Persistent organ damage and cancer production in rats and mice.
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Animals↗
Biomedical subjects
P Grasso
Publications and source records attributed to P Grasso.
Experimental liver tumours in animals.
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Animals↗
Transglutaminase activity in bovine calf testicular membranes: evidence for a possible role in the interaction of follicle-stimulating hormone with its receptor.
Transglutaminase (TGase) activity, determined by incorporation of [1,4-14C]putrescine into casein, was demonstrated in a light membrane fraction derived from bovine calf testicular homogenates. In common with other TGases thus far identified, testicular TGase is calcium dependent. A concentration-dependent relationship was observed between decreasing TGase activity and increasing concentrations of N-ethylmaleimide and bacitracin, inhibitors of TGase, and the TGase substrate monodansylcadaverine. Although NEM at all concentrations tested had no effect on [125I]human (h) FSH-receptor binding, dissociation of [125I]hFSH from its receptor was enhanced when [125I]hFSH-receptor complexes were formed in the presence of NEM. Dissociation of [125I]hFSH from its receptor also increased in the presence of bacitracin or monodansylcadaverine. Reduced TGase activity always paralleled increases in hormone-receptor dissociation. The inverse relationship between TGase activity and [125I]hFSH-receptor dissociation observed in this study suggests that TGase activity may be involved in the interaction of FSH with its receptor and that protein cross-linking (via peptide bond formation) may be a mechanism whereby some FSH-receptor complexes are stabilized in the bovine testis.
Animals↗
Time and dose study on the response of rats to the hypolipidaemic drug fenofibrate.
Groups of male Wistar albino rats were administered diets containing sufficient fenofibrate to ensure intakes of either 200, 60 or 13 mg/kg/day or sufficient clofibrate to ensure an intake of 400 mg/kg/day. Four rats from each experimental group and 6 control rats were killed, 3, 7, 14 and 28 days, 8, 12 and 20 weeks and 6, 9, 12 and 18 months after commencement of treatment. At all time points livers were subjected to histological, electron microscopic and biochemical examination, the other major abdominal organs were removed for histological examination. A more extensive necropsy was carried out on rats killed after 12 and 18 months. The major alterations were observed in the liver, although there were also morphological changes in the thyroid, pancreas and kidney after prolonged treatment. The hepatic changes followed a distinct time course. Within 24 h of offering diets containing the compounds to the rats there was accumulation of small droplets of lipid, induction of peroxisomal enzymes and of the specific cytochrome P-450 catalysing omega-hydroxylation of fatty acids and an increase in the number of mitotic figures. More slowly developing changes were loss from the centrilobular zone of fat, glycogen and of glucose 6-phosphatase activity. Here maximal changes were observed after 14 days of treatment. A still more slowly developing change was accumulation of enlarged lipid-loaded lysosomes, which was maximal at 26 weeks, accompanied by the development of lipofuscin bodies. Finally, in animals treated for 12 months or more there was evidence for increasing cell turnover as indicated by an increased number of mitotic figures, more dark cells and induction of serum alanine transaminase. The last 2 groups of changes were not observed in rats treated with 13 mg/kg/day of fenofibrate. In general the degree of change in rats treated with 400 mg/kg/day of clofibrate was similar to those found in rats treated with 60 mg/kg/day of fenofibrate.
Animals↗
Effects of phthalic acid esters on the liver and thyroid.
The effects, over periods from 3 days to 9 months of administration, of diets containing di-2-ethylhexyl phthalate are very similar to those observed in rats administered diets containing hypolipidemic drugs such as clofibrate. Changes occur in a characteristic order commencing with alterations in the distribution of lipid within the liver, quickly followed by proliferation of hepatic peroxisomes and induction of the specialized P-450 isoenzyme(s) catalyzing omega oxidation of fatty acids. There follows a phase of mild liver damage indicated by induction of glucose-6-phosphatase activity and a loss of glycogen, eventually leading to the formation of enlarged lysosomes through autophagy and the accumulation of lipofuscin. Associated changes are found in the kidney and thyroid. The renal changes are limited to the proximal convoluted tubules and are generally similar to changes found in the liver. The effects on the thyroid are more marked. Although the levels of thyroxine in plasma fail to about half normal values, serum triiodothyronine remains close to normal values while the appearance of the thyroid varies, very marked hyperactivity being noted 7 days after commencement of treatment, this is less marked at 14 days, but even after 9 months treatment there is clear cut evidence for hyperactivity with colloid changes which indicate this has persisted for some time. Straight chain analogs of di-2-ethylhexyl phthalate, di-n-hexyl phthalate and di-n-oxtyl phthalate differ entirely in their short-term effects on the liver and kidney but have similar effects on the thyroid. The short-term in vivo hepatic effects of the three phthalate esters can be reproduced in hepatocytes in tissue culture. All three phthalate esters, as well as clofibrate, have early marked effects on the metabolism of fatty acids in isolated hepatocytes. The nature of these changes is such as to increase storage of lipid in the liver. A hypothesis is presented to explain the progress from these initial metabolic effects to the final formation of liver tumors.
Animals↗
Cancer hazard from exposure to solvents.
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Animals↗
Nuclear enlargement--an early change produced in mouse epidermis by carcinogenic chemicals applied topically in the presence of a promoter.
A series of 27 compounds, including 11 polycyclic hydrocarbons, 8 proximate carcinogens and 8 other compounds, were tested for their ability to induce epidermal nuclear enlargement in mouse skin. The materials were applied topically in methyl ethyl ketone containing 0.1% croton oil, repetitively for 3 days. Nuclear enlargement present on the fourth day was determined with the aid of a Quantiment image-analysing computer. The results obtained indicated a close correlation between the topical carcinogenicity of the test compounds and their ability to induce epidermal nuclear enlargement in the presence of croton oil.
Administration, Topical↗
Comparison of the short-term effects of di(2-ethylhexyl) phthalate, di(n-hexyl) phthalate, and di(n-octyl) phthalate in rats.
This study compares changes in the livers of rats treated with di(2-ethylhexyl) phthalate (DEHP) and its straight-chain analogs di(n-hexyl) phthalate (DnHP) and di(n-octyl phthalate (DnOP). Groups of rats were fed diets containing 20,000 ppm of one of these compounds. Subgroups were killed after 3, 10, and 21 days, and the livers were examined by histological, cytological, and biochemical methods. The results show considerable differences between the effects of the branched-chain phthalate ester DEHP and its straight-chain analogs. The major effects on the liver following administration of diets containing DEHP were midzonal and periportal accumulation of small droplets of lipid, hepatomegaly accompanied by an initial burst of mitosis, proliferation of hepatic peroxisomes and of smooth endoplasmic reticulum accompanied by induction of peroxisomal fatty acid oxidation, damage to the peroxisomal membranes as evidenced by increased leakage of catalase to the cytosol, and centrilobular loss of glycogen and falls in glucose-6-phosphatase activity and in low-molecular-weight reducing agents. In contrast, diets containing DnHP or DnOP induced accumulation of large droplets of fat around central veins leading, by 10 days, to mild centrilobular necrosis and a very slight induction of one peroxisomal enzyme and an increase in liver weight, but no significant changes in any other parameters which were affected by DEHP.
Administration, Oral↗
Time and dose-response study of the effects on rats of the plasticizer di(2-ethylhexyl) phthalate.
Groups of male and groups of female Wistar albino rats were administered diets containing sufficient di(2-ethylhexyl) phthalate (DEHP) to ensure intakes of either 1000, 200, or 50 mg/kg/day. Four rats from each experimental group and six control rats of the same sex were killed 3, 7, 14, and 28 days and 9 months after commencement of treatment. At all time points the major abdominal organs were removed and subjected to histological examination. A more extensive necropsy was performed on those rats killed after 9 months of treatment. At all time points the livers of the rats were subjected to extensive histologic, electron microscopic, and biochemical examination. Changes could be grouped according to their time course. Two early and transient alterations were noticed. First, there were morphologic changes in the bile canaliculi of male rats treated with 1000 mg/kg/day of DEHP. Second, there was a burst of mitosis immediately after the start of administration of the compound. The time course of this mitotic burst varied; the increase in mitosis was greatest at 3 days in rats treated with 1000 mg/kg/day of DEHP and was smaller but more prolonged in rats treated with 200 or 50 mg/kg/day. Other changes, namely, a midzonal to periportal accumulation of fat, induction of peroxisomal enzymes, and induction of the P-450 isoenzyme also developed rapidly but were sustained throughout the study. The maximal change was usually attained within 7 days of commencement of treatment. More slowly developing changes were hypertrophy of the hepatocytes, centrilobular loss of glycogen, and a fall in glucose-6-phosphatase activity. Here maximal changes were not attained until 28 days after commencement of treatment. These three effects were clearly observed in rats treated with 200 or 1000 mg/kg/day of DEHP but were only marginally altered in rats treated with 50 mg/kg/day. Finally accumulation of lipid-loaded lysosomes assessed by light and electron microscopy and by assay of beta-galactosidase activity was only apparent in rats treated with DEHP for 9 months with 200 or 1000 mg/kg/day of DEHP. Changes in female rats were qualitatively similar to those observed in male rats. The alterations were, however, less pronounced than in male rats treated with an equal dose of DEHP and the degree of liver enlargement was much less because, although the initial hyperplasia was clearly apparent, there was a much smaller degree of hypertrophy.
Administration, Oral↗
Peroxisome proliferation and hepatotoxicity in rodents.
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Animals↗
In vitro responses of luteinizing rat granulosa cells to human thyroid-stimulating hormone.
The effect of human thyroid-stimulating hormone (hTSH) on progesterone (P4) secretion during initial luteinization and subsequent prolactin (Prl)-mediated steroidogenesis by cultured rat granulosa cells was studied. Granulosa cells, obtained from pregnant mare's serum gonadotropin (PMSG)-treated immature female rats, were preincubated for 1, 3, 6, 12, or 24 h in control medium lacking added hormones or in medium containing 1.0 microgram/ml human chorionic gonadotropin (hCG) or hTSH, and maintained subsequently for 6 days in medium containing 1.0 microgram/ml bovine (bPrl). Indices of luteotropic stimulation were provided by: 1) elevated P4 concentrations determined by radioimmunoassay of spent media samples; and 2) cytoplasmic lipid accumulation assessed by osmium tetroxide staining following fixation after 7 days of culture. Progesterone levels in media from cultures exposed to hCG for 24 h were twofold higher than control cultures, whereas those in media from cultures preincubated in hTSH for 24 h were fourfold higher than control levels. Cultures preincubated in 1.0 microgram/ml hCG for as little as 1 h and then maintained for 6 days in Prl secreted significantly more P4 than did control cultures also maintained with Prl for 6 days. Cultures preincubated in hTSH required a 24-h exposure before a significant increase in Prl-mediated P4 secretion was observed. Intensity of cytoplasmic osmiophilia correlated directly with P4 concentration. These results suggest that: 1) hTSH has the ability to promote P4 secretion during initial luteinization and to regulate subsequent Prl-mediated steroidogenesis by cultured rat granulosa cells; and 2) the mechanism by which hTSH stimulates Prl-mediated P4 secretion in this model system may differ from that of hCG.
Animals↗
Prolactin-mediated progesterone secretion by cultured rat granulosa cells: regulation by purified human glycoprotein hormones and their subunits.
The effects of purified alpha- and beta-subunits of human glycoprotein hormones on initial luteinization and subsequent prolactin-mediated progesterone responses of cultured rat granulosa cells were studied. Granulosa cells, obtained from immature female rats 50 h after PMSG treatment, were incubated for 24 h in control medium lacking added hormones or in medium containing hCG or the alpha- or beta-subunit of human (h) FSH, LH, CG, or TSH at 0.1, 0.5, and 1.0 microgram/ml. Cultures were maintained subsequently for 6 days in medium containing 1.0 microgram/ml bovine PRL (bPRL), with medium changes every 48 h. Indices of luteotropic stimulation in response to bPRL were provided by 1) elevated progesterone concentrations determined by RIA of spent media samples, and 2) cytoplasmic lipid accumulation assessed by osmium tetroxide staining following fixation of monolayers after 7 days of culture. Progesterone concentrations in media from cultures incubated in 0.5 or 1.0 microgram/ml hCG were 6-fold higher than in cultures incubated in control medium, while those in media from cultures incubated in 0.5 or 1.0 microgram/ml hFSH alpha, hLH alpha, hCG alpha, hTSH alpha, hLH beta, or hCG beta (but not in hFSH beta or hTSH beta) were from 2- to 4-fold higher than those in control cultures. This enhancement was not evident when subunits were added to the incubation media at the lowest concentration. Progesterone secretion corresponded directly with the degree of cytoplasmic osmiophilia. These results suggest that the alpha-subunit of each of the glycoprotein hormones as well as the beta-subunit of hLH and hCG have the ability to promote progesterone secretion during initial luteinization and to regulate subsequent PRL-mediated steroidogenesis by rat granulosa cells in vitro. Furthermore, these effects are greater than can be accounted for by potential contamination of subunit preparations with undissociated hormones, as demonstrated by dose-response curves.
Animals↗
Genotoxicity studies on di-(2-ethylhexyl) phthalate and adipate and toxicity studies on di-(2-ethylhexyl) phthalate in the rat and marmoset.
These studies have provided evidence that DEHP and DEHA do not bind covalently to DNA and do not therefore possess the characteristics of a genotoxic agent (Lutz, 1982). This suggests that the tumours induced in the rodent liver may result from some non-genotoxic mechanism and supports the view that the weakly positive dominant lethal test seen on administration of DEHP by the ip (but not the oral) route (Singh et al. 1974) is unlikely to have resulted from a direct effect on the genome of the sperm cells. Although the mechanism responsible for the induction of tumours by high doses of DEHP in rodents is not clear, it would appear both from these studies and from work on hypolipidaemic agents, that peroxisomal proliferation and the induction of enzymes associated with this organelle are in some way implicated (Cohen & Grasso, 1981). Other studies have shown that changes of this type are produced by doses of hypolipidaemic agents that induce liver cancer in rodents (Cohen & Grasso, 1981) and our investigations have indicated that they were also prominent at dose levels of DEHP similar to those that induced liver cancer in the NCI study (National Toxicology Program, 1982). No cancer induction would be expected to occur in the absence of these changes. In our dose-response study in rats it was shown that at the lowest dose (50 mg/kg body weight/day, approximately equivalent to a dietary level of 1000 ppm) several effects seen with higher doses were not apparent and others differed only slightly from normal control values. This is particularly relevant to assessments of the risk posed by DEHP and DEHA present as contaminants in foods, since human exposure via the food chain has been estimated by Shiota, Chou & Nishimura (1980) as 30 micrograms/kg body weight/day, several orders of magnitude less than the lowest exposure level used in these experiments. In addition, our studies indicate that none of the changes found in the rat were observed in the marmoset, suggesting that rodents and primates differ fundamentally in their hepatic and testicular response to DEHP. Previous studies by other authors (reviewed by Cohen & Grasso, 1981) indicated that morphological changes in the endoplasmic reticulum and the proliferation of peroxisomes are not features of the response of monkeys and man to high doses of hypolipidaemic agents.(ABSTRACT TRUNCATED AT 400 WORDS)
Adipates↗
Neurophysiological and psychological disorders and occupational exposure to organic solvents.
A number of reports, particularly from Scandinavian countries, claim that painters and workers in other trades in which prolonged occupational exposure to organic solvents may occur develop a type of mental illness characterized principally by impairment of memory and co-ordination and some deterioration of personality. The condition, called 'organic solvent disease', is recognized as a cause of premature retirement and is classed as an occupational disease in certain countries. The conclusions of these reports have been contested and the existence of such a disease entity has been questioned. The publications reporting adverse neurological, neurophysiological and psychological disorders in solvent-exposed workers, and the methods used to determine adverse effects, have therefore been evaluated. In addition, data from animal behavioural studies have been examined but were found to have little or no to have little or no relevance to the reported human disease. The human data indicate that, of the solvents studied, only CS2 provided clear evidence of neurotoxic damage detectable by clinical and pathological examination as well as by neurophysiological measurements (e.g. nerve conduction velocity and nerve action potentials) or neuropsychological techniques (e.g. Rorschach inkblot test and WAIS intelligence tests). In the case of several other solvents and mixtures of solvents commonly used in industry, the evidence of CNS impairment, based principally on the response to questionnaires and the results of neuropsychological and neurophysiological examinations was questionable. A critical evaluation of the reliability of these methods in detecting minor deviations from normal and of their ability to provide acceptable evidence of CNS dysfunction or damage leaves little doubt that these methods are of value in investigating personality, intelligence and memory in the clinical examination of individual patients. However, evidence indicates that they are not suitable for use in epidemiological studies, principally because the variability of response in normal individuals is ill-defined and insufficiently investigated. The same conclusion was arrived at in evaluating the contribution of electroencephalography, computerized axial tomography scanning and other electrophysiological examinations to the diagnosis of brain changes in groups of solvent-exposed and unexposed workers. Furthermore, the personality changes identified (by neuropsychological tests) in painters and other workers exposed to solvents could well be produced by ageing, exposure to lead or mercury, excessive alcohol intake, psychoactive drugs or the ordinary stresses of everyday life.(ABSTRACT TRUNCATED AT 400 WORDS)
Aging↗
Response of the upper respiratory tract of the rat to white spirit vapour.
The threshold limit value (TLV) adopted by industry for white spirit is that of Stoddard's solvent and is currently set at 100 ppm (525 mg/m3). Exposure of rats to atmospheres containing an average of 214 mg/m3, 4 h/day, for 4 consecutive days resulted in irritation of the membranes lining the upper respiratory tract but only minimal signs of lung injury. Histopathological changes were observed in the nasal cavity, trachea and larynx and included loss of cilia, mucous and basal cell hyperplasia and squamous metaplasia. These results suggest that in man exposure to white spirit at or around the TLV may result in irritation of the upper respiratory tract.
Animals↗
Aspartame: succession of false alarms.
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Animals↗
Nitrosamine carcinogenesis in 5120 rodents: chronic administration of sixteen different concentrations of NDEA, NDMA, NPYR and NPIP in the water of 4440 inbred rats, with parallel studies on NDEA alone of the effect of age of starting (3, 6 or 20 weeks) and of species (rats, mice or hamsters).
A Weibull analysis is presented of the dose and time relationships for the effects on 4 080 inbred rats of chronic ingestion in the drinking water of 16 different doses of N-nitroso-dimethylamine (NDMA) and of N-nitrosodiethylamine (NDEA). The sites chiefly affected were the liver (by both agents) and the oesophagus (by NDEA only). Since the experiment continued into extreme old age, effects became clearly measurable even at a dose of only 0.01 mg/kg per day, which is an order of magnitude lower than previously achieved. (After only two years of treatment, however, the 'TD50' doses needed to halve the proportion of tumourless survivors would have been about 0.06 mg/kg per day of NDEA and about 0.12 mg/kg per day of NDMA.) The general pattern of response was that the log probability of remaining tumourless was given by the product of two terms, the first (the 'Weibull b-value') depending on the dose-rate but not on the duration of exposure, and the second depending not on dose at all but on (approximately the seventh power of) duration. For oesophageal tumours, the 'Weibull b-value' was approximately proportional to the cube of the dose-rate of NDEA (males 21 d3, females 11 d3, where d = dose-rate in mg/kg adult body weight/day, and the background incidence was unmeasurably low. For liver tumours induced by NDEA, the b-value was approximately proportional to the fourth power of dose-rate + 0.04 mg/kg per day (males, 19 (d + 0.04)4; females, 32 (d + 0.04)4), although the relationships were slightly different for the different subsites of liver tumour. This one formula implies both approximate linearity at low doses and an approximately cubic relationship within the higher range of doses that was studied. For liver tumours induced by NDMA, the Weibull b-value was approximately proportional to the sixth power of dose rate + 0.1 mg/kg per day (males, 37 (d + 0.1)6; females, 51 (d + 0.1)6), again with variation between liver subsites, and again implying approximate linearity at low doses. The difference between the latter two relationships may represent differences in the induction of particular DNA repair enzymes by NDMA and NDEA, or in the effects of those enzymes on methylated and on ethylated DNA. These formulae should, of course, be trusted only in the range of doses from which they were derived, and particularly not for those above it.(ABSTRACT TRUNCATED AT 400 WORDS)
Age Factors↗
Changes in medullary glycosaminoglycan histochemistry and microvascular filling during the development of 2-bromoethanamine hydrobromide-induced renal papillary necrosis.
A single dose of 2-bromoethanamine (BEA) hydrobromide induced an apex limited renal papillary necrosis (RPN) at low doses (50 mg/kg, ip and sc, or 100 mg/kg, po), and the total ablation of the medulla at high doses (greater than 150 mg/kg, ip and sc, or 1000 mg/kg, po) in both male and female Wistar rats. BEA (at levels of 50, 100, and 150 mg/kg ip to male rats) caused transient hydropic changes. By 12 to 24 hr the anatomical elements of the medulla were necrosed, and active regeneration and reepithelialization of the viable areas was apparent. The only cortical changes up to 120 hr were dilatation of the nephron and the Bowman's spaces. The medullary glycosaminoglycan matrix (assessed by histochemical staining with colloidal iron) gave an intense granular appearance 2 hr after BEA, which was more marked and diffuse in the apex of the medulla from 8 to 12 hr. The staining was lost from those areas where early necrotic changes had occurred by 12 to 24 hr, and there was no subsequent staining of matrix material up to 120 hr. Vascular filling with colloidal carbon particles suggested an early shift of blood flow to the outer medulla at 2 to 4 hr, followed by the sustained filling of the microvasculature of the medulla well into the period when frank necrotic change was apparent. These data suggest that changes in the medullary matrix might play a previously unrecognized role in the development of 2-bromoethanamine hydrobromide-induced renal papillary necrosis, and these changes may give rise to loss of the material that supports the medullary capillaries.
Animals↗