Morphological changes in the kidney of dogs chronically exposed to exogenous growth hormone.

Porcine growth hormone was administered subcutaneously to beagle dogs at doses of 0.025, 0.1, and 1 IU/kg/d for 14 weeks, markedly elevating serum growth hormone (GH) and insulin-like growth factor-1 (IGF-1) levels. This was accompanied by a significant increase in body weight gain and kidney weights in both male and female dogs. The increase in kidney weight (6 to 54%) was slightly greater than the increase in body weight (6 to 40%). By light microscopy, glomerular deposits, mesangial thickening, and very slight cellular infiltration in glomeruli were seen in mid- and high-dose groups. Based on morphometric evaluation, there was an increase in the renal glomerular area, which was statistically significant (p < or = 0.05) in the mid- and high-dose males and in the high-dose females. This was associated with a statistically significant (p < or = 0.05) increase in the number of total glomerular cells in the mid- and high-dose males. By transmission electron microscopy, thickening of the glomerular basal lamina and diffuse increase of the mesangial matrix were observed in both male and female dogs in the mid- and high-dose groups. Immunohistochemical reactions were negative for IgG, IgM, and C3. The morphological changes in the kidney of dogs resemble the diffuse glomerulosclerosis described in human diabetic nephropathy.

Morphological changes in the pituitary gland of dogs chronically exposed to exogenous growth hormone.

Growth hormone (GH) synthesis and release from the pituitary is regulated by hypothalamic releasing hormone, insulin-like growth factor-1 (IGF-1), and somatostatin. However, the potential effects of pharmacological doses of exogenous GH on the pituitary are not well studied. To determine the potential chronic effects of exogenous GH on pituitary morphology in dogs, porcine GH (pGH) was administered subcutaneously to 3 groups of dogs (4 animals/sex/group) at doses of 0.025, 0.1, and 1.0 IU/kg/day for 14 wk. A group (4/sex) of dogs served as the vehicle control. The pituitaries from all dogs were weighed and fixed in appropriate fixatives for light and electron microscopic examination; in addition, cells of the pars distalis were quantitated by a point counting method following immunostaining to identify cells containing GH, prolactin (PRL), and adrenocorticotrophic (ACTH) hormones. Administration of pGH resulted in a statistically significant (p < or = 0.05) increased pituitary weight through the high dose. By light microscopy (LM), hypertrophy of pars distalis cells was evident in mid- and high-dose female dogs. The pituitaries of dogs given the lowest dose (0.025 IU/kg/day) of pGH were not remarkable based on weight and LM findings. In addition, transmission electron microscopic (TEM) examination of the pituitary gland of high-dose demonstrated, in both sexes, pituitary cells with variably dilated rough endoplasmic reticulum and decreased numbers of secretory granules; some of these cells reacted positively to GH immunostaining. Quantitative analysis of the pituitary gland of high-dose males and females showed an increase in the absolute volume of all cell populations studied: GH-, PRL-, and ACTH-positive cells. Based on the LM and TEM findings, the increased volume of the cell populations studied is likely related to cellular hypertrophy. The expected elevation in serum GH levels following repeated administration of pGH and an associated elevation in serum IGF-1 levels resulted in morphologic changes in the pituitary gland of dogs given high doses (> or = 0.1 IU/kg/day) of pGH; these observations differed from the reported findings in pituitaries of transgenic mice secreting large quantities of bovine GH.

Effect of chronic growth hormone administration on skeletal muscle in dogs.

Administration of growth hormone (GH) results in increased body weight gain in dogs. Increased body weight gain is believed to be a result of the trophic effect of GH on the musculoskeletal system. However, edema is one of the side effects described in man following exogenous GH administration. Thus, the objective of this study was to determine if the expected increased weight gain in GH-treated dogs is a result of increased muscle mass. Porcine growth hormone (pGH), administered subcutaneously to beagle dogs at doses of 0.025, 0.1, and 1 IU/kg/day for 14 wk, resulted in elevated serum GH and insulin-like growth factor-1 (IGF-1) levels (see accompanying paper, Prahalada et al). This was associated with a significant increase in body weight gain and weights of the cranial tibialis muscle in both male and female dogs. The increased muscle mass likely contributed to the significant increase in body weight gain seen in both sexes. Quantitative analysis of skeletal muscle sections stained for ATPase activity showed increases in type I (slow twitch) and type II (fast twitch) myofiber sizes in mid- and high-dose males and in high-dose females. The ratio of type I and type II muscle fibers remained unchanged. Hypertrophic myofibers were enlarged but had a normal histologic and ultrastructural organization when observed by light and transmission electron microscopy. The results of this study have demonstrated that increased muscle mass in pGH-treated dogs is related to hypertrophy of muscle fibers and not due to edema. Exogenous GH administration has an anabolic effect on skeletal muscle in dogs.

Effect of early body weight and moderate dietary restriction on the survival of the Sprague-Dawley rat.

The effects of ad libitum (AL) feeding, moderate dietary restriction (DR), and initial (6-week) and one-year body weights on the two-year survival of the Sprague-Dawley (SD) rat were evaluated. DR-fed rats were given approximately 75 percent of the adult AL food intake. At two years, body weights of DR-fed males and females were approximately 69 and 58 percent of the AL-fed male and female body weights, respectively. The 2-year survival rate was 80 and 74 percent in DR-fed males and females, respectively, and 28 and 38 percent in AL-fed males and females, respectively. This increase in longevity indicates that DR-fed males and females in carcinogenicity studies would have 14.8 and 9.1 additional weeks of exposure in a 2-year period to test compounds, respectively, compared to AL-fed animals. There was no correlation between initial body weight and 2-year survival in DR or AL-fed rats. There was no association between 1-year body weight and 2-year survival among DR-fed rats. However, AL-fed rats with the greatest 1-year body weight had a lower 2-year average survival compared with the lightest AL-fed rats; this trend was statistically significant only in males. Body weights between the first and second years were statistically significantly correlated for both genders and feeding regimens but no correlation was observed between pretest and 2-year body weights. These findings demonstrate that initial body weight is not the determining factor of 2-year survival, but that the total adult food (caloric) intake is important. In conclusion, moderate dietary restriction prevented excessive body weight gain and greatly increased the 2-year survival of the SD rat. Initial body weights did not correlate to 2-year body weight gain and were not a predictive biomarker of 2-year SD rat survival.

Lipidosis induced in the dog gallbladder by a direct 5-lipoxygenase inhibitor.

The direct 5-lipoxygenase inhibitor L-739,010 was administered at a dose of 60 mg/kg/day per os to beagle dogs for 15 days. Histopathological examination of gallbladders from treated dogs showed epithelial vacuolation and submucosal infiltration by foamy macrophages that were positive for lipids in Sudan Black-and/or Oil Red O-stained sections. Scanning electron microscopic examination of gallbladder mucosa showed thickening of epithelial folds and multifocal epithelial membrane disruptions. Transmission electron microscopy confirmed these findings and showed mucosal epithelial cell lipid droplet accumulation and submucosal infiltration of macrophages filled with lipid droplets, myelin figures, heterophagosomes, and cholesterol clefts. These changes resembled those reportedly seen in the human gallbladder with cholesterolosis and/or chronic cholecystitis.

Quantitative evaluation of glandular and stromal compartments in hyperplastic dog prostates: effect of 5-alpha reductase inhibitors.

The objective of this study was to determine the effects of 2 different 5-alpha reductase inhibitors (finasteride and MK-0434) on the glandular and stromal compartments of hyperplastic canine prostates. In this study, dogs received 1 of the 2 compounds orally, at a dose of 1 mg/kg/day for 16 weeks; control dogs received a placebo. The morphological changes in the glandular and stromal compartments in the prostate were quantitated by a point-counting method on Masson's trichrome-stained sections. Treatment with 5-alpha reductase inhibitors resulted in significant (P < or = 0.05) decreases in mean prostatic volumes, microscopic evidence of prostatic atrophy, and significant (P < or = 0.05) decreases in the absolute volumes of the prostatic glandular and stromal compartments compared to controls. In finasteride-treated dogs, the mean percent change from baseline was: epithelium, -52; lumens, -58; fibrovascular stroma, -41; and smooth muscle, -29. In MK-0434-treated dogs, the mean percent change from baseline was: epithelium, -77; lumens, -58; fibrovascular stroma, -38; and smooth muscle, -42. The effect on the glandular compartment in dogs treated with MK-0434 was slightly greater than in dogs treated with finasteride; however, the effect on the stroma was similar. These results clearly demonstrate that inhibition of 5-alpha reductase enzyme activity affects growth and maintenance of both glandular and stromal compartments of dog hyperplastic prostates. It is likely that the decrease in size of the prostate in finasteride-treated (Proscar) men is due to shrinkage of both glandular and stromal compartments.

Effects of chronic oral administration of a selective 5 alpha-reductase inhibitor, finasteride, on the dog prostate.

Young mature dogs received finasteride, a selective 5 alpha-reductase inhibitor, orally at 0, 5, 15, and 45 mg/kg/day for 27 or 53 weeks. The effect of finasteride administration on prostatic size and morphology was evaluated macroscopically and microscopically. Changes in glandular and fibromuscular compartments were quantitated by a point counting method on trichrome-stained sections. Finasteride administration induced a decrease of mean prostatic weights and epithelial atrophy in all treated groups. No changes in testicular weights and morphology were observed. The greatest prostatic shrinkage was obtained in the group receiving 45 mg/kg/day for 53 weeks; compared to placebo controls, the percent decreases in absolute volumes occupied by epithelium, lumens, fibrovascular stroma, and smooth muscle were 88, 97, 51 and 72, respectively. These results clearly demonstrate that prostatic shrinkage following finasteride administration results from a decrease in both glandular and fibromuscular compartments.

Rat urinary bladder hyperplasia induced by oral administration of carbonic anhydrase inhibitors.

The carbonic anhydrase inhibitors, acetazolamide and MK-0927, were given by oral route to male Sprague-Dawley rats at 200 mg/kg/day and 25 mg/kg/day, respectively, for up to 4 weeks. Sequential necropsies were performed and urinary bladders were examined by light microscopy (LM), scanning electron microscopy (SEM), and transmission electron microscopy (TEM). Similar urinary bladder changes were seen with both compounds. SEM evidenced slight multifocal urothelial changes consisting of cell swelling, dissociation, degeneration, and exfoliation after 3 and 5 days of treatment. After 2 and 4 weeks of treatment, elevated or leafy microridges on the luminal cell surfaces were seen together with foci of swollen cells. After a 2-month-recovery-period, the urothelial surfaces were normal. LM and TEM showed multifocal vacuolation of the urothelium associated with inflammation of the underlying lamina propria after 3 and 5 days of treatment. Cellular hypertrophy and hyperplasia of the transitional epithelium was seen after a 5-day treatment, persisted without increasing severity after 2 and 4 weeks of treatment, and totally regressed after the recovery period. It was concluded that, in the rat urinary bladder, oral administration of acetazolamide and MK-0927 induced early degeneration and inflammation followed by epithelial regeneration, resulting in a reversible hyperplasia of the transitional epithelium.

Corneal toxicity studies in rabbits and dogs with hydroxyethyl cellulose and benzalkonium chloride.

Hydroxyethyl cellulose (HEC) is used as a viscosity-enhancing agent in ophthalmic formulations to prolong corneal contact time and increase intraocular drug levels. Benzalkonium chloride (BAK) is the preservative most frequently used in ophthalmic formulations. Corneal epithelial changes were seen by slit lamp and light microscopic examination in rabbits but not dogs after multiple instillations of an ophthalmic vehicle containing 0.01% BAK and 0.5% HEC. Microscopically, there was sloughing of superficial epithelial cells and a slight loss of polarity of the basal cells. Formulations with 0.01% BAK and HEC, at concentrations between 0.3 and 0.8%; caused these changes but not with BAK or HEC alone. It was concluded that hydroxyethyl cellulose increased the viscosity and prolonged the contact time of BAK with cornea resulting in corneal epithelial damage in the rabbit. Physiological and anatomical features of the rabbit combined with the increased contact time were concluded to favor these changes in this species. The results confirm that the rabbit is a sensitive and unique species in studies of ocular toxicity of drugs.

Drug-induced changes in dogs after long term application of a beta-blocker.

Ocular irritation studies are important in the safety evaluation of ocular formulations. There are many reports on acute ocular irritation studies, but almost nothing about prolonged ocular instillation of ophthalmic formulations. This report discusses the predominantly lymphocytic infiltrates seen in the limbus corneae and eyelids of dogs treated with 1 and 2% solutions of L-653,328, an ocular hypotensive beta-adrenoceptor antagonist, for up to 53 weeks. The number of animals affected and severity of the lesions increased with time and concentration. A minimal effect was seen microscopically with the 2% solution as early as 14 weeks. Rabbits treated similarly for 14 weeks had no such changes. The first and only clinical sign in dogs was diffuse pinkness of the bulbar conjunctiva seen from Drug Week 22 onwards. Although not seen with similar molecular structures, given the equivocal results in sensitization studies and the long time required for the development of change, delayed contact hypersensitivity was suspected as the cause of the ocular infiltrates. Simple chronic irritation was not ruled out, however. These findings suggest that delayed contact hypersensitivity in dogs may be a phenomenon not limited to skin, but may also involve the eye after repeated ocular instillation.

Corneal endothelial changes with azone, a penetration enhancer.

Azone has been used to enhance percutaneous absorption. Its ability to improve penetration makes it an attractive candidate for incorporation into ophthalmic formulations to increase therapeutic action of a drug or achieve an equivalent effect with a lower concentration of the active ingredients. Ophthalmic vehicles containing 0, 1, or 2% Azone were studied to determine their ocular irritation potential in the rabbit. The vehicle ingredients were poloxamer 188, hydroxy-ethylcellulose, benzalkonium chloride and phosphate buffer. Rabbits received 30mcl topically of each of the test products three times daily for 29 days. Clinical and histopathological evidence of ocular toxicity occurred in eyes treated with the vehicle containing 1 or 2% Azone, but not in the vehicle without Azone. Clinical signs of ocular irritation were transient and included redness of conjunctivae and iris, discharge and corneal edema. Scanning electron microscopy and semi-thin sections revealed corneal changes characterized by ballooning and vacuolation of endothelial cells resulting in distortion of the typical polygonal appearance. These results indicate that instillation of ophthalmic vehicles containing 1 or 2% Azone damages corneal endothelial cells of the rabbit. It is not clear, however, if this irritation is due to the direct action of Azone on the endothelium or the enhanced penetration of potential irritants in the formulation such as benzalkonium chloride.

[Paris V and vertical dimension]. / Paris V et la dimension verticale.

For pedagogic reasons, the department of dento-facial orthopedics of the university of Paris V thinks that: the diagnosis would be based on the evaluation of some characteristics; the prognosis would appreciate possible dento-alveolar compensations; the orthopedic, orthodontic and surgical treatment would coincide perfectly with these diagnosis and prognosis data. The authors' opinion is based on records of dysmorphic and normal persons.

The phagocytic activity of Lumbricus terrestris leukocytes is enhanced by the vertebrate opsonins: IgG and complement C3b fragment.

The effect of the vertebrates opsonins: IgG and complement C3 fragments on phagocytic activity of Lumbricus terrestris leukocytes towards sheep erythrocytes was studied. Sheep erythrocytes were previously sensitized with specific IgG or IgM and coated with one of the third component fragments (C3b, C3bi, C3d) of human complement. Our results show that leukocyte phagocytosis was enhanced by vertebrate IgG and C3b complement fragments but not by IgM and fragment C3d. Because opsonization in vertebrates is related to the presence of receptors on the surface of phagocyte membrane, our results suggest that similar receptors exist on earthworm leukocyte surfaces. These new data strengthen the arguments in favour of the presence of components in Lumbricus terrestris which partially share common structures and functions with components of vertebrate humoral immune reactions.

Leiomyoma of the iris in a Sprague-Dawley rat.

A leiomyoma of the iris was composed of bundles of spindle-shaped cells with abundant myoglial fibres and represents the first observation of this rare tumour in a rat.

Evidence of synthesis by Lumbricus terrestris of specific substances in response to an immunization with a synthetic hapten.

Lumbricus terrestris (Annelid, Oligocheta) is capable of cellular- and humoral-specific reactions against natural antigens. Is this earthworm able to elaborate a response of antibody type against a synthetic hapten? L. terrestris have been immunized with conjugates made of one of two different synthetic haptens (400 mw) and a carrier protein: bovine serum albumin (BSA) or keyhole limpet haemocyanin (KLH). The presence of anti-hapten substances in coelomic fluid was tested against each iodinated hapten derivative (125I-hapten). The 125I-hapten-substance complexes were separated from the free derivatives by polyethylene glycol (PEG) 6000 precipitation or by gel filtration. The experiments showed that L. terrestris synthesized specific substances against the immunizing hapten. The importance of the response depended on the carrier protein and on the amount of introduced immunogen. A kinetic study of first and second immunization showed that these substances, elaborated in response to the immunization with the synthetic hapten, were synthesized by cells which kept the immunological memory. These data are discussed in relation to the humoral protection mechanisms already established in the invertebrates.