Jack E. Gray

Ultrastructural studies of the hepatic changes brought about by Clindamycin and Erythromycin in animals.

Abstract In the hepatocytes of purebred beagles, treated po with 600 mg/kg of clindamycin, whorled figures formed in lysosomes within 24 hr. The greatest abundance (100 or more) and maximum development (2–4 μ) of these pericanalicular dense bodies occurred after 7–10 days of treatment. Such inclusions were shown to be transient and formed rarely or not at all when daily treatments of 300 mg/kg or less were given for as long as 1 yr. After a single po dose of 1000 mg/kg was given to immature male rats of the Upjohn Sprague-Dawley line, whorled figures formed within 30 min in the peribiliary areas of the hepatocytes. With continued treatment, the whorled figures were more frequently multicentric but were not as numerous as in the dog. An apparent increase in autophagic vacuoles and minor changes in the endoplasmic reticulum and mitochondria (at 1500 mg/kg) suggested that the turnover of worn-out organelles may have been accelerated. A single po dose of 1000 mg/kg of erythromycin in the rat produced nonprogressive degenerative changes in a minor portion of hepatic mitochondria in 24 hr. Membrane-bound whorled figures of the type derived from autophagic vacuoles were formed in small numbers. Simultaneous administration of phenobarbital and erythromycin for 10 days resulted in two apparently unrelated ultrastructural responses. Whorled figures were formed in autophagic vacuoles, mitochondria, and secondary lysosomes when erythromycin was given po to beagles at a level of 600 mg/kg for 10–30 days.

The parenteral toxicity of clindamycin 2-phosphate in laboratory animals☆

Abstract The ip and iv LD50 values of 1145 and 855 mg/kg of clindamycin 2-phosphate in the Swiss white mouse were approximately 3 times higher than those of clindamycin hydrochloride. The lesion produced by single injection of 50 or 100 mg/ml of clindamycin 2-phosphate in the loin muscles of the New Zealand White rabbit was graded as slight. The 24-hr serum creatine phosphokinase value was 1500 IU/liter which was less than one-half that of the parent antibiotic. Body weight gains and food conversion ratios in groups of 10 Sprague-Dawley rats injected sc with 120 mg/kg for 6 days were comparable to those of the control group; 90 mg/kg was tolerated in these terms nearly as well for 30 days as no treatment or doses of 30 and 60 mg/kg. From 22 to 33 injections (each equivalent to 30, 60 or 90 mg/kg) were made bilaterally in the posterior thigh muscles of groups of 3 beagle dogs. The terminal elevations of serum glutamic-oxaloacetic transaminase varied from 54 to 400 Reitman-Frankel units. The characteristic pathologic change resulting from the superimposed injections was dose-related progressive scarring of the muscle bundles. Intravenous administration of 60 and 120 mg/kg daily in divided doses in 2 groups of 4 dogs each for 30 days produced no detectable irritation in the peripheral veins or drug-related hemolysis. Tests for drug-induced hemolysis and changes in erythrocyte fragility by in vitro methods were negative. In dogs treated iv with 120 mg/kg for 1 week, as light increase in neutral lipid droplets was present in hepatocytes from 3 hr to 3 days when examined by electron microscopy. This transient change was not observed in dogs injected im with 90 mg/kg for the same period.

The oral toxicity of clindamycin in laboratory animals

Abstract The potoxicities of clindamycin hydrochloride and clindamycin palmitate have been extensively studied in mice, rats and dogs. For the hydrochloride, the ip mouse and po rat LD50 values were 361 and 2618 mg/kg, respectively. The values for the palmitate ester were >2500 and >5000 mg/kg; in a formulated syrup the po LD50 was reduced to 1950 mg/kg. The maximum daily tolerated dose of the hydrochloride in the dog and rat for as long as 1 yr was greater than 300 mg/kg but less than 600 mg/kg. Significant changes in bioclinical data in dogs treated with 300 mg/kg were confined to sporadic elevations in serum transaminase levels. At 600 mg/kg, in short-term studies, the effects of focal irritation were recognized in the mucosa of the stomach and gallbladder of the dog. No teratogenic effect or developmental retardation in the offspring of mouse or rat dams, or effect on the breeding performance of male and female rats was observed. In a comparable series of studies with the palmitate ester, essentially no effects due to the antibiotic were detected. A daily dose of 600 mg/kg was well tolerated in the dog and rat for 6 mo.

Effects of tetracycline on ultrastructure and lipoprotein secretion in the rat hepatocyte

Abstract Fat droplets, 0.1–0.5 μm, accumulated in the space of Disse of hepatocytes in rats treated ip with 300 mg/kg of tetracycline hydrochloride. This change occurred in both pregnant and unbred females and in males but was not observed in similar rats treated with 100 mg/kg which showed typical steatosis induced by tetracycline. Ultrastructural changes in mitochondria, rough and smooth endoplasmic reticulum (SER) were similar to those previously reported. Uptake of iv administered palmitic [9-10-3H]acid in rats treated with 300 mg/kg was not appreciably impaired. One hour after an injection of 300 mg/kg, the antibiotic concentration was 10 times greater in the liver than in the serum. From 1 to 6 hr the concentrations increased from roughly 300 to 800 μg/g of liver. In rats treated with 100 mg/kg, no concentrations above 200 μg/g were observed during a 48-hr postinjection period. Under in vitro conditions, tetracycline complexed with lipoprotein in a precipitate when a concentration greater than 200 μg/ml was obtained in rat blood serum. In the hepatocyte of rats treated with 300 mg/kg it was presumed that as the antibiotic concentration rose above 200 μg/g nascent very low density lipoprotein complexed rapidly with it. The complexed lipoprotein was channeled in dilated SER in the manner of liposomes to the sinusoidal surface and discharged from vesicles into the space of Disse. The plasma membrane of the hepatocyte showed loss of microvilli and surface distortion. The space of Disse became distended with insoluble fat droplets from 6 hr to 3 days after injection.

Appraisal of the Intramuscular Irritation Test in the Rabbit

Musculoirritancy testing in the rabbit has been recognized as sufficiently sensitive to be highly predictive of human tolerance for intramuscularly administered drugs. A three-part study consisting of gross characterization of the lesion, microscopic observation and determination of serum activity of the enzyme creatine phosphokinase (CPK) has evolved as a standard test for evaluating single injection injury in the muscle. Injection and tissue processing procedures are described as well as a revised grading system for gross examination based on degrees of necrosis and hemorrhage, respectively. Based on 213 determinations, the control mean serum CPK activity was 262 +/- 113 I.U./liter. The importance of pretest acclimation of rabbits in establishing this value was emphasized. Designations of slight, moderate, or marked irritation are based on accumulative scoring of all rabbits at postinjection intervals of 1, 3, and 7 days. At 24 hr postinjection, serum CPK activities up to and including a range of 2000-3000 I.U./liter have been shown to be compatible with slight irritation and are considered predictive of human tolerance.

Foam cell response in the lung and lymphatic tissues during long-term high-level treatment with erythromvcin

Abstract Collections of foam cells in the lungs and lymphatic tissues have not been previously recognized in dogs and rats treated with high oral doses (500–1000 mg/kg) of erythromycin. With extended periods of treatment with either the octadecylsulfamate derivative or the antibiotic base, slow progressive transformation of macrophages became apparent. Foam cells in the lung showed a repetitive pattern of small whorls in vacuoles similar to that previously recognized with other drugs that induce phospholipidosis. The foam cells in the lymphatic tissues of the dog were confined principally to the germinal centers, whereas those in the rat lymph nodes were more diffusely scattered and were scored with cholesterol clefts. The cholesterol content of the mesenteric lymph nodes of treated rats was significantly increased. Residual antibiotic activity was assayed in the lungs and mesenteric lymph nodes of rats 5 days after treatment. The mobilization of macrophages in the lymphatic tissues appeared to be related secondarily to the low-grade formation of antibiotic-induced myeloid bodies in lymphoid cells.

Teratogenicity and neonatal toxicity of clindamycin 2-phosphate in laboratory animals

Abstract Subcutaneous injections of clindamycin 2-phosphate at 100 and 180 mg/kg on gestation days 6 through 15 in Upjohn Sprague-Dawley rats, Upjohn ICR and CF1 mice had no detremental effect on reproductive performance. Teratogenic effects were not observed. A concurrent low incidence of spontaneous cleft palate was confirmed in the Upjohn ICR strain of mouse. This condition was not eliminated by vitamin B supplementation. The sc LD50 of clindamycin-2-phosphate has been shown to be 179 mg/kg in the neonatal rat and > 2000 mg/kg in the adult rat. The newborn/adult toxicity ratio was calculated at 1:11.2, which is comparable to potassium penicillin G. Subcutaneous injections of clindamycin-2-phosphate at 120 mg/kg in nearly mature and recently weaned Gunn rats did not alter the partially impaired bilirubin metabolism in this neonate model. Simultaneous treatment of immature Gunn rats with 120 mg/kg of clindamycin-2-phosphate and therapeutic levels of sulfadiazine and sulfamerazine did not produce sufficient displacement of bilirubin from albumin binding in the serum to produce clinical signs of kernicterus.

Comparative effect of acetylsalicylic acid and acetylsalicylic acid anhydride on the non-glandular portion of the rat stomach

Abstract A method has been developed for the evaluation of gastric irritation induced by acetylsalicylic acid in the rat. Acute pustular inflammation of the nonglandular portion of the rat stomach resulted from single oral sublethal dosage of acetylsalicylic acid; no significant involvement of the fundic mucosa was observed. Acetylsalicylic acid anhydride was determined to be appreciably less irritating than acetylsalicylic acid upon the basis of morphologic evidence obtained at comparable dosages in 182 rats.

Comparative Responses of Vaginal Mucosa to Chronic Pyrimidinone-induced Irritation*

Intravaginal studies in guinea pigs, Cebus monkeys (Cebus apella), and beagles were made with an anti-herpes drug, 2-amino-5-bromo-6-phenyl-4(3H)-pyrimidinone (ABPP). Concentrations of 5 to 40 mg/ml, suspended in saline or polyethylene glycol, were given for periods of 2 1/2 to 90 days. Very little gross evidence of injury was observed. Histologically, the regimens brought about increasing degrees of mononuclear cell infiltrations (histiocytes, lymphocytes, and plasma cells) in the vaginal lamina propria of the respective species. In the Cebus monkey and beagle the nonkeratinized vaginal lining became progressively thinned. The integrity of focal denuded areas was preserved by a process of homogenization in the lamina propria of the Cebus monkey. In contrast, the most severely affected areas in the canine mucosa were covered by what appeared to be a coagulated exudative layer. The effect of 90 days of treatment in the beagle did not noticeably increase the severity of the findings beyond that observed after 30 days. Considerable reversibility of the lesions occurred in guinea pigs held for 1 week and in Cebus monkeys held for 3 weeks posttreatment. Local cell-mediated immunity was believed to be implicit with the mononuclear cell infiltration in the lamina propria of the vagina.

Hepatic changes produced by 30-day administration of a novel aminocyclitol antibiotic, trospectomycin sulfate, to laboratory animals

The studies described here were done to characterize the hepatic response to a new aminocyclitol antibiotic, trospectomycin sulfate, administered intravenously (beagle dog) or subcutaneously (Sprague-Dawley rat) at a variety of dose levels, to investigate reversibility of observed changes, and to document any untoward effects of subchronic trospectomycin sulfate administration. Both species showed significant elevations in serum levels of alanine and aspartate transaminases in higher dose groups. In the dog only, a transient neuromuscular blockade was also observed within higher dose groups. No other functional, morphological, or serum chemical changes were observed. Examination of liver by electron microscopy revealed the presence of cytoplasmic lamellar inclusion bodies, concentrated in the bile canalicular region of the hepatocytes. Occurrence of the lamellar bodies and coincident transaminase increases were found to be reversible upon discontinuance of treatment (studied in the dog). Electron microscopy of acid phosphatase cytochemistry in the rat indicated that most, but not all, of the lamellar bodies contained this enzyme. This observation suggests that they may be derived from the lysosome, or once formed become lysosomal.