JOURNAL
OF SURGICAL
RRSRARCH
51.253-258
(19%)
Urinary PGE, in Rats with Chronic Partial Unilateral Ureteral Obstruction’** BASHIR R. SANKARI, M.D., GEORGE F. STEINHARDT, M.D.,3 LUIS SALINAS-MADRIGAL, M.D., AND LESLIE A. SPRY, M.D.4 Division
of Urology, Department Department of Pathology,
of Surgery and Division of Nephrology, Department of Internal Medicine, St. Louis University School of Medicine, St. Louis, Missouri 63104 Submitted
for publication
effects of complete ureteral obstruction upon renal physiology and pathophysiology. Acute and chronic complete ureteral obstruction, with and without release, results in a remarkable decrease in glomerular filtration rate (GFR) with variable recovery over time. The most common form of obstructive uropathy seen in the pediatric age group is chronic partial obstruction. A method of partially obstructing the ureters was described in dogs in 1962 [4] and was successfully applied to newborn rats [5]. Many studies have addressed the functional and morphological renal changes that occur in chronic partial unilateral ureteral obstruction (UUO) [6-g]. From these hemodynamic studies, it was evident that local intrarenal mechanisms govern the glomerular adaptation to ureteral obstruction. Prostaglandins (PGS) have been the subject of intense investigation as modulators of renal hemodynamics and tubular function [ 10-131. Renal PGS have been postulated to participate in the functional changes in renal blood flow (RBF) and GFR observed in ureteral obstruction [6, 141, and to play a role in the postobstructive diuresis and natriuresis observed following relief of complete bilateral ureteral obstruction (BUO) [ 151. The purpose of this experiment was to examine the role of urinary prostaglandin E, (PGE,) in chronic partial UUO in a young rat model followed into adulthood.
Urinary prostaglandin E, (PGE,) was measured in Munich-Wistar rats with surgically created chronic partial unilateral ureteral obstruction (UUO). Mean values of bladder urine PGE, were higher in sham than in UUO (24.5 * 14.4 vs 12.9 & 8.2 ng/mg creatinine, respectively, P < 0.05). Following diuresis, both ureters were cannulated and urine was collected. PGEz excretion was increased in sham (66.5 + 34.4 and 70.1 + 44.5 ng/mg creatinine, left and right, respectively). But in UUO, the obstructed kidney excreted less PGE, than the contralateral kidney (32.1 + 6.0 vs 62.3 + 40.4 ng/mg creatinine, obstructed vs contralateral, respectively, P = 0.08). PGE, synthesis was then determined in separated renal medullary and cortical slices. Renal medullary slices from kidneys with severe obstruction synthesized less PGE, than the contralateral unobstructed side (3.30 f 1.22 vs 10.52 + 3.23 ng/mg wet wt-30 min, respectively, P < 0.05) and failed to respond to arachidonic acid stimulation with any significant increase in PGE, synthesis (3.30 f 1.22 vs 4.47 f 1.04 ng/mg wet wt-30 min, baseline vs stimulated). In contrast, contralateral unobstructed kidney slices responded with a significant increase in PGE, synthesis (10.62 + 3.23 vs 21.10 + 2.50 ng/mg wet wt-30 P -C 0.05). We conclude min, baseline vs stimulated, that chronic partial UUO in the Munich-Wistar rats o 1991 resulted in significantly less PGE, elaboration. Academic
Press,
May 29, 1990
Inc.
MATERIALS INTRODUCTION
AND
METHODS
Thirty weanling Munich-Wistar rats were anesthetized with intraperitoneal pentobarbital(80 mg/kg body wt.). Fifteen animals constituted the control group and underwent sham surgery. Partial unilateral ureteral obstruction was accomplished in 15 animals. The obstruction was created by burying the isolated left ureter in a l-cm groove in the psoas muscle according to the technique of Ulm and Miller [4]. The muscle was closed over the ureter using 5-O Prolene. The rats were fed standard rat chow (Purina) and given free access to water. At 6 months of age, the animals were anesthetized with intra-
Obstructive uropathy is a common cause of renal failure and end-stage renal disease in the pediatric population [l]. Many previous reports [2,3] have examined the
1 Presented at the AUA Annual Meeting, Dallas, TX, May 1989. ’ Supported by a Fleur De Lis Grant, St. Louis, MO. 3 To whom reprint requests should be addressed at Pediatric Urology, 1465 South Grand Boulevard, St. Louis, MO 63104. ’ Current address: 7520 North Hampton, Lincoln, NE 68506. 253
0022-4804/91
$1.50
Copyright 0 1991 by Academic Press, Inc. All rights of reproduction in any form reserved.
254
JOURNAL
OF SURGICAL
RESEARCH:
muscular ketamine and innovar. Bladder urine was collected by aspiration on opening the abdominal cavity. This urine was assumed to represent steady-state conditions. After the bladder urine was obtained, 5 cc of normal saline was infused in the vena cava to promote diuresis. Small tapered polyethylene tubes were passed up each ureter and tied in place. The initial urine obtained by passage of the catheter into a dilated ureter was discarded. Urine was then collected via the polyethylene tubes for a period of 2 to 4 hr. This urine was assumed to represent a period of diuresis. In some instances, an additional 2.5 cc of intravenous saline was given to facilitate urine collection. The magnitude of ureteral obstruction was graded on the basis of visual inspection of the ureter proximal to the obstruction. Mild obstruction was defined as a dilated ureter with a diameter less than three times the diameter of the contralateral untouched ureter. Severe obstruction was defined as a ureter with a diameter more than three times the diameter of the contralateral untouched ureter. Ureters with no visible dilatation were graded as zero obstruction. Following urine collection, blood was aspirated from the vena cava for creatinine and blood urea nitrogen (BUN) measurements. The kidneys were then removed and stored in formalin for histologic analysis. The collected urine was labeled and frozen for later PGE, assay. Urinary
PGE, Assay
Urine was stored at -20°C until the time of radioimmunoassay. PGE, was measured in unextracted urine by radioimmunoassay. PGE, antisera were purchased from Regis Chemical Co. (Morton Grove, IL). Goat anti-rabbit antibody was obtained from Antibodies, Inc. (Davis, CA). The details of the radioimmunoassay have been described elsewhere [ 161. Urinary and serum creatinine levels were determined by the alkaline picrate method and results for urinary PGE, were expressed in nanograms of PGE, per milligram of creatinine (ng PGE,/mg treat). The values were expressed as means f SD. Paired and unpaired Student t tests were used to compare mean differences. Renal Slice Preparation
VOL.
51, NO. 3, SEPTEMBER
1991
TABLE Urinary
Prostaglandin Control group 1 (n = 15)
Weight (g) BUN Creatinine Bladder (PGE,/mg treat) Left ureteral (PCE,/mg treat) Right ureteral (PGE,/mg treat)
1 E, Values Unilateral obstruction group 2 (n = 13)
P value
299 2 36 30.4 _t 5 0.92 + 0.2
3 13 * 33 34.3 k 5.3 0.85 t 0.27
N.S. N.S. N.S.
24.5 t 14.4
12.9 + 8.2
OF SURGICAL
RRSRARCH
51.253-258
(19%)
Urinary PGE, in Rats with Chronic Partial Unilateral Ureteral Obstruction’** BASHIR R. SANKARI, M.D., GEORGE F. STEINHARDT, M.D.,3 LUIS SALINAS-MADRIGAL, M.D., AND LESLIE A. SPRY, M.D.4 Division
of Urology, Department Department of Pathology,
of Surgery and Division of Nephrology, Department of Internal Medicine, St. Louis University School of Medicine, St. Louis, Missouri 63104 Submitted
for publication
effects of complete ureteral obstruction upon renal physiology and pathophysiology. Acute and chronic complete ureteral obstruction, with and without release, results in a remarkable decrease in glomerular filtration rate (GFR) with variable recovery over time. The most common form of obstructive uropathy seen in the pediatric age group is chronic partial obstruction. A method of partially obstructing the ureters was described in dogs in 1962 [4] and was successfully applied to newborn rats [5]. Many studies have addressed the functional and morphological renal changes that occur in chronic partial unilateral ureteral obstruction (UUO) [6-g]. From these hemodynamic studies, it was evident that local intrarenal mechanisms govern the glomerular adaptation to ureteral obstruction. Prostaglandins (PGS) have been the subject of intense investigation as modulators of renal hemodynamics and tubular function [ 10-131. Renal PGS have been postulated to participate in the functional changes in renal blood flow (RBF) and GFR observed in ureteral obstruction [6, 141, and to play a role in the postobstructive diuresis and natriuresis observed following relief of complete bilateral ureteral obstruction (BUO) [ 151. The purpose of this experiment was to examine the role of urinary prostaglandin E, (PGE,) in chronic partial UUO in a young rat model followed into adulthood.
Urinary prostaglandin E, (PGE,) was measured in Munich-Wistar rats with surgically created chronic partial unilateral ureteral obstruction (UUO). Mean values of bladder urine PGE, were higher in sham than in UUO (24.5 * 14.4 vs 12.9 & 8.2 ng/mg creatinine, respectively, P < 0.05). Following diuresis, both ureters were cannulated and urine was collected. PGEz excretion was increased in sham (66.5 + 34.4 and 70.1 + 44.5 ng/mg creatinine, left and right, respectively). But in UUO, the obstructed kidney excreted less PGE, than the contralateral kidney (32.1 + 6.0 vs 62.3 + 40.4 ng/mg creatinine, obstructed vs contralateral, respectively, P = 0.08). PGE, synthesis was then determined in separated renal medullary and cortical slices. Renal medullary slices from kidneys with severe obstruction synthesized less PGE, than the contralateral unobstructed side (3.30 f 1.22 vs 10.52 + 3.23 ng/mg wet wt-30 min, respectively, P < 0.05) and failed to respond to arachidonic acid stimulation with any significant increase in PGE, synthesis (3.30 f 1.22 vs 4.47 f 1.04 ng/mg wet wt-30 min, baseline vs stimulated). In contrast, contralateral unobstructed kidney slices responded with a significant increase in PGE, synthesis (10.62 + 3.23 vs 21.10 + 2.50 ng/mg wet wt-30 P -C 0.05). We conclude min, baseline vs stimulated, that chronic partial UUO in the Munich-Wistar rats o 1991 resulted in significantly less PGE, elaboration. Academic
Press,
May 29, 1990
Inc.
MATERIALS INTRODUCTION
AND
METHODS
Thirty weanling Munich-Wistar rats were anesthetized with intraperitoneal pentobarbital(80 mg/kg body wt.). Fifteen animals constituted the control group and underwent sham surgery. Partial unilateral ureteral obstruction was accomplished in 15 animals. The obstruction was created by burying the isolated left ureter in a l-cm groove in the psoas muscle according to the technique of Ulm and Miller [4]. The muscle was closed over the ureter using 5-O Prolene. The rats were fed standard rat chow (Purina) and given free access to water. At 6 months of age, the animals were anesthetized with intra-
Obstructive uropathy is a common cause of renal failure and end-stage renal disease in the pediatric population [l]. Many previous reports [2,3] have examined the
1 Presented at the AUA Annual Meeting, Dallas, TX, May 1989. ’ Supported by a Fleur De Lis Grant, St. Louis, MO. 3 To whom reprint requests should be addressed at Pediatric Urology, 1465 South Grand Boulevard, St. Louis, MO 63104. ’ Current address: 7520 North Hampton, Lincoln, NE 68506. 253
0022-4804/91
$1.50
Copyright 0 1991 by Academic Press, Inc. All rights of reproduction in any form reserved.
254
JOURNAL
OF SURGICAL
RESEARCH:
muscular ketamine and innovar. Bladder urine was collected by aspiration on opening the abdominal cavity. This urine was assumed to represent steady-state conditions. After the bladder urine was obtained, 5 cc of normal saline was infused in the vena cava to promote diuresis. Small tapered polyethylene tubes were passed up each ureter and tied in place. The initial urine obtained by passage of the catheter into a dilated ureter was discarded. Urine was then collected via the polyethylene tubes for a period of 2 to 4 hr. This urine was assumed to represent a period of diuresis. In some instances, an additional 2.5 cc of intravenous saline was given to facilitate urine collection. The magnitude of ureteral obstruction was graded on the basis of visual inspection of the ureter proximal to the obstruction. Mild obstruction was defined as a dilated ureter with a diameter less than three times the diameter of the contralateral untouched ureter. Severe obstruction was defined as a ureter with a diameter more than three times the diameter of the contralateral untouched ureter. Ureters with no visible dilatation were graded as zero obstruction. Following urine collection, blood was aspirated from the vena cava for creatinine and blood urea nitrogen (BUN) measurements. The kidneys were then removed and stored in formalin for histologic analysis. The collected urine was labeled and frozen for later PGE, assay. Urinary
PGE, Assay
Urine was stored at -20°C until the time of radioimmunoassay. PGE, was measured in unextracted urine by radioimmunoassay. PGE, antisera were purchased from Regis Chemical Co. (Morton Grove, IL). Goat anti-rabbit antibody was obtained from Antibodies, Inc. (Davis, CA). The details of the radioimmunoassay have been described elsewhere [ 161. Urinary and serum creatinine levels were determined by the alkaline picrate method and results for urinary PGE, were expressed in nanograms of PGE, per milligram of creatinine (ng PGE,/mg treat). The values were expressed as means f SD. Paired and unpaired Student t tests were used to compare mean differences. Renal Slice Preparation
VOL.
51, NO. 3, SEPTEMBER
1991
TABLE Urinary
Prostaglandin Control group 1 (n = 15)
Weight (g) BUN Creatinine Bladder (PGE,/mg treat) Left ureteral (PCE,/mg treat) Right ureteral (PGE,/mg treat)
1 E, Values Unilateral obstruction group 2 (n = 13)
P value
299 2 36 30.4 _t 5 0.92 + 0.2
3 13 * 33 34.3 k 5.3 0.85 t 0.27
N.S. N.S. N.S.
24.5 t 14.4
12.9 + 8.2
