Observed rates of surgical instrument errors point to visualization tasks as being a critically vulnerable point in sterile processing and a significant cause of lost chargeable OR minutes.

BACKGROUND: The reporting of surgical instrument errors historically relies on cumbersome, non-automated, human-dependent, data entry into a computer database that is not integrated into the electronic medical record. The limitations of these reporting systems make it difficult to accurately estimate the negative impact of surgical instrument errors on operating room efficiencies. We set out to determine the impact of surgical instrument errors on a two-hospital healthcare campus using independent observers trained in the identification of Surgical Instrument Errors. METHODS: This study was conducted in the 7 pediatric ORs at an academic healthcare campus. Direct observations were conducted over the summer of 2021 in the 7 pediatric ORs by 24 trained student observers during elective OR days. Surgical service line, error type, case type (inpatient or outpatient), and associated length of delay were recorded. RESULTS: There were 236 observed errors affecting 147 individual surgical cases. The three most common errors were Missing+ (n = 160), Broken/poorly functioning instruments (n = 44), and Tray+ (n = 13). Errors arising from failures in visualization (i.e. inspection, identification, function) accounted for 88.6% of all errors (Missing+/Broken/Bioburden). Significantly more inpatient cases (42.73%) had errors than outpatient cases (22.32%) (p = 0.0129). For cases in which data was collected on whether an error caused a delay (103), over 50% of both IP and OP cases experienced a delay. The average length of delays per case was 10.16 min. The annual lost charges in dollars for surgical instrument associated delays in chargeable minutes was estimated to be between $6,751,058.06 and $9,421,590.11. CONCLUSIONS: These data indicate that elimination of surgical instrument errors should be a major target of waste reduction. Most observed errors (88.6%) have to do with failures in the visualization required to identify, determine functionality, detect the presence of bioburden, and assemble instruments into the correct trays. To reduce these errors and associated waste, technological advances in instrument identification, inspection, and assembly will need to be made and applied to the process of sterile processing.

Using Preference Cards to Support a Thoughtful, Evidence-based Orthopaedic Surgery Practice.

INTRODUCTION: When orthopaedic surgeons begin or relocate their careers, they must communicate effectively about their instrumentation and equipment needs. 'Preference Cards' or 'Pick Lists' are generated by and for individual surgeons at the time of hire and can be updated over time to reflect their needs for common cases. Currently, such decisions are made without formal guidance or preparation. BODY: Surgeons must consider and plan for their operating room needs. Health system and industry factors affect these decisions, as do surgeons' unique interests, preferences, and biases. Orthopaedic surgeons currently face challenges: formal education is deficient in this space, material and reprocessing costs are not transparent, relationships and contracts with industry are complex, and few health systems have mechanisms to support preference card optimization. This complex landscape influences utilization decisions and leaves opportunities for integration, collaboration, and innovation. SUMMARY: Choices about instrument and resource utilization in the OR have wide-reaching impacts on costs, waste generation, OR efficiency, sterile processing, and industry trends. Surgeons and their teams have much to gain by making intentional choices and pursuing both individual and systematic improvements in this space.

Introduction of pediatric laparoscopic inguinal hernia repair in Guatemala.

PURPOSE: Introducing new surgical techniques in a developing country can be challenging. Inguinal hernias in children are a common surgical problem, and open repair is the standard surgical approach. Laparoscopic repair has gained popularity in developed countries because of similar results. This study aimed to determine the outcomes following the introduction of laparoscopic repairs in Guatemala. METHODS: This retrospective analysis of prospectively collected data from all patients under 18 years who underwent laparoscopic repair at Corpus Christi Hospital in Patzun, Guatemala, from September 5th to September 8th, 2022. RESULTS: A total of 14 patients were included in the study. A board-certified pediatric surgeon and a Guatemalan physician performed all cases. The mean patient age was 7.6 years; 7 boys and 7 girls. All patients were interviewed at 7 days, 30 days, and 6 months. There were no postoperative infections, pain requiring re-evaluation, gonadal atrophy, or hernia recurrence. CONCLUSION: Under controlled circumstances with limited but proper equipment and disposables, laparoscopic inguinal hernia repairs can be introduced and performed in a developing country with a risk complication profile comparable to that in developed countries. This study provides promising evidence of laparoscopic repair feasibility and safety where surgical resources are limited.

Reducing cost and waste in pediatric laparoscopic procedures.

BACKGROUND: In 2017 the healthcare cost in the United States accounted for 17.9% of the Gross Domestic Product (GDP). Furthermore, healthcare facilities produce more than 4 billion pounds of waste annually. Interhospital and intersurgeon variabilities in surgical procedures are some of the drivers of high healthcare cost and waste. We sought to determine the effect of a monthly surgeon report card detailing the utilization and cost of disposable and reusable surgical supplies on cost and waste reduction for pediatric laparoscopic procedures. METHODS: Starting in July 2017, surgeons were provided with an individual report with supply cost per case, high cost, and disposable supply utilization, and clinical outcomes. Cost, utilization, and clinical outcomes six quarters before and after the intervention were compared. RESULTS: A total of 998 pediatric laparoscopic procedures were analyzed. We reduced the median supply cost per case by 43% after the intervention with total cost savings of $71,035 for the first four quarters. We also reduced the use of disposable trocars by 56% and the use of disposable harmonics and staplers by 33%. CONCLUSIONS: Using a periodic surgeon report card, we significantly reduced supply cost and utilization of disposable items for all pediatric laparoscopic procedures performed at the University of Wisconsin American Family Children's Hospital. TYPE OF STUDY: Cost effectiveness study. LEVEL OF EVIDENCE: Level III.

Intercostal nerve cryoablation is associated with lower hospital cost during minimally invasive Nuss procedure for pectus excavatum.

Minimally invasive repair of pectus excavatum (Nuss procedure) is associated with significant pain, and efforts to control pain impact resource utilization. Bilateral thoracic intercostal nerve cryoablation has been proposed as a novel technique to improve post-operative pain control, though the impact on hospital cost is unknown. METHODS: We conducted a retrospective study of patients undergoing a Nuss procedure from 2016 to 2019. Patients who received cryoablation were compared to those that received traditional pain control (patient-controlled analgesia or epidural). Outcome variables included postoperative opioid usage (milligram morphine equivalents, MME), length of stay (LOS), and hospital cost. RESULTS: Thirty-five of 73 patients studied (48%) received intercostal nerve cryoablation. LOS (1.0 vs 4.0 days, p < 0.01) and total hospital cost ($21,924 versus $23,694, p = 0.04) were decreased in the cryoablation cohort, despite longer operative time (152 vs 74 min, p < 0.01). Cryoablation was associated with decreased opioid usage (15.0 versus 148.6 MME, p < 0.01) during the 24 h following surgery and this persisted over the entire postoperative period, including discharge opioid prescription (112.5 vs 300.0 MME, p < 0.01). CONCLUSION: Bilateral intercostal nerve cryoablation is associated with decreased postoperative opioid usage and decreased resource utilization in pediatric patients undergoing a minimally invasive Nuss procedure for pectus excavatum. LEVEL OF EVIDENCE: Retrospective comparative study, level III.

Lack of discreet colocalization of epithelial apoptosis to the atretic precursor in the colon of the Fibroblast growth factor receptor 2IIIb mouse and staining consistent with cellular movement suggest a revised model of atresia formation.

BACKGROUND: Colonic atresias in the Fibroblast growth factor receptor 2IIIb (Fgfr2IIIb) mouse model have been attributed to increased epithelial apoptosis and decreased epithelial proliferation at embryonic day (E) 10.5. We therefore hypothesized that these processes would colocalize to the distal colon where atresias occur (atretic precursor) and would be excluded or minimized from the proximal colon and small intestine. RESULTS: We observed a global increase in intestinal epithelial apoptosis in Fgfr2IIIb -/- intestines from E9.5 to E10.5 that did not colocalize to the atretic precursor. Additionally, epithelial proliferations rates in Fgfr2IIIb -/- intestines were statistically indistinguishable to that of controls at E10.5 and E11.5. At E11.5 distal colonic epithelial cells in mutants failed to assume the expected pseudostratified columnar architecture and the continuity of the adjacent basal lamina was disrupted. Individual E-cadherin-positive cells were observed in the colonic mesenchyme. CONCLUSIONS: Our observations suggest that alterations in proliferation and apoptosis alone are insufficient to account for intestinal atresias and that these defects may arise from both a failure of distal colonic epithelial cells to develop normally and local disruptions in basal lamina architecture.

RW-2018-Research Workshop: The Effect of Nutrition on Epigenetic Status, Growth, and Health.

The goal of the 2018 American Society for Parenteral and Enteral Nutrition (ASPEN) Research Workshop was to explore the influence of nutrition and dietary exposure to xenobiotics on the epigenome during critical periods in development and how these exposures influence both disease incidence and severity transgenerationally. A growing compendium of research indicates that the incidence and severity of common and costly human diseases may be influenced by dietary exposures and deficiencies that modify the epigenome. The greatest periods of vulnerability to these exposures are the periconception period and early childhood. Xenobiotics in the food chain, protein malnutrition, and methyl donor deficiencies could have a profound bearing on the risk of developing heart disease, diabetes, obesity, hypertension, and mental illness over multiple generations. The financial impact and the life burden of these diseases are enormous. These and other aspects of nutrition, epigenetics, and health are explored in this research workshop.

Management of asymptomatic pediatric umbilical hernias: a systematic review.

INTRODUCTION: Uncomplicated pediatric umbilical hernias are common and most close spontaneously. No formal practice guidelines exist regarding the optimal timing and indications for repair. The objective of this review is to examine the existing literature on the natural history of pediatric umbilical hernias, known complications of repair and non-operative approaches, and management recommendations. STUDY DESIGN: A systematic literature search was performed to identify publications relating to pediatric umbilical hernias. Inclusion criteria comprised studies addressing recommendations for optimal timing of repair, evidence examining complications from hernias not operatively repaired, and research exploring the likelihood of pediatric umbilical hernias to close spontaneously. In addition, the websites of all pediatric hospitals in the United States were examined for recommendations on operative timing. RESULTS: A total of 787 manuscripts were reviewed, and 28 met criteria for inclusion in the analysis. Studies examined the likelihood of spontaneous closure based on child's age and size of hernia defect, complications of unrepaired umbilical hernias including incarceration, strangulation and evisceration based on child's age and size of defect, incidence of postoperative complications and current recommendations for timing of repair. In addition, 63 (27.5%) of the United States pediatric hospital websites published a wide range of management recommendations. CONCLUSION: Despite the high prevalence of pediatric umbilical hernias, there is a paucity of high quality data to guide management. The literature does suggest that expectant management of asymptomatic hernias until age 4-5years, regardless of size of hernia defect, is both safe and the standard practice of many pediatric hospitals. TYPE OF STUDY: Review Article. LEVEL OF EVIDENCE: Level IV.

Synchronous Presentation of Renal Cell Carcinoma and Hodgkin Lymphoma in an Adolescent.

BACKGROUND: Coincidence of renal cell carcinoma (RCC) and hematologic malignancies has been reported in adults but not in children. OBSERVATION: We report a case of a 16-year-old girl in whom RCC was incidentally discovered on the computed tomography scan that was performed to stage her underlying Hodgkin lymphoma. Analysis of constitutional cytogenetics for common genetic aberrations that predispose to RCC did not reveal any mutations or genetic variations. However, cytogenetics on the RCC tumor demonstrated a rare reciprocal translocation between chromosomes 6 and 11, t(6;11)(p21;q12). After undergoing partial nephrectomy with regional lymphadenectomy and treatment with multiagent chemotherapy, patient is cancer-free, now 33 months from end of therapy. CONCLUSIONS: This case highlights the importance for histologic confirmation of a renal mass when concurrently discovered during the diagnostic evaluation of other malignancies.

Outcomes from a 12-Week, Open-Label, Multicenter Clinical Trial of Teduglutide in Pediatric Short Bowel Syndrome.

OBJECTIVE: To determine safety and pharmacodynamics/efficacy of teduglutide in children with intestinal failure associated with short bowel syndrome (SBS-IF). STUDY DESIGN: This 12-week, open-label study enrolled patients aged 1-17 years with SBS-IF who required parenteral nutrition (PN) and showed minimal or no advance in enteral nutrition (EN) feeds. Patients enrolled sequentially into 3 teduglutide cohorts (0.0125 mg/kg/d [n = 8], 0.025 mg/kg/d [n = 14], 0.05 mg/kg/d [n = 15]) or received standard of care (SOC, n = 5). Descriptive summary statistics were used. RESULTS: All patients experienced ≥1 treatment-emergent adverse event; most were mild or moderate. No serious teduglutide-related treatment-emergent adverse events occurred. Between baseline and week 12, prescribed PN volume and calories (kcal/kg/d) changed by a median of -41% and -45%, respectively, with 0.025 mg/kg/d teduglutide and by -25% and -52% with 0.05 mg/kg/d teduglutide. In contrast, PN volume and calories changed by 0% and -6%, respectively, with 0.0125 mg/kg/d teduglutide and by 0% and -1% with SOC. Per patient diary data, EN volume increased by a median of 22%, 32%, and 40% in the 0.0125, 0.025, and 0.05 mg/kg/d cohorts, respectively, and by 11% with SOC. Four patients achieved independence from PN, 3 in the 0.05 mg/kg/d cohort and 1 in the 0.025 mg/kg/d cohort. Study limitations included its short-term, open-label design, and small sample size. CONCLUSIONS: Teduglutide was well tolerated in pediatric patients with SBS-IF. Teduglutide 0.025 or 0.05 mg/kg/d was associated with trends toward reductions in PN requirements and advancements in EN feeding in children with SBS-IF. TRIAL REGISTRATION: ClinicalTrials.gov:NCT01952080; EudraCT: 2013-004588-30.

A call for a standardized definition of perforated appendicitis.

BACKGROUND: Abscess rates have been reported to be as low as 1% and as high as 50% following perforated appendicitis (PA). This range may be because of lack of universal definition for PA. An evidence-based definition (EBD) is crucial for accurate wound classification, risk-stratification, and subsequent process optimization. ACS NSQIP-Pediatric guidelines do not specify a definition of PA. We hypothesize that reported postoperative abscess rates underrepresent true incidence, as they may include low-risk cases in final calculations. METHODS: Local institutional records of PA patients were reviewed to calculate the postoperative abscess rate. The ACS NSQIP-Pediatric participant use file (PUF) was used to determine cross-institutional postoperative abscess rates. A PubMed literature review was performed to identify trials reporting PA abscess rates, and definitions and rates were recorded. RESULTS: 20.9% of our patients with PA developed a postoperative abscess. The ACS NSQIP-Pediatric abscess rate was significantly lower (7.61%, p<0.001). In the eighteen published studies analyzed, average abscess rate (14.49%) was significantly higher than ACS NSQIP-Pediatric (p<0.001). There was significantly more variation in trials that do not employ an EBD of perforation (Levene's test F-value =6.980, p=0.018). CONCLUSIONS: A standard EBD of perforation leads to lower variability in reported postoperative abscess rates following PA. Nonstandard definitions may be significantly altering the aggregate rate of postoperative abscess formation. We advocate for adoption of a standard definition by all institutions participating in ACS NSQIP-Pediatric data submission. LEVEL OF EVIDENCE: III.

A Single Institution Review of Initial Application of a 5-mm Stapler.

BACKGROUND: Operating in small spaces presents physical constraints that can be even more challenging in minimally invasive operations. Recently, a 5-mm stapler was approved for use in general surgery and pediatric surgery. Here, we present our initial experience using the 5-mm stapler in pediatric general surgery. MATERIALS AND METHODS: A retrospective chart review was conducted to identify cases using the 5-mm stapler at our institution. Demographic data included age (in months) and weight (in kilograms). Operative data included indication for use, number of loads used, complications related to 5-mm stapler use, and interventions to address complications. A second review focused on patients undergoing the same operations, but using a 10-mm stapler. RESULTS: A total of 60 staple loads were deployed in 32 procedures. There were four adverse outcomes, all recognized intraoperatively. One bleed resulted from application on irradiated tissue and another bleed from application to a small noninflamed mesoappendix. A bronchial staple line leak resulted from improper stapler loading, and a bowel anastomosis leak was oversewn with a single stitch. When compared with 32 matched cases using a 10-mm stapler, there was no difference in age (5-mm = 39.11 months, 10-mm = 50.21 months, P = .49) or weight (5-mm = 16.34 kg, 10-mm = 19.93 kg, P = .51). A total of 60 staple applications were used, with one bleed noted. There was no significant difference in overall complication rate (5-mm rate = 4/60, 10-mm rate = 1/60; P = .36). CONCLUSION: Our initial experience suggests that although there were more complications with the 5-mm stapler, there is no statistically significant difference in complication rates when compared with the 10-mm stapler. Furthermore, the 5-mm stapler complications can be corrected with device training and proper patient selection. In appropriately selected pediatric surgery cases with size limitations, the 5-mm stapler can be used to minimize the invasiveness of the operation.

Long-Term Outcomes of Patients with Tracheoesophageal Fistula/Esophageal Atresia: Survey Results from Tracheoesophageal Fistula/Esophageal Atresia Online Communities.

Introduction Outcome studies of tracheoesophageal fistula (TEF) and/or esophageal atresia (EA) are limited to retrospective chart reviews. This study surveyed TEF/EA patients/parents engaged in social media communities to determine long-term outcomes. Materials and Methods A 50-point survey was designed to study presentation, interventions, and ongoing symptoms after repair in patients with TEF/EA. It was validated using a test population and made available on TEF/EA online communities. Results In this study, 445 subjects completed the survey during a 2-month period. Mean age of patients when surveyed was 8.7 years (0-61 years) and 56% were male. Eighty-nine percent of surveys were completed by the parent of the patient. Sixty-two percent of patients underwent repair in the first 7 days of life. Standard open repair was most common (56%), followed by primary esophageal replacement (13%) and thoracoscopic repair (13%). Out of 405, 106 (26%) patients had postoperative leak. Postoperative leak was least likely in primary esophageal replacement (18%) and standard open repair (19%). Leak occurred in 32% of patients who had thoracoscopic repair; 31% (128/413) reported long-gap atresia, which was significantly associated with increased risk of postoperative leak (54/128, 42%) when compared with standard short-gap atresia (odds ratio, 3.5; p = 0.001). Out of 409, 221 (54%) patients reported dysphagia after repair, with only 77/221 (34.8%) reporting resolution by age 5. Out of 381, 290 (76%) patients reported symptoms of gastroesophageal reflux disease (GERD). There was no difference in dysphagia rates or GERD symptoms based on type of initial repair. Antireflux surgery was required in 63/290, 22% of patients with GERD (15% of all patients) and 27% of these patients who had surgery required more than one procedure antireflux procedure. The most common was Nissen fundoplication (73%), followed by partial wrap (14%). Reflux recurred in 32% of patients after antireflux surgery. Conclusion TEF/EA patients have long-term dysphagia and GERD that may be under reported. Retrospective studies of outcomes after TEF/EA repair may underestimate long-term esophageal dysmotility, dysphagia, GERD, and strictures that occur regardless of the type of repair and adversely affect quality of life. Fifteen percent of all TEF/EA patients surveyed required an antireflux procedure during childhood, and more than one-quarter of those required repeat surgery. These data demonstrate the need for long-term follow-up as pediatric patients transition to adult care.

Evolution in management of adolescent blunt aortic injuries--a single institution 22-y experience.

BACKGROUND: In children, severe, life-threatening traumatic injuries of the thoracic aorta can be seen after motor vehicle collisions (MVCs) resulting in a sudden deceleration. Concurrent injuries in the thorax and abdomen can make treatment prioritization difficult and require early recognition and prompt intervention. With the increased utilization of minimally invasive endovascular approaches to traumatic aortic (TA) injuries, patients are often spared the increased surgical morbidity (spinal cord ischemia and renal insults) that can be seen with an open technique. The aim of this study was to evaluate a single American College of Surgeons level 1 pediatric trauma center's 22-y experience with TA injuries in children. METHODS: After the Institutional Review Board approval, a 22-y (January 1990-April 2013) retrospective review of all pediatric trauma patients admitted with TA injuries was performed. Patient demographics including age, injury detail, treatment, and outcomes were recorded for analysis. RESULTS: 17 children (<21-y old) were identified with ages ranging from 13-20 y old. The most common mechanism of injury was MVC with all 17 children sustaining TA injuries. The traumatic injuries included aortic transection (9), intimal flap (5), pseudoaneurysm (2), and contained thoracic rupture (1). All children were managed operatively with those before 2008 using an open technique. The endovascular approach was used in 7/17 (41%) cases with the median length of hospitalization 12 d versus 22.5 d using the open approach (P < 0.05). No child required conversion from an endovascular to an open technique for treatment of the aortic injury. There were no operative deaths, no procedure-related paraplegia and all children were discharged home from the hospital. Two children had mild mental deficits as a result of head trauma. CONCLUSIONS: TA injuries are an uncommon injury in children and can result from MVCs or other sudden deceleration mechanisms. Surgical intervention is required in most of the cases and can be performed safely and effectively with low morbidity using an endovascular approach, which is the evolving approach of choice for thoracic aortic injuries. Lengthy follow-up care is recommended in children treated with an endovascular device to monitor for endoleaks and device complications.

National trends in pediatric blunt spleen and liver injury management and potential benefits of an abbreviated bed rest protocol.

PURPOSE: Recent reports suggest that an abbreviated bed rest protocol (ABRP) may safely reduce length of stay (LOS) and resource utilization in pediatric blunt spleen and liver injury (BSLI) patients. This study evaluates national temporal trends in BLSI management and estimates national reduction in LOS using an ABRP. METHODS: Pediatric patients (<18 years old) sustaining BLSI were identified in the Kids' Inpatient Database from 2000 to 2009. Yearly rates of injury and operative intervention were examined and stratified by type of injury. APSA guidelines and the reported ABRP were applied based on abbreviated injury score (AIS) and compared with actual LOS. RESULTS: 22,153 patients were identified. Over the study period, operative rates for spleen and liver injuries and overall mortality significantly declined: LOS=3.1 days (±1.6) and 2.7 days (±1.9) for spleen and liver, respectively. If APSA guidelines were followed, the rates were LOS=3.7 days (±1.1) and 3.4 days (±0.7), respectively. Application of the ABRP would result in LOS=1.3 days (±0.5) for all BSLI patients. An ABRP could potentially save 1.7 hospital days/patient or 36,964 patient hospital days nationally. CONCLUSION: Our study confirms a significant national decrease in operative intervention and overall mortality in patients with BSLI. Additionally, it appears that a shorter observation period than the APSA guidelines is being utilized. The implementation of ABRP holds potential in further reducing LOS and resource utilization.

Exogenous Sonic hedgehog protein does not rescue cultured intestine from atresia formation.

BACKGROUND: The mechanism of intestinal atresia formation remains undefined. Atresia in fibroblast growth factor receptor 2IIIb (Fgfr2IIIb(-/-)) mutant mouse embryos is preceded by endodermal apoptosis and involution of the surrounding mesoderm. We have observed that involution of the atretic segment is preceded by the downregulation of Sonic hedgehog (SHH) in the endoderm, which is a critical organizer of the intestinal mesoderm. We hypothesized that supplementation of Fgfr2IIIb(-/-) intestinal tracts with exogenous SHH protein before atresia formation would prevent involution of the mesoderm and rescue normal intestinal development. METHODS: In situ hybridization was performed on control and Fgfr2IIIb(-/-) intestinal tracts for Shh or forkhead box protein F1 (FoxF1) between embryonic (E) day 11.5 and E12.0. Control and Fgfr2IIIb(-/-) intestinal tracts were harvested at E10.5 and cultured in media supplemented with fibroblast growth factor (FGF) 10 + SHH, or FGF10 with a SHH-coated bead. In situ hybridization was performed at E12.5 for Foxf1. RESULTS: SHH and Foxf1 expression were downregulated during intestinal atresia formation. Media containing exogenous FGF10 + SHH did not prevent colonic atresia formation (involution). A SHH protein point source bead did induce Foxf1 expression in controls and mutants. CONCLUSIONS: Shh and Foxf1 expression are disrupted in atresia formation of distal colon, thereby serving as potential markers of atretic events. Application of exogenous SHH (in media supplement or as a point source bead) is sufficient to induce Foxf1 expression, but insufficient to rescue development of distal colonic mesoderm in Fgfr2IIIb(-/-) mutant embryos. Shh signal disruption is not the critical mechanism by which loss of Fgfr2IIIb function results in atresia formation.

Utility and limits of Hprt-Cre technology in generating mutant mouse embryos.

BACKGROUND: Hprt-Cre doubles the prevalence of homozygous null embryos per litter versus heterozygous breedings without decreasing litter size. Resulting mutant embryos are genotypically and phenotypically equivalent between strategies. We set out to confirm the effectiveness of this approach with other alleles and hypothesized that it would increase efficiency in generating compound mutants. MATERIALS AND METHODS: Null mutants for Cyp26b1, Pitx2, and Shh were generated with Hprt-Cre from conditional alleles as were double and triple allelic combinations of Fgfr2IIIb, Raldh2, and Cyp26b1. Embryos were genotyped and phenotyped by whole mount photography, histology, and immunohistochemistry. RESULTS: Fifty percent of Hprt-Cre litters were homozygous null for Cyp26b1 (15/29) and Pitx2 (75/143), with phenotypic and genotypic equivalence to mutants from standard heterozygous breedings. In multi-allele breedings, mutant embryos constituted half of litters without significant embryo loss. In contrast, Shh breedings yielded a smaller ratio of embryos carrying two recombined alleles (6 of 16), with a significant litter size reduction because of early embryonic lethality (16 live embryos from 38 deciduae). CONCLUSIONS: Hprt-Cre can be used to efficiently generate large numbers of mutant embryos with a number of alleles. Compound mutant generation was equally efficient. However, efficiency is reduced for genes whose protein product potentially interacts with the Hprt pathway (e.g., Shh).

Haploinsufficiency of retinaldehyde dehydrogenase 2 decreases the severity and incidence of duodenal atresia in the fibroblast growth factor receptor 2IIIb-/- mouse model.

BACKGROUND: Homozygous null mutation of the fibroblast growth factor receptor 2IIIb (Fgfr2IIIb) gene in mice results in 42% of embryos developing duodenal atresias. Retinaldehyde dehydrogenase 2 (Raldh2, a gene critical for the generation of retinoic acid) is expressed in the mouse duodenum during the temporal window when duodenal atresias form. Raldh2 is critical for the normal development of the pancreatoduodenal region; therefore, we were interested in the effect of a Raldh2 mutation on duodenal atresia formation. To test this, we rendered Fgfr2IIIb(-/-) embryos haploinsufficient for the Raldh2 and examined these embryos for the incidence and severity of duodenal atresia. METHODS: Control embryos, Fgfr2IIIb(-/-) mutants, and Fgfr2IIIb(-/-); Raldh2(+/-) mutants were harvested at embryonic day 18.5, genotyped, and fixed overnight. Intestinal tracts were isolated. The type and severity of duodenal atresia was documented. RESULTS: A total of 97 Fgfr2IIIb(-/-) embryos were studied; 44 had duodenal atresias, and 41 of these presented as type III. In the 70 Fgfr2IIIb(-/-); Raldh2(+/-) embryos studied, a lesser incidence of duodenal atresia was seen (15 of 70; P = .0017; Fisher exact test). Atresia severity was also decreased; there were 12 embryos with type I atresias, 3 with type II atresias, and 0 with type III atresias (P < 2.81E-013; Fisher exact test). CONCLUSION: Haploinsufficiency of Raldh2 decreases the incidence and severity of duodenal atresia in the Fgfr2IIIb(-/-) model. The ability to alter defect severity through manipulation of a single gene in a specific genetic background has potentially important implications for understanding the mechanisms by which intestinal atresias arise.