Lead poisoning: rapid formation of intranuclear inclusions.

A single dose of lead (0.05 milligram per gram of body weight) induced characteristic intranuclear inclusions in the epithelium of proximal tubules in rat kidney within 1 to 6 days. The development of the intranuclear inclusions is thus an acute manifestation of lead poisoning, not a delayed one, as has been thought hitherto. Cytoplasmic structures resembling the intranuclear inclusions and situated in the vicinity of endoplasmic reticulum were regularly found in cells bearing the pathognomonic intranuclear inclusions. The latter and the cytoplasmic structures may be derived from a common precursor, perhaps a soluble protein-lead complex.

Non-histone chromosomal proteins of chemically transformed neoplastic cells in tissue culture.

Chromatin proteins from control and dimethylnitrosamine-transformed baby hamster kidney cells were comparbd by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Our results indicate that non-histone chromosomal proteins from transformed cells contained protein components of low and intermediate electrophoretic mobility, which were deficient in normal cells. Comparison of the relative amount of incorporation of labeled amino acids into non-histone chromosomal proteins showed that protein components with a molecular weight of about 60,000 M.W. had a markedly increased labeling activity in the chemically transformed cells. These results suggest that changes in non-histone chromosomal proteins are associated with neoplastic transformation by chemical carcinogens.

Long-term association of N-(phosphonacetyl)-L-aspartate with bone.

By means of enzymatic and autoradiographic techniques, it has been demonstrated that, 24 hr after a single dose of the antitumor amino acid N-phosphonacetyl-L-aspartic acid (PALA), (400 mg/kg i.p.; 1.15 mmol/kg) to C57BL x DBA/2 F1 mice, the agent accumulates in bone to a concentration of approximately 400 microM; this is 3000 times greater than the Ki of PALA for its target enzyme, aspartate carbamoyltransferase. However, disproportionately low inhibition of enzyme activity was demonstrated in homogenates of bone from these recipients, suggesting that the drug was sequestered from its target in this tissue. Autoradiography of sections of femoral shafts from mice treated with 14C-labeled drug demonstrated that autoradiogram density due to [14C]PALA equivalents was confined to the bony matrix, with no label above background resolvable in bone marrow. Following in vivo administration of PALA (400 mg/kg i.p.), the half-life of the drug in the bone was approximately 23 days. In vitro, with equilibrium dialysis at pH 7.4, it was demonstrated that: (a) normal pulverized and decalcified bone bound PALA with capacities of 3.5 nmol/mg and 0.1 nmol/mg bone, respectively, at a PALA concentration of 5 mM; (b) binding of PALA to normal bone reached saturation at a concentration of 200 mM; and (c) PALA functions as a solubilizer of bone at concentrations above this. Since administration of PALA was shown to produce long-lasting inhibition of aspartate carbamoyltransferase in liver and tumor and since its ultimate half-life in the plasma of mice, following a single 400-mg/kg administration of the drug, is 8 days, it is suggested that bone serves as a reservoir from which PALA is released at a slow rate into plasma and other tissues.

Inhibition of capping of immunoglobulin and concanavalin A receptors by cis-dichlorodiammine-platinum-(II) in mouse spleen cells.

Effects of cis-dichlorodiammineplatinum-(II) (cis-DDP), an anticancer agent, on capping of membrane receptors were investigated in mouse spleen cells in vitro, using fluorescein labeled ligands. Cap formations of surface immunoglobulin and of concanavalin A receptors were significantly reduced by cis-DDP in relation to dose, while cell viability remained unchanged. The binding of the ligands to specific receptors was not affected by cis-DDP. The results suggested that cis-DDP inhibited cap formation by blocking membrane movement of the ligand-receptor complexes.

Mitigation of intestinal cytotoxicity of cisplatin by EDTA in rats.

Cisplatin (cis-dichlorodiammineplatinum-II), an antitumor agent containing platinum, produced acute necrotizing enteritis in rats. This toxic side effect was significantly reduced by oral treatment with CaNa2EDTA. The protective effect of CaNa2EDTA against intestinal cytotoxicity of cisplatin was dose-related. Based on histopathological evaluation of cell necrosis and mitotic index, it was estimated that pretreatment with CaNa2EDTA in drinking water (75 mM) and by gavage (1.5 g/kg) prevented intestinal cytotoxicity by up to 90% compared with cisplatin controls.

G2 sub-population in mouse liver induced into mitosis by lead acetate.

A single intracardiac dose of lead acetate (40 microgram lead/g body weight) induced a 25-fold increase in mitosis of mouse hepatocytes 5 hr after injection, as determined by autoradiography. The prompt appearance of a mitotic wave and the relatively large number of mitoses suggest that the mitotic cells were derived from a hepatocyte sub-population arrested in the G2 phase. The injection of lead also stimulated a small increase in labeled hepatocytes within 6 hr. Analysis of grain counts gave no evidence for unscheduled DNA synthesis. The incremental labeled cells may have originated from a small fraction of the G1 population that was ready to enter the S phase without the usual pre-synthetic delay.

Cell proliferation in rat kidneys after prolonged treatment with lead.

Effects of chronic lead intoxication on cellular proliferation in rat kidneys were investigated by autoradiography. The rats were given intraperitoneal injections of lead acetate in aqueous solution for 6 months. At the end of this period, the proliferative activity of proximal tubular epithelial cells was about 15 times greater in rats given lead than in untreated controls. In the leaded rats approximately 40% of the proximal tubular cells contained intranuclear inclusions, approximately 0.5% of proximal tubular epithelial cells were labeled and approximately 6% of labeled cells contained intranuclear inclusions. Thus, cells with intranuclear inclusions can replicate DNA. Effects of chronic lead poisoning on the replication of proximal tubular cells in rats subjected to left uninephretomy before inception of treatment with lead were substantially the same. No renal carcinomas were found after 6 months of treatment with lead, but there was epithelial hyperplasia in some proximal tubules, with occasional atypia. The presence of increased synthesis of DNA and epithelial hyperplasia in the kidneys of rats chronically poisoned with lead suggests that the renal carcinogenicity of lead, observed by others, is related to lead-induced stimulation of renal cell proliferation.

Cell proliferation in rat kidney induced by lead acetate and effects of uninephrectomy on the proliferation.

Effects of a single dose of lead (0.04 mg lead g body weight) on the proliferation of proximal tubular epithelium in rat kidneys were investigated by autoradiography over a period of 72 hours, using (3)H-thymidine as a label. The results demonstrate that cell proliferation was greatly stimulated within 2 days after lead was injected. The increase in DNA synthesis began about 20 hours after intraperitoneal injection of lead, reached a sharp peak at 30 hours, and declined rapidly thereafter. At the peak, the mean labeling activity was 40 times that observed in control rats. Cumulatively, an average of 14.5% of the proximal tubular epithelial cells were labeled 72 hours after lead was injected. When uninephrectomy was followed immediately by injection of lead, the stimulation of DNA synthesis in the remaining kidney was, on the average, greater than the sum of the separate effects of the two treatments. This indicates that the stimulatory effects of uninephrectomy and injection of lead on renal cell proliferation were additive.

Biogenesis of intranuclear lead-protein inclusions in mouse kidney.

A single dose of lead (5 mug/g body weight), given as lead acetate by intracardiac injection, produces, within 8 hours, characteristic fibrillar intranuclear and intracytoplasmic inclusions in proximal tubular epithelial cells in mouse kidneys; after a dose of 40 mug/g body weight, such inclusion appeared within 6 hours. Their development was completely prevented by cycloheximide (one or more intraperitoneal injections of 20 mug/g body weight). The development of intranuclear inclusions was also noted in tubular epithelial cells explanted from normal mice and grown in vitro for 15 hours in a medium containing 20 mug lead/ml. Thus, the development of the characteristic fibrillar inclusion bodies depends upon de novo synthesis of inclusion body protein, induced by lead.

Acute and chronic cisplatin nephropathy in rats.

Cisplatin (cis-dichlorodiammineplatinum-II) is a new class of platinum coordination compounds showing a potent antitumor activity, but it also produces adverse effects on renal function. In this study, cisplatin nephropathy and renal accumulation of platinum were analyzed in rats after acute chronic treatment. A single intraperitoneal dose of cisplatin (6 mg. per kg.) induced marked focal necrosis in the proximal and distal tubules with a maximal lesion on day 7. The tubular damage was localized mainly in the corticomedullary region, where the concentration of platinum was the highest within the kidney. Repeated treatment with cisplatin (1 mg. per kg., intraperitoneally, twice weekly) for 11 weeks resulted in massive tubular basement membranes. Some glomeruli appeared fibrotic, indicating that chronic treatment with cisplatin could cause irreversible renal damage. The results indicated that the toxicity of platinum was probably a major factor for cisplatin nephropathy.