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Effects of progesterone treatment on expression of genes involved in uterine quiescence.

ABSTRACT: An important action of progesterone during pregnancy is to maintain the uterus in a quiescent state and thereby prevent preterm labor. The causes of preterm labor are not well understood, so progesterone action on the myometrium can provide clues about the processes that keep the uterus from contracting prematurely. Accordingly, we have carried out Affymetrix GeneChip analysis of progesterone effects on gene expression in immortalized human myometrial cells cultured from a patient near the end of pregnancy. Progesterone appears to inhibit uterine excitability by a number of mechanisms, including increased expression of calcium and voltage-operated K(+) channels, which dampens the electrical activity of the myometrial cell, downregulation of agents, and receptors involved in myometrial contraction, reduction in cell signal components that lead to increased intracellular Ca(2+) concentrations in response to contractile stimuli, and downregulation of proteins involved in the cross-linking of actin and myosin filaments to produce uterine contractions.

SUBMITTER: Soloff MS

PROVIDER: S-EPMC4051400 | biostudies-literature | 2011 Aug

REPOSITORIES: biostudies-literature

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Effects of progesterone treatment on expression of genes involved in uterine quiescence.

Soloff Melvyn S MS Jeng Yow-Jiun YJ Izban Michael G MG Sinha Mala M Luxon Bruce A BA Stamnes Susan J SJ England Sarah K SK

Reproductive sciences (Thousand Oaks, Calif.) 20110801 8

An important action of progesterone during pregnancy is to maintain the uterus in a quiescent state and thereby prevent preterm labor. The causes of preterm labor are not well understood, so progesterone action on the myometrium can provide clues about the processes that keep the uterus from contracting prematurely. Accordingly, we have carried out Affymetrix GeneChip analysis of progesterone effects on gene expression in immortalized human myometrial cells cultured from a patient near the end o ...[more]

PMID: 21795739

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Project description:The present investigation examined the spatiotemporal expression of estrogen receptors (ER-alpha and ER-beta) and progesterone receptor (PR) in the periimplantation mouse uterus (days 1-8). ER-alpha messenger RNA (mRNA) was detected at much higher levels in the periimplantation uterus compared with that of ER-beta mRNA, the levels of which were very low in all uterine cells during this period. Results of in situ hybridization demonstrated expression of ER-alpha mRNA primarily in the luminal and glandular epithelia on days 1 and 2 of pregnancy. On days 3 and 4, the accumulation was localized primarily in stromal cells in addition to its presence in the epithelium. Following implantation on day 5, the accumulation of this mRNA was more condensed in the luminal and glandular epithelia, but declined in the subluminal epithelial stroma at the sites of implanting embryos. On days 6-8, the accumulation of ER-alpha mRNA was primarily localized in the secondary decidual zone (SDZ) with more intense localization in the subepithelial cells at the mesometrial pole. In contrast, signals were very low to undetectable in the primary decidual zone (PDZ), and no signals were detected in implanting embryos. The undifferentiated stroma underneath the myometrium also showed positive signals. The immunolocalization of ER-alpha protein correlated with the mRNA localization. Western blot analysis showed down-regulation of ER-alpha in day 8 decidual cell extracts consistent with the down-regulation of ER-alpha mRNA in decidual cells immediately surrounding the embryo on this day. The expression pattern of PR was also dynamic in the periimplantation uterus. On day 1, the accumulation of PR mRNA was very low to undetectable, whereas only a modest level of accumulation in the epithelium was noted on day 2. On days 3 and 4, the accumulation of this mRNA was detected in both the epithelium and stroma. In contrast, the expression was restricted only to the stroma with increased signals at the sites of implantation on day 5. On days 6-8, PR mRNA accumulation increased dramatically throughout the deciduum. The localization of immunoreactive PR correlated with the mRNA distribution in the periimplantation uterus. Taken together, the results demonstrate that the expression of ER-alpha, ER-beta, and PR is differentially regulated in the periimplantation mouse uterus. This compartmentalized expression of ER and PR provides information regarding the sites of coordinated effects ofestrogen and progesterone in the preparation of the uterus for implantation and decidualization during early pregnancy.

Project description:BackgroundIn the non-pregnant dog, ovarian cyclicity is independent of a uterine luteolysin. This is in contrast to pregnant animals where a prepartum increase of luteolytic PGF2α occurs, apparently originating in the pregnant uterus. Recently, the placenta as a source of prepartum prostaglandins (PGs) was investigated, indicating fetal trophoblast cells as the likely main source. However, the possible contribution of uterine interplacental tissues to the production of these hormones has not yet been thoroughly examined in the dog.MethodsSeveral key factors involved in the production and/or actions of PGs were studied: cyclooxygenase 2 (COX2, PTGS2), PGF2α-synthase (PGFS/AKR1C3), PGE2-synthase (PGES), and the respective receptors FP (PTGFR), EP2 (PTGER2) and EP4 (PGTER4), 15-hydroxyprostaglandin dehydrogenase (HPGD), PG-transporter (PGT, SLCO2A1) and progesterone receptor. Their expression and localization patterns were assessed by Real Time PCR and immunohistology in the interplacental uterine sites from pregnant dogs during the pre-implantation period (days 8-12), post-implantation (days 18-25), mid-gestation (days 35-40) and during antigestagen-induced luteolysis/abortion.ResultsWhereas only low COX2 expression was observed in uterine samples at all the selected time points, expression of PGFS/AKR1C3 strongly increased post-implantation. A gradual increase in PGES-mRNA expression was noted towards mid-gestation. FP-mRNA expression decreased significantly with the progression of pregnancy until mid-gestation. This was associated with clearly detectable expression of HPGD, which did not change significantly over time. The expression of FP and EP2-mRNA decreased significantly over time while EP4-mRNA expression remained unaffected. The antigestagen-treatment led to a significant increase in expression of COX2, PGES, EP2 and PGT (SLCO2A1) mRNA. COX2 was localized predominantly in the myometrium. The expression of PGFS/AKR1C3, which was unchanged, was localized mostly to the surface luminal epithelium. The expression of EP4, PGT and HPGH did not change during treatment, they were co-localized with PGES and EP2 in all uterine compartments.ConclusionsThe data clearly demonstrate the basic capability of the canine pregnant uterus to produce and respond to PGs and suggests their functions both as local regulatory factors involved in the establishment and maintenance of pregnancy, as well as potential contributors to the process of parturition, supporting the myometrial contractility associated with fetal expulsion.

Project description:This study evaluated the effects of treatment with meloxicam (a non-steroidal anti-inflammatory drug), parity, and blood progesterone concentration on the dynamics of the uterine microbiota of 16 clinically healthy postpartum dairy cows. Seven primiparous and 9 multiparous postpartum Holstein cows either received meloxicam (0.5 mg/kg SC, n = 7 cows) once daily for 4 days (10 to 13 days in milk (DIM)) or were untreated (n = 9 cows). Endometrial cytology samples were collected by cytobrush at 10, 21, and 35 DIM, from which the microbiota analysis was conducted using 16S rRNA gene sequence analysis. A radioimmunoassay was used to measure progesterone concentration in blood serum samples at 35 DIM and cows were classified as ˃ 1 ng/mL (n = 10) or ≤ 1 ng/mL (n = 6). Alpha diversity for bacterial genera (Chao1, Shannon-Weiner, and Camargo's evenness indices) were not affected by DIM, meloxicam treatment, parity, or progesterone category. For beta diversity (genera level), principal coordinate analysis (Bray-Curtis) showed differences in microbiota between parity groups. At the phylum level, the relative abundance of Actinobacteria was greater in primiparous than multiparous cows. At the genus level, there was lesser relative abundance of Bifidobacterium, Lactobacillus, Neisseriaceae, Paracoccus, Staphylococcus, and Streptococcus and greater relative abundance of Bacillus and Fusobacterium in primiparous than multiparous cows. Bray-Curtis dissimilarity did not differ by DIM at sampling, meloxicam treatment, or progesterone category at 35 DIM. In conclusion, uterine bacterial composition was not different at 10, 21, or 35 DIM, and meloxicam treatment or progesterone category did not affect the uterine microbiota in clinically healthy postpartum dairy cows. Primiparous cows presented a different composition of uterine bacteria than multiparous cows. The differences in microbiota associated with parity might be attributable to changes that occur consequent to the first calving, but this hypothesis should be investigated further.

Project description:The myometrium is an important reproductive tissue composed primarily of smooth muscle cells. Contractility of the myometrial smooth muscle cells during pregnancy and labour is modulated by hormones. Despite much research, little is known about the molecular mechanism by which estrogen and progesterone regulate myometrial contractility. This study investigates global gene expression profile of cultured human uterine smooth muscle cells (hUtSMCs) following 17Î²-estradiol (E2) and/or progesterone treatments using cDNA microarray technology. Furthermore the effects of addition of the progesterone inhibitor RU486 were investigated. Many genes were regulated in the presence of P4 or E2 alone but almost six times these numbers were regulated in the presence of the combination. The majority of these genes returned to near baseline levels (control samples) on addition of RU486. In total 796 annotated genes were significantly differentially expressed in the combined presence of P4 and E2 (relative to untreated levels). Furthermore, 666 annotated genes were significantly regulated by the addition of RU486 to the P4/E2 combination (relative to P4/E2 levels). Gene ontology analysis of these differentially expressed genes revealed a strong association of P4/E2 treatment with the downregulated expression of genes involved in cell communication, signal transduction, channel activity, inflammatory response and differentiation. Upregulated processes included cell survival (anti-apoptosis), gene transcription, steroid hormone biosynthesis, muscle development, insulin receptor signaling and cell growth. Functional withdrawal of progesterone using RU486 effectively reversed the processes induced by P4/E2 treatment. The hUtSMC system is an additional useful model to investigate steroid effects on smooth muscle cells in isolation from other myometrial cell types. In this study human myometrial smooth muscle cells (hUtSMCs) purchased from Lonza were cultured and treated with 17beta estradiol and progesterone either alone or in combination. In addition the 17beta estradiol and progesterone combined treated samples were also treated with the progesterone inhibitor RU486 (mifepristone). RNA was isolated after 72h from both treated and untreated control cells. The experiment was repeated 3 times. Please note that slides were scanned three times successively (i.e. 3 .gpr files per sample) and the median raw values were used as input raw data for GeneSpring analysis (Median_Table_InputRAW.xlsx).

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Effects of progesterone treatment on expression of genes involved in uterine quiescence.

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Effects of progesterone treatment on expression of genes involved in uterine quiescence.

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