Uterine Connective Tissue Research Articles

Influence of genistein and diadizine on regularity of estrous cycle in cyclic female Wistar rat: interaction with estradiol receptors and vascular endothelial growth factor.

Isoflavones are estrogenic compounds that exist in soy, clover, and peanuts. They are selective estrogen receptor modulators. The study was planned to explain the interactions of isoflavones with estrogen receptors alpha (ERα), beta (ERβ), and vascular endothelial growth factor (VEGF) expressions in ovarian and uterine tissues during different stages of the estrous cycle of regular cyclic female Wistar rats. Thirty-two regular cyclic females were divided equally into control group: fed casein-based diet and isoflavones group: fed casein-based diet and gavaged 50 mg/kg/day soy isoflavones extract 40%. The regularity of estrus cycles was monitored. Final body weight (FBW), weight gain (BWG), and ovarian and uterine weights were estimated. Histopathology and immunohistochemistry for ERα, Erβ, and VEGF in ovarian and uterine tissues were performed. All females (100%, n = 16) in control group showed regularity in estrous cycle compared to 62.5% (n = 10) in isoflavones group. Estrus and diestrus phases revealed prolongation and shortening in isoflavones rats than control, respectively. Nonsignificant variation was noted in the duration of the whole cycle of both groups. FBW and BWG significantly decreased however, ovarian and uterine weights increased significantly in all estrous phases of isoflavones group than control. Histopathology demonstrated an increase in number of follicles/ovaries besides, hyperplasia and proliferation of luminal epithelium with hydropic degeneration in the isoflavones group. Also, uterine connective tissue stroma showed edema in the isoflavones group during all estrous phases. Immunostaining percentages of ERα, Erβ, and VEGF protein expression were significantly elevated in the isoflavones group during all estrous phases. Isoflavones induced irregularity of the estrous cycle that was encountered by increased and altered ERα, Erβ, and VEGF expressions in ovarian and uterine tissues.

The fetal-maternal interface of the nine-banded armadillo: endothelial cells of maternal sinus are partially replaced by trophoblast.

BackgroundThe evolution of invasive placentation in the stem lineage of eutherian mammals entailed resolution of the incompatibility between a semi-allogenic fetus and the maternal immune system. The haemochorial placenta of nine-banded armadillo (Dasypus novemcinctus) is thought to conceal itself from the maternal immune system to some degree by developing inside a preformed blood sinus, with minimal contact with the uterine connective tissue. In the present study, we elucidate the micro-anatomical relationship between fetal and maternal tissue of the nine-banded armadillo using histochemical and immunohistochemical tools.ResultsWe conclude that the chorio-allantoic villi are separated from the myometrium by a vascular endothelial layer, as previously proposed. However, we also observe that the trophoblast cells establish direct contact with the endometrial stroma on the luminal side of the endometrium by partially replacing the endothelial lining of the sinus. Further, we demonstrate the presence of leukocytes, perhaps entrapped, in the placental fibrinoids at the interface between the intervillous space and the endometrial arcade.ConclusionsThe trophoblast of the armadillo invades the uterine tissue to a greater extent than was previously believed. We discuss the implications of this finding for the fetal-maternal immune tolerance.

Immunohistochemical expression of collagen types I, III, IV and α-actin in the uterine horns of nulliparous and multiparous beagles

Collagen and smooth muscle cells play essential roles in the remodelling of uterine tissue during pregnancy and involution. To investigate the immunoreactivity and distribution pattern of collagen types I, III, IV and smooth muscle alpha-actin resulting from these processes, two homogenous groups of nulliparous and multiparous beagles were evaluated by immunohistochemistry. Immunostaining patterns of collagens I and III delineated the uterine connective tissue fibers and revealed their dual presence within fibers of both beagle groups. Collagen III staining, in particular, was more pronounced and especially evident in superficial fiber sections. The numerous, large arteries in the myometrial stratum vasculare of multiparous uteri exhibited a highly thickened intima, which distinctly expressed type I and III collagens. Intense collagen IV immunolabeling was discernable in the basement membranes of vascular endothelia and smooth muscle cells. Staining of the basement membranes of the luminal and glandular epithelia, conversely, was either absent or very weak. No difference in the immunoreactivity and distribution of the assessed collagens and actin could be detected between nulliparous and multiparous dogs. Overall, and with the exception of sclerotic arteries, immunohistochemical analysis revealed that the expression of uterine collagens and actin does not change in the uterus of multiparous beagles, even after seven elapsed pregnancies.

Analysis of obstetric complications and uterine connective tissue in tenascin-X-deficient humans and mice

Tenascin-X (TNX) is a large, multi-domain, extracellular matrix glycoprotein. Complete deficiency of TNX in humans leads to a recessive form of Ehlers-Danlos syndrome (EDS), and TNX haploinsufficiency is a cause of hypermobility type EDS. EDS patients appear to have a higher risk of several complications during pregnancy, such as pelvic instability, premature rupture of membranes, and postpartum hemorrhage. Here, we present a study of genitourinary and obstetric complications in TNX-deficient women of reproductive age. We have found complications, such as uterus prolapses, that are in agreement with previous findings in other EDS types. In TNX knockout (KO) mice, we have observed mild pregnancy-related abnormalities. Morphological and immunohistological analysis of uterine tissues has not revealed obvious quantitative or spatial differences between TNX KO and wildtype mice with respect to collagen types I, III, V, and XII or elastic fibers. We conclude that TNX-deficient women are at risk of obstetric complications, but that TNX KO mice show only a mild phenotype. Furthermore, we show that TNX is involved in the stability of elastic fibers rather than in their initial deposition.

The effect of dystocia and previous cesarean uterine scar on the tensile properties of the lower uterine segment

The remodeling of uterine connective tissue during labor can lead to the reorganization of the extracellular matrix that, in turn, may influence the biomechanical properties of the myometrial wall. We hypothesized that the stretching of the lower uterine segment in laboring women with dystocia changes the viscoelastic properties of the uterine wall. We tested the tensile strength of lower uterine segment myometrium in 68 pregnant women at term. The biomechanical, structural, and biochemical properties were compared among 3 groups: (1) 39 laboring women who underwent primary low-transverse cesarean delivery for labor dystocia, (2) 12 nonlaboring women who underwent primary elective low-transverse cesarean delivery and (3) 17 women who underwent an elective repeat low-transverse cesarean delivery at term. The tensile properties were quantitated with a stretching regimen that was designed to mimic the conditions of labor. Parameters such as slope, yield point, and break point were recorded, analyzed, and interpreted. Biochemical properties were determined by the measurement of the sulfated glycosaminoglycans, hydroxyproline, and pyridinoline-deoxypyridinoline. Histologic properties of the connective tissue were assessed by collagen birefringence. Lastly, the association between these properties and biomechanical responses were compared among groups. Lower uterine segment myometrium specimens obtained from laboring women were stiffer compared with specimens from women who were not in labor (P = .013) or had scarred myometrium (P < .001). The force that was required to reach the yield point was similar between labor and nonlabor groups (P = .216). Likewise, a previous lower uterine segment scar did not alter the yield point. The break point was similar among all groups (P = .317). Sulfated glycosaminoglycan levels were unaffected by labor or scarring (P = .354). Scarred lower uterine segment myometrium had a higher collagen content compared with unscarred myometrium specimens that were obtained during labor (P = .025). Although there were similar degrees of collagen cross-linking among groups (P = .212), there was lower collagen birefringence in myometrium from laboring women compared with nonlaboring women (P < .001). Labor alters the viscoelastic properties of myometrium. Lower uterine segment myometrium is stiffest in women with dysfunctional labor compared with nonlabor control subjects. Labor and scarring also alter the pattern of collagen birefringence. Similar collagen cross-linking among the study groups may explain the reason that the breaking strength of the tissue is not altered by the state of labor and the reason that the rupture of the uterine scar is a rare event.

Light and electron microscope immunocytochemical studies of the distribution of pregnancy associated glycoproteins (PAGs) throughout pregnancy in the cow: possible functional implications

Pregnancy associated glycoproteins (PAGs) comprise a large group of placental antigens of the aspartic proteinase family. Phylogenetic analysis indicated that the PAGs form two distinct groups, one of ancient origin and one produced by a more recent series of gene duplications. This paper summarises the molecular biological and biochemical studies which have been used to purify and raise antibodies against specific PAGs and groups of related PAGs and their use in light and electron microscope immunocytochemistry to demonstrate that the ancient PAG group has a similar distribution at the placental fetomaternal interface (microvillar junction, MVJ) in cows and pigs. This localization suggests either a possible role in binding the surfaces together and/or in establishment of an immunological barrier. The more recently evolved PAG group, absent in the pig, exhibited no significant localization to the MVJ but was restricted to the trophoblast binucleate cell (BNC) granules in the cow. Furthermore, these PAGs bind to newly formed maternal uterine connective tissue to which they are delivered by BNC migration and fusion with uterine epithelial cells. At this location in the developing maternal villi of the placentomes, they are ideally positioned to manipulate the maternal immune system to facilitate a successful pregnancy.

Changes of the uterine proteoglycan distribution at term pregnancy and during labour

Objective: To characterise uterine proteoglycans and changes therein during pregnancy and labour. Study Design: Uterine samples were collected from 6 non-pregnant, 10 term-pregnant and from 10 women in active labour. The proteoglycans were extracted by 4M guanidine hydrochloride and precipitated with Alcian Blue. They were separated by electrophoresis and identified by Western blotting. Results: Decorin was the dominating proteoglycan and smaller amounts of biglycan was found. A considerable amount of heparan sulphate proteoglycans was also detected. Decorin and biglycan decreased by 40% until term. The amount of heparan sulphate proteoglycans increased by 46% during active labour. Conclusion: Our data indicate that a considerable remodelling of the uterine connective tissue occurs during pregnancy and labour. The decrease of decorin and biglycan and the increase of heparan sulphate proteoglycans may be important for normal myometrial contractions during labour.

Interaction of estradiol, progesterone and corticosterone on uterine connective tissue degrading enzymes

The impact of ovarian hormones and corticosterone acetate on uterine connective tissue degrading enzymes were studied in mature albino rats. Ovariectomy resulted in a significant increase in the activities of α- and ß- galactosidases and glucosidases in the uterus. Administration of estradiol to ovariectomized rats brought back the activities of α-galactosidase and α-glucosidase to normalcy. While ß-galactosidase and ß- glucosidase were significantly decreased. Administration of progesterone to ovariectomized rats resulted in the increase of α- and ß-galactosidases and glucosidases. Administration of corticosterone to ovariectomized rats produced a further increase in α-and ß-galactosidases and glucosidases in the uterus. Adrenalectomy in ovary intact rats produced a decrease in α-galactosidase however, ß-glucosidase was significantly increased. Administration of corticosterone to ovary intact rats significantly increased the activities of α- and ß-galactosidases, while α- and ß-glucosidases were found to be decreased. Ovariectomy resulted in a significant increase in the activities of cathepsin-D and cathepsin-E. Administration of estradiol to ovariectomized rats brought back the activity of cathepsin-D to normalcy, whereas cathepsin-E was significantly increased. Administration of progesterone as well as estradiol to ovariectomized rats significantly increased the levels of cathepsin-E, however, cathepsin-D was brought back to normalcy. Administration of corticosterone to ovariectomized rats as well as Ovariectomy + adrenalectomy significantly increased the activity of cathepsin-D and cathepsin-E. Adrenalectomy significantly decreased the activity of cathepsin-D, while administration of corticosterone increased the cathepsin-D and cathepsin-E in the uterus. Therefore, these results suggest that estradiol is a potent ovarian steroid protecting the extra cellular matrix components. The effect of progesterone appears to modulate and act hand in hand with estradiol. Corticosterone appears to have an opposite effect to that of estradiol.

The influence of estrogens, progesterone and their antagonists on the collagen content in the pregnant rat uterus.

The influence of progesterone, estrogens and their antagonists: mifepristone and tamoxifen on connective tissue metabolism in pregnant rat uterus near term was evaluated. Total collagen and its soluble fraction as well as water concentration was measured in a population of 65 pregnant rats; treated with progesterone (n = 10), mifepristone (n = 13), estradiol (n = 17) and tamoxifen (n = 12). The results suggest the important influence of sex steroids on the uterine connective tissue metabolism during pregnancy.

Changes in rat cervical collagen during gestation and after antiprogesterone treatment as measured in vivo with light-induced autofluorescence

OBJECTIVE: This study was conducted to evaluate use of light-induced autofluorescence for measuring changes in collagen of cervical connective tissue during gestation. STUDY DESIGN: In a conventional laboratory setting light-induced autofluorescence from the cervix of Charles-Dawley timed pregnant rats was measured in vivo. Eight distinct groups at various times of gestation were examined. Measurements were also performed on rats at day 17 of gestation 24 hours after treatment with RU 38486. RESULTS: The amount of light-induced autofluorescence decreases significantly as pregnancy approaches term (means of the arbitrary total counts per spectrum on day 19 vs day 21 = 2.8 × 10 5 vs 1.9 × 10 5, p < 0.01) and reaches its lowest point when the pups are engaged in the cervix (day 19 vs day 22 = 2.8 × 10 5 vs 0.95 × 10 5, p < 0.001) and during delivery (day 19 vs delivery = 2.8 × 10 5 vs 0.83 × 10 5, p < 0.001). Preterm treatment with RU 38486 also caused a significant decrease in the native fluorescence compared with vehicle-injected controls (day 17 control vs day 17 RU 38486 = 2.8 × 10 5 vs 1.5 × 10 6, p < 0.001). CONCLUSION: Significant decreases in cervical collagen at parturition parallel the results of previous studies that used various indirect methods to analyze collagen content. These data support the use of light-induced autofluorescence for detecting and studying the changes in cervical and uterine connective tissue during gestation in vivo.

3 Biochemistry and physiology of preterm labour and delivery

Human parturition is associated with profound changes in uterine connective tissue affecting mainly the cervix, but the endocrine control of cervical ripening remains obscure. Connective tissue changes are also implicated in premature rupture of the membranes, a problem often associated with preterm delivery, and it is believed that local inflammatory infiltration may play a role in both this condition and cervical ripening, but it is difficult to define which changes precede parturition and which are a consequence of the trauma of labour. Chorioamnionitis can cause preterm labour by provoking the release of inflammatory mediators in the decidua/fetal membranes area and it is likely that activation of prostaglandin release by decidual macrophages is involved in triggering labour. However, the role of macrophages and other bone marrow derived cells in normal labour and in labour associated with chorioamnionitis needs to be defined. It is likely that treatment with a combination of antibiotics and prostaglandin synthase inhibitors and/or other anti-inflammatory drugs is the most appropriate therapeutic approach. Idiopathic preterm labour and spontaneous labour at term are probably due to changes in the sensitivity of the myometrium to endogenous agonists. Recent progress in cell signalling pathways, such as the characterization of regulatory G proteins and the cloning of hormone receptors, should clarify the mechanism of action of relaxing and contracting agents on myometrial cells and should provide the means for the development of new therapeutic agents of high effectiveness and selectivity. This approach should result in better management of both term and preterm labour.

Insufficient Remodelling of the Uterine Connective Tissue in Women With Protracted Labour

Department of Obstetrics and Gynaecology, Karolinaka Institutet, Danderyd Hospital, Danderyd, Sweden: and Department of Physiological Chemistry, University of Lund, Lund, Sweden

Insufficient remodelling of the uterine connective tissue in women with protracted labour

To investigate the association between a slow progress of labour and insufficient remodelling of the uterine connective tissue. An open comparative study. Danderyd Hospital, Department of Obstetrics and Gynaecology, referral centre. Eleven women (study group) in oxytocin augmented labour but with an unripe cervix in whom vaginal delivery could not be accomplished and 12 women (normal labour group) in normally progressing spontaneous labour and a favourable cervix but who needed to be delivered by caesarean section due to signs of fetal distress. At caesarean section tissue specimens were obtained from the fundus, the isthmus and the cervix uteri. Collagen concentration and extractability, collagenolytic activity expressed as DNP-peptide hydrolytic activity and the concentrations of sulphated glycosaminoglycans (S-GAG) and hyaluronic acid (HA) in the tissue specimens. Statistically significantly higher concentrations and lower extractability of collagen in the isthmus and the cervix uteri was found in women with slow progress of labour compared with those with normal labour. An insufficient remodelling of the connective tissue in the cervix and isthmus uteri may contribute to slow progress of labour.

Identification of a Uterine Elastase in the Pregnant Rat Uterus

During development of the pregnant rat uterus there is a several fold increase in elastin content. Using Verhoeff's elastic fiber stain, we have shown that a significant proportion of these elastin fibers are in the extracellular matrix of the myometrium. They do not appear as an organized structure but rather in a variety of partially extended, random configurations. An elastase was identified in both the pregnant and the postpartum uterus. Partial characterization of the enzyme indicated that it is a serine protease with a molecular weight around 24,500 and a pH optimum of 8.5. In addition to the enzyme, relatively high levels on an elastase inhibitor were found in the uterine extracts. The inhibitor did not inhibit trypsin, indicating that it was not alpha-1-antiprotease. The data suggest that the elastase and inhibitor are uterine tissue derived and perhaps important in the normal remodeling process of uterine connective tissue.

Hormonal influence on glycosaminoglycan synthesis in uterine connective tissue of term pregnant women.

Adaptation of the uterus to the growing fetus necessitates remodelling of the uterine connective tissue. Proteoglycans, being a main constituent of the extracellular matrix, influence the physical properties of the tissue and play an important regulatory role for a number of functional events. The synthesis of glycosaminoglycans (GAGs), the carboxyhydrate side chains of proteoglycans, in tissue from the lower uterine segment of term pregnant women was investigated in vitro by measurement of 35SO4 and [14C]glucosamine incorporation. Prostaglandin E2 and oestradiol-17 beta significantly increased the synthesis of sulphated GAGs but decreased the incorporation of [14C]glucosamine, while relaxin, prostaglandin F2 alpha and oxytocin had no significant effect. To further explore the influence of prostaglandin E2, tissue specimens were incubated with [14C]glucosamine and GAGs separated into three fractions on cetylpyridinium chloride cellulose micro columns. Prostaglandin E2 was found to significantly reduce the synthesis of components recovered in the glycoprotein and hyaluronate fractions, whereas synthesis of components in the sulphated GAG fraction was increased. The results indicate that prostaglandin E2 and oestradiol-17 beta have differential effects on different GAGs whereas relaxin, oxytocin and prostaglandin F2 alpha have no effect.

Antagonistic effects of estradiol dipropionate and progesterone on the histology of the vagina and uterus of the mouse.

Administration of estradiol dipropionate (20 micrograms/day; 7 days) to ovariectomized mice caused heavy epithelial proliferation and intense cornification in the vagina and cellular as well as glandular proliferation in uterine tissues. Endometrial hypertrophy with cystlike appearance of uterine glands was seen in response to a long-term (14 days) administration of estradiol dipropionate. Daily injection of progesterone (2 mg; 7 days) to ovariectomized mice resulted in desquamating mucosa, without any trace of vaginal cornification, and the presence of dense uterine connective tissue in the stromal region with typical uterine glands. However, treatment of estradiol depropionate in combination with progesterone at 1:100 dose ratio for 7 days produced vaginal histology similar to that in proestrus and uterine histology equivalent to the ovariectomized condition. The results revealed that progesterone antagonized the estrogenic effects and also that estradiol dipropionate antagonized the effects of progesterone. The effects of the two female sex steroids (estradiol dipropionate and progesterone) in vivo appeared to be more potent in the uterus than in the vagina.

Effects of oxytocin on cervical and uterine connective tissue.

The effect of oxytocin on collagen metabolism in the cervix and lower uterine segment of pregnant women was studied by measuring the incorporation of [3H]proline in vitro. Oxytocin had a concentration related inhibitory effect on the labelling with [3H]proline. Addition of indomethacin did not influence the response to oxytocin indicating that the effect was not directly mediated by prostaglandins. Oestradiol-17 beta potentiated the effect of oxytocin. Vasopressin decreased the incorporation of [3H]proline slightly but the action of this hormone was significantly less than that of oxytocin. The results suggest that oxytocin under in vitro experimental conditions influences cervical connective tissue metabolism which is in contrast to current clinical experiences.

Regulatory influence of relaxin on human cervical and uterine connective tissue.

Cervical tissue was obtained from women undergoing legal abortion in the 7th-15th week of gestation and tissue from the lower uterine segment was excised at elective Casearean section in the 38th-40th week. The specimens were incubated with [3H]proline in the presence of relaxin or prostaglandin E2 (PGE2). Relaxin had a concentration related inhibitory effect on the radiolabelling in the 7th-9th week but failed to influence the amino acid uptake in the 10th-15th week of pregnancy. PGE2 had the inverse effect, i.e. no influence in the former group but reduced incorporation of proline in the latter group of patients. Incubation of tissue from the lower uterine segment showed a similar response as that of the early pregnant cervix. It is concluded that relaxin has a significant influence on [3H]proline incorporation by cervical and uterine tissue under in vitro experimental conditions.

Degradation of extracellular matrix by mouse trophoblast outgrowths: a model for implantation

During implantation the embryo attaches to the endometrial surface and trophoblast traverses the uterine epithelium, anchoring in the uterine connective tissue. To determine whether trophoblast can facilitate invasion of the uterus by degrading components of normal uterine extracellular matrix, mouse blastocysts were cultured on a radio-labeled extracellular matrix that contained glycoproteins, elastin, and collagen. The embryos attached to the matrix, and trophoblast spread over the surface. Starting on day 5 of culture there was a release of labeled peptides into the medium. The radioactive peptides released from the matrix by the embryos had molecular weights ranging from more than 25,000 to more than 200. By day 7 there were areas where individual trophoblast cells had separated from one another, revealing the underlying substratum that was cleared of matrix. When trophoblast cells were lysed with NH(4)OH on day 8, it was apparent that the area underneath the trophoblast outgrowth had been cleared of matrix. Scanning electron microscopy and time-lapse cinemicrography confirmed that the digestion of matrix was highly localized, taking place only underneath the trophoblast, with no evidence of digestion of the matrix beyond the periphery of the trophoblast outgrowth. The sharp boundaries of degredation observed may be due to localized proteinase secretion by trophoblast, to membrane proteinases on the surface of trophoblast, or to endocytosis. Digestion of the matrix was not dependent on plasminogen, thus ruling out a role for plasminogen activator. Digestion was not inhibited by a variety of hormones and inhibitors, including progesterone, 17beta-estradiol, leupeptin, EDTA, colchicine, NH(4)Cl, or epsilon-aminocaproic acid. This system of culturing embryos on extracellular matrix may be useful in determining the processes that regulate trophoblast migration and invasion into the maternal tissues during implantation.0

The role of connective tissue in the maintenance of epithelial differentiation in the adult.

Specimens of buccal mucosa and ear skin were introduced into the uteri of isologous rats and the animals maintained under estrogen stimulation for ten weeks, whereupon the uteri were removed and processed for light- and electron-microscopic examination. The majority of implants were successful with the epithelium having migrated to replace the adjacent uterine epithelium. Epidermis and oral epithelium growing on its own connective tissue in the uterus showed a normal pattern of histodifferentiation, including the formation of a thickened keratinized surface and appendages such as hair and sebaceous glands. Ultrastructurally, maturation was similar to that of normal tissue. Epidermis growing on uterine connective tissue did not form appendages but the ultrastructural pattern of cytodifferentiation was similar to control tissue. Buccal epithelium on the uterine stroma was markedly altered in its pattern of maturation, the epithelium appearing thinner than usual and showing a pattern of maturation resembling that of non-keratinization. These results suggest that normal histodifferentiation in epidermis and oral epithelium requires the presence of the appropriate connective tissue; in its absence epidermis has an intrinsic capacity for more or less normal cytodifferentiation but this is lacking in the buccal epithelium.