DNA Damage-Induced Nucleosome Depletion Enhances Homology Search Independently of Local Break Movement.

To determine whether double-strand break (DSB) mobility enhances the physical search for an ectopic template during homology-directed repair (HDR), we tested the effects of factors that control chromatin dynamics, including cohesin loading and kinetochore anchoring. The former but not the latter is altered in response to DSBs. Loss of the nonhistone high-mobility group protein Nhp6 reduces histone occupancy and increases chromatin movement, decompaction, and ectopic HDR. The loss of nucleosome remodeler INO80-C did the opposite. To see whether enhanced HDR depends on DSB mobility or the global chromatin response, we tested the ubiquitin ligase mutant uls1Δ, which selectively impairs local but not global movement in response to a DSB. Strand invasion occurs in uls1Δ cells with wild-type kinetics, arguing that global histone depletion rather than DSB movement is rate limiting for HDR. Impaired break movement in uls1Δ correlates with elevated MRX and cohesin loading, despite normal resection and checkpoint activation.

POT-3 preferentially binds the terminal DNA-repeat on the telomeric G-overhang.

Eukaryotic chromosomes typically end in 3' telomeric overhangs. The safeguarding of telomeric single-stranded DNA overhangs is carried out by factors related to the protection of telomeres 1 (POT1) protein in humans. Of the three POT1-like proteins in Caenorhabditis elegans, POT-3 was the only member thought to not play a role at telomeres. Here, we provide evidence that POT-3 is a bona fide telomere-binding protein. Using a new loss-of-function mutant, we show that the absence of POT-3 causes telomere lengthening and increased levels of telomeric C-circles. We find that POT-3 directly binds the telomeric G-strand in vitro and map its minimal DNA binding site to the six-nucleotide motif, GCTTAG. We further show that the closely related POT-2 protein binds the same motif, but that POT-3 shows higher sequence selectivity. Crucially, in contrast to POT-2, POT-3 prefers binding sites immediately adjacent to the 3' end of DNA. These differences are significant as genetic analyses reveal that pot-2 and pot-3 do not function redundantly with each other in vivo. Our work highlights the rapid evolution and specialisation of telomere binding proteins and places POT-3 in a unique position to influence activities that control telomere length.

Surgical Outcomes and Complications of Laparoscopic Hysterectomy for Endometriosis: A Multicentric Cohort Study.

STUDY OBJECTIVE: To investigate the postoperative morbidity of laparoscopic hysterectomy (LH) for endometriosis/adenomyosis in terms of operative outcomes and complications. DESIGN: Retrospective multicentric cohort study. SETTING: Eight European minimally invasive referral centers. PATIENTS: Data from 995 patients with pathologically confirmed endometriosis and/or adenomyosis who underwent LH without concomitant urological and/or gastroenterological procedures from January 2010 to December 2020. INTERVENTIONS: Total LH. MEASUREMENTS AND MAIN RESULTS: Demographic patients' characteristics, surgical outcomes, and intraoperative and postoperative complications were evaluated. We considered major postoperative surgical-related complications, any grade 2 or more events (Clavien-Dindo score) that occurred within 30 days from surgery. Univariate analysis and multivariable models fit with logistic regression were used to estimate the adjusted odds ratio (OR) and corresponding 95% confidence interval (CI) for major complications. Median age at surgery was 44 years (28-54), and about half of them (505, 50.7%) were on medical treatment (estro-progestins, progestin, or Gonadotropin hormone-releasing hormone-analogues) at the time of surgery. In association with LH, posterior adhesiolysis was performed in 387 (38.9%) cases and deep nodule resection in 302 (30.0%). Intraoperative complications occurred in 3% of the patients, and major postoperative complications were registered in 93 (9.3%). The multivariable analysis showed an inverse correlation between the occurrence of Clavien-Dindo >2 complications and age (OR 0.94, 95% CI 0.90-0.99), while previous surgery for endometriosis (OR 1.62, 95% CI 1.01-2.60) and intraoperative complications (OR 6.49, 95% CI 2.65-16.87) were found as predictors of major events. Medical treatment at the time of surgery has emerged as a protective factor (OR 0.50, 95% CI 0.31-0.81). CONCLUSION: LH for endometriosis/adenomyosis is associated with non-negligible morbidity. Knowing the factors associated with higher risks of complications might be used for risk stratification and could help clinicians during preoperative counseling. The administration of estro-progestin or progesterone preoperatively might reduce the risks of postoperative complications following surgery.

Characterization of technical skill progress in a standardized rabbit model for training in laparoscopic duodenal atresia repair.

BACKGROUND: Laboratory skills training is an essential step before conducting minimally invasive surgery in clinical practice. Our main aim was to develop an animal model for training in clinically highly challenging laparoscopic duodenal atresia repair that could be useful in establishing a minimum number of repetitions to indicate safe performance of similar interventions on humans. MATERIALS AND METHODS: A rabbit model of laparoscopic duodenum atresia surgery involving a diamond-shaped duodeno-duodenostomy was designed. This approach was tested in two groups of surgeons: in a beginner group without any previous clinical laparoscopic experience (but having undergone previous standardized dry-lab training, n = 8) and in an advanced group comprising pediatric surgery fellows with previous clinical experience of laparoscopy (n = 7). Each participant performed eight interventions. Surgical time, expert assessment using the Global Operative Assessment of Laparoscopic Skills (GOALS) score, anastomosis quality (leakage) and results from participant feedback questionnaires were analyzed. RESULTS: Participants in both groups successfully completed all eight surgeries. The surgical time gradually improved in both groups, but it was typically shorter in the advanced group than in the beginner group. The leakage rate was significantly lower in the advanced group in the first two interventions, and it reached its optimal level after five operations in both groups. The GOALS and participant feedback scores showed gradual increases, evident even after the fifth surgery. CONCLUSIONS: Our data confirm the feasibility of this advanced pediatric laparoscopic model. Surgical time, anastomosis quality, GOALS score and self-assessment parameters adequately quantify technical improvement among the participants. Anastomosis quality reaches its optimal value after the fifth operation even in novice, but uniformly trained surgeons. A minimum number of wet-lab operations can be determined before surgery can be safely conducted in a clinical setting, where the development of further non-technical skills is also required.

Laparoscopic Treatment of Benign Adnexal Masses During Pregnancy: A Comprehensive Evaluation of Treatment Basics, Efficacy and Surgical Techniques.

This review summarizes the evidence-based recommendations for how to approach and laparoscopically treat adnexal masses during pregnancy. We conducted a comprehensive review of studies related to the laparoscopic management of adnexal masses during pregnancy. Selected studies were independently reviewed by two authors. The overall incidence of ovarian tumors in pregnancy ranges between 0.05% and 5.7%, of which less than 5% are malignant. Diagnosis is based mainly on routine transvaginal ultrasound. More than 64% of simple cysts, less than 6 cm in diameter, will spontaneously resolve in less than 16 weeks. However, for persistent and complex tumors, the risk of acute complications can reach up to 9%. Surgical indications are similar to those in the non-gravidic setting, and include acute complications (torsion, rupture, hemorrhage), suspected malignancy and large (over 6 cm) persistent masses. Surgery must be scheduled between 16 and 20 weeks to allow for the spontaneous resolution of functional cysts. Furthermore, within that period, pregnancy becomes independent of the corpus luteum and enlargement of the uterus gives sufficient exposure for the surgery to be performed safely. A recent meta-analysis found that, compared to open surgery, laparoscopy is associated with significantly less preterm labor, blood loss and hospital stay, without differences in pregnancy loss or preterm birth rate. Since the main concerns about maternal-fetal safety are related to increased intraperitoneal pressure and the effects of hypercarbia (maternal hypertensive complications, fetal acidosis), a lower CO2 pressure (10 to 12 mmHg) and reduced operative times (less than 30 minutes) are recommended.

The ATP-dependent chromatin remodelling enzyme Uls1 prevents Topoisomerase II poisoning.

Topoisomerase II (Top2) is an essential enzyme that decatenates DNA via a transient Top2-DNA covalent intermediate. This intermediate can be stabilized by a class of drugs termed Top2 poisons, resulting in massive DNA damage. Thus, Top2 activity is a double-edged sword that needs to be carefully controlled to maintain genome stability. We show that Uls1, an adenosine triphosphate (ATP)-dependent chromatin remodelling (Snf2) enzyme, can alter Top2 chromatin binding and prevent Top2 poisoning in yeast. Deletion mutants of ULS1 are hypersensitive to the Top2 poison acriflavine (ACF), activating the DNA damage checkpoint. We map Uls1's Top2 interaction domain and show that this, together with its ATPase activity, is essential for Uls1 function. By performing ChIP-seq, we show that ACF leads to a general increase in Top2 binding across the genome. We map Uls1 binding sites and identify tRNA genes as key regions where Uls1 associates after ACF treatment. Importantly, the presence of Uls1 at these sites prevents ACF-dependent Top2 accumulation. Our data reveal the effect of Top2 poisons on the global Top2 binding landscape and highlights the role of Uls1 in antagonizing Top2 function. Remodelling Top2 binding is thus an important new means by which Snf2 enzymes promote genome stability.

Neovagina creation in congenital vaginal agenesis: New mini-laparoscopic approach applying intraoperative indocyanine green fluorescence.

Background. There is still no standardized treatment for vaginal agenesis; surgical repair using a minimally invasive technique is a good option, as it offers rapid results along with a fast postoperative recovery. Objective. To describe a new modified Vecchietti procedure by a mini-laparoscopic approach with intraoperative use of indocyanine green fluorescence technology for the creation of a neovagina in the setting of congenital vaginal agenesis. Methods. Retrospective review of all cases of congenital vaginal agenesis submitted to a mini-laparoscopic modified Vecchietti procedure with intraoperative use of indocyanine green (ICG) fluorescence (between June and September 2019). Patient relevant medical history, surgical technique, postoperative care, outcomes, and complications are described. Results. Four patients with Mayer-Rokitansky-Küster-Hauser syndrome (mean age: 19 years; mean preoperative vaginal length: 1.5 cm) were included in the study. The procedure was completed successfully and uneventfully in all patients. External traction device was removed 5-7 days after surgery and replaced by external vaginal silicone dilators. This technique was able to create 10-11 cm neovaginas in 1 week; six weeks after surgery all patients had an epithelized 10.5-12 cm length and 2.5-3 cm diameter neovagina. Complications consisted of one case of isolated fever at the ninth postoperative day, solved with empiric antibiotic treatment. Conclusion. The mini-laparoscopic modified Vecchietti approach, associated with ICG fluorescence imaging, is feasible and effective in the treatment of congenital vaginal agenesis, as it is a simple and safe procedure with promising outcomes.

Diaphragmatic endometriosis minimally invasive treatment: a feasible and effective approach.

The present review aims to analyse the current data available on the feasibility, safety and effectiveness of the minimally invasive surgical (MIS) treatment of diaphragmatic endometriosis (DE). Through the use of PubMed and Google Scholar database, we conducted a literature review of all available research related to diagnosis and treatment of DE, focussed on the minimally invasive techniques. The studies were selected independently by two authors according to the aim of this review. DE is an under-diagnosed disease affecting between 0.1% and 1.5% of fertile women. It is predominantly multiple, asymptomatic and highly associated with pelvic disease in about 50-90%. MIS techniques seems to be safe, effective and feasible in tertiary advanced endometriosis centre, offering definitive advantages in terms of hospital stay, post-operative pain and return to normal activity by using several surgical techniques as hydro-dissection plus resection, laser CO2 vaporisation, electrical fulguration, Sugarbaker peritonectomy, partial (shaving) and full-thickness diaphragmatic resection. Symptoms control range from 85% to 100%, with less than 3% of conversion, peri-operative complications and recurrence rate. All cases must be performed by multidisciplinary teams including at least a gynaecologist, thoracic surgeon and anaesthetist. The lack of prospective evaluation of DE interferes with the understanding about the natural history of disease and treatment results. Therefore, the development of adequate evidence-based recommendations about diagnosis, management and follow-up is difficult at this moment.

Ghost Ileostomy in Anterior Resection for Bowel Endometriosis: Technical Description.

OBJECTIVE: To demonstrate our application of the ghost ileostomy in the setting of laparoscopic segmental bowel resection for symptomatic bowel endometriosis nodule. DESIGN: Technical step-by-step surgical video description (educative video) SETTING: University Tertiary Hospital. Institutional Review Board ruled that approval was not required for this study. Endometriosis affects the bowel in 3% to 37% of all cases, and in 90% of these cases, the rectum or sigmoid colon is also involved. Infiltration up to the rectal mucosa and invasion of >50% of the circumference have been suggested as an indication for bowel resection [1]. Apart from general risks (bleeding, infection, direct organ injuries) and bowel and bladder dysfunctions, anastomotic leakage is one of the most severe complications. In women with bowel and vaginal mucosa endometriosis involvement, there is a risk of rectovaginal fistula after concomitant rectum and vagina resections. Hence, for lower colorectal anastomosis, the use of temporary protective ileostomy is usually recommended to prevent these complications but carries on stoma-related risks, such as hernia, retraction, dehydration, prolapse, and necrosis. Ghost ileostomy is a specific technique, first described in 2010, that gives an easy and safe option to prevent anastomotic leakage with maximum preservation of the patient's quality of life [2]. In case of anastomotic leakage, the ghost (or virtual) ileostomy is converted, under local anesthesia, into a loop (real) ileostomy by extracting the isolated loop through an adequate abdominal wall opening. In principle, avoiding readmission for performing the closure of the ileostomy, with all the costs related, means a considerable saving for the hospital management. Also, applying a protective rectal tube in intestinal anastomosis may have a beneficial effect [3]. These options are performed by general surgeons in oncological scenarios, but their use in endometriosis has never been described. INTERVENTIONS: In a 32-year-old woman with intense dysmenorrhea, deep dyspareunia, dyschesia, and cyclic rectal bleeding, a complete laparoscopic approach was performed using blunt and sharp dissection with cold scissors, bipolar dissector and a 5-mm LigaSure Advance (Covidien, Valley lab, Norwalk, Connecticut). An extensive adhesiolysis restoring the pelvic anatomy and endometriosis excision was done. Afterward, the segmental bowel resection was performed using linear and circular endo-anal stapler technique with immediate end-to-end bowel anastomosis and transit reconstitution. Once anastomosis was done, the terminal ileal loop was identified, and a window was made in the adjacent mesentery. Then, an elastic tape (vessel loop) was passed around the ileal loop, brought out of the abdomen through the right iliac fossa 5-mm port site incision and, fixed to the abdominal wall using nonabsorbable stitches. Finally, a trans-anal tube was placed for 5 days. The patient was discharged on the fifth day postoperatively without any complications. The tape was removed 10 days after surgery, and the loop dropped back. Two months after the intervention, the patient remains asymptomatic. CONCLUSION: Ghost ileostomy is a simple, safe, and feasible technique available in the setting of lower colorectal anastomosis following bowel endometriosis resection.

Application of Indocyanine Green in Gynecology: Review of the Literature.

The present review aims to analyze the current data available on the different applications of indocyanine green (ICG) in gynecology. A semantic review of English-language publications was performed by searching for MeSH terms and keywords in the PubMed and Google Scholar databases. The studies were finally selected by one author according to the aim of this review. ICG is a highly water-soluble tricarbocyanine dye that fluoresces in the NIR spectrum. Approved by the FDA in 1959, it can be administered either IV (usual dose of 5 mg) or locally/submucosally (usual dose of 5-6.25 mg) according to the pathology or indication. It is used most often in the setting of oncology, endometriosis and other gynecological conditions. In oncological applications, ICG is used to identify sentinel lymph nodes (SLN) using near-infrared light in endometrial, cervical and vulvar cancers. The main advantages that it offers include a reduction of surgical time, improved SLN detection rates, and the ability to avoid radioactivity. In cases of endometrial (submucosal or hysteroscopic applications) or cervical (intracervical administration) cancer, ICG can detect SLN at an accuracy of 95% to 98%. For vulvar cancer, the SLN detection rate can reach 100%. In endometriosis, the lack of good evidence hinders the final evaluation of this method in both diagnostic and therapeutic scenarios. An analytical, well-designed, prospective study is currently underway.

Vesico-Vaginal Fistula: Nature and Evidence-Based Minimally Invasive Surgical Treatment.

The present review aims to analyze the current information available on the pathophysiology, clinical presentation and treatment of vesico-vaginal fistulas (VVF), with particular focus on the safety and efficacy of minimally invasive surgical (MIS) techniques. Through the use of the PubMed and Google Scholar databases, we conducted a literature review of all available studies related to MIS treatment of VVF, focusing on laparoscopic techniques. After abstracts were read to identify pertinent studies, full manuscripts were reviewed by two authors according to the aim of the review. Vesico-vaginal fistula is defined as an abnormal passage that connects the bladder to the vagina and affects over 3 million women worldwide. It can be classified according to its complexity (simple or complex) and mechanism (obstetric-related or iatrogenic). Laparoscopic treatment of VVF started in 1994 and is currently the gold-standard approach for this pathology. No differences in terms of efficacy or safety have been reported between MIS (laparoscopy, robotic-assisted laparoscopy and laparoscopic single-site) using extra-vesical and trans-vesical approaches, with success rates of 80% to 100%, and low rates of conversion (1.9%), recurrence (less than 1%) and intra- or post-operative complications (3%). Surgical principles for fistula repair, described independently by Angioli and Couvelaire, must always be followed. A bladder fill and integrity test with at least 300 mL should be performed before ending surgery, since this increases the success rate by about 6%. Other interventions such as flap interposition, number of layers in closure and expectant management (spontaneous closure with a Foley catheter alone) remain controversial. To date, no differences have been seen among the laparoscopic surgical techniques. The lack of prospective evaluations has hindered a better understanding of the natural history of the disease and the development of evidence-based recommendations regarding diagnosis, management and follow-up. Since no differences were found compared to a trans-vesical approach, extra-vesical repair is recommended to avoid bladder bi-valving.

Endometriosis of the Bladder: Clinical and Surgical Outcomes after Laparoscopic Surgery.

BACKGROUND/AIMS: Urinary tract endometriosis is rare. The bladder is the most common site affected. The nonspecific symptoms can make a diagnosis difficult. The aim of this study was to evaluate the clinical and surgical outcomes in women who underwent surgical treatment for bladder endometriosis (BE). METHODS: Ten patients who underwent surgical treatment for BE from January 2012 to November 2016 were retrospectively reviewed. Pre- and postoperative data, intraoperative findings, type of surgical procedure, and intra- and postoperative complications were analyzed. RESULTS: Two women were treated by laparoscopic shaving of the bladder lesion and 8 underwent laparoscopic partial cystectomy. Simultaneous resection of coexisting pelvic nodules was performed. No conversions to laparotomy were observed. There was only one intraoperative complication. No major or minor postoperative complications were observed and none of the patients required repeated interventions. Improvements in clinical symptoms were reported and there was no increase in long-term urinary frequency after surgery. There was 1 case of urinary symptom recurrence. CONCLUSION: Laparoscopic partial cystectomy and shaving of the bladder lesion seem to improve urinary symptoms, with a low rate of intra- and postoperative complications and a low rate of recurrence, without affecting long-term bladder capacity. This surgical approach requires an experienced gynecologist and urologist team.

Rectovaginal Splenosis: An Unexpected Cause of Dyspareunia Approached by Laparoscopy.

STUDY OBJECTIVE: To demonstrate the technique of laparoscopic approach in a rare case of rectovaginal splenosis with severe dyspareunia and dyschesia. DESIGN: A step-by-step explanation of the patient's condition, diagnosis, surgical technique, and postoperative results (Canadian Task Force classification II-3). SETTING: Splenosis consists of ectopic functioning splenic tissue that can be located anywhere within the abdomen or pelvis. Fragments are often multiple and range in diameter from a few millimeters to a few centimeters. They are reddish-blue and are sessile or pedunculated. Their appearance can mimic that of neoplasms or endometriosis, which are the main differential diagnoses. Trauma and subsequent splenectomy is the cause in most cases. Splenosis is a benign condition usually found incidentally and is usually asymptomatic. The need for therapy is controversial, and treatment is suggested only in symptomatic cases, primarily those related to pelvic or abdominal lesions, as in our patient. The diagnosis of splenosis in a woman complaining of pelvic pain may present diagnostic difficulties. The splenic tissue has the macroscopic appearance of endometriosis, and its position in the pelvis also may suggest this diagnosis. Where excision of splenosis is considered necessary, the approach should be laparoscopic, unless this is considered too risky owing to the proximity of vital structures. INTERVENTION: A 40-year-old woman was referred to our department for severe dyspareunia and dyschezia. The gynecologic examination revealed a painfull nodularity on the posterior vaginal cul de sac. Further evaluation with 2- and 3-dimensional ultrasound and magnetic resonance imaging revealed several soft tissue nodules in the pouch of Douglas (POD), which were enhanced on contrast administration. She had undergone a splenectomy 15 years earlier after a car accident. A laparoscopic approach to a rectovaginal nodularity was performed. Under general anesthesia, the patient was placed in the dorsal decubitus position with her arms alongside her body and her legs in abduction. Pneumoperitoneum was achieved using a Veres needle placed at the umbilicus. Four trocars were placed: a 10-mm trocar at the umbilicus for the 0-degree laparoscope; a 5-mm trocar at the right anterosuperior iliac spine; a 5-mm trocar in the midline between the umbilicus and the pubic symphysis, approximately 8 to 10 cm inferior to the umbilical trocar; and a 5-mm trocar at the left anterosuperior iliac spine. The entire pelvis was inspected for endometriotic lesions. In the pelvis, hypervascular and bluish nodules were visible with extension from the POD into the deep rectovaginal space. The macroscopic appearance was atypical for endometriotic implants. The nodularities were carefully dissected and excised, and histological assessment revealed splenic tissue. At the time of this report, the patient had been asymptomatic for 6 months after surgery. CONCLUSION: Rectovaginal splenosis may mimic endometriosis. The laparoscopic approach to rectovaginal splenosis avoids an abdominal incision, with its associated pain and possible adhesion formation. It also provides a better view for dissection. In this patient, the splenosis was removed by laparoscopy, with no postoperative dyspareunia or dyschesia.

Yeast telomerase and the SUN domain protein Mps3 anchor telomeres and repress subtelomeric recombination.

Telomeres form the ends of linear chromosomes and protect these ends from being recognized as DNA double-strand breaks. Telomeric sequences are maintained in most cells by telomerase, a reverse transcriptase that adds TG-rich repeats to chromosome ends. In budding yeast, telomeres are organized in clusters at the nuclear periphery by interactions that depend on components of silent chromatin and the telomerase-binding factor yeast Ku (yKu). In this study, we examined whether the subnuclear localization of telomeres affects end maintenance. A telomere anchoring pathway involving the catalytic yeast telomerase subunits Est2, Est1, and Tlc1 is shown to be necessary for the perinuclear anchoring activity of Yku80 during S phase. Additionally, we identify the conserved Sad1-UNC-84 (SUN) domain protein Mps3 as the principal membrane anchor for this pathway. Impaired interference with Mps3 anchoring through overexpression of the Mps3 N terminus in a tel1 deletion background led to a senescence phenotype and to deleterious levels of subtelomeric Y' recombination. This suggests that telomere binding to the nuclear envelope helps protect telomeric repeats from recombination. Our results provide an example of a specialized structure that requires proper spatiotemporal localization to fulfill its biological role, and identifies a novel pathway of telomere protection.

Adnexal Torsion in the First Trimester of Pregnancy: Diagnosis, Laparoscopic Management, and Review of the Literature.

We report a case of an adnexal torsion in a 27-year-old woman in her 12th week of gestation. She presented with hypogastric and lumbar pain with biliary vomiting and nausea. Upon physical examination, tenderness in the right lower quadrant with rebound tenderness was apparent, and gynecological examination revealed right adnexal tenderness with absence of abnormal cervical discharge. The transvaginal ultrasonography demonstrated an enlarged cystic right ovary with no flow detected on color and power Doppler mode. We performed a laparoscopy and the diagnosis of adnexal torsion was confirmed. Detorsion of the right adnexa and cystectomy of the ovary was achieved without surgical or anesthetic complications. The patient was supplemented with progesterone during pregnancy and gave birth to a healthy child at term. Adnexal torsion is an unusual cause of abdominal pain in gestation with an incidence of 1-5:10.000, and it is more often observed during the first and early second trimesters of gestation. The clinical signs are often unspecific, and an ultrasound is the most commonly used imaging method for this diagnosis. The laparoscopy is the preferred method of diagnosis and treatment can be safely used in pregnancy if the guidelines are respected.

Resection of Sentinel Lymph Nodes by an Extraperitoneal Minilaparoscopic Approach Using Indocyanine Green for Uterine Malignancies: A Preclinical Comparative Study.

Background The sentinel lymph node (SLN) concept might minimize surgical aggressiveness in cervical and endometrial malignancies. The aim of the study was to test the feasibility and reliability of minilaparoscopic extraperitoneal SLN excision after indocyanine green (ICG) cervical injection using a high-definition near infrared (NIR) imaging system in an in vivo porcine model. The same procedure was performed using conventional laparoscopic instruments and both outcomes were compared. Methods Twenty-four animals were equally and randomly divided into a minilaparoscopic group (group A) and a 5-mm conventional laparoscopic group (group B). A high-definition NIR imaging system and a 30° ICG endoscope were used. First, ICG (0.5 mL) was injected in the paracervical region. The SLN coloring time was recorded. An extraperitoneal approach to the SLN was executed with the same CO2 retropneumoperitoneum pressures (10 mm Hg). In both groups, the times for SLN localization and excision, as well as complications, were registered. Finally, a laparotomy was then done to evaluate whether any stained SLN still remained. The same surgical team performed all experiments. Results SLNs were identified and extraperitoneally excised in all animals without major complications. The SLN localization varied between animals from external iliac to preaortic regions. The surgical times were shorter with minilaparoscopy (39.3 ± 13 minutes) than with conventional 5-mm instruments (51.3 ± 14.17 minutes; P = .042). In group B, one stained SLN remained and was only detected by laparotomy. Conclusions We confirmed the feasibility and reliability of extraperitoneal minilaparoscopic approach for identification, dissection, and excision of SLN using an NIR imaging system and ICG.

Regulating repression: roles for the sir4 N-terminus in linker DNA protection and stabilization of epigenetic states.

Silent information regulator proteins Sir2, Sir3, and Sir4 form a heterotrimeric complex that represses transcription at subtelomeric regions and homothallic mating type (HM) loci in budding yeast. We have performed a detailed biochemical and genetic analysis of the largest Sir protein, Sir4. The N-terminal half of Sir4 is dispensable for SIR-mediated repression of HM loci in vivo, except in strains that lack Yku70 or have weak silencer elements. For HM silencing in these cells, the C-terminal domain (Sir4C, residues 747-1,358) must be complemented with an N-terminal domain (Sir4N; residues 1-270), expressed either independently or as a fusion with Sir4C. Nonetheless, recombinant Sir4C can form a complex with Sir2 and Sir3 in vitro, is catalytically active, and has sedimentation properties similar to a full-length Sir4-containing SIR complex. Sir4C-containing SIR complexes bind nucleosomal arrays and protect linker DNA from nucleolytic digestion, but less effectively than wild-type SIR complexes. Consistently, full-length Sir4 is required for the complete repression of subtelomeric genes. Supporting the notion that the Sir4 N-terminus is a regulatory domain, we find it extensively phosphorylated on cyclin-dependent kinase consensus sites, some being hyperphosphorylated during mitosis. Mutation of two major phosphoacceptor sites (S63 and S84) derepresses natural subtelomeric genes when combined with a serendipitous mutation (P2A), which alone can enhance the stability of either the repressed or active state. The triple mutation confers resistance to rapamycin-induced stress and a loss of subtelomeric repression. We conclude that the Sir4 N-terminus plays two roles in SIR-mediated silencing: it contributes to epigenetic repression by stabilizing the SIR-mediated protection of linker DNA; and, as a target of phosphorylation, it can destabilize silencing in a regulated manner.

Screening for Diabetic Retinopathy in the Central Region of Portugal. Added Value of Automated 'Disease/No Disease' Grading.

Purpose: To describe the procedures of a nonmydriatic diabetic retinopathy (DR) screening program in the Central Region of Portugal and the added value of the introduction of an automated disease/no disease analysis. Methods: The images from the DR screening program are analyzed in a central reading center using first an automated disease/no disease analysis followed by human grading of the disease cases. The grading scale used is as follows: R0 - no retinopathy, RL - nonproliferative DR, M - maculopathy, RP - proliferative DR and NC - not classifiable. Results: Since the introduction of automated analysis in July 2011, a total of 89,626 eyes (45,148 patients) were screened with the following distribution: R0 - 71.5%, RL - 22.7%, M - 2.2%, RP - 0.1% and NC - 3.5%. The implemented automated system showed the potential for human grading burden reduction of 48.42%. Conclusions: Screening for DR using automated analysis allied to a simplified grading scale identifies DR vision-threatening complications well while decreasing human burden. © 2014 S. Karger AG, Basel.

Minilaparoscopic sacrocolpopexy for vaginal prolapse after hysterectomy.

Genital prolapse repair is one of the most common indications for benign gynecologic surgery. The lifetime risk of undergoing a single operation for prolapse in the female population is rising. Many different surgical techniques have been described. We report 4 cases of minilaparoscopic sacrocolpopexy to correct vaginal apical prolapse after previous total hysterectomy. For each patient we collected some socio-demographic data, vaginal apical prolapse grade using the Pelvic Organ Prolapse Quantification (POP-Q), intraoperative details and postoperative outcomes. Operative time was recorded as well as difficulties and complications (Clavien-Dindo Classification) at each step of the procedure. The range of women's ages was from 57 to 71 years old. The mean BMI was 24.75 ± 3.2 Kg/m2. Three patients had a stage III POP-Q prolapses and there was one case of a symptomatic stage II POP-Q prolapse. The mean surgical time was 119 minutes and there were no intraoperative complications. The postoperative pain assessment revealed very positive recovery in every patient. An ambulatory consult and an anatomic assessment were done 1 and 3 months after surgery. The incision scars were almost invisible after 1 month, and the anatomic cure rate was 100%. We confirmed the feasibility of a minilaparoscopic surgical approach for vaginal vault prolapse after total hysterectomy.

MRI of endometriosis in correlation with the #Enzian classification: applicability and structured report.

Endometriosis represents one of the most common causes of life-impacting chronic pelvic pain and female infertility. Magnetic resonance imaging (MRI) plays an increasing role in the diagnosis and mapping of endometriosis, while diagnostic laparoscopy currently tends to be reserved for the patients with negative imaging results. The #Enzian, published in 2021, proposes a new comprehensive classification system of endometriosis, combining a complete staging of deep infiltrative endometriosis with the evaluation of peritoneal/ovarian/tubal localizations and the presence of adenomyosis. This article addresses in detail the applicability of the #Enzian classification, primarily based on surgical findings, to the MRI evaluation of the endometriosis. Overall, there is a significant matching between MRI features and the #Enzian classification criteria, two different perspectives of endometriosis mapping, with different goals and levels of detail. The main discrepancy lies in the evaluation of tubo-ovarian condition, which is not fully assessable by MRI. Furthermore, as endometriosis is a complex disease, usually multifocal, that can present with a myriad of imaging findings, MRI reporting should be clear and well organized. The authors group, both radiologists and gynecologists, propose a structured MRI report of endometriosis in correlation with the #Enzian classification, merging the detailed anatomical and pre-operative information provided by the MRI with the benefits of a comprehensive classification system of endometriosis in the clinical practice and research field.Critical relevance statement This article addresses in detail the applicability of the #Enzian classification, primarily based on surgical findings, to the MRI evaluation of the endometriosis and proposes a #Enzian-based structured MRI report.