Sandra L. Garber

Effect of Relaxin in Two Models of Renal Mass Reduction

Background: Relaxin (Rlx), a 6-kD protein hormone, belongs to the insulin growth factor family. We have previously shown that Rlx reduces interstitial fibrosis in a model of chronic papillary necrosis. Hypothesis: The purpose of this study was to extend these observations to a model of renal injury induced by mass reduction. Material and Methods: Renal mass was reduced by either infarction or surgical excision of both poles, with removal of the contralateral kidney. Two weeks later, creatinine clearance was done and animals from both groups implanted with osmotic pumps delivering either Rlx (2 µg/h) or vehicle (Veh). Treatment was continued for 4 weeks. The severity of the glomerular injury was quantified by planimetric measurements. Renal function was assessed by creatinine clearance and plasma creatinine. Results: Rlx significantly decreased systolic blood pressure in animals with infarction. This was accompanied by a decrease in serum creatinine and a slight improvement in creatinine clearance. The severity of the glomerular lesion was reduced by Rlx (sclerosis index, Veh 1.16 ± 0.13 vs. Rlx 0.74 ± 0.16, p = 0.037). In the excision group the animals were normotensive. In this group, Rlx treatment was accompanied by a decrease in serum creatinine (Veh 1.01 ± 0.03 vs. Rlx 0.81 ± 0.05 mg/dl, p = 0.02) and an increase in GFR (Veh 0.90 ± 0.14 vs. Rlx 1.33 ± 0.11 ml/min, p = 0.03). The sclerosis index was also reduced. Conclusion: Rlx decreases renal injury by at least two mechanisms, one by lowering blood pressure as seen in the infarction model, the other independent of blood pressure as seen in the normotensive excision model where there was also a significant functional improvement.

Evolution of experimentally induced papillary necrosis to focal segmental glomerulosclerosis and nephrotic proteinuria

Bromoethylamine (BEA)-induced papillary necrosis is a reproducible model for analgesic nephropathy. We induced this lesion in groups of male Sprague-Dawley rats and followed the functional and histological changes for 1 year. We found that by 1 month, necrosis of the papilla was complete, glomerular filtration rate was depressed, and urine albumin excretion was increased. There was an extensive interstitial fibrosis characterized by a mononuclear cell infiltrate and patchy tubular atrophy. By 6 months, there was re-epithelialization of the papillary stump accompanied by a marked increase in albuminuria and an improvement in concentrating ability. Changes seen at 9 months were more advanced. There was extensive cortical fibrosis manifested by pitting of the surface of the kidney. At 1 year, renal function remained impaired (creatinine clearance reduced by 65% to 0.26 mL/min/100 g), and the animals were now markedly nephrotic, with albuminuria of 254 mg of albumin/24 h. In the BEA rats, there was selective destruction of the deep nephrons leading to an increase in the volume-ratio of superficial to deep nephrons. Glomerular changes, affecting approximately 60% of the glomeruli, were characteristic of focal segmental glomerular sclerosis. This model of papillary necrosis/interstitial fibrosis is associated with chronic renal insufficiency and leads to the development of focal glomerular sclerosis and nephrotic proteinuria by 6 to 12 months after its induction.

Xanthopterin-induced renal dysfunction: A reversible model of crystal nephropathy

Xanthopterin (XPT), an unconjugated pteridine compound, affects cell growth and differentiation. When injected into rats, XPT has caused changes that have been interpreted as renal growth and hypertrophy. In the present study, we investigated the effect of intraperitoneal administration of XPT on the renal function in the rat. XPT administration was associated with polyuria and a reversible form of nonoliguric acute renal failure (ARF), with renal function declining maximally after 2 days and returning to normal after 7 days. The polyuria was due, at least in part, to a concentrating defect that was vasopressin resistant. The ability of XPT to induce ARF was modulated by dietary salt intake, being enhanced by a low-sodium diet and prevented by a high sodium intake. Histological examination of the kidneys showed intratubular crystal deposition and acute tubule necrosis, suggesting that XPT induces crystal nephropathy. There was an increase in wet and dry weights of the kidney and an increased DNA/protein ratio, compatible with a hyperplastic response. Because the severity of other crystal nephropathies may be modulated by urine flow rate and pH, we studied the ability of water diuresis or alkaline diuresis to protect against XPT-induced ARF. Both water diuresis and HCO3 loading blunted the ability of XPT to decrease renal function. The change in renal function induced by XPT in the various groups was paralleled by corresponding changes in the levels of XPT-like substances in the kidney and by the amount of crystal deposition. Thus, XPT injection induces crystal nephropathy, the severity of which can be modulated by dietary salt intake, urine pH, and urine flow rate.

Effect of acute and chronic glutathione depletion on renal function in the rat

Renal function was evaluated in normal and acid-loaded rats following acute and chronic depletion of glutathione (GSH) by buthionine sulfoximine (BSO). Creatinine clearance and fractional excretion of electrolytes were normal. There was no acidification or concentration defect detected in animals with acute or chronic GSH depletion.

Differential effect of xanthopterin and biopterin on cell growth

1. Xanthopterin inhibited proliferation of primary renal proximal tubule cells (RPTC) and LLC-PK1 cells while in a growth phase but when incubated at confluence the cells were relatively insensitive. 2. The growth of malignant human prostate PC-3 cells was also inhibited by xanthopterin in a concentration and time dependent manner. 3. Dunning R3327 AT-3 rat prostate tumor cells which were exposed to xanthopterin in vitro before their in vivo inoculation resulted in smaller tumours while in vivo administration of xanthopterin following implantation also resulted in smaller tumors. 4. Xanthopterin exerts differential effects on cell growth dependent upon the cell origin and their state of proliferation.

Persistence of the immunoprotective effects of leukemia × fibroblast hybrid cells toward leukemia in histocompatible mice☆

Hybrids of ASL-1 murine leukemia cells and LM(TK-) cells, a cultured line of mouse fibroblast origin, stimulate partial immunity toward ASL-1 cells in (A/J X C3H/HeJ)F1 mice (F1 mice). Such mice ordinarily exhibit no resistance to the malignant proliferation of ASL-1 cells. Unprotected animals invariably die within 14-18 days after receiving as few as 200 ASL-1 cells. The hybrid cells, the mice used in the experimental studies and the leukemia cells used for challenge all share the same histocompatibility antigens. ASL-1 cells are H-2a; LM(TK-) cells are H-2k, both ASL-1 X LM(TK-) hybrid cells and A/J X C3H/HeJ)F1 mice are H-2a/k. The long-term persistence of the immunoprotective properties of the hybrid cells toward murine leukemia was investigated by using cells that had been in continuous culture for approx. 36 months. (A/J X C3H/HeJ)F1 mice injected previously with hybrid cells in continuous culture and then challenged with up to 10(7) ASL-1 cells survived longer (p less than 0.001) than mice who had not received hybrid cells previously. Some mice challenged with lesser number of ASL-1 cells survived indefinitely (greater than 200 days). The median survival time of F1 mice injected simultaneously with 10(7) hybrid cells and 200 or 2000 ASL-1 cells was significantly (p less than 0.001) prolonged as well, although the differences between experimental and control groups are less pronounced than if the hybrid cells were injected before challenge with ASL-1 cells. The hybrid cells like those freshly prepared continue to be rejected by histocompatible precipients. In no instance has there been evidence of a progressively growing tumor of hybrid cells in immunocompetent F1 mice. Hybrid cells like those investigated previously do form rapidly growing metastasizing tumors in immunodeficient nu/nu (BALB/c) or X-irradiated (550 R) F1 mice. The cells possess approx. 70 chromosomes (reduced from 85) including chromosomes identified as having originated in each parental source. Like (A/J X C3H/HeJ)F1 animals, they continue to express both H-2a and H-2k antigenic determinants.

Differential effects of enalapril and irbesartan in experimental papillary necrosis

This study was undertaken to determine whether angiotensin receptor blockers are as renoprotective as angiotensin-converting enzyme inhibitors in an experimental model of chronic interstitial renal disease. Groups of rats received one of the following treatments for 1 week: (1) enalapril, (2) diltiazem, (3) a cocktail of hydralazine, reserpine, and hydrochlorothiazide, or (4) irbesartan (an AT1 antagonist). The animals were injected with bromoethylamine (200 mg/kg), and antihypertensive treatment continued for 1 month. All drugs were effective in lowering the mean arterial pressure. The bromoethylamine-treated rats developed albuminuria and sustained a 40–50% decrease in creatinine clearance. Enalapril and irbesartan reduced albuminuria, but only enalapril partially prevented the decline in creatinine clearance and lowered the number of ED-1-positive cells. Diltiazem and cocktail had no effect on proteinuria, creatinine clearance, or ED-1 cells. In this experimental model, the effects of enalapril and irbesartan were not identical. Both drugs reduced proteinuria, but enalapril was more effective in protecting the renal function. The fact that the AT1 antagonist protected against albuminuria but did not affect the clearance of creatinine implies that the results seen with angiotensin-converting enzyme inhibition may be in part due to an effect on angiotensin II via AT2 receptor blockade or through an effect on bradykinin.

Circadian Rhythm of Blood Ethanol Clearance Rates in Rats: Response to Reversal of the L/D Regimen and to Continuous Darkness and Continuous Illumination

Phase relationships of the circadian rhythms of blood ethanol clearance (metabolic) rates and body temperature were studied in rats successively exposed to 4 illumination regimens: LD (light from 0800-2000 hr), DL (light from 2000-0800 hr), constant darkness (DD) and, lastly, constant light (LL). After a 4-wk standardization to each regimen, body temperatures were taken at 9 X 4-hr intervals to establish baseline circadian profiles. One week later, groups (N = 8) received 1.5 g/kg ethanol (i.p.) at 6 equally spaced timepoints during a 24-hr span, when temperatures were again measured. Ethanol clearance rates were estimated from decreasing blood ethanol levels sampled every 20 min from 60-200 min after dosing, and the resultant elimination curves were subjected to cosinor analysis. These studies show for the first time that the high amplitude circadian rhythm in ethanol metabolism persists under constant conditions of illumination (DD and LL), demonstrating that it may well be a truly internal circadian rhythm and not a response to exogenous cues of the light/dark cycle. During both LD and DL, maximal and minimal ethanol clearance rates fell near the end of the dark and light phases, respectively, and followed circadian peak and trough control temperatures by approximately 6 hr. A fixed internal phase relationship between the core body temperature and the circadian rhythm in ethanol metabolism is demonstrated, thus establishing the rhythm in body temperature as a suitable and convenient internal marker rhythm for studies of the metabolism of low-to-moderate ethanol doses. These studies demonstrate that the phase relationships of blood ethanol clearance rate and body temperature can be manipulated by the illumination regimen selected, an observation of both basic and practical importance.

Relaxin decreases renal interstitial fibrosis: A novel treatment to slow the progression of kidney disease?

Interstitial fibrosis is a characteristic finding in numerous pathologies. In the kidney this condition is manifested by a disturbance in tubular architecture such that there is tubular atrophy accompanied by tubular dilation. The normally sparse intercellular space is expanded and filled with extracellular matrix composed mainly of collagen and fibronectin as well as mononuclear cells. If left untreated this may progress to encompass the glomeruli and renal insufficiency may result.