OBJECTIVE: This study was to investigate the expression of CDK inhibitors, p27(kip1) and p57(kip2) in mouse endometrium during the estrus cycle and pregnant period. METHODS: Total RNA and protein were extracted from endometrium of mouse sacrificed at diestrus, proestrus, estrus, and metestrus cycle, and at day 1-6 post-coitum (p.c.), then semi-quantitative RT-PCR and western blotting of p27(kip1) and p57(kip2) was carried out. RESULTS: p27(kip1) and p57(kip2) mRNA was highly expressed in diestrus and proestrus stage than estrus and metestrus stage. In comparison with estrus cycle, p27(kip1) and p57(kip2) mRNA level was highly maintained in gestational endometrium (except p27(kip1) of day 5 p.c). p57(kip2) protein level was relatively low from day 1 p.c. to day 4 p.c. But it was significantly increased in day 5 p.c. and day 6 p.c. CONCLUSION: These results show that p27(kip1) and p57(kip2) may play a role in endometrial differentiation for regular estrus cycle and implantation, and especially p57(kip2) may play an essential role in endometrial differentiation for maintenance of implantation.
Animals ; Blotting, Western ; Diestrus ; Endometrium* ; Estrus ; Female ; Metestrus ; Mice* ; Proestrus ; RNA ; RNA, Messenger

OBJECTIVE: This study was to investigate the localization of CDK inhibitor, p57(kip2) in mouse endometrium during the estrus cycle and pre- and peri-implantation periods. METHODS: The p57(kip2) protein was immunostained from endometrium of mouse sacrificed at diestrus, proestrus, estrus, and metestrus cycle, and at day 1-6 post-coitum (p.c.). RESULTS: The staining in the luminal epithelium was very weak in comparison with glandular and stromal cells. In diestrus stage, immunoreactivity of p57(kip2) was heterogeneously strong in parts of decidualized or degenerated stromal cells. In proestrus stage, strong immunoreactivity p57(kip2) was largely found in stromal cells. But, p57(kip2) was showed low immunoreactivity in estrus stage. In metestrus stage, immunoreactivity of p57(kip2) was heterogeneously strong in decidualized stromal cells. In day 1-2 p.c., immunoreactivity of p57(kip2) was low in some endometrial stromal cells. In day 3-4 p.c., immunoreactivity of p57(kip2) was strong in some endometrial stromal cells. In day 5-6 p.c., immunoreactivity of p57(kip2) was strong in decidual cells. CONCLUSION: These suggest that p57(kip2) may play an essential role in endometrial differentiation for maintenance of implantation, especially decidualization of endometrial stromal cells.
Animals ; Diestrus ; Endometrium* ; Epithelium ; Estrus* ; Female ; Metestrus ; Mice* ; Phenobarbital ; Proestrus ; Stromal Cells

The objective of this experiment was to assess the relationship between electrical resistance of the vaginal mucosa and plasma progesterone for optimal mating time in the bitch. Eight mature beagle bitches were examined, and we observed eight times of estrus. Vaginal electric resistance was recorded weekly using a Draminski ovulation detector in anestrus, and daily in estrus. Plasma progesterone concentration was estimated by radioimmunoassay. In the bitch, incline in vaginal electric resistance (376.20 +/- 105.63 units) showed a closely association with the onset of proestrus. Ovulation day was determined as the first day when plasma progesterone concentration increased above 5.0 ng/ml (Day 0). On Day 0, vaginal mucous electric resistance was 438 +/- 132 units. Vaginal mucous electric resistance showed a slight decrease or was maintained until Day 0. However, it showed an explosive increase, and peaked on Day 1~3, which was above 600 units. Two of eight cases peaked on Day 1, three of eight cases were revealed on Day 2, and others were revealed on Day 3. After Day 4, resistance showed a rapid drop to below 600 units and reached 200 units on Day 8. The optimal mating time was determined when vaginal mucous electric resistance was above 600 units.
Anestrus ; Electric Impedance ; Estrus ; Female ; Mucous Membrane ; Ovulation ; Plasma ; Proestrus ; Progesterone ; Radioimmunoassay

Estrous cycle -related histological and histochemical changes in the vaginal epithelium of mature female rats were studied with PAS (periodic acid Schiff) alcian blue pH 2.5 and biotinylated lectins (DBA, SBA, PNA, BSL -1, sWGA, UEA -1, RCA -1, Con A and LCA).The prominent characteristic changes that occured during the estrous cycle were mucinous transformation in proestrus and cornification in estrus. In proestrus, the superficial mucinous cells of the epithelium were increased in number and enlarged in size, and the amount of acid and neutral mucosubstances was more increase in proestrus than in diestrus and metestrus. About the binding pattern of all lectins examined to the superficial mucinous cells, in diestrus, the binding pattern of these cells showed a similar affinity as in metestrus with intense DBA and UEA -1 reactivity. In proestrus, however, these cells were reactive with seven lectins examined except LCA and PNA, and DBA, SBA, BSL -1, RCA -1 and UEA -1 reacted more strongly than in diestrus and metestrus. In estrus, the superficial cornified cell layers showed a weak reactivity of SBA, BSL -1 and PNA. In diestrus and metestrus, the mucinous cells in the intermediate layers of the basal portion of vaginal fold stained with eight lectins examined except LCA and showed the same binding pattern to the superficial mucinous cells. About the distribution of glycoconjugates in the intermediate layer, the upper spindle cells showed different binding pattern according to the estrous stages. In diestrus, estrus, and metestrus, these cells showed a affinity for all lectins examined. In proestrus, however, DBA and PNA staining were not observed, and stained more intensely with sWGA, SBA and UEA -1, and less intensely with BSL -1 and RCA - 1. In estrus, DBA and PNA reactivity reappeared as trace, and RCA -1 and sWGA reactivity increased. In metestrus, sWGA reactivity reduced and BSL -1 and UEA -1 increased continually. The lower rounded cells of the intermediate layers stained with all lectins examined in estrus, with six lectins examined except Con A, DBA and UEA -1 in proestrus and with five lectins examined except DBA, UEA -1, sWGA and BSL -1 in diestrus and metestrus. BSL -1 reactivity for the layers increased in proestrus, estrus and metestrus, and PNA reactivity increased in estrus and reduced in metestrus. The basal layer of the vaginal epithelium showed different binding pattern to the different portion of vagina, and showed faint staining of BSL -1, SBA and RCA -1, and moderately staining of BSL -1 in proestrus and estrus. In conclusion, alpha /-N -acetyl -D -galactosamine, alpha /-D -galactose and alpha -L -fucose participate in the mucinous transformation of the vaginal epithelium, and beta -N -acetyl -D -glucosamine participates in the cornification of the vaginal epithelium.
Alcian Blue ; Animals ; Diestrus ; Epithelium* ; Estrous Cycle* ; Estrus ; Female ; Glycoconjugates ; Humans ; Hydrogen-Ion Concentration ; Lectins ; Metestrus ; Mucins ; Proestrus ; Rats* ; Vagina

The estrogen receptor (ER) is present in a wide variety of mammalian tissues and is required for the physiological responses of estrogen, including estrogen-induced tissue-specific changes in gene expression. But most of our knowledge on the regulation of ER mRNA levels comes from in vivo steroid replacement experiments or cancer cell lines that express the ER. Thus the present study was attempted to determine 1) the anterior pituitary ER mRNA levels during rat estrous cycle 2) if estradiol itself directly modulates the ER mRNA levels in cultured rat anterior pituitary using RT-PCR method. In rats with 4 day estrous cycle, the ER mRNA levels in anterior pituitary gland reached to maximum at proestrus 11:00h just before serum estradiol concentration showed the highest. From then, the ER mRNA levels gradually declined during the rest of the proestrus. On the other hands, in cultured rat anterior pituitary cells, the ER mRNA levels were significantly decreased by the treatment of estradiol. These results indicate that the surge of estradiol was proceeded by the increase in pituitary ER mRNA levels during the proestrus and in cultured anterior pituitary cells, estrogen might be involved in the down-regulation of the ER mRNA levels.
Animals ; Cell Line ; Down-Regulation ; Estradiol ; Estrogens* ; Estrous Cycle ; Gene Expression ; Hand ; Pituitary Gland, Anterior* ; Proestrus ; Rats* ; RNA, Messenger*

The objective of the present study was to monitor and characterize morphological alterations in ovaries of agouti (Dasyprocta prymnolopha), reared in captivity, by using abdominal ultrasonography. All animals underwent daily vaginal cytological examination to identify the current cycle phase. For each phase of the estrous cycle, ultrasound examinations were carried out to identify and describe the morphology of both ovaries. Topographic parameters in an ultrasound window were established to locate the ovaries. The agouti estrous cycle lasted an average of 29.94 ± 6.77 days. During vaginal cytology examinations, all cell types were identified, and each phase of the estrous cycle was established by cell counts. No significant alterations were observed in the assessed ovarian morphometry measurements. In 75% of the animals examined, ovarian follicle presence was observed in the proestrus phase.
Animals ; Cell Count ; Dasyproctidae ; Epithelium ; Estrous Cycle ; Female ; Gonads ; Investigative Techniques ; Ovarian Follicle ; Ovary ; Proestrus ; Reproduction ; Ultrasonography

OBJECTIVE: Bok, Bcl-2-related ovarian killer, is a proapoptotic Bcl-2 family protein identified in the ovary based on its dimerization with the antiapoptotic protein Mcl-1. The present study examined the hormonal regulation and localization of Bok messenger RNA levels in the rat ovary during the follicle development. METHODS: We used the female rats of SD strain. Bok mRNA levels in the ovary were determined by Northern blot analysis. In situ hybridization were performed to determine the specific ovarian cell type expressing Bok mRNA. RESULTS: Northern blot analysis of ovaries obtained from immature rats revealed increasing levels of Bok mRNA during postnatal development. The major cell types expressing Bok mRNA were the granulosa cells of preantral and atretic follicles. Treatment of immature rats with diethylstilbestrol (DES) for 24-48 h increased ovarian Bok mRNA levels. Bok mRNA was remained the same levels in rats removed DES for 24- 48 h to induce apoptosis. High signals of Bok mRNA after DES treatment were detected in granulosa cells of early antral follicles. Treatment of immature rats with pregnant mare's serum gonadotropin (PMSG) for 12 h increased markedly ovarian Bok mRNA expression which was detected mainly in preantral and atretic follicles. Interestingly, low levels of Bok mRNA were also expressed in granulosa cells of preovulatory follicles. Treatment of PMSG-primed rats with human chorionic gonadotropin (hCG) stimulated strongly ovarian Bok mRNA expression at 6-9 h. At that time, Bok mRNA was expressed in granulosa cells of atretic and small growing follicles. In adult estrus cyclic ovaries, Bok gene expression was higher on proestrus and estrus than metaestrus and diestrus. Moreover, the highly increased expression of Bok mRNA was found in rat ovaries at 48 h after hypophysectomy. CONCLUSION: These results demonstrate that Bok is one of proapoptotic Bcl-2 members expressed in early growing and atretic follicles during the ovarian follicular development. Gonadotropins induce a transient increase of Bok gene expression in granulosa cells of preantral and preovulatory follicles indicating some role in the ovulatory process.
Adult ; Animals ; Apoptosis* ; Blotting, Northern ; Chorionic Gonadotropin ; Diestrus ; Diethylstilbestrol ; Dimerization ; Estrus ; Female ; Gene Expression ; Gonadotropins ; Granulosa Cells ; Humans ; Hypophysectomy ; In Situ Hybridization ; Ovarian Follicle ; Ovary* ; Proestrus ; Rats* ; RNA, Messenger

Protein expression patterns of matrix metalloproteinases (MMPs) were examined in mouse reproductive organs during estrous cycle. Estrous cycle was classified into diestrus, proestrus, estrus or metestus and MMP expression was analyzed by zymography using gelatin as a substrate. Uterine fluid (UF) obtained both at diestrus and proestrus exhibited 4 major MMPs including 106kDa, 64kDa, 62kDa and 59kDa gelatinases. However, in UF at estrus, the gelatinolytic activity of 64kDa MMP disappeared and that of 106kDa and 62kDa MMPs dramatically decreased. At metestrus, 64kDa MMP activity reappeared and 106kDa and 62kDa MMP exhibited increased activities such that the band intensity of 106kDa was comparable to that in UF at diestrus. Gelatinolytic activity of 59kDa MMP was not changed throughout the cycle. Both ovarian and oviductal tissue homogenate revealed 4 MMPs which corresponded to the 4 MMPs of UF. However, unlike UF MMPs, gelatinolytic activity of these MMPs did not show distinct changes throughout the cycle. Either an inhibitor of MMP, 1, 10-phenanthroline, or a metal chelator, EDTA, abolished the appearance of the above MMP activities in gelatinated gel whereas a serine proteinase inhibitor, phenylmethylsulfonyl fluoride, failed to inhibit the appearance of MMP activities, proving that gelatinolytic activity of the above reproductive tissues were due to the enzymatic activity of MMP. When gelatinolytic activity of mouse serum was examined, it revealed 5 MMPs (131kDa, 106kDa, 89kDa, 64kDa and 62kDa bands) and one gelatinase (84kDa) band. From these results, it is concluded that the protein expression of MMPs of mouse reproductive organs, particularly uterus, is temporally regulated during estrous cycle and uterine 106kDa, 64kDa and 62kDa MMP,3 are suggested to play an important role in cyclic tissue remodeling of mouse uterus.
Animals ; Diestrus ; Edetic Acid ; Estrous Cycle* ; Estrus ; Female ; Gelatin ; Gelatinases ; Matrix Metalloproteinases* ; Metestrus ; Mice* ; Oviducts ; Phenylmethylsulfonyl Fluoride ; Proestrus ; Serine Proteases ; Uterus

PURPOSE: We established a rat model to investigate the female vaginal arousal response and the effects of the estrous cycle on vaginal blood flow in vivo. MATERIALS AND METHODS: Laser Doppler flowmetry was used to measure changes in vaginal blood flow induced by pelvic nerve stimulation(PNS) in 30 female Sprague-Dawley rats. Frequency response data were obtained in each animal. In addition, the rat's stage in the estrous cycle was determined according to the cell types observed in a vaginal smear. Changes in blood flow at 10 Hz frequency caused maximal response, and these responses were evaluated by comparing the relative peak flow, time to peak, duration of response, and relative area under the curve(AUC) according to the estrous cycle. RESULTS: Reproducible frequency dependent increases in vaginal blood flow were observed in response to PNS. Relative peak flow, time to peak, duration of response and relative AUC were slightly greater in proestrus and estrus groups(relatively higher estradiol level, n=17) than those in metestrus and diestrus groups(relatively lower estradiol level, n=13). However, these differences were not statistically significant. CONCLUSIONS: Our data suggest that the rat is a useful and reliable animal model for investigating the vaginal arousal response. In addition, the estrous cycle of the animal does not seem to be an important confounding factor in this animal model.
Animals ; Area Under Curve ; Arousal* ; Diestrus ; Estradiol ; Estrous Cycle ; Estrus ; Female* ; Humans ; Laser-Doppler Flowmetry ; Metestrus ; Models, Animal* ; Proestrus ; Rats* ; Rats, Sprague-Dawley ; Vagina ; Vaginal Smears

Opinions on ovariohysterectomy (OHE) of bitches vary depending on region and country. In this descriptive, prospective cross-sectional study, uterine tracts and ovaries exhibiting gross pathologic findings (n = 76) were collected post-surgery from a reference population of 3,600 bitches (2.11% incidence) that underwent elective OHE during September to November 2013 and evaluated by histopathology examination. Data were evaluated by using descriptive statistics and chi-squared tests. Bitches were of crossbred background with average age 5 years (range 0.6–8.0 years) and most were nulliparous (69.7%) with no anamnesis of reproductive diseases (81.6%). Frequencies of proestrus, estrus, and diestrus were 42.1%, 6.6%, and 19.7%, respectively. The presence of mammary gland masses (5.3%) significantly correlated with histopathologic findings in ovaries and age of the bitch (p < 0.05). Predominant uterine histopathologies included cystic endometrial hyperplasia, periglandular fibrosis, lymphoplasmocytary endometritis, and adenomyosis (19.7%, 14.5%, 4.0%, and 2.6%, respectively). In ovaries, hyperplasia of rete ovarii, follicular cysts, oophoritis, adenoma of the rete ovarii, cysts of superficial structures, and granulosa cell tumors (10.5%, 10.5%, 7.9%, 4.0%, 2.6%, and 2.6%, respectively) were observed. The results reveal the presence of subclinical pathologies in healthy bitches, suggesting that OHE at an early age is beneficial for prevention of reproductive pathologies.
Adenoma ; Adenomyosis ; Animals ; Cross-Sectional Studies* ; Diestrus ; Dogs* ; Endometrial Hyperplasia ; Endometritis ; Estrus ; Female ; Fibrosis ; Follicular Cyst ; Granulosa Cell Tumor ; Hyperplasia ; Mammary Glands, Human ; Oophoritis ; Ovary* ; Pathology ; Proestrus ; Prospective Studies ; Uterus*