Nancy L. Miller

Characterization of heterogeneity in the molecular pathogenesis of lupus nephritis from transcriptional profiles of laser-captured glomeruli

The molecular pathogenesis of focal/diffuse proliferative lupus glomerulonephritis was studied by cDNA microarray analysis of gene expression in glomeruli from clinical biopsies. Transcriptional phenotyping of glomeruli isolated by laser-capture microscopy revealed considerable kidney-to-kidney heterogeneity in increased transcript expression, resulting in four main gene clusters that identified the presence of B cells, several myelomonocytic lineages, fibroblast and epithelial cell proliferation, matrix alterations, and expression of type I IFN-inducible genes. Glomerulus-to-glomerulus variation within a kidney was less marked. The myeloid lineage transcripts, characteristic of those found in isolated activated macrophages and myeloid dendritic cells, were widely distributed in all biopsy samples. One major subgroup of the samples expressed fibrosis-related genes that correlated with pathological evidence of glomerulosclerosis; however, decreased expression of TGF-beta1 argued against its role in lupus renal fibrosis. Expression of type I IFN-inducible transcripts by a second subset of samples was associated with reduced expression of fibrosis-related genes and milder pathological features. This pattern of gene expression resembled that exhibited by activated NK cells. A large gene cluster with decreased expression found in all samples included ion channels and transcription factors, indicating a loss-of-function response to the glomerular injury.

Hepatic aluminum accumulation in children on total parenteral nutrition

Five children receiving long-term total parenteral nutrition (TPN) containing casein hydrolysate as the protein source underwent percutaneous liver biopsies because of the development of cholestasis and abnormal liver function tests. All five demonstrated moderate to severe histopathologic changes. In addition, hepatic aluminum content was determined to be markedly elevated in all cases. Although the hepatotoxicity of aluminum is as yet undetermined, deposition of other metals has been associated with liver damage, and aluminum has been associated with pathology in other tissues. Thus, the possibility that aluminum deposition may play a role in the pathogenesis or exacerbate the course of liver dysfunction associated with TPN should be considered.

Aluminum-associated hepatobiliary dysfunction in rats: relationships to dosage and duration of exposure.

ABSTRACT: Aluminum may contaminate parenteral nutrition solutions and accumulate in bone and liver of patients receiving this therapy. Although aluminum exposure is associated with low-turnover osteomalacia, there are few studies of hepatotoxicity. We therefore studied the effects of aluminum given to rats on total serum bile acid concentration and bile flow to determine if aluminum administration could produce abnormalities. Aluminum was given intravenously as follows: 5 mg/kg daily for 7 or 14 days and 1 mg/kg for 14 days. Hepatic aluminum was high in treated rats and undetectable in controls. Total serum bile acid concentrations were significantly higher in treated rats than in pair-fed controls with higher concentrations after 14 days than after 7 days. Bile flow was reduced by 33% in rats given 5 mg/kg but not in rats given 1 mg/kg. Hepatic aluminum correlated inversely with bile flow but not with serum bile acid concentration. Aluminum exposure in rats is associated with elevated serum bile acid concentration and diminished bile flow and may play a role in the pathogenesis of parenteral nutrition-induced hepatobiliary dysfunction.

Bone Magnesium Pools in Uremia

Bone magnesium pools were studied in vitro in bone specimens obtained from control subjects, from patients with chronic renal failure before and after renal transplantation, and in a patient with chronic hypomagnesemia. 30% of bone magnesium is in a surface limited pool present either within the hydration shell or else on the crystal surface. The larger fraction of bone magnesium was shown not to be associated with bone matrix but rather to be an integral part of the bone crystal. With incineration this pool was mobilized at the same temperature that sudden enlargement of bone crystal size occurred. It is suggested that heating causes surface calcium to displace magnesium from the apatite crystal. Both magnesium pools are increased in patients with chronic renal failure. The major factor determining magnesium concentration in bone would appear to be the serum magnesium level. Following renal transplantation, in association with the fall in serum magnesium, surface magnesium was within the normal range; whereas, residual magnesium was not different from the other urenic bones. Both magnesium pools were significantly reduced in a patient with chronic hypomagnesemia. The in vitro studies would suggest that surface magnesium should rapidly reflect changes in serum magnesium levels, whereas, the deeper magnesium pool is probably deposited at time of bone formation with mobilization being dependent upon the resorptive processes. Since magnesium can influence crystal size and stability it seems possible that excess bone magnesium may play a role in renal osteodystrophy.

Effect of Age and Magnesium Depletion on Bone Magnesium Pools in Rats

In vivo and in vitro studies were carried out to characterize the exchangeable bone magnesium pool and determine what effect age and magnesium depletion has on bone magnesium. A highly significant correlation was found between the size of the in vitro elutable and in vivo exchangeable bone magnesium (r=0.97). To show that the exchangeable bone magnesium was the surface-limited bone magnesium, elution studies were performed 4 h after the in vivo administration of radiomagnesium. Specific activity in the eluant was 85% of that found in the serum at time of death, suggesting that the elutable and exchangeable bone magnesium pools were largely the same pool. Bone magnesium concentration fell with increasing age. The entire fall in bone magnesium was a result of a decrease in the surface-limited fraction. Since bone crystals have been shown to enlarge with aging with resulting contraction of the surface area, this would be the most apparent explanation for this finding. During magnesium depletion, magnesium concentration in both the exchangeable and nonexchangeable pools decreased. The fractional change in the exchangeable pool was much larger than the change in total or nonexchangeable bone magnesium, suggesting that the surface-limited magnesium pool is available during magnesium depletion. The change in size of the nonexchangeable bone magnesium pool appeared to be more related to the duration of magnesium depletion than the change in serum magnesium levels. The fall in magnesium concentration in this pool is probably a consequence of continuing formation of low magnesium bone during the depletion period.

Hepatic Abnormalities Associated with Aluminum Loading in Piglets

Cholestasis is a common complication of total parenteral nutrition (TPN) in infants. A contributing factor to the hepatic dysfunction may be a contaminant of the TPN solution, such as aluminum, that accumulates in liver and may act as a hepatotoxin. To study the hepatic effects of aluminum, growing piglets were given daily intravenous injections of aluminum, 1.5 mg/kg, for 50 days; pair-fed controls were given heparinized saline. At sacrifice, liver and serum were obtained. Liver was analyzed for histopathology and for aluminum content and localization. The hepatocyte lysosomes of the experimental group showed aluminum peaks by x-ray microanalysis, whereas the control group did not. No differences in ultrastructure were noted between the two groups when examined by electron microscopy. Mean serum total bile acid levels (27.8 +/- 15.9 SD vs 6.3 +/- 1.5 mumol/liter, p less than 0.05), mean alkaline phosphatase (309 +/- 108 vs 180 +/- 27 IU/liter, p = NS), and mean hepatic copper content (24.8 +/- 4.5 vs 14.4 +/- micrograms/g dry weight, p less than 0.01), were elevated in the aluminum-loaded piglets, indicating that cholestasis may have been produced. Also, a small but significant reduction in serum levels of 25 hydroxy-vitamin D was found in the aluminum-loaded piglets, suggesting that vitamin D hydroxylation may be impaired. Inasmuch as lysosomal contents are excreted into the bile, aluminum accumulation in lysosomes may alter lysosomal function and possibly affect bile flow or content.

Urinary iron speciation in nephrotic syndrome

In nephrotic syndrome, iron is presented to the tubule fluid in a nonreactive form in association with transferrin as a result of the glomerular protein leak. At an alkaline pH, iron remains bound to transferrin throughout the nephron and is excreted as such in the urine. As urine pH decreases below 6, iron is dissociated from transferrin. In the dissociated form, iron exists in the urine in a soluble, ultrafiltrable, and labile state. It is suggested that iron is maintained in this state by chelation to a relatively small organic compound, such as citrate. This non-transferrin-bound iron is capable of catalyzing bleomycin degradation of DNA, suggesting that this labile form of iron is able to catalyze free radical formation and cause tubule cell injury. Urine from proteinuric states represents one of the few, if not only, biologic fluids containing large amounts of reactive iron species. This may explain the mechanism by which proteinuric states cause tubulointerstitial disease and renal failure.

Effects of aluminum on the liver following high-dose enteral administration to rats.

Parenteral administration of aluminum (Al) to animals can result in hepatobiliary dysfunction, including elevated total serum bile acid concentration, reduced bile flow, and reduction of mixed function oxidase activities. Despite substantial hepatic Al accumulation, biliary Al excretion is negligible. We studied the effects of enteral administration of pharmacologic doses of Al to rats in order to see if by this route Al also produced hepatobiliary dysfunction or if biliary Al excretion was enhanced following enteral administration, protecting the liver from the effects of Al. Six rats were given 100 mg/kg/day of Al for 14 days as Al citrate by duodenal cannula. Pair-fed littermate controls were given sodium citrate. Serum Al and urinary Al/creatinine were significantly higher in Al-fed rats than in controls. Liver Al was significantly increased in the Al-fed group, but very low when compared to liver Al concentration with intravenous Al administration. Biliary Al was only 2 +/- 1% of urinary Al in the experimental group. Serum bile acid concentration and bile flow were not different between groups. We conclude that Al given in pharmacologic doses is absorbed but does not accumulate in the liver. We hypothesize that a slow rate of Al absorption may not overwhelm plasma transferrin carrying capacity or renal Al excretory capacity.

The Role of Aluminum in the Pathogenesis of Anemia in an Outpatient Hemodialysis Population

Anemia is a well-defined complication of aluminum overload in chronic dialysis patients which may be present before other manifestations of aluminum toxicity are obvious. Causes of anemia in chronic renal failure are multiple, and at the present time there is no marker for aluminum-induced anemia. Deferoxamine (DFO) treatment can correct aluminum-related anemia and microcytosis, but may be associated with side effects. Because of the possible role of aluminum in red blood cells in causing the anemia associated with aluminum overload, we attempted to test red blood cell (RBC) aluminum as a marker for aluminum-associated anemia and to assess the prevalence of aluminum-associated anemia in an outpatient dialysis population. Both random plasma aluminum and RBC aluminum correlated well with the increase in plasma aluminum seen following DFO challenge. However, RBC aluminum was affected less by changes in oral aluminum intake than plasma aluminum. There were strong correlations of RBC and plasma aluminum to corpuscular volume (MCV) in our patients. Moreover, patients within the highest quartile of RBC aluminum had a lower mean MCV (82.1 +/- 1.7 vs 89.6 +/- 1.7, p less than .01) and hematocrit (HCT) (24.3 +/- 4 vs 28.2 +/- 1.5, p less than .05) than those within the lowest quartile. These data suggest that aluminum toxicity is an important cause of microcytic anemia in outpatient hemodialysis patients. Prospective long-term studies are needed to further define the usefulness of RBC aluminum in diagnosing and following hemodialysis patients with aluminum-induced anemia.

THE EFFECTS OF 1,25(OH) 2 D 3 ON TISSUE AND BONE METABOLISM IN THE UREMIC RAT MODEL

Therapy with 1,25(OH)2D3 to prevent osteodystrophy in renal failure patients can cause hypercalcemia, soft tissue calcification, and decreased creatinine clearance. To test whether low dose therapy with 1,25(OH)2D3 could prevent decreased mineralization in uremia without concomitant hypercalcemia, we studied 2 groups (A & B) of 13 rats with remnant kidney and 8 non-remnant controls (C). Groups A & B received 0 μg and .01 μg/Kg/day of 1,25(OH)2D3 respectively. Results: Weekly serum calcium (Ca), phosphorus, creatinine, body weight (wt) were not significantly different between Groups A & B. Analysis after sacrifice at 16 weeks (mean ± 1 SD) showed:Creatinine correlated inversely with % ash (r=0.77), and femur wt correlated with body wt (r=0.95) in A but not in B. Conclusion: 1,25(OH)2D3 prevented decreased bone mineralization associated with uremia without a significant increase of hypercalcemia, soft tissue calcification, or serum creatinine.