Neutrophil extracellular traps and DNases orchestrate formation of peritoneal adhesions.

Peritoneal adhesions are poorly understood but highly prevalent conditions that can cause intestinal obstruction and pelvic pain requiring surgery. While there is consensus that stress-induced inflammation triggers peritoneal adhesions, the molecular processes of their formation still remain elusive. We performed murine models and analyzed human samples to monitor the formation of adhesions and the treatment with DNases. Various molecular analyses were used to evaluate the adhesions. The experimental peritoneal adhesions of the murine models and biopsy material from humans are largely based on neutrophil extracellular traps (NETs). Treatment with DNASE1 (Dornase alfa) and the human DNASE1L3 analog (NTR-10), significantly reduced peritoneal adhesions in experimental models. We conclude that NETs serve as essential scaffold for the formation of adhesions; DNases interfere with this process. Herein, we show that therapeutic application of DNases can be employed to prevent the formation of murine peritoneal adhesions. If this can be translated into the human situation requires clinical studies.

Therapeutic Targeting of Neutrophil Extracellular Traps Improves Primary and Secondary Intention Wound Healing in Mice.

Background: Neutrophils are the first responders in wound healing after injury that mediate pro- and anti-inflammatory activities i.a. through the formation of extracellular traps (NETs). However, excessive NETs presence in wound tissue can cause local hyperinflammation and -coagulation resulting in delayed wound healing. To improve wound healing, we aimed to examine the role of NETs and DNase1 on primary and secondary wound healing. Methods: The study included 93 C57BL/6 mice, with 3 different genotypes: wildtype, Pad4-, and DNase1-Knockout (KO). Pad4-KO mice show limited NETs formation, while DNase1-KO mice cannot disintegrate them. All 3 genotypes were included in (1) a laparotomy group and (2) a thermal injury group. Animals in both groups either received DNase1 or a vehicle i.p. post wound induction and wound assessment and euthanasia were conducted. Laparotomy and burn scars were assessed using the stony brook scar evaluation scale and modified Yeong scale respectively. Tissue was analyzed histologically using H&E staining. Ly6g, Collagen I and III, SMA, and Fibrinogen were visualized and neutrophils activation (NE, MPO) and NETs (H3cit) formation assessed. Results: All animals survived with no complications. DNase1 treatment led to a significantly improved scar appearance in both groups, which was also seen in Pad4-KO mice. In the laparotomy group DNase1 improved collagen deposition and fibrin concentration was significantly reduced by DNase1 treatment. Markers of neutrophil activation were significantly reduced in the treatment and Pad4-KO group. In the thermal injury group wound closure time was significantly reduced after DNase1 treatment and in the Pad4-KO group. Even though inflammation remained high in the thermal injury model over time, neutrophil activation and NETs formation were significantly reduced by DNase1 treatment compared to controls. Discussion: Primary and secondary intention wound healing is improved by targeting NETs through DNase1 treatment or genetic KO, as assessed by wound closure time and scar appearances. Additionally, wound stability was not affected by DNASE treatment. The results suggest that overall wound healing is accelerated and DNase1 appears to be a promising option to reduce scar formation; which should be evaluated in humans.

The BAL-Score Almost Perfectly Predicts Testicular Torsion in Children: A Two-Center Cohort Study.

Introduction: Testicular torsion (TT) is a common emergency that warrants immediate exploration to prevent infertility or testicular loss. To improve diagnostic reliability, various scoring systems have been published. The aim of this study was to evaluate and validate different testicular torsion scores in a large cohort of children with acute scrotum. Methods: Retrospective analysis of all male children that were admitted for acute scrotum at the Pediatric Surgery Department of the Altonaer Kinderkrankenhaus and University medical Center Hamburg-Eppendorf from 01/2013 to 03/2019. Two testicular torsion scores (Boettcher Alert Score, Testicular Workup for Ischemia and Suspected Torsion Score) were applied to all data sets. Furthermore, an artificial intelligence (AI)-based score was developed and compared to the two current scores. Results: In total, 460 boys were included in the study. Of those, 48 (10.4%) had TT. Children with TT suffered most often from short duration of pain, nausea and vomiting, high riding testicle and absent cremasteric reflex. The BALS and the AI-based score had excellent predictive values and all patients with TT would have been detected. Conclusion: The BAL and the AI score show excellent predictive capabilities and may be used to identify all cases of TT in a pediatric population. The scores are easy to apply. As the BALS was slightly better, we advocate to use this score but to validate our findings in prospective multicenter studies.

The Modified Heidelberg and the AI Appendicitis Score Are Superior to Current Scores in Predicting Appendicitis in Children: A Two-Center Cohort Study.

Background: Acute appendicitis represents the most frequent reason for abdominal surgery in children. Since diagnosis can be challenging various scoring systems have been published. The aim of this study was to evaluate and validate (and improve) different appendicitis scores in a very large cohort of children with abdominal pain. Methods: Retrospective analysis of all children that have been hospitalized due to suspected appendicitis at the Pediatric Surgery Department of the Altonaer Children's Hospital and University Medical Center Hamburg-Eppendorf from 01/2018 until 11/2019. Four different appendicitis scores (Heidelberg Appendicitis Score, Alvarado Score, Pediatric Appendicitis Score and Tzanakis Score) were applied to all data sets. Furthermore, the best score was improved and artificial intelligence (AI) was applied and compare the current scores. Results: In 23 months, 463 patients were included in the study. Of those 348 (75.2%) were operated for suspected appendicitis and in 336 (96.6%) patients the diagnosis was confirmed histopathologically. The best predictors of appendicitis (simple and perforated) were rebound tenderness, cough/hopping tenderness, ultrasound, and laboratory results. After modifying the HAS, it provided excellent results for simple (PPV 95.0%, NPV 70.0%) and very good for perforated appendicitis (PPV 34.4%, NPV 93.8%), outperforming all other appendicitis score. Discussion: The modified HAS and the AI score show excellent predictive capabilities and may be used to identify most cases of appendicitis and more important to rule out perforated appendicitis. The new scores outperform all other scores and are simple to apply. The modified HAS comprises five features that can all be assessed in the emergency department as opposed to current scores that are relatively complex to utilize in a clinical setting as they include of up to eight features with various weighting factors. In conclusion, the modified HAS and the AI score may be used to identify children with appendicitis, yet prospective studies to validate our findings in a large mutli-center cohorts are needed.

Markers of NETosis Do Not Predict Neonatal Early Onset Sepsis: A Pilot Study.

Introduction: Early-onset sepsis in neonates potentially results in substantial morbidity and mortality. A key player in sepsis a neutrophil extracellular traps (NETs) to limit dissemination of pathogens. Aim of this study was to evaluate markers of NET formation in umbilical cord blood as a predictor of neonatal sepsis. Methods: Prospective study including term and preterm neonates. Umbilical cord blood samples were obtained immediately after birth and following markers of inflammation and NET formation were assessed: complete blood count, C-reactive protein (CRP), interleukin 6 (IL-6), levels of cell-free DNA (cfDNA), neutrophil elastase (NE), and myeloperoxidase (MPO). The study population included neonates with confirmed early-onset sepsis and propensity score matched controls. Results: Umbilical cord blood samples of 491 neonates were obtained, of whom 17 neonates (n = 17) presented clinical and laboratory signs of infection within the first 72 h postpartum. Seventeen neonates without infection were matched as controls. IL-6 differed significantly between both groups, whereas other infection parameters such as CRP and neutrophil levels, and in particular the NET surrogate markers (cfDNA, NE, MPO), did not show any significant differences. Conclusion: NET markers in umbilical cord blood appear to not predict the onset of neonatal sepsis. These findings probably result from the neonates' inability or delayed ability to form NETs, which is suspected to be a main reason for the increased risk of severe infections in neonates, but is also assumed to prevent negative NET-mediated consequences during perinatal adaptation.