Histopathological changes in ewe lambs exposed to prolonged diet on lucerne

Transcription

1 Histopathological changes in ewe lambs exposed to prolonged diet on lucerne M Barberan, J Valderrabano, Ja Bascuas To cite this version: M Barberan, J Valderrabano, Ja Bascuas. Histopathological changes in ewe lambs exposed to prolonged diet on lucerne. Annales de Recherches Vétérinaires, INRA Editions, 1990, 21 (2), pp <hal > HAL Id: hal Submitted on 1 Jan 1990 HAL is a multi-disciplinary open access archive for the deposit and dissemination of scientific research documents, whether they are published or not. The documents may come from teaching and research institutions in France or abroad, or from public or private research centers. L archive ouverte pluridisciplinaire HAL, est destinée au dépôt et à la diffusion de documents scientifiques de niveau recherche, publiés ou non, émanant des établissements d enseignement et de recherche français ou étrangers, des laboratoires publics ou privés.

2 Short note Histopathological changes in ewe lambs exposed to prolonged diet on lucerne M Barberan J Valderrabano JA Bascuas! Departamento Patologia Animal, Facultad de Veterinaria, Miguel Servet 177, Zaragoza; 2 Servicio de Investigacion Agraria, DGA Apdo 727, Zaragoza, Spain (Received 23 March 1989; accepted 24 October 1989) Summary &horbar; Histopathological changes were observed in 10 ewe lambs reared during nine months on lucerne as compared with 10 controls reared on ryegrass. These changes consisted of periglandular oedema and subacute endometritis in the uterus, secretory-cell hyperplasia with prevalence of neutral mucopolysaccharides in the cervix and hyperkeratinisation and colpitis in the vagina. The mammary glands of lambs reared on lucerne were significantly heavier and showed secretory activity. lucerne / ewe lamb / phyto-oestrogen / oestrogenic diet / reproductive system Résumé &horbar; Modifications histopathologiques chez des agnelles nourries à la luzerne. Des modifications histopathologiques ont été étudiées sur 2 groupes de 10 agnelles nourries pendant 9 mois soit à la luzerne, soit à l herbe (groupe témoin). Chez les agnelles nourries à la luzerne ont été observés un oedème périglandulaire et une endométrite subaiguë de l utérus, une hyperplasie des cellules secrétoires avec prédominance des mucopolysaccharides neutres dans le col utérin, une vaginite et une hyperkératose du vagin. Les mammelles des mêmes agnelles étaient significativement plus lourdes et présentaient une activité sécrétoire. luzerne / agnelle / phyto-oestrogène / diète!strogénique / appareil génital INTRODUCTION Ingestion of highly oestrogenic clovers may cause alterations in the female reproductive system that may lead to temporary or permanent infertility in the ewe (Rizzoli and Moran, 1977). Adams (1977) and Heydon and Adams (1977) have studied the pathological effects of some highly oestrogenic clovers in ewes, but little information is available on the morphological changes that other feedstuffs, containing moderate amounts of phyto-oestrogens, may cause in the organs of ewes after being consumed for long periods of time, as can be the case for lucerne, widely used under mediterranean conditions. The present paper describes the morphological changes in the genital tract, ovaries, adrenal, thyroid and mammary * Correspondence and reprints

3 glands of ewes exposed to prolonged grazing on lucerne. MATERIALS AND METHODS Twenty Fleischschaf x (Romanov x Rasa Aragonesa) ewe lambs aged 3 months at the beginning of the experiment were randomly distributed to 2 equal groups. Ten animals were reared on lucerne containing moderate amounts of isoflavones (68 mg/kg DM in autumn, 98 mg/kg DM in winter and 127 mg/kg DM in spring) and 10 on italian ryegrass containing only traces of isoflavones. All ewes were slaughtered at 12 months of age. Both groups of animals were grazing in spring and autumn, but were kept indoors and given ad libitum either lucerne or Italian ryegrass hay in winter. All the animals received anthelmintic treatment at regular intervals and had free access to water and mineral blocks. Acetate fluorogestone intravaginal sponges were applied to synchronize the reproductive cycle of the ewes. The ewes were slaughtered 48 h after removal of vaginal sponges, at the time of oestrus. The genital tract, ovaries, adrenal, thyroid and mammary glands were removed, weighed and measured. Tissue samples of all the organs obtained were fixed in 10% formol saline solution, routinely processed and stained with haematoxylin and eosin (HE). Mucins from the re- tract were studied histochemically productive using periodic acid-schiff (PAS) and Alcian blue, ph 2.5 (AB). Staining was carried out as described by McManus and Mowry (1968). Comparisons between treatments at slaugh - ter were carried out by ttests. RESULTS The mean growth rate in both groups of ewe lambs was 90 g/day, and their final weight was not significantly affected by treatment. Ewes fed with lucerne presented greater mean weights in the genital tract, adrenal, thyroid and mammary glands than ewes fed with ryegrass, although the differences were only significant for the mammary gland (P < 0.001) and thyroid (P < 0.05). The genital tract of ewes fed with lucerne was significantly longer (18.1 cm vs 16.5 cm; P < 0.05) than that of ewes fed with ryegrass (fig 1 but no changes were observed in the mucosa, and there were no exudates inside the genital tract. The ovaries of ewe lambs fed on lucerne were significantly smaller (P < 0.05) than those of ewes fed on ryegrass. The average number of growing and atretic follicles was slightly lower in ewes fed with lucerne (4.9 vs 7.0 and 11.7 vs 14.6). However the average number of corpora lutea was slightly higher (1.0 vs 0.4) in this group of ewes. Numerous intraepithelial cysts lined with a cuboid or squamous epithelium occurred throughout the oviduct mucosa in both groups of ewes. Furthermore, 2 ewes from each group presented mild salpingitis consisting of vascular congestion, lymphocytic infiltration of the lamina propia, and slight epithelial desquamation. The uterus of ewes reared with lucerne presented extensive periglandular and myometrial oedema and focal subacute endometritis with neutrophilic and lymphocytic infiltration of the lamina propia and intraepithelial necrotic debris. The cervical epithelium in both groups of animals was columnar in the rostral cervix, squamous in the caudal cervix and of both types in the mid-cervix (columnar in the villi and squamous in the crypts). However, secretory cells were more numerous in ewes fed with lucerne and were located preferentially in the mid-cervix, lining the villi as well as forming one or more layers over the squamous epithelium of the

4 crypts. The cervical secretory cells were PAS-AB positive, with prevalence of neutral mucopolysaccharides PAS positive) in ewes fed with lucerne. One ewe fed with lucerne presented extensive plasmocytic and lymphocytic infiltrates in the lamina propia of the caudal cervix, necrosis and desquamation of lining epithelium and flattening of longitudinal villi. The vaginal epithelium was of similar thickness in both groups of ewes; nevertheless; keratinisation and keratohyaline synthesis in parabasal layers was greater in ewes fed with lucerne. Four animals in this group presented a colpitis, characterised by the presence of round cells and neutrophils in the lamina propia and epithelium (fig 2). Due to an increase in the secretory tissue, the mammary gland of ewes fed with lucerne presented a lower proportion of connective and adipose tissues (fig 4) than that of ewes fed with ryegrass (fig 3). Mammary glands from ewes fed with lucerne had ducts filled with PAS-positive material and obvious development of alveoli, both signs of secretory activity. The adrenal glands of ewes fed with lucerne presented slight cortical hyperplasia and foci of dense atrophic cells occurred in the 3 zones of the cortex. Thyroids of both ewe groups presented a mild form of parenchymatous goitre. Histologically the follicular epithelium was taller and &dquo;resorption vacuoles&dquo; were seen in the colloid.

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6 DISCUSSION The occurrence of intra-epithelial cysts in the oviduct of both affected and control ewes was also noted by Adams (1976) in a survey with oestrogenic clover, but their effect on reproduction should be studied in more detail. The uterine lesions observed in ewes fed on lucerne are similar, though less intense to those produced by oestrogenic clover (Adams, 1976, 1977; Rizzoli and Moran, 1977), but may produce reproductive deficiencies by restricting nesting (Adams, 1976). The secretory-cell hyperplasia observed in the cervical epithelium of ewes fed with lucerne has also been noted in ewes exposed to highly oestrogenic diets (Lightfoot and Adams, 1979). The prevalence of neutral mucopolysaccharides in the chemical composition of cervical mucus observed in ewes fed with lucerne suggests a decrease in the amount of acidic radicals, a fact that has been related to a decrease in mucus viscosity (Hamana et al, 1971); these conditions are known to impair sperm transport through the cervix, with the subsequent failure of fertilisation (Adams, 1976). Mammary development and lactation is a constant finding in non-pregnant ewes exposed to oestrogenic products (Adams, 1976, 1977; Jubb et al, 1985). The changes in the adrenal glands of ewes fed with lucerne are similar to those

7 noted by Adams and Nairn (1983) in clover diseased ewes. The presence of atrophic cells in the adrenal cortex of ewes exposed to oestrogenic diets was considered by Gardiner and Nairn (1969) as evidence of stress. The mild form of parenchymatous goitre observed in the thyroids of both groups of ewes should not be considered significant as the experiment was carried out in an iodine-deficient region. The larger amount of colloid contained in the follicles of ewes fed on ryegrass may explain the presence of significantly heavier thyroid glands. Results obtained suggest that the intake of lucerne containing moderate amounts of phyto-oestrogens for long periods of time may give rise to similar but less severe morphological alterations in to those ewe lamb reproductive organs produced by oestrogenic clover, which could alter their reproductive performance. Further studies are therefore necessary to determine how far these morphological alterations resulting from phyto-oestrogen intake could affect their future reproductive lives. ACKNOWLEDGMENT This work was supported by CICYT project No PA $ REFERENCES Adams NR (1976) Pathological changes in the tissues of infertile ewes with clover disease. J Comp Pathol86, Adams NR (1977) Morphological changes in the organs of ewes grazing oestrogenic subterranean clover. Res Vet Sci 22, Adams NR, Nairn ME (1983) The nature of dystokia in ewes after grazing oestrogenic subterranean clover. Aust Vet J 60, Gardiner MR, Nairn ME (1969) Studies on the effect of cobalt and selenium in clover disease of ewes. Aust Vet J 45, Hamana K, El-Banna AA, Hafez ESE (1971) Sialic acid and some physicochemical characteristics of bovine cervical mucus. Cornell Vet 61, Heydon RA, Adams NR (1977) Histochemical studies on cervical glands in ewes with clover disease. J Comp Pathol 87, Jubb KVF, Kennedy PC, Palmer N (1985) Pathology of Domestic Animals. Academic Press, London, 328 Lightfoot RJ, Adams NR (1979) Changes in cervical histology in ewes following prolonged grazing on oestrogenic subterranean clover. J Comp Pathol89, McManus JFA, Mowry RW (1968) Tecnica Histol6gica. Atika SA, Madrid, Rizzoli DJ, Moran AR (1977) Permanent clover infertility in ewes. Aust Vet J 53,