Effect of contraceptive suture on biochemical composition of uterine fluid of protein deficient rats

To study the effect of a protein deficient diet on the biochemical composition and the volume of uterine fluid, 18 rats were maintained on a protein deficient diet and 18 rats on a normal diet. A surgical silk suture was inserted into the right uterine horn of each rat. After ovariectomy followed by rest of 7 days and injection of 50 mgm estradiol dipropionate daily for 7 days, the animals were sacrificed and the luminal fluid aspirated and studied. The presence of the silk suture led to an increase in the amount of albumin present in the fluid. There were no other alterations of biochemical composition of fluid attributable to the IUD or dietary protein deficiency. In vivo incorporation of d1-valine-1-carbon-14 into protein lactic acid glycogen, acid-soluble phosphorus, and phosphatase activity was not affected by the presence of the sutures.     

Biochemical changes in the developing rat central nervous system due to hyperthermia

Rat embryos at 10 days of gestation were exposed to 43 degrees C for 8 minutes by submerging the exteriorized right uterine horn in heated saline solution and then reinserting the uterine horn into the abdominal cavity. At 15 days, the fetuses were removed, and cells from the cerebral hemispheres were dissociated and grown as primary cultures. Embryos from the left uterine horn served as controls. No morphological changes were observed between the cultures of cells from control and heat-exposed embryos at different days in culture. However, exposure of embryos to hyperthermia at 10 days significantly affected the developmental pattern of activities of acetylcholine esterase associated with cholinergic neurons and of 2',3'-cyclic nucleotide phosphohydrolase associated with oligodendrocytes and myelin membrane formation. These results suggest that hyperthermia at 10 days of gestation in the rat may lead to an impairment in the development of neurons and oligodendrocytes in the central nervous system.     

Prostaglandin mediated action of IUD's (II) F-prostaglandins (PGF) in the uterine horn of pregnant rats and hamsters with intrauterine devices

The present investigation clearly shows that the presence of an IUD is associated with uterine hypertrophy and an increased production of F-prostaglandins.In the hamster, a bilateral IUD produced a significant increase in peripheral plasma PGF, presumably of uterine origin and an increased uterine PGF content at 2230 – 2300 h of day 4 of pregnancy. Earlier sampling (1530 – 1600 h) on the same day of pregnancy did not reveal any significant difference.In the rat, uterine PGF content and concentration, as well as PGF levels in uterine venous plasma were all increased in the IUD bearing horn from day 4 pregnant animals at 1530 – 1630 h in comparison to the contralateral control horn.     

An indwelling catheter for the collection of cat uterine fluids

A catheter system is described which allows for repeated samplings of uterine fluids in an awake animal. At laparotomy, catheters were inserted surgically into the ovarian pole of each uterine horn. The catheter ports then were passed through the abdominal wall through a stab incision and tunneled subcutaneously to a midlumbar position on each flank. The common uterine body was ligated to form a closed compartment. Uterine fluids were collected by saline lavage. Approximately 80% of the saline injected into one port was collected from the contralateral port and the catheters functioned successfully in each animal for approximately 45 repeated samplings.     

Death of sperm in the cervix of ewes bearing intrauterine plastic spirals or threads

A plastic spiral intrauterine device (IUD) in the ewe inhibits sperm transport through the cervix. In Exp. 1, plastic spirals were inserted surgically into the lumen of one or both uterine horns. In Exp. 2, Dacron threads were placed in the lumen of each horn. At a subsequent estrus, ewes were mated and necropsied 2 hours later. Sperm were washed from the uterine body and from the anterior, middle and posterior one-third of the cervix and examined. Plastic spirals significantly reduced the percentage of sperm recovered from the uterine body and each segment of the cervix that were motile and live and had normal acrosomes. In the anterior one-third of the cervix, e.g., 49% of the sperm were motile and 66% were live in control ewes but only 17% were motile and 23% were live in IUD ewes. Intrauterine threads also reduced the percentages of sperm that were motile and live in the anterior cervix. Failure of sperm transport in IUD-bearing ewes may be caused by conditions which result in loss of sperm motility in the cervix, thereby inhibiting the establishment of a reservoir of live sperm for transport to the oviducts.     

The rat uterine oestrogen receptor in relation to intra-uterine devices and the oestrous cycle [proceedings

There are changes in the nuclear content of the estrogen receptor in the rat uterus during the estrous cycle that are associated with changes in its physiology. The changes correlate with the concentrations of circulating estradiol. It appears that uterotrophic response to estradiol is a function of the nuclear receptor. The insertion of an IUD leads to changes in the treated uterine horn which appear to be the result of an increased responsitivity to circulating estradiol. The presence of an IUD did not alter the estrous cycle, gonadotropin, or corpus luteum function. The intracellular distribution of the estrogen receptor was investigated in normal uterine horns and in the horns with devices throughout the estrous cycle. Groups of 30 Wistar rats had a silk suture fitted in the lumen of 1 uterine horn. After 14 days the progress of these estrous cycles was determined. Rats were grouped according to the stage of the cycle on the 4th day. Rats were then killed and the uteri removed. Cytosol receptors were measured. The capacity of the cytosol estrogen receptor to bind to oligo(dT)-cellulose was determined. Cytosol protein, nuclear protein, and DNA were measured. At all stages of the estrous cycle, the wet weight and cytosol receptor of the treated horns were greater than the control horns. A slight increase in the capacity of cytosol receptor to bind to oligo(dT)-cellulose was noted at proestrus. The response elicited by the IUD was not considered to be due to an estrogenic response since the changes observed were not accompanied by a corresponding increase in the content of nuclear receptor.     

Effects of a copper-bearing intrauterine device on the ovarian function, body weight gain and pregnancy rate of nulliparous heifers

A copper-bearing intrauterine device (IUD), designed to cause a slight distention of the uterus, was inserted through the cervix into each uterine horn of 230 heifers; an additional 230 heifers served as the control group. Blood was drawn at 0, 1, 2, 20 and 120 d for progesterone and testosterone assays. The heifers were checked twice daily for estrus and examined at 0, 60 and 120 d for weight gain. Thereafter they were bred over a 120-d period. The IUD caused anestrus in 98% of the heifers, with a daily weight gain 25.5 % higher than in the control heifers. Moreover, the device was 100% effective in preventing pregnancy. At 20 and 120 d after IUD insertion progesterone levels averaged 0.7 ng/ml, which was 4 to 5 times lower than in the control animals, suggesting a failure in ovulation or in corpus luteum (CL) formation due to the IUD. Simultaneously, testosterone values were increased up to 8 times in IUD-treated heifers, reaching a mean concentration of 163 pg/ml. Associated histological evaluations of the ovaries from UD-treated heifers revealed the presence of 2 or more cysts per ovary, with marked hyperthecosis in many antral follicles in which the granulosa cell layers were either thinned or lacking. The results suggest that the action of the copper-releasing IUD used in this study resulted in high contraceptive efficiency but also in disturbance of ovarian function. Our findings further raise the possibility of a cause and effect relationship between hyperandrogenism and the higher body weight gain observed in heifers treated with the IUD.     

Fertilization rates and embryonic development in superovulated cattle inseminated in different sites within the reproductive tract

Superovulated Jersey and Holstein heifers and cows were bred 9.7 ± 2.7 h after the first observation of estrus with a single dose of frozen semen. Animals were grouped by site of insemination: 1) right uterine horn (n = 5), 2) left uterine horn (n = 4), 3) mid- uterine body (n = 5), and 4) mid-cervix (n = 6). The number of unfertilized ova, normal and abnormal embryos were recorded for each horn at slaughter 115.8 ± 18.6 h after insemination. All viable embryos were cultured in vitro and assessed for development. The overall fertilization rate was 76.8%, with the ipsilateral horn being higher than that of the contralateral horn (P < 0.05). Similar fertilization rates resulted among all treatments except those inseminated in the contralateral horn (P < 0.05). In vivo development of embryos was higher for the ipsilateral horn inseminations than those of the body of the uterus or cervical inseminations (P < 0.05), but it was not higher than the contralateral horn inseminations (P > 0.05). Under the conditions of this study with superovulated cows, these results suggest than 1) spermatozoa migrate from one horn to the other and 2) inseminating too deep into one horn may reduce the chances for concention when ovulation occurs contralaterally.     

Cytoplasmic oestradiol-binding sites and their relationship to oestradiol content in the endometrium of cattle.

Twenty-three cows and heifers were killed at known times during the oestrous cycle or during the first 35 days of pregnancy. Duplicate cytosol preparations were made from the endometrium of each uterine horn and both the binding-site concentration and the oestradiol level were determined for each sample. During the cycle, the oestradiol concentration was only 0-2 to 1-7% of the concentration of binding sites which varied considerably between Days 19 and 5 (47,665 +/- 7538 sites/cell, mean +/- S.E.M.) and Days 6 to 18 (7060 +/- 444 sites/cell). The concentration of binding sites remained low in pregnant animals (6689 +/- 492), although the oestradiol concentration was high about 20 days after insemination, resulting in almost 14% of the sites being occupied. Five inseminated animals in which no conceptus was found when they were slaughtered 19 to 22 days later had low concentrations of binding sites but two animals had high levels of oestradiol with 13% and 15%, respectively, of their cytoplasmic sites being occupied. It is suggested that these animals had recently lost their conceptuses. Two ovariectomized cows and one non-cyclic animal contained high concentrations of oestradiol-binding sites in the uterine cytoplasm. No significant difference was found between the uterine horn adjacent to the ovary with the CL and the contralateral horn in early pregnancy or during the luteal phase of the oestrous cycle. An animal killed 1 week after parturition contained fourfold more sites in the involuting horn than in the opposite horn. It is suggested that progesterone plays a major role in regulating oestrogen-induced replacement of cytoplasmic binding sites.     

Effect of prostacyclin inhibition by tranylcypromine on uterine 6-keto-pgf 1 alpha levels during estrogen hyperemia in rats

A role for prostacyclin (PGI2) as a mediator of estrogen-induced increases in uterine blood volume (UBV) was investigated by measuring uterine tissue levels of 6-keto-prostaglandin F1 alpha (6-keto-PGF 1 alpha), and testing estrogen responses in rats pretreated with the PGI2 synthesis inhibitor, tranylcypromine (TCP). Uterine 6-keto-PGF1 alpha content was determined by radioimmunoassay of tissue extracts purified through the use of high-performance liquid chromatography (HPLC). Estrogen treatment of castrate rats resulted in a significant increase of uterine 6-keto-PGF 1 alpha was compared to saline treated controls (9.3 ng/uterine horn vs 6.7 ng/uterine horn, p=0.01). Pretreatment with TCP (20 mg/kg) markedly reduced the uterine content of 6-keto-PGF 1 alpha (2.5 ng/uterine horn). The typical 50% increase in UBV observed after estrogen was unaffected by tranylcypromine pretreatment. It was concluded that the increased PGI2 synthesis, as indicated by elevated levels of 6-keto-PGF1 alpha, may function as an amplifying mechanism for the uterine vasodilation-induced by estrogen in castrate rats, but that production of this prostanoid is not essential for the estrogen response.     

The effect of bone-marrow transplantation on decidua formation in pseudopregnant rats

A main cause of failed pregnancy is insolvency of the decidual reaction of endometrial cells. It is believed that bone-marrow cells (BMCs) are a partial source of decidual cells in endometrial tissue. In the present work, the effect is studied of transplantation of BMCs on the desidualization processes using the model of pseudopregnancy in rats. BMCs were flushed from rat femurs and tibias. On the fifth day of pseudopregnancy, a suspension of single BMCs was injected into one of the uterine horns. PBS injection into the contralateral horn without cells served as control. Rats were sacrificed on the 11th day of pseudopregnancy. The diameter in the meso-antimesometral direction in the experimental uterine horn increased by 1.5–2 times compared to the control horn. The weight of decidual tissue in the experimental horn was three times greater than the weight of the control horn. The presence of transplanted BMCs in decidual tissue was documented by preliminary double-staining of BMCs with membrane dye PKH26 Red and nuclear dye Hoechst 33342. Histological analysis of decidua sections after transplantation did not reveal any alterations in cell differentiation or tissue structure. We concluded that transplantation of BMCs stimulated decidualization in animals.     

Unilateral luteotropic effect of uterine venous effluent of a gravid uterine horn in sheep.

The involvement of the main uterine vein in the unilateral maintenance of CL was studied in bilaterally ovulating unilaterally pregnant ewes. Ewes were mated at estrus (Day 0) and bilaterally ovulating ewes were randomized into three groups at surgery on Day 5. In all ewes, the uterine horns were separated through the intercornual area and one was ligated and transected near the internal bifurcation to produce a nongravid horn. One group served as controls (five ewes). In the other two groups the main uterine vein on one side was surgically anastomosed (end to side) to the corresponding vein of the opposite side (gravid side to nongravid side in one group--three ewes, and nongravid side to gravid side in the other--three ewes). Necropsies were done on Day 20. Mean CL weight was less, (P less than .01) on the nongravid side in control ewes than on the gravid side in control ewes or for either side in the other two groups. There were no significant differences among mean weights of CL on the gravid side in control ewes and either side in the other two groups. The CL regressed when the ipsilateral uterine vein contained blood from only the nongravid horn whereas the CL was maintained when the ipsilateral uterine vein contained venous blood from a gravid horn, whether or not it also contained blood from a nongravid horn. Results indicate that the uterine venous effluent from a gravid uterine horn in sheep has a luteotropic effect on the ipsilateral CL.     

Efficacy of a Y-Design Silastic Elastomer Intrauterine Device as a Horse Contraceptive

To successfully manage population growth rates of free-roaming horses (Equus ferus caballus), an effective, long-acting contraceptive could be beneficial. We evaluated the efficacy and safety of an intrauterine device (IUD) in a breeding trial using seasonal pasture assemblages of 2 males individually pastured in large enclosures (~81 ha) each with 10 females per male. The study took place at a large animal research facility at Oklahoma State University in Stillwater, Oklahoma, USA. Specific objectives of the study were to evaluate the effectiveness of IUDs in preventing pregnancy over 20.5 months with return to fertility assessed over the subsequent 6 months and to evaluate potential effects of this IUD on ovarian function and uterine health (e.g., endometrial inflammation, infection, fibrosis). We developed a Y-design IUD, made of silastic elastomer, and set the desired retention rate at 75%. After procurement of endometrial biopsies, we inserted IUDs into the uteri of 20 females and examined them via transrectal ultrasound every 2 weeks for detection of the device and assessment of uterine health. After 20.5 months we removed the remaining 15 IUDs, obtained a second endometrial biopsy for comparision, and returned the females to their respective males for continued breeding in efforts to assess uterine return to health as indicated by pregnancy rates. The Y-design IUD met the 75% retention rate goal, with 100% contraception in females that retained the device; 12 of 20 treated females became pregnant (i.e. returned to fertility) after removal of the IUD. Based on these results, we conclude that the Y-design IUD is a safe and effective device to control fertility in free-roaming horses. © 2021 The Wildlife Society.     

Morphometry of implantation in sheep. I. Trophoblast attachment, modification of the uterine lining, conceptus size and embryo location

The degree of trophoblastic attachment, modification of the uterine lining epithelium during implantation, possible differences in size of the conceptus, and preferential zones of nidation were evaluated. Eight ewes at 14 (n=3), 17 (n=2), 20 (n=1), and 24 (n=2) d of gestation were used, and 6 to 8 equidistant samples of either uterine o uterine plus embryonic membranes tissues were taken from each uterine horn for histomorphometric analysis. Trophoblastic attachment varied from 1.5 to 100% at 14 and 24 d of gestation, respectively. Modification of the uterine lining epithelium ranged from 0% at 14 d to 90% at 24 d. The conceptuses occupied a variable area of the uterine cavity; 15.1% at 14 d, 100% at 17 d, 70.6% at 20 d and 100% at 24 d of gestation. All the embryos were located caudal to the center of the uterine horn. Attachment of the trophoblast and modification of the uterine lining epithelium occurred in the area surrounding the embryos. Implantation was a gradual and long-lasting process which commenced near the embryo and extended peripherally. Morphological variations need to be considered when evaluating embryonic viability.     

Sequential uterine horn versus simultaneous total uterine flush to recover bovine embryos nonsurgically

Beef cows were superovulated with follicle stimulating hormone (FSH) to compare two nonsurgical methods of embryo recovery from the uterus. In the first method each uterine horn was independently flushed with physiological saline solution (PSS) through a Foley catheter passed through the cervix and into the uterine horn. In the second method both uterine horns were simultaneously flushed with PSS by passing the catheter into the uterine body. In both methods, the numbers of ovulations were determined after embryo collection by counting the corpora lutea (CL) on both ovaries of each cow through a flank incision. Independent flushing (n = 19) averaged 6.4 embryos and 16.1 CL per cow for a recovery rate of 40%. Simultaneous flushings (n = 22) averaged 5.4 embryos and 17.7 CL per cow for a recovery rate of 31%. This difference between the recovery rates of the two flushing methods was not significant (P>0.05).     

The bovine intercaruncular placenta throughout gestation

The attachment of chorion to uterine epithelium which results in a functional placenta began in the middle of the ipsilateral horn near the embryonic disc and spread towards the tip of each uterine horn. The intercaruncular union was fragile but perfusion and careful tissue processing preserved contact between chorion and intercaruncular epithelium from the fourth week until late in gestation. The adhesion-attachment process included binucleate cell migration from the chorion and remodelling of the maternal tissue. In areas where attachment was occurring or maintained, maternal epithelial height was reduced from columnar to cuboidal or squamous. Multinucleated cells and syncytium, which resulted from fusion of migrant binucleate cells with uterine epithelium, were common in the earlier stages of placentation. Extensive syncytium did not persist beyond the second month but some multinucleates were found throughout gestation. Attachment involved substantial areas in the bodies and middle regions of both horns but was not universal. Very limited attachment was found in the tip of the ipsilateral horn and none in the tip of the contralateral horn. In sections where contact could not be preserved the uterine epithelium was composed of tall columnar cells and no binucleate cell migration was detected. Intraepithelial lymphocytes were common in the tall columnar epithelium but almost totally absent where attachment and migration were occurring. Preservation of contact between the chorion and intercaruncular uterine epithelium throughout gestation demonstrated the diffuse nature of bovine placental attachment.     

The sensitivity of the uterus of the mouse and rat to intraluminal instillation

The tissue accumulation of 125I-labelled human serum albumin 30 min after intravenous injection was used as an index of uterine vascular permeability. In ovariectomized mice, all sham and experimental instillation procedures produced a 6-10-fold increase in vascular permeability. Some effects were also manifest in the contralateral, control horn. In ovariectomized rats, instillation of saline and arachis oil increased vascular permeability 3-7-fold. After 3 or more days of progesterone treatment following oestradiol priming, fluorocarbon and arachis oil instillation produced marked vascular responses, but these were not restricted to the transient period in which the uterus would respond with decidualization. An IUD prevented the response to arachis oil instillation. These results indicate that the uterus is very sensitive to any manipulation and are consistent with decidualization representing a specialization of a normal uterine inflammatory response.     

Histological studies on the therapeutic effect of sustained-release microspheres of a potent LHRH agonist (leuprorelin acetate) in an experimental endometriosis model in rats

The therapeutic effect of sustained-release microspheres of a potent LHRH agonist (leuprorelin acetate) on experimental endometriosis in female rats was examined histologically. Endometriosis was produced in rats by autotransplantation of endometrial tissue obtained from the left uterine horn into the renal subcapsular space. In the nontreated rats, the transplants were well established and had formed large cysts containing fluid. The walls of the cysts were composed of epithelium and stroma resembling that of normal endometrium. In the rats which received the microspheres of leuprorelin acetate, growth of the transplant was markedly suppressed as evidenced by the reduced size of the cystic cavity and the flattened and pyknotic epithelium. Also, the uterine and ovarian weight decreased significantly. In the ovariectomized rats, growth of the transplant was also markedly suppressed, and the uterine weight decreased. The present results clearly indicate that a single injection of the sustained-release microspheres of leuprorelin acetate markedly suppresses growth of the transplant and produces uterine and ovarian atrophy in the rats.     

A new non-uremic rat model of long-term peritoneal dialysis

Together with the development of peritoneal dialysis (PD), appropriate animal models play an important role in the investigation of physiological, pathophysiological and clinical aspects of PD. However, there is still not an ideal experimental PD animal model. In this study, 45 Sprague-Dawley rats were divided into three groups. Group 1 (n=15) was receiving daily peritoneal injection through the catheter connected to the abdominal cavity, using PD solution containing 3.86 % D-glucose. Group 2 (n=15) was receiving daily peritoneal injection of 0.9 % physiological saline through a catheter. Group 3 (n=15), which was subjected to sham operation, served as controls. Our results showed that WBC counts in peritoneal effluent of Group 1 were slightly higher than those of Group 2 and control group, respectively (p<0.05). However, there was no episode of infection in any group. In addition, there was no significant difference in neutrophils fractions among these three groups. Hematoxylin-eosin and Masson's trichrome staining demonstrated a dramatic increase in thickness of the mesothelium-to-muscle layer of peritoneum exposed to high glucose (Group 1) compared to Group 2 and controls (p<0.01). These data indicated that we established a novel rat model of PD with a modified catheter insertion method. This model is more practical, easy to operate, not too expensive and it will facilitate the investigate of long-term effects of PD.     

Progesterone inhibits rejection of xenogeneic transplants in the sheep uterus

OBJECTIVES: One of the proposed roles of progesterone is to prevent maternal immunological destruction of the allogeneic conceptus. Here, it was demonstrated that progesterone allows survival of a xenotransplant placed in the uterine lumen. METHODS: Ovariectomized ewes, surgically prepared to have ligatures around each uterine horn, were given daily subcutaneous injections of 50 mg progesterone or vehicle (sesame oil). After 30 days of treatment, mouse hybridoma cells were transplanted to one ligated uterine horn and phosphate-buffered saline was injected into the other horn. The uterus was flushed after an additional 14 days of treatment and hybridoma cells were identified by immunofluorescence. RESULTS: Overall, hybridoma cells were recovered from 4 of 5 progesterone-treated ewes and 1 of 5 vehicle-treated ewes. Immunohistochemical analysis of intercaruncular endometrium using antibodies towards CD8, gammadelta, and CD45R lymphocyte markers revealed that local presence of hybridoma cells caused a significant increase in CD8+ cells in all tissue compartments. While not significant, the numbers of CD8+ cells in the luminal and glandular epithelium were lower for progesterone-treated ewes. Progesterone tended to increase gammadelta T cell numbers in the glandular epithelium. CONCLUSIONS: Results demonstrate that xenograft rejection in the uterus is associated with an increase in CD8+ cells in the endometrium and that progesterone can inhibit uterine tissue graft rejection responses sufficiently to allow survival or delay rejection of xenograft tissue.