Myometrial Growth Research Articles

8636 Characterization Of The Role Of Neutrophils In Promoting Progression Of Lymphangioleiomyomatosis

Abstract Disclosure: T.R. Henson: None. B. Minor: None. S.R. Hammes: None. Lymphangioleiomyomatosis (LAM) is a rare, multisystemic disease that is associated with cystic lung destruction. LAM is caused by small smooth muscle cell-like tumors throughout the lung that contain deleterious mutations in the tuberous sclerosis complex genes TSC1 or TSC2. This attenuates the activity of the mammalian target of rapamycin complex 1 (mTORC1) causing increased proliferation of the smooth muscle cells. LAM is a sexually dimorphic disease that primarily affects genetic females. Clinical observations implicate the role of estrogen in disease progression, as it is exacerbated during pregnancy and by use of exogenous estrogens in birth control. LAM tumor-derived cells lines have shown that direct estrogen stimulation of cell proliferation is modest compared to the dramatic in vivo effects of estradiol (E2) stimulation in a uterine-specific TSC2-null mouse model. Creation of a monoclonal cell line from these mice resulted in the generation of the LTM3 cell line, which also demonstrates low response to direct estrogen stimulation in vitro. This indicates that direct E2 stimulation may act upon other cell types in the tumor microenvironment. Immunophenotyping of uterine-specific TSC2-null mice has shown that the tumor microenvironment is enriched with myeloid cells specifically neutrophils. Functional analysis of neutrophil suppression in vivo has shown a reduction in myometrial growth in the uterine-specific TSC2-null mice, indicating a key role for neutrophil-mediated LAM tumor growth. Altered transcriptional regulation of neutrophils further highlights the importance of neutrophils in LAM progression. Bulk messenger RNA sequencing revealed E2 dependent increase in the neutrophil derived serine protease neutrophil elastase (NE). NE in vitro has been shown to increase migration, invasion, and proliferation of LAM cell lines, while inhibition in vivo has shown a reduction to the myometrial growth. Previously published data by the lab has shown that E2 increases tumor induced expansion of neutrophils in the bone marrow. Isolation and purification of Neutrophils from mouse bone marrow stimulated with tumor conditioned media from the LTM3 cells has shown signs of activation via upregulated gene expression. The focus of the current research has been the characterization of bone marrow derived neutrophils. Currently working with an in vitro system to study the proliferation of LTM3 and the rat leiomyoma (ELT3) cell lines when they are co-cultured with neutrophils. Understanding the mechanism by which neutrophils are activated and interact with LAM cells can give understanding to disease progression may provide insight into future treatment options. Presentation: 6/3/2024

Reproductive Functions of the Mitochondrial Progesterone Receptor (PR-M).

Classic transcriptional regulation by progesterone via the nuclear progesterone receptors A and B (PR-A, PR-B) has been recognized for decades. Less attention has been given to a mitochondrial progesterone receptor (PR-M) responsible for non-nuclear activities. PR-M is derived from the progesterone receptor (PR) gene from an alternate promoter with the cDNA encoding a unique 5' membrane binding domain followed by the same hinge and hormone-binding domain of the nPR. The protein binds to the mitochondrial outer membrane and functions to increase cellular respiration via increased beta-oxidation and oxidative phosphorylation with resulting adenosine triphosphate (ATP) production. Physiologic activities of PR-M have been studied in cardiac function, spermatozoa activation, and myometrial growth, all known to respond to progesterone. Progesterone via PR-M increases cardiomyocyte cellular respiration to meet the metabolic demands of pregnancy with increased contractility. Consequential gene changes associated with PR-M activation include production of proteins for sarcomere development and for fatty acid oxidation. Regarding spermatozoa function, progesterone via PR-M increases cellular energy production necessary for progesterone-dependent hyperactivation. A role of progesterone in myometrial and leiomyomata growth may also be explained by the increase in necessary cellular energy for proliferation. Lastly, the multi-organ increase in cellular respiration may contribute to the progesterone-dependent increase in metabolic rate reflected by an increase in body temperature through compensatory non-shivering thermogenesis. An evolutionary comparison shows PR-M expressed in humans, apes, and Old World monkeys, but the necessary gene sequence is absent in New World monkeys and lower species. The evolutionary advantage to PR-M remains to be defined, but its presence may enhance catabolism to support the extended gestation and brain development found in these primates.

Identification of uterine telocytes and their architecture in leiomyoma.

Uterine leiomyoma is the most widespread benign tumor affecting women of childbearing age. ere are still gaps in the understanding of its pathogenesiss. Telocytes are unique cells described in greater than 50 different locations inside the human body. The functional relationship of cells could clarify the pathogenesis of leiomyomata. In the current study, we focused on the identification of telocytes in all regions of the human uterus to explain their involvement in leiomyoma development. Tissue samples from a healthy and myomatous uterus were stained for c-kit, tryptase, CD34 and PDGFRα to identify telocytes. Routine histology was performed to analyze tissue morphology and collagen deposits. Telocytes were detected in the cervix, corpus of the uterus and leiomyoma. The density of telocytes in fibroid foci was reduced compared with normal myometrium. Our results demonstrated the existence of telocytes in all parts of the human body affected and unaffected by leiomyoma of the uterus. In addition, telocytes were also present in leiomyoma foci. Our results suggest that the reduced density of telocytes is important for the pathomechanisms of myometrial growth, demonstrating its value as a main component of the myomatous architecture.

In Vivo Evidence of the Androgen Receptor in Association With Myometrial Cell Proliferation and Apoptosis.

The androgen receptor (AR) plays essential roles in female reproductive physiology. In vitro studies have shown that AR exerts 2 distinct but related functions in myometrial cells. The AR is required for myometrial cell cycling ligand independently, thereby controls myometrial cell proliferation. The AR also exerts antiapoptotic functions through myeloid cell leukemia 1 (MCL1) in both ligand-dependent and ligand-independent manners. However, these AR functions have not yet been confirmed in vivo. This study provides in vivo evidence that AR expression is associated with myometrial cell proliferation and apoptosis. Myometrial tissue was collected on day 6 of pregnancy from mice bearing an intact AR (AR+/+) or mice with AR heterozygous (AR-/+) or homozygous (AR-/-) functional knockout. Human uterine leiomyoma and paired myometrial tissue biopsies (n = 14 patients) were collected during hysterectomy. Immunohistochemistry was used to assess the protein expression of AR, MCL1, and Ki-67 index, as well as the functional associations of AR with MCL1 and Ki-67 index. In AR-/- mice, MCL1 protein levels were reduced ∼50% in both circular and longitudinal myometrial cells when compared with that in AR+/+ and AR-/+ mice. By contrast, Ki-67 index remained unchanged. The AR protein expression and Ki-67 index in leiomyoma were ∼2- to 3-fold higher than that in normal myometrium. Although MCL1 expression was not coordinately increased in leiomyoma, strong positive correlations between AR and MCL1 expression were observed in leiomyoma but not in normal myometrium tissue. These results suggest that AR is an important regulator of myometrial growth in vivo during pregnancy and in leiomyoma.

A Study of Myometrial Growth and Development

Study ObjectiveTo evaluate myometrial growth and development. DesignThirty-five autopsy uteri, ranging from 10 weeks' gestation to age 18 years, acquired over 3 decades from 2 hospitals, were studied based on specimen availability, photographed for documentation, and reviewed at the end of the study. Most were embedded in toto, with 1 block and 1 slide per case. Some were immunostained for actin, CD10, MIB-1, and/or trichrome stain for collagen and muscle. Myometrial thickness was measured by ocular micrometry when sections were nontangential and analyzed by paired-sample t tests and bivariate linear regression. SettingTwo university-affiliated hospitals. ResultsFrom 20 to 34 weeks, lateral wall corpus thickness increased 6-fold, with a 4- to 6-fold perinatal burst of growth (P < .01) and a drop in thickness after the neonatal period (P = .013). The corpus was thicker than the dome (P < .01) but less thick than the lower uterine segment (P = .087). The lower uterine segment was fully muscular in the second trimester, becoming more fibrous near term. Intramural, subserosal, and inframucosal myometaplasia were observed, as primitive stromal cells turned into muscle cells. Myometrial proliferation was brisk in the second trimester but greatly diminished in the perinatal period. Pressure effects from myometrial tone were observed during development. There was a pubertal burst of inframucosal myometaplasia. ConclusionsMyometaplasia accounted for most myometrial growth, especially in the perinatal and pubertal bursts of growth. Pressure effects, related to myometrial tone, appeared to affect myometrial development. True endocervix, with a fibrous wall and mucinous epithelium, appeared late in development.

Polymorphic stability of progesterone under stress conditions

Progesterone (PRG) is an essential hormone in regulating the female reproductive system. It is used in gynecology, including contraception, medical abortion, and treatment of conditions related to endometrial and myometrial growth [1]. This drug exhibits two different crystal structures: forms 1 (PRG-1) and 2 (PRG-2); both with equal pharmacological effects. PRG-2 is the metastable form and it is 1.1 kcal/mol less stable than PRG-2 [2]. Polymorphic instability occurs when a metastable solid form (PRG-2) transforms into the most thermodynamically stable (PRG-1) [3]. It is well-known the therapeutic efficacy of a drug substance or drug product can be altered by its polymorphic or chemical transformation [4]. In order to study the polymorphic stability both polymorphs, PRG-1 and PRG-2, were placed under the same mechanical, thermal and high humidity stress conditions. X-ray powder diffraction (XRPD) analysis showed PRG-2 maintained its crystal structure after 3 months of exposure to dark storage condition at 400C/75% relative humidity and 1 month to light. High performance liquid chromatography (HPLC) showed no degradation of PRG-1 and PRG-2. However, mechanical stress (grinding) on PRG-2 produced seeds of PRG-1 that induced its polymorphic transformation (Figure 1A). DSC curve showed and endothermic events at 120.70C corresponding to the melting point of PRG-2 followed by the recrystallization at 123.80C of PRG as PRG-1, finally the fusion of PRG-1 at 127.90C. The Ea average value was higher in PRG-2 than in PRG-1; it suggested PRG-2 is more thermally stable than PRG-1 and, the decomposition behavior is different between these two polymorphs. Degradation products from thermal stability studies were monitored by coupled TGA-FT-IR analyses and different analyses (Table 2 inset in Figure 1). PRG-1 shows a high maximal rate of degradation (Tdmax) and initial temperature of thermal decomposition (To) compares to PRG-2.

The androgen receptor mediates antiapoptotic function in myometrial cells

During pregnancy, myometrial phenotype is programmed into three characteristic stages referred to as the early proliferative, the midterm hypertrophic, and the late contractile stage. Increased myometrial growth in the early and midterm of pregnancy involves a complex process of cell proliferation, antiapoptosis and differentiation. We have previously demonstrated that the androgen receptor (AR) is required for myometrial cell proliferation by modulating IGF-1 signaling during early pregnancy. Here, we report that AR also exerts its antiapoptotic function in human myometrial cells. Enhanced AR expression protects, whereas AR silencing sensitizes myometrial cells to both intrinsic and extrinsic apoptotic stimuli. AR agonist inhibits, whereas AR antagonist induces myometrial cells to undergo apoptotic cell death. Gene microarray analysis confirms that the central functions of AR in myometrial cells are to regulate cell cycling and apoptosis through three major gene groups involving the epidermal growth factor (EGF) signaling, RNA splicing and DNA repair processes. AR mediates its antiapoptotic function through two distinct pathways. In the receptor-dependent pathway, AR is required for the expression of several protein factors within the EGF signaling pathway. Through the PI3K/Akt pathway, AR enhances the expression of the antiapoptotic protein Mcl-1. In the ligand-dependent pathway, AR agonist triggers the activation of Src kinase, which in turn phosphorylates STAT3 to increase Mcl-1 expression. We conclude from these results that the AR signaling exerts antiapoptotic function in myometrial cells, further supporting its key role in programming of myometrial phenotype.

A Case of Misdiagnosed Cesarean Scar Pregnancy with a Viable Birth at 28 Weeks

We report our experience with a case of presumptive cesarean scar pregnancy, based on detection of a gestational sac (GS) in early pregnancy at the site of a previous cesarean scar. The GS grew into the uterine cavity as the pregnancy progressed, showing an ultrasound image similar to that of a normal pregnancy. Thus, the pregnancy continued, resulting in a viable birth at 28 weeks of gestation. Cesarean scar pregnancy is classified as myometrial implantation or implantation growth into the uterine cavity. In the latter type, the gestational sac moves upward with increasing gestational weeks and it shows the same ultrasound image as a normal pregnancy. Therefore, the diagnosis must be made in the early pregnancy.

Uterine-Specific Loss of Tsc2 Leads to Myometrial Tumors in Both the Uterus and Lungs

Lymphangioleiomyomatosis (LAM) is a rare disease characterized by proliferation of abnormal smooth-muscle cells in the lungs, leading to functional loss and sometimes lung transplantation. Although the origin of LAM cells is unknown, several features of LAM provide clues. First, LAM cells contain inactivating mutations in genes encoding Tsc1 or Tsc2, proteins that limit mTORC1 activity. Second, LAM tumors recur after lung transplantation, suggesting a metastatic pathogenesis. Third, LAM is found almost exclusively in women. Finally, LAM shares features with uterine leiomyomas, benign tumors of myometrial cells. From these observations, we proposed that LAM cells might originate from uterine leiomyomas containing Tsc mutations. To test our hypothesis, and to develop mouse models for leiomyoma and LAM, we targeted Tsc2 deletion primarily in uterine cells. In fact, nearly 100% of uteri from uterine-specific Tsc2 knockout mice developed myometrial proliferation and uterine leiomyomas by 12 and 24 weeks, respectively. Myometrial proliferation and mTORC1/S6 activity were abrogated by the mTORC1 inhibitor rapamycin or by elimination of sex steroid production through ovariectomy or aromatase inhibition. In ovariectomized Tsc2 null mice, mTORC1/S6 activity and myometrial growth were restored by estrogen but not progesterone. Thus, even without Tsc2, estrogen appears to be required for myometrial mTORC1/S6 signaling and proliferation. Finally, we found Tsc2 null myometrial tumors in lungs of older Tsc2 uterine-specific knockout females, suggesting that lung LAM-like myometrial lesions may indeed originate from the uterus. This mouse model may improve our understanding of LAM and leiomyomas and might lead to novel therapeutic strategies for both diseases.

Current Perspectives in Uterine Research: Reviews From the Society for Gynecologic Investigation Myometrium and Parturition Satellite Symposia

It is difficult to imagine examples of greater physiological adaptations than those of the uteroplacental unit during pregnancy. The sheer scale of molecular, cellular and tissue remodeling, organogenesis, and organism development in a relatively short period of time seems the epitome of ‘‘systems biology.’’ The processes are challenging to comprehend, and investigate experimentally, but undoubtedly fascinating. Efforts to understand the physiology and pathology of these phenomena are of prime importance due to the considerable health and socioeconomic pressures brought to bear by the high incidences of compromised pregnancies and, especially, preterm birth (PTB). The World Health Organization-sponsored studies equate the worldwide incidence of PTB to be approximately 13 million births per year with approximately 1 million neonatal deaths attributable to PTB. If improved diagnosis and therapeutic treatment of PTB are to be achieved, an integrated understanding of the physiological basis of pregnancy and parturition is required. With this in mind, each year the Myometrium and Parturition Satellite Symposium of the Society for Gynecologic Investigation meeting gathers international experts in this field to share their experimental findings. We are delighted that several plenary speakers from the 2011 and 2012 symposia have provided reviews on their topics for this issue of Reproductive Sciences. The articles provide an up-to-date compendium that integrates many of the important facets of uteroplacental remodeling and function leading to term and preterm labor. Jerome Strauss (Virginia) introduces the importance of dynamic remodeling of the extracellular matrix (ECM) and its molecular components in uteroplacental tissues. With a focus on fetal membrane interfaces, he summarizes the ECMs’ multiplicity of roles including receptor signaling, cytokine/ growth factor sequestration, and tensile strength. Among the important molecular components are fetal fibronectin, hyaluronan, and up to 28 collagens, regulated by different metallomatrix proteinases. Mutations in many genes encoding such ECM proteins are associated with increased incidence of preterm premature rupture of membranes and the roles of environmental and/or epigenetic factors are being increasingly elucidated. In a similar vein, Shynlova et al (Toronto, Canada) describe pregnancy in terms of progressive changes of tissue remodeling under the control of localized inflammatory events (physiological inflammation). The authors posit a carefully structured theory whereby spatially restricted inflammatory events are under the influence of ever-changing endocrine (and a focus is given to the role of progesterone) and mechanical inputs activating a rich milieu of chemokine/cytokine signaling. These, in turn, serve to coordinate diverse processes such as decidualization, myometrial cell proliferation and growth, leukocyte migration, cervical remodeling, and postparturient involution. Olcese et al (Tallahassee, USA) review an oft-neglected consideration of the mechanisms responsible for the timing of parturition. They too give credence to the notion that pregnancy and parturition involve dynamic physiological adaptations—in this case that uterine contractions of labor may be initiated by activity of the central circadian oscillator of the maternal hypothalamic suprachiasmatic nuclei via the feedback ‘‘gating’’ actions of neuroendocrine outputs. Of key importance may well be circadian secretion of melatonin given recent interest in the uterotonic actions of this hormone and uterine expression of melatonin receptors. Regulation of contractile activity of the uterus is also the focus of reviews by Europe-Finner et al (Newcastle, UK) and Wim Lammers (Al Ain, UAE). The former concentrates on intracellular signaling and describes the possibility that posttranslational modification by acetylation may be a mechanism of wider significance to the uterus than first realized when considering ‘‘‘only’’ nuclear events of histone modification, chromatin remodeling, and gene transcription. They suggest

Mechanical stretch regulates hypertrophic phenotype of the myometrium during pregnancy

The adaptive growth of the uterus is a critical event that involves changes in cellular phenotypes throughout pregnancy. In early pregnancy, uterine growth is due to hyperplasia of uterine smooth muscle cells (SMCs) within the myometrium; however, the major component of myometrial growth occurs after mid-gestation. This study sought to test the hypothesis that increase in myometrial growth seen during late pregnancy is due to SMC hypertrophy caused by mechanical stretch of uterine tissue by a growing fetus(es) by providing direct measurements of individual SMC size. We employed a stereological approach to calculate the average cell volumes of uterine myocytes through diameter measurements using the Stereoinvestigator statistical software. Uterine tissues were collected from nonpregnant Wistar rats, as well as from gravid and nongravid horns of unilaterally pregnant animals on gestational days (d) 8 (early gestation), 14 (mid-gestation), 19 (late gestation), 22 (term), and 4 days post partum. Anti-caveolin-1 immunostaining was used to clearly delineate SMC boundaries. The stereological analysis revealed that the dramatic increase in myometrial growth seen during late gestation (d19-22) is due to a threefold increase in the size of uterine myocytes. A significant increase in SMC volumes was detected in the gravid uterine horn as compared with the corresponding empty horn of unilateral term pregnant animals (day 22, mean cell volume 1114 vs 361 microm(3), P<0.05), indicating the effect of uterine occupancy. The restriction of the hypertrophy to cells within the gravid horn suggests that it may be a response to the biological mechanical stretch of uterine walls by the growing fetus(es) and placenta(s).

Insulin-like Growth Factors and Their Binding Proteins Define Specific Phases of Myometrial Differentiation During Pregnancy in the Rat1

While the insulin-like growth factor (IGF) system is known to regulate uterine function during the estrous cycle, there are limited data on its role in myometrial growth and development during pregnancy. To address this issue, we defined the expression of the Igf hormones (1 and 2), their binding proteins (Igfbp 1-6), and Igf1r receptor genes in pregnant, laboring, and postpartum rat myometrium by real-time PCR. IGF family genes were differentially expressed throughout gestation. Igf1 and Igfbp1 mRNA levels were upregulated during proliferative phase (Days 6-12) of rat gestation. Igfbp3 gene expression also was elevated in proliferating smooth muscle cells (SMCs) and was highest at the time of transition between proliferative and synthetic phases (Days 12-15). Igfbp6 gene expression profile paralleled plasma progesterone (P4) concentrations, peaking during the synthetic phase (Days 17-19) and decreasing thereafter. Administration of P4 at late pregnancy (starting from Day 20) to maintain elevated plasma P4 concentrations blocked the onset of labor and prevented the fall in Igfbp6 mRNA levels. In contrast, the treatment of pregnant rats with the P4 receptor antagonist RU486 on Day 19 induced preterm labor and the premature decrease of Igfbp6 gene expression. Igfbp2 gene expression was transiently upregulated during the contractile phase of gestation (Days 21-23) solely in the gravid horn of unilaterally pregnant rats, but it was not affected in P4- or RU486-treated animals, supporting a role for mechanical stretch imposed by the growing fetuses. Igfbp5 gene was induced during postpartum involution. Our results suggest the importance of the IGF system in phenotypic and functional changes of myometrial SMCs throughout gestation in preparation for labor.

Myometrial Apoptosis: Activation of the Caspase Cascade in the Pregnant Rat Myometrium at Midgestation1

In the present study, we determined the contribution of myometrial hyperplasia, hypertrophy, and apoptosis to uterine growth during pregnancy. The changes in two endogenous markers of cell replication, proliferating cell nuclear antigen (PCNA) protein expression and bromodeoxyuridine (BrdU) incorporation, were studied. Myocyte hypertrophy was assessed by measuring the protein:DNA ratio. The expression levels of antiapoptotic regulatory proteins (BCL2 and BCL2L1) and enzymes involved in apoptosis (caspases 3, 6, 7, 9, and 10) were assessed by immunoblotting throughout gestation and postpartum. Myometrial cell apoptosis was determined by TUNEL staining and DNA fragmentation assays. Both BrdU incorporation and PCNA labeling were elevated in early pregnant myometrium and decreased dramatically after midgestation, with a simultaneous increase in cellular hypertrophy. Levels of BCL2 were high during early gestation, followed by significantly elevated levels of BCL2L1 at midgestation. The expression of caspase 10 in myometrial samples declined from a high nonpregnant level to a complete loss at early gestation. The cleaved forms of caspases (CC) 3, 6, 7, and 9, as well as poly(ADP-ribose)polymerase-1, were undetectable in the myometrial samples at early or late gestation but were transiently elevated at midgestation. Immunohistochemical staining of CC3 confirmed the activation of the caspase cascade, but TUNEL-positive staining or the increase in DNA fragmentation was not detected. Collectively, two distinct phases of myometrial growth were observed: myocyte hyperplasia associated with an increase in antiapoptotic proteins during the first half of gestation, and cellular hypertrophy during the second part of gestation. The transition between these phases was associated with transient activation of the caspase cascade that triggered the differentiation of uterine smooth muscle.

Comparative effects of heparin-binding epidermal growth factor-like growth factor on the growth of cultured human uterine leiomyoma cells and myometrial cells

The objective of this study was to investigate the comparative effects of heparin-binding epidermal growth factor-like growth factor (HB-EGF) on the growth of cultured human leiomyoma cells and myometrial cells. Isolated cells were subcultured in Phenol Red-free Dulbecco's modified Eagle's medium supplemented with 10% fetal bovine serum for 120 h and then stepped down to serum-free conditions for an additional 24 and 48 h in the presence or absence of graded concentrations of HB-EGF (0.1, 1, 10 and 100 ng/ml). These cells were used for immunocytochemical analysis for Ki67, western blot analysis for proliferating cell nuclear antigen (PCNA) and human EGF receptor (HER1), and TdT (terminal deoxynucleotidyl transferase)-mediated dUDP nick-end labelling (TUNEL) assay. Treatment with HB-EGF at concentrations >1 ng/ml significantly increased the Ki67-positive rate of cultured leiomyoma cells and myometrial cells. Treatment with HB-EGF also resulted in a dose-dependent increase in PCNA expression in both cells compared with untreated control cultures. A significant increase in PCNA expression in cultured myometrial cells was noted following treatment with HB-EGF at concentrations >1 ng/ml, whereas an increase in PCNA expression in cultured leiomyoma cells was noted following treatment with HB-EGF at concentrations >10 ng/ml. HER1 expression was significantly higher in untreated myometrial cells than in untreated leiomyoma cells. A significant increase in HER1 expression in myometrial cells was observed when treated with HB-EGF at concentrations >10 ng/ml, whereas a significant increase in HER1 expression in leiomyoma cells was noted only by the treatment with HB-EGF at concentrations >100 ng/ml. Treatment with HB-EGF decreased the TUNEL-positive rate of those cells with no significant differences between the two cell types. The results obtained suggest that HB-EGF plays a role in stimulating the proliferation of leiomyoma cells and myometrial cells and in inhibiting apoptosis of those cells through augmentation of HER1 expression. Since the proliferative potential of myometrial cells responded better to HB-EGF than that of leiomyoma cells, HB-EGF may play a more vital role in myometrial growth than leiomyoma growth.

Progesterone and gravidity differentially regulate expression of extracellular matrix components in the pregnant rat myometrium.

Myometrial growth and remodeling during pregnancy depends on increased synthesis of interstitial matrix proteins. We hypothesize that the presence of mechanical tension in a specific hormonal environment regulates the expression of extracellular matrix (ECM) components in the uterus. Myometrial tissue was collected from pregnant rats on Gestational Days 0, 12, 15, 17, 19, 21, 22, 23 (labor), and 1 day postpartum and ECM expression was analyzed by Northern blotting. Expression of fibronectin, laminin beta2, and collagen IV mRNA was low during early gestation but increased dramatically on Day 23 during labor. Expression of fibrillar collagens (type I and III) peaked Day 19 and decreased near term. In contrast, elastin mRNA remained elevated from midgestation onward. Injection of progesterone (P4) on Days 20-23 (to maintain elevated plasma P4 levels) delayed the onset of labor, caused dramatic reductions in the levels of fibronectin and laminin mRNA, and prevented the fall of collagen III mRNA levels on Day 23. Treatment of pregnant rats with the progesterone receptor antagonist RU486 on Day 19 induced preterm labor on Day 20 and a premature increase in mRNA levels of collagen IV, fibronectin, and laminin. Analysis of the uterine tissue from unilaterally pregnant rats revealed that most of the changes in ECM gene expression occurred specifically in the gravid horn. Our results show a decrease in expression of fibrillar collagens and a coordinated temporal increase in expression of components of the basement membrane near term associated with decreased P4 and increased mechanical tension. These ECM changes contribute to myometrial growth and remodeling during late pregnancy and the preparation for the synchronized contractions of labor.

Myometrial thickness during human labor and immediately post partum

Objective: Morphologic studies suggest dramatic, asymmetric uterine growth during pregnancy that is caused by muscle cell hypertrophy. This growth is most marked at the fundus. Our objective was to evaluate sonographically the in vivo changes in myometrial thickness during active labor, second-stage labor, and after delivery. Study design: Abdominal ultrasound scans were performed on 52 term pregnant women to investigate the dynamic changes in myometrial thickness during the active and second stages of labor and immediately after delivery. Twenty-six women (mean ± SEM gestational age, 39.09 ± 0.3 weeks) were in active labor (cervical dilatation >4 cm with regular uterine contractions). An additional 26 nonlaboring women (gestational age, 39.8 ± 0.2 weeks) provided control measurements. The myometrium was defined sonographically as the echo homogeneous layer between the serosa and the decidua. Myometrial thickness was measured at the low segment and mid anterior, fundal, and posterior uterine walls by the same observer. Myometrial thickness was also measured during uterine contractions and after artificial rupture of the amniotic membranes. All laboring women had uncomplicated labor patterns when studied and were delivered spontaneously. Results: The myometrium was significantly thinner during active labor compared with nonlabor at each site studied: midanterior (mean [±SEM] myometrial thickness, 5.8 ± 0.27 vs 8.83 ± 0.51 mm; t test, P < .001), fundus (mean myometrial thickness, 6.78 ± 0.32 vs 8.49 ± 0.35 mm; P = .0015), and posterior (mean myometrial thickness, 6.22 ± 0.34 vs 8.12 ± 0.30 mm; P < .001). However, myometrial thickness did not differ among sites within the two groups. The thickness of the low segment was not affected by labor status (nonlabor, 4.68 ± 0.48 vs labor, 4.66 ± 0.37 mm; P = .97). Similarly, the myometrial thickness of the anterior uterine wall was unaffected by contractions (no contractions, 5.56 ± 0.2 vs contractions, 5.68 ± 0.22 mm; t test, P = .654). There was no change in myometrial thickness measured immediately before and after rupture of the amniotic membranes, despite a significant decrease of the amniotic fluid index. There was significant thickening of the anterior and fundal myometrium during the second stage of labor after the fetal head descended to +3 station by digital examination (anterior, 12.99 ± 0.60 vs 5.8 ± 0.27 mm; t test, P < .001; fundus, 10.61 ± 1.63 vs 6.78 ± 0.32 mm; t test, P = .04). Valsalva maneuver (pushing) during contractions did not affect myometrial thickness at the fundus (between contractions, 10.61 ± 1.63 vs pushing, 10.76 ± 1.95 mm; t test, P = .99). Immediately after delivery, the myometrial thickness at the placental insertion site was the thinnest. After completion of the third stage of labor, the uterine fundus remained significantly thinner than the anterior and posterior walls (fundus, 27.37 ± 3.5 mm vs anterior, 40.94 ± 3.5 vs posterior, 42.34 ± 2.44; one-way analysis of variance, P = .02). Conclusion: There is significant and widespread thinning of the myometrium during active labor. Descent of the fetal head during the second stage of labor is associated with a significant relative thickening of the anterior and fundal myometrium. After delivery, the relationship reverses. These findings suggest the directionality of the expulsive force vectors (fundal dominance) is not determined by asymmetric myometrial growth but, rather, may be a function of increased “myometrial mass” that results from increased surface area at the fundus. (Am J Obstet Gynecol 2003;188:553-9.)

Protein kinase Calpha is required for endothelin-1-induced proliferation of human myometrial cells.

The role of protein kinase C (PKC)-alpha in endothelin-1 (ET-1)-induced proliferation of human myometrial cells was investigated. Inhibition of conventional PKC with Gö 6976 eliminated the proliferative effect of ET-1. Treatment of myometrial cells with an antisense oligonucleotide against PKCalpha efficiently reduced PKCalpha protein expression without effect on other PKC isoforms and resulted in the loss of ET-1-induced cell growth. Immunocytochemistry using an antibody against PKCalpha revealed that there was no PKCalpha immunoreactivity in the nuclei of quiescent nonconfluent untreated cells, whereas it is evenly distributed throughout the cytoplasm. Exposure of myometrial cells to ET-1 for 15 min caused the PKCalpha to shift towards the perinuclear area, and incubation for 60 min caused a shift towards the nucleus. These results reveal that PKCalpha is required for ET-1-induced human myometrial cell growth and suggest that targeting of PKCalpha by antisense nucleotides might be an important approach for the development of anticancer treatments.

Differential Regulation of Protein Kinase C Isoforms in Human Uterine Leiomyoma

The protein kinase C (PKC) isoenzyme expression pattern in human uterine leiomyoma was compared with that obtained in homologous myometrium distal from the tumor. The six PKC isoforms (PKCα, PKCβ1, PKCβ2, PKCδ, PKCΕ and PKCζ) evidenced in the myometrium were found to be similarly expressed in leiomyoma. Quantitative immunoblotting revealed that all PKC isoforms were preferentially localized in the particulate fraction. To gain insight into the possible functional consequences of PKC expression patterns, subcellular redistribution in response to the mitogenic peptide endothelin-1 (ET-1) was studied. After stimulation with ET-1, differential redistribution occurred in leiomyoma and myometrium, suggesting a selective role of PKC isoforms in the myometrial growth process.

Corticotrophin Releasing Hormone: Its Potential for a Role in Human Myometrium

Aside from its role as a hypothalamic stress hormone, corticotrophin releasing hormone (CRH) is also a placental hormone, at least in primates. Although the function of placentally derived CRH remains to be fully elucidated, elevated CRH levels have been associated with premature labour, suggesting that the hormone may be involved in regulating the duration of pregnancy. Indeed, pregnant human myometrium expresses functional CRH receptors (CRH R1 and CRH R2 subtypes) thought to signal predominantly via the second messenger cAMP. Thus, like other cAMP-producing hormones in the myometrium such as beta(2) agonists, CRH may play a part in maintaining uterine quiescence. However, several of the CRH receptor isoforms identified to date have a reduced ability to activate adenylate cyclase, raising the question as to whether they are linked to other signal transduction pathways. Here, we discuss critically the evidence for the peptide's role in regulating contractility, both directly at the myometrium and indirectly via the fetal membranes and decidua. The possibility of a role in myometrial growth modulation is also described. Experimental Physiology (2001) 86.2, 273-281.

Uterine myoma in postmenopause: a comparison between two therapeutic schedules of HRT

Objectives: It is still controversial whether hormone replacement therapy (HRT) can affect the onset of uterine myomas or their growth in postmenopause. It is likely that some therapeutic schedules can influence the myometrial growth differently, due to a more potent stimulation of the uterine receptors. The aim of the present study is to evaluate the effects of two different hormonal treatment schedules on the risk of uterine myoma onset or progression. Methods: In a 2 year prospective randomised study we compared an oral cyclic association of oestradiol valerate and cyproterone acetate versus a sequential combination of transdermal E 2 and per oral medrossiprogesterone acetate on 240 postmenopausal women with and without uterine myomas. Results: Among the patients without uterine myomas treated with the transdermal–oral combination we noted the onset of myomas in 5% of cases after 24 months of treatment, while no new uterine formation was observed for the orally treated women ( P<0.01). Among the patients with uterine myomas at the begining of the study, in the group transdermally treated we found a mean increase in myoma volumes of 25.3% in the following 24 months, which was significantly different compared with the initial volume of myomas. On the other hand, women treated with the oral combination showed no significant modification of myoma volumes at the end of the study. Conclusions: Percutaneous-oral schedule of HRT seems to affect the growth of uterine myomas more than a single oral combination of oestradiol valerate and cyproterone acetate.