M. Webb

Influence of dietary cadmium on the distribution of the essential metals copper, zinc and iron in tissues of the rat

The effect of long-term dietary cadmium treatment upon the distribution of the metals copper, iron and zinc has been compared in various organs of male and female rats. The renal accumulation of cadmium was similar in both sexes without a plateau being reached. In contrast, the hepatic accumulation of cadmium was higher in the female than in the male rat and a plateau was observed after 30-35 weeks of dietary cadmium treatment. Most of the cadmium which accumulated in these organs was recovered in the metallothionein fraction andthe concentration of hepatic cadmiumthionein in the female rat was correspondingly higher than in the male rat. Accumulation of cadmium was associated with an increased zinc concentration in the liver and an increased copper concentration in the kidney; these increases were correlated with increases in liver and kidney metallothioneins induced by cadmium. Uptake of cadmium into organs other than liver and kidney occurred to a small extent but was not associated with changes in the concentration of copper and zinc. Cadmium also accumulated in the intestinal mucosa where it could be recovered in a fraction corresponding to metallothionien. A loss of iron from the liver and kidney was also observed following dietary cadmium treatment and involved mainly a loss of iron from ferritin.

Localisation of the MRC OX‐2 Glycoprotein on the Surfaces of Neurones

Abstract: The MRC OX‐2 monoclonal antibody recognises membrane glycoproteins of Mr 41,000 in rat brain and 47,000 on thymocytes. It also reacts with follicular dendritic cells in lymphoid organs, endothelium, smooth muscle, and B‐lymphocytes. Indirect immunoperoxidase staining of cryostat sections showed that OX‐2 antigen was present throughout the cerebellum, with staining of both grey and white matter. Blood vessels were also stained. The Purkinje cell layer appeared to be unlabelled. Double‐immunofluorescence staining of cerebellar interneurone cultures with MRC OX‐2 antibody and tetanus toxin showed that all tetanus‐positive cells (neurones) were MRC OX‐2‐positive. Glial fibrillary acidic protein‐positive astrocytes were not labelled by MRC OX‐2 antibody. Thus OX‐2 antigen is one of the few biochemically characterised components of neuronal membranes and its properties are compared with those of the neuronal membrane glycoprotein Thy‐1.

Effects of dimethoxyethyl phthalate, monomethoxyethyl phthalate, 2-methoxyethanol and methoxyacetic acid on post implantation rat embryos in culture

The secondary metabolite of dimethoxyethyl phthalate (DMEP), methoxyacetic acid (MAA), but neither the diester nor either of its primary metabolites, monomethoxyethyl phthalate (MMEP) and 2-methoxyethanol (ME), interferes with normal growth and development of organogenesis phase rat embryos in culture. These in vitro observations suggest that the teratogenicity of DMEP in vivo is due to enzymic cleavage of the diester to ME, followed by oxidation of the latter to MAA in the maternal compartment.

The teratogenicity of methoxyacetic acid in the rat

Intraperitoneal (i.p.) administration of methoxyacetic acid (MAA) to rats on Day 8, 10, 12 or 14 of pregnancy was embryolethal and teratogenic. Skeletal anomalies, hydrocephalus and dilatation of the kidney pelvis were the most common malformations. Embryonic response to MAA varied with gestational age and with dosage (0.1 to 2.5 mmol/kg). These actions are similar to those previously reported for 2-methoxyethanol (ME) and dimethoxyethyl phthalate (DMEP). Embryos were also examined on Day 12, 48 h following i.p. administration of 2.5 mmol/kg MAA. Abnormalities were comparable to those previously observed following MAA treatment of rat conceptuses in culture. These data support the conclusion that MAA is the proximal teratogenic metabolite of ME and DMEP.

The teratogenic potential of alkoxy acids in post-implantation rat embryo culture: Structure-activity relationships

Alkoxy acids are the active metabolites of teratogenic glycol ethers. To examine the relationship of chemical structure to embryotoxicity, the effects of 6 acids on the development of 9.5-day rat embryos over 48 h in culture were studied. Methoxyacetic acid (MAA) and ethoxyacetic acid (EAA) (5 mM) were growth-retarding and induced gross structural defects, with MAA being more effective. n-Propoxyacetic acid (n-PAA) and n-butoxyacetic acid (n-BAA) (5 mM) were markedly less embryotoxic and produced only minor anomalies. Thus, the activities of these substituted acetic acids decreased with the increase in the length of the alkoxy chain. 3-Methoxypropionic acid (3-MPA) and 4-methoxybutyric acid (4-MBA) (5 mM) were much less active than MAA and induced only minor defects. Therefore in this series: RO(CH2)nCOOH, an increase in the value of n caused a greater reduction in embryotoxicity than did an increase in chain length of the alkyl group R.

Complex formation between selenium and methylmercury

Methylmercury, after incubation at 3k7 degrees C and pH 7.0 with selenite in the presence of rat erythrocytes, can be extracted into benzene as an unstable 2 : 1 complex with selenium. The same complex, possibly bis-methylmercury selenide, is formed when methylmercury is treated with hydrogen selenide at pH 7.0 in the absence of erythrocytes.

The interaction of cadium-induced rat renal metallothionein with bivalent mercury in vitro

Addition of Hg2+ in vitro to metallothionein (Cd : Cu : Zn = 6.5 : 4 : 1) from the kidneys of Cd2+ exposed rats appears to result initially in the replacement of Zn2+, then Cd2+ and finally copper. The ionic stoichiometries between Hg2+-binding and the release of Cd2+ (or Zn2+) and copper are 3 : 2 and 1 : 1 respectively. After treatment with amounts of Hg2+ sufficient to displace completely either the bound Zn2+ and Cd2+, or all of the original cations, the metallothionein has little or no negative charge at pH 8.0 and is not resolved into the two isometallothioneins, which characterize the (Cd, Cu, Zn)-thionein, by ion exchange chromatography at this pH. Cation substitution occurs in both isometallothioneins and there is no evidence that Hg2+ interacts preferentially with one of them. Treatment of the metallothionein with increasing amounts of Hg2+, equivalent to approx. 25% and 50% of the bound cations gives rise to a range of molecular species of progressively decreasing negative charge. The consistent stiochiometry between Hg2+ uptake and Cd2+ or Zn2+ release at all concentrations of Hg2+ indicates that free thiol groups are not formed during the substitution reaction.

The toxicity and teratogenicity of mercuric mercury in the pregnant rat

Mercuric mercury (Hg2+), when injected IV into the pregnant Wistar rat, is retained mainly in the maternal compartment and uptake by the conceptuses is small. Thus if the dose is based on total body weight, the maternal body burden, particularly in late gestation, is greater than the whole body burden in the non-pregnant animal. The LD50 of Hg2+ (mg/kg total body weight), however, remains essentially constant (1.0–1.2 mg Hg2+/kg) throughout pregnancy. It seems, therefore, that the rat becomes more resistant to Hg2+ with increasing gestational age. This increased resistance does not correlate with differences in (a) the uptake of Hg2+ by the kidneys, the target organs of toxicity, (b) the severity of the histopathologically detected renal damage and (c) the inhibition of glomerular filtration. Biochemical measurements, however, suggest that kidney function may become less susceptible to Hg2+ as pregnancy advances from conception to near term. During mid-gestation the minimum effective teratogenic dose of Hg2+ (0.79 mg/kg total body weight) is high in relation to the maternal LD50 and the incidence of foetal malformations, mainly brain defects (23% in all live foetuses), is low. In rats of different gestational ages uptake of Hg2+ by the embryo/foetus at this dose level decreases sharply between day 12 and day 13. The teratogenic effects in the foetus and both the structural and functional damage to the maternal kidneys, however, are essentially the same in animals that are dosed with Hg2+ either immediately before, or immediately after these gestational ages. It is probable, therefore, that foetal defects result not from any direct action of Hg2+ on the conceptuses, but from either the inhibition of the transport of essential metabolites from the mother, or the maternal kidney dysfunction.

The stimulation and inhibition of the exhalation of volatile selenium

Administration of methylmercury (1.5-24 mumol kg-1; s.c.) to female rats simultaneously with Na2 75SO3 (0.25 or 24 mumol kg-1; s.c.) causes a dose-dependent increase in the exhalation of dimethylselenide. At the low selenite dose level, exhalation of 75Se over a 24 hr period is about fourfold greater after treatment with 24 mumol kg-1 methylmercury than that (approximately 0.75% of the dose) in the controls, but excretion by other routes (urine, faeces) and the liver and kidney contents of 75Se are not affected significantly. At the higher selenite dose level (24 mumol kg-1) exhalation of 75Se is correlated with the log dose of methylmercury. The faecal and urinary excretion remains essentially unaffected, and in rats treated with 24 mumol kg-1 methylmercury the 75Se contents of the liver, kidneys and blood are reduced by 78%, 86% and 18% respectively. The effects of the alkylmercurial are not specific since, at this selenite dose level, ethylmercury increases the exhalation and decreases the liver and kidney contents of 75Se approximately to the same extent as an equimolar dose of methylmercury. In methylmercury-treated and control animals dosed with 24 mumol kg-1 Na 75SeO3 the exhalation of 75Se is inhibited to the same extent by periodate-oxidized adenosine (PAD; 15 mumol kg-1, i.p.) in the first 6 hr. Later inhibition is less pronounced in methylmercury-treated rats. Under these conditions PAD has little effect on the renal content, but increases the hepatic content of 75Se. It seems, therefore, that the methylation of selenite occurs mainly in the liver and in both control and methylmercury-treated animals, S-adenosylmethionine is the major methyl donor. It is possible that methylmercury does not affect directly the methylation enzyme system but, by competition for protein sulphydryl groups, increases the availability of the intermediary selenide anion.

Endogenous metal binding proteins in relation to the differences in absorption and distribution of mercury in newborn and adult rats

Both in the intestinal copper complex and hepatic zinc-thionein of the newborn rat provide immediately available, high-affinity binding sites for Hg2+. Retention of orally administered Hg2+ in the gastrointestinal tract is much greater in the 5-day old rat than in the weanling and accounts for most (70–90%) of the body burden at 18 h. At a dose level of 790 μg Hg2+/kg body weight, 41% of the Hg2+ retained in the gastrointestinal tract of the newborn animal is located in the soluble fraction of the mucosa and 48% of this is bound by the copper-complex. This complex, therefore, seems to limit the transfer of Hg2+ from the mucosa to the serosa. It does not appear, however, to be a primary acceptor of Hg2+ in the neonatal intestine since, after administration of 100 μgHg2+/kg body weight, its binding of Hg2+ is less (33% of the soluble Hg2+) than that (67% of the soluble Hg2+) of the high molecular weight protein fraction.At 18 h after the intraperitoneal administration of Hg2+ (790 μg Hg2+/kg body weight), whole body retention is greater in the adult (80% of the dose) than in the suckling or weanling animal (63–68% of the dose). In the suckling animal the inter- and intra-organ distributions of the retained Hg2+ also differ from those in the weanling and/or adult rat. Thus the ratio of the renal: hepatic Hg2+ contents, after either intragastric or intraperitoneal doses of Hg2+, increases with age. Also most (approximately 90%) of the Hg2+ retained in the liver of the newborn animal after an intragastric dose is bound by the metallothionein whereas, in the adult liver, only 4% of the total Hg2+ is in this form. The presence of the metallothionein, however, is not the cause of the increased Hg2+ uptake in the former, since induction of hepatic zinc-thionein in the adult by pretreatment with Zn2+ does not affect the distribution of Hg2+ between the liver and kidneys.These findings are discussed in relation to the known differences in intestinal absorption and tissue distribution of Cd2+ and Hg2+ in newborn and adult animals and the greater resistance of the former to Hg2+ nephrotoxicity.