Differentiation of patient-specific void urine-derived human induced pluripotent stem cells to fibroblasts and skeletal muscle myocytes.

Cell-based therapy is a major focus for treatment of stress urinary incontinence (SUI). However, derivation of primary cells requires tissue biopsies, which often have adverse effects on patients. A recent study used human induced pluripotent stem cells (iPSC)-derived smooth muscle myocytes for urethral sphincter regeneration in rats. Here, we establish a workflow using iPSC-derived fibroblasts and skeletal myocytes for urethral tissue regeneration: (1) Cells from voided urine of women were reprogrammed into iPSC. (2) The iPSC line U1 and hESC line H9 (control) were differentiated into fibroblasts expressing FSP1, TE7, vinculin, vimentin, αSMA, fibronectin and paxillin. (3) Myogenic differentiation of U1 and H9 was induced by small molecule CHIR99021 and confirmed by protein expression of myogenic factors PAX7, MYOD, MYOG, and MF20. Striated muscle cells enriched by FACS expressed NCAM1, TITIN, DESMIN, TNNT3. (4) Human iPSC-derived fibroblasts and myocytes were engrafted into the periurethral region of RNU rats. Injected cells were labelled with ferric nanoparticles and traced by Prussian Blue stain, human-specific nuclear protein KU80, and human anti-mitochondria antibody. This workflow allows the scalable derivation, culture, and in vivo tracing of patient-specific fibroblasts and myocytes, which can be assessed in rat SUI models to regenerate urethral damages and restore continence.

Pilot implementation of Bukhali: A preconception health trial in South Africa.

OBJECTIVES: This article describes the learnings from the pilot phase of the Healthy Life Trajectories Initiative, a preconception health trial for 18- to 25-year-old women in Soweto, South Africa. METHODS: The study compares two arms focussed on either physical and mental health (intervention; delivered by community health workers - 'Health Helpers') or standard of care plus (control; standard access to healthcare plus additional telephonic input on 'life skills'; delivered by call centre assistants). These are collectively referred to as Bukhali. Data on the pilot implementation of the Bukhali trial (n = 1655) were collected from (1) weekly team meetings, (2) two focus groups (one with the intervention team Health Helpers, n = 7; one with intervention participants, n = 8) and one paired interview with control call centre assistants (n = 2), (3) notes from eight debrief sessions with Health Helpers and (4) quantitative trial monitoring data. Qualitative data were thematically analysed. RESULTS: The findings clustered within three themes: (1) challenges for young women in Soweto, (2) priorities for young women in Soweto and (3) implementation challenges and perceptions of the intervention. Challenges were mostly related to tough socioeconomic circumstances and less prioritisation of living a healthier life. The priorities of employment and educational opportunities reflected the socioeconomic challenges, where health was not recognised as priority. The main challenge to participation and compliance with the trial was that young women in Soweto generally wanted a tangible and preferably financial and immediate benefit. Community peer sessions, despite being recommended by young women as part of the intervention development, were not successful. Many women also moved between multiple households within Soweto, which flagged concerns for a cluster trial and risk of contamination. CONCLUSION: Preconception health trials should consider socioeconomic challenges present in urban poor contexts. Learnings from the pilot phase significantly affected the design and implementation of the main Bukhali trial.

Urban young women's preferences for intervention strategies to promote physical and mental health preconception: A Healthy Life Trajectories Initiative (HeLTI).

This study aimed to qualitatively investigate young women's preferences for preconception intervention strategies to promote physical and mental health in a rapidly transitioning, urban setting. Four semi-structured focus group discussions were conducted with young women (n = 29, 18-24 years old) from Soweto, South Africa. Qualitative data were thematically analysed. Two main themes were identified: 1) challenges and needs of intervention beneficiaries; and 2) preferences for intervention strategies (content and delivery). The challenges participants mentioned could be classified as those relating to social pressure, identity, and socioeconomic circumstances. Mental health support appeared to be a greater need than physical health, and this featured in their preferences for intervention content, although a number of physical health topics were also mentioned (healthy eating and contraception). Participants had mixed preferences for intervention materials, ranging from printed to electronic and mobile resources. Their preferences for intervention activities ranged from educational sessions, to fun and interactive practical activities, and activities they could take home. Community health workers (CHWs) were the preferred agent of delivery for interventions, though participants emphasised the importance of CHWs having appropriate interpersonal skills and own life experience. Some women preferred one-on-one sessions with a CHW, while others preferred group sessions. While recognising the value of family sessions, young women were less enthusiastic about this approach. These findings provide valuable formative data for developing effective interventions to optimise young women's preconception health in urban Africa. These contextual realities should be acknowledged when addressing key physical and mental health issues facing young women.

Depressive symptoms, body composition and bone mass in young adults: a prospective cohort study.

BACKGROUND: An association between depression and obesity is well recognised, but longitudinal studies of depressive symptoms in adolescents as a predictor of body composition are lacking. OBJECTIVE: We examined depressive symptoms at age 14, 17 and 20 years as predictors of lean, fat and bone mass at age 20 years in a birth cohort. SUBJECTS/METHODS: In 1161 participants (569 females) in the Western Australia Pregnancy Cohort (Raine) Study, depressive symptoms were assessed using the Beck Depression Inventory for Youth at age 14 and 17 years, and the Depression, Anxiety and Stress Scale 21 at age 20 years. Participants were further classified into two trajectories using latent class analysis: no/transient and persistent/recurrent depression. At age 20 years, lean body mass (LBM), fat body mass (FBM) and total body bone mass were measured by dual-energy X-ray absorptiometry. RESULTS: In females, accounting for age and lifestyle factors, depression scores at age 14 and 20 years were positively associated with body weight, body mass index (BMI), FBM and % FBM (r=0.110-0.184, P<0.05) but negatively correlated with % LBM (r=-0.120, P<0.05) at age 20 years. Females in the persistent/recurrent depression trajectory (n=99) had significantly higher body weight (+5.1 kg), BMI (+1.8 kg m-2), FBM (+3.9 kg) and % FBM (+2.2%) and significantly lower % LBM (-2.2%) at age 20 years than those with no/transient depression (n=470; all P<0.05). In males, depression scores at age 17 and 20 years were negatively associated with LBM but not weight or BMI, and depression trajectory was not a predictor of body composition at age 20 years. Depression scores and trajectories did not predict bone mass in either males or females. CONCLUSIONS: Depressive symptoms and persistent/recurrent depression in adolescence are predictors of greater adiposity at age 20 years in females, but not males, but do not predict bone mass in either gender.

ATP-binding cassette transporters in reproduction: a new frontier.

BACKGROUND: The transmembrane ATP-binding cassette (ABC) transporters actively efflux an array of clinically relevant compounds across biological barriers, and modulate biodistribution of many physiological and pharmacological factors. To date, over 48 ABC transporters have been identified and shown to be directly and indirectly involved in peri-implantation events and fetal/placental development. They efflux cholesterol, steroid hormones, vitamins, cytokines, chemokines, prostaglandins, diverse xenobiotics and environmental toxins, playing a critical role in regulating drug disposition, immunological responses and lipid trafficking, as well as preventing fetal accumulation of drugs and environmental toxins. METHODS: This review examines ABC transporters as important mediators of placental barrier functions and key reproductive processes. Expression, localization and function of all identified ABC transporters were systematically reviewed using PubMed and Google Scholar websites to identify relevant studies examining ABC transporters in reproductive tissues in physiological and pathophysiological states. Only reports written in English were incorporated with no restriction on year of publication. While a major focus has been placed on the human, extensive evidence from animal studies is utilized to describe current understanding of the regulation and function of ABC transporters relevant to human reproduction. RESULTS: ABC transporters are modulators of steroidogenesis, fertilization, implantation, nutrient transport and immunological responses, and function as 'gatekeepers' at various barrier sites (i.e. blood-testes barrier and placenta) against potentially harmful xenobiotic factors, including drugs and environmental toxins. These roles appear to be species dependent and change as a function of gestation and development. The best-described ABC transporters in reproductive tissues (primarily in the placenta) are the multidrug transporters p-glycoprotein and breast cancer-related protein, the multidrug resistance proteins 1 through 5 and the cholesterol transporters ABCA1 and ABCG1. CONCLUSIONS: The ABC transporters have various roles across multiple reproductive tissues. Knowledge of efflux direction, tissue distribution, substrate specificity and regulation of the ABC transporters in the placenta and other reproductive tissues is rapidly expanding. This will allow better understanding of the disposition of specific substrates within reproductive tissues, and facilitate development of novel treatments for reproductive disorders as well as improved approaches to protecting the developing fetus.

Dynamic changes in maternal decidual leukocyte populations from first to second trimester gestation.

INTRODUCTION: Decidual leukocytes are critical to the development of the fetomaternal interface, regulating tolerance to the semi-allogeneic fetus and vascular transformation of the uterine spiral arteries. Despite the continuation of these processes beyond the first trimester of pregnancy, the second trimester has largely been unstudied, with investigation focusing on early gestation and term tissues. We sought to characterize changes in decidual leukocyte populations from first to second trimester. METHODS: Multicolor flow cytometry was performed on isolated decidual leukocytes from elective terminations of pregnancy between 6 and 20 weeks of gestation for study of first (6-12 weeks) and second trimesters (13-20 weeks). Specific subpopulations were identified by comparison to isotype and fluorescent-minus-one (FMO) controls. RESULTS: Decidual natural killer cells (CD56(+)CD16(-)CD3(-)) did not change in number, although a population of dNK with decreased CD56 brightness was observed in second trimester decidua. CD14(+)HLA-DR(+) macrophage numbers declined from first to second trimester (p = 0.031), yet a CD163(+)CD206(+) subset designating alternatively activated M2-like macrophages increased during the same period (p = 0.015). Intermediate CD205(+) dendritic cells demonstrated significant decline (p = 0.022), but immature CD209(+) and mature CD83(+) dendritic cells did not differ between trimesters. Total CD3(+) and CD3(+)CD4(+) T lymphocytes increased (p = 0.0079, p = 0.0028); CD3(+)CD8(+) T cells trended towards increase but did not differ significantly. CONCLUSION: Several changes in leukocyte subsets are observed in the second trimester that promote a tolerogenic and angiogenic decidual microenvironment through mid-gestation.

Endocrine gland-derived endothelial growth factor (EG-VEGF) is a potential novel regulator of human parturition.

EG-VEGF is an angiogenic factor that we identified as a new placental growth factor during human pregnancy. EG-VEGF is also expressed in the mouse fetal membrane (FM) by the end of gestation, suggesting a local role for this protein in the mechanism of parturition. However, injection of EG-VEGF to gravid mice did not induce labor, suggesting a different role for EG-VEGF in parturition. Here, we searched for its role in the FM in relation to human parturition. Human pregnant sera and total FM, chorion, and amnion were collected during the second and third trimesters from preterm no labor, term no labor, and term labor patients. Primary human chorion trophoblast and FM explants cultures were also used. We demonstrate that circulating EG-VEGF increased toward term and significantly decreased at the time of labor. EG-VEGF production was higher in the FM compared to placentas matched for gestational age. Within the FM, the chorion was the main source of EG-VEGF. EG-VEGF receptors, PROKR1 and PROKR2, were differentially expressed within the FM with increased expression toward term and an abrupt decrease with the onset of labor. In chorion trophoblast and FM explants collected from nonlaboring patients, EG-VEGF decreased metalloproteinase-2 and -9 activities and increased PGDH (prostaglandin-metabolizing enzyme) expression. Altogether these data demonstrate that EG-VEGF is a new cytokine that acts locally to ensure FM protection in late pregnancy. Its fine contribution to the initiation of human labor is exhibited by the abrupt decrease in its levels as well as a reduction in its receptors.

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.

Design, fabrication, and characterization of thermoplastic microlenses for fiber-optic probe imaging.

Microlens-ended fibers could find great usefulness in future biomedical applications, particularly in endoscopic imaging applications. In this context, this paper focuses on microlens-attached specialty optical fibers such as imaging fiber that can be used for probe imaging applications. Stand-alone self-aligned polymer microlenses have been fabricated by microcompression molding. The fabrication parameters have been optimized for different materials, such as poly(methyl methacrylate) (PMMA), polycarbonate (PC Lexan 123R), Zeonor 1060R (ZNR), and Topas COC. A comparison study of the focusing and spatial resolution of the fabricated lenses is performed prior to employing them for fiber-optic fluorescence imaging applications.

Gender differences in the relationships between lean body mass, fat mass and peak bone mass in young adults.

UNLABELLED: The relationships between fat mass and bone mass in young adults are unclear. In 1,183 young Australians, lean body mass had a strong positive relationship with total body bone mass in both genders. Fat mass was a positive predictor of total body bone mass in females, with weaker association in males. INTRODUCTION: Body weight and lean body mass are established as major determinants of bone mass, but the relationships between fat mass (including visceral fat) and peak bone mass in young adults are unclear. The aim of this study was to evaluate the associations between bone mass in young adults and three body composition measurements: lean body mass, fat mass and trunk-to-limb fat mass ratio (a surrogate measure of visceral fat). METHODS: Study participants were 574 women and 609 men aged 19-22 years from the Raine study. Body composition, total body bone mineral content (TBBMC), bone area and areal bone mineral density (TBBMD) were measured using DXA. RESULTS: In multivariate linear regression models with height, lean body mass, fat mass and trunk-to-limb fat mass ratio as predictor variables, lean mass was uniquely associated with the largest proportion of variance of TBBMC and TBBMD in males (semi-partial R(2) 0.275 and 0.345, respectively) and TBBMC in females (semi-partial R(2) 0.183). Fat mass was a more important predictor of TBBMC and TBBMD in females (semi-partial R(2) 0.126 and 0.039, respectively) than males (semi-partial R(2) 0.006 and 0.018, respectively). Trunk-to-limb fat mass ratio had a weak, negative association with TBBMC and bone area in both genders (semi-partial R(2) 0.004 to 0.034). CONCLUSIONS: Lean body mass has strong positive relationship with total body bone mass in both genders. Fat mass may play a positive role in peak bone mass attainment in women but the association was weaker in men; different fat compartments may have different effects.

Effect of oxygen on multidrug resistance in the first trimester human placenta.

INTRODUCTION: The multidrug resistance proteins, P-glycoprotein (P-gp, encoded by the ABCB1 gene) and breast cancer resistance protein (BCRP, encoded by ABCG2) are highly expressed in the first trimester placenta. These transporters protect the fetus from exposure to maternally derived toxins and xenobiotics. Since oxygen is a regulator of multidrug resistance in various tissues, we hypothesized that changes in oxygen tension alter placental ABCB1/P-gp and ABCG2/BCRP expression in the first trimester. METHODS: Placental specimens were collected from first (n = 7), second (n = 5) and term pregnancies (n = 5). First trimester placental villous explants were incubated (24 or 48 h) in different oxygen tension (3-20%). ABCB1, ABCG2 and VEGFA mRNA expression levels were assessed by RT-PCR and protein was localized by IHC. RESULTS: ABCB1 is expressed most highly in the first trimester placenta (p < 0.05), whereas ABCG2 expression does not change significantly over pregnancy. P-gp and BCRP staining is present in the syncytiotrophoblast and in cytotrophoblasts. ABCG2 mRNA is increased in hyperoxic (20%) conditions after 48 h (p < 0.05). In contrast, hypoxia (3%) did not change ABCB1 mRNA expression but significantly increased VEGFA mRNA (p < 0.05). Hypoxia resulted in increased BCRP staining in cytotrophoblasts and in the microvillous membrane of the syncytium. Whereas, hypoxia resulted in increased P-gp staining in proliferating cytotrophoblasts. CONCLUSION: We conclude that placental multidrug resistance expression, specifically ABCG2, is regulated by oxygen tension in the first trimester. It is possible that changes in placental oxygen supply are capable of altering fetal drug exposure especially during early pregnancy.

Polymorphisms in genes within the IGF-axis influence antenatal and postnatal growth.

Two pregnancy cohorts were used to investigate the association between single-nucleotide polymorphisms (SNPs) in genes within the insulin-like growth factor (IGF)-axis and antenatal and postnatal growth from birth to adolescence. Longitudinal analyses were conducted in the Raine pregnancy cohort (n = 1162) using repeated measures of fetal head circumference (HC), abdominal circumference (AC) and femur length (FL) from 18 to 38 weeks gestation and eight measures of postnatal height and weight (1-17 years). Replications of significant associations up to birth were undertaken in the Generation R Study (n = 2642). Of the SNPs within the IGF-axis genes, 40% (n = 58) were associated with measures of antenatal growth (P ⩽ 0.05). The majority of these SNPs were in receptors; IGF-1R (23%; n = 34) and IGF-2R (13%; n = 9). Fifteen SNPs were associated with antenatal growth (either AC or HC or FL) in Raine (P ⩽ 0.005): five of which remained significant after adjusting for multiple testing. Four of these replicated in Generation R. Associations were identified between 38% (n = 55) of the IGF-axis SNPs and postnatal height and weight; 21% in IGF-1R (n = 31) and 9% in IGF-2R (n = 13). Twenty-six SNPs were significantly associated with both antenatal and postnatal growth; 17 with discordant effects and nine with concordant effects. Genetic variants in the IGF-axis appear to play a significant role in antenatal and postnatal growth. Further replication and new analytic methods are required in order to better understand this key metabolic pathway integrating biologic knowledge about the interaction between IGF-axis components.

A novel pathogenic variant of the LDLR gene in the Asian population and its clinical correlation with familial hypercholesterolemia.

Familial hypercholesterolemia (FH) is a disease implicated with defects in either, Low density lipoprotein receptor gene (LDLR), Apolipoprotein B-100 gene (APOB), the Proprotein convertase subtilisin/kexin type 9 gene (PCSK9) or other related genes of the lipid metabolism pathway. The general characterization of heterozygous FH is by elevated low-density lipoprotein (LDL) cholesterol and early-onset cardiovascular diseases, while the more severe type, the homozygous FH results in extreme elevated levels of LDL cholesterol and usually death of an affected individual by early twenties. We present here a novel non-synonymous, missense mutation in exon 14 of the LDLR gene in two siblings of the Malay ethnicity discovered during an in-house genetic test. We postulate that their elevated cholesterol is due to this novel mutation and they are positive for homozygous FH. This is the first report of a C711Y mutation in patients with elevated cholesterol in Asia.

Fat mass and obesity-associated obesity-risk genotype is associated with lower foetal growth: an effect that is reversed in the offspring of smoking mothers.

Fat mass and obesity-associated (FTO) gene variants are associated with childhood and adult obesity; however, the influence of FTO polymorphisms on foetal growth is unknown. Associations between the FTO variant rs9939609 and the foetal growth trajectories, maternal pregnancy weight gain, anthropometric measures at birth and body mass index (BMI) at age 14 years were assessed in 1079 singleton-birth Australian Caucasians. Analyses were repeated in 3512 singleton-birth Dutch Caucasians. The rs9939609 obesity-risk AA genotype was associated with symmetrical intrauterine growth restriction; an effect reversed in mothers who smoked during pregnancy. The effect increased over time and was modified by maternal smoking for head circumference (P = 0.007), abdominal circumference (P = 0.007), femur length (P = 0.02) and estimated foetal weight (P = 0.001). The modification of the association between the AA genotype and birth anthropometrics by maternal smoking was consistent across birth weight (P = 0.01) and birth length (P = 0.04) and neonatal day 2 anthropometry. Consistent associations were replicated in the Generation R cohort. Maternal pregnancy weight gain matched the pattern of birth weight and was independent of placental weight. In adolescents, the AA genotype was associated with increased BMI-adjusted-for-age in males (P = 0.00009), but no effect was detected in females. A variant in the FTO gene influences foetal growth trajectories in the third trimester, early postnatal growth and adiposity in adolescence. Maternal smoking during pregnancy reversed the direction of association of rs9939609 on foetal growth, which was probably mediated by maternal energy intake. The detection of genetic variants associated with foetal growth has the potential to identify novel molecular mechanisms underlying growth and targeted early life intervention.

Heparin promotes soluble VEGF receptor expression in human placental villi to impair endothelial VEGF signaling.

BACKGROUND: Severe preeclampsia is characterized by hypertension, renal injury and placental dysfunction. Prothrombotic disorders are discovered in 10-20% of women with preeclampsia, providing the rationale for prescribing low-molecular-weight heparin (LMWH) in future pregnancies. Heparin has diverse molecular actions and appears to reduce the recurrence risk of preeclampsia in women without prothrombotic disorders. The placenta-derived anti-angiogenic splice-variant protein soluble vascular endothelial growth factor (VEGF) receptor-1 (sFLT1) is strongly implicated in the pathogenesis of the underlying endothelial dysfunction. As the placental syncytiotrophoblast is the principal source of sFLT1, we tested the hypothesis that heparin suppresses placental sFLT1 secretion. METHODS AND RESULTS: First trimester placental villi exposed to LMWH (0.25-25 IU mL(-1)) in an in vitro explant model significantly increased the expression and release of sFLT1 by the syncytiotrophoblast into culture media, reducing phosphorylation of FLT1 and KDR receptors in cultured human umbilical vein endothelial cells. This response was significantly diminished in placental villi from healthy term pregnancies. Placental villi from severely preeclamptic pregnancies had a higher baseline sFLT1 release, compared with first trimester placental villi and did not respond to LMWH treatment. LMWH promoted villous cytotrophoblast proliferation (BrdU incorporation) and impaired syncytial fusion-differentiation, causing syncytiotrophoblast apoptosis (by caspase 3&7 activity and TUNEL staining) and necrosis (ADP/ATP ratio). CONCLUSION: LMWH promotes sFLT1 synthesis and release from first trimester placental villi in a manner similar to that of severely preeclamptic placental villi, which antagonizes VEGF signaling in endothelial cells. These effects in part are mediated by an interaction between heparin and the cytotrophoblasts that regenerates the overlying syncytiotrophoblast responsible for sFLT1 secretion into the maternal blood.

HER1 signaling mediates extravillous trophoblast differentiation in humans.

This study examines the role of HER1 signaling in the differentiation of proliferative extravillous trophoblast (EVT) into invasive EVT. Using the JAR choriocarcinoma cell line and placental villous explants as experimental models and immunohistochemical assessment of protein markers of EVT differentiation (downregulation of HER1 and Cx40 and upregulation of HER2 and alpha1 integrin), we show that the ability of decidual conditioned medium (DCM) to induce HER1/2 switching was abrogated in the presence of the HER1 antagonist, AG1478. Similarly, epidermal growth factor (EGF) treatment resulted in the downregulation of HER1 and an upregulation of HER2 expression, whereas co-incubation of EGF with AG1478 inhibited this response. However, EGF did not downregulate Cx40 or induce migration of EVT. In contrast, heparin-binding epidermal-like growth factor (HBEGF) stimulated dose-dependent JAR cell migration, which was inhibited by both AG1478 and AG825 (HER2 antagonist). Western blot analysis of HER1 activation demonstrated that HBEGF-mediated phosphorylation of the HER1 Tyr992 and Tyr1068 sites, while EGF activated the Tyr1045 site. Moreover, HBEGF induced a stronger and more sustained activation of both the mitogen-activated protein kinase and phosphoinositol 3 kinase (PIK3) signaling pathways. Migration assays using a panel of signaling pathway inhibitors demonstrated that the HBEGF-mediated migration was dependent on the PIK3 pathway. These results demonstrate that HBEGF-mediated HER1 signaling through PIK3 is an important component of EVT invasion.

Spatiotemporal expression of alpha(1), alpha(3) and beta(1) integrin subunits is altered in rat myometrium during pregnancy and labour.

Integrins are transmembrane extracellular matrix (ECM) receptors composed of alpha- and beta-subunits. Integrins can cluster to form focal adhesions and, because there is significant ECM remodelling and focal adhesion turnover in the rat myometrium during late pregnancy, we hypothesised that the expression of alpha(1), alpha(3) and beta(1) integrin subunits in the rat myometrium would be altered at this time to accommodate these processes. Expression of alpha(1) and beta(1) integrin subunit mRNA was significantly increased on Days 6-23 of pregnancy compared with non-pregnant (NP) and postpartum (PP) time points (P < 0.05). In contrast, alpha(3) integrin subunit mRNA expression was significantly increased on Days 14, 21 and 22 compared with NP, Day 10, 1 day PP and 4 days PP (P < 0.05). A relative gene expression study revealed that, of the integrins studied, the expression of beta(1) integrin mRNA was highest in pregnant rat myometrium. The alpha(1), alpha(3) and beta(1) integrin subunit proteins became immunolocalised to myocyte membranes in situ by late pregnancy and labour in both myometrial muscle layers. Increased alpha(1), alpha(3) and beta(1) integrin gene expression during gestation and the specific detection of these subunits in myocyte membranes during late pregnancy and labour may contribute to the cell-ECM interactions required for the development of a mechanical syncytium.

Glial cell missing-1 transcription factor is required for the differentiation of the human trophoblast.

Mammalian placentation is a highly regulated process and is dependent on the proper development of specific trophoblast cell lineages. The two major types of trophoblast, villous and extravillous, show mitotic arrest during differentiation. In mice, the transcription factor, glial cell missing-1 (Gcm1), blocks mitosis and is required for syncytiotrophoblast formation and morphogenesis of the labyrinth, the murine equivalent of the villous placenta. The human homolog GCM1 has an analogous expression pattern, but its function is presently unknown. We studied GCM1 function in the human-derived BeWo choriocarcinoma cell line and in first trimester human placental villous and extravillous explants. The GCM1 expression was either inhibited by siRNA and antisense oligonucleotides methods or upregulated by forskolin treatment. Inhibition of GCM1 resulted in an increased rate of proliferation, but prevented de novo syncytiotrophoblast formation in syncytially denuded floating villous explants. GCM1 inhibition prevented extravillous differentiation along the invasive pathway in extravillous explants on matrigel. By contrast, forskolin-induced expression of GCM1 reduced the rate of proliferation and increased the rate of syncytialization in the floating villous explant model. Our studies show that GCM1 has a distinct role in the maintenance, development and turnover of the human trophoblast. Alterations in GCM1 expression or regulation may explain several aspects of two divergent severe placental insufficiency syndromes, namely preeclampsia and intrauterine growth restriction, which cause extreme preterm birth.