum at the time the experiments were completed (Table 2). There was also a substantial uptake of aluminum into bone. The results of

quantitative histomorphometry of trabecular bone and the relationship between aluminum administration and the development of osteomalacia in these animals are reported elsewhere [29]. Three of the four dogs that exhibited a marked impairment of renal function demonstrated the highest levels of aluminum in renal tissue (> 1000 mg/kg).

The histopathology of the kidneys at the conclusion of the studies revealed variable changes that were not seen uniformly in all animals. Thus, focal tubular necrosis was present only in dog 6; focal infiltrates with acute inflammatory cells occurred in

365

Kinetics of aluminum in the dog Table 1. Analysis of plasma kinetics in individual dogs following the injection of aluminuma

—

Urine

1

2 3 4 5 6

mi/mm

liters

67.3 55.7 72.4 59.7

1.32 1.63 2.21

1.38 1.36

54.1

1.03

mm

4.7 5.7 6.3 6.3 6.8 5.7 7.2 7.9

198

9.6

470 317 295 287 88 95 88

15.0 18.4 13.0 11.4

6'

86.2 31.3 43.8

Mean ±SE

65.9

1.30

5.95

±5.0

±0.17

±0.30

5c

1.44 1.47

% dose in 150 mm

% body wt

Renal

Total

Al

Plasma t 1/2

Total

GFR

Dog no.

Plasma clearanc&'

.

.

Volume of distribution

mi/mm

1.09 1.76

4.63 2.48 4.82 3.24 3.28 8.15

21.3

— —

3.20 1.54 1.24

2.83 4.19 7.23

10.5 11.6

276

14.8

4.43

1.94

±52.8

±1.8

±0.83

±0.36

a Means ± SE are limited to initial studies carried out in the six dogs. b See Results for discussion of renal contribution to plasma aluminum clearance. Studies in these animals were repeated after 3 weeks.

80

j

60

II-..U)

E .c

20

0

0

150

Time, mm

Fig. 3. Serum iPTH levels before and 150 mm after the initial injection of aluminum, 1 mg/kg. The fall in serum iPTH was significant, P < 0.05. The following symbols represent individual dogs by number: I, •; 2, 0;

3, X; 4, ; 5, A; 6, LII. 0

30

60

90

120

150

Time, mm

Fig. 2. The changes in serum calcium, phosphorous, and urinary calcium excretion in dogs after the initial injection of aluminum, I mg/kg. Values are mean ±SE. The asterisk (*) indicates values that differ significantly from control values, by paired analysis P < 0.05.

dogs 3, 4, and 5, and scattered areas of degenerated tubules were present in dogs 3 and 6. There were small focal areas of calcium deposition only in dog 3. Of particular interest was the minimal nature of calcium deposition in the renal parenchema, in view of the hypercalcemia that developed in this animal. The serial values of serum iPTH are shown in Figure 8. Four

of the six dogs, those with the greatest increments in serum creatinine, showed a rise in serum iPTH to values above normal

for this assay in the dog. During the overall period of study

there was a significant correlation between serum iPTH and serum creatinine (r = 0.51, P < 0.01). The changes in serum iPTH in response to EDTA-induced hypocalcemia prior to and following the parenteral aluminum loading are shown in Figure 9. Following the infusions of EDTA, serum ionized calcium decreased from 4.87 ± 0.056 to 3.68 ± 0.25 mgldl during the control study before aluminum administration and from 5.04 ± 0.20 to 3.47 ± 0.20 mgldl after 10 days of aluminum treatment. Aluminum administration did not cause a significant decrease in either the control, pre-EDTA

serum levels of ionized calcium, or the percent decrement in ionized calcium produced by the sodium-EDTA. With these similar decrements of ionized serum calcium, the peak serum

iPTH levels, expressed as a percent of the control value obtained prior to the EDTA infusion, were not different before

or after aluminum loading. There was no evidence that the

Henry et at

366 Aluminum injections 20

N-

14-

Aluminum injections

20 15 10 0)

•0 0)

Sc

5 4 3

a)

2

a)

15.

C

a) C-)

a>

C)

E

E 1.0

)l)

'1)

U)

0.5

1

10

—7 10

0

10

20

30

Time, days

•

20

30

Time, days

Fig. 4. Serial levels of total serum calcium following repeated aluminum injections, 1 mg/kg, given intravenously on 5 days of each week. Lines

connect the values obtained in the same animal. The symbols are the same as those used in Figure 3.

infusion of sodium EDTA on days 10 and 20 had any effect on

Fig.

5. Serial levels of serum creatinine in dogs receiving repeaten

aluminum loading as in Figure 4 (note log scale of serum creatinine). The symbols are the same as those used in Figure 3.

half of its disappearance from plasma. Data reported elsewhere [16, 23] indicate that there is little gastrointestinal excretion of

aluminum that has been given parenterally. Thus, the extra-

the basal levels of either serum calcium or aluminum, as renal clearance of aluminum may be accounted for, in large

part, by the slow continued uptake of aluminum into tissues. Since aluminum can accumulate in the body's with repeated exposure and since there is a limited capacity for the kidney to Discussion excrete aluminum, it can be anticipated that a considerable Considerable evidence indicates that aluminum may be interval without aluminum exposure may be needed for the pathogenic in both dialysis dementia and dialysis-associated kidney to reduce the body's burden of aluminum significantly. osteomalacia. Aifrey, Hegg, and Craswell [17] demonstrated This contention is supported by clinical studies which show that substantial tissue retention of aluminum in patients with renal the urinary excretion of aluminum by renal transplant recipients insufficiency, regardless of whether or not long-term dialysis may be increased for up to 6 months following renal transplanhad been used in their treatment regimen. The results of the tation [30]. The disturbances in calcium homeostasis and renal function present study not only support the earlier findings which show tissue accumulation of aluminum but also provide additional observed in the present study were unexpected. Moderate insight into the kinetics of aluminum in plasma and its renal hypercalcemia developed within minutes after single bolus excretion. The results obtained following both acute single injections of aluminum, whereas repeated aluminum injections injections and short-term aluminum loading indicate that renal produced hypercalcemia that was sustained for several weeks. excretion contributes substantially to the total plasma clearance Factitious hypercalcemia was excluded as an explanation for of aluminum. In contrast to previous studies [23], the clearance these findings; thus, the addition of aluminum to plasma did not of aluminum from plasma by the kidney was quite low in the interfere with serum calcium determinations as measured by present experiments. Only 10 to 20% of injected aluminum was atomic absorption spectroscopy over a wide range of plasma excreted in the urine over 150 mm. This finding, the very high aluminum concentrations. Unfortunately, serum ionized calciplasma aluminum levels achieved after the bolits injection, the um levels were not measured throughout the experiment. When small calculated initial volume of distribution of aluminum, and ionized calcium levels were measured after 10 and 20 days, the measurement of ultrafilterable aluminum in one animal respectively, of aluminum administration, they had not insuggest that a large fraction of administered aluminum is creased significantly despite the mean increase in total serum determined by biweekly measurements.

promptly bound to plasma proteins. The conclusion that little of

the plasma aluminum is ultrafilterable is supported by work demonstrating very limited plasma clearances of aluminum across hemodialysis membranes [24] and by observations in patients treated with long-term total parenteral nutrition [16].

calcium of 1.1 mg/dl and the highest total serum calcium of 12.7 mgldl. The lack of long-term suppression of serum iPTH levels

suggests that serum ionized calcium was not increased, although no conclusion can be reached in the three animals that developed marked hypercalcemia just before the studies were The present observations provide strong support for the view completed. The modest decrease in serum iPTH levels obthat there is slow, progressive accumulation of administered served during the initial infusion of aluminum suggests that aluminum in tissues. The plasma kinetic data show that the serum ionized calcium may have increased transiently during renal contribution to aluminum clearance accounts for less than the time of this acute study. Further investigation is needed to

367

Kinetics of aluminum in the dog luminum injections

6000

Co

5000 4000

C0

E0)

10,000

.

0)

E

3000

r=

I.

0.90

P

Co

a

2000

Co

0)

100

10

Co

m

1000

—7

0

10

20

30

Time, days

0 4 2 Renal clearance, mi/mm

6

Fig. 7. Serial values of plasma aluminum before and during the repeated loading with aluminum; the latter measurements were made

at least 24 hr after the previous injection of aluminum. Symbols represent individual dogs and are the same as those used in Figure 3.

is most likely that serum calcium levels increased acutely due to the mobilization of skeletal calcium arising as a consequence of

6000

.

5000

the aluminum injection. The precise mechanism of this response remains uncertain. With repeated injections of aluminum, hypercalcemia became sustained and progressive; indeed, it was very marked in some animals. It is likely that pathophysiologic mechanisms other than calcium release from bone contributed to the striking

0)

.

4000

4 Co

E (0

a

3000

r=

Co

2000

increase in total serum calcium. As serum calcium and its

0.79 S

Co CO

Co

1000

.

.

0

0

5

10

Total plasma clearance, mi/mm

Fig. 6. The relationship between morning, basal plasma aluminum levels, and both the renal contribution to plasma clearance (top panel) and the total plasma clearances (bottom panel) of aluminum. These clearance measurements were determined after the intravenous infusion of a single dose of aluminum, 1 mg/kg (see Table I); the basal

serum aluminum level was measured 4 days later, 24 hr after the

previous injection of aluminum. Renal clearance calculations utilized total serum aluminum measurements (see Results).

clarify the effects of aluminum on the partition of calcium in plasma.

Overt hypercalcemia or a predisposition to its development

with either calcium or vitamin D therapy has occurred in patients with the syndrome of dialysis osteomalacia that is associated with aluminum accumulation in bone [10, 12]. The present data suggest that this tendency toward hypercalcemia could represent a direct effect of aluminum, per se. Despite our lack of definitive data regarding changes in the distribution of calcium within its various components in plasma, the present observations do permit speculation regarding the mechanisms of calcium transfer or flux during the administration of aluminum. With a single bolus injection of aluminum, the

rise in total serum calcium level was prompt and sustained without evidence for excessive PTH secretion. The time course of this change is too brief to incriminate an intestinal mechanism for the development of hypercalcemia, and the increase in urinary calcium excretion excludes a renal mechanism. Thus, it

filtered load increase, the renal excretion of calcium represents an important mechanism for protection against worsening hypercalcemia [31]. With the continued entry of calcium into the extracellular fluid, hypercalcemia might be aggravated if renal function is significantly reduced. Under these conditions, the entry of calcium into the skeleton may be critical in the defense against further increments in serum calcium. With the repeated

injections of aluminum into these dogs, there was a close correlation between the total serum calcium concentration and the serum creatinine level; moreover, marked hypercalcemia (serum calcium levels above 15 mg/dl) developed only in the face of an abrupt and striking reduction in renal function noted late in the course of the study. This degree of hypercalcemia cannot be explained only on the basis of reduced renal function; the calcium entering the extracellular fluid probably originated from the skeleton as an example of "disequilibrium hypercalce-

mia" which was aggravated by renal failure [31, 32]. Other factors could also contribute to the hypercalcemia in the present studies. Although it is possible that intestinal calcium absorption could have been increased, this seems unlikely in the face of the reduced serum levels of 1 ,25(OH)2D observed [291. Moreover, there were variable reductions in food intake in some of the dogs. The present data do not permit delineation of the relative contributions of excessive calcium mobilization versus reduced calcium uptake by bone to the pathogenesis of hypercalcemia. Bone biopsy specimens following aluminum administration showed evidence of osteomalacia with reduced rates of bone apposition and poor uptake of tetracycline into bone [29]. These findings would clearly indicate reduced entry of calcium into

bone. On the other hand, the severity of the hypercalcemia makes it unlikely that serum calcium increased entirely as a consequence of reduced calcium entry into bone. Calcium

368

Henry et a!

Table 2. Tissue levels of aluminum in dogs receiving repeated aluminum injections

Tissue aluminum, mg/kg/FFDW Dog no.

lb 2b 3

4 5 6

Bone'

.

Brain

Liver

Kidney

330 462 563 440 334

1234 1089 275 1512

231

104

209

Muscle

3.0

Thyroid

Parathyroid

Spleen — — 594 559 440

g

Pre

Post

5.2 7.2

ND

11.2

ND

91 136 129

2.8 3.8 3.4

1.5 1.0

4.5

—

20.0

23.0

2.5 3.5 2.5

13.5

31.0 79.0 48.0 40.0

6.2 8.7

16.0 19.0

21.9 ±9.5

38.8

554

5.6

±9.4

±35

±1.3

Mean

393

737

6.0

±SE

±48

±249

±2.8

58.0

581

1.2

94

4.3

78 90

1.5

103

±0.58

±9.6

Abbreviations: ND, not detectable (<0.5 mg/kg); FFDW, fat free dry weight. a Pre and Post refer to aluminum administration. This animal received aluminum loading for 3 weeks; other animals received aluminum for 5 weeks. Aluminum-loaded

Control 1200

> F- O

LU

1000

C, 0=

.2.5

2

I

800

600

(0

E

400

S

x 200 100 Basal

C

20

10

30

Time, days

8. Serial values of serum immunoreactive parathyroid hormone (iPTH), measured with a carboxyl terminal antiserum, during the Fig.

EDTA

Basal

EDTA

Fig. 9. The maximum increment in serum iPTH, represented as a percent of the control value, after hypocalcemia induced with sodiumEDTA, 75mg/kg. Studies were before (control) and after 10 days of the

repeated aluminum injections. In two dogs (*) the response was evaluated on a second occasion after 4 weeks of aluminum loading.

repeated loading with aluminum as described in Figure 4.

could be liberated into the extracellular fluid as a consequence of increased bone resorption, although the data supporting this mechanism in the current study are only indirect. In an exten-

studied in vitro [19]. Thus, it is possible that aluminum may have a direct inhibitory effect on the parathyroid glands. The results of the present investigation neither demonstrate marked aluminum accumulation by parathyroid tissue compared to

sion of the present studies [29], there was a reduction in other types of tissue nor do they indicate that aluminum trabecular bone volume without an increase in resorptive produced a defect in the secretory reserve for PTH in response surface of bone, suggesting that bone resorption may not have to acute hypocalcemia. However, the present studies were occurred through normal bone remodeling processes. However, this calcium mobilization might also involve a more rapidly

conducted after the administration of aluminum for relatively short periods. Therefore, an effect of more prolonged aluminum

exchangeable pool of calcium in bone. There are no data

loading on PTH secretion cannot be excluded. Indeed, the

available to distinguish between these possibilities, although preliminary evidence suggests that aluminum loading may increase resorptive activity in the tibial diaphysis of the rat [331. Another feature of the clinical syndrome of dialysis osteomalacia is the presence of serum iPTH levels which are lower than expected in advanced renal failure [10]. Moreover, there are

present results do not exclude the possibility that a reduction in parathyroid activity may in some way predispose to the accu-

mulation of aluminum in bone and the occurrence of the syndrome of dialysis osteomalacia.

The dose of aluminum given to these animals was high; however, on a body weight basis, it was generally far less than that given to rats [13, 34, 351. Thus, the serum aluminum levels

data suggesting that aluminum may accumulate preferentially in the parathyroid glands [18], and other observations indicate that measured were substantially greater than those observed in

aluminum can reduce PTH secretion by parathyroid cells clinical conditions of aluminum loading. However, the tissue

369

Kinetics of aluminum in the dog

levels achieved, particularly those in bone, were comparable to those observed in clinical conditions associated with exposure to aluminum [10, 17, 161. Acknowledgments Presented, in part, before the 15th Annual Meeting of the American Society of Nephrology, Chicago, Illinois, December 12—14, 1982. This work was supported in part by grants AM29926 and AM14750 and by research funds from the Veterans Administration. Dr. R. K. Nudelman is the recipient of a Veterans Administration Associate Investigator

award. Dr. A. C. Alfrey and Dr. J. W. Coburn are recipients of the Medical Investigator Award of the Veterans Administration. Technical assistance was provided by D. Mishler, J. Gilligan, N. Miller, J. Steppe, W. van Buren, and E. Tallos. Secretarial assistance was provided by A. Sato.

Reprint requests to Dr. J. W. Coburn, Nephrology Section 691/JIlL,

Veterans Administration Wadsworth Medical Center, Wilshire and Sawtelle Blvds., Los Angeles, California 90073, USA

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