Performance of polymyxin B agar-based tests among carbapenem-resistant Enterobacterales.

Therapeutic options for infections caused by Carbapenem-resistant Enterobacterales (CRE) are restricted and include polymyxins-centred schemes. Evaluation of in vitro susceptibility is difficult and time consuming. Agar-based methodologies are an alternative to broth microdilution (BMD) and we aimed to evaluate the accuracy of those methods among Enterobacterales. A total of 137 non-duplicated CRE were subjected to polymyxin B BMD, agar screening test (Mueller Hinton plates containing 3 µg ml-1 of polymyxin B) and agar dilution (antibiotic serially diluted 0·25-64 µg ml-1 ). CRE of 42·3% were resistant to polymyxin B (MICs range: 0·25->64 µg ml-1 ) and 16·8% presented borderline MICs. Sensitivity, specificity, PPV and NPV were 86·2, 98·7, 98 and 90·7% for screening test and 86·2, 97·5, 96·1 and 90·6% for agar dilution. ME was 0·73 and 1·5% for screening and agar dilution respectively; VME was 5·8% for both techniques. In general, agar-based methods had a good performance. As far as we know, this is the first study to propose an agar screening test using polymyxin B instead of colistin.

A randomised clinical pilot trial to test the effectiveness of parent training with video modelling to improve functioning and symptoms in children with autism spectrum disorders and intellectual disability.

BACKGROUND: Poor eye contact and joint attention are early signs of autism spectrum disorder (ASD) and important prerequisites for developing other socio-communicative skills. Teaching parents evidence-based techniques to improve these skills can impact the overall functioning of children with ASD. We aimed to analyse the impact of conducting a group parent-training intervention with video modelling to improve the intelligent quotient (IQ), social and communication functioning and to minimise symptoms in children with ASD and intellectual disability (ID). METHODS: Study design: A multicentre, single-blinded, randomised clinical pilot trial of parent training using video modelling was conducted. SAMPLE: Sixty-seven parents of children with ASD, aged between 3 and 6 years and with IQs between 50 and 70, were randomised: 34 to the intervention group and 33 to the control group. Intervention program: The intervention group received parent training over 22 sessions, and the control group received the standard community treatment. INSTRUMENTS: Pre-evaluation and post-evaluation (week 28), the following were used: Autism Diagnostic Interview, Vineland Adaptive Behaviour Scale I, Snijders-Oomen Nonverbal Intelligence Test, Autism Behaviour Checklist and Hamilton Depression Rating Scale. DATA ANALYSIS: Intention to treat and complier-average causal effect (CACE) were used to estimate the effects of the intervention. RESULTS: There was a statistically significant improvement in the Vineland standardized communication scores in CACE (Cohen's d = 0.260). There was a non-statistically significant decrease in autism symptomatology (Autism Behaviour Checklist total scores) and a significant increase in the non-verbal IQ in the intervention group. After the false discovery rate correction was applied, IQ remained statistically significant under both paradigms. The effect size for this adjusted outcome under the intention-to-treat paradigm was close to 0.4, and when considering adherence (CACE), the effect sizes were more robust (IQ's Cohen's d = 0.433). CONCLUSIONS: Parent training delivered by video modelling can be a useful technique for improving the care given to children with ASD and ID, particularly in countries that lack specialists.

Emergence of serotype 19A Streptococcus pneumoniae after PCV10 associated with a ST320 in adult population, in Porto Alegre, Brazil.

Use of pneumococcal conjugate vaccines has caused emergence of non-vaccine serotypes. No Brazilian data specifically about serotype 19A are available. We aimed to evaluate the frequency of occurrence, susceptibility profile and molecular epidemiology of serotype 19A before and after vaccine introduction in Brazil. Pneumococcal identification was performed by the conventional method. Strain serotype was determined by multiplex polymerase chain reaction (PCR) and/or Quellung reaction. Resistance was determined by Etest® and PCR was performed to determine the presence of macrolide resistance genes, ermB and/or mefA. Pneumococci were typed by Multilocus Sequence Typing. Thirty-eight serotype 19A Streptococcus pneumoniae were recovered, mostly from invasive diseases. Prevalence of serotype 19A increased following vaccination (from 3.5% before vaccination to 8.1% after, p = 0.04196). Non-susceptibility increased to most antimicrobials after vaccine introduction and was associated with clonal complex (CC)320. MLST showed nine different STs, which were grouped in one main CC: CC320 (63.9%). During the post-vaccination era, the frequency of this serotype increased significantly from 1.2% in 2011 to 18.5% in 2014 (p = 0.00001), with a concomitant decrease in the genetic variability: ST320 consistently predominated after vaccine-introduction (61.1%). Overall, our results showed a post-PCV10 increase in the frequency of serotype 19A. This was accompanied by a selection of CC320 and antimicrobial resistance.

Androgen signaling is a confounding factor for ß-catenin-mediated prostate tumorigenesis.

Emerging evidence has demonstrated the critical roles for both androgen and Wnt pathways in prostate tumorigenesis. A recent integrative genomic analysis of human prostate cancers (PCas) has revealed a unique enrichment of androgen and Wnt signaling in early-onset PCas, implying their clinical significance in the disease. Additionally, interaction between the androgen receptor (AR) and ß-catenin has long been detected in PCa cells. However, the consequence of this interaction in prostate tumorigenesis is still unknown. Because mutations in adenomatous polyposis coli, ß-catenin and other components of the destruction complex are generally rare in PCas, other mechanisms of aberrant Wnt signaling activation have been speculated. To address these critical questions, we developed Ctnnb1(L(ex3)/+)/R26hAR(L/+):PB-Cre4 mice, in which transgenic AR and stabilized ß-catenin are co-expressed in prostatic epithelial cells. We observed accelerated tumor development, aggressive tumor invasion and a decreased survival rate in Ctnnb1(L(ex3)/+)/R26hAR(L/+):PB-Cre4 compound mice compared with age-matched Ctnnb1(L(ex3)/+):PB-Cre4 littermate controls, which only have stabilized ß-catenin expression in the prostate. Castration of the above transgenic mice resulted in significant tumor regression, implying an essential role of androgen signaling in tumor growth and maintenance. Implantation of the prostatic epithelial cells isolated from the transgenic mice regenerated prostate intraepithelial neoplasias and prostatic adenocarcinoma lesions. Microarray analyses of transcriptional profiles showed more robust enrichment of known tumor- and metastasis-promoting genes: Spp1, Egr1, c-Myc, Sp5, and Sp6 genes, in samples isolated from Ctnnb1(L(ex3)/+)/R26hAR(L/+):PB-Cre4 compound mice than those from Ctnnb1(L(ex3)/+):PB-Cre4 and R26hAR(L/+):PB-Cre4 littermate controls. Together, these data demonstrate a confounding role of androgen signaling in ß-catenin-initiated oncogenic transformation in prostate tumorigenesis.

Inflammation, neurotrophism and oxidative stress and childhood psychopathology in a large community sample.

OBJECTIVE: To investigate the association between peripheral biomarkers and child psychopathology in a large community sample. METHOD: A total of 625 aged 6- to 13-year old subjects were recruited from a community school-based study. Psychopathology was assessed using the Child Behaviour Checklist (CBCL). Psychiatric diagnosis was evaluated using the Development and Well-Being Assessment. The following biomarkers were examined in peripheral blood: brain-derived neurotrophic factor, cytokines (IL-2, IL-4, IL-6, IL-10, IL-17, IFN-g, and TNF-α), chemokines (eotaxin/CCL11, IP-10, MCP-1), cytokine receptors (sTNFR1 and sTNFR2), and the oxidative stress marker TBARS. RESULTS: We found significant associations between sTNFR2, eotaxin/CCL11 and CBCL total score, as well as with specific dimensions of psychopathology. There were different patterns of association between these biomarkers and psychological and behavioural symptoms in children with and without a mental disorder. TBARS, IL-6 and MCP-1 were more specific to some clusters of symptoms in children with a psychiatric diagnosis. CONCLUSION: Our data support the potential use of biomarkers, especially those involved in immune-inflammatory pathways, in investigating neurodevelopmental psychopathology. Their association with different dimensions of symptoms might be of useful when analyzing illness severity and clusters of symptoms within specific disorders.

The power and perils of animal models with urogenital anomalies: handle with care.

Congenital abnormalities of the urogenital tracts form a major part of clinical practice for paediatric urologists, but their knowledge of normal and abnormal development is often limited. Advances in understanding frequently come from studying experimental findings from animal models, however, most clinicians underestimate both the power and perils of extrapolating scientific knowledge from animals. In this review, the key issues that urologists need to understand in order to link animal studies to clinical practice are discussed. Urologists must avoid the traps of anthropomorphism (assuming humans are always the same as animal models) or anthropocentrism (assuming humans are too different from animal models). This review used two common disorders: hypospadias and undescended testes.

Endocrine differentiation of fetal ovaries and testes of the spotted hyena (Crocuta crocuta): timing of androgen-independent versus androgen-driven genital development.

Female spotted hyenas (Crocuta crocuta) have an erectile peniform clitoris and a pseudoscrotum but no external vagina, all established by day 35 of a 110-day gestation. Recent studies indicate that these events are androgen-independent, although androgen secretion by fetal ovaries and testis was hypothesized previously to induce phallic development in both sexes. We present the first data relating to the capacity of the ovaries and testes of the spotted hyena to synthesize androgens at different stages of fetal life. Specifically, spotted hyena fetal gonads were examined by immunohistochemistry at GD 30, 45, 48, 65, and 95 for androgen-synthesizing enzymes, as related to the morphological development. Enzymes included 17alpha-hydroxylase/17,20-lyase cytochrome P450 (P450c17), cytochrome b5, 3beta-hydroxysteroid dehydrogenase (3betaHSD), and cholesterol side-chain cleavage cytochrome P450 (P450scc). Anti-Müllerian-hormone (AMH) expression was also examined. AMH was strongly expressed in fetal Sertoli cells from GD 30 and after. P450c17 expression was detected in Leydig cells of developing testes and surprisingly in Müllerian duct epithelium. Fetal ovaries began to organize and differentiate by GD 45, and medullary cells expressed P450c17, cytochrome b5, 3betaHSD, and P450scc. The findings support the hypothesis that external genital morphology is probably androgen-independent initially, but that fetal testicular androgens modify the secondary, male-specific phallic form and accessory organs. Fetal ovaries appear to develop substantial androgen-synthesizing capacity but not until phallic differentiation is complete, i.e. after GD 45 based on circulating androstenedione concentrations. During late gestation, fetal ovaries and testes synthesize androgens, possibly organizing the neural substrates of aggressive behaviors observed at birth in spotted hyenas. These data provide an endocrine rationale for sexual dimorphisms in phallic structure and reveal a potential source of androgenic support for neonatal aggression in female and male C. crocuta.

Induction of apoptosis by castration in epithelium of the mouse seminal vesicles.

Castration on days 0, 5, 10, 20, 40, and 60 caused increases in an apoptotic index (% of apoptotic cells) in seminal vesicle (SV) epithelium, peaking 1-3 days after castration. The peak apoptotic indices after castration on days 0, 5, 10, and 20 were significantly lower than peak apoptotic indices observed after castration on days 40 and 60. DNA extracted from mouse SVs 2 days after castration on days 0, 5, 10, and 60 showed a ladder pattern on agarose gel electrophoresis. The secretion of androgen by testes was confirmed by the growth retardation of the SVs after castration on days 0, 5, 10, and 20. It would appear that a proportion of SV epithelial cells dependent on testicular androgens for survival is smaller before day 20 than after day 20.

Variation in ovarian morphology in four species of New World moles with a peniform clitoris.

Female moles of the Old World genus Talpa display a curious suite of reproductive features that include a peniform clitoris and ovaries with a discrete interstitial gland or testis-like region (so-called 'ovotestes'). The masculinization of the female external genitalia in Talpa has accordingly been linked with secretion of androgens from the interstitial gland region of the fetal gonad. Although their ovarian morphology has received less attention, some species of New World moles also have ovaries with a pronounced interstitial gland (for example star-nosed mole, Condylura cristata), whereas females of other species do not (for example eastern mole, Scalopus aquaticus). Although it is difficult to determine the sex of both Old and New World moles, published accounts describing the external genitalia of female moles are available only for Talpa. The hypothesis that masculinization of the female external genitalia in moles is associated with the presence of an ovarian interstitial gland (OIG) was tested in the present study by using a comparative approach to determine whether these features are ever found in isolation of one another. Three genera of North American moles (Scapanus, Condylura and Neurotrichus) were studied and a peniform clitoris was found in all three species, but OIG were found in only two of three genera. The ovaries of S. latimanus and S. orarius were unremarkable, with no evidence of a discrete interstitial gland or testis-like region. Mapping these results onto recent talpid phylogenies indicates that loss of the bipolar ovarian morphology is a derived trait in Scapanus, and conclusively demonstrates that masculinization of the external genitalia in female moles can develop in the presence or absence of 'ovotestes'.

Malignant transformation in a nontumorigenic human prostatic epithelial cell line.

The human prostatic epithelial cell line BPH-1 is normally nontumorigenic in nude mice. The present report demonstrates that this cell line can be permanently transformed by its microenvironment to become tumorigenic. The establishment of a series of tumorigenic sublines based on this parental cell line is described. BPH-1 cells were induced to form tumors either by recombination with human prostatic carcinoma-associated fibroblasts (CAFs) or by exposure to carcinogenic doses of testosterone and estradiol (T+E2) after recombination with rat urogenital sinus mesenchyme. Epithelial cells isolated from these tumors were established as cell strains in culture. When regrafted to nude mouse hosts epithelial cells isolated from CAF- or T+E2-induced tumors were found to be consistently tumorigenic even in the absence of CAF or T+E2. The T+E2-induced cell strains have been designated BPH1(TETD)-A and -B and the CAF-induced strains are designated BPH1(CAFTD)-01 through -08. In vitro, the cells had an epithelial morphology with a less well-defined cobblestone pattern than the parental line. They express SV40 large T antigen, confirming their derivation from the parental BPH-1 line. The BPH1(CAFTD) strains formed colonies in soft agar, whereas the parental BPH-1 cells and the BPH1(TETD) sublines did not. There was no immunocytochemically detectable expression of androgen (AR), alpha-estrogen (ERalpha), or progesterone (PR) receptors by the parental BPH-1 cell line or by any of the tumor-derived cell strains. The cells uniformly coexpressed both basal and luminal cell-type cytokeratins and the basal cell marker p63. When grafted beneath the renal capsule of athymic mouse hosts, all of the tumor-derived cell strains consistently formed tumors. These were predominantly poorly or moderately differentiated squamous or adenosquamous tumors, similar in organization to the primary tumors from which the cell strains were derived. The cell strains continued to express both basal- and luminal-type cytokeratins in vivo. Some of the cell strains also coexpressed vimentin. E-cadherin expression was absent from many of the cells, although patches of cells expressing this marker were seen. The cells continued to express SV40T antigen. These cell strains, which are all derived from a common nontumorigenic progenitor, represent a useful resource for examining genetic and phenotypic changes during carcinogenesis.

The consequences of chromosomal aneuploidy on gene expression profiles in a cell line model for prostate carcinogenesis.

Here we report the genetic characterization of immortalized prostate epithelial cells before and after conversion to tumorigenicity using molecular cytogenetics and microarray technology. We were particularly interested to analyze the consequences of acquired chromosomal aneuploidies with respect to modifications of gene expression profiles. Compared with nontumorigenic but immortalized prostate epithelium, prostate tumor cell lines showed high levels of chromosomal rearrangements that led to gains of 1p, 5, 11q, 12p, 16q, and 20q and losses of 1pter, 11p, 17, 20p, 21, 22, and Y. Of 5700 unique targets on a 6.5K cDNA microarray, approximately 3% were subject to modification in expression levels; these included GRO-1, -2, IAP-1,- 2, MMP-9, and cyclin D1, which showed increased expression, and TRAIL, BRCA1, and CTNNA, which showed decreased expression. Thirty % of expression changes occurred in regions the genomic copy number of which remained balanced. Of the remainder, 42% of down-regulated and 51% of up-regulated genes mapped to regions present in decreased or increased genomic copy numbers, respectively. A relative gain or loss of a chromosome or chromosomal arm usually resulted in a statistically significant increase or decrease, respectively, in the average expression level of all of the genes on the chromosome. However, of these genes, very few (e.g., 5 of 101 genes on chromosome 11q), and in some instances only two genes (MMP-9 and PROCR on chromosome 20q), were overexpressed by > or =1.7-fold when scored individually. Cluster analysis by gene function suggests that prostate tumorigenesis in these cell line models involves alterations in gene expression that may favor invasion, prevent apoptosis, and promote growth.

Urethral seam formation and hypospadias.

Knowledge of the formation of the normal male urethra may elucidate the etiology of hypospadias. We describe urethral formation in the mouse, show the similarities and relevance to human urethral development, and introduce the concept of the epithelial seam formation and remodeling during urethral formation. Three mechanisms may account for epithelial seam formation: (1) epithelial-mesenchymal transformation similar to that described in the fusion of the palatal shelves, (2) apoptosis, and/or (3) tissue remodeling via cellular migration. Urethral development in the embryonic mouse (14-21 days of gestation) was compared with urethral formation in embryonic human specimens (8-16 weeks of gestation) by using histology, immunohistochemistry, and three-dimensional reconstruction. The urethra forms by fusion of the epithelial edges of the urethral folds, giving a midline epithelial seam. The epithelial seam is remodeled via cellular migration into a centrally located urethra and ventrally displaced remnant of epithelial cells. The epithelial seam is remodeled by narrowing approximately at its midpoint, with subsequent epithelial migration into the urethra or penile skin. The epithelial cells are replaced by mesenchymal cells. This remodeling seam displays a narrow band (approximately 30 microns wide) of apoptotic activity corresponding to the mesenchymal cells and not to epithelial cells. No evidence was seen of the co-expression of cytokeratin and mesenchymal markers (actin or vimentin). Urethral seam formation occurs in both the mouse and the human. Our data in the mouse support the hypothesis that seam transformation occurs via cellular migration and not by epithelial mesenchymal transformation or epithelial apoptosis. We postulate that disruption of epithelial fusion remodeling, and cellular migration leads to hypospadias.

A human prostatic epithelial model of hormonal carcinogenesis.

The effects of stromal and hormonal environment on the immortalized but nontumorigenic human prostatic epithelial cell line BPH-1 were investigated in an in vivo model. BPH-1 cells were recombined with rat urogenital sinus mesenchyme (UGM), and the tissue recombinants were grafted to the renal capsule of adult male athymic mouse hosts. BPH-1 + UGM recombinants formed solid branching epithelial cords with a well-defined basement membrane. The cords canalized to form ductal structures. The mesenchymal cells formed thick sheets of well-differentiated smooth muscle surrounding the epithelium, reinforcing the idea that the epithelium dictates the patterning of prostatic stromal cells. When hosts carrying BPH-1 + UGM tissue recombinants were exposed to testosterone propionate and 17-beta-estradiol (T + E2), the tissue recombinants responded by forming invasive carcinomas, demonstrating mixed, predominantly squamous as well as adenocarcinomatous (small acinar and mucinous) differentiation. When either untreated or T + E2-treated hosts were castrated, epithelial apoptosis was observed in the grafts. When tumors were removed and regrafted to fresh hosts they grew rapidly. Tumors were serially regrafted through six generations. Histologically these tumors consisted largely of focally keratinizing squamous cell carcinoma with high-grade malignant cytological features. BPH-1 cells grown in the absence of UGM survived at the graft site but did not form tumors or organized structures. This behavior was not influenced by the presence or absence of T + E2 stimulation. These data show that an immortalized, nontumorigenic human prostatic epithelial cell line can undergo hormonal carcinogenesis in response to T + E2 stimulation. In addition, the data demonstrate that the stromal environment plays an important role in mediating hormonal carcinogenesis.

Regulation of prostate branching morphogenesis by activin A and follistatin.

Ventral prostate development occurs by branching morphogenesis and is an androgen-dependent process modulated by growth factors. Many growth factors have been implicated in branching morphogenesis including activins (dimers of beta(A) and beta(B) subunits); activin A inhibited branching of lung and kidney in vitro. Our aim was to examine the role of activins on prostatic development in vitro and their localization in vivo. Organ culture of day 0 rat ventral prostates for 6 days with activin A (+/- testosterone) inhibited prostatic branching and growth without increasing apoptosis. The activin-binding protein follistatin increased branching in vitro in the absence (but not presence) of testosterone, suggesting endogenous activins may reduce prostatic branching morphogenesis. In vivo, inhibin alpha subunit was not expressed until puberty, therefore inhibins (dimers of alpha and beta subunits) are not involved in prostatic development. Activin beta(A) was immunolocalized to developing prostatic epithelium and mesenchymal aggregates at ductal tips. Activin beta(B) immunoreactivity was weak during development, but was upregulated in prostatic epithelium during puberty. Activin receptors were expressed throughout the prostatic epithelium. Follistatin mRNA and protein were expressed throughout the prostatic epithelium. The in vitro evidence that activin and follistatin have opposing effects on ductal branching suggests a role for activin as a negative regulator of prostatic ductal branching morphogenesis.

Paracrine regulation of apoptosis by steroid hormones in the male and female reproductive system.

In males, androgens are essential in maintaining the integrity of the prostate. Androgen-ablation induces apoptosis of the prostatic epithelium. In females, ovariectomy induces apoptosis in uterine epithelium while progesterone inhibits this process. The objective of this study was to determine whether androgen and progesterone inhibit apoptosis, respectively, in mouse prostatic and uterine epithelia via steroid receptors in the epithelium or in the stroma. To address this question, prostatic tissue recombinants were prepared with rat urogenital sinus mesenchyme plus bladder epithelium from wild-type or testicular feminization mutant (Tfm) mice. Thus, prostatic tissue was generated having androgen receptor (AR) in both epithelium and stroma or in the stroma only. Castration of hosts induced apoptosis in the AR-negative Tfm prostatic epithelium with an epithelial apoptotic index virtually identical to prostatic tissue recombinants containing wild-type epithelium. Moreover, this castration-induced prostatic epithelial apoptosis was blocked by testosterone and dihydrotestosterone in both wild-type and Tfm prostatic tissue recombinants. Likewise, uterine tissue recombinants were prepared in which epithelium and/or stroma was devoid of progesterone receptor (PR) by using uterine epithelium and stroma of wild-type and PR knockout mice. Progesterone inhibited uterine epithelial apoptosis only in tissue recombinants prepared with PR-positive stroma. The PR status of the epithelium did not affect epithelial apoptotic index. Therefore, the apoptosis in prostatic and uterine epithelia is regulated by androgen and progesterone via stromal AR and PR, respectively. In both cases, epithelial AR or PR is not required for hormonal regulation of epithelial apoptosis in prostatic and uterine epithelium.

fucosyltransferase1 and H-type complex carbohydrates modulate epithelial cell proliferation during prostatic branching morphogenesis.

The prostate undergoes branching morphogenesis dependent on paracrine interactions between the prostatic epithelium and the urogenital mesenchyme. To identify cell-surface molecules that function in this process, monoclonal antibodies raised against epithelial cell-surface antigens were screened for antigen expression in the developing prostate and for their ability to alter development of prostates grown in serum-free organ culture. One antibody defined a unique expression pattern in the developing prostate and inhibited growth and ductal branching of cultured prostates by inhibiting epithelial cell proliferation. Expression cloning showed that this antibody binds fucosyltransferase1, an alpha-(1,2)-fucosyltransferase that synthesizes H-type structures on the complex carbohydrate modifications of some proteins and lipids. The lectin UEA I that binds H-type 2 carbohydrates also inhibited development of cultured prostates. These data demonstrate a previously unrecognized role for fucosyltransferase1 and H-type carbohydrates in controlling the spatial distribution of epithelial cell proliferation during prostatic branching morphogenesis. We also show that fucosyltransferase1 is expressed by epithelial cells derived from benign prostatic hyperplasia or prostate cancer; thus, fucosyltransferase1 may also contribute to pathological prostatic growth. These data further suggest that rare individuals who lack fucosyltransferase1 (Bombay phenotype) should be investigated for altered reproductive function and/or altered susceptibility to benign prostatic hyperplasia and prostate cancer.

Epithelial-stromal interactions and tumor progression: meeting summary and future directions.

The Epithelial-Stromal Interactions Workshop was organized with the purpose of accelerating progress in understanding the interrelationship between tumor cells and their microenvironment and applying this knowledge to the control of tumor progression. The format of the meeting was the presentation of brief reports that focused on concepts rather than specifics, with extensive discussion periods to identify the issues and barriers hindering progress in this area. This report summarizes the findings of this meeting, highlighting the intimate relationship between tumor cells and their environment and addressing the opportunities that manipulation of host-tumor interactions has for therapeutic intervention. Several specific recommendations are made to advance knowledge and progress in this field.

Regulation of progesterone receptors and decidualization in uterine stroma of the estrogen receptor-alpha knockout mouse.

Regulation of progesterone receptor (PR) in uterine stroma (endometrial stroma plus myometrium) by estrogen was investigated in estrogen receptor-alpha (ERalpha) knockout (alphaERKO) mice. 17 beta-Estradiol (E(2)) increased PR levels in uterine stroma of ovariectomized alphaERKO mice, and ICI 182 780 (ICI) inhibited this E(2)-induced PR expression. Estrogen receptor-beta(ER beta) was detected in both uterine epithelium and stroma of wild-type and alphaERKO mice by immunohistochemistry. In organ cultures of alphaERKO uterus, both E(2) and diethylstilbestrol induced stromal PR, and ICI inhibited this induction. These findings suggest that estrogen induces stromal PR via ERbeta in alphaERKO uterus. However, this process is not mediated exclusively by ERbeta+, because in ERbeta knockout mice, which express ERalpha, PR was up-regulated by E(2) in uterine stroma. In both wild-type and alphaERKO mice, progesterone and mechanical traumatization were essential and sufficient to induce decidual cells, even though E(2) and ERalpha were also required for increase in uterine weight. Progesterone receptor was strongly expressed in decidual cells in alphaERKO mice, and ICI did not inhibit decidualization or PR expression. This study suggests that up-regulation of PR in endometrial stroma is mediated through at least three mechanisms: 1) classical estrogen signaling through ERalpha, 2) estrogen signaling through ERbeta, and 3) as a result of mechanical stimulation plus progesterone, which induces stromal cells to differentiate into decidual cells. Each of these pathways can function independently of the others.