Histological changes in the oviduct of rabbits after administration of cadmium

The effects of cadmium on the oviduct of 32 rabbits were studied by light microscopy with morphometry. Cadmium was administered intraperitoneally (i.p.) to group A in a single dose of 1.5 mg/kg body weight (BW), per oss (p.o.) to group B in daily doses of 1.0 mg/kg BW for a 5-month period, and also p.o., 1.0 mg/kg BW; for a 5-month period daily followed by a 7-month interval in group C. All experimental groups were compared to the control group (K), as well as to other experimental groups. Animals were examined directly after the experimental period. The amount of epithelium in the oviduct ranged from 19.67 to 25.50%, with no significant differences. Significantly higher amounts of stroma were found in group A in comparison with group C. In the stroma, dilatation of blood vessels was evident with subsequent oedematization and diapedesis. No significant differences were found in the muscular layer.


INTRODUCTION
Cadmium is chemically similar to zinc and occurs naturally with zinc and lead in sulphide ores.Some cadmium has been found in all natural materials that have been analyzed.High concentrations in air, water and soil, however, are commonly associated with industrial emission sources, particularly with non-ferrous mining and metal refining (Friberg et al., 1986).
Cd AFFECTS CHANGES IN RABBIT'S OVIDUCT Generally, it is well known that cadmium accumulates very intensively in the liver and kidneys (Renon et al, 1980;Friberg et al., 1986;Bires et al., 1990;Massanyi et al., 1995a;Nad et al., 1995;Sokol et al., 1998).Cadmium alters the growth of experimental animals (Massanyi and Uhrin, 1996a), has effects on the growth of offspring (Massanyi et al., 1995a), effects on the blood serum metabolic profile (Massanyi et al., 1995d,e), and is responsible for changes in ovarian structure (Massanyi and Uhrin, 1996b).Klaassen and Kotsonis (1977) found, in rats, a very high relationship between the concentration of cadmium in the liver and the administered dose.The highest levels were reported in the kidneys and in the liver.In our previous study (Massanyi et al., 1995a), we reported that the concentration of cadmium in rabbits not treated with cadmium was 0.138 mg/kg in liver, 0.458 mg/kg in kidneys, 0.033 mg/kg in the ovaries, and 0.040 mg/kg wet tissue in the uterus.Bires et al. (1991) have reported that the cadmium level in ewe ovaries is 1.6 ±0.36 mg/kg.Cadmium affects reproduction.In males, cadmium produces testicular injury (Kotsonis and Klaassen, 1977).Exposure to cadmium causes vascular damage and interstitial cell oedema, leading to hemorrhage, reduced androgen production by Leydig cells, necrosis of the Sertoli cells, inhibition of spermatogenesis, and eventually testicular atrophy (Gunn et al., 1963;Toman and Massanyi, 1997).Also motility of bovine spermatozoa is inhibited by cadmium chloride (Massanyi et al., 1996).In females, it affects the ovaries (follicular atresia, interstitial oedema, altered hormone production) as well as the uterus (interstitial oedema, capillary modification) (Peereboom-Stegeman et al., 1987;Rehm and Waalkes, 1988;Paksy et al., 1992;Massanyi andUhrin, 1996b, 1997;Massanyi et al., 1997).The histological changes and structure of the oviduct and ovary are well described (Bires et al., 1995;Kolodzieyski and Danko, 1995;Cigankovaet al., 1996;Jantosovicova et al., 1996;Danko, 1997).
The purpose of this study was to analyze oviduct structure after experimental administration of cadmium as there are no previous reports on changes in the oviduct.Our previous studies have found the highest cadmium accumulation in the oviduct in comparison with other reproductive organs (Massanyi et al., 1995b,c).

MATERIAL AND METHODS
All experiments were conducted on rabbits (Hyla, VUZV Nitra), an animal with spontaneous ovulation, housed individually in cages.The animals (32) were divided into four groups (K, A, B, C).Eight rabbits received cadmium i.p. (1.5 mg/kg BW) in a single dose.These animals (group A) were killed 48 h after administration of cadmium (CdCl2, Sigma, St. Louis, MO, USA).A continuous experiment (group B) was carried out on the same number (8) of animals.Cadmium was administered daily at a dose of 1.0 mg/kg BW for 5 months in pelletized food (PD Cataj, Slovak Republic).After this period the animals were sacrificed.In group C, eight female rabbits received a cadmium dose of 1.0 mg/kg BW for 5 months daily, then administration was stopped for 7 months.To obtain histological samples, all animals in group C were killed after the experiment.Food and water were available for all animals ad libitum.The administered cadmium was diluted with a physiological solution to the appropriate concentration.The last group (K) was the control, and received no cadmium.
Oviducts (mid part of the ampulla) of the cadmium-treated and untreated rabbits were fixed in 10% formol (48 h), dehydrated in a graded series of ethanol and embedded in paraffin.Oviducts were sectioned by a microtome.Serial sections (5 from each animal) 10 |j,m thick were stained with haematoxylin and eosin.From photographs (Docuval, Carl Zeiss Jena) based on micromorphological criteria (Weibel et al., 1966;Uhrin, 1992) the percentage of epithelium, stroma, and muscular layer were evaluated with respect to each oviduct.
The final data was subjected to basic statistical analysis (mean, SD), and to analysis of variance by Sheffe's test for each variable.

RESULTS
In evaluating the percentage of epithelium, stroma, and muscular layer in oviducts (Table 1), we found that the highest amount of epithelium was in the group receiving cadmium continuously (25.5%).In the other groups the percentage was very similar (19.67-23.13%).No significant differences were determined.In comparing the stroma, we report a significantly lower amount of this parameter in the group with a 5 month daily p.o. administration with later cessation of administration in comparison with i.p. application.This result describes oedematization of the oviduct tissue, caused by disintegration of the capillary wall, after a high level of cadmium intake.Dense material on the top of well-vascularized papillary mucosa is evident after cadmium administration (Figures 1 and 2).On the other hand, we also found a relatively lower amount of stroma in group K and B compared to group A, but this decrease was not significant.The amount of muscular layer was, in all studied groups, in the range of 52.94 to 63.55% with no significant difference.

DISCUSSION
The biological effects of cadmium deserve attention for many reasons.Its use in different industrial processes and products has increased its presence in the environment.
Cadmium exposure in our experiment resulted in changes in oviduct structure, mainly the stroma and epithelium, with changes in the capillary wall structure.A similar result was found in the endometrium of the uterus where the percentage of stroma was significantly higher after i.p. administration of cadmium (Massanyi and Uhrin, 1997).Oedematization of the uterus with disintegration in the blood vessels and subsequent diapedesis were evident.
There are no previous reports about structural changes in the oviduct after administration of cadmium.It is believed that cadmium bound to carrier proteins is able to enter the oviduct lumen only in small quantities (Paksy et al., 1992).Moreover, due to its poor blood supply, the oviduct contains relatively lower levels of cadmium compared to the corpora lutea in pseudopregnant rats treated with cadmium at a dose of 3.5 mg/kg (Paksy et al., 1990).Possible indirect influence of cadmium on oviduct function is perhaps caused by a dose-dependent drop in ovarian secretion of progesterone (Paksy et al., 1992).
Changes in oviduct structure are caused by changes in the blood vessel wall, and subsequent diapedesis.Similar alterations are reported in the uterus (Massanyi and Uhrin, 1997).Peereboom-Stegeman et al. (1987) have studied the small vessels in the myometric layer of the uteri in rats exposed subcutaneously to the cadmium.The thickness of the media was analyzed and an inventory was made of the morphology of the media, the endothelial layer, and the perivascular connective tissue.A dose-and time-related increase of the thickness of the media was demonstrated.At the higher dose, signs of a perivascular inflammatory reaction were observed.From these, as well as from our data it is evident that cadmium causes disorders in the blood vessels wall with significant oedematization.

Figure 1 .
Figure 1.Histological structure of the oviduct of control.In the oviductal wall surface epithelium (a) is present, under which lays lamina propria mucosae (b) formed by connective tissue and muscular layer (c) (H-E, 200x)