RESUMO
Since the early 1960s, researchers began culturing placental cells to establish an in vitro model to study the biology of human trophoblasts, including their ability to differentiate into syncytiotrophoblasts and secrete steroid and peptide hormones that help sustain a viable pregnancy. This task was addressed by testing different serum concentrations, cell culture media, digestive enzymes, growth factors, substrate coating with diverse proteins from the extracellular matrix, and so on. Among the many methodological challenges, the contamination of trophoblasts with other cell types, such as immune and stromal cells, was a matter of concern. However, introducing the Percoll gradient to isolate cytotrophoblasts was an excellent contribution, and later, the depletion of contaminating cells by using magnetic bead-conjugated antibodies also helped increase the purity of cytotrophoblasts. Herein, with some modifications, we describe a rapid and easy method for cytotrophoblast isolation from the term human placenta based on the previously reported method by Harvey Kliman et al. (Endocrinology 118:1567-1582, 1986). This method yields about 40-90 million cells from a single placenta, with a purity of around 85-90%.
Assuntos
Gonadotropina Coriônica , Placenta , Humanos , Gravidez , Feminino , Gonadotropina Coriônica/metabolismo , Células Cultivadas , TrofoblastosRESUMO
AIM: miR-141-5p expression in patients with Early Spontaneous Abortion (ESA) and its correlation with hormone levels during pregnancy were investigated. METHODS: A total of 70 pregnant women with ESA were selected as the research group, and 70 normal pregnant women who chose abortion for non-medical reasons were selected as the Con group. Serum ß-HCG, Progesterone (P), and Estrogen (E2) were detected by enzyme-linked immunosorbent assay. Differentially expressed miRNAs were screened by miRNA microarray analysis. miR-141-5p expression was detected by RT-qPCR, and its correlation with serum ß-HCG, P, and E2 levels was analyzed. The diagnostic value of miR-141-5p for ESA was evaluated by the ROC curve. RESULTS: Serum ß-HCG, P, and E2 were decreased and serum miR-141-5p was increased in patients with ESA. Pearson correlation analysis showed that serum ß-HCG, P, and E2 levels were negatively correlated with miR-141-5p expression levels. ROC curve showed that miR-141-5p had a diagnostic value for ESA. CONCLUSIONS: miR-141-5p is related to hormone levels during pregnancy and is expected to become a new candidate diagnostic marker for ESA.
Assuntos
Aborto Espontâneo , MicroRNAs , Humanos , Feminino , Gravidez , Aborto Espontâneo/diagnóstico , Relevância Clínica , MicroRNAs/genética , Biomarcadores , ProgesteronaRESUMO
OBJECTIVE: To study the predictive value of serum ß-hCG on day 5 after frozen-thawed embryo transfer (FET) to predict pregnancy outcomes and to establish cut-off values for very early biochemical pregnancy diagnosis. METHODS: This retrospective cohort study was performed at a private reproductive medicine centre and we reviewed the medical records of women who underwent FET cycles from January 2018 to June 2019. A total of 116 evaluated cycles had serum ß-hCG levels measured on days 5 and 10 after FET. The predictive value of serum ß-hCG levels measured on day 5 after FET was investigated for very early biochemical pregnancy diagnosis. RESULTS: The standard biochemical pregnancy diagnosis was defined as a ß-hCG ≥25 IU/L on day 10 after FET. We then generated a receiver operating characteristic curve, and the cut-off value of ß-hCG on day 5 for predicting biochemical pregnancies was 4.0 IU/L, with 93.4% sensitivity and 92.7% specificity (AUC, 0.960; 95% confidence interval, 0.923-0.997). CONCLUSIONS: Values for ß-hCG at day 5 after FET ≥4.0IU/L are accurate for the diagnosis of biochemical pregnancy. The use of very early biochemical pregnancy diagnosis in clinical practice enables earlier management, patient counselling, and appropriate follow-up.
Assuntos
Gonadotropina Coriônica Humana Subunidade beta , Complicações na Gravidez , Transferência Embrionária , Feminino , Humanos , Gravidez , Resultado da Gravidez , Estudos RetrospectivosRESUMO
BACKGROUND: An imbalance between anti- and angiogenic factors during early placentation is key for the development of preeclampsia. Nevertheless, the majority of studies addressing this issue relate to maternal blood and not the fetal circulation. OBJECTIVE: To measure placental growth factor (PlGF), free beta human chorionic gonadotropin (ß-hCG), and pregnancy-associated plasma protein-A (PAPP-A) levels in the fetal circulation of near-term pregnancies complicated with severe preeclampsia (n = 20), and their controls matched for parity, and maternal and gestational age. METHOD: Upon delivery, a blood sample was withdrawn from the umbilical artery and vein of each case and its control in order to measure the proposed analytes using direct fluoroimmunoassay. RESULTS: Preeclampsia cases showed significantly lower median PlGF levels in fetal circulation as compared to controls (25.2 versus 36.9 and 23.6 versus 33.9 pg/mL, artery and vein, respectively, p < 0.05). Contrarily, cases displayed higher concentrations of PAPP-A (1024.0 versus 720.9 [median] and 1027.0 ± 298.4 versus 690.3 ± 401.9 mIU/L, artery and vein, respectively, p < 0.05), and free ß-hCG (mean: 33.9 ± 4.3 versus 17.2 ± 4.0 and 30.1 ± 5.2 versus 13.7 ± 3.3 ng/mL, artery, and vein respectively, p < 0.05). CONCLUSION: Lower PlGF and higher PAPP-A and free ß-hCG levels were found in the fetal circulation of near-term severe preeclamptic pregnancies. There is a need for more research in this regard.