Brief Resolved Unexplained Events Symptoms Frequently Result in Inappropriate Gastrointestinal Diagnoses and Treatment.

OBJECTIVE: To determine associations between presenting symptoms and oropharyngeal dysphagia diagnoses, gastroesophageal reflux disease (GERD) diagnoses, and treatment with acid suppression medication in infants with brief resolved unexplained event (BRUE). STUDY DESIGN: We performed a prospective cohort study of infants with BRUE to review presenting symptoms and their potential impact on testing and treatment. Videofluoroscopic swallow study (VFSS) results and explanatory diagnoses were obtained from medical record review; acid suppression use was determined by parental survey. Binary and multivariable logistic regression models were used to evaluate associations between presenting symptoms and obtaining VFSS, VFSS results, GERD diagnoses, and acid suppression medication. RESULTS: Presenting symptoms were varied in 157 subjects enrolled at 51.0 ± 5.3 days of age, with many symptoms that may be related to GERD or dysphagia. Of these, 28% underwent VFSS with 71% abnormal. Overall, 42% had their BRUE attributed to GERD, and 33% were treated with acid suppression during follow-up. Presenting symptoms were significantly associated with the decision to obtain VFSS but not with abnormal VFSS results. Presenting symptoms were also associated with provision of GERD explanatory diagnoses. Both presenting symptoms and GERD explanatory diagnoses were associated with acid suppression use (aOR 2.3, 95% CI 1.03-5.3, P = .04). CONCLUSIONS: Presenting symptoms may play a role in clinicians' decisions on which BRUE patients undergo VFSS but are unreliable to make a diagnosis of oropharyngeal dysphagia. Presenting symptoms may also influence assignment of GERD explanatory diagnoses that is associated with increased acid suppression medication use.

Breastfeeding in infants who aspirate may increase risk of pulmonary inflammation.

OBJECTIVE: To evaluate management strategies and pulmonary outcomes for breastfed infants with oropharyngeal dysphagia. STUDY DESIGN: We performed a retrospective cohort study of breastfed infants diagnosed with oropharyngeal dysphagia with documented aspiration or laryngeal penetration on videofluoroscopic swallow study (VFSS). Medical records were reviewed for VFSS results and speech-language pathologist recommendations following VFSS, results of chest x-ray, results of bronchoalveolar lavage (BAL) within 1 year of VFSS, and aspiration-related hospitalizations occurring before or within 1 year of VFSS. Subjects were categorized as cleared or not cleared to breastfeed based on the VFSS. Proportions were compared with Chi-square and Fisher's exact tests and means with Student's t-tests. RESULTS: Seventy-six infants (4.7 ± 0.4 months old) were included; 50% (38) had aspiration and 50% (38) had laryngeal penetration. After VFSS, 70% (53) were cleared to breastfeed while 30% (23) were not cleared to breastfeed. Patients with aspiration were less likely to be cleared to breastfeed (p = .006); however, 55% (21/38) of those with aspiration were still cleared to breastfeed. Infants cleared to breastfeed had significantly more pulmonary hospitalizations (p = .04) and were also at increased risk of elevated neutrophil count (p = .02) and culture growth on BAL (p = .01). Significantly increased abnormal neutrophil count was also found in those cleared to breastfeed with laryngeal penetration (p = .01). CONCLUSIONS: Infants with oropharyngeal dysphagia counseled to continue breastfeeding had increased risk of BAL inflammation and more pulmonary hospitalizations compared to those that were told to stop breastfeeding.

Pediatric Aerodigestive Medicine: Advancing Collaborative Care for Children With Oropharyngeal Dysphagia.

OBJECTIVES: Aerodigestive disorders encompass various pathological conditions affecting the lungs, upper airway, and gastrointestinal tract in children. While advanced care has primarily occurred in specialty centers, many children first present to general pediatric gastroenterologists with aerodigestive symptoms necessitating awareness of these conditions. At the 2021 Annual North American Society for Pediatric Gastroenterology, Hepatology and Nutrition meeting, the aerodigestive Special Interest Group held a full-day symposium entitled, Pediatric Aerodigestive Medicine: Advancing Collaborative Care of Children with Aerodigestive Disorders. The symposium aimed to underline the significance of a multidisciplinary approach to achieve better outcomes for these complex patients. METHODS: The symposium brought together leading experts to highlight the growing aerodigestive field, promote new scientific and therapeutic strategies, share the structure and benefits of a multidisciplinary approach in diagnosing common and rare aerodigestive disorders, and foster multidisciplinary discussion of complex cases while highlighting the range of therapeutic and diagnostic options. In this article, we showcase the diagnostic and therapeutic approach to oropharyngeal dysphagia (OPD), one of the most common aerodigestive conditions, emphasizing the role of a collaborative model. CONCLUSIONS: The aerodigestive field has made significant progress and continues to grow due to a unique multidisciplinary, collaborative model of care for these conditions. Despite diagnostic and therapeutic challenges, the multidisciplinary approach has enabled and greatly improved efficient, high-quality, and evidence-based care for patients, including those with OPD.

A Prospective Study of Parental Experience with Thickening Feeds for Children with Oropharyngeal Dysphagia and Gastroesophageal Reflux.

Thickening is efficacious and commonly recommended for oropharyngeal dysphagia and gastroesophageal reflux. Little is known about parental experience with this practice. Results of this cross-sectional questionnaire study suggest attitudes are positive, but parents frequently adjust recipes/nipple sizes, which might increase aspiration risk. Clinical follow-up is essential to ensure safe feeding.

Gastrointestinal factors associated with risk of bronchiectasis in children.

OBJECTIVE: To evaluate gastrointestinal (GI) risk factors for bronchiectasis in children. We hypothesized that upper GI tract dysmotility would be associated with increased risk of bronchiectasis. STUDY DESIGN: Subjects in this retrospective cohort study included those evaluated for persistent pulmonary symptoms in the Aerodigestive Center at Boston Children's Hospital who underwent chest computed tomography (CT) between 2002 and 2019. To determine gastrointestinal predictors of bronchiectasis, baseline characteristics, comorbidities, enteral tube status, medications received, gastroesophageal reflux burden, adequacy of swallow function, esophageal dysmotility, gastric dysmotility, and neutrophil count on bronchoalveolar lavage (BAL) were compared between patients with and without bronchiectasis. Proportions were compared with Fisher's exact test and binary logistic regression with stepwise selection was used for multivariate analysis. ROC analyses were utilized to compare BAL neutrophils and bronchiectasis. RESULTS: Of 192 subjects, 24% were found to have evidence of bronchiectasis on chest CT at age 7.9 ± 0.5 years. Enteral tubes (OR 5.77, 95% CI 2.25-14.83, p < 0.001) and increased BAL neutrophil count (OR 5.79, 95% CI 1.87-17.94, p = 0.002) were associated with increased risk while neurologic comorbidities were associated with decreased risk (OR 0.24, 95% CI 0.09-0.66, p = 0.006). Gastroesophageal reflux was not found to be a significant risk factor. Neutrophil counts >10% had 72% sensitivity and 60% specificity for identifying bronchiectasis. CONCLUSIONS: Enteral tubes were associated with significantly increased risk of bronchiectasis but gastroesophageal reflux was not. Providers should consider obtaining chest CT to evaluate for bronchiectasis in children found to have unexplained elevated BAL neutrophil count.

A Prospective Study of Brief Resolved Unexplained Events: Risk Factors for Persistent Symptoms.

OBJECTIVE: The risk of persistent symptoms after a brief resolved unexplained event (BRUE) is not known. Our objective was to determine the frequency and risk factors for persistent symptoms after BRUE hospitalizations. METHODS: We conducted a prospective longitudinal cohort study of infants hospitalized with an admitting diagnosis of BRUE. Caregiver-reported symptoms, anxiety levels, and management changes were obtained by questionnaires during the 2-month follow-up period. Clinical data including repeat hospitalizations were obtained from a medical record review. Multivariable analyses with generalized estimating equations were conducted to determine the risk of persistent symptoms. RESULTS: Of 124 subjects enrolled at 51.6 ± 5.9 days of age, 86% reported symptoms on at least 1 questionnaire after discharge; 65% of patients had choking episodes, 12% had BRUE spells, and 15% required a repeat hospital visit. High anxiety levels were reported by 31% of caregivers. Management changes were common during the follow-up period and included 30% receiving acid suppression and 27% receiving thickened feedings. Only 19% of patients had a videofluoroscopic swallow study while admitted, yet 67% of these studies revealed aspiration/penetration. CONCLUSIONS: Many infants admitted with BRUE have persistent symptoms and continue to access medical care, suggesting current management strategies insufficiently address persistent symptoms. Future randomized trials will be needed to evaluate the potential efficacy of therapies commonly recommended after BRUE.

Acid Suppression Does Not Improve Laryngomalacia Outcomes but Treatment for Oropharyngeal Dysphagia Might Be Protective.

OBJECTIVE: To determine whether the use of acid suppression and thickened feeds impact laryngomalacia outcomes in infants, including supraglottoplasty risk, time to supraglottoplasty, and hospitalization risk. STUDY DESIGN: We performed a retrospective cohort study to compare risk and time with supraglottoplasty and frequency and duration of hospitalizations for infants diagnosed with laryngomalacia at Boston Children's Hospital between January 1 and December 31, 2017. The primary outcomes were supraglottoplasty requirement, time to supraglottoplasty, and hospitalization risk. Multivariate analyses were performed to determine predictors of supraglottoplasty and hospitalization risk after adjusting for laryngomalacia severity and comorbidities in addition to propensity score adjustment. Kaplan-Meier curves were created to determine the impact of acid suppression use on time to supraglottoplasty. RESULTS: In total, 236 subjects with mean age 62.6 ± 4 days were included in the analysis; 55% were treated with acid suppression. Subjects treated with acid suppression had a greater risk of supraglottoplasty (hazard ratio 3.36, 95% CI 1.36-8.29, P = .009), shorter time to supraglottoplasty (5.64 ± 0.92 vs 7.98 ± 1.92 months, P = .006), and increased respiratory hospitalization risk (relative risk 1.97, 95% CI 1.01-3.85, 0.047), even after adjustment for covariates. Subjects receiving thickening had fewer respiratory hospitalization nights and longer time to supraglottoplasty (9.3 ± 1.7 vs 4.56 ± 0.73 months, P = .004), even after adjustment. CONCLUSIONS: Acid suppression use does not reduce the frequency of supraglottoplasty and related hospitalizations compared with untreated subjects. However, patients treated with thickening have decreased hospitalization and longer time to supraglottoplasty, suggesting that thickening of feeds may be a preferred intervention over acid suppression.

Overlapping Symptoms of Gastroesophageal Reflux and Aspiration Highlight the Limitations of Validated Questionnaires.

OBJECTIVES: Infants frequently present with feeding difficulties and respiratory symptoms, which are often attributed to gastroesophageal reflux but may be because of oropharyngeal dysphagia with aspiration. The Infant Gastroesophageal Reflux Questionnaire Revised (I-GERQ-R) is a clinical measure of gastroesophageal reflux disease but now there is greater understanding of dysphagia as a reflux mimic. We aimed to determine the degree of overlap between I-GERQ-R and evidence of dysphagia, measured by Pediatric Eating Assessment Tool-10 (Pedi-EAT-10) and videofluoroscopic swallow study (VFSS). METHODS: We performed a prospective study of subjects <18 months old with feeding difficulties. All parents completed Pedi-EAT-10 and I-GERQ-R as a quality initiative to address parental feeding concerns. I-GERQ-R results were compared with Pedi-EAT-10 and, whenever available, results of prior VFSS. Pearson correlation coefficients were calculated to determine the relationship between scores. Groups were compared with 1-way ANOVA and Fisher exact test. ROC analysis was completed to compare scores with VFSS results. RESULTS: One hundred eight subjects with mean age 7.1 ±â€Š0.5 months were included. Pedi-EAT-10 and I-GERQ-R were correlated (r = 0.218, P = 0.023) in all subjects and highly correlated in the 77 subjects who had prior VFSS (r = 0.369, P = 0.001). The blue spell questions on I-GERQ-R had relative risk 1.148 (95% confidence interval [CI] 1.043-1.264, P = 0.142) for predicting aspiration/penetration on VFSS, with 100% specificity. Scores on the question regarding crying during/after feedings were also higher in subjects with abnormal VFSS (1.1 ±â€Š0.15 vs 0.53 ±â€Š0.22, P = 0.04). CONCLUSIONS: I-GERQ-R and the Pedi-EAT-10 are highly correlated. I-GERQ-R results may actually reflect oropharyngeal dysphagia and not just gastroesophageal reflux disease in infants.

Clinical Aspects of Thickeners for Pediatric Gastroesophageal Reflux and Oropharyngeal Dysphagia.

PURPOSE OF REVIEW: The purpose of this review is to discuss current knowledge and recent findings regarding clinical aspects of thickeners for pediatric gastroesophageal reflux and oropharyngeal dysphagia. We review evidence for thickener efficacy, discuss types of thickeners, practical considerations when using various thickeners, and risks and benefits of thickener use in pediatrics. RECENT FINDINGS: Thickeners are effective in decreasing regurgitation and improving swallowing mechanics and can often be used empirically for the treatment of infants and young children. Adverse effects have been reported, but with careful consideration of appropriate thickener types, desired thickening consistency, and follow-up in collaboration with feeding specialists, most patients have symptomatic improvements. Thickeners are typically well tolerated and with few side effects, but close follow-up is needed to make sure patients tolerate thickeners and have adequate symptom improvement.

The Impact of the American Academy of Pediatrics Brief Resolved Unexplained Event Guidelines on Gastrointestinal Testing and Prescribing Practices.

OBJECTIVES: To determine if hospitalization, testing, diagnosis, and management of suspected gastroesophageal reflux, and follow-up visits decreased since introduction of American Academy of Pediatrics guidelines for brief resolved unexplained events (BRUEs). STUDY DESIGN: We performed a retrospective cohort study of infants with BRUE evaluated at Boston Children's Hospital in the year before and after guideline implementation to determine if practice patterns have changed. Outcomes included hospitalization rates, frequency of swallow assessments, other diagnostic testing, and reflux diagnoses, cost of care, and number of repeat visits. Groups were compared based on whether they presented before or after guideline implementation. RESULTS: In total, 359 subjects (186 pre-, 173 post-guidelines) were identified. There were no significant differences in practice patterns or outcomes before or after guideline implementation. Subjects had mean age 2.53 ± 0.15 months, and 80% were hospitalized for 2.49 ± 0.26 days. Each subject had 2.47 diagnostic tests performed, and 89% were noncontributory. Despite only 13% having videofluoroscopic swallow study performed, 72% showed aspiration/penetration. No subject had gastroesophageal reflux testing, yet reflux was implicated as the cause for admission in 40% of subjects, resulting in increased odds of discharge on acid suppressing medications (OR 2.88, 95% CI 1.68-4.92, P = .0001). In follow-up, 28% of subjects had repeat hospitalizations or emergency department visits for persistent symptoms. CONCLUSIONS: Infants with BRUE continue to undergo low-yield diagnostic testing and after admission remain symptomatic and frequently re-present to medical care. Swallow testing remains infrequent despite its high-yield, reflux continues to be implicated and children are still being discharged on acid suppression despite lack of efficacy.

Feeding Interventions Are Associated With Improved Outcomes in Children With Laryngeal Penetration.

OBJECTIVES: To determine if children with laryngeal penetration on videofluoroscopic swallow study (VFSS) who received feeding interventions (thickened liquids, change in liquid flow rate, and/or method of liquid delivery) had improved symptoms and decreased hospitalizations compared with those without intervention. METHODS: We performed a retrospective cohort study of children under 2 years with laryngeal penetration on VFSS at our institution in 2015 to determine initial and follow-up VFSS findings, symptom improvement at follow-up, and hospitalization risk before and after VFSS. Proportions were compared with Fisher exact test and hospitalizations with paired t tests. RESULTS: We evaluated 137 subjects with age 8.93 ±â€Š0.59 months who had laryngeal penetration without aspiration on VFSS. Fifty-five percent had change in management, with 40% receiving thickening and 15% a change in flow rate. There was significant improvement in symptoms for children that had feeding intervention and this improvement was the greatest with thickening (OR 41.8, 95% CI 12.34-141.69, P < 0.001). On repeat VFSS, 26% had evidence of aspiration that was not captured on initial VFSS. Subjects had decreased total and pulmonary hospitalizations with feeding intervention and decreased pulmonary nights with thickening (P < 0.05). CONCLUSIONS: Laryngeal penetration appears to be clinically significant in children with oropharyngeal dysphagia and interventions to decrease its occurrence are associated with improved outcomes including decreased symptoms of concern and hospitalization nights. Thickening or other feeding intervention should be considered for all symptomatic children with laryngeal penetration on swallow study.

Association of Proton Pump Inhibitors With Hospitalization Risk in Children With Oropharyngeal Dysphagia.

Importance: Proton pump inhibitors (PPI) are commonly prescribed to children with oropharyngeal dysphagia and resultant aspiration based on the assumption that these patients are at greater risk for reflux-related lung disease. There is little data to support this approach and the potential risk for increased infections in children treated with PPI may outweigh any potential benefit. Objective: The aim of this study was to determine if there is an association between hospitalization risk in pediatric patients with oropharyngeal dysphagia and treatment with PPI. Design, Setting, and Participants: We performed a retrospective cohort study to compare the frequency and length of hospitalizations for children who had abnormal results on videofluoroscopic swallow studies that were performed between January 1, 2015, and December 31, 2015, and who were or were not treated with PPI, with follow-up through December 31, 2016. Records were reviewed for children who presented for care at Boston Children's Hospital, a tertiary referral center. Participants included 293 children 2 years and younger with evidence of aspiration or penetration on videofluoroscopic swallow study. Exposures: Groups were compared based on their exposure to PPI treatment. Main Outcomes and Measures: The primary outcomes were hospital admission rate and hospital admission nights and these were measured as incident rates. Multivariable analyses were performed to determine predictors of hospitalization risk after adjusting for comorbidities. Kaplan-Meier curves were created to determine the association of PPI prescribing with time until first hospitalization. Results: A total of 293 patients with a mean (SD) age of 8.8 (0.4) months and a mean (SD) follow-up time of 18.15 (0.20) months were included in the analysis. Patients treated with PPI had higher admission rates (Incidence rate ratio [IRR], 1.77; 95% CI, 1.16-2.68) and admission nights (IRR, 2.51; 95% CI, 1.36-4.62) even after adjustment for comorbidities. Patients with enteral tubes who were prescribed PPIs were at the highest risk for admission (hazard ratio [HR], 2.31; 95% CI, 1.24-4.31). Conclusions and Relevance: Children with aspiration who are treated with PPI have increased risk of hospitalization compared with untreated patients. These results support growing concern about the risks of PPI use in children.

Presenting Signs and Symptoms do not Predict Aspiration Risk in Children.

OBJECTIVES: To determine if any presenting symptoms are associated with aspiration risk, and to evaluate the reliability of clinical feeding evaluation (CFE) in diagnosing aspiration compared with videofluoroscopic swallow study (VFSS). STUDY DESIGN: We retrospectively reviewed records of children under 2 years of age who had evaluation for oropharyngeal dysphagia by CFE and VFSS at Boston Children's Hospital and compared presenting symptoms, symptom timing, and CFE and VFSS results. We investigated the relationship between symptom presence and aspiration using the Fisher exact test and stepwise logistic regression with adjustment for comorbidities. CFE and VFSS results were compared using the McNemar test. Intervals from CFE to VFSS were compared using the Student t test. RESULTS: A total of 412 subjects with mean (±SD) age 8.9 ± 6.9 months were evaluated. No symptom, including timing relative to meals, predicted aspiration on VFSS. This lack of association between symptoms and VFSS results persisted even in the adjusted multivariate model. The sensitivity of CFE for predicting aspiration by VFSS was 44%. Patients with a reassuring CFE waited 28.2 ± 8.5 days longer for confirmatory VFSS compared with those with a concerning CFE (P < .05). CONCLUSIONS: Presenting symptoms are varied in patients with aspiration and cannot be relied upon to determine which patients have aspiration on VFSS. The CFE does not have the sensitivity to consistently diagnose aspiration so a VFSS should be performed in persistently symptomatic patients.

Oropharyngeal Dysphagia Is Strongly Correlated With Apparent Life-Threatening Events.

OBJECTIVES: The aim of the present study was to investigate the prevalence of oropharyngeal dysfunction with resultant aspiration in patients admitted after apparent life-threatening events (ALTE) and to determine whether historical characteristics could predict this oropharyngeal dysphagia and aspiration risk. METHODS: We retrospectively reviewed the records of all patients admitted to Boston Children's Hospital between 2012 and 2015 with a diagnosis of ALTE to determine the frequency of evaluation for oropharyngeal dysphagia using video fluoroscopic swallow studies (VFSS) and clinical feeding evaluations, to determine the prevalence of swallowing dysfunction in subjects admitted after ALTE and to compare presenting historical characteristics to swallow study results. RESULTS: A total of 188 children were admitted with a diagnosis of ALTE of which 29% (n = 55) had an assessment of swallowing by VFSS. Of those who had a VFSS, 73% (n = 40) had evidence of aspiration or penetration on VFSS. Of all of the diagnostic tests ordered on patients with ALTEs, the VFSS had the highest rate of abnormalities of any test ordered. None of the historical characteristics of ALTE predicted which patients were at risk for aspiration. In patients who had both clinical feeding evaluations and VFSS, observed clinical feedings incorrectly identified 26% of patients as having no oropharyngeal dysphagia when in fact aspiration was present on VFSS. CONCLUSIONS: Oropharyngeal dysphagia with aspiration is the most common diagnosis identified in infants presenting with ALTEs. The algorithm for ALTE should be revised to include an assessment of VFSS as clinical feeding evaluations are inadequate to assess for aspiration.

Gastroesophageal Reflux Burden, Even in Children That Aspirate, Does Not Increase Pediatric Hospitalization.

OBJECTIVES: Gastroesophageal reflux is common but remains a controversial disease to diagnose and treat and little is known about the role of reflux testing in predicting clinical outcomes, particularly in children at risk for extraesophageal reflux complications. The aim of this study was to determine if rates of hospitalization were affected by reflux burden even after adjusting for aspiration risk. METHODS: We prospectively recruited, between 2009 and 2014, a cohort of pediatric patients with suspected extraesophageal reflux disease who were referred for reflux testing and underwent both multichannel intraluminal impedance with pH (pH-MII) and modified barium swallow studies. A subset of patients also underwent bronchoalveolar lavage with pepsin analysis. We determined their rates of hospitalization for a minimum of 1 year following pH-MII testing. RESULTS: We prospectively enrolled 116 pediatric patients who presented for care at Boston Children's Hospital and underwent both pH-MII and modified barium swallow studies. There was no statistically significant relationship between reflux burden measured by pH-MII or bronchoalveolar pepsin and total number of admissions or number of admission nights even after adjusting for aspiration status (P > 0.2). There were no statistically significant relationships between reflux burden by any method and the number or nights of urgent pulmonary admissions before or after adjusting for aspiration risk (P > 0.08). CONCLUSIONS: Even in aspirating children, reflux burden did not increase the risk of hospitalization. Based on these results, routine reflux testing cannot be recommended even in aspirating children, because the results do not impact clinically significant outcomes.

Evaluation of the use of an induced puripotent stem cell sheet for the construction of tissue-engineered vascular grafts.

OBJECTIVE: The development of a living, tissue-engineered vascular graft (TEVG) holds great promise for advancing the field of cardiovascular surgery. However, the ultimate source and time needed to procure these cells remain problematic. Induced puripotent stem (iPS) cells have recently been developed and have the potential for creating a pluripotent cell line from a patient's own somatic cells. In the present study, we evaluated the use of a sheet created from iPS cell-derived vascular cells as a potential source for the construction of TEVG. METHODS: Male mouse iPS cells were differentiated into embryoid bodies using the hanging-drop method. Cell differentiation was confirmed by a decrease in the proportion of SSEA-1-positive cells over time using fluorescence-activated cell sorting. The expression of endothelial cell and smooth muscle cell markers was detected using real-time polymerase chain reaction (PCR). The differentiated iPS cell sheet was made using temperature-responsive dishes and then seeded onto a biodegradable scaffold composed of polyglycolic acid-poly-l-lactide and poly(l-lactide-co-ε-caprolactone) with a diameter of 0.8 mm. These scaffolds were implanted as interposition grafts in the inferior vena cava of female severe combined immunodeficiency/beige mice (n = 15). Graft function was serially monitored using ultrasonography. The grafts were analyzed at 1, 4, and 10 weeks with histologic examination and immunohistochemistry. The behavior of seeded differentiated iPS cells was tracked using Y-chromosome fluorescent in situ hybridization and SRY real-time PCR. RESULTS: All mice survived without thrombosis, aneurysm formation, graft rupture, or calcification. PCR evaluation of iPS cell sheets in vitro demonstrated increased expression of endothelial cell markers. Histologic evaluation of the grafts demonstrated endothelialization with von Willebrand factor and an inner layer with smooth muscle actin- and calponin-positive cells at 10 weeks. The number of seeded differentiated iPS cells was found to decrease over time using real-time PCR (42.2% at 1 week, 10.4% at 4 weeks, 9.8% at 10 weeks). A fraction of the iPS cells were found to be Y-chromosome fluorescent positive at 1 week. No iPS cells were found to co-localize with von Willebrand factor or smooth muscle actin-positive cells at 10 weeks. CONCLUSIONS: Differentiated iPS cells offer an alternative cell source for constructing TEVG. Seeded iPS cells exerted a paracrine effect to induce neotissue formation in the acute phase and were reduced in number by apoptosis at later time points. Sheet seeding of our TEVG represents a viable mode of iPS cell delivery over time.