Ovarian follicles of young patients with Turner's syndrome contain normal oocytes but monosomic 45,X granulosa cells.

STUDY QUESTION: What is the X chromosomal content of oocytes and granulosa cells of primordial/primary (small) follicles and stromal cells in ovaries of young patients with Turner's syndrome (TS)? SUMMARY ANSWER: Small ovarian follicles were detected in one-half of the patients studied, and X chromosome analysis revealed that most oocytes were normal, granulosa cells were largely monosomic, while stromal cells showed a high level of mosaicism. WHAT IS KNOWN ALREADY: Most women with TS experience a premature reduction or complete loss of fertility due to an accelerated loss of gametes. To determine whether fertility preservation in this group of patients is feasible, there is a strong need for information on the X chromosomal content of ovarian follicular and stromal cells. STUDY DESIGN, SIZE, DURATION: Small follicles (<50 µm) and stromal cells were isolated from ovarian tissue of young TS patients and analysed for their X chromosomal content. In addition to ovarian cells, several other cell types from the same patients were analysed. PARTICIPANTS/MATERIALS, SETTING, METHODS: After unilateral ovariectomy, ovarian cortex tissue was obtained from 10 TS patients (aged 2-18 years) with numerical abnormalities of the X chromosome. Ovarian cortex fragments were prepared and cryopreserved. One fragment from each patient was thawed and enzymatically digested to obtain stromal cells and primordial/primary follicles. Stromal cells, granulosa cells and oocytes were analysed by FISH using an X chromosome-specific probe. Extra-ovarian cells (lymphocytes, buccal cells and urine cells) of the same patients were also analysed by FISH. Ovarian tissue used as control was obtained from individuals undergoing oophorectomy as part of their gender affirming surgery. MAIN RESULTS AND THE ROLE OF CHANCE: Ovarian follicles were detected in 5 of the 10 patients studied. A method was developed to determine the X chromosomal content of meiosis I arrested oocytes from small follicles. This revealed that 42 of the 46 oocytes (91%) that were analysed had a normal X chromosomal content. Granulosa cells were largely 45,X but showed different levels of X chromosome mosaicism between patients and between follicles of the same patient. Despite the presence of a low percentage (10-45%) of 46,XX ovarian cortex stromal cells, normal macroscopic ovarian morphology was observed. The level of mosaicism in lymphocytes, buccal cells or urine-derived cells was not predictive for mosaicism in ovarian cells. LIMITATIONS, REASONS FOR CAUTION: The results are based on a small number (n = 5) of TS patient samples but provide evidence that the majority of oocytes have a normal X chromosomal content and that follicles from the same patient can differ with respect to the level of mosaicism of their granulosa cells. The functional consequences of these observations require further investigation. WIDER IMPLICATIONS OF THE FINDINGS: The results indicate that despite normal ovarian and follicular morphology, stromal cells and granulosa cells of small follicles in patients with TS may display a high level of mosaicism. Furthermore, the level of mosaicism in ovarian cells cannot be predicted from the analysis of extra-ovarian tissue. These findings should be considered by physicians when offering cryopreservation of ovarian tissue as an option for fertility preservation in young TS patients. STUDY FUNDING/COMPETING INTEREST(S): Unconditional funding was received from Merck B.V. The Netherlands (Number A16-1395) and the foundation 'Radboud Oncologie Fonds' (Number KUN 00007682). The authors have no conflicts of interest. TRIAL REGISTRATION NUMBER: NCT03381300.

Vaginal Colpectomy in Transgender Men: A Retrospective Cohort Study on Surgical Procedure and Outcomes.

BACKGROUND: Colpectomy, removal of the vaginal epithelium, may be performed in transgender men because of a disturbed male self-image, to reduce vaginal discharge, or to reduce the risk of fistula formation at the urethral-neourethral junction in future phalloplasty or metaidoioplasty. AIM: To demonstrate that vaginal colpectomy in transgender men, either alone or in combination with, for example, laparoscopic hysterectomy, metaidoioplasty, scrotoplasty, or urethroplasty, is a feasible procedure. METHODS: This single-center retrospective cohort study included 143 transgender men who underwent vaginal colpectomy between January 2006 and April 2018. Surgical details and clinical outcomes were collected from all patients. OUTCOMES: The primary outcome was the number of perioperative and postoperative complications, including intraoperative blood loss. Secondary outcomes were operating time, change in hemoglobin level, and duration of hospital stay. RESULTS: In 109 patients (76%), the procedure consisted of colpectomy only, whereas in 34 patients (23%), colpectomy was combined with other procedures. In the whole group (combined procedures included), the median blood loss was 300 mL (interquartile range [IQR] = 250 mL), the mean operating time was 132 ± 62 minutes, and the mean duration of hospital admission was 3.6 ± 1.9 days. In the colpectomy-only group, the median blood loss was 300 mL (IQR = 250 mL), mean operating time was 112 ± 40 minutes, and mean duration of hospital admission was 3.2 ± 1.5 days. For the total group, 15 patients (10%) experienced a major perioperative complication (ie, bowel injury, ureter injury, urethra injury, bladder injury, hemorrhage requiring transfusion and/or intervention and conversion to laparoscopy), and 1 patient (0.7%) had a minor perioperative complication (hemorrhage). Major postoperative complications (hemorrhage, hematoma, fistula, wound infection and prolonged pain complaints) were reported in 17 patients (12%), and minor postoperative complications (urinary tract infection, urinary retention, hemorrhage, and hematoma) occurred in 50 patients (35%). CLINICAL IMPLICATIONS: This study provides a detailed description of our technique and comprehensive reporting on perioperative and postoperative complications and reintervention rate. STRENGTHS & LIMITATIONS: Study strengths include the large number of patients included and the detailed reporting on the complications of vaginal colpectomy. The main limitation is the retrospective design, which can cause data to go missing during extraction and is prone to bias. CONCLUSION: Vaginal colpectomy is a procedure with a high complication rate, but its advantages seem to outweigh its disadvantages. In all but 1 case, no long-term sequelae were reported. However, the high complication rate and reintervention rate should be discussed with patients who are considering undergoing this procedure. Nikkels C, van Trotsenburg M, Huirne J, et al. Vaginal Colpectomy in Transgender Men: A Retrospective Cohort Study on Surgical Procedure and Outcomes. J Sex Med 2019;16:924-933.

Colpectomy Significantly Reduces the Risk of Urethral Fistula Formation after Urethral Lengthening in Transgender Men Undergoing Genital Gender Affirming Surgery.

PURPOSE: We assessed the effect of performing colpectomy before (primary) or after (secondary) gender affirming surgery with single stage urethral lengthening on the incidence of urethral fistula in transgender men. MATERIALS AND METHODS: We retrospectively reviewed the charts of all transgender men who underwent gender affirming surgery with urethral lengthening between January 1989 and November 2016 at VU University Medical Center. Patient demographics, surgical characteristics, fistulas and fistula management, and primary and secondary colpectomy were recorded. Descriptive statistics were calculated and incidence rates were compared. RESULTS: A total of 294 transgender men underwent gender affirming surgery with urethral lengthening. A urethral fistula developed in 111 of the 232 patients (48%) without colpectomy and in 13 of the 62 (21%) who underwent primary colpectomy (p <0.01). Secondary colpectomy resulted in 100% fistula closure when performed in 17 patients with recurrent urethral fistula at the proximal urethral anastomosis and the fixed part of the neourethra. CONCLUSIONS: Primary colpectomy decreases the incidence rate of urethral fistulas. Secondary colpectomy is also an effective treatment of fistulas at the proximal urethral anastomosis and the fixed part of the neourethra.

Robot-assisted laparoscopic colpectomy in female-to-male transgender patients; technique and outcomes of a prospective cohort study.

BACKGROUND: Gender-affirming surgeries in female-to-male (FtM) transgender patients include mostly hysterectomy, bilateral salpingo-oophorectomy and mastectomy. Sometimes further surgery is performed, such as phalloplasty. Colpectomy may be performed to overcome gender dysphoria and disturbing vaginal discharge; furthermore, it may be important in reducing the risk of fistulas due to the phalloplasty procedure with urethral elongation. Colpectomy prior to the reconstruction of the neourethra seems to reduce fistula rates on the very first anastomosis. Therefore, at our center, colpectomy has become a standard procedure prior to phalloplasty and metoidioplasty with urethral elongation. Colpectomy is known as a procedure with potentially serious complications, e.g., extensive bloodloss, vesicovaginal fistula or rectovaginal fistula. Colpectomy performed via the vaginal route can be a challenging procedure due to lack of exposure of the surgical field, as many patients are virginal. Therefore, we investigated whether robot-assisted laparoscopic hysterectomy with bilateral salpingo-oophorectomy (TLH-BSO) followed by robot-assisted laparoscopic colpectomy (RaLC) is an alternative for the vaginal approach. METHODS: Robot TLH/BSO and RaLC as a single-step procedure was performed in 36 FtM patients in a prospective cohort study. RESULTS: Median length of the procedure was 230 min (197-278), which reduced in the second half of the patients, median blood loss was 75 mL (30-200), and median discharge was 3 days (2-3) postoperatively. One patient with a major complication (postoperative bleeding with readmission and transfusion) was reported. CONCLUSION: To our knowledge, this is the first report of RaLC. Our results show that RaLC combined with robot TLH-BSO is feasible as a single-step surgical procedure in FtM transgender surgery. Future studies are needed to compare this technique to the two-step surgical approach and on its outcome and complication rates of subsequent phalloplasty.

Completing or Abandoning Radical Hysterectomy in Early-Stage Lymph Node-Positive Cervical Cancer: Impact on Disease-Free Survival and Treatment-Related Toxicity.

INTRODUCTION: Management regarding completing hysterectomy in case of intraoperative finding of positive lymph nodes in early-stage cervical cancer differs between institutions. The aim of this study was to compare survival and toxicity after completed hysterectomy followed by adjuvant (chemo-)radiotherapy versus abandoned hysterectomy and primary treatment with chemoradiotherapy (CRT). METHODS: A retrospective multicenter cohort study was performed. All patients were scheduled for radical hysterectomy with pelvic lymphadenectomy (RHL). In the RHL group, hysterectomy was completed followed by adjuvant (chemo-)radiotherapy. In the second group, hysterectomy was abandoned, and CRT was conducted. Primary outcomes were disease-free survival (DFS) and overall survival. A multivariable analysis on DFS was performed. Toxicity was scored according to the National Cancer Institute CTCAE (Common Terminology Criteria for Adverse Events) v4.03. RESULTS: A total of 121 patients were included (RHL, n = 89; CRT, n = 32). There was no difference in overall survival (84% vs 77%). Five-year DFS was in favor of completing RHL (81% vs 67%). Multivariable analysis showed that, corrected for lymph node variables, treatment regimen was not associated with DFS. After RHL, pelvic recurrence rate was significantly lower compared with CRT (2% vs 16%). CTCAE grade 3-4 toxicity rates were higher in the CRT compared with the RHL group (59% vs 30%), mainly because of differences in chemotherapy-related hematologic toxicity. CONCLUSIONS: In patients with clinically N0 early-stage cervical cancer with intraoperative detection of positive nodes, completing RHL followed by adjuvant (chemo-)radiotherapy may result in a better pelvic control compared with abandoning hysterectomy and treatment with chemoradiotherapy. However, if corrected for lymph node variables, treatment (RHL or CRT) was not associated with DFS.

Hypoxia-inducible factor-1 stimulates postnatal lung development but does not prevent O2-induced alveolar injury.

This study investigated whether hypoxia-inducible factor (HIF)-1 influences postnatal vascularization and alveologenesis in mice and whether stable (constitutive-active) HIF could prevent hyperoxia-induced lung injury. We assessed postnatal vessel and alveolar formation in transgenic mice, expressing a stable, constitutive-active, HIF1α-subunit (HIF-1αΔODD) in the distal lung epithelium. In addition, we compared lung function, histology, and morphometry of neonatal transgenic and wild-type mice subjected to hyperoxia. We found that postnatal lungs of HIF-1αΔODD mice had a greater peripheral vessel density and displayed advanced alveolarization compared with control lungs. Stable HIF-1α expression was associated with increased postnatal expression of angiogenic factors, including vascular endothelial growth factor, angiopoietins 1 and 2, Tie2, and Ephrin B2 and B4. Hyperoxia-exposed neonatal HIF-1αΔODD mice exhibited worse lung function but had similar histological and surfactant abnormalities compared with hyperoxia-exposed wild-type controls. In conclusion, expression of constitutive-active HIF-1α in the lung epithelium was associated with increased postnatal vessel growth via up-regulation of angiogenic factors. The increase in postnatal vasculature was accompanied by enhanced alveolar formation. However, stable HIF-1α expression in the distal lung did not prevent hyperoxia-induced lung injury in neonates but instead worsened lung function.

Tissue Engineering Neovagina for Vaginoplasty in Mayer-Rokitansky-Küster-Hauser Syndrome and Gender Dysphoria Patients: A Systematic Review.

Background: Vaginoplasty is a surgical solution to multiple disorders, including Mayer-Rokitansky-Küster-Hauser syndrome and male-to-female gender dysphoria. Using nonvaginal tissues for these reconstructions is associated with many complications, and autologous vaginal tissue may not be sufficient. The potential of tissue engineering for vaginoplasty was studied through a systematic bibliography search. Cell types, biomaterials, and signaling factors were analyzed by investigating advantages, disadvantages, complications, and research quantity. Search Methods: A systematic search was performed in Medline, EMBASE, Web of Science, and Scopus until March 8, 2022. Term combinations for tissue engineering, guided tissue regeneration, regenerative medicine, and tissue scaffold were applied, together with vaginoplasty and neovagina. The snowball method was performed on references and a Google Scholar search on the first 200 hits. Original research articles on human and/or animal subjects that met the inclusion (reconstruction of vaginal tissue and tissue engineering method) and no exclusion criteria (not available as full text; written in foreign language; nonoriginal study article; genital surgery other than neovaginal reconstruction; and vaginal reconstruction with autologous or allogenic tissue without tissue engineering or scaffold) were assessed. The Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) checklist, the Newcastle-Ottawa Scale, and the Gold Standard Publication Checklist were used to evaluate article quality and bias. Outcomes: A total of 31 out of 1569 articles were included. Data extraction was based on cell origin and type, biomaterial nature and composition, host species, number of hosts and controls, neovaginal size, replacement fraction, and signaling factors. An overview of used tissue engineering methods for neovaginal formation was created, showing high variance of cell types, biomaterials, and signaling factors and the same topics were rarely covered multiple times. Autologous vaginal cells and extracellular matrix-based biomaterials showed preferential properties, and stem cells carry potential. However, quality confirmation of orthotopic cell-seeded acellular vaginal matrix by clinical trials is needed as well as exploration of signaling factors for vaginoplasty. Impact statement General article quality was weak to sufficient due to unreported cofounders and incomplete animal study descriptions. Article quality and heterogenicity made identification of optimal cell types, biomaterials, or signaling factors unreliable. However, trends showed that autologous cells prevent complications and compatibility issues such as healthy cell destruction, whereas stem cells prevent cross talk (interference of signaling pathways by signals from other cell types) and rejection (but need confirmation testing beyond animal trials). Natural (orthotopic) extracellular matrix biomaterials have great preferential properties that encourage future research, and signaling factors for vascularization are important for tissue engineering of full-sized neovagina.

Effectiveness, complications, and reproductive outcomes after cesarean scar pregnancy management: a retrospective cohort study.

BACKGROUND: There is a dramatic rise in cesarean deliveries worldwide, leading to higher complication rates in subsequent pregnancies. One of these complications is a cesarean scar pregnancy. During the last decades, treatment options for cesarean scar pregnancies have changed, and less invasive interventions have been employed to preserve fertility and reduce morbidity. However, the optimal treatment approach and the influence of various treatments on reproductive outcomes have to be determined. OBJECTIVE: This study aimed to evaluate the short- and long-term outcomes after cesarean scar pregnancy management. STUDY DESIGN: We conducted a retrospective cohort study of women determined to have a cesarean scar pregnancy from 2010 to 2021 at a tertiary referral center, the Amsterdam University Medical Center, in the Netherlands. Outcomes of the following management strategies were compared: expectant management, methotrexate, curettage with temporary cervical cerclage, or a laparoscopic niche resection. We performed a curettage if the cesarean scar pregnancy did not cross the serosal line of the uterus, and a laparoscopic niche resection was performed if the cesarean scar pregnancy crossed the serosal line. The main outcomes were treatment efficacy and time to subsequent ongoing pregnancy or pregnancy leading to a live birth. RESULTS: Of the 60 included women, 5 (8.3%) were managed expectantly, 8 (13.3%) were treated with methotrexate, 31 (51.8%) were treated with a curettage, and 16 (26.7%) with a laparoscopic niche resection. The groups were not comparable. The gestational age and human chorionic gonadotropin levels were generally higher in women who received methotrexate or a laparoscopic niche resection. Additional treatment in the conservative group was needed for 4 (80%) women after expectant management and for 7 (87.5%) women after methotrexate treatment. In the surgical group, all 31 women treated with a curettage and all 16 treated with a laparoscopic niche resection did not require additional treatment. The subsequent ongoing pregnancy rate after cesarean scar pregnancy management was 81.1% (30/37) among women who wished to conceive, with a live birth rate of 78.4% (29/37); 1 woman was in her third trimester of pregnancy at the time of analyses. The time between cesarean scar pregnancy management and subsequent ongoing pregnancy was 4 months (interquartile range, 3-6; P=.02) after expectant management, 18 months (interquartile range, 13-22) after initial methotrexate treatment, 5 months (interquartile range, 3-14; P=.01) after a curettage, and 6 months (interquartile range, 4-15; P=.03) after a laparoscopic niche resection. CONCLUSION: Surgical treatment of a cesarean scar pregnancy led to a high success rate without additional interventions, high pregnancy rates with a short time interval between treatment, and subsequent pregnancy leading to an ongoing pregnancy or live birth. Conservative management, both with expectant management and methotrexate treatment, led to high (re)intervention rates. Different management approaches are indicated for different types of cesarean scar pregnancies.

Creation of a decellularized vaginal matrix from healthy human vaginal tissue for potential vagina reconstruction: experimental studies.

BACKGROUND: When a disorder causes the absence of a healthy, full-size vagina, various neovaginal creation methods are available. Sometimes dilation or stretching of the vaginal cavity is sufficient, but intestinal or dermal flap tissue is generally required. However, different inherent tissue properties cause complications. Therefore, a lost body part should be replaced with a similar material. The use of organ-specific acellular vaginal tissue carries great potential, as its similar architecture and matrix composition make it suitable for vaginal regeneration. METHODS: The authors developed an optimized protocol for decellularization of healthy human vaginal tissue. Resected colpectomy tissue from 12 healthy transgender patients was used. Successful decellularization was confirmed by applying acellular criteria from in-vivo remodeling reports. Suitability as a tissue-mimicking scaffold for vaginal reconstruction was determined by visible structural features, biocompatibility during stretching, and the presence of visible collagen, elastin, laminin, and fibronectin. RESULTS: Histological examination confirmed the preservation of structural features, and minimal cellular residue was seen during fluorescence microscopy, DNA and RNA quantification, and fragment length examination. Biomechanical testing showed decreased peak load (55%, P <0.05), strain at rupture (23%, P <0.01), and ultimate tensile stress (55%, P <0.05) after decellularization, while the elastic modulus (68%) did not decrease significantly. Fluorescence microscopy revealed preserved Fibronectin-I/II/III and Laminin-I/II, while Collagen-I and Ficolin-2B were decreased but mostly retained. CONCLUSIONS: The absence of cellular residue, moderately altered biomechanical extracellular matrix properties, and mostly preserved structural proteins appear to make our decellularized human vaginal matrix a suitable tissue-mimicking scaffold for vagina transplantation when tissue survival through vascularization and innervation are accomplished in the future.

Effectiveness of laparoscopic niche resection versus expectant management in patients with unexplained infertility and a large uterine caesarean scar defect (uterine niche): protocol for a randomised controlled trial (the LAPRES study).

INTRODUCTION: A uterine niche is a defect at the site of the uterine caesarean scar that is associated with gynaecological symptoms and infertility. Promising results are reported in cohort studies after a laparoscopic niche resection concerning reduction of gynaecological symptoms in relation to baseline and concerning pregnancy outcomes. However, randomised controlled trials to study the effect of a laparoscopic niche resection on reproductive outcomes in infertile women are lacking. This study will answer the question if laparoscopic niche resection in comparison to expectant management improves reproductive outcomes in infertile women with a large uterine niche. METHODS AND ANALYSIS: The LAPRES study is a randomised, non-blinded, controlled trial, including 200 infertile women with a total follow-up of 2 years. Women with the presence of a large niche in the uterine caesarean scar and unexplained infertility of at least 1 year or failed IVF will be randomly allocated to a laparoscopic niche resection within 6 weeks or to expectant management for at least 9 months. A large niche is defined as a niche with a depth of >50% of the myometrial thickness and a residual myometrium of ≤3 mm on transvaginal ultrasound. Those receiving expectant management will be allowed to receive fertility therapies, including assisted reproductive techniques, if indicated. The primary outcome is time to ongoing pregnancy, defined as a viable intrauterine pregnancy at 12 weeks' gestation. Secondary outcome measures are time to conception leading to a live birth, other pregnancy outcomes, received fertility therapies after randomisation, menstruation characteristics, patient satisfaction, quality of life, additional interventions, and surgical and ultrasound outcomes (intervention group). Questionnaires will be filled out at baseline, 6, 12 and 24 months after randomisation. Ultrasound evaluation will be performed at baseline and at 3 months after surgery. ETHICS AND DISSEMINATION: The study protocol was approved by the medical ethics committee of the Amsterdam University Medical Centre. (Ref. No. 2017.030). Participants will sign a written informed consent before participation. The results of this study will be submitted to a peer-reviewed journal for publication. TRIAL REGISTRATION NUMBER DUTCH TRIAL REGISTER REF NO NL6350 : http://www.trialregister.nl.

The effect of colpectomy on lower urinary tract function in transgender men.

Background: In transgender men, effects of colpectomy on voiding function are unknown, except for the incidence rates of urinary tract infections and urinary retention. Aims: To provide insight into the effect of colpectomy on Lower Urinary Tract Function (LUTF) in transgender men. Methods: A retrospective chart review was conducted among transgender men who underwent colpectomy between January 2018 and October 2020. Primary outcomes were objective and subjective changes in voiding. Secondary outcomes were transurethral catheterization length and the need for clean intermittent self-catheterization (CISC). Results: Of 132 men, 89 (67%) underwent Robot-assisted Laparoscopic Colpectomy (RaLC) and 43 (33%) Vaginal Colpectomy (VC). Maximum flow rate on uroflowmetry decreased following RaLC (mean of 29.1 vs. 38.3 mL/s, p = 0.002) and VC (mean of 29.2 vs. 40.3 mL/s, p < 0.001) after a median of four months postoperatively. An increase in total International Prostate Symptom Score was seen more frequently following VC compared to RaLC. Subjective changes were indicated by 39%, more often by men who underwent VC, of which the majority improved during the first months postoperatively. Trial without catheter (TWOC) on the first postoperative day was more successful after RaLC (79/89, 89%) than VC (24/43, 56%). Secondary TWOC was successful in 22/132 (17%) patients after a median of eight days postoperatively. In 5/132 (4%) men (three VC and two RaLC), temporary CISC was necessary for a period ranging from 5 to 21 days. The last 2/132 (2%) men after RaLC were still performing CISC at end of follow-up. Eventually, 5% (two VC and four RaLC) had to refrain from genital gender-affirming surgery with urethral lengthening due to voiding dysfunction. Discussion: After colpectomy, most objective and subjective worsening in LUTF is of a temporary nature, however, 5% had to refrain from genital gender-affirming surgery with urethral lengthening due to persistent voiding dysfunction, despite the desire to void while standing.

Assessing the use of tumor-specific DARPin-toxin fusion proteins for ex vivo purging of cancer metastases from human ovarian cortex before autotransplantation.

OBJECTIVE: To assess the use of tumor-specific designed ankyrin repeat proteins (DARPins) fused to a domain of Pseudomonas aeruginosa exotoxin A for purging of cancer metastases from the ovarian cortex. DESIGN: Experimental study. SETTING: University medical center. PATIENT(S): Human ovarian cortex. INTERVENTION(S): Ovarian cortex harboring artificially induced breast cancer metastases was treated with DARPins targeted to epithelial cell adhesion molecule (EpCAM) and human epidermal growth factor receptor 2 (HER2). MAIN OUTCOME MEASURE(S): The presence of any remaining cancer cells after purging was analyzed by (immuno)histochemistry and reverse transcriptase polymerase chain reaction. Effects on the viability of the ovarian cortex were determined by (immuno)histology, a follicular viability assay, and an assay to determine the in vitro growth capacity of small follicles. RESULT(S): After purging with EpCAM-targeted DARPin, all EpCAM-positive breast cancer cells were eradicated from the ovarian cortex. Although treatment had no effect on the morphology or viability of small follicles, a sharp decrease in oocyte viability during in vitro growth was observed, presumably due to low-level expression of EpCAM on oocytes. The HER2-targeted DARPins had no detrimental effects on the morphology, viability, or in vitro growth of small follicles. HER2-positive breast cancer foci were fully eliminated from the ovarian cortex, and the reverse transcriptase polymerase chain reaction showed a decrease to basal levels of HER2 mRNA after purging. CONCLUSION(S): Purging cancer metastases from ovarian cortex without impairing ovarian tissue integrity is possible by targeting tumor cell surface proteins with exotoxin A-fused DARPins. By adapting the target specificity of the cytotoxic DARPin fusions, it should be possible to eradicate metastases from all types of malignancies.

Perineal Cyst in Transgender Men: A Rare Complication Following Gender Affirming Surgery - A Case Series and Literature Overview.

INTRODUCTION: Genital gender affirming surgery (gGAS) is usually the final stage in the medical transition for transgender men and consists of creating a neophallus and neo-scrotum, with or without urethral lengthening(UL). To reduce the complication risks of UL, a mandatory colpectomy is performed prior to UL. Colpectomy is considered a complex surgery, which may lead to various perioperative complications. There are few long-term complications reported. AIM: To describe the clinical presentation and management of 3 consecutive transgender men presenting with a perineal cyst following gGAS. METHODS: After obtaining informed consent all clinical data was collected, including medical history, current symptoms, imaging, as well as surgery and histological outcomes. Furthermore, a literature search was performed. MAIN OUTCOME MEASURE: To hypothesize the aetiology of the perineal cyst based on current published literature. RESULTS: Three otherwise healthy transgender men, ages 26-46 with a similar medical history, presented with a perineal cyst several months or years following colpectomy and gGAS with UL. All patients underwent surgery to remove the cyst. Several theories regarding aetiology of this perineal cyst are discussed in this report. CONCLUSION: There remain several gaps in our knowledge regarding the aetiology and management of this perineal cyst. Therefore, further research is necessary. Asseler JD, Ronkes BL, Groenman FA, et al. Perineal Cyst in Transgender Men: A Rare Complication Following Gender Affirming Surgery - A Case Series and Literature Overview. J Sex Med 2021;9:100415.

Combining total laparoscopic hysterectomy and bilateral salpingo-oophorectomy with subcutaneous mastectomy in trans men: The effect on safety outcomes.

Background: Masculinizing mastectomy is the most requested gender affirming surgery (GAS) in trans men, followed by genital GAS. Mastectomy and total laparoscopic hysterectomy, with or without bilateral salpingo-oophorectomy (TLH ± BSO), can both be performed in one single operation session. However, data on complication rates of the combined procedure is scarce and no consensus exists on the preferred order of procedures. Aims: To compare safety outcomes between mastectomy performed in a single procedure with those when performed in a combined procedure and assess whether the order of procedures matters when they are combined. Methods: A retrospective chart review was performed of trans men who underwent masculinizing mastectomy with or without TLH ± BSO in a combined session. The effects of the surgical procedure on complication and reoperation rate of the chest were assessed using logistic regression. Results: In total, 480 trans men were included in the study. Of these, 212 patients underwent the combined procedure. The gynecological procedure was performed first in 152 (71.7%) patients. In the total sample, postoperative hematoma of the chest occurred in 11.3%; 16% in the combined versus 7.5% in the single mastectomy group (p = 0.001). Reoperations due to hematoma of the chest were performed in 7.5% of all patients; 10.8% in the combined versus 4.9% in the single mastectomy group (p = 0.017). The order of procedures in the combined group had no significant effect on postoperative hematoma of the chest (p = 0.856), and reoperations (p = 0.689). Conclusion: Combining masculinizing mastectomy with TLH ± BSO in one session was associated with significantly more hematoma and reoperations compared with separately performing mastectomy. This increased risk of complications after a combined procedure should be considered when deciding on surgical options. The order of procedures in a combined procedure did not have an effect on safety outcomes.

Role of angiogenesis in adenomyosis-associated abnormal uterine bleeding and subfertility: a systematic review.

BACKGROUND: Adenomyosis commonly occurs with abnormal uterine bleeding (AUB) and is associated with subfertility and a higher miscarriage rate. Recent evidence showed abnormal vascularization in the endometrium in patients with adenomyosis, suggesting a role of angiogenesis in the pathophysiology of AUB and subfertility in adenomyosis and providing a possible treatment target. OBJECTIVE AND RATIONALE: We hypothesized that the level of abnormal vascularization and expression of angiogenic markers is increased in the ectopic and eutopic endometrium of adenomyosis patients in comparison with the endometrium of control patients. This was investigated through a search of the literature. SEARCH METHODS: A systematic search was performed in PubMed and Embase until February 2019. Combinations of terms for angiogenesis and adenomyosis were applied as well as AUB, subfertility or anti-angiogenic therapy. The main search was limited to clinical studies carried out on premenopausal women. Original research articles focusing on markers of angiogenesis in the endometrium of patients with adenomyosis were included. Studies in which no comparison was made to control patients or which were not published in a peer-reviewed journal were excluded. A second search was performed to explore the therapeutic potential of targeting angiogenesis in adenomyosis. This search also included preclinical studies. OUTCOMES: A total of 20 articles out of 1669 hits met our selection criteria. The mean vascular density (MVD) was studied by quantification of CD31, CD34, von Willebrand Factor (vWF) or factor-VIII-antibody-stained microvessels in seven studies. All these studies reported a significantly increased MVD in ectopic endometrium, and out of the six articles that took it into account, four studies reported a significantly increased MVD in eutopic endometrium compared with control endometrium. Five articles showed a significantly higher vascular endothelial growth factor expression in ectopic endometrium and three articles in eutopic endometrium compared with control endometrium. The vascular and pro-angiogenic markers α-smooth muscle actin, endoglin, S100A13, vimentin, matrix metalloproteinases (MMPs), nuclear factor (NF)-kB, tissue factor (TF), DJ-1, phosphorylated mammalian target of rapamycin, activin A, folli- and myostatin, CD41, SLIT, roundabout 1 (ROBO1), cyclooxygenase-2, lysophosphatidic acid (LPA) 1,4-5, phospho signal transducer and activator of transcription 3 (pSTAT3), interleukin (IL)-6, IL-22 and transforming growth factor-ß1 were increased in ectopic endometrium, and the markers S100A13, MMP-2 and -9, TF, follistatin, myostatin, ROBO1, LPA1 and 4-5, pSTAT3, IL-6 and IL-22 were increased in eutopic endometrium, compared with control endometrium. The anti-angiogenic markers E-cadherin, eukaryotic translation initiation factor 3 subunit and gene associated with retinoic-interferon-induced mortality 19 were decreased in ectopic endometrium and IL-10 in eutopic endometrium, compared with control endometrium. The staining level of vWF and two pro-angiogenic markers (NF-κB nuclear p65 and TF) correlated with AUB in patients with adenomyosis. We found no studies that investigated the possible relationship between markers of angiogenesis and subfertility in adenomyosis patients. Nine articles reported on direct or indirect targeting of angiogenesis in adenomyosis-either by testing hormonal therapy or herbal compounds in clinical studies or by testing angiogenesis inhibitors in preclinical studies. However, there are no clinical studies on the effectiveness of such therapy for adenomyosis-related AUB or subfertility. WIDER IMPLICATIONS: The results are in agreement with our hypothesis that increased angiogenesis is present in the endometrium of patients with adenomyosis compared with the endometrium of control patients. It is likely that increased angiogenesis leads to fragile and more permeable vessels resulting in adenomyosis-related AUB and possibly subfertility. While this association has not sufficiently been studied yet, our results encourage future studies to investigate the exact role of angiogenesis in the etiology of adenomyosis and related AUB or subfertility in women with adenomyosis in order to design curative or preventive therapeutic strategies.

Morphometric analysis of the lung vasculature after extracorporeal membrane oxygenation treatment for pulmonary hypertension in newborns.

Persistent pulmonary hypertension in the newborn (PPHN) is characterised by increased medial and adventitial thickness in the lung vasculature. This study describes morphometry of lung vasculature after extracorporeal membrane oxygenation (ECMO) in newborns with PPHN, due to meconium aspiration syndrome, sepsis or idiopathic persistent pulmonary hypertension of the newborn (i-PPHN). Three groups were studied: newborns with PPHN treated with ECMO (n=9), newborns with PPHN not treated with ECMO (n=12) and age-matched controls without PPHN (n=11). In pulmonary arteries with an external diameter of less than 150 microm, arterial media, adventitia and total wall thickness, expressed as a percentage of the external diameter, and their cross-sectional areas were calculated. Newborns with PPHN, compared with controls, demonstrated increased percentage of media thickness, adventitia thickness and total wall thickness, and increased medial, adventitial and total wall cross-sectional area. Newborns treated with ECMO, compared with those not treated so, showed a decreased percentage of media thickness and medial cross-sectional area in arteries with an external diameter less than 75 microm, and decreased percentage of media thickness and decreased medial, adventitial and total wall cross-sectional area in arteries with an external diameter of 75-150 microm. ECMO for persistent PPHN, due to meconium aspiration syndrome, sepsis or i-PPHN, reduces the abnormal morphometry of small pulmonary arteries. The underlying mechanisms contributing to this improved morphometry are yet unknown.

Effect of chemical stabilizers of hypoxia-inducible factors on early lung development.

Low oxygen stimulates pulmonary vascular development and airway branching and involves hypoxia-inducible factor (HIF). HIF is stable and initiates expression of angiogenic factors under hypoxia, whereas normoxia triggers hydroxylation of the HIF-1alpha subunit by prolyl hydroxylases (PHDs) and subsequent degradation. Herein, we investigated whether chemical stabilization of HIF-1alpha under normoxic (20% O(2)) conditions would stimulate vascular growth and branching morphogenesis in early lung explants. Tie2-LacZ (endothelial LacZ marker) mice were used for visualization of the vasculature. Embryonic day 11.5 (E11.5) lung buds were dissected and cultured in 20% O(2) in the absence or presence of cobalt chloride (CoCl(2), a hypoxia mimetic), dimethyloxalylglycine (DMOG; a nonspecific inhibitor of PHDs), or desferrioxamine (DFO; an iron chelator). Vascularization was assessed by X-gal staining, and terminal buds were counted. The fine vascular network surrounding the developing lung buds seen in control explants disappeared in CoCl(2)- and DFO-treated explants. Also, epithelial branching was reduced in the explants treated with CoCl(2) and DFO. In contrast, DMOG inhibited branching but stimulated vascularization. Both DFO and DMOG increased nuclear HIF-1alpha protein levels, whereas CoCl(2) had no effect. Since HIF-1alpha induces VEGF expression, the effect of SU-5416, a potent VEGF receptor (VEGFR) blocker, on early lung development was also investigated. Inhibition of VEGFR2 signaling in explants maintained under hypoxic (2% O(2)) conditions completely abolished vascularization and slightly decreased epithelial branching. Taken together, the data suggest that DMOG stabilization of HIF-1alpha during early development leads to a hypervascular lung and that airway branching proceeds without the vasculature, albeit at a slower rate.

Angiogenic factors stimulate tubular branching morphogenesis of sonic hedgehog-deficient lungs.

Sonic Hedgehog (Shh)-deficient mice have a severe lung branching defect. Recent studies have shown that hedgehog signaling is involved in vascular development and it is possible that the diminished airway branching in Shh-deficient mice is due to abnormal pulmonary vasculature formation. Therefore, we investigated the role of Shh in pulmonary vascular development using Shh/Tie2lacZ compound mice, which exhibit endothelial cell-specific LacZ expression, and Pecam-1 immunohistochemistry. In E11.5-13.5 Shh-deficient mice, the pulmonary vascular bed is decreased, but appropriate to the decrease in airway branching. However, when E12.5 Shh-deficient lungs were cultured for 4-6 days, the vascular network deteriorated compared to wild-type lungs. The expression of vascular endothelial growth factor (Vegf) or its receptor Vegfr2 (KDR/Flk-1) was not different between E12.5-13.5 Shh-deficient and wild-type lungs. In contrast, angiopoietin-1 (Ang1), but not Ang2 or the angiopoietin receptor Tie2, mRNA expression was downregulated in E12.5-E13.5 lungs of Shh null mutants. Recombinant Ang1 alone was unable to restore in vitro branching morphogenesis in Shh-deficient lungs. Conversely, the angiogenic factor fibroblast growth factor (Fgf)-2 alone or in combination with Ang1, increased vascularization and tubular growth and branching of Shh-deficient lungs in vitro. The angiogenic factors did not overcome the reduced smooth muscle cell differentiation in the Shh null lungs. These data indicate that early vascular development, mediated by Vegf/Vegfr2 signaling proceeds normally in Shh-deficient mice, while later vascular development and stabilization of the primitive network mediated by the Ang/Tie2 signaling pathway are defective, resulting in an abnormal vascular network. Stimulation of vascularization with angiogenic factors such as Fgf2 and Ang1 partially restored tubular growth and branching in Shh-deficient lungs, suggesting that vascularization is required for branching morphogenesis.

Iroquois genes influence proximo-distal morphogenesis during rat lung development.

Lung development is a highly regulated process directed by mesenchymal-epithelial interactions, which coordinate the temporal and spatial expression of multiple regulatory factors required for proper lung formation. The Iroquois homeobox (Irx) genes have been implicated in the patterning and specification of several Drosophila and vertebrate organs, including the heart. Herein, we investigated whether the Irx genes play a role in lung morphogenesis. We found that Irx1-3 and Irx5 expression was confined to the branching lung epithelium, whereas Irx4 was not expressed in the developing lung. Antisense knockdown of all pulmonary Irx genes together dramatically decreased distal branching morphogenesis and increased distention of the proximal tubules in vitro, which was accompanied by a reduction in surfactant protein C-positive epithelial cells and an increase in beta-tubulin IV and Clara cell secretory protein positive epithelial structures. Transmission electron microscopy confirmed the proximal phenotype of the epithelial structures. Furthermore, antisense Irx knockdown resulted in loss of lung mesenchyme and abnormal smooth muscle cell formation. Expression of fibroblast growth factors (FGF) 1, 7, and 10, FGF receptor 2, bone morphogenetic protein 4, and Sonic hedgehog (Shh) were not altered in lung explants treated with antisense Irx oligonucleotides. All four Irx genes were expressed in Shh- and Gli(2)-deficient murine lungs. Collectively, these results suggest that Irx genes are involved in the regulation of proximo-distal morphogenesis of the developing lung but are likely not linked to the FGF, BMP, or Shh signaling pathways.

Alveolar capillary dysplasia: a logical approach to a fatal disease.

PURPOSE: The aim of the study was to review the authors' experience with alveolar capillary dysplasia (ACD), a cause of persistent pulmonary hypertension of the neonate (PPHN) caused by decreased alveolar units, dilated anomalous pulmonary veins, thick-walled arterioles, and thickened interalveolar septa. METHODS: The records of all neonates with ACD were reviewed from Children's Hospital, Columbus, Ohio, and Sophia's Children's Hospital, Rotterdam, The Netherlands. The clinical characteristics and pathological findings are discussed. RESULTS: Eight neonates were diagnosed with ACD from 1994 to 2002. Twenty-five percent (2/8) experienced respiratory distress immediately after birth, whereas 75% (6/8) had normal Apgar scores but deteriorated 1.5 hours to 30 days after birth. All infants required conventional ventilation initially; 50% (4/8) were placed on high-frequency oscillating ventilation and 87% (7/8) on extracorporeal membrane oxygenation. A premorbid diagnosis was established in 3 patients by open lung biopsy. The diagnosis of ACD was confirmed at autopsy in all patients. CONCLUSIONS: ACD is a fatal disease that should be suspected in all neonates with respiratory failure and PPHN who fail conventional therapy. Prompt diagnosis helps to avoid prolongation of costly treatment modalities in a uniformly fatal disease. An algorithm is proposed in which neonates with PPHN who fail treatment with extracorporeal membrane oxygenation are managed by open lung biopsy.