Experimental Papillary Necrosis of the Kidney: IV. Medullary Plasma Flow

To test the thesis that vasoconstriction plays a significant role in the pathogenesis of papillary necrosis caused by bromoethylamine hydrobromide (BEA), medullary plasma flow was determined in rats treated with BEA. Medullary blood flow was normal ½ to 1 hour after BEA treatment, and was actually elevated 6 hours after BEA. There was no increase in plasma levels of prostaglandins A and E, which would have been expected if there had been medullary ischemia. Pretreatment with reserpine, which inhibited the development of papillary necrosis, had little effect on medullary plasma flow. These observations do not support the notion that vasoconstriction is the mechanism by which BEA causes papillary necrosis.     

Haloalkylamine-induced renal papillary necrosis: a histopathological study of structure-activity relationships.

The haloalkylamine 2-bromoethanamine (BEA) causes necrosis of renal papillae of rats within 24 h of a single intraperitoneal dose greater than or equal to 100 mg/kg. Nine structural analogues of BEA, differing by halide substitution, alkyl chain elongation or amine substitution, were tested for their ability to induce renal papillary lesions in rats. Three compounds (2-chloroethanamine, 3-bromopropanamine and 2-chloro-N,N-dimethylethanamine) induced lesions which were morphologically indistinguishable from those of BEA. All the molecular structural variations investigated reduced papillotoxicity compared with BEA, the parent compound. A variety of non-renal lesions including hepatic, adrenal, testicular and lymphoid necroses were also encountered. The most toxic compound was 2-fluorethanamine, a 5 mg/kg dose of which was lethal and induced renal corticomedullary mineralization and centrilobular hepatic necrosis. One analogue, 3-bromo-2-hydroxypropanamine, caused rapid and extensive necrosis of the adrenal pars fasciculata and reticularis, simulating human Waterhouse Friderichsen syndrome. The three newly identified renal papillotoxins are all theoretically capable of generating direct-acting alkylating species in solution and their activity as direct-acting mutagens in the Ames bacterial mutagenicity test with TA100 (indicating base pair substitution) closely correlated with their potency as papillotoxins. We therefore hypothesize that non-enzymically formed direct-acting alkylating species mediate these papillary lesions, and that the target selectivity of haloalkylamine toxicity most probably results from the accumulation of these alkylating species in papillary tissue.     

The effects of chronic papillary necrosis on acid excretion

Complete papillary necrosis in rats can be induced within 1 month following a single injection of 2-bromoethylamine hydrobromide (BEA) (50 mg, i.v.). Utilizing a combination of clearance and balance techniques the effects of complete absence of the papilla was examined as regards urinary acidification, whole kidney glomerular filtration rate (GFR), single nephron GFR, and morphology. Whole kidney GFR was not different from control, however, the percent filtering juxtamedullary nephrons was markedly diminished (87.2±2.1 vs. 31.5±3.6% filtering, control vs. BEA, respectively,P<0.001) and significantly reduced in the superficial nephrons (80.6±3.6 vs. 62.2±6.1% filtering, control vs. BEA, respectively,P<0.05). There was a significant decrease in juxtamedullary single nephron GFR and an increase in the superficial single nephron GFR as assessed by the quantitative Hanssen's technique in the animals with chronic papillary necrosis. Complete papillary necrosis was associated with normal arterial bicarbonate concentration, pH, and plasma electrolyte concentrations. At the same degree of acidemia (induced by NH₄Cl administration) minimal urinary pH, ammonium excretion, and titratable acid excretion were not different than seen in age matched controls. The response to Na₂SO₄ infusion and phosphate infusion was the same in both groups of animals. The urineblood (U-B)pCO₂, an index of urinary acidification, was identical in BEA and control animals. Scanning electron microscopy showed scarring of the juxtamedullary glomeruli one month after BEA. The papilla was sloughed and lying free in the renal pelvis in every experimental animal. These data demonstrate that complete papillary necrosis is not associated with acidosis nor a defect in urinary acidification.     

Ultrastructural appearances of acute renal papillary lesions induced by aspirin

Renal papillary necrosis develops 16 hr after intravenous administration of aspirin to the uninephrectomised homozygous Gunn rat. Ultrastructural studies show the papillary interstitial cells to be most severely affected, the first changes being visible at 1 hr. Changes in capillaries are late in onset, and this suggests that the lesions are due to a direct toxic effect rather than to ischaemia.     

Early pathophysiological features in canine renal papillary necrosis induced by nefiracetam

To ascertain the early pathophysiological features in canine renal papillary necrosis (RPN) caused by the neurotransmission enhancer nefiracetam, male beagle dogs were orally administered nefiracetam at 300 mg/kg/day for 4 to 7 weeks in comparison with ibuprofen, a non-steroidal anti-inflammatory drug (NSAID), at 50 mg/kg/day for 5 weeks. During the dosing period, the animals were periodically subjected to laboratory tests, light-microscopic, immunohistochemical, and electron-microscopic examinations and/or cyclooxygenase (COX)-2 mRNA analysis. In laboratory tests, a decrease in urinary osmotic pressure and increases in urine volume and urinary lactate dehydrogenase (LDH) level were early biomarkers for detecting RPN. Light-microscopically, nefiracetam revealed epithelial swelling and degeneration in the papillary ducts in week 7, while ibuprofen displayed degeneration and necrosis in the papillary interstitium in week 5. In immunohistochemical staining with COX-2 antibody, nefiracetam elicited a positive reaction within interstitial cells around the affected epithelial cells in the papillary ducts (upper papilla) in week 7, and ibuprofen positively reacted within interstitial cells adjacent to the degenerative and/or necrotic lesions in week 5. Ultrastructurally, nefiracetam exhibited reductions of intracellular interdigitation and infoldings of epithelial cells in the papillary ducts, whereas ibuprofen showed no changes in the identical portions. Thus, the early morphological change in the papilla brought about by nefiracetam was quite different from that elicited by ibuprofen. By the renal papillary COX-2 mRNA expression analysis, nefiracetam exceedingly decreased its expression in week 4, but markedly increased it in week 7, suggesting an induction of COX-2 mRNA by renal papillary lesions. These results demonstrate that the epithelial cell in the papillary ducts is the primary target site for the onset of RPN evoked by nefiracetam.     

Experimental renal papillary necrosis. Effects of beta adrenergic receptor blockade, heparin and hydrocortisone.

Renal papillary necrosis was induced in rats by daily subcutaneous injection of 15 mg 2-bromoethylamine hydrobromide (BEA) per 100 g of body weight for 2 successive days. This dose was 50% higher than that reported previously. Beta adrenergic receptor blockade with oxyprenolol did not influence the kidney damage.The administration of heparin did not show any effect. The doses applied did not induce the incoagulability for a sufficient period of time. On the contrary, treatment with hydrocortisone decreased papillary necrosis without inducing increased diuresis.     

Experimental analgesic nephropathy: changes in renal structure and urinary concentrating ability in Fischer 344 rats given continuous low doses of aspirin and paracetamol

Long-term treatment with aspirin and paracetamol produced renal papillary necrosis in female Fischer 344 rats. Aspirin (230 mg/kg body weight/day) and paracetamol (380 mg/kg body weight/day) were dissolved in drinking water and given continuously for up to 65 weeks. Renal morphological changes were examined between 21 weeks and 65 weeks of commencement of analgesic treatment using light and electron microscopy, and were compared with age-matched controls. Structural damage initially occurred in the mid-papillary region, and specifically involved the interstitial cells and interstitial matrix. Necrosis of the epithelium of the thin limbs of the loop of Henle was present only after interstitial changes were well established. Cortical interstitial fibrosis and tubular atrophy occurred after renal papillary changes were observed. There was no evidence of significant vascular damage. Urinary concentrating ability was measured sequentially during the period of analgesic treatment. A decrease in urine concentrating ability was present when early changes to the interstitial cells and matrix were observed, and concentrating ability continued to decrease in parallel with increasing morphological damage. This study describes an animal model of analgesic-induced nephropathy, enabling early morphological changes to be studied and correlated with renal functional changes.     

Experimental Papillary Necrosis of the Kidney: I. Morphologic and Functional Data

Papillary necrosis was produced in rats by a single intravenous injection of bromoethylamine hydrobromide (BEA). The earliest changes as seen by light microscopy were necroses of the limbs of Henle and eosinophilic droplets in collecting ducts. Complete necrosis of the papilla took place between 4 and 7 days and the dead papilla was usually sequestered completely by 21 days. Cortical changes occurred secondary to papillary necrosis. Tubular atrophy and loss was greatest in the deeper parts of the central cortex, the more superficial nephrons frequently being spared. The perihilar cortex was the least involved. This distribution was considered to be related to the respective lengths of the limbs of Henle, nephrons with limbs extending into the papilla being those undergoing change. Increased urine output occurred during the first day and continued thereafter. There was a profound defect in concentrating ability.     

Lesions of the renal papilla induced by paracetamol

The acute nephrotoxic effects of paracetamol in the uninephrectomized homozygous Gunn rat are different from those of aspirin. Both compounds induce renal papillary necrosis but paracetamol produces accumulation of non-cellular material in the interstitial space, less damage to interstitial cells, more damage to tubular epithelium, and more severe necrosis of proximal convoluted tubules. In both cortex and papilla only a small fraction of the cells at risk are affected. It is concluded that the findings are consistent with a synergistic nephrotoxic effect between the two compounds, but that the lesions are not sufficiently severe for the natural history of analgesic nephropathy to be wholly explicable by such synergism.     

Renomedullary and intestinal hyaluronan content during body water excess: a study in rats and gerbils

Our previous studies in rats have suggested a role for renomedullary hyaluronan (HA) in water homeostasis. The gerbil is known for its unique ability to conserve water. In the present study renal papillary and intestinal HA were compared between groups of anaesthetized gerbils and rats before and after up to 6 h of i.v. water loading. Baseline papillary HA in gerbils was only 37 % of that in the rat. Water loading in rats increased the papillary HA content. Elevation was maximal (+27 %,   P < 0.05  ) after 2 h of water loading and then declined to control levels after 6 h of water loading (+3 %, n.s.). In contrast, the gerbil responded with a decreased papillary HA content during water loading. The depression was maximal after 2 h (-49 %,   P < 0.05  ) and was still 41 % below the control values after 6 h (   P < 0.05  ). The urine flow rate increased rapidly in the rat and its maximum, 21 times above the control level (   P < 0.05  ), occurred at the HA peak, i.e. after 2 h of water loading while in the gerbil, the urine flow rate increased slowly and slightly and was only six times above control values after 6 h of water loading (   P < 0.05  ). The HA content along the intestine was similar in the two species: lowest in the duodenum and jejunum and highest in the distal colon. To conclude, in the rat, the elevation of papillary interstitial HA during acute water loading would counteract water reabsorption by changing the physico-chemical characteristics of the interstitial matrix favouring rapid water diuresis. This would work as a complement to the powerful regulation by ADH. The gerbil has a diametrically different regulation of papillary HA turnover during water loading. The decreased papillary HA level during water loading and the slow and small diuretic response may represent a genetic difference in adaptation to enhance the ability to conserve water in an arid environment.     

Experimental renal papillary necrosis in the rat: The selective vulnerability of medullary structures to injury

Summary Acute renal papillary necrosis was produced in rats by the administration of ethyleneimine. Low doses resulted in necrosis of interstitial cells, thin limbs of the loops of Henle and vasa recta, while collecting ducts were spared (subtotal renal papillary necrosis). High doses resulted in necrosis of all elements of the papilla (total renal papillary necrosis). Although the ranges of the doses that produced these two patterns of necrosis overlapped, it is clear that there is a dose dependent selective vulnerability of renal medullary structures to injury by the toxic agent studied.     

Spontaneous papillary necrosis in the heterozygous Gunn rat

Spontaneous papillary necrosis develops in aging heterozygous non-jaundiced Gunn rats. The lesion is situated in the subapical or mid papilla and in its earliest stages is manifest by the appearance of amorphous material in the interstitial space. This is seen in plastic-embedded sections taken from rats 6 months old. In its later stages, the accumulation of amorphous material is accompanied by loss of interstitial cells and cyst formation, but there is no associated inflammatory reaction. The largest lesions are found in the oldest rats, but even in these animals the macroscopic appearance of the papilla is normal. Similar papillary changes were not found in albino or homozygous Gunn rats, but in aging albino rats there was loss of papillary interstitial cells without accumulation of amorphous material.     

Glomerular lesions in experimental renal papillary necrosis. I. Ultrastructural aspects and some pathogenic considerations.

Papillary necrosis was induced in rats by a single intravenous injection of bromoethylamine hydrobromide (BEA). From 7 days on glomerular lesions were recognized. They consisted of electron dense deposits mainly subepithelial in location; mild mesangial hypercellularity and matrix increase. Immunofluorescence with anti-rat gammaglobulin was positive, showing granular fluorescence in relation with basement membrane and mesangium. The possibility is raised that these lesions are due to the pathogenic action of immune complexes, the antigen being one arising during the necrosis of the renal papilla. It is also suggested that this mechanism can be operative in ths human being in cases of papillary necrosis of the kidney.     

Interferon-gamma induced lethality in the late phase of Plasmodium vinckei malaria despite effective parasite clearance by chloroquine.

A combination therapy was tested consisting of chloroquine and interferon-gamma (IFN-gamma) in the late phase of blood-stage Plasmodium vinckei malaria in BALB/c mice. When mice were treated with three times 300 micrograms chloroquine at 24-h intervals starting at a parasitemia of 30%-50%, only 5 of 14 mice (36%) died 2-4 days after initiation of therapy. However, when infected mice received chloroquine plus 1 microgram IFN-gamma at the same time, 14 of 18 mice (78%) died 0.5-3 days after start of therapy (p < 0.05) despite clearance of parasitemia. The histopathology from mice dying after combination therapy revealed interstitial leukocyte infiltration of lung tissue, severe liver cell necrosis and kidney tubular necrosis. Pretreatment of P. vinckei-infected mice with pentoxifylline, a phosphodiesterase inhibitor, led to a significant decrease of IFN-gamma-induced lethality (p < 0.05). In contrast, pretreatment with neutralizing antibodies to tumor necrosis factor or with L-N-monomethyl arginine, the latter an inhibitor of the nitric oxide synthase, significantly increased lethality (p < 0.05).     

Healed experimental renal papillary necrosis and cortical scarring in the rat from 2-bromoethylamine hydrobromide

Summary Male Sprague-Dawley rats were each given a single subcutaneous injection of an aqueous solution of bromoethylamine hydrobromide (BEA) at dose levels of 80 mg/kg (16 rats), 125 mg/kg (15 rats) and 250 mg/kg (16 rats) or a single subcutaneous injection of water (controls, 15 rats). The dose levels were chosen so as to cause renal papillary injury varying from minor necrotic foci to necrosis and subsequent sloughing of the entire papilla. The animals were killed after 5 months and the kidneys were weighed and examined macroscopically and microscopically for the presence of RPN and cortical scarring. Macroscopically evident RPN occurred in 18 of 43 surviving BEA-treated rats, bilaterally in 16 and unilaterally in 2. Bilateral asymmetry of the extent of sloughing was evident. All kidneys with macroscopic RPN exhibited cortical scarring. Asymmetry of the extent of atrophy and scarring in some animals resulted in a significant unilateral reduction in renal weight and a significant contralateral compensatory hypertrophy. Twenty-three of the 25 BEA-treated rats without macroscopically evident RPN exhibited minor, histologically visible lesions of the renal papillae including necrosis of loops of Henle in the presence of intact collecting ducts. Only 2 of these animals exhibited tiny, unilateral cortical scars, and renal weights did not differ significantly from those of controls. It may therefore be concluded, contrary to certain published proposals: that experimental RPN may be followed by severe renal cortical scarring, reduction in renal size and (in the presence of asymmetrical lesions) compensatory renal hypertrophy, and that necrosis of thin limbs of loops of Henle does not appear to lead to frequent or severe cortical scarring.     

Changes of sodium and urea concentrations in the renal papillary interstitial fluid on dehydration of rats

1. The isolated renal papillae of a rat were centrifuged in a two tube assembly which allowed fluid from the tissue to separate into the lower tube.2. The papillae were centrifuged for 15 min at 300 g and 1500 g consecutively.3. After intraperitoneal injection of Na (131)I-diatrizoate, the activities of urine, and fluid samples obtained from the papilla, were compared. It was found at 1500 g that the median value for papillary fluid activity was 1.52% of the activity of urine. This is evidence that the papillary fluid was virtually free from any contamination from the terminal collecting ducts.4. It is considered that the fluid sample obtained from the papillae by centrifugation at 1500 g is a representative and reproducible sample of interstitial fluid.5. The method was used to demonstrate changes in solute concentrations in the renal papillary fluid, following dehydration of rats.     

PAX2-siRNA在梗阻性肾病大鼠体内的作用研究

目的 通过RNA干扰和基因转染技术,研究沉默PAX2基因对UUO大鼠肾间质纤维化的影响.方法 将PAX2-siRNA转染至UUO大鼠肾脏被膜下,将64只幼年雄性大鼠随机分为:阴性对照组(NC)32只;沉默组(RNAi) 32只,于转染后3、5、7、14天分为4组,每组8只,留取肾组织标本,应用Real-Time PCR及Western blot检测肾皮质PAX2 mRNA及其蛋白的沉默情况,HE染色和Masson染色方法在光镜下观察肾小管损伤情况和肾间质纤维化程度.结果 ①沉默组与对照组比较PAX2 mRNA和PAX2蛋白表达量降低,有统计学意义(P＜0.05),②HE和Masson染色观察到对照组随梗阻时间延长,肾小管损伤逐渐明显,肾间质中可见胶原蛋白逐渐增加.沉默组与对照组比较在7d以前无明显变化(P＞0.05);在梗阻14d沉默组与对照组比较肾小管损伤减轻、肾间质病变减低、胶原纤维沉积减少(P＜0.05).结论 在UUO大鼠模型中,梗阻早期沉默PAX2基因对肾间质纤维化进程无明显影响,梗阻晚期可减轻肾小管损伤及肾间质纤维化病变,对肾间质纤维化有明显治疗作用.     

Intratubular calcium phosphate deposition in acute analgesic nephropathy in rabbits.

Earlier chronic studies using both animal and human autopsy material have suggested that the initial lesion of analgesic nephropathy is papillary necrosis with secondary cortical interstitial nephritis. The present study was designed to define ultrastructural changes in renal tubules exposed to high levels of analgesics. Female New Zealand White rabbits were given 5-7 g APC and sacrificed after periods of 6 to 36 hours. As early as 6 hours after treatment, hydroxyapatite crystallite aggregates were seen impacted in Segment III (straight segment) of the proximal tubule at the cortico-medullary junction. Ultrastructural changes included selective calcification of the brush border, mitochondrial calcification, and peroxisomal changes. It was concluded, on the basis of ultrastructural changes, that calcification of altered tubular cells may be an initial event in analgesic nephropathy.     

Association between congenital defects in papillary outgrowth and functional obstruction in Crim1 mutant mice

Crim1 hypomorphic (Crim1(KST264/KST264)) mice display progressive renal disease characterized by glomerular defects, leaky peritubular vasculature, and progressive interstitial fibrosis. Here we show that 27% of these mice also present with hydronephrosis, suggesting obstructive nephropathy. Dynamic magnetic resonance imaging using Magnevist showed fast development of hypo-intense signal in the kidneys of Crim1(KST264/KST264) mice, suggesting pooling of filtrate within the renal parenchyma. Rhodamine dextran (10 kDa) clearance was also delayed in Crim1(KST264/KST264) mice. Pyeloureteric peristalsis, while present, was less co-ordinated in Crim1(KST264/KST264) mice. However, isolated renal pelvis preparations suggest normal pelvic smooth muscle contractile responses. An analysis of maturation during the immediate postnatal period [postnatal day (P) 0-15] revealed defects in papillary extension in Crim1({KST264/KST264) mice. While Crim1 expression is weak in pelvic smooth muscle, strong expression is seen in the interstitium and loops of Henle of the extending papilla, commencing at the tip of the P1 papilla and disseminating throughout the papilla by P15. These results, as well as implicating Crim1 in papillary extension and pelvic smooth muscle contractility, highlight the previously unrecognized association between defects in papillary development and progression to chronic kidney disease later in life.