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Introduction The purpose of this study was to develop a multi-institutional registry to characterize the demographics, management, and outcomes of a contemporary cohort of children undergoing congenital lung malformation (CLM) resection. Methods After central reliance IRB approval, a web-based, secure database was created to capture retrospective cohort data on pathologically-confirmed CLMs performed between 2009 and 2015 within a multi-institutional research collaborative. Results Eleven children's hospitals contributed 506 patients. Among 344 prenatally diagnosed lesions, the congenital pulmonary airway malformation volume ratio was measured in 49.1%, and fetal MRI was performed in 34.3%. One hundred thirty-four (26.7%) children had respiratory symptoms at birth. Fifty-eight (11.6%) underwent neonatal resection, 322 (64.1%) had surgery at 1–12 months, and 122 (24.3%) had operations after 12 months. The median age at resection was 6.7 months (interquartile range, 3.6–11.4). Among 230 elective lobectomies performed in asymptomatic patients, thoracoscopy was successfully utilized in 102 (44.3%), but there was substantial variation across centers. The most common lesions were congenital pulmonary airway malformation (n = 234, 47.3%) and intralobar bronchopulmonary sequestration (n = 106, 21.4%). Conclusion This multicenter cohort study on operative CLMs highlights marked disease heterogeneity and substantial practice variation in preoperative evaluation and operative management. Future registry studies are planned to help establish evidence-based guidelines to optimize the care of these patients. Level of evidence Level II.

A detailed understanding of protective immunity against SARS-CoV-2 is incredibly important in fighting the pandemic. Central to protective immunity is the ability of the immune system to recall previous exposures. Although antibody and T cell immunity have gained considerable attention, the contribution of the NK cell compartment to immune recall and protection from SARS-CoV-2 has not been explored. In this study, we investigate the NK cell responses to stimulation with SARS-CoV-2 in previously exposed and non-exposed individuals. We show that NK cells demonstrate an enhanced CD4+ T cell dependent response when re-exposed to SARS-CoV-2 antigen. The enhanced response is dependent on T cells and correlates with the number of SARS-CoV-2 specific CD4 T cells. We find that IL-2 is a critical mediator of NK cell function. These findings suggest that NK cells contribute to the protective responses against SARS-CoV-2 through a cooperation with antigen-specific CD4 T cells and have significant implications on our understanding of protective immunity in SARS-CoV-2.

Background: The factors that contribute to success as a pediatric surgeon–scientist are not well defined. The purpose of this study is to define a group of NIH-funded pediatric surgeons, assess their academic productivity, and elucidate factors that have contributed to their success. Methods: Pediatric surgeons were queried in the NIH report database to determine NIH funding awarded. Academic productivity was then assessed. An online survey was then targeted to NIH-funded pediatric surgeons. Results: Since 1988, 83 pediatric surgeon–investigators have received major NIH funding. Currently, there are 37 pediatric surgeons with 43 NIH-sponsored awards. The mean h -index of this group of pediatric surgeons was 18±1.1, mean number of publications (since 2001) was 21±2.1, and both increase commensurate with academic rank. In response to the survey, 81% engaged in research during their surgical residency, and 48% were mentored by a pediatric surgeon–scientist. More than 60% of respondents had significant protected time and financial support. Factors felt to be most significant for academic success included mentorship, perseverance, and protected time. Conclusions: Mentorship, perseverance, institutional commitment to protected research time, and financial support are considered to be important to facilitate the successes of pediatric surgeon–scientists. These results will be useful to aspiring pediatric surgeon–scientists and departments wishing to develop a robust research program.

Anti-inflammatory cytokine interleukin (IL)-10 has been shown to induce regenerative healing in postnatal wounds. A viral homolog of IL-10 produced by human cytomegalovirus (CMV IL-10) similarly generates potent immunoregulatory effects, but its effects on wound healing have not been investigated. Currently, there are limited cost-effective methods of screening vulnerary therapeutics. Taken together, we aim to develop and validate a novel human ex vivo dermal wound model and hypothesize that CMV IL-10 will enhance dermal wound healing.Full-thickness circular (6-mm) explants were taken from surgical skin samples and 3-mm full-thickness wounds were created. Explants were embedded in collagen I matrix and maintained in specially formulated media with the epidermis at air-liquid interface, and treated with human IL-10 or CMV IL-10 (200 ng/mL). The viability of cultured explants was validated by histology and lactate dehydrogenase (LDH) activity. Epithelial gap, epithelial height, basal keratinocyte migration, vascular endothelial growth factor levels, and neovascularization were measured at days 3 and 7 to determine IL-10 effects on wound healing.Culture explants at day 7 appeared similar to fresh skin in morphology, cell, and vessel density. By day 14, the epidermis separated from the dermis and the cell density diminished. Day 7 wounds appeared viable with advancing epithelial and basal keratinocyte migration with no evidence of necrosis. Cytotoxicity analysis via the quantification of LDH revealed no differences between controls and treated groups. There was a slight increase in the quantity of LDH in media at day 3; however, this decreased at day 5 and continued to decline up to day 21. CMV IL-10 treatment resulted in a significant decrease in the epithelial gap and an increase in epithelial height. There were no differences in the rates of basal keratinocyte migration at day 7 between treated and control groups. Interestingly, human IL-10 increased vascular endothelial growth factor expression and neovascularization compared with controls.The human ex vivo wound model provides a simple and viable design to study dermal wound healing. Both IL-10 homologs demonstrate vulnerary effects. The viral homolog demonstrates enhanced effects on wound closure compared with human IL-10. These data represent a novel tool that can be used to screen therapeutics, such as CMV IL-10, before preclinical studies.

Background Chronic wounds are characterized by a wound healing and neovascularization deficit. Strategies to increase neovascularization can significantly improve chronic wound healing. Insulin-like growth factor (IGF)-1 is reported to be a keratinocyte mitogen and is believed to induce angiogenesis via a vascular endothelial growth factor (VEGF)-dependent pathway. Using a novel ex vivo human dermal wound model and a diabetic-impaired wound healing murine model, we hypothesized that adenoviral overexpression of IGF-1 (Ad-IGF-1) will enhance wound healing and induce angiogenesis through a VEGF-dependent pathway. Methods Ex vivo: 6-mm full-thickness punch biopsies were obtained from normal human skin, and 3-mm full-thickness wounds were created at the center. Skin explants were maintained at air liquid interface. Db/db murine model: 8-mm full-thickness dorsal wounds in diabetic (db/db) mice were created. Treatment groups in both human ex vivo and in vivo db/db wound models include 1 × 108 particle forming units of Ad-IGF-1 or Ad-LacZ, and phosphate buffered saline (n = 4–5/group). Cytotoxicity (lactate dehydrogenase) was quantified at days 3, 5, and 7 for the human ex vivo wound model. Epithelial gap closure (hematoxylin and eosin; Trichrome), VEGF expression (enzyme-linked immunosorbent assay), and capillary density (CD 31 + CAPS/HPF) were analyzed at day 7. Results In the human ex vivo organ culture, the adenoviral vectors did not demonstrate any significant difference in cytotoxicity compared with phosphate buffered saline. Ad-IGF-1 overexpression significantly increases basal keratinocyte migration, with no significant effect on epithelial gap closure. There was a significant increase in capillary density in the Ad-IGF-1 wounds. However, there was no effect on VEGF levels in Ad-IGF-1 samples compared with controls. In db/db wounds, Ad-IGF-1 overexpression significantly improves epithelial gap closure and granulation tissue with a dense cellular infiltrate compared with controls. Ad-IGF-1 also increases capillary density, again with no significant difference in VEGF levels in the wounds compared with control treatments. Conclusions In two different models, our data demonstrate that adenoviral-mediated gene transfer of IGF-1 results in enhanced wound healing and induces angiogenesis via a VEGF-independent pathway. Understanding the underlying mechanisms of IGF-1 effects on angiogenesis may help produce novel therapeutics for chronic wounds or diseases characterized by a deficit in neovascularization.

Background The current research environment for academic surgeons demands that extramural funding be obtained. Financial support from the National Institutes of Health (NIH) is historically the gold standard for funding in the biomedical research community, with the R01 funding mechanism viewed as indicator of research independence. The NIH also supports a mentor-based career development mechanism (K-series awards) in order to support early-stage investigators. The goal of this study was to investigate the grants successfully awarded to pediatric surgeon-scientists and then determine the success of the K-series award recipients at achieving research independence. Methods In July 2012, all current members of the American Pediatric Surgery Association (APSA) were queried in the NIH database from 1988–2012 through the NIH Research Portfolio Online Reporting Tools. The following factors were analyzed: type of grant, institution, amount of funding, and funding institute or center. Results Among current APSA members, there have been 83 independent investigators receiving grants, representing 13% of the current APSA membership, with 171 independent grants funded through various mechanisms. Six percent currently have active NIH funding, with $7.2 million distributed in 2012. There have been 28 K-series grants awarded. Of the recipients of expired K08 awards, 39% recipients were subsequently awarded an R01 grant. A total of 63% of these K-awarded investigators transitioned to an independent NIH award mechanism. Conclusions Pediatric surgeon-scientists successfully compete for NIH funding. Our data suggest that although the K-series funding mechanism is not the only path to research independence, over half of the pediatric surgeons who receive a K-award are successful in the transition to independent investigator.

Gastroschisis is a congenital abdominal wall defect in which the intestines develop outside the abdomen and are exposed to amniotic fluid. When the defect is small, lymphatic, venous, and intestinal obstruction may occur and contribute to the formation of intestinal edema, atresia, ischemia, and a thick inflammatory peel. Treatment requires early coverage of abdominal contents either by primary closure or by the placement of temporary Silastic silo followed by abdominal wall closure. Currently, both traditional suture closure and the sutureless plastic closure are being employed to repair the gastroschisis defect. The goal of the current study is to evaluate plastic closure. We predict no difference will be found in clinical outcomes between plastic closure and traditional suture closure.A retrospective review of 80 patients treated between 2000 and 2009 was performed. Plastic closure was used in 52 (65%) and traditional suture closure in 28 (35%) babies. The surgical procedure was determined by surgeon preference. Of the 31(39%) babies who required silos, 15 (19%) were treated with plastic closure and 16 (20%) underwent traditional closure. We collected the following demographic data and clinical progression data. Using SAS 9.2 (SAS Institute Inc, Cary, NC), we conducted linear regression, logistic regression, and time to event models to compare the following outcomes: days on ventilator, days to start enteral feeds, days to reach goal enteral feeds, days on total parenteral nutrition, hospital charges, duration of stay, mortality, and complications.The mean duration of follow-up was 11.4 months. Patients spent an average of 6 days on the ventilator. There were 2 mortalities. A multivariate analysis demonstrated that no differences were found between the 2 closures with most of the outcomes; however, when compared with traditional suture closure, those babies treated with plastic closure spent 4 days fewer days on the ventilator (P.01). Those babies who underwent suture closure were more likely to have an infection or sepsis (odds ratio, 5.15; P.001). When the entire cohort was considered, no significant difference was found between plastic and suture closure in time to start feeds, time to reach goal feeds, time on parenteral nutrition, hospital charges, duration of stay, or complications. Ventral hernias were noted in 46 (58%) patients, 32 (62%) after plastic closure and 14 (50%) after suture closure (P = .32). Hernia repair was required in 16 (20%) patients, 11 (21%) after plastic closure, and 5 (18%) after traditional repair (P = .32). In the silo cohort, children treated with plastic closure required 7.5(P.01) fewer days to start enteral feeds than those treated with suture closure.Plastic closure of abdominal wall defects in gastroschisis is effective both as a primary procedure and after silo placement. A multivariate analysis shows plastic closure to be associated with fewer days of mechanical ventilation and less likelihood of developing infection or sepsis.

Background Clerkship directors (CDs) are key educators and active clinicians. In 2003, the Alliance for Clinical Education published standards for CD resources and responsibilities, but how reality compares is unknown. Methods Representatives from each core clinical disciplines' CD organizations created an electronic survey that CDs received in 2006–2007. Results More than 500 CDs responded, including 71 surgeons. Surgeons reported spending approximately 27% of professional time on education. Most have codirectors, so total CD effort approximates the greater than 50% Alliance for Clinical Education guidelines. No disciplines' CDs have more than one support staff as recommended. Surgeons have the least clinic time, but the most inpatient weeks and many publications. Surgery CD concerns are curricula and simulation; few believe being a CD impairs academic advancement and more than 95% believe it enhances work satisfaction. Conclusions Surgery CDs are clinically active and academically productive. Although few surgery CDs have the recommended support staff, more than 95% report being a CD enhances work satisfaction.

Background/Purpose Spinal muscular atrophy (SMA) in children leads to progressive muscle weakness, dysphagia, aspiration, and death. We hypothesized that early laparoscopic fundoplication and gastrostomy in infants with SMA type I could be performed safely perhaps leading to fewer aspiration events and improved nutritional status. Methods Children diagnosed with SMA type I from 2002 through 2005 were included (n = 12). All children underwent laparoscopic Nissen fundoplication with gastrostomy shortly after diagnosis. Postoperative respiratory management and discharge criteria were standardized. Results All patients were extubated immediately postoperatively. There were no significant complications. Average time to full feeding and inpatient length of stay were 42 ± 4.9 hours (range, 30-48 hours) and 78 ± 22.5 hours (range, 44-120 hours), respectively. Mean weight-for-length percentile was doubled at 1 year postoperatively ( P = .03). The number of respiratory-related hospitalizations in the cohort decreased by almost 50% in the ensuing 12 months after surgery, although this did not reach statistical significance in this small cohort ( P = .34). Conclusions Early laparoscopic fundoplication and gastrostomy is safe and is associated with improved nutritional status. A trend toward fewer significant long-term aspiration-related events was seen after fundoplication. To better assess the long-term benefits of performing an antireflux procedure in these high-risk patients, a larger prospective trial comparing current nutritional support practices is needed.

The development of hydrops fetalis and maternal mirror syndrome in the setting of fetal sacrococcygeal teratoma (SCT) carries a grave prognosis. Fetal surgery is not typically offered for hydropic fetuses beyond 26 weeks of gestational age. Few options exist for these families, and they are typically counseled to continue pregnancy until delivery is necessary for maternal indications or the fetus is 30 weeks old. Although a number of series report the survival of hydropic SCT infants delivered after 30 weeks of gestation, a paucity of information is available regarding the outcome of hydropic SCT infants delivered between 26 and 30 weeks of gestation. We now report successful postpartum resection and survival of a hydropic fetus delivered at 27 weeks for maternal mirror syndrome that may assist in counseling families in similar situations.

In the absence of immunodeficiency, only microchimerism (0.1%) has been achieved in human fetal recipients or nonhuman primates following in utero hematopoietic cell transplantation (IUHCT). We hypothesized that enhanced long-term engraftment might be more reliably achieved in microchimeric systems if higher levels of chimerism existed during development of adaptive immunity. To evaluate this hypothesis, we stimulated the donor cells with vascular endothelial growth factor (VEGF) and stem cell factor (SCF) prior to IUHCT in a chimerism-resistant murine strain combination.Donor Balb/c marrow was cultured in media with or without VEGF and SCF supplementation for 12 hours prior to IUHCT into B6 fetuses at 14 days postcoitum (dpc). Donor cell phenotype, homing, and chimerism were assessed at short and long-term time points and transplanted animals received skin allografts at 8 weeks.In pretreated allogeneic recipients, early chimerism rates were more than double that of controls (71% vs 33%, p = 0.01). These differences were associated with higher numbers of pretransplant donor cell colony-forming cells without change in donor cell homing. Despite prolonged skin allograft survival for pretreated recipients compared with controls (mean survival = 20.8 vs 8.2 days, p0.001), long-term engraftment was unchanged.These findings demonstrate that higher levels of early chimerism in recipients of cytokine-stimulated marrow result in improved short-term chimerism and tolerance. Future studies are needed to confirm the existence of a "threshold" level of chimerism necessary to sustain long-term engraftment.

Background. The primary barrier to clinical application of in utero hematopoietic stem cell (HSC) transplantation is limited donor cell engraftment. We hypothesized that limited engraftment was due to competition between host and donor cells for available niches. We reasoned that increased engraftment might be achieved by performing multiple transplants separated by brief intervals to allow time for formation of new niches. To test this we performed multiple transplants in a congenic combination to avoid confounding immunologic effects. Materials and methods. C57Pep3B (H2K b , CD45.1) mice were used as donors of adult bone marrow and C57B1/6 (H2K b , CD45.2) mice were used as 14-day-gestation fetal recipients. All fetuses were injected intraperitoneally with 1 × 10 6 mononuclear cells. Boosted neonates were injected at Days 2, 4, and 7 of life with 5 × 10 6 cells. All animals were analyzed for donor cell engraftment by dual-color flow cytometry using CD45 and CD45.1 antigens. Results are reported as the mean ± SD. Statistical analysis was performed using the two-tailed Student t test with P < 0.05 considered significant. Results. Postnatally boosted animals demonstrated significantly elevated levels of donor cell engraftment (3.30 ± 0.8%; n = 8; P < 0.00001) when compared to the control animals (0.69 ± 0.5%; n = 9) as determined by peripheral blood analysis at 6 weeks of age. This elevated level of engraftment was stable long term. Conclusions. Our results demonstrate a significant increase in donor cell engraftment with postnatal booster injections after in utero transplantation. This supports the hypothesis that a limited number of niches may be a major component of the barrier to engraftment. It also suggests that postnatal booster injections may be a viable therapeutic strategy for improving donor cell engraftment after in utero HSC transplantation.

Background/Purpose: Immune responses to both vector and transgene antigens have limited the efficacy of postnatal gene therapy. We hypothesize that the fetal period may offer immunologic and developmental advantages for successful gene therapy. In this study we examined the efficacy, persistence, and immunologic effects of recombinant adenovirus after intramuscular delivery into fetal mice. Methods: E1-deleted adenovirus (AdCMVlacZ) containing the β-galactosidase marker gene was used for injection. Fetal Balb/c mice (14 to 15 days' gestation) were injected with AdCMVlacZ in 10-μL volume in either the shoulder or hindlimb musculature. Animals were killed at 18 to 20 days' gestation and up to 4 months postnatally for analysis of transgene expression and adenoviral genome persistence. Results: Fetuses were injected with doses of AdCMVlacZ from 1 × 10 8 to 2 × 10 10 viral particles (n = 80). Optimal survival rate was 83% at 18 to 20 days' gestation and 55% at 4 weeks of age using a dose of 1 × 10 9 particles. Expression of β-galactosidasae at 18 to 20 days localized to multiple muscle groups surrounding the site of injection, as well as bone marrow stroma, liver, lung, and dorsal root ganglia. Persistent muscle and liver transgene expression was observed for as long as 16 and 8 weeks, respectively, after injection. The pattern of liver expression was confined to discrete foci of hepatocytes, which appeared to increase in size in older animals. No histological evidence of muscle or liver inflammation was observed at any time after injection. No neutralizing antibodies were observed postnatally. Conclusions: Our results confirm that gene therapy in the fetus may be advantageous. Distribution of vector in the fetus at the site of injection is clearly broader than in the adult setting. Furthermore, the absence of immune response and persistence of transgene expression suggests that fetal exposure to foreign transgene and vector antigens may induce tolerance. Although we have not proven genomic integration, the histological appearance of transgene expression in the liver supports this conclusion. By understanding the mechanisms that underlie persistent transgene expression, fetal gene therapy may become a feasible strategy for the treatment of fatal genetic diseases.

Aortopexy is the therapeutic modality of choice for severe primary tracheomalacia. The thoracoscopic approach has been used with good results in infants and toddlers, but little information exists on the use of aortopexy in older children. We present the case of a boy with a lifelong history of refractory, a steroid-dependent reactive airway disease, and who was found to have severe primary tracheomalacia. He subsequently underwent thoracoscopic aortopexy with immediate resolution of the tracheomalacia as demonstrated by serial bronchoscopy and long-term resolution of his clinical respiratory symptoms at 1 year.

Donor-specific tolerance has been induced after both fetal and neonatal hematopoietic stem cell (HSC) transplantation in mice. However, the relationship between hematopoietic microchimerism and tolerance in these models has not been defined due to the insensitivity of donor cell detection methodology. To address this problem we developed a semiquantitative polymerase chain reaction (PCR)-based assay for detection of microchimerism after major histocompatibility (MHC) class I disparate HSC transplantation. This assay was used to examine the relationship between microchimerism and tolerance after fetal and neonatal transplantation of fully allogeneic bone marrow cells.C57BL/6 mice (H2-Kb) were used as adult bone marrow donors and Balb/c mice (H2-Kd) were used as fetal or newborn recipients. A dose of 10(10) BM cells/kg was injected intraperitoneally into recipient animals. Peripheral blood of animals which survived beyond 3 weeks of age was analyzed by PCR for the presence of donor MHC class I DNA. Tolerance was tested by placement of donor-specific skin grafts after determination of chimerism status.Our assay was found to be specific for H2-Kb donor cells in an H2-Kd background with a sensitivity of0.0001%. Of 49 animals injected in utero 19 (38%) had donor DNA present in peripheral blood at low levels (0.1%) whereas only 1 of 18 neonatally injected animals had detectable donor cells (P0.01). Tolerance to donor-specific skin grafts was found in 6 of 9 animals which were chimeric after in utero HSC transplantation whereas none of the 18 neonatally injected animals including the chimeric animal were tolerant.Our results indicate the following. (CITEREF RID="JR975255RF1"1) Hematopoietic microchimerism can be detected by PCR in peripheral blood after in utero injection of fully allogeneic HSCs. (CITEREF RID="JR975255RF2"2) Fetal injections yield a higher incidence of microchimerism than newborn injections. (CITEREF RID="JR975255RF3"3) Tolerance can be induced across a fully allogeneic barrier by in utero HSC transplantation and this is associated with the presence of peripheral blood microchimerism.