Patrón de decidualización en mujeres con preeclampsia mediante la aplicación de métodos de transcriptómica y proteómica

Los defectos en la decidualización de las células del estroma endometrial humanas (CEE) están relacionados con la alteración en la invasión trofoblástica y en las alteraciones fibrinolíticas, siendo una posible causa de la preeclampsia. El objetivo de esta tesis doctoral es investigar el proceso de decidualización en pacientes que han sufrido preeclampsia severa y la implicación de la anexina A2 (ANXA2) como regulador clave de la invasión defectuosa del trofoblasto y las alteraciones fibrinolíticas de esta enfermedad. Material y Métodos: CEE procedentes de mujeres que han sufrido preeclampsia grave (PE) (N = 13) y pacientes con antecedentes de embarazos sanos (Non-PE) (N = 13) fueron decidualizadas in vitro. La expresión de PRL, IGFBP-1 y ANXA2 fueron evaluadas. Se investigó el fenómeno de expansión trofoblástica en un modelo in vitro utilizando un co-cultivo heterólogo con blastocitos de ratón en CEE decidualizadas, silenciándose una de las líneas de cultivo para la expresión de ANXA2 con siRNA. La invasión del trofoblasto humando se analizó utilizando JEG-3 línea celular en cámaras de invasión de colágeno. Resultados: La dedidualización in vitro fue defectuosa en CEE procedentes de mujeres con antecedente de PE severa vs mujeres con antecedente de gestaciones sanas. Durante la decidualización, la PRL secretada y IGFBP-1 se redujeron significativamente en PE vs no-PE (2,98 ± 0,6 vs 15,3 ± 4,6 ng / ml para la PRL, y 5,5 ± 1,7 frente a 14 , 59 ± 7,2 ng / ml para IGFBP1). La proteína ANXA2 aumentó significativamente en cultivo decidual en mujeres sanas vs mujeres con PE (65,76 ± 21,2 mg / mL vs 13,53 ± 16,9; 7,8 ± 4,1 vs 2,49 ± 2 g / mL niveles intra y extracelulares, respectivamente). Tratando de imitar la dinámica de ANXA2 en CEE procedentes de pacientes con PE, ANXA2 siRNA se usó como silenciado, mostrando una marcada disminución en el área de difusión de trofoblasto en comparación con las células control (33,6% ± 15,3 vs 81,3 ± 13,5%), como así como la invasión del trofoblasto (49,2% ± 9,1 vs 86,5 ± 11,6%). Conclusiones: las CEE procedentes de pacientes que han sufrido PE severa en algunas de sus gestaciones mostraron una capacidad limitada para decidualizarse y una regulación a la baja en la producción de ANXA2 tanto intra como extracelular. Además, se valora que la inhibición in vitro de CEE silenciadas para la expresión de ANXA2, afecta a los fenómenos de difusión e invasión trofoblástica, como un mecanismo implicado en la participación endometrial en el origen de la preeclampsia.Defects in human endometrial stromal cell (hESC) decidualization are related with trophoblast invasion alteration as a possible cause of preeclampsia. The aim of this work was to investigate the decidualization process in patients that have suffered severe preeclampsia and the implication of annexin A2 (ANXA2) as key regulator of defective trophoblast invasion. Methods: hESC of women that have suffered severe preeclampsia (PE) (N=13) and healthy patients (Non-PE) (N=13) were in vitro decidualized and PRL, IGFBP-1 and ANXA2 assessed. Trophoblast spreading was investigated in an in vitro model using hatched mouse embryos cocultured on decidual ANXA2 siRNA inhibited vs control hESCs. Trophoblast invasion was analyzed using JEG-3 cell line on collagen transwell invasion chambers. Results: In vitro decidualization was successful at a reduced extent in hESC obtained from PE compared to Non-PE patients. During decidualization, secreted PRL and IGFBP-1 were significantly reduced in PE vs non-PE (2,98±0,6 vs 15,3±4,6 ng/mL for PRL; and 5,5±1,7 vs 14,59±7,2 ng/mL for IGFBP1). ANXA2 protein significantly increased in decidual vs non-decidual hESCs obtained from non-PE patients (65,76±21,2 µg/mL vs 13,53±16,9; 7,8±4,1 vs 2,49±2 µg/mL intra- and extracellular levels, respectively). Interestingly, this regulation was abolished during decidualization compared with non-decidualizated hESC in PE patients. Trying to mimic the dynamics of ANXA2 in hESC from PE patients, ANXA2 siRNA transfected hESC showed a marked decrease in trophoblast spreading area compared to control cells (33,6%±15,3 vs 81,3%±13,5), as well as trophoblast invasion (49,2%±9,1 vs 86,5%±11,6). Conclusions: hESCs from patients that have suffered early severe PE showed a limited ability to decidualize and intra/extracellular ANXA2 regulation was abolished during decidualization. We further demonstrate in vitro that ANXA2 inhibition in hESC affects trophoblast spreading and invasion as a mechanism involved in the onset of the endometrial origin of PE

Accuracy and Survival Outcomes after National Implementation of Sentinel Lymph Node Biopsy in Early Stage Endometrial Cancer

Background. Sentinel lymph node (SLN) biopsy has recently been accepted to evaluate nodal status in endometrial cancer at early stage, which is key to tailoring adjuvant treatments. Our aim was to evaluate the national implementation of SLN biopsy in terms of accuracy to detect nodal disease in a clinical setting and oncologic outcomes according to the volume of nodal disease. Patients and Methods. A total of 29 Spanish centers participated in this retrospective, multicenter registry including patients with endometrial adenocarcinoma at preoperative early stage who had undergone SLN biopsy between 2015 and 2021. Each center collected data regarding demographic, clinical, histologic, therapeutic, and survival characteristics. Results. A total of 892 patients were enrolled. After the surgery, 12.9% were suprastaged to FIGO 2009 stages III-IV and 108 patients (12.1%) had nodal involvement: 54.6% macrometastasis, 22.2% micrometastases, and 23.1% isolated tumor cells (ITC). Sensitivity of SLN biopsy was 93.7% and false negative rate was 6.2%. After a median follow up of 1.81 years, overall surivial and disease-free survival were significantly lower in patients who had macrometastases when compared with patients with negative nodes, micrometastases or ITC. Conclusions. In our nationwide cohort we obtained high sensitivity of SLN biopsy to detect nodal disease. The oncologic outcomes of patients with negative nodes and low-volume disease were similar after tailoring adjuvant treatments. In total, 22% of patients with macrometastasis and 50% of patients with micrometastasis were at low risk of nodal metastasis according to their preoperative risk factors, revealing the importance of SLN biopsy in the surgical management of patients with early stage EC

Defective decidualization during and after severe preeclampsia reveals a possible maternal contribution to the etiology

In preeclampsia (PE), cytotrophoblast (CTB) invasion of the uterus and spiral arteries is often shallow. Thus, the placenta's role has been a focus. In this study, we tested the hypothesis that decidual defects are an important determinant of the placental phenotype. We isolated human endometrial stromal cells from nonpregnant donors with a previous pregnancy that was complicated by severe PE (sPE). Compared with control cells, they failed to decidualize in vitro as demonstrated by morphological criteria and the analysis of stage-specific antigens (i.e., IGFBP1, PRL). These results were bolstered by global transcriptional profiling data that showed they were transcriptionally inert. Additionally, we used laser microdissection to isolate the decidua from tissue sections of the maternal-fetal interface in sPE. Global transcriptional profiling revealed defects in gene expression. Also, decidual cells from patients with sPE, which dedifferentiated in vitro, failed to redecidualize in culture. Conditioned medium from these cells failed to support CTB invasion. To mimic aspects of the uterine environment in normal pregnancy, we added PRL and IGFBP1, which enhanced invasion. These data suggested that failed decidualization is an important contributor to down-regulated CTB invasion in sPE. Future studies will be aimed at determining whether this discovery has translational potential with regard to assessing a woman's risk of developing this pregnancy complication

Accuracy and survival outcomes after national implementation of sentinel lymph node biopsy in early stage endometrial cancer

Background Sentinel lymph node (SLN) biopsy has recently been accepted to evaluate nodal status in endometrial cancer at early stage, which is key to tailoring adjuvant treatments. Our aim was to evaluate the national implementation of SLN biopsy in terms of accuracy to detect nodal disease in a clinical setting and oncologic outcomes according to the volume of nodal disease. Patients and Methods A total of 29 Spanish centers participated in this retrospective, multicenter registry including patients with endometrial adenocarcinoma at preoperative early stage who had undergone SLN biopsy between 2015 and 2021. Each center collected data regarding demographic, clinical, histologic, therapeutic, and survival characteristics. Results A total of 892 patients were enrolled. After the surgery, 12.9% were suprastaged to FIGO 2009 stages III-IV and 108 patients (12.1%) had nodal involvement: 54.6% macrometastasis, 22.2% micrometastases, and 23.1% isolated tumor cells (ITC). Sensitivity of SLN biopsy was 93.7% and false negative rate was 6.2%. After a median follow up of 1.81 years, overall surivial and disease-free survival were significantly lower in patients who had macrometastases when compared with patients with negative nodes, micrometastases or ITC. Conclusions In our nationwide cohort we obtained high sensitivity of SLN biopsy to detect nodal disease. The oncologic outcomes of patients with negative nodes and low-volume disease were similar after tailoring adjuvant treatments. In total, 22% of patients with macrometastasis and 50% of patients with micrometastasis were at low risk of nodal metastasis according to their preoperative risk factors, revealing the importance of SLN biopsy in the surgical management of patients with early stage EC.FUNDING. Open Access Funding provided by Universitat Autonoma de Barcelon

Impact of uterine manipulator on oncological outcome in endometrial cancer surgery.

There are limited data available to indicate whether oncological outcomes might be influenced by the uterine manipulator, which is used at the time of hysterectomy for minimally invasive surgery in patients with endometrial cancer. The current evidence derives from retrospective studies with limited sample sizes. Without substantial evidence to support its use, surgeons are required to make decisions about its use based only on their personal choice and surgical experience. To evaluate the use of the uterine manipulator on oncological outcomes after minimally invasive surgery, for apparent early-stage endometrial cancer. We performed a retrospective multicentric study to assess the oncological safety of uterine manipulator use in patients with apparent early-stage endometrial cancer, treated with minimally invasive surgery. The type of manipulator, surgical staging, histology, lymphovascular space invasion, International Federation of Gynecology and Obstetrics stage, adjuvant treatment, recurrence, and pattern of recurrence were evaluated. The primary objective was to determine the relapse rate. The secondary objective was to determine recurrence-free survival, overall survival, and the pattern of recurrence. A total of 2661 women from 15 centers were included; 1756 patients underwent hysterectomy with a uterine manipulator and 905 without it. Both groups were balanced with respect to histology, tumor grade, myometrial invasion, International Federation of Gynecology and Obstetrics stage, and adjuvant therapy. The rate of recurrence was 11.69% in the uterine manipulator group and 7.4% in the no-manipulator group (P In this study, the use of a uterine manipulator was associated with a worse oncological outcome in patients with uterus-confined endometrial cancer (International Federation of Gynecology and Obstetrics I-II) who underwent minimally invasive surgery. Prospective trials are essential to confirm these results

Lymphovascular Space Invasion in Early-Stage Endometrial Cancer (LySEC): Patterns of Recurrence and Predictors. A Multicentre Retrospective Cohort Study of the Spain Gynecologic Oncology Group

The main aim is to compare oncological outcomes and patterns of recurrence of patients with early-stage endometrioid endometrial cancer according to lymphovascular space invasion (LVSI) status. The secondary objective is to determine preoperative predictors of LVSI. We performed a multicenter retrospective cohort study. A total of 3546 women diagnosed with postoperative early-stage (FIGO I-II, 2009) endometrioid endometrial cancer were included. Co-primary endpoints were disease-free survival (DFS), overall survival (OS), and pattern of recurrence. Cox proportional hazard models were used for time-to-event analysis. Univariate and multivariate logistical regression models were employed. Positive LVSI was identified in 528 patients (14.6%) and was an independent prognostic factor for DFS (HR 1.8), OS (HR 2.1) and distant recurrences (HR 2.37). Distant recurrences were more frequent in patients with positive LVSI (78.2% vs. 61.3%, p < 0.01). Deep myometrial invasion (OR 3.04), high-grade tumors (OR 2.54), cervical stroma invasion (OR 2.01), and tumor diameter ≥ 2 cm (OR 2.03) were independent predictors of LVSI. In conclusion, in these patients, LVSI is an independent risk factor for shorter DFS and OS, and distant recurrence, but not for local recurrence. Deep myometrial invasion, cervical stroma invasion, high-grade tumors, and a tumor diameter ≥ 2 cm are independent predictors of LVSI