Efficacy of an anti-fertility vaccine based on mammalian gonadotrophin releasing hormone (GnRH-I)--a histological comparison in male animals.

A N-terminal modified gonadotrophin releasing hormone (GnRH-I, tetanus toxoid-CHWSYGLRPG-NH2) conjugate was evaluated histologically in a number of male animal species (mice, dogs and sheep). The immunogen has previously been shown to be highly effective in rats, by suppressing both steroidogenesis and spermatogenesis. However, cross-species efficacy of peptide vaccines is known to be highly variable. Therefore, a comparative evaluation of reproductive tissues from animals immunized against this immunogen adsorbed onto an alum-based adjuvant was made. The sheep and dogs were chosen, as use of anti-fertility vaccines in these species is important in farming and veterinary practice. Changes in testicular size were measured during the immunization period and the greatest alteration (attributed to gonadal atrophy) was observed in the rat. Following euthanasia, the testicular tissue was evaluated for spermatogenesis. The most susceptible species to GnRH-I ablation was the rat, which showed significant (P < 0.0001) arrest in spermatogenesis compared with untreated controls. Testicular sections taken from treated animals were completely devoid of spermatozoa or spermatids, in comparison with 94% of the untreated controls showing evidence of spermatogenesis. The immunized mice and rams also showed significant arrest (P < 0.0001). There was a 30-45% decrease in spermatogenesis and total azoospermia was not apparent. However, the least responsive were the dogs, which showed little significant variation compared to untreated animals and only a 5% decrease in activity. A comparison of the specific IgG response to GnRH-I indicated that in sheep and dogs the response was not maintained, unlike in rodents, suggesting that suppression of fertility may be due to differences in immune responses in different animal species.

Ovarian function and FSH receptor characteristics during canine anoestrus.

Ovaries of bitches are relatively inactive during anoestrus despite apparently adequate circulating FSH concentrations. Alternative FSH receptor (FSH-R) transcripts in bitches might hinder the follicular response to gonadotrophins, which may account for anoestrus. The expression of the full length FSH-R and novel isoforms in bitches was examined using in situ hybridization and RT-PCR analysis. Various PCR primers to the FSH-R were used and its expression was assessed in ovarian tissue at different stages of the oestrous cycle. RT-PCR amplification of the extracellular domain (exon 1-10) was generally successful, indicating that cFSH-R expression (> 90%) occurs throughout the oestrous cycle. Two FSH-R isoforms were sequenced, but there were no clear differences in the pattern of expression between anoestrus and other stages of the oestrous cycle, except that isoform expression was less frequent (30% occurrence) in prepubertal bitches. Data from in situ hybridization showed clear expression of the FSH-R in secondary and antral follicles, and corpora lutea. It was concluded that there is no evidence of a change in the expression of the FSH-R specific to anoestrus.

Measurement of plasma oestradiol after an injection of a gonadotrophin as a test for neutered bitches.

Twenty-eight bitches with unknown reproductive histories were injected intravenously with either human chorionic gonadotrophin (hCG) or equine chorionic gonadotrophin (eCG) (pregnant mare's serum gonadotrophin) and their oestradiol responses were measured at the time of the injection and 90 minutes later. They were at various stages of the oestrous cycle as determined by histology and a progesterone assay for luteal function. Twenty-six of them were considered to be entire because they showed either an increase in plasma oestradiol over preinjection values or steady high values. The ovaries were removed from 25 of these animals and the other probably had a remnant of ovary because it came into oestrus some weeks later. In two remaining bitches no oestradiol could be detected either before or after the injection of gonadotrophin and they were predicted to have been neutered, which was confirmed at laparotomy. In the entire bitches, the highest plasma oestradiol concentration was measured during metoestrus and the lowest during anoestrus.

Cadmium concentrations in human kidney in the UK: 1978-1993.

Almost 2700 samples of human renal cortex have been collected from throughout the UK over a 16 year period from 1978 to 1993. The mean Cd concentration was 19 micrograms g-1 and the median 16 micrograms g-1. Smokers were, on average, about 5 micrograms g-1 higher than non-smokers. Cd increased from low concentration in the young to a maximum of 23 micrograms g-1 in middle age followed by a decrease in old age. Subjects who had died of renal disease had lower Cd concentrations. Geographical variations in the UK are small and the concentrations appear to be static over the 16 year period. Some 3.9% of the population had Cd concentrations > 50 micrograms g-1, the critical level at which beta 2-microglobulin appears in urine.

Coronary endothelial dysfunction increases the severity of ischaemia-induced ventricular arrhythmias in rat isolated perfused hearts.

In order to determine the role of coronary vascular endothelial cells in generating cardioprotective substances during myocardial ischaemia, rat isolated hearts, perfused at constant flow by the Langendorff technique, were subjected to treatment with the detergent Triton X100 and the responses of these hearts to a 30 or 60 min period of coronary artery occlusion was determined. Endothelial damage or denudation was shown both by histological examination and by the altered vasodilator response to the endothelium-dependent vasodilator bradykinin (100 nM), which was reversed to vasoconstriction in hearts treated with Triton X-100. In contrast, the responses to sodium nitroprusside (100 microM) were unimpaired in these hearts and were not different from control responses. Ventricular ectopic activity was much more pronounced in hearts with endothelial dysfunction (e.g., 3329 +/- 361 ventricular premature beats over a 30 min occlusion period compared to 243 +/- 34 in controls; P < 0.01), and the duration of ventricular tachycardia was greatly increased (1162 +/- 391 s v 9 +/- 12 s in the controls; P < 0.01). Ventricular ectopic activity was still marked when the occlusion was prolonged to 1 h and was still apparent at the end of this 1 h occlusion period. Reperfusion arrhythmias (ventricular tachycardia and ventricular fibrillation) were marked in endothelium-damaged hearts (50%); whereas there were no such arrhythmias after a 30 or 60 min occlusion period in control hearts. Hearts were also preconditioned by a 3 min coronary artery occlusion period 10 min prior to a 30 min coronary artery occlusion. This reduced ventricular ectopic activity in both control and endothelium-damaged hearts to about the same extent (between 80 and 90% suppression). The results suggest that under normal conditions substances generated from endothelial cells protect the myocardium against ventricular arrhythmias both during ischaemia and reperfusion. However, in this species, preconditioning is still possible in hearts from which the coronary vascular endothelium has been removed. If these results can be extrapolated to the clinical situation, it suggests that in patients with endothelial dysfunction ventricular arrhythmias may be more pronounced following a period of ischaemia and especially of reperfusion.

Sensitivity of the early luteal phase ovine cervix to prostaglandin E2 (PGE2) and expression of EP3 receptor mRNA.

The effects and mechanism of action of prostaglandin E2 (PGE2) on the ovine cervix are largely unknown in the luteal phase. In these studies we have shown that low levels of EP3-receptor (EP3R) mRNA are present in the ovine cervix and that the PGE2 induces activation of polymorphonuclear leukocytes in the ovine cervix on day 6 of the oestrous cycle. It is possible, therefore, that PGE2 acts on the ovine cervix through coupling to EP3 receptors.

Effects of L-canavanine, an inhibitor of inducible nitric oxide synthase, on endotoxin mediated shock in rats.

The effects of L-canavanine, an inhibitor of nitric oxide synthase, on endotoxin-induced shock was investigated in the pentobarbitone anesthetized rat. Endotoxin infusion (2.5 mg kg-1 h-1 over 6 h) produced progressive and marked hypotension and hypoglycemia. Electron microscopy showed marked changes in the kidney, comprising severe endothelial cell disruption and the accumulation of platelets in the blood vessels. In the lung, there was marked accumulation of polymorphonuclear leukocytes in small blood vessels and endothelial disruption. Treatment with L-canavanine (10 mg kg-1 by bolus injection each hour starting 70 min after endotoxin or saline infusion) significantly reduced endotoxin-induced hypotension, without any effect on the hypoglycemia. This treatment markedly reduced the endotoxin-induced electron microscopical changes in the kidneys and lungs. Although L-canavanine, like L-NAME, inhibited both cerebellar constitute and splenic inducible nitric oxide synthase in vitro, in contrast to L-NAME it did not modify either arterial blood pressure or carotid artery blood flow in control rats. The data are consistent with L-canavanine being a selective inhibitor of inducible nitric oxide synthase, at least in vivo, and suggest that inhibitors of this enzyme may be beneficial in endotoxin-induced shock.

Cadmium concentrations in human kidneys from the UK.

The concentration of cadmium in the cortex and medulla of nearly 1000 kidneys obtained at autopsy was determined by atomic absorption spectrophotometry. Particular care was taken over the analytical procedures to ensure the accuracy and precision of the results; intrakidney variability was examined and found to be small. The frequency distributions of cadmium concentrations were approximately lognormal. There was much variability in cadmium concentrations between individuals, with geometric coefficients of variation for both cortex and medulla of around 100%. Cadmium concentrations varied with age and smoking habits, but not sex. Values were successively higher than those in the previous age-group up to 50-59 years (geometric mean 19 micrograms/g), after which they were successively lower; on average, heavy smokers had cadmium concentrations some 15-20% higher than those in light smokers who in turn had values some 15-20% higher than those in non-smokers. No definite conclusions could be reached on the variation of cadmium concentrations with either cause of death or location, or over time. The present study, the first of its kind in the UK, has provided benchmark data; its results are broadly in line with those from previous but smaller studies in other countries.

An ultrastructural study of the kidney of normal, copper poisoned and thiomolybdate-treated sheep.

Histological, ultrastructural and kidney function techniques were used to assess changes in the kidney of sheep given either copper (Cu) or Cu and the Cu complexing agent thiomolybdate (TM), or TM alone. Kidney function was normal in sheep given Cu and TM together or TM alone. In these animals the cells lining cortical tubules accumulated Cu within numerous, large, electron-dense lysosomes. Sheep given Cu alone developed haemolysis, impaired kidney function and a variety of morphological defects including an increase in number of large lysosomes in cells of the cortical tubules. There was a breakdown of the glomerular endothelial lining and fusion of foot processes. Cells of the cortical tubules showed degeneration and necrosis and an increase in microbodies and rough endoplasmic reticulum. Cortical and medullary blood vessels were dilated, with evidence of breakdown of the endothelial lining. Copper appeared to injure kidney tissue at three sites, tubular epithelium, glomerular basal lamina and capillary blood vessels. Changes reported here are similar to the renal lesions in cadmium toxicity.

Histopathology of early effects of oral cadmium in the rat kidney.

Adult male Wistar rats were given 50 ppm Cd in drinking water over a period of 1-24 weeks. The rats were killed and the cadmium concentration of whole blood, blood plasma red cells, liver and kidneys estimated. The plasma metallothionein concentration was measured by radioimmunoassay. Kidney samples were taken for light, transmission and scanning electron microscopic examination. The accumulation of cadmium in the tissues was shown by a linear increase with time, after exposure for 12 weeks. Plasma Cd concentrations showed a clear increase after 3 weeks and preliminary investigation suggests that most is present as Cd-thionein. Early pathological changes in the rat kidney were seen around the 4-6 week period which coincided with the distinct rise in plasma Cd. At 12 weeks, signs of tubular necrosis, interstitial fibrosis and glomerular epithelial cell hypertrophy were present in small areas of the cortex. By 24 weeks, the renal cortex showed clear evidence of tubulo-interstitial nephritis at a Cd concentration of 60 micrograms Cd/g wet weight.

Pregnancy-associated changes in the physical and microscopic characteristics of the ovine cervix.

Ovine cervices were obtained at slaughter from 18 sheep comprising six groups of three--cycling ewe lambs, non-pregnant, and four groups of pregnant animals of progressively longer gestation PI to PIV. Physical and microscopic analyses carried out on these cervices showed consistent results throughout any individual tissue. A number of factors changed with increasing gestational age and some of these changes were noted from early gestation. No significant change in water content was observed through pregnancy. With increasing length of gestation the cervix increased in length and width, there were relative increases in fibroblasts, smooth muscle and softening of the tissue, and relative decreases in collagen and fibrocytes. An increased vascularity was observed in the tissues from late gestation sheep, and a change in the nature of the epithelial secretion, from neutral to acidic mucins, was also associated with late pregnancy.

Renal cadmium content in the West of Scotland.

The trace element content of kidneys has been documented in very few international centres. Significant differences can be demonstrated between the cortical and medullary levels of calcium, zinc and cadmium. For the West of Scotland the values for the latter are similar to those found in Scandinavia. There are significantly higher cadmium contents in subjects who smoke and who have evidence of ischaemic heart disease.

An unusual abnormality of human spermatozoa.

A defect affecting 90% of the spermatozoa of an infertile man consisted of round heads and a loss of tails, probably caused by a developmental abnormality at the spermatid stage.

Methyl cholanthrene and cadmium induced changes in rat prostate.

Cadmium workers have been thought to be more likely to develop prostatic cancer. The present study compares the effects of cadmium on rat prostate as compared with a known carcinogen, 20 methyl cholanthrene. The various histological effects produced are discussed. Possibly the most important aspect is the stratification and primitive clear cells found in rat prostatic acini in cadmium injected rats.

Histological and ultrastructural changes in rat kidney following cadmium injection.

Cadmium injection in rats affects the genito-urinary system. The renal effects are generally considered to be mainly tubular. Light microscopy of 29 specimens confirms cellular damage in the glomeruli with changes in glomerular capillaries. Electron microscopic changes in the glomerulari confirm changes in the podocytes, thickening of the basement lamina, the occurrence of electron dense deposits in association with the capillary endothelium with platelet and polymorphonuclear cell accumulations which suggests opening of the endothelial junction of the glomerular vessels.

The distribution and retention of cadmium 115m in the rat following injection into the prostate.

The gonadal effects produced by injection of cadmium in the rats are well known. The effects of direct injection into the prostate are less well understood. The present experiments indicated the retention of cadmium by the various lobes of the rat prostate following direct injection of radioactive cadmium into the organ. The cells of the prostate are capable of retaining cadmium in measurable amounts up to at least 6 weeks following the injection. The subcellular fractions were measured with respect to cadmium activity and these are indicated. In addition measurements were made in organs which are known to be affected by cadmium, that is, kidney and the testicle. The affinity of the kidney for cadmium is confirmed. The areas of the prostate gland with a high zinc content appear to retain less cadmium than areas of the gland which are known to have a low zinc content. This suggests a possible zinc protective mechanism.