Morphologic alterations of the endometrial surface of rats induced by ethinyl estradiol, lynestrenol and lyndiol

의학과/박사[한글] 인구의 증가가 세계적인 심각한 문제로 대두됨에 따라 여러가지의 피임방법이 개발되게 되었고 이들중 경구피임제는 사용이 간편하고 효과가 정확하므로 널리 사용하게 되었다. 경구피임제의 주 작용은 난소에서의 배란을 억제하는 것이지만 이외에도 자궁경관, 자궁내막 및 수란관 등에 특이하게 작용함으로서 피임효과를 가져오게 된다(Vandenberg et al., 1974;Perez-Lopez et al., 1975: Mishell et al., 1977; Moghissi, 1980). 수정란이 자궁내막에 착상하여 자라기 위하여는 난소에서 분비되는 호르몬들이 적당량으로 적당한 시기에 자궁내막을 자극하여 형태학적 및 기능적 변화를 가져오게 된다. 이러한 현상중 자궁내막의 선상피 및 기질의 형태학적 변화에 관한 연구는 많이 되어 있으나, 수정란이 처음으로 접촉하여 착상을 시작하는 단계에 관여하는 자궁내막 표층상피세포의 변화에 대한 연구는 드문 실정이다. 이에 저자는 난소를 제거하지 않은 상태의 백서와 난소를 제거한 상태하의 백서에 경구피임제인 Lyndiol의 주성분인 ethinyl estradiol 및 Iynestrenol을 단독 또는 복합으로 투여하여 초래되는 백서자궁내막 표층상피세포의 변화를 광학현미경, 투시 및 주사전자현 미경적으로 관찰하여 다음과 같은 결과를 얻었다. 1. 정상 자성백서의 성 주기에 따른 자궁내막표층의 광학현미경적 특징은 혈중 estrogen농도가 높은 proestrous 및 estrous기에는 내막 표면의 굴곡이 심하고, 표층 상피세포는 높고 세포질 내에 다수의 대소부동한 공포형성을 보였으며 estrogen양이 낮은 metestrou s 맞 diestrous기에 있어서는 내막표면이 평활하고 상피세포가 낮으며 공포형성을 볼 수 없었다. 난소 존재하에 estradiol을 단독 투여한군에 있어서는 자궁 내막표면의 굴곡이 proestrous기보다 현저하여 졌으며 상피세포의 모양도 길어졌고 다수의 대소부동한 공포형성을 보임으로써 proestrous기의 소견이 증가된 양상을 보였다. 이에 비하여 Iynestrenol의 투여는 자궁 내막표면을 평활하게 하였으며 표층의 상피세포의 키가 낮고 공포형성은 없었으나 간혹 소낭포형성을 보였다. 한편 두약물의 복합투여는 주로 estradiol투여로 인한 변화를 보였다. 2. 투시전자현미경 소견상 정상 자성백서의 성주기에 따른 자궁내막 표층상피세포는 혈중 estrogen 농도가 높은 proestrous 및 estrous기에 있어서 키가 높고 핵이 불규칙하며 세포질내에 각종 미세구조의 발달이 현저하고 특히 RER이 심한 확장을 일으켜 매우 활동적인 양상을 보였으며 표면에 다수의 조밀하고 긴 융모가 잘 발달되어 있었으며 estrogen농도가 낮은 metestrous 및 diestroos기에서는 키가 낮고 핵은 비교적 평활하며 세포질내 에 미세구조의 발달이 미약하여 휴식상태의 소견을 보였긴 표면의 융모가 짧고 감소된 양상을 보였다. 난소 존재하에 estradiol을 단독 투여한 군에서는 proestrous기에 있어서의 소견이 보다 증강되었고 Iynestrenol 투여군에서는 metestrous 또는 diestrous기의 소견을 보였으며 복합하여 투여한 군에서는 광학현미경 소견에서와 같이 estradiol의 영향이 보다 현저 하게 나타났다. 3. 주사 전자현미경 소견상 정상 자성백서 자궁내막의 표층은 proestrous 및 estrous기에 있어서 융모가 매우 잘 발달되어 길고 조밀한 분포를 보였으며 세포윤곽은 다각형이고 세포간격은 벌어져 있었다. 이에 비하여 metestrous 및 diestrous기에 있어서는 융모의 발달이 적으며 짧고 불규칙한 분포를 보였으며 세포윤곽은 원형내지 타원형이고 세포간격이 밀착된 상태를 보였다. 난소존재하에 estradiol의 투여는 proestrous기의 소견이 더욱 증강된 양상을 보였으며 Iynestrenol의 단독투여는 diestrous기의 소견을 보였고 복합제 투여군에서는 estradiol의 영향이 보다 더 현저히 나타났다. 4. 난소적출은 광학현미경, 투시 및 주사전자현미경적 소견상 위축상을 보였고 표면상피세포는 dieetrous기의 소견을 보였다. 난소적출후 estradiol투여군 에서는 proeatrous기의 소견을 보였고 IyneBtrenol투여에서는 표층상피세포가 다소 활동성을 보였으나 그 변화는 미약하였고 복합투여한군에서는 estradiol의 효과가 주로 나타났다. Morphologic Alterations of the Endometrial Surface of Rats Induced by Ethinyl estradiol, Lynestrenol and Lyndiol Young Jae Hong Department of Medical Science, The Graduate School, Yonsei University (Directors: Prof. Yoo Bock Lee, M.D. and Chung Sook Kim, M.D.) Population control is a world wide problem and various contraceptive measures are being sought to meet this problem, and the oral contraceptives have been proved to be a choice of methods among vaious contraceptive means. The principal action of the oral contraceptive is ovulation inhibition by depressing the secretion of gonadotropins from the anterior pituitary gland. However, the morphologic and the functional changes of the fallopian tubas, the uterine endometrium and the uterine cervix induced by oral contraceptives knave also teen considered as the possible contributors to the action of the drugs (Vandenberg et al., 1974: Perez-Lopez et al., 1975; Mishell et al., 1977; Moghissi, 1980). Implantation is an initial process for pregnancy. Therefore it has been regarded as a promising target for contraception, becaus it involves two quite independent and separately controlled targets, the uterus and the embryo (McLaren, 1978). The successful implantation of the blastocyst need optimal condition of the endometrium in which a high degree of structural changes are brought about by the influence of ovarian hormones. As might he suspected, the use of oral contraceptives containing synthetic or derivative of ovarian steroids induces a wide variety of endometrial changes and the most of the studies were focusd on the epithelium of the endometrial glands and stroma, although the luminal surface epithelium is also vital for implantation and pregnancy (Song et al., 1970; Ober, 1977). The Present investigation, therefore, is aimed at study on the morphologic alterations of the uterine luminal surface epithelium by light and electronmicroscopic methods following the administration of oral contraceptives. A total of 108 rats weighing about 200gm were usd and divided into three groups. The first was normal control group. The second was experimental group without oophorectomy. The third was experimental group after oophorectomy. The drubs were dissolved in the vegetable oil and given by intramuscular injection every day. Animals were sacrificed with ether anesthesia at the each please of sexual cycle in the normal control group, and 4th, 8th days after oophorectomy in the oophorectomized control group. The animals in the second group were sacrificed after 2, 4, 8, 12, 16, 20, 24, and 28 days of injection. In the third experimental group, the drug injections were started at the 4 th days after oophorectomy, and sacrificed at the 4th and 8th days after injection. Microsections were made from paraffin block and were stained with hematoxylin-eosin, periodic acid-Schiff and alcian blue. For transmission electron microscopic examination, a portion of endometrium was cut in 1 mm**3 size fixed in 1% osmium tetroxide, and embedded in Epon 812. Seotions were cue by grass knife, stained with uranyl acetate and lead citrate, and axamined with Hitachi H-500 transmission electron microscope. For scanning election microscopic examination, the mid portion of the uterus was excised and opened to expose the epithelial surface and fixed in 3% glutaraldehyde followed by emersion in 0.2 M sucrose solution in 0.1 M phosphate buffer at pH 7.2 for 24 hours to dissolve the mucus on the surface and post fixed 1% osmium tetroxide. The tissue was dried With the critical Point drier, coated with gold palladium and examined with Hitachi S-450 Scanning electron microcope at 10∼20 KV. Based on the results obtained, following conclusions are made. 1. Light microscopic characteristics of normal cyclic changes of rat's endometrial surface consisted of prominent infoldings lined by tall columnar epithelial cells containing many cytoplasmic vacuoles at proestrous and estrous phases when blood concentration of estrogen is most high, while flattening of surface lined by low columnar or cuboidal epithelial cells without vacuoles is noted at metestrous and diestrous phases when blood estrogen is the lowest. Administration of estradiol induced increase of surface foldings, epithelial height and cytoplasmic vacuoles indicating an accentuation of proestrous phase, while lynestrenol induced flattening of surface, descrease of epithelial cell height, and loss of vauoles, and combined administration of estradiol and lynestrenol accompanied predominantly estradiol effect. 2. Transmission electron microscopic features of endometrial surface epithelium at proestrous and estrous phases were characterized by tall columnar cells containing large irregular shaped nuclei, abundant cytoplasmic organelles an many dilated rough endoplasmic reticulum, and abundant regularly arranged Bong microvilli on surface, indicating active state of the cell, while at metestrous and diestrous phases epithelial cells were low cuboidal containing relatively small and round nucleus, and very poorly developed cytoplasmic organelles, with sparse and short inicrovilli on cell surface, indicating resting state of the cell. Administration of estradiol induced inerease of epithelial cells 7eight, more irregular nuclear outline, more pronounced dilatation of rough endoplasmic reticulum, and mere abundant and longer surfacs microvilli indicating an accentuation of proestrous phase, while administration of lynestrenol induced the features of metestrous and diestrous phases, and the combined administration of two drugs showed predominantly estradiol effect. 3. Scanning electron microscopic observations of endometrial surface showed polygonal outline of epithelial cells with loose intercellular border, and abundant amount of long and regularly arranged surface microvilli at proestrous and estrous phases, while round epithelial cell outline with tight cell herders, and short and sparse microvilli on the surface were characteristic features at metestrous and diestrous phases. Administration of estradiol induced an accentuated features of proestrous and estrous phases, while lynestrenol induced a picture of metestrous and diestrous phases, and combined administration induced predominantly estradiol effect. 4. Oophorectomy induced atrophy of the surface epithelial cells and exhibited features of diestrous phase in light microscapic, transmission and scanning electron microscopic findings. Administration of estradiol, lynestrenol, or combination after oophorectomy inducted features of proestrous, diestrous, and proestrous phases respectively. In summary, estradiol stimulates rat's endometrial surface epithelium to proliferate inducing surface foldings, active state of subcellular structures. and development. of surface microvilli, while lynestrenol induces atrophy and inactive resting state of surface epithelium, and a combined administration induces predominantly estradiol effect. [영문] Population control is a world wide problem and various contraceptive measures are being sought to meet this problem, and the oral contraceptives have been proved to be a choice of methods among vaious contraceptive means. The principal action of the oral contraceptive is ovulation inhibition by depressing the secretion of gonadotropins from the anterior pituitary gland. However, the morphologic and the functional changes of the fallopian tubas, the uterine endometrium and the uterine cervix induced by oral contraceptives knave also teen considered as the possible contributors to the action of the drugs (Vandenberg et al., 1974: Perez-Lopez et al., 1975; Mishell et al., 1977; Moghissi, 1980). Implantation is an initial process for pregnancy. Therefore it has been regarded as a promising target for contraception, becaus it involves two quite independent and separately controlled targets, the uterus and the embryo (McLaren, 1978). The successful implantation of the blastocyst need optimal condition of the endometrium in which a high degree of structural changes are brought about by the influence of ovarian hormones. As might he suspected, the use of oral contraceptives containing synthetic or derivative of ovarian steroids induces a wide variety of endometrial changes and the most of the studies were focusd on the epithelium of the endometrial glands and stroma, although the luminal surface epithelium is also vital for implantation and pregnancy (Song et al., 1970; Ober, 1977). The Present investigation, therefore, is aimed at study on the morphologic alterations of the uterine luminal surface epithelium by light and electronmicroscopic methods following the administration of oral contraceptives. A total of 108 rats weighing about 200gm were usd and divided into three groups. The first was normal control group. The second was experimental group without oophorectomy. The third was experimental group after oophorectomy. The drubs were dissolved in the vegetable oil and given by intramuscular injection every day. Animals were sacrificed with ether anesthesia at the each please of sexual cycle in the normal control group, and 4th, 8th days after oophorectomy in the oophorectomized control group. The animals in the second group were sacrificed after 2, 4, 8, 12, 16, 20, 24, and 28 days of injection. In the third experimental group, the drug injections were started at the 4 th days after oophorectomy, and sacrificed at the 4th and 8th days after injection. Microsections were made from paraffin block and were stained with hematoxylin-eosin, periodic acid-Schiff and alcian blue. For transmission electron microscopic examination, a portion of endometrium was cut in 1 mm**3 size fixed in 1% osmium tetroxide, and embedded in Epon 812. Seotions were cue by grass knife, stained with uranyl acetate and lead citrate, and axamined with Hitachi H-500 transmission electron microscope. For scanning election microscopic examination, the mid portion of the uterus was excised and opened to expose the epithelial surface and fixed in 3% glutaraldehyde followed by emersion in 0.2 M sucrose solution in 0.1 M phosphate buffer at pH 7.2 for 24 hours to dissolve the mucus on the surface and post fixed 1% osmium tetroxide. The tissue was dried With the critical Point drier, coated with gold palladium and examined with Hitachi S-450 Scanning electron microcope at 10∼20 KV. Based on the results obtained, following conclusions are made. 1. Light microscopic characteristics of normal cyclic changes of rat's endometrial surface consisted of prominent infoldings lined by tall columnar epithelial cells containing many cytoplasmic vacuoles at proestrous and estrous phases when blood concentration of estrogen is most high, while flattening of surface lined by low columnar or cuboidal epithelial cells without vacuoles is noted at metestrous and diestrous phases when blood estrogen is the lowest. Administration of estradiol induced increase of surface foldings, epithelial height and cytoplasmic vacuoles indicating an accentuation of proestrous phase, while lynestrenol induced flattening of surface, descrease of epithelial cell height, and loss of vauoles, and combined administration of estradiol and lynestrenol a companied predominantly estradiol effect. 2. Transmission electron microscopic features of endometrial surface epithelium at proestrous and estrous phases were characterized by tall columnar cells containing large irregular shaped nuclei, abundant cytoplasmic organelles an many dilated rough endoplasmic reticulum, and abundant regularly arranged Bong microvilli on surface, indicating active state of the cell, while at metestrous and diestrous phases epithelial cells were low cuboidal containing relatively small and round nucleus, and very poorly developed cytoplasmic organelles, with sparse and short inicrovilli on cell surface, indicating resting state of the cell. Administration of estradiol induced inerease of epithelial cells 7eight, more irregular nuclear outline, more pronounced dilatation of rough endoplasmic reticulum, and mere abundant and longer surfacs microvilli indicating an accentuation of proestrous phase, while administration of lynestrenol induced the features of metestrous and diestrous phases, and the combined administration of two drugs showed predominantly estradiol effect. 3. Scanning electron microscopic observations of endometrial surface showed polygonal outline of epithelial cells with loose intercellular border, and abundant amount of long and regularly arranged surface microvilli at proestrous and estrous phases, while round epithelial cell outline with tight cell herders, and short and sparse microvilli on the surface were characteristic features at metestrous and diestrous phases. Administration of estradiol induced an accentuated features of proestrous and estrous phases, while lynestrenol induced a picture of metestrous and diestrous phases, and combined administration induced predominantly estradiol effect. 4. Oophorectomy induced atrophy of the surface epithelial cells and exhibited features of diestrous phase in light microscapic, transmission and scanning electron microscopic findings. Administration of estradiol, lynestrenol, or combination after oophorectomy inducted features of proestrous, diestrous, and proestrous phases respectively. In summary, estradiol stimulates rat's endometrial surface epithelium to proliferate inducing surface foldings, active state of subcellular structures. and development. of surface microvilli, while lynestrenol induces atrophy and inactive resting state of surface epithelium, and a combined administration induces predominantly estradiol effect.restrictio