Growth plate cartilage formation and resorption are differentially depressed in growth retarded uremic rats

To characterize the modifications of growth plate in individuals with growth impairment secondary to chronic renal failure, young rats were made uremic by subtotal nephrectomy (NX) and, after 14 d, their tibial growth plates were studied and compared with those of sham-operated rats fed ad libitum (SAL) or pair-fed with NX (SPF). NX rats were growth retarded and severely uremic. Growth plate height (mean +/- SD) was much greater (P<0.05) in NX (868.4+/-85.4 microm) than SAL (570.1+/-93.5 microm) and SPF (551.9+/-99.7 microm) rats as a result of a higher (P<0.05) hypertrophic zone (661.0+/-89.7 versus 362.8+/-71.6 and 353.0+/-93.9 microm, respectively). The increased size of the growth plate was associated with a greater number of chondrocytes and modifications in their structure, particularly in the hypertrophic zone adjacent to bone. In this zone, chondrocytes of NX animals were significantly (P<0.05) smaller (12080.4+/-1158.3 microm3) and shorter (34.1+/-2.5 microm) than those of SAL (16302.8+/-1483.4 microm3 and 37.8+/-2.0 microm) and SPF (14465.8+/-1521.0 microm3 and 36.3+/-1.8 microm). The interface between the growth plate cartilage and the metaphyseal bone appeared markedly irregular in NX rats. Kinetics of chondrocytes was also modified (P<0.05) in the NX rats, which had lower cell turnover per column per day (5.4+/-0.9), longer duration of hypertrophic phase (89.0+/-15.2 h), and reduced cellular advance velocity (7.4+/-2.2 microm/h) compared with SAL (8.0+/-1.6, 32.1+/-6.7 h, and 11.3+/-2.7 microm/h) and SPF (7.2+/-1.1, 34.8+/-5.1 h, and 10.1+/-2.5 microm/h). Cell proliferation was no different among the three groups. Because the growth plates of SPF and SAL rats were substantially not different, modifications observed in the NX rats cannot be attributed to the nutritional deficit associated with renal failure. These findings indicate that chronic renal failure depresses both the activity of the growth plate cartilage by altering chondrocyte hypertrophy and the replacement of cartilage by bone at the metaphyseal end. The two processes are differentially depressed since cartilage resorption is more severely lowered than cartilage enlargement and this leads to an accumulation of cartilage at the hypertrophic zone.     

Growth hormone secretion from pituitary cells in chronic renal insufficiency.

To examine whether growth hormone (GH) secretion is adversely affected by chronic renal insufficiency (CRI), the GH secretory response of dispersed anterior pituitary cells perifused with GH-releasing hormone (GHRH) was investigated in 5/6 nephrectomized (CRI, N = 18) and sham-operated (N = 18) rats. Two weeks after nephrectomy, during a period of stable uremia, CRI rats had significantly higher serum concentrations (mean +/- SEM) of urea nitrogen and creatinine than sham rats, 16.8 +/- 1.4 mmol/liter (47 +/- 4 mg/dl) and 79.6 +/- 0.0 mumol/liter (0.9 +/- 0.0 mg/dl) versus 6.1 +/- 0.4 mmol/liter (17 +/- 1 mg/dl) and 35.4 +/- 0.0 mumol/liter (0.4 +/- 0.0 mg/dl), respectively (P less than 0.0001). Incremental gains in body weight and nose to tail-tip length of CRI rats over two weeks were also significantly depressed, 53.3 +/- 5.38 g (CRI) versus 87.0 +/- 3.78 g (sham; P less than 0.0001) and 3.2 +/- 0.2 cm (CRI) versus 3.6 +/- 0.1 cm (sham; P less than 0.05). The cumulative food intake as well as food efficiency (g food consumed/g weight gain) were also adversely influenced by the uremic state: food intake 304 +/- 1 g (CRI) versus 397 +/- 6 g (sham; P less than 0.0001) and food efficiency 0.173 +/- 0.013 g/g of weight gain (CRI) versus 0.219 +/- 0.008 g/g of weight gain (sham). No significant difference in GH secretory rate (ng/min/10(7) cells) was found between the uremic and sham animals under basal conditions, 65.2 +/- 2.1 (CRI) and 67.9 +/- 2.2 (sham) or in response to GH-releasing hormone, 282.8 +/- 42.4 (CRI) versus 306.2 +/- 42.6 (sham).(ABSTRACT TRUNCATED AT 250 WORDS)     

Growth hormone and growth hormone-related mRNA in uremic rats: effect of a growth hormone secretagogue

In experimental animals, the decreased growth during mild uremia is not accompanied by a loss in the capacity of the pituitary gland to secrete growth hormone (GH). With the development of orally administered GH "secretagogues" (GHS), it might be possible to stimulate growth during uremia without injections. This study was designed to determine the effects of the GHS, L-163,255. Uremia was induced by 5/6 nephrectomy (NX). GHS was given orally, 3 mg/kg, twice a week. Four groups of animals included: (1) sham-operated, (2) sham-operated, pair-fed, (3) uremic (NX), and (4) uremic, GHS-treated (NX+GHS). Blood sampling was conducted via intra-atrial catheters, and GH was quantitated. Pituitary GH mRNA was measured by Northern blot, and liver GH receptor and insulin-like growth factor-I mRNAs by RNAase protection. Untreated NX animals had a specific decrease in the "mass" of the GH pulses. A burst of GH was induced by GHS, but the pulsatile pattern of GH secretion over 6 h was not affected. An increase or a return to non-uremic levels of GH-related mRNAs occurred after GHS. Thus, GHS stimulated an acute burst of GH secretion and increased specific mRNAs encoding GH-related proteins in uremic animals.     

Cortisone effects on growth, food efficiency, and in vitro growth hormone release

These studies were designed to investigate the cause of growth retardation during glucocorticoid treatment in rats. In young animals, body weights and amounts of food consumed were measured at two-day intervals, beginning at 29 days of age. Average food intake and food efficiency were calculated. Animals were treated with cortisone (CORT, 5 mg/rat/day, s.c.) or saline (SAL) for eight days between 37 and 44 days. Growth hormone (GH) release by dispersed pituitary cells in response to nine concentrations of GH-releasing hormone (GHRH) were tested by in vitro perifusion at 45 and 73 days. As previously shown, CORT caused a cessation of growth during the treatment period, and body weight failed to catch up. Food efficiency was decreased during CORT treatment. All parameters of in vitro GH release including basal GH secretory rate, overall GH response to GHRH, and the GHRH concentration-response curves were significantly increased by CORT in the 45-day-old animals. An age-related increase in GH release was also observed between the 45 and 73 day saline-treated animals. These results support the hypothesis that glucocorticoids inhibit growth by induction of changes in food metabolism and GH secretion. The effect on the pituitary gland itself paradoxically involves an increase in GH secretory capacity in response to GHRH.     

Improvement of growth and food utilization by human recombinant growth hormone in uremia

We compared growth rate, food conversion ratio and morphology of the growth zone in female Sprague-Dawley rats with subtotal nephrectomy or sham operation. Both groups were either given vehicle or 1.4 IU/day recombinant human growth hormone (GH) by s.c. osmotic minipump, or 2.5 IU twice daily intraperitoneally for 14 or 20 days, respectively. Compared to uremic rats infused with vehicle, infusion of GH significantly (P less than 0.01) improved growth; that is, it increased gain of weight (delta 27.0 +/- 7.7 g vs. 11.6 +/- 4.9 g) and length (delta 1.8 +/- 0.3 cm vs. 1.12 +/- 0.44 cm) in ad libitum fed uremic rats. This was accompanied by increased food utilization ratio (0.146 vs. 0.065 g weight gain per g food intake). A similar increment of growth and food utilization ratio was also observed in GH versus solvent infused controls, either pairfed as for the uremic animals or fed ad libitum. Despite administration of GH, growth was not completely restored to normal in uremic animals. Circulating immunoreactive IGF I was not significantly increased by GH infusion in either uremic animals or controls. Histological analyses of the proximal tibia showed increased rate of longitudinal growth, as evaluated by tetracyclin-labeling, and increased volumetric density of primary spongiosa with unchanged width of primary spongiosa trabecules when GH was infused in uremic animals. The data suggest that growth impairment in the uremic rat is partially responsive to GH, and this is not accompanied by an increase of circulating IGF I. Therapeutic trials with recombinant GH in uremic children appear justified.     

Administration of ghrelin to young uraemic rats increases food intake transiently, stimulates growth hormone secretion and does not improve longitudinal growth.

BACKGROUND: Ghrelin administration stimulates appetite and growth hormone (GH) secretion. Whether these effects are preserved in young individuals with chronic renal failure (CRF) and their potential benefit on growth is questioned. METHODS: Three experiments were performed in subtotally nephrectomized young rats (Nx). (i) Food intake was monitored in CRF rats receiving saline intraperitoneally or a ghrelin dose (30 nmol) shown to increase food intake over 2 and 24 h in rats with normal renal function. (ii) Plasma levels of GH were measured after a dose of intravenous ghrelin (3 nmol) was given to three groups of five rats each: Nx, sham-operated fed ad libitum (SAL) and sham-operated pair-fed with Nx (SPF). (iii) Growth of Nx rats treated with intraperitoneal ghrelin (3 nmol) for 7 days (Nx-Ghr) was compared with that of SAL and Nx groups receiving saline (n=8-10 per group). RESULTS: In CRF rats, the dose of 30 nmol of ghrelin increased food consumption for 2 h (1.3+/-0.2 g vs 0.5+/-0.2 g, P<0.05) but not 24-h food intake (12.5+/-0.6 g vs 12.2+/-0.5 g). Ghrelin (3 nmol) increased plasma levels of GH, which were maximal 10 min after injection, no differences being observed among groups (SAL: 666.2+/-104.6 ng/ml; Nx: 691.6+/-90.7 ng/ml; SPF: 577.8+/-125.4 ng/ml). Return to basal GH levels was delayed in Nx. Ghrelin did not improve body length and weight gains, longitudinal bone growth rate or food intake in the Nx-Ghr group. CONCLUSIONS: In young uraemic rats, ghrelin increases appetite but not 24-h food intake, stimulates GH secretion and does not improve growth.     

A new experimental model for secondary hyperparathyroidism

We have developed an animal model to study the pathogenesis of secondary hyperparathyroidism by inducing stable uremia in Sprague-Dawley rats by selective microligation of terminal branches of the left renal artery, followed by right nephrectomy. After 4 weeks the animals were killed, the parathyroid glands were removed and weighed, and blood samples were obtained. Of 30 rats, uremia developed in 22 (73%; uremic group) and eight (27%) died or did not become uremic. A sham-operated group of 15 rats served as control (control group). Creatinine levels were 1.8 +/- 0.5 mg/dl in the uremic group versus 0.5 +/- 0.1 mg/dl in the control group (p less than 0.0001). Parathyroid glands were hyperplastic in all rats with uremia and were heavier than parathyroid glands of control animals (70.3 +/- 26 vs 19.1 +/- 8 micrograms; p less than 0.0001). In the group with uremia, parathyroid hormone levels were increased over those of the control group (112.6 +/- 13 vs 28.9 +/- 6.2 pg/ml; p less than 0.0001), whereas osteocalcin levels were similar (36.6 +/- 11 vs 37.5 +/- 1 ng/ml). Serum calcium, phosphate, and alkaline phosphatase levels were similar in both groups. Our model can be used to test hypotheses concerning the treatment of secondary hyperparathyroidism and the relative pathogenetic relevance of vitamin D deficiency and phosphate retention.     

Effect of growth hormone on kidney growth and glomerular structure

The effect of growth hormones (GH) on renal growth was measured in growing uremic rats using a five-sixths nephrectomy model and GH, 5 mg/kg per day. At the end of 8 weeks, somatic size was significantly smaller in the untreated uremic rats. The uremic rats given GH were the same size as the non-uremic control animals. Organ size (heart, liver and kidney) differed in that only untreated uremic animals had a significantly smaller kidney weight. Despite a five-sixths nephrectomy, the uremic animals receiving GH had kidneys the same size as shamoperated control animals. Renal function was not changed by GH therapy in either control or uremic animals. DNA content expressed as milligrams per kilogram kidney tissue was low only in the untreated uremic rats. Glomerular volume and proximal tubular area were elevated in both groups of uremic animals but were elevated to a significantly greater degree in those receiving GH. GH given in large doses to growing animals appears to induce both somatic and renal growth.     

Intermittent administration of parathyroid hormone (1-37) improves growth and bone mineral density in uremic rats

BACKGROUND: Parathyroid hormone (PTH) is secreted in a pulsatile fashion. Continuous infusion of PTH(1-84) resulted in a net decrease in trabecular bone volume. Differential effects have been reported following an intermittent application of PTH. We investigated the effects of a continuous infusion and of an intermittent (2 times daily subcutaneously) administration of PTH(1-37) on growth and bone mineral density (BMD) in healthy and uremic rats. METHODS: Two-stage subtotal nephrectomy was performed on 130 g female Sprague-Dawley rats. PTH(1-37) or solvent was administered through minipumps in sham-operated and uremic rats (60 microg/kg x day for 2 weeks). The effect of intermittent administration was tested with a subcutaneous injection of solvent: 30 microg/kg PTH(1-37) two times per day, 100 pmol calcitriol (C)/kg two times per day, or both. The length (snout-tailtip) and BMD were measured at the start of uremia and at sacrifice. BMD was measured by peripheral quantitative computer tomography at the proximal tibia, 6 and 12 mm distal of the kneejoint space. Femur bone morphology was assessed by x-rays, and calcium content was measured by atomic absorption spectrophotometry. RESULTS: Length gain was not altered by the continuous infusion of PTH. In contrast, it was significantly increased by intermittent PTH (control solvent 5.35 +/- 0.37 cm vs. control PTH 6.19 +/- 0.47 cm; uremia solvent 4.78 +/- 0.20 cm vs. uremia PTH 6.17 +/- 0.36 cm; P < 0.05). Intermittent PTH but not C increased BMD in uremic rats (Delta total BMD 134 + 13.3 vs. 76.3 +/- 11.5 mg/mL; P < 0.05). X-rays revealed increased bone mass following treatment with PTH but not with C. Uremia decreased bone calcium content (64 +/- 0.3 vs. 73. 3 +/- 2.5 mg/mL), which was normalized by PTH (80 +/- 3.6 mg/mL, P < 0.05) but not by C (69 +/- 1.9 mg/mL). CONCLUSION: Pulsatile administration of PTH does not adversely affect, but improves longitudinal growth independent of concomitant treatment with C. At the same time PTH increases BMD and the calcium content of bone.     

Decreased calcium-sensing receptor expression in hyperplastic parathyroid glands of uremic rats: role of dietary phosphate

BACKGROUND: The abnormal control of parathyroid hormone secretion in chronic renal failure is attributed, in part, to down-regulation of the calcium-sensing receptor (CaR) in hyperplastic parathyroid tissue. The cause of this down-regulation is unknown. Here we examined the roles of uremia and parathyroid hyperplasia on parathyroid gland (PTG) CaR expression in the rat model of renal failure. METHODS: Rats made uremic by 5/6 nephrectomy were maintained for one month on diets containing 0.2% P (low phosphate), 0.5% P (normal phosphate) or 1.2% P (high phosphate); intact rats (controls) were maintained on the normal-phosphate diet. RESULTS: CaR mRNA was reduced only in uremic rats fed the high-phosphate diet (55% less than in controls, P < 0.05). Immunohistochemical staining revealed decreased CaR protein expression in uremic high-phosphate rat PTG compared with controls (41% decrease as determined by computer-assisted quantitation, P < 0.01). PTG size was increased in uremic rats fed the high-phosphate diet compared with controls (2.77 +/- 0.95 vs. 0.77 +/- 0.16 microgram/g body wt, P < 0.0001). There was no increase in PTG size in uremic rats fed the low-phosphate and normal-phosphate diets (0.92 +/- 0.31 and 1.01 +/- 0.31 micrograms/g) compared with controls (0.77 +/- 0.16 microgram/g body wt). Immunohistochemical staining for proliferating cell nuclear antigen in hyperplastic PTG from uremic rats showed that CaR was decreased primarily in areas of active cell proliferation. CONCLUSION: These results suggest that CaR down-regulation cannot be attributed to uremia per se, but rather, is associated with parathyroid cell proliferation. Furthermore, dietary phosphate restriction prevents both the parathyroid hyperplasia and decreased CaR expression in renal failure.     

Evidence for reduced catabolism by the antiglucocorticoid RU 38486 in acutely uremic rats

Previous studies suggested that increased blood levels of, or increased tissue sensitivity to, glucocorticoids may contribute to catabolism in acute uremia. To examine this possibility we determined urea nitrogen (urea-N) appearance, plasma levels of Nt-methylhistidine and the activity of the alkaline myofibrillar proteinase in acutely uremic rats with and without treatment with RU 38486, a selective antiglucocorticoid. Forty-eight hours after bilateral nephrectomy, the rats had markedly elevated serum levels of urea-N, creatinine, potassium and phosphorus. In uremic rats receiving RU 38486, comparable levels of serum creatinine were found, but the serum levels of urea-N (221 +/- 4 vs. 259 +/- 5 mg/dl) and phosphorus (6.5 +/- 0.3 vs. 8.5 +/- 0.4 mmol/l) were significantly decreased as compared to uremic animals without RU 38486. In comparison to sham-operated rats, urea-N appearance (net urea production) was increased by 56% 48 h after bilateral nephrectomy. This increment was almost completely reversed in uremic animals receiving the antiglucocorticoid. In untreated uremic rats, plasma levels of Nt-methylhistidine were 10.3 +/- 0.9 microgram/dl, whereas the administration of RU 38486 caused a significant decline in the levels of this amino acid (7.6 +/- 0.5 microgram/dl). This reduction in Nt-methylhistidine was associated with a concomitant decrease of myofibrillar proteinase activity in muscle tissue homogenates. Compared to sham-operated animals, this proteinase activity was increased by 30% in uremic rats, but was normal in those given RU 38486. Taken together, these data support the view that in acute uremia accelerated ureagenesis occurs, while enhanced muscle protein breakdown, owing to an increment in myofibrillar proteinase activity, provides the necessary amino acid precursors.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of cholinergic muscarinic blockade on growth hormone responses to growth hormone-releasing hormone in uraemic patients.

BACKGROUND: Several alterations in growth hormone (GH) secretion have been reported in patients with chronic renal insufficiency. However, cholinergic modulation of somatotopic cell function has not been fully clarified in uraemic patients. To gain further insight into the disrupted mechanism of GH regulation in chronic renal failure, we investigated whether the blockade of cholinergic muscarinic receptor with pirenzepine could modify the response of GH to its physiological releasing hormone. METHODS: Eight uraemic male patients on peritoneal dialysis and six normal controls were studied. All subjects underwent two endocrine tests in random order. In one of them placebo was administered 60 min before the injection of GH-releasing hormone (GHRH, 100 microg, i.v. in bolus at 0 min). In another the muscarinic blocking agent pirenzepine, 100 mg p.o., was administered at that time. Blood samples for GH were collected at -60, 0, 15, 30, 45, 60 and 90 min. RESULTS: Baseline plasma GH concentrations were similar in patients and controls. GH responses to GHRH were characterized by great interindividual variability in uraemic patients with regard to the amount and the time to maximal peak. In the placebo plus GHRH test, the maximum GH concentrations in patients (14.0 +/- 3.2 microg/l) were similar to those reached by controls (18.0 +/- 3.1 microg/l), although GH secretion was more sustained in patients. The area under the secretory curve (AUC) of GH secretion in patients was also similar to that found in controls (14.4 +/- 2.9 vs 15.4 +/- 3.3 microg/h/l). When subjects were given pirenzepine before GHRH injection an abolishment of GHRH-induced GH release was observed in all controls and in all but one of the uraemic patients. The AUC of GH secretion was, therefore, significantly reduced both in uraemic patients (4.1 +/- 2.0 microg/h/l, P<0.05) and in control subjects (2.0 +/- 0.3 microg/h/l, P<0.05). CONCLUSION: These results suggest that GH secretion in uraemic patients is modulated, at least in part, by a cholinergic mechanism. The muscarinic blockade, possibly acting via an increase in somatostatin release, is able to inhibit GH release in response to direct pituitary stimulation with GHRH.     

Growth hormone and the kidney: the use of recombinant human growth hormone (rhGH) in growth-retarded children with chronic renal insufficiency.

Hypothalamic production of growth hormone releasing hormone stimulates the anterior pituitary gland to release growth hormone (GH). The clinical manifestations of GH on tissues are either direct or are mediated by insulin-like growth factors (IGF). Both the somatic effects of GH and the renal manifestations of an increase in glomerular filtration rate and renal plasma flow are mediated by IGF. The increase in glomerular filtration rate/renal plasma flow that occurs with either exogenous or endogenous GH is not apparent in patients with chronic renal failure (CRF); therefore, it is unlikely that recombinant human growth hormone (rhGH) treatment of patients with CRF will result in glomerular hyperfiltration. Longitudinal studies are required to determine if the glomerulosclerosis and renal functional impairment occurring in GH and growth hormone releasing hormone transgenic mice occurs after rhGH treatment of growth-retarded uremic rats with GH resulted in an improvement in growth velocity. This led to preliminary studies in growth-retarded children with CRF by using rhGH. The acceleration of growth velocity was dramatic despite the fact that GH levels are elevated in uremia. The elevated IGF carrier proteins in uremic children may contribute to the growth retardation. Treatment with rhGH may be efficacious by stimulating a net increase in the free (unbound) IGF levels. Hyposecretion of GH may contribute to the failure to achieve optimal growth after successful renal transplantation. Treatment with rhGH may be efficacious in improving the growth velocity of renal allograft recipients.     

Plasma and pituitary content of growth hormone luteinizing hormone, and prolactin in uremic rats. Effects of chronic infusion of insulin

The influence of chronic renal failure on pituitary content and on serum concentrations of growth hormone (GH), prolactin (PRL), and luteinizing hormone (LH) was studied in chronically uremic rats by comparison with control rats fed ad libitum and diet-restricted rats pair-fed with uremic rats. A decrease of pituitary GH content was found in uremic and diet-restricted rats, in association with a normal circulating GH level. A decrease of pituitary PRL and LH content with respectively high and normal serum values was observed in uremic but not in diet-restricted rats. These data strongly suggest that GH disturbances are related to malnutrition, whereas PRL and LH abnormalities are related to the uremic state per se. As hypoinsulinemia was observed in uremic rats, and as insulin is largely implicated in growth, we have investigated the effects of chronic infusion of insulin, using miniosmotic pumps, on pituitary hormone content. In spite of normalization of circulating insulin levels in uremic rats treated with insulin, pituitary GH, LH, and PRL contents were unaffected. Thus, insulin deficiency did not appear to be responsible for the diminished pituitary reserve of these hormones.     

Reduction of urea generation and muscle protein degradation by adrenalectomy in acutely uremic rats

The effect of adrenalectomy on the enhanced protein degradation in acute uremia was investigated. Therefore, serum urea nitrogen, urea N appearance and Nt-methylhistidine were followed in bilaterally nephrectomized rats. At 48 h after induction of uremia the animals displayed serum urea nitrogen levels of 223 +/- 9.5 mg/dl as compared to 26.0 +/- 1.0 mg/dl in sham-treated rats. This increment was significantly attenuated in acutely uremic, adrenalectomized animals (176 +/- 6.0 mg/dl). When these rats were substituted with corticosterone (5 mg/kg body weight), serum urea nitrogen readily increased to levels of acutely uremic animals with intact adrenal glands (225 +/- 6.0 mg/dl). The net generation of urea, as determined by the urea N appearance, was significantly increased during acute uremia (370 +/- 26 mg/48 h) as compared to SHAM animals (220 +/- 15 mg/48 h). This increment of urea formation could almost be completely reversed by simultaneous adrenalectomy (238 +/- 20 mg/48 h). When these rats were substituted with corticosterone, the urea N appearance rebounded to values quite comparable to acutely uremic rats with intact adrenal glands (363 +/- 30 mg/48 h). To determine whether skeletal muscle proteins might serve as a source for the enhanced protein degradation in acute uremia, plasma levels of Nt-methylhistidine were measured. Bilaterally nephrectomized rats had Nt-methylhistidine values of 9.6 +/- 1.0 micrograms/ml. In acutely uremic rats without adrenal glands, Nt-methylhistidine levels were found to be significantly decreased (6.0 +/- 0.4 micrograms/ml).(ABSTRACT TRUNCATED AT 250 WORDS)     

mRNA expression of the IGF system in the kidney of the hypersomatotropic rat

AIM: To examine the response of the insulinlike growth factor (IGF) system in the kidney during a state of extreme growth. METHODS: We studied the mRNA expression of IGF-I, IGF-I receptor, and IGF-binding proteins (BP) using sensitive RNase protection assays following subcutaneous implantation of growth hormone pituitary cells (GH(3)) in rats. RESULTS: Within 5 weeks, the serum GH levels increased from 18.0 +/- (SE) 5.0 ng/ml in control animals to 389.8 +/- 30.3 ng/ml in GH(3) rats (n = 5, p < 0.001). The circulating IGF-I levels were also elevated. The kidney weights increased from 0.74 +/- 0.01 g in controls to 1.06 +/- 0.03 g in GH(3) animals (n = 5, p < 0.001). Similar changes were observed at week 10. The renal IGF-I mRNA averaged 1.0 +/- (SD) 0.33 relative densitometry units in controls (n = 4) and increased to 2.11 +/- 0.13 relative densitometry units in GH(3) rats (n = 5, p < 0.001). On the other hand, mRNA for the type I IGF receptor decreased in hypersomatotropic rats. Messenger RNAs for IGFBP-1 and IGFBP-4, which have been localized to renal tubules, both decreased significantly following growth induction, while IGFBP-3, the mRNA of which has an interstitial localization, was increased at week 10. CONCLUSION: These data suggest that there is a dynamic relationship between tubular and interstitial compartments with regard to the IGF system in the kidney which may be important in the regulation of the cell mass.     

Persistent downregulation of calcium-sensing receptor mRNA in rat parathyroids when severe secondary hyperparathyroidism is reversed by an isogenic kidney transplantation

Experimental severe secondary hyperparathyroidism (HPT) is reversed within 1 wk after reversal of uremia by an isogenic kidney transplantation (KT) in the uremic rats. Abnormal parathyroid hormone (PTH) secretion in uremia is related to downregulation of CaR and vitamin D receptor (VDR) in the parathyroid glands (PG). The aim of this investigation was to examine the expression of CaR and VDR genes after reversal of uremia and HPT in KT rats. 5/6 nephrectomized rats were kept on a normal or high-phosphorus (hP) diet for 8 wk to induce severe HPT (n = 8 in each group). In another group of seven uremic hP rats, uremia was reversed by an isogenic KT and PG were harvested within 1 wk posttransplant. Plasma urea, creatinine, total calcium, phosphorus, and PTH levels were measured. Parathyroid CaR and VDR mRNA were measured by quantitative PCR. Uremic hP rats had significantly elevated levels of creatinine, urea, and phosphorus (P < 0.001) and developed significant hypocalcemia (plasma calcium 1.83 +/- 0.2 mmol/L; P < 0.001) compared with normal control rats. After KT, the levels were normalized from day 3 to 7: creatinine from 0.117 +/- 0.016 to 0.050 +/- 0.002 mmol/L; urea from 23 +/- 4 to 7 +/- 0.3 mmol/L; phosphorus from 3.9 +/- 0.6 to 1.5 +/- 0.06 mmol/L; calcium from 1.8 +/- 0.2 to 2.5 +/- 0.02 mmol/L. Plasma PTH levels fell from 849 +/- 224 to a normal level of 38 +/- 9 pg/ml (P < 0.01). In uremic rats on a standard diet, CaR mRNA was similar to that of normal control rats, whereas VDR mRNA was significantly decreased. In uremic rats kept on hP diet, CaR mRNA was significantly decreased to 26 +/- 7% of control rats (P = 0.01) and VDR mRNA reduced to 36 +/- 11% (P < 0.01). In KT, previously hP uremic rats, both CaR mRNA and VDR mRNA remained severely reduced (CaR, 39 +/- 7%; VDR, 9 +/- 3%; P < 0.01) compared with normal rats. In conclusion, circulating plasma PTH levels normalized rapidly after KT, despite persisting downregulation of CaR and VDR gene expression. This indicates that upregulation of CaR mRNA and VDR mRNA is not necessary to induce the rapid normalization of PTH secretion from hyperplastic parathyroid glands.     

Diminished response of ovarian cAMP to luteinizing hormone in experimental uremia

In prepuberal female rats with acute bilateral nephrectomy or chronic subtotal nephrectomy, the increase of ovarian cAMP concentration in response to submaximal doses of luteinizing hormone (LH 10 micrograms) and human chorionic gonadotropine (hCG 2.5 IU) was diminished (CO + 2.5 IU hCG 488 +/- 49 pmoles cAMP/mg protein; NX + 2.5 IU hCG 366 +/- 56. P less than 0.05). The cAMP response to follicle stimulating hormone (FSH) was unchanged. The abnormality was found both after administration of LH in vivo and incubation of ovaries with LH in vitro. Similarly, plasma estradiol concentrations in response to submaximal hCG stimulation were diminished. Basal cAMP concentrations and cAMP concentrations after maximal stimulation were unchanged. The defect was observed both in ovaries of untreated prepuberal rats, of pregnant mare serum (PMS)-treated rats (follicular phase) and PMS/hCG-treated rats (luteal phase). Diminished ovarian cAMP response to LH was observed both in parathyroid intact and in parathyroidectomized rats. Administration of 1,25(OH)2D3 in physiological doses (60 ng/kg) to acutely uremic rats restored diminished ovarian cAMP response to submaximal LH stimulation irrespective of parathyroid status. The effect of 1,25(OH)2D3 could not be reproduced by hypercalcemia resulting from intraperitoneal calcium injection. In vivo administration of indomethacin further diminished ovarian cAMP response in uremic animals and had no effect in control animals. Incubation of ovaries with PGE1 and PGE2 increased basal and stimulated cAMP concentrations and abolished the difference between control and uremic animals. The diminished response of ovarian cAMP content to submaximal doses of hCG was not corrected by bromocriptine (1 mg/kg) despite normalization of hyperprolactinemia. The present study shows diminished ovarian cAMP and plasma estradiol response to LH in experimental uremia. It documents a role of 1,25(OH)2D3 and prostaglandins in the genesis of this abnormality.     

Mitochondrial granulation in the proximal renal tubule in uremia

Mitochondria contain electron-dense particles, partly composed of an amorphous form of calcium phosphate. We have used electron microscopy from percutaneous renal biopsy material to analyze mitochondrial granulation in the proximal renal tubule of nonuremic and uremic children. Based on a technique of cutting mitochondria from ten electron micrographs per biopsy, counting the granules in each mitochondrion and weighing the paper, we found that mitochondria of nonuremic children averaged 23.7 +/- 1.2 granules/g paper while uremic children had only 11.8 +/- 1.1 granules/g. The number of granules per gram was unrelated to the serum calcium phosphate solubility product. A significant decrease in calcium granulation in uremia can also be produced experimentally in rats. Control rats averaged 14.7 +/- 1.5 granules/g, while rats made uremic by partial nephrectomy had 6.0 +/- 0.7 granules/g. Treatment of uremic rats with a pharmacological dose of vitamin D restored granulation to normal within 24 h. The significant decrease in calcium phosphate granulation in the renal proximal tubule in uremic children and in experimental animals is probably related to the documented loss of 1 alpha-hydroxylation of vitamin D in uremia.