Charlotte Ngô

Reactive Oxygen Species Controls Endometriosis Progression

Endometriosis is associated with chronic inflammation, and reactive oxygen species (ROS) are proinflammatory mediators that modulate cell proliferation. We have investigated whether the dysregulation of ROS production in endometriotic cells correlates with a pro-proliferative phenotype and can explain the spreading of this disease. Stromal and epithelial cells were purified from ovarian endometrioma and eutopic endometrium from 14 patients with endometriosis to produce four primary cell lines from each patient. ROS production, detoxification pathways, cell proliferation, and mitogen-activated protein kinase pathway activation were studied and compared with epithelial and stromal cell lines from 14 patients without endometriosis. Modulation of the proliferation of endometriosis by N-acetyl-cysteine, danazol, and mifepristone was tested in vitro and in 28 nude mice implanted with endometriotic tissue of human origin. Endometriotic cells displayed higher endogenous oxidative stress with an increase in ROS production, alterations in ROS detoxification pathways, and a drop in catalase levels, as observed for tumor cells. This increase in endogenous ROS correlated with increased cellular proliferation and activation of ERK1/2. These phenomena were abrogated by the antioxidant molecule N-acetyl-cysteine both in vitro and in a mouse model of endometriosis. Human endometriotic cells display activated pERK, enhanced ROS production, and proliferative capability. Our murine model shows that antioxidant molecules could be used as safe and efficient treatments for endometriosis.

Questioning patients about their adolescent history can identify markers associated with deep infiltrating endometriosis

OBJECTIVE To investigate whether the clinical history, particularly of the adolescence period, contains markers of deeply infiltrating endometriosis (DIE). DESIGN Cross-sectional study. SETTING Universitary tertiary referral center. PATIENT(S) Two hundred twenty-nine patients operated on for endometriosis. Endometriotic lesions were histologically confirmed as non-DIE (superficial peritoneal endometriosis and/or ovarian endometriomas) (n = 131) or DIE (n = 98). INTERVENTION(S) Surgical excision of endometriotic lesions with pathological analysis of each specimens. MAIN OUTCOME MEASURE(S) Epidemiological data, pelvic pain scores, family history of endometriosis, absenteeism from school during menstruation, oral contraceptive (OC) pill use. RESULT(S) Patients with DIE had significantly more positive family history of endometriosis (odds ratio [OR] = 3.2; 95% confidence interval [CI]: 1.2-8.8) and more absenteeism from school during menstruation (OR = 1.7; 95% CI: 1-3). The OC pill use for treating severe primary dysmenorrhea was more frequent in patients with DIE (OR = 4.5; 95% CI: 1.9-10.4). Duration of OC pill use for severe primary dysmenorrhea was longer in patients with DIE (8.4 ± 4.7 years vs. 5.1 ± 3.8 years). There was a higher incidence of OC pill use for severe primary dysmenorrhea before 18 years of age in patients with DIE (OR = 4.2; 95% CI: 1.8-10.0). CONCLUSION(S) The knowledge of adolescent period history can identify markers that are associated with DIE in patients undergoing surgery for endometriosis.

The mTOR/AKT Inhibitor Temsirolimus Prevents Deep Infiltrating Endometriosis in Mice

Deep infiltrating endometriosis (DIE) is a particular clinical and histological entity of endometriosis responsible for chronic pelvic pain and infertility. Here we characterize the proliferative phenotype of DIE cells, to explore the cellular and molecular mechanisms that could explain their aggressive potential. In addition, the inhibition of mTOR/AKT pathway was tested, as a potential treatment of DIE. Included were 22 patients with DIE and 12 control patients without endometriosis. Epithelial and stromal cells were extracted from biopsies of eutopic endometrium and deep infiltrating endometriotic nodules from patients with DIE. Cell proliferation was determined by thymidine incorporation. Oxidative stress was assayed by spectrofluorometry. The ERK and mTOR/AKT pathways were analyzed in vitro by Western blot and for AKT in vivo in a mouse model of DIE. The proliferation rate of eutopic endometrial cells and of deep infiltrating endometriotic cells from DIE patients was higher than that of endometrial cells from controls. The hyperproliferative phenotype of endometriotic cells was associated with an increase in endogenous oxidative stress, and with activation of the ERK and mTOR/AKT pathways. mTOR/AKT inhibition by temsirolimus decreased endometriotic cell proliferation both in vitro and in vivo in a mouse model of DIE. Blocking the mTOR/AKT pathway offers new prospects for the treatment of DIE.

Protein kinase inhibitors can control the progression of endometriosis in vitro and in vivo.

Endometriosis affects 6–10% of women in their reproductive years, causing chronic pelvic pain and infertility. Its pathogenesis remains poorly understood and current treatments, based on hormonal therapy or surgery, are often insufficient. The purpose of our study was to investigate the role of the ERK pathway in the development of endometriosis and to test the effects of protein kinase inhibitors on the proliferation of endometriotic cells in vitro and in vivo. We studied ex vivo human endometrial and endometriotic cells in culture. Stromal and epithelial cells were extracted from endometrial and endometriotic biopsies from patients with endometriosis and from patients without endometriosis. The ERK pathway was explored by western blot on cell lysates and by ELISA on total crushed specimens of endometrium. Cells in culture were treated with A771726, PD98059, and U0126. Human endometriotic lesions were implanted in nude mice. Mice were treated with A771726, leflunomide, PD98059, U0126 or PBS during 2 weeks before sacrifice and extraction of the endometriotic implants for histological examination. We found that the ERK pathway was significantly activated in endometriotic cells and in endometrial cells from patients with endometriosis compared to endometrial cells of control patients, both by ELISA and by western blot. This phenomenon was associated with an increased proliferation of endometriotic cells compared to endometrial cells. Treating endometriotic cells with A771726, PD98059 or U0126 abrogated the phosphorylation of ERK and significantly decreased the cellular proliferation in vitro. In vivo, A771726, leflunomide, PD98059, and U0126 controlled the growth of endometriotic implants in the mouse model of endometriosis. Our study shows that protein kinase inhibitors could be new candidates to treat endometriosis. However, further studies are needed to evaluate their effects and tolerability in humans. Copyright

Smoking habits of 411 women with histologically proven endometriosis and 567 unaffected women

Smoking habits did not influence either the risk of any form of endometriosis (superficial peritoneal endometriosis, ovarian endometriomas, and deep infiltrating endometriosis) and did not correlate with the revised American Fertility Society stages or scores.

Antiproliferative Effects of Cannabinoid Agonists on Deep Infiltrating Endometriosis

Deep infiltrating endometriosis (DIE) is characterized by chronic pain, hyperproliferation of endometriotic cells and fibrosis. Since cannabinoids are endowed with antiproliferative and antifibrotic properties, in addition to their psychogenic and analgesic effects, cannabinoid agonists have been evaluated in DIE both in vitro and in vivo. The in vitro effects of the cannabinoid agonist WIN 55212-2 were evaluated on primary endometriotic and endometrial stromal and epithelial cell lines extracted from patients with or without DIE. Cell proliferation was determined by thymidine incorporation and production of reactive oxygen species by spectrofluorometry. ERK and Akt pathways were studied by immunoblotting. Immunoblotting of α-smooth muscle actin was studied as evidence of myofibroblastic transformation. The in vivo effects of WIN 55212-2 were evaluated on Nude mice implanted with human deep infiltrating endometriotic nodules. The in vitro treatment of stromal endometriotic cells by WIN 55212-2 decreased cell proliferation, reactive oxygen species production, and α-smooth muscle actin expression. The decrease in cell proliferation induced by WIN 55212-2 was not associated with a decrease in ERK activation, but was associated with the inhibition of Akt activation. WIN 55212-2 abrogated the growth of endometriotic tissue implanted in Nude mice. Cannabinoid agonists exert anti-proliferative effects on stromal endometriotic cells linked to the inhibition of the Akt pathway. These beneficial effects of cannabinoid agonists on DIE have been confirmed in vivo.

Antiproliferative effects of anastrozole, methotrexate, and 5-fluorouracil on endometriosis in vitro and in vivo

OBJECTIVE To investigate the effects of antiproliferative drugs (anastrozole, methotrexate, and 5-fluorouracil [5-FU]) on the proliferation of endometriotic cells in vitro and in vivo. DESIGN Ex vivo study on human endometrial and endometriotic cells in culture; establishment of a murine model using mice implanted with human endometriosis. SETTING University research center. PATIENT(S) Ten patients with ovarian endometrioma, 10 patients with deep infiltrating endometriosis, and 10 patients without endometriosis. INTERVENTION(S) Stromal and epithelial cells were extracted from endometrial and endometriotic biopsies from patients with endometriosis and from patients without endometriosis. Cells were treated in vitro with anastrozole, methotrexate, progesterone, or 5-FU. Human endometriotic lesions were implanted in nude mice. Mice were treated with 5-FU or phosphate-buffered saline during 4 weeks before sacrifice and extraction of the endometriotic implants. MAIN OUTCOME MEASURE(S) Stromal and epithelial cell proliferation and pathology score of endometriotic implants. RESULT(S) Although anastrozole, methotrexate, and progesterone were ineffective, 5-FU significantly decreased the proliferation of endometriotic cells in vitro and controlled the growth of both cells from ovarian endometrioma and deep infiltrating endometriosis. CONCLUSION(S) Considering common features between endometriotic cells and tumor cells, the use of 5-FU could be an option in the management of severe endometriosis.

Real-Time Imaging of Resident T Cells in Human Lung and Ovarian Carcinomas Reveals How Different Tumor Microenvironments Control T Lymphocyte Migration

T cells play a key role in the battle against cancer. To perform their antitumor activities, T cells need to adequately respond to tumor antigens by establishing contacts with either malignant cells or antigen-presenting cells. These latter functions rely on a series of migratory steps that go from entry of T cells into the tumor followed by their locomotion in the tumor stroma. Our knowledge of how T cells migrate within tumors mainly comes from experiments performed in mouse models. Whereas such systems have greatly advanced our understanding, they do not always faithfully recapitulate the disease observed in cancer patients. We previously described a technique based on tissue slices that enables to track with real-time imaging microscopy the motile behavior of fluorescent T cells plated onto fresh sections of human lung tumors. We have now refined this approach to monitor the locomotion of resident tumor-infiltrating CD8 T cells labeled with fluorescently coupled antibodies. Using this approach, our findings reveal that CD8 T cells accumulate in the stroma of ovarian and lung carcinomas but move slowly in this compartment. Conversely, even though less populated, tumors islets were found to be zones of faster migration for resident CD8 T cells. We also confirm the key role played by collagen fibers, which, by their orientation, spacing and density, control the distribution and migration of resident CD8 T cells within the tumor stroma. We have subsequently demonstrated that, under some physical tissue constraints, CD8 T cells exhibited a mode of migration characterized by alternate forward and backward movements. In sum, using an ex vivo assay to track CD8 T cells in fresh human tumor tissues, we have identified the extracellular matrix as a major stromal component in influencing T cell migration, thereby impacting the control of tumor growth. This approach will aid in the development and testing of novel immunotherapy strategies to promote T cell migration in tumors.

Clinical practice guidelines from the French College of Gynecologists and Obstetricians (CNGOF): benign breast tumors – short text

Screening with breast ultrasound in combination with mammography is needed to investigate a clinical breast mass (Grade B), colored single-pore breast nipple discharge (Grade C), or mastitis (Grade C). The BI-RADS system is recommended for describing and classifying abnormal breast imaging findings. For a breast abscess, a percutaneous biopsy is recommended in the case of a mass or persistent symptoms (Grade C). For mastalgia, when breast imaging is normal, no MRI or breast biopsy is recommended (Grade C). Percutaneous biopsy is recommended for a BI-RADS category 4-5 mass (Grade B). For persistent erythematous nipple or atypical eczema lesions, a nipple biopsy is recommended (Grade C). For distortion and asymmetry, a vacuum core-needle biopsy is recommended due to the risk of underestimation by simple core-needle biopsy (Grade C). For BI-RADS category 4-5 microcalcifications without any ultrasound signal, a minimum 11-G vacuum core-needle biopsy is recommended (Grade B). In the absence of microcalcifications on radiography cores additional samples are recommended (Grade B). For atypical ductal hyperplasia, atypical lobular hyperplasia, lobular carcinoma in situ, flat epithelial atypia, radial scar and mucocele with atypia, surgical excision is commonly recommended (Grade C). Expectant management is feasible after multidisciplinary consensus. For these lesions, when excision margins are not clear, no new excision is recommended except for LCIS characterized as pleomorphic or with necrosis (Grade C). For grade 1 phyllodes tumor, surgical resection with clear margins is recommended. For grade 2 phyllodes tumor, 10mm margins are recommended (Grade C). For papillary breast lesions without atypia, complete disappearance of the radiological signal is recommended (Grade C). For papillary breast lesions with atypia, complete surgical excision is recommended (Grade C).

Evidence for the use of robotically assisted surgery in gynecologic cancers.

Purpose of review Robotically assisted laparoscopy has been introduced in the armamentarium of gynaecologic oncology surgeons. A lot of studies compared robotic surgery and laparotomy when the real issue is to demonstrate the interest and added value of robotically assisted laparoscopy versus standard laparoscopy. In this review, we will describe the most meaningful indications and advantages of robotically assisted laparoscopy in gynaecologic oncology. Recent findings The learning curve for advanced procedures in robot-assisted laparoscopy is shorter and easier than with the standard laparoscopy, especially for beginners. In most of the series, operating time is longer with robot, but complication rates are often decreased, especially in obese patients with a conversion rate to laparotomy that is decreased compared with standard laparoscopy. Robot-assisted laparoscopy can be used for surgery of high-risk endometrial cancer, staging of early-ovarian cancer, and pelvic exenteration in case of recurrent malignancies. Furthermore, more recent robots allow performing sentinel node biopsy in endometrial or cervical cancer using fluorescence detection with indocyanine green. Summary The spreading of robotic surgery led to an enhancement of minimal invasive surgical approach in general, and to the development of new indications in gynaecologic oncology. The superiority of robot-assisted laparoscopy still has to be demonstrated with properly designed trials.