Modulation of LIN28B/Let-7 Signaling by Propranolol Contributes to Infantile Hemangioma Involution.

OBJECTIVE: Infantile hemangiomas (IHs) are the most common benign vascular neoplasms of infancy, characterized by a rapid growth phase followed by a spontaneous involution, or triggered by propranolol treatment by poorly understood mechanisms. LIN28/let-7 axis plays a central role in the regulation of stem cell self-renewal and tumorigenesis. However, the role of LIN28B/let-7 signaling in IH pathogenesis has not yet been elucidated. APPROACH AND RESULTS: LIN28B is highly expressed in proliferative IH and is less expressed in involuted and in propranolol-treated IH samples as measured by immunofluorescence staining and quantitative RT-PCR. Small RNA sequencing analysis of IH samples revealed a decrease in microRNAs that target LIN28B, including let-7, and an increase in microRNAs in the mir-498(46) cistron. Overexpression of LIN28B in HEK293 cells induced the expression of miR-516b in the mir-498(46) cistron. Propranolol treatment of induced pluripotent stem cells, which express mir-498(46) endogenously, reduced the expression of both LIN28B and mir-498(46) and increased the expression of let-7. Furthermore, propranolol treatment reduced the proliferation of induced pluripotent stem cells and induced epithelial-mesenchymal transition. CONCLUSIONS: This work uncovers the role of the LIN28B/let-7 switch in IH pathogenesis and provides a novel mechanism by which propranolol induces IH involution. Furthermore, it provides therapeutic implications for cancers in which the LIN28/let-7 pathway is imbalanced.

Growth Hormone Induces Recurrence of Infantile Hemangiomas After Apparent Involution: Evidence of Growth Hormone Receptors in Infantile Hemangioma.

Infantile hemangiomas (IHs) are the most common benign tumor of infancy, characterized by a natural history of early proliferation in the first months of life to eventual involution during childhood, often with residual fibrofatty tissue. Once involution has been achieved, IHs do not typically recur. We present two cases of exogenous growth hormone therapy resulting in the recurrence of IHs in late childhood, supported by radiological, immunohistochemical, in vitro, and in vivo evidence.

Clinical spectrum of capillary malformation-arteriovenous malformation syndrome presenting to a pediatric dermatology practice: a retrospective study.

Capillary malformation-arteriovenous malformation syndrome (CM-AVM) is an autosomal dominant disorder caused by RASA1 mutations. The prevalence and phenotypic spectrum are unknown. Evaluation of patients with multiple CMs is challenging because associated AVMs can be life threatening. The objective of this study was to describe the clinical characteristics of children presenting with features of CM-AVM to an academic pediatric dermatology practice. After institutional review board approval was received, a retrospective chart review was performed of patients presenting between 2009 and 2012 with features of CM-AVM. We report nine cases. Presenting symptoms ranged from extensive vascular stains and cardiac failure to CMs noted incidentally during routine skin examination. All demonstrated multiple CMs, two had Parkes Weber syndrome, and two had multiple infantile hemangiomas. Seven patients had family histories of multiple CMs; three had family histories of large, atypical CMs. Six had personal or family histories of AVMs. Genetic evaluation was recommended for all and was pursued by six families; four RASA1 mutations were identified, including one de novo. Consultations with neurology, cardiology, and orthopedics were recommended. Most patients (89%) have not required treatment to date. CM-AVM is an underrecognized condition with a wide clinical spectrum that often presents in childhood. Further evaluation may be indicated in patients with multiple CMs. This study is limited by its small and retrospective nature.

Propranolol promotes accelerated and dysregulated adipogenesis in hemangioma stem cells.

BACKGROUND: Infantile hemangiomas (IHs) are the most common tumor of infancy, yet there are no Food and Drug Administration-approved therapeutics to date. Recently, the nonselective ß-adrenergic-blocker propranolol has been shown to be a safe and effective means of treating IHs, although its mechanism has yet to be elucidated. We have previously demonstrated that propranolol induces early and incomplete adipogenesis in stem cells derived from hemangiomas. We hypothesize that propranolol promotes dysregulated adipogenesis via the improper regulation of adipogenic genes. METHODS: Hemangioma stem cells (HemSCs) isolated from resected IH specimens were treated with adipogenic medium for 1 or 4 days in either propranolol or vehicle. Cell death was measured by the incorporation of annexin V and propidium iodide by flow cytometry. Adipogenesis was assessed by visualizing lipid droplet formation by Oil Red O staining. Proadipogenic genes C/EBPα, C/EBPß, C/EBPδ, PPARδ, PPARγ, RXRα, and RXRγ were analyzed by quantitative reverse transcription and polymerase chain reaction. RESULTS: Hemangioma stem cells treated with propranolol increased lipid droplet formation compared to vehicle-treated cells indicating increased adipogenesis. Cell death as measured by FACS analysis indicated that the propranolol-treated cells died due to necrosis and not apoptosis. During adipogenesis, transcript levels of PPARδ, PPARγ, C/EBPß, and C/EBPδ were significantly increased (P<0.01) in propranolol-treated cells relative to control cells. In contrast, RXRα and RXRγ levels were significantly decreased (P<0.05), and C/EBPα, a gene required for terminal adipocyte differentiation, was strongly suppressed by propranolol when compared to vehicle-treated cells (P<0.01). CONCLUSIONS: In HemSCs, propranolol accelerated dysregulated adipogenic differentiation characterized by improper adipogenic gene expression. Consistent with accelerated adipogenesis, propranolol significantly increased the expression of the proadipogenic genes, PPARγ, C/EBPß, and C/EBPγ compared to control. However, propranolol treatment also led to improper induction of PPARδ and suppression of C/EBPα, RXRα, and RXRγ. Taken together these data indicate that propranolol promoted dysregulated adipogenesis and inhibited the HemSCs from becoming functional adipocytes, ultimately resulting in cell death. Understanding this mechanism behind propranolol's effectiveness will be a vital factor in producing more effective therapies in the future.

Infantile Hemangiomas Lose Vascular Endothelial Cadherin During Involution: Potential Role in Cell Death?

Background: Infantile hemangiomas (IHs) are benign endothelial cell (EC) tumors that undergo a predictable natural history, with rapid proliferation, stabilization, and involution. However, mechanisms regulating these transitions are not well understood. We have observed loss of vascular endothelial cadherin (VECAD) in involuting/involuted IHs. VECAD plays a critical role in angiogenesis, cell cycle progression, and EC survival. We hypothesize that loss of VECAD is associated with apoptosis occurring during IH involution. Methods: Resected IH samples were clinically categorized as proliferating (n = 4), stable (n = 4), or involuting/involuted (n = 5). Neonatal dermal tissues were used as controls (n = 5). Immunohistochemistry was conducted on sectioned specimens using antibodies against EC markers VECAD and CD31. Apoptosis was assessed with terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling assay. Results: CD31 signal intensity in proliferating, stable, and involuting/involuted IH ECs was unchanged relative to each other and to control ECs. VECAD signal significantly and progressively diminished as IHs progressed from proliferation to involution. Involuting/involuted IHs had significantly reduced VECAD expression compared with control ECs (P < 0.0001), proliferating IHs (P < 0.0001), and stable IHs (P < 0.001). As expected, the number of terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling-positive ECs was significantly higher in involuting/involuted IHs (P < 0.05) relative to control ECs and proliferating IHs. Conclusions: Loss of VECAD expression in IH endothelium corresponded to IH involution and increased apoptosis. It is unclear whether loss of VECAD is causative of IH involution; further studies are needed to elucidate the role of VECAD function in EC survival.

Trends in orthoplastic operative exposure for plastic surgery residents in the United States.

BACKGROUND: Recent trials have demonstrated clinical benefits to a combined orthoplastic approach for complex reconstructive surgery of the hand, upper and lower extremity. PURPOSE: We sought to assess recent trends in exposure to orthoplastic-type procedures among plastic surgery residents training in the United States. METHODS: Independent plastic surgery residents' case logs were extracted from the Accreditation Council for Graduate Medical Education (2011-2022). Select reconstructive procedure were taken as proxies for orthoplastic-type cases and analyzed by descriptive statistical analysis. RESULTS: The average number of orthoplastic-type cases completed per resident per year increased from 168.2 to 189.2 (12.5% increase) between 2011-2022. The greatest increase was in exposure to peripheral nerve injury repair of the hand and upper extremity (22.6 to 39.1, 73% increase). As a proportion of total procedures during the study period, orthoplastic-type procedures remained relatively unchanged (range 9.5-10.4%). CONCLUSIONS: Our findings suggest that plastic surgery residents may be increasingly well-prepared to contribute to orthoplastic care during and following their training. The steady proportion of cases that orthoplastic-type procedures represented over the study period suggests the increase in relevant orthoplastic case volume may be incidental and secondary to an overall rise among all procedures. Given evidence of the benefits of an orthoplastic approach, we recommend consideration of explicit benchmarks for orthoplastic training among plastic surgery residents.

Congenital cutaneous hemangioma causing cardiac failure: a case report and review of the literature.

We present a case of a large congenital hemangioma (CH) on the neck causing cardiac failure and thrombocytopenia in a female neonate. A trial of medical therapy with corticosteroids and propranolol was attempted, but the patient ultimately underwent definitive treatment with embolization and surgical resection with a positive outcome. A review of the English language literature revealed 16 previously reported cases of CHs complicated by congestive heart failure. This series supports known demographic features of CHs, including a lack of gender discrepancy and a predilection to affect the head and neck. These CHs are rarely diagnosed in utero; most patients present with a mass at birth. Cardiac failure is identified prenatally or in the first days of life. A mild to moderate thrombocytopenia and coagulopathy, which is likely transient and distinct from classic Kasabach-Merritt phenomenon, accompanies many of these cases. There is a 30% associated mortality rate. Both medical and interventional treatment modalities have been reported. Steroids are the most commonly used medication, but without any clear benefit. We hypothesize that, based on its possible mechanisms of action,propranolol may be a more effective treatment for CHs requiring treatment. As surgical intervention may be necessary, we recommend a multidisciplinary approach to treating patients with problematic CHs.

Pathophysiology of Slow-Flow Vascular Malformations: Current Understanding and Unanswered Questions.

Background: Slow-flow vascular malformations include venous, lymphatic, and lymphaticovenous malformations. Recent studies have linked genetic variants hyperactivating either the PI3K/AKT/mTOR and/or RAS/RAF/MAPK signaling pathways with slow-flow vascular malformation development, leading to the use of pharmacotherapies such as sirolimus and alpelisib. It is important that clinicians understand basic and translational research advances in slow-flow vascular malformations. Methods: A literature review of basic science publications in slow-flow vascular malformations was performed on Pubmed, using search terms "venous malformation," "lymphatic malformation," "lymphaticovenous malformation," "genetic variant," "genetic mutation," "endothelial cells," and "animal model." Relevant publications were reviewed and summarized. Results: The study of patient tissues and the use of primary pathogenic endothelial cells from vascular malformations shed light on their pathological behaviors, such as endothelial cell hyperproliferation and disruptions in vessel architecture. The use of xenograft and transgenic animal models confirmed the pathogenicity of genetic variants and allowed for preclinical testing of potential therapies. These discoveries underscore the importance of basic and translational research in understanding the pathophysiology of vascular malformations, which will allow for the development of improved biologically targeted treatments. Conclusion: Despite basic and translation advances, a cure for slow-flow vascular malformations remains elusive. Many questions remain unanswered, including how genotype variants result in phenotypes, and genotype-phenotype heterogeneity. Continued research into venous and lymphatic malformation pathobiology is critical in understanding the mechanisms by which genetic variants contribute to vascular malformation phenotypic features.

Propranolol Therapy for Congenital Chylothorax.

Congenital chylothorax is a rare and often severe anomaly without well-established medical therapies. Previously, propranolol use in patients with lymphatic malformations and secondary chylothorax was associated with improvement in clinical signs. We hypothesized that propranolol treatment would be beneficial for severe congenital chylothorax. We reviewed medical records of neonates born from 2015 to 2019 at our tertiary center with a prenatal diagnosis of congenital chylothorax for whom either prenatal or postnatal propranolol therapy was initiated. Inclusion was limited to fetuses diagnosed with severe congenital chylothorax without significant genetic, infectious, or cardiac anomalies, and who underwent prenatal interventions to mitigate consequences of the condition. Propranolol was administered orally to pregnant women at 20 mg 4 times daily and increased to a maximum dose of 40 mg 4 times daily, or to infants at 0.3 mg/kg/d and increased to 1 to 2 mg/kg/d. Primary outcomes were the time course of resolution of ultrasonographical, clinical, and/or radiologic signs of chylothorax after treatment with propranolol. Four neonates met the inclusion criteria. In 2 cases, prenatal initiation of propranolol led to resolution of the chylothoraxes before delivery (38 and 32 days after treatment) on a dose of 40 mg/day 4 times daily. Neonates had a normal postnatal course. Postnatal propranolol was initiated in 2 neonates with respiratory failure when chylothoraces were refractory to standard management. Stabilization and improvement of their pleural effusion was observed by imaging at 29 and 13 days after initiation of propranolol. There were no significant maternal or neonatal complications from prenatal or postnatal propranolol use. Propranolol may be efficacious in treating severe fetal congenital chylothorax.

Utilization of Healthcare Resources by Vascular Anomaly Patients: An Assessment of Healthcare Burden by Lesion Complexity.

Background: Vascular anomalies (VAs) are heterogeneous lesions. Symptoms vary widely by lesion type and complexity. VA patients often require life-long interdisciplinary care; however, there is a paucity of data on the healthcare utilization of VA patients, and their burden on the healthcare system remains largely unquantified. We hypothesize that healthcare utilization by complex lymphatic malformation (LM) and venous malformation (VM) patients will be significantly higher compared with simple LM and VM patients. Methods: A retrospective, longitudinal study was performed of LM/VM patients seen through multidisciplinary VA clinics between January 1, 2019 and December 31, 2020. Data were collected from each patient's first presentation through December 31, 2021 and included number of office visits, imaging studies, specialists involved, procedures, hospitalization data, and approximate costs, normalized to per year utilization. Patients were divided into "simple" and "complex" LMs/VMs. Involvement of the airway, more than one anatomic area, and/or complex lymphatic anomalies were defined as "complex." Results: In total, 28 simple and 29 complex LM patients and 51 simple and 18 complex VM patients were identified. Complex LM and VM patients had significantly higher numbers of imaging studies, specialists involved, procedures and hospitalizations, and costs incurred. Complex LM patients also had significantly higher per year office visits. Conclusions: VA care is chronic and costly, especially for complex LM/VM patients. LM/VM complexity was a predictor for increased inpatient and outpatient healthcare utilization and higher costs. Better awareness of the healthcare utilization trends of LM/VM patients will allow for improved counseling for these patients regarding prognosis and expectations.

JAGGED1 signaling regulates hemangioma stem cell-to-pericyte/vascular smooth muscle cell differentiation.

OBJECTIVE: The aim of our study is to determine the cellular and molecular origin for the pericytes in infantile hemangioma (IH) and their functional role in the formation of pathological blood vessels. METHODS AND RESULTS: Here we show that IH-derived stem cells (HemSCs) form pericyte-like cells. With in vitro studies, we demonstrate that HemSC-to-pericyte differentiation depends on direct contact with endothelial cells. JAGGED1 expressed ectopically in fibroblasts was sufficient to induce HemSCs to acquire a pericyte-like phenotype, indicating a critical role for JAGGED1. In vivo, we blocked pericyte differentiation with recombinant JAGGED1, and we observed reduced formation of blood vessels, with an evident lack of pericytes. Silencing JAGGED1 in the endothelial cells reduced blood vessel formation and resulted in a paucity of pericytes. CONCLUSIONS: Our data show that endothelial JAGGED1 controls HemSC-to-pericyte differentiation in a murine model of IH and suggests that pericytes have a fundamental role in formation of blood vessels in IH.

Notch1 deficiency results in decreased inflammation during wound healing and regulates vascular endothelial growth factor receptor-1 and inflammatory cytokine expression in macrophages.

We investigated whether Notch signaling plays a role in regulating macrophage responses to inflammation. In a wound healing assay, macrophage recruitment was decreased in Notch1(+/-) mice, and the wounds were characterized by decreased TNF-α expression. As wound healing progressed, Notch1(+/-) wounds had increased vascularization and collagen deposition compared with wild-type wounds. In mice with myeloid-specific Notch1 deletion, wounds had decreased macrophage recruitment as well as decreased TNF-α expression, indicating the specific role of Notch1 in the inflammatory response in these cells. In vitro, we found that vascular endothelial growth factor receptor-1 (VEGFR-1) was upregulated in macrophages in response to LPS/IFN-γ and that this upregulation depended on Notch signaling. Furthermore, macrophages from Notch1(+/-) mice had decreased expression of VEGFR-1 compared with macrophages from wild-type mice, whereas VEGFR-1 expression in Notch4(-/-) macrophages was normal. Inhibition of Notch signaling decreased induction of the inflammatory cytokines IL-6, IL-12, CXCL10, MCP-1, monokine induced by IFN-γ, and TNF-α in macrophages in response to LPS/IFN-γ. Additionally, macrophages from Notch1(+/-) mice demonstrated decreased induction of IL-6, IL-12, and TNF-α in response to stimulation compared with wild-type mice. Thus, both pharmacological inhibition of Notch and genetic analysis demonstrate that Notch1 regulates VEGFR-1 and cytokine expression in macrophages. We have also established that Notch1 is important for the inflammatory response during wound healing in mice.

Propranolol treatment for chylothorax after congenital cardiac surgery.

OBJECTIVE: Postoperative chylothorax causes significant morbidities in pediatric patients with cardiac disease. New treatment approaches based on evolving understanding of underlying lymphatic dysfunction are being developed. We hypothesized that propranolol reduces morbidities and duration of chest tube requirement in high-output chylous effusion. METHODS: The postoperative courses of 50 pediatric patients with cardiac disease and high-output chylous effusion (control, n = 25; propranolol-treated, n = 25) were reviewed, including morbidities, length of hospitalization, and duration of chest tube requirement. Statistical analysis was performed using Welch's t test, Kruskal-Wallis tests for continuous variables, and chi-square and Fisher exact tests for categorical variables. Univariable logistic regression was used to determine predictors of response. RESULTS: Propranolol response was defined as 80% or more drainage reduction in 9 days or less. Treated patients were grouped into responders (<9 days) and nonresponders (>10 days). Neither initial amount of drainage (P = .12) nor day of propranolol initiation (P = .17) correlated with response. When compared with controls and nonresponders, responders had significantly fewer days with chest tube requirement (P < .01), infection (P < .0002), and thrombus (P = .005), and shorter hospitalization (P < .05). All patients had low serum albumin, although nonresponders had significantly decreased serum albumin when compared with responders and control patients (P < .002), and were more likely to receive albumin replacement (P < .01). Malnutrition was prevalent in all patient groups. CONCLUSIONS: Responders to propranolol had significantly less morbidity and duration of chest tube requirement when compared with control patients and nonresponders. Nonresponders did not have worse outcomes than control patients. We conclude that propranolol may be an effective treatment of patients with refractory chylothorax.

Interparietal bone (Os Incae) in craniosynostosis.

The interparietal bone, Os Incae, is formed in a persistent mendosal suture. This suture is a normal variant in the human skull, well-known in anatomy and radiology textbooks. We report 11 children with craniosynostosis in the presence of an interparietal bone, five from Children's Hospital at Montefiore and six children from Children's Hospital Boston. The true incidence of an interparietal bone in patients with craniosynostosis or craniofacial anomalies is not known; nor are there recognized sequelae of an interparietal bone (bathrocephaly). Hypotheses regarding mechanisms that may contribute to the formation of an interparietal bone are discussed.

Craniofacial growth in patients with FGFR3Pro250Arg mutation after fronto-orbital advancement in infancy.

BACKGROUND: The facial features of children with FGFR3Pro250Arg mutation (Muenke syndrome) differ from those with the other eponymous craniosynostotic disorders. We documented midfacial growth and position of the forehead after fronto-orbital advancement (FOA) in patients with the FGFR3 mutation. METHODS: We retrospectively reviewed all patients who had an FGFR3Pro250Arg mutation and craniosynostosis. Only patients who had FOA in infancy or early childhood were included. The clinical records were evaluated for type of sutural fusion; midfacial hypoplasia and other clinical data, including age at operation; type of procedures and fixation (wire vs resorbable plate); frequency of frontal readvancement, forehead augmentation, midfacial advancement; and complications. Preoperative and postoperative sagittal orbital-globe relationship was measured by direct anthropometry. Outcome of FOA was graded according to the Whittaker classification as category I, no revision; category II, minor revisions, that is, foreheadplasty; category III, alternative bony work; category IV; redo of initial procedure (ie, secondary FOA). Midfacial position was determined by clinical examination and lateral cephalometry. RESULTS: A total of 21 study patients with Muenke syndrome (8 males and 13 females) were analyzed. The types of craniosynostosis were bilateral coronal (n=15), of which 3 also had concurrent sagittal fusion, and unilateral coronal (n=5). Two patients had early endoscopic suturectomy, but later required FOA. Mean age at FOA was 22.9 months (range, 3-128 months). Secondary FOA was necessary in 40% of patients (n=8), and secondary foreheadplasty in 25% (n=5) of patients. No frontal revisions were needed in the remaining 35% of patients (n=7). Mean age at initial FOA was significantly younger in the group requiring repeat FOA or foreheadplasty compared with patients who did not require revision (P<0.05). Location of synostosis, type of fixation, and bone grafting did not significantly affect the need for revision. Only 30% (n=6) of patients developed midfacial retrusion. CONCLUSIONS: The frequency of frontal revision in patients with Muenke syndrome who had FOA in infancy and early childhood is lower than previously reported. Age at forehead advancement inversely correlated with the incidence of relapse and need for secondary frontal procedures. Midfacial retrusion is relatively uncommon in FGFR3Pro250Arg patients.

Home Field Advantage: Assessing the Geographic Trends of the Plastic Surgery Residency Match during the COVID-19 Pandemic.

OBJECTIVE: In previous plastic surgery residency match cycles, in-person activities at other institutions, such as away rotations, have facilitated matches outside of an applicant's home program or region. The COVID-19 pandemic, however, limited these in-person opportunities. Therefore, we hypothesized that applicants of the 2021 cycle would be more likely to match into programs with which they have existing geographic connections when compared to previous years. DESIGN: Residency websites and social media accounts were searched for resident names and educational information for those matching in 2021 and 2015 to 2020. Outcomes included proportion of applicants matching at the program affiliated with their medical school ("home program"), or matching in the same state or United States Census Map region as their medical school or undergraduate institution. Subgroup analyses were stratified by program region, incoming resident class size, and Doximity residency reputation ranking. SETTING: Columbia University (New York). PARTICIPANTS: For the 2015 to 2020 residency cycles, 963 residents were identified from 78 (95.1%) programs. For 2021, 159 incoming interns were identified from 70 (82.3%) programs. RESULTS: 2021 applicants matched into their home program at higher rates than 2015-2020 applicants (36.0% vs. 24.1%, p  = 0.019). This trend was similar regardless of program region or size. This increase was significant for programs ranked outside of the top 30 (41.5% vs. 26.4%, p = 0.032), but not for the top 30 programs (32.1% vs. 22.3%, p = 0.128). Excluding those who matched at their home program, 2015 to 2020 and 2021 applicants matched in the same state or region of their medical school or undergraduate institution at similar rates (p > 0.05 for all). CONCLUSIONS: During the COVID-19 pandemic, plastic surgery residency programs matched more applicants from affiliated medical schools than in previous years. This may result from lack of in-person opportunities for applicants at other programs. Alternative relationship-building opportunities may facilitate broader geographic connections in the 2022 cycle.

Propranolol Therapy in Infantile Hemangioma: It Is Not Just About the Beta.

BACKGROUND: Propranolol, a nonselective ß-adrenergic receptor antagonist, is approved by the U.S. Food and Drug Administration to treat problematic infantile hemangiomas, but a subset of patients experience treatment complications. Parents wary of long-term use and side effects consult plastic surgeons on surgical options or as a second opinion. Understanding the mechanism(s) of action of propranolol will allow plastic surgeons to better inform parents. METHODS: A systemic literature search was performed to query published translational and basic science studies on propranolol effects on infantile hemangiomas and cells derived from these lesions. RESULTS: In experimental studies, propranolol was antiproliferative and cytotoxic against hemangioma endothelial and stem cells and affected infantile hemangioma perivascular cell contractility. Propranolol inhibited migration, network formation, vascular endothelial growth factor A production, and vascular endothelial growth factor receptor 2 activation and down-regulated PI3K/AKT and mitogen-activated protein kinase signaling in hemangioma endothelial cells, but it increased ERK1/2 activity in hemangioma stem cells. At effective clinical doses, measured propranolol plasma concentration is 100 times higher than necessary for complete ß-adrenergic receptor blockade, yet was 10 to 100 times less than required to induce hemangioma stem cell death. CONCLUSIONS: Propranolol targets multiple cell types in infantile hemangiomas by means of ß-adrenergic receptor-dependent and -independent mechanisms. Plasma concentration played a significant role. At clinically relevant doses, incomplete infantile hemangioma suppression may explain the rebound phenomenon and worsening ulceration, and propranolol off target effects may lead to commonly reported adverse effects, such as sleep and gastrointestinal disturbances. Propranolol limitations and complications underscore the importance of surgical treatment options in cases of rebound and severe adverse effects. Surgical intervention remains an important treatment choice when parents are hesitant to use propranolol.

Venous malformation vessels are improperly specified and hyperproliferative.

Venous malformations (VMs) are slow-flow malformations of the venous vasculature and are the most common type of vascular malformation with a prevalence of 1%. Germline and somatic mutations have been shown to contribute to VM pathogenesis, but how these mutations affect VM pathobiology is not well understood. The goal of this study was to characterize VM endothelial and mural cell expression by performing a comprehensive expression analysis of VM vasculature. VM specimens (n = 16) were stained for pan-endothelial, arterial, venous, and endothelial progenitor cell proteins; proliferation was assessed with KI67. Endothelial cells in the VM vessels were abnormally orientated and improperly specified, as seen by the misexpression of both arterial and endothelial cell progenitor proteins not observed in control vessels. Consistent with arterialization of the endothelial cells, VM vessels were often surrounded by multiple layers of disorganized mural cells. VM endothelium also had a significant increase in proliferative endothelial cells, which may contribute to the dilated channels seen in VMs. Together the expression analysis indicates that the VM endothelium is misspecified and hyperproliferative, suggesting that VMs are biologically active lesions, consistent with clinical observations of VM progression over time.

Lymphatic Endothelial Cell Defects in Congenital Cardiac Patients With Postoperative Chylothorax.

OBJECTIVES: Chylothorax following cardiac surgery for congenital cardiac anomalies is a complication associated with severe morbidities and mortality. We hypothesize that there are intrinsic defects in the lymphatics of congenital cardiac patients. METHODS: Postsurgical chylothorax lymphatic endothelial cells (pcLECs) (n = 10) were isolated from the chylous fluid from congenital cardiac defect patients, and characterized by fluorescent-activated cell sorting, immunofluorescent staining, and quantitative RT-PCR. Results were compared to normal human dermal lymphatic endothelial cells (HdLECs). pcLECs (n = 3) and HdLECs were xenografted into immunocompromised mice. Implants and postoperative chylothorax patient's pulmonary tissues were characterized by immunostaining for lymphatic endothelial proteins. RESULTS: pcLECs expressed endothelial markers VECADHERIN, CD31, VEGFR2, lymphatic endothelial markers PROX1, podoplanin, VEGFR3, and progenitor endothelial markers CD90 and CD146. However, pcLECs had key differences relative to HdLECs, including altered expression and mislocalization of junctional proteins (VECADHERIN and CD31), and essential endothelial proteins, VEGFR2, VEGFR3, and PROX1. When xenografted in mice, pcLECs formed dilated lymphatic channels with poor cell-cell association. Similar to congenital lymphatic anomalies, the pulmonary lymphatics were dilated in a patient who developed postoperative chylothorax after cardiac surgery. CONCLUSIONS: Recent studies have shown that some postoperative chylothoraces in congenital cardiac anomalies are associated with anatomical lymphatic defects. We found that pcLECs have defects in expression and localization of proteins necessary to maintain lymphatic specification and function. This pcLEC phenotype is similar to that observed in lymphatic endothelial cells from congenital lymphatic anomalies. Co-existence of lymphatic anomalies should be considered as a feature of congenital cardiac anomalies.

A switch in Notch gene expression parallels stem cell to endothelial transition in infantile hemangioma.

BACKGROUND: Infantile hemangioma (IH) is the most common benign tumor of infancy, yet its pathogenesis is poorly understood. Notch family members are known to play a role in vascular development during embryogenesis and postnatal tumor angiogenesis, yet the role of Notch signaling in the pathogenesis of IH has not been investigated. This study aims to survey Notch expression in IH. MATERIALS AND METHODS: RNA from resected hemangioma tissue and hemangioma-derived stem cells (HemSCs) and endothelial cells (HemECs) was used for gene expression analyses by real-time PCR. Results were confirmed with immunofluorescence for protein expression in tissue. RESULTS: Real-time PCR showed that Notch family gene expression in IH is distinct from placenta and skin. Notch3 is expressed in HemSCs, but not in HemECs, indicating Notch3 is downregulated as HemSCs differentiate into HemECs. Moreover, expression of endothelial-associated Notch proteins, Notch1, Notch4, and Jagged-1 are increased in involuting hemangiomas and HemECs, suggesting that as hemangioma progresses toward involution, it acquires more differentiated endothelium. A subset of cells stained double positive for Notch3 and CD31, pointing to a potential intermediate between the HemSC cellular differentiation into HemEC. CONCLUSION: HemSCs have distinct Notch expression patterns from differentiated HemECs and from normal human endothelial cells. Notch3 is expressed in HemSCs, while Notch1, Notch4, and Jagged-1 have higher expression levels in HemECs. Notch3 was localized to the interstitial cells outside of the nascent vascular channels in proliferating IH tissue sections, but became more apparent in the perivascular cells in involuting IH. In summary, the pattern of Notch gene expression mirrors the progression from immature cells to endothelial-lined vascular channels (i.e., endothelial differentiation) that characterizes the growth and involution of IH.