A mouse model for endometrioid ovarian cancer arising from the distal oviduct.

Ovarian cancer is the deadliest gynecological malignancy in Western countries. Early detection, however, is hampered by the fact that the origin of ovarian cancer remains unclear. Knowing that in a high percentage of endometrioid ovarian cancers Wnt/ß-catenin signaling is activated, and in view of the hypothesis that ovarian cancer may originate from the distal oviduct, we studied mice in which Wnt/ß-catenin signaling was activated in Müllerian duct-derived tissues. Conditional adenomatous polyposis coli (Apc) knockout mice were used to study the activation of Wnt/ß-catenin signaling in Müllerian duct-derived organs. These Pgr(Cre/+);Apc(ex15lox/lox) mice (n = 44) were sacrificed at 10, 20, 40 and 80 weeks and uterus, oviduct, ovaries and surrounding fat tissues were assessed using immunohistochemistry. Using nuclear ß-catenin staining, Wnt/ß-catenin signaling activation was confirmed in the entire epithelium of the adult Müllerian duct (fimbriae, oviduct and endometrium), but was absent in ovarian surface epithelium cells (OSEs). Besides endometrial hyperplasia, in 87.2% of mice intraepithelial lesions of the distal oviduct were found, whereas OSEs remained unaffected. In addition, 62.5% of mice developed tumors in the distal and fimbrial part of the oviduct. In the ovaries, mainly at young age, in 16.3% of mice, simple epithelial cysts were noted, which developed further into endometrioid ovarian tumors, resembling human endometrioid ovarian cancer (27.9% of mice). Next to this, locoregional growth in the utero-ovarian ligament was also shown. Here, for the first time, mutations (activation of Wnt/ß-catenin) in the distal oviduct result in precursor lesions that develop into ovarian tumors, resembling human endometrioid ovarian cancer.

The prevention of colorectal anastomotic leakage with tissue adhesives in a contaminated environment is associated with the presence of anti-inflammatory macrophages.

BACKGROUND: Colorectal anastomoses created in a contaminated environment result in a high leakage rate. This study investigated whether using anastomotic sealants (TissuCol(®), Histoacryl(®) Flex, and Duraseal(®)) prevents leakage in a rat peritonitis model. STUDY DESIGN: Sixty-seven Wistar rats were divided into control and experimental groups (TissuCol, Histoacryl, and Duraseal groups). Peritonitis was induced 1 day before surgery with the cecal ligation puncture model. On day 0, colonic anastomosis was constructed with sutures and then sealed with no adhesive (control group) or one select adhesive (experimental groups). Bursting pressure, abscess formation, and adhesion severity were evaluated on day 3 or day 14. Hematoxylin and eosin staining and immunohistochemical staining for CD4, CD8, CD206, and iNOS were performed. RESULTS: On day 3, bursting pressures of the TissuCol group (120.1 ± 25.3 mmHg), Histoacryl group (117.3 ± 20.2 mmHg), and Duraseal group (123.6 ± 35.4 mmHg) were significantly higher than the that of the control group (24.4 ± 31.7 mmHg, p < 0.001). Abscesses around the anastomosis were found in the control group (6/7) and Duraseal group (2/9) but not in the TissuCol group or Histoacryl group. A higher number of CD206+ cells (M2 macrophages), a lower number of iNOS+ cells (M1 macrophages), a higher M2/M1 index, and a higher CD4+/CD8+ index were seen at the anastomotic site in all experimental groups compared with the control group on day 3. On day 14, abscesses were only found in the control group. Adhesion severity in the Duraseal group was significantly lower than that in the control group (p = 0.001). CONCLUSIONS: Anastomotic sealing using TissuCol(®), Histoacryl(®) Flex, or Duraseal(®) seems to be an effective and safe option to prevent leakage in contaminated colorectal surgery. The presence of large numbers of anti-inflammatory macrophages seems to be involved in preventing the leakage.

Quality and Quantity of Memories in Patients Who Undergo Awake Brain Tumor Resection.

BACKGROUND: Awake craniotomy is performed with increasing frequency for brain tumor surgery in eloquent areas; however, little is known about patients' memories of this procedure. Here we retrospectively analyzed the quality and quantity of memories in a series of patients treated following a standardized protocol. METHODS: We treated 61 consecutive patients within 3 years, 48 of whom were alive when the study was performed. Each of these patients received a questionnaire eliciting information about their perioperative memories and perceptions. The perioperative process was broken down into steps, and for each step the patient was to judge the quantity (nothing-everything) and quality (very negative-very positive) of his or her memories. RESULTS: Thirty-six of the 48 patients completed the questionnaire (75%). The quantity of memories was quite incomplete, even for intraoperative moments when patients were awake and cooperative. On average, the quality of memories was neutral or positive. A higher quantity of memories was associated with a higher quality of memories. The most commonly reported sources of discomfort were placement of the Mayfield clamp, followed by laying on the operating room table with movement restriction, and irritation by the urinary catheter in situ. CONCLUSIONS: Awake craniotomy can be performed following our protocol in such a way that it is experienced as (very) comfortable. However, there are moments of discomfort, which can be managed by the team. Extensive preoperative preparation may be considered a crucial part of the procedure. Less amnesia seems to improve patient satisfaction. The results of this study can help guide protocol optimization, expectation management, and information for future patients.

Clinical, mechanical, and immunohistopathological effects of tissue adhesives on the colon: An in-vivo study.

BACKGROUND: Tissue adhesives may be useful for sealing bowel anastomoses by preventing anastomotic leakage. Prior to clinical implementation, an in-depth analysis of the clinical and immunohistopathological effects of tissue adhesives on the target tissue and of the mechanical strength of the adhesive bond in an in vivo model is needed. MATERIALS AND METHODS: In 84 rats, two bowel segments were glued using one of the following tissue adhesive: Bioglue, Gelatin-resorcinol-formaldehyde (GRF), Glubran 2, Histoacryl Flex, Omnex, Duraseal Xact, or Tissucol. Rats were followed for 7 or 28 days. Endpoints were clinical complication rate, mechanical strength, and immunohistopathological reactions. RESULTS: Of the seven tissue adhesives, GRF and Bioglue showed the highest rates of bowel wall destruction and ileus and the most severe immunohistopathological tissue reactions at 7 and 28 days. Cyanoacrylates (Histoacryl Flex, Omnex, Glubran 2) showed high mechanical strength and mild immunohistopathological reactions at 7 and 28 days. Duraseal Xact and Tissucol were the most inert tissue adhesives, but exhibited low mechanical strength. At 28 days, mechanical strength was significantly correlated to CD8, CD68, and Ki67 cell counts. CONCLUSION: Based on the clinical and immunohistopathological outcomes, GRF and Bioglue were found to be the least suitable tissue adhesives for colonic use. Duraseal Xact and Tissucol were inert but also showed low mechanical strength. Cyanoacrylates exhibited mild clinical and immunohistopathological effects while maintaining high strength, which makes them promising as colonic sealants. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 846-854, 2017.

Colorectal anastomotic leakage caused by insufficient suturing after partial colectomy: a new experimental model.

BACKGROUND: Colorectal anastomotic leakage (CAL) is the most important complication of colorectal surgery, accounting for one third of post-operative deaths. To prevent it, many interventions have been tested in animal models, mostly rats. However, few of these models have been validated. We aimed to develop a reproducible rat CAL model by creating an anastomosis with insufficient suturing after partial colectomy. METHODS: To establish the number of sutures that would create an appropriate leakage rate for research (20%-50%), partial colectomy was performed in 60 Wistar rats using a 12-suture anastomosis in the control group and an anastomosis with insufficient suturing in the experimental group, starting with five sutures. Seven days later, the rats were examined for the occurrence and severity of CAL, the presence of adhesions, and anastomotic bursting pressure. When an acceptable leakage rate was achieved, the experimental and control studies were repeated twice to confirm the adequacy of the chosen technique. RESULTS: On day 7, five-suture and 12-suture anastomoses had leakage rates of 50% vs. 30%, 44.4% vs. 20%, and 50% vs. 20%, respectively, in the various series. Overall, the five-suture group had a significantly higher CAL rate than did the 12-suture group (48.3% vs. 23.3%; p=0.045). It also had higher CAL severity and more adhesions (p for both<0.05). The bursting pressure of these anastomoses was significantly lower than that in the 12-suture group (116.8±58.9 mm Hg vs. 150.4±50.3 mm Hg; p=0.041). CONCLUSION: Anastomosis with five sutures after partial colectomy provides a suitable rat CAL model. Its future applications may help to improve the consistency of CAL studies.

Interaction between sex hormones and WNT/ß-catenin signal transduction in endometrial physiology and disease.

Wnt/ß-catenin signalling plays a rate-limiting role in early development of many different organs in a broad spectrum of organisms. In the developing Müllerian duct, Wnt/ß-catenin signalling is important for initiation, outgrowth, patterning and differentiation into vagina, cervix, uterus and oviducts. In adult life, sex hormones modulate Wnt/ß-catenin signalling in the endometrium to maintain the monthly balance between estrogen-induced proliferation and progesterone-induced differentiation, and enhanced Wnt/ß-catenin signalling seems to be involved in endometrial carcinogenesis. However, early in pregnancy enhanced Wnt/ß-catenin signalling is prerequisite for proper implantation and invasion of trophoblast cells into endometrium and myometrium thus helping to form a placenta. Overall, it seems that tight control of Wnt/ß-catenin signalling in time and space is important for initiation, development and normal function of the female reproductive tract. However, if Wnt/ß-catenin signalling is not kept in check, it easily seems to initiate or contribute to development of a number of uterine disorders.

Identification of quiescent, stem-like cells in the distal female reproductive tract.

In fertile women, the endometrium undergoes regular cycles of tissue build-up and regression. It is likely that uterine stem cells are involved in this remarkable turn over. The main goal of our current investigations was to identify slow-cycling (quiescent) endometrial stem cells by means of a pulse-chase approach to selectively earmark, prospectively isolate, and characterize label-retaining cells (LRCs). To this aim, transgenic mice expressing histone2B-GFP (H2B-GFP) in a Tet-inducible fashion were administered doxycycline (pulse) which was thereafter withdrawn from the drinking water (chase). Over time, dividing cells progressively loose GFP signal whereas infrequently dividing cells retain H2B-GFP expression. We evaluated H2B-GFP retaining cells at different chase time points and identified long-term (LT; >12 weeks) LRCs. The LT-LRCs are negative for estrogen receptor-α and express low levels of progesterone receptors. LRCs sorted by FACS are able to form spheroids capable of self-renewal and differentiation. Upon serum stimulation spheroid cells are induced to differentiate and form glandular structures which express markers of mature mullerian epithelial cells. Overall, the results indicate that quiescent cells located in the distal oviduct have stem-like properties and can differentiate into distinct cell lineages specific of endometrium, proximal and distal oviduct. Future lineage-tracing studies will elucidate the role played by these cells in homeostasis, tissue injury and cancer of the female reproductive tract in the mouse and eventually in man.

Progesterone inhibits epithelial-to-mesenchymal transition in endometrial cancer.

BACKGROUND: Every year approximately 74,000 women die of endometrial cancer, mainly due to recurrent or metastatic disease. The presence of tumor infiltrating lymphocytes (TILs) as well as progesterone receptor (PR) positivity has been correlated with improved prognosis. This study describes two mechanisms by which progesterone inhibits metastatic spread of endometrial cancer: by stimulating T-cell infiltration and by inhibiting epithelial-to-mesenchymal cell transition (EMT). METHODOLOGY AND PRINCIPAL FINDINGS: Paraffin sections from patients with (n = 9) or without (n = 9) progressive endometrial cancer (recurrent or metastatic disease) were assessed for the presence of CD4+ (helper), CD8+ (cytotoxic) and Foxp3+ (regulatory) T-lymphocytes and PR expression. Progressive disease was observed to be associated with significant loss of TILs and loss of PR expression. Frozen tumor samples, used for genome-wide expression analysis, showed significant regulation of pathways involved in immunesurveillance, EMT and metastasis. For a number of genes, such as CXCL14, DKK1, DKK4, PEG10 and WIF1, quantitive RT-PCR was performed to verify up- or downregulation in progressive disease. To corroborate the role of progesterone in regulating invasion, Ishikawa (IK) endometrial cancer cell lines stably transfected with PRA (IKPRA), PRB (IKPRB) and PRA+PRB (IKPRAB) were cultured in presence/absence of progesterone (MPA) and used for genome-wide expression analysis, Boyden- and wound healing migration assays, and IHC for known EMT markers. IKPRB and IKPRAB cell lines showed MPA induced inhibition of migration and loss of the mesenchymal marker vimentin at the invasive front of the wound healing assay. Furthermore, pathway analysis of significantly MPA regulated genes showed significant down regulation of important pathways involved in EMT, immunesuppression and metastasis: such as IL6-, TGF-ß and Wnt/ß-catenin signaling. CONCLUSION: Intact progesterone signaling in non-progressive endometrial cancer seems to be an important factor stimulating immunosurveilance and inhibiting transition from an epithelial to a more mesenchymal, more invasive phenotype.