Growth capacity of a Wharton's Jelly derived mesenchymal stromal cells tissue engineered vascular graft used for main pulmonary artery reconstruction in piglets.

Background: Surgical treatment of congenital heart defects affecting the right ventricular outflow tract (RVOT) often requires complex reconstruction and multiple reoperations due to structural degeneration and lack of growth of currently available materials. Hence, alternative approaches for RVOT reconstruction, which meet the requirements of biocompatibility and long-term durability of an ideal scaffold, are needed. Through this full scale pre-clinical study, we demonstrated the growth capacity of a Wharton's Jelly derived mesenchymal stromal cells (WJ-MSC) tissue engineered vascular graft used in reconstructing the main pulmonary artery in piglets, providing proof of biocompatibility and efficacy. Methods: Sixteen four-week-old Landrace pigs were randomized to undergo supravalvar Main Pulmonary Artery (MPA) replacement with either unseeded or WJ-MSCs-seeded Small Intestinal Submucosa-derived grafts. Animals were followed up for 6 months by clinical examinations and cardiac imaging. At termination, sections of MPAs were assessed by macroscopic inspection, histology and fluorescent immunohistochemistry. Results: Data collected at 6 months follow up showed no sign of graft thrombosis or calcification. The explanted main pulmonary arteries demonstrated a significantly higher degree of cellular organization and elastin content in the WJ-MSCs seeded grafts compared to the acellular counterparts. Transthoracic echocardiography and cardiovascular magnetic resonance confirmed the superior growth and remodelling of the WJ-MSCs seeded conduit compared to the unseeded. Conclusion: Our findings indicate that the addition of WJ-MSCs to the acellular scaffold can upgrade the material, converting it into a biologically active tissue, with the potential to grow, repair and remodel the RVOT.

Wharton's Jelly-Mesenchymal Stem Cell-Engineered Conduit for Pulmonary Artery Reconstruction in Growing Piglets.

Surgical treatment of congenital heart defects affecting the right ventricular outflow tract often requires complex reconstruction and multiple reoperations. With a randomized controlled trial, we compared a novel tissue-engineered small intestine submucosa-based graft for pulmonary artery reconstruction (seeded with mesenchymal stem cells derived from Wharton's Jelly) with conventional small intestine submucosa in growing piglets. Six months after implantation, seeded grafts showed integration with host tissues at cellular level and exhibited growth potential on transthoracic echocardiography and cardiovascular magnetic resonance. Our seeded graft is a promising biomaterial for pulmonary artery reconstruction in pediatric patients with right ventricular outflow tract abnormalities.

Reconstruction of the Swine Pulmonary Artery Using a Graft Engineered With Syngeneic Cardiac Pericytes.

The neonatal heart represents an attractive source of regenerative cells. Here, we report the results of a randomized, controlled, investigator-blinded preclinical study, which assessed the safety and effectiveness of a matrix graft cellularized with cardiac pericytes (CPs) in a piglet model of pulmonary artery (PA) reconstruction. Within each of five trios formed by 4-week-old female littermate piglets, one element (the donor) was sacrificed to provide a source of CPs, while the other two elements (the graft recipients) were allowed to reach the age of 10 weeks. During this time interval, culture-expanded donor CPs were seeded onto swine small intestinal submucosa (SIS) grafts, which were then shaped into conduits and conditioned in a flow bioreactor. Control unseeded SIS conduits were subjected to the same procedure. Then, recipient piglets were randomized to surgical reconstruction of the left PA (LPA) with unseeded or CP-seeded SIS conduits. Doppler echocardiography and cardiac magnetic resonance imaging (CMRI) were performed at baseline and 4-months post-implantation. Vascular explants were examined using histology and immunohistochemistry. All animals completed the scheduled follow-up. No group difference was observed in baseline imaging data. The final Doppler assessment showed that the LPA's blood flow velocity was similar in the treatment groups. CMRI revealed a mismatch in the average growth of the grafted LPA and contralateral branch in both treatment groups. Histology of explanted arteries demonstrated that the CP-seeded grafts had a thicker luminal cell layer, more intraparietal arterioles, and a higher expression of endothelial nitric oxide synthase (eNOS) compared with unseeded grafts. Moreover, the LPA stump adjacent to the seeded graft contained more elastin and less collagen than the unseeded control. Syngeneic CP engineering did not accomplish the primary goal of supporting the graft's growth but was able to improve secondary outcomes, such as the luminal cellularization and intraparietal vascularization of the graft, and elastic remodeling of the recipient artery. The beneficial properties of neonatal CPs may be considered in future bioengineering applications aiming to reproduce the cellular composition of native arteries.

Wharton's Jelly-Mesenchymal Stem Cell-Engineered Conduit for Pediatric Translation in Heart Defect.

The materials available for the right ventricular outflow tract (RVOT) reconstruction in patients with tetralogy of fallot (TOF)/pulmonary atresia come with the severe limitation of long-term degeneration and lack of growth potential, causing right ventricular dysfunction, aneurysm formation, and arrhythmias, thus necessitating several high-risk reoperations throughout patients' lives. In this study, we evaluated the capacity of mesenchymal stem cells (MSCs) derived from the Wharton's Jelly (WJ-MSCs), the gelatinous inner portion of the umbilical cord, to grow and recellularize an extracellular matrix (ECM) graft in our optimized xeno-free, good manufacturing practice-compliant culture system. WJ-MSCs were phenotypically and functionally characterized by flow cytometry and multilineage differentiation capacity, respectively. The typical MSC immunophenotype and functional characteristics were retained in our xeno-free culture system, as well as the capacity to grow and engraft onto a naturally occurring scaffold. WJ-MSCs, from both human and swine source, showed excellent capacity to recellularize ECM graft producing a living cell-seeded construct. In addition, we have provided an in vivo proof of concept of feasibility of the cellularized conduit, engineered with swine WJ-MSCs, to be used in a novel porcine model of main pulmonary artery reconstruction, where it showed good integration within the host tissue. Our study indicates that the addition of WJ-MSCs to the ECM scaffold can upgrade the material, converting it into a living tissue, with the potential to grow, repair, and remodel the RVOT. These results could potentially represent a paradigm shift in pediatric cardiac intervention toward new modalities for effective and personalized surgical restoration of pulmonary artery and RVOT function in TOF/pulmonary atresia patients. Impact Statement The materials available for pulmonary artery reconstruction in pediatric patients with Congenital Heart Defect come with the limitation of long-term degeneration and lack of growth, thus necessitating several reoperations. Here, we describe a novel approach combining perinatal stem cells and naturally occurring scaffold to create a living tissue engineered conduit that showed good growth potential in a pulmonary artery reconstruction porcine model. We envision this approach is of great interest and relevance in tissue engineering field applied to cardiovascular regenerative medicine, as it may open up new avenues for correction of congenital cardiac defects, with remarkable medical and social benefits.

A low threshold for neonatal intervention yields a high rate of biventricular outcomes in pulmonary atresia with intact ventricular septum.

AIMS: Management strategies for pulmonary atresia with intact ventricular septum are variable and are based on right ventricular morphology and associated abnormalities. Catheter perforation of the pulmonary valve provides an alternative strategy to surgery in the neonatal period. We sought to assess the long-term outcome in terms of survival, re-intervention, and functional ventricular outcome in the setting of a 26-year single-centre experience of low threshold inclusion criteria for percutaneous valvotomy. METHODS AND RESULTS: Retrospective analysis of patients diagnosed with pulmonary atresia with intact ventricular septum from 1990 to 2016 at a tertiary referral centre, was performed. Of 71 patients, 48 were brought to the catheterisation laboratory for intervention. Catheter valvotomy was successful in 45 patients (94%). Twenty-three patients (51%) also underwent ductus arteriosus stenting. The length of intensive care and hospital stay was significantly shorter, and early re-interventions were significantly reduced in the catheterisation group. There were eight deaths (17%); all within 35 days of the procedure. Of the survivors, only one has required a Fontan circulation. Twenty-eight patients (74%) have undergone biventricular repair and nine patients (24%) have one-and-a-half ventricle circulation. Following successful valvotomy, 80% of patients required further catheter-based or surgical interventions. CONCLUSIONS: A low threshold for initial interventional management yielded a high rate of successful biventricular circulations. Although mortality was low in patients who survived the peri-procedural period, the rate of re-intervention remained high in all groups.

Visualization of coronary arteries in paediatric patients using whole-heart coronary magnetic resonance angiography: comparison of image-navigation and the standard approach for respiratory motion compensation.

AIMS: To investigate the use of respiratory motion compensation using image-based navigation (iNAV) with constant respiratory efficiency using single end-expiratory thresholding (CRUISE) for coronary magnetic resonance angiography (CMRA), and compare it to the conventional diaphragmatic navigator (dNAV) in paediatric patients with congenital or suspected heart disease. METHODS: iNAV allowed direct tracking of the respiratory heart motion and was generated using balanced steady state free precession startup echoes. Respiratory gating was achieved using CRUISE with a fixed 50% efficiency. Whole-heart CMRA was acquired with 1.3 mm isotropic resolution. For comparison, CMRA with identical imaging parameters were acquired using dNAV. Scan time, visualization of coronary artery origins and mid-course, imaging quality and sharpness was compared between the two sequences. RESULTS: Forty patients (13 females; median weight: 44 kg; median age: 12.6, range: 3 months-17 years) were enrolled. 25 scans were performed in awake patients. A contrast agent was used in 22 patients. The scan time was significantly reduced using iNAV for awake patients (iNAV 7:48 ± 1:26 vs dNAV 9:48 ± 3:11, P = 0.01) but not for patients under general anaesthesia (iNAV = 6:55 ± 1:50 versus dNAV = 6:32 ± 2:16; P = 0.32). In 98% of the cases, iNAV image quality had an equal or higher score than dNAV. The visual score analysis showed a clear difference, favouring iNAV (P = 0.002). The right coronary artery and the left anterior descending vessel sharpness was significantly improved (iNAV: 56.8% ± 10.1% vs dNAV: 53.7% ± 9.9%, P < 0.002 and iNAV: 55.8% ± 8.6% vs dNAV: 53% ± 9.2%, P = 0.001, respectively). CONCLUSION: iNAV allows for a higher success-rate and clearer depiction of the mid-course of coronary arteries in paediatric patients. Its acquisition time is shorter in awake patients and image quality score is equal or superior to the conventional method in most cases.

A novel real-time computational framework for detecting catheters and rigid guidewires in cardiac catheterization procedures.

PURPOSE: Catheters and guidewires are used extensively in cardiac catheterization procedures such as heart arrhythmia treatment (ablation), angioplasty, and congenital heart disease treatment. Detecting their positions in fluoroscopic X-ray images is important for several clinical applications, for example, motion compensation, coregistration between 2D and 3D imaging modalities, and 3D object reconstruction. METHODS: For the generalized framework, a multiscale vessel enhancement filter is first used to enhance the visibility of wire-like structures in the X-ray images. After applying adaptive binarization method, the centerlines of wire-like objects were extracted. Finally, the catheters and guidewires were detected as a smooth path which is reconstructed from centerlines of target wire-like objects. In order to classify electrode catheters which are mainly used in electrophysiology procedures, additional steps were proposed. First, a blob detection method, which is embedded in vessel enhancement filter with no additional computational cost, localizes electrode positions on catheters. Then the type of electrode catheters can be recognized by detecting the number of electrodes and also the shape created by a series of electrodes. Furthermore, for detecting guiding catheters or guidewires, a localized machine learning algorithm is added into the framework to distinguish between target wire objects and other wire-like artifacts. The proposed framework were tested on total 10,624 images which are from 102 image sequences acquired from 63 clinical cases. RESULTS: Detection errors for the coronary sinus (CS) catheter, lasso catheter ring and lasso catheter body are 0.56 ± 0.28 mm, 0.64 ± 0.36 mm, and 0.66 ± 0.32 mm, respectively, as well as success rates of 91.4%, 86.3%, and 84.8% were achieved. Detection errors for guidewires and guiding catheters are 0.62 ± 0.48 mm and success rates are 83.5%. CONCLUSION: The proposed computational framework do not require any user interaction or prior models and it can detect multiple catheters or guidewires simultaneously and in real-time. The accuracy of the proposed framework is sub-mm and the methods are robust toward low-dose X-ray fluoroscopic images, which are mainly used during procedures to maintain low radiation dose.

Large Calibre Self-Expanding Stents for Pulmonary Stenosis After the Arterial Switch, a Low-Risk Solution to a Low-Flow Situation.

Branch pulmonary artery stenosis is one of the most common complications late after the arterial switch operation. The pathophysiology of stenosis in these circumstances is very different to that encountered in with normally related great vessels. The LeCompte manoeuvre leaves the pulmonary arteries straddled and stretched over the aortic root which contributing significantly to the degree of stenosis encountered. Unilateral branch pulmonary artery stenosis rarely leads to significant symptoms in young patients with biventricular repairs; however, significant discrepancies in the split flows between right and left lung may place the patient in an unfavourable physiological position as they move in to middle age. Balloon expandable stent implantation distorts the anatomical arrangement of the LeCompte and is associated with negative interactions with the often dilated aortic root. We present three cases of the use of flexible self-expanding stents to improve the physiological flow distributions to each lung whilst hopefully decreasing the risk of erosion and perforation or fistula formation related to the pulmonary artery. All three patients had uncomplicated procedures with significant improvements in vessel diameter and flow distribution noted on follow-up MRI scans. This is the first report of the use of self-expanding stents in the context of branch pulmonary artery stenosis after the LeCompte manoeuvre. Theoretically and based on our early experience, this technique provides potential safety advantages over the use of more traditional balloon expandable stents.

Matrix-Assisted Ionization on a Portable Mass Spectrometer: Analysis Directly from Biological and Synthetic Materials.

Matrix-assisted ionization (MAI)-mass spectrometry (MS) eliminates the need for high voltage, a heat source, lasers, and compressed gases in the ionization process and uses minimal solvents in sample preparation, thus making MAI ideal for field-portable mass spectrometers. The broad applicability of MAI is demonstrated by simple, rapid, and robust positive and negative detection mode analyses of low and high mass compounds including some pesticides, dyes, drugs, lipids, and proteins (186 Da to 8.5 kDa) from various materials including urine, biological tissue sections, paper, and plant material on a low pumping capacity, single-quadrupole mass spectrometer. Different sample introduction methods are applicable, including the use of a pipet tip or glass melting point tube, allowing integration of sample preparation with sample introduction for increased analytical utility and ease of operation, even when sampling directly from surfaces.

Reproducibility and quantification of illicit drugs using matrix-assisted ionization (MAI) mass spectrometry.

Matrix-assisted ionization (MAI) mass spectrometry (MS) is a simple and sensitive method for analysis of low- and high-mass compounds, requiring only that the analyte in a suitable matrix be exposed to the inlet aperture of an atmospheric pressure ionization mass spectrometer. Here, we evaluate the reproducibility of MAI and its potential for quantification using six drug standards. Factors influencing reproducibility include the matrix compound used, temperature, and the method of sample introduction. The relative standard deviation (RSD) using MAI for a mixture of morphine, codeine, oxymorphone, oxycodone, clozapine, and buspirone and their deuterated internal standards using the matrix 3-nitrobenzonitrile is less than 10% with either a Waters SYNAPT G2 or a Thermo LTQ Velos mass spectrometer. The RSD values obtained using MAI are comparable to those using ESI or MALDI on these instruments. The day-to-day reproducibility of MAI determined for five consecutive days with internal standards was better than 20% using manual sample introduction. The reproducibility improved to better than 5% using a mechanically assisted sample introduction method. Hydrocodone, present in a sample of undiluted infant urine, was quantified with MAI using the standard addition method.

Second stage after initial hybrid palliation for hypoplastic left heart syndrome: Arterial or venous shunt?

OBJECTIVE: Hybrid palliation for hypoplastic left heart syndrome has been developed as an alternative to neonatal Norwood surgery. At the second stage, a source of pulmonary blood flow has to be established. Options include an arterial modified Blalock-Taussig or a venous superior cavopulmonary shunt. METHODS: We retrospectively reviewed patients who received second-stage palliation after the initial hybrid. Patients were stratified according to the source of pulmonary blood supply into the arterial shunt (n = 17 patients) or venous shunt (n = 26 patients). RESULTS: Age and weight at second stage were lower in the arterial group (85 [45-268] days vs 152.5 [61-496] days, P = .001 and 3.6 [2.7-9.4] kg vs 5.1 [2.97-9.4] kg, P = .001, respectively). All recorded surgical times were shorter in the arterial group. Mechanical ventilation and intensive care stay were shorter in the venous group (5.82 [2.01-14.9] days vs 2.42 [0.56-13.67] days, P = .005 and 8.5 [3.6-23.7] vs 5.75 [0.8-17.6] days, P = .036, respectively) There was no difference in mortality (2/17 vs 5/26; P = .685) or incidence of complications between the 2 groups. There was a tendency toward a higher need for intervention in the immediate postoperative period in the venous group, but this did not reach significance (6/17 vs 13/26, P = .342). The arterial group has shown better development of the branch pulmonary arteries with a higher lower lobe index (158.38 ± 39.43 mm(2)/m(2) vs 113.33 ± 43.96 mm(2)/m(2), respectively, P = .037). CONCLUSIONS: Both arterial and venous shunts are viable options with mortality and morbidity results comparable to those in the literature. The arterial shunt pathway (2-stage Norwood I) may offer better pulmonary arterial growth than the venous shunt (comprehensive/combined Norwood I and II).

High-throughput characterization of small and large molecules using only a matrix and the vacuum of a mass spectrometer.

Matrix assisted ionization vacuum (MAIV) rapidly generates gas-phase analyte ions from subliming solid-phase matrix:analyte crystals for analysis by mass spectrometry (MS). Ionization from the solid-phase allows the use of a variety of surfaces for introducing matrix:analyte samples to the vacuum of a mass spectrometer, including common laboratory materials, such as disposable pipet tips, filter paper, tooth picks, and nylon mesh. MAIV is shown here to be capable of analyses as fast as 3 s per sample with achievable sensitivities in the low femtomole range. MAIV-MS coupled with ion mobility spectrometry (IMS)-MS and tandem mass spectrometry (MS/MS) is shown to be especially powerful for analysis and characterization of a wide range of molecules ranging from small molecules such as drugs and metabolites (∼300 Da) to intact proteins (25.6 kDa). Automated sample introduction is demonstrated on two different commercial mass spectrometers using a programmable XYZ stage. A MAIV high-throughput nontargeted MS(E) approach is also demonstrated utilizing IMS for rapid characterization of small molecules and peptides from standard solutions, as well as drug spiked human urine.

Long-term follow-up is not indicated after routine interventional closure of persistent arterial ducts.

BACKGROUND: Little is known about the necessity for long-term follow-up after interventional closure of persistent arterial duct (PDA). Potential side effects and complications include residual shunts, haemolysis, device embolization, and obstruction to flow in the adjoining vessels. METHODS: Single centre retrospective study of paediatric patients undergoing interventional PDA occlusion. RESULTS: 315 patients who underwent interventional occlusion of a PDA between November 2002 and September 2013 were included. Of these, eight needed re-intervention (three for device embolization, five for residual shunt). Seven had mild obstruction to flow in the adjoining vessels, but did not require any intervention. All sequelae were found latest at the first follow-up appointment after the procedure (usually within 3 months); whilst none developed during further follow-up. CONCLUSION: Complications of interventional closure of PDA were apparent immediately after the procedure or by three months of follow-up. Long-term follow-up is not indicated in cases when no complications are seen early after the procedure.

Anaesthetic management of previously non-diagnosed phaeochromocytoma: Clinical vigilance, the ultimate saviour of anaesthesiologist.

A 39-year-old male, post nephrectomy and adrenalectomy (right), was planned for adrenalectomy (left) and radiofrequency ablation of left renal mass. Clinical evaluation indicated a possibility of phaeochromocytoma, whereas biochemical parameters were found to be within normal limits. Intraoperatively, massive fluctuations in haemodynamic parameters were noticed while the tumour was being handled. Patient was stabilised with inotropes, vasopressors, fluids and careful titration of anaesthetic agents. Preoperatively diagnosed coronary disease could have complicated anaesthetic care. Optimum and modern anaesthetic care leads to safe execution of surgery.

Amlodipine prevents apoptotic cell death by correction of elevated intracellular calcium in a primary neuronal model of Batten disease (CLN3 disease).

CLN3 disease (Spielmeyer-Vogt-Sjogren-Batten disease) is a severe pediatric neurodegenerative disorder for which there is currently no effective treatment. The disease is characterized by progressive neuronal death, which may be triggered by abnormal intracellular calcium levels leading to neuronal apoptosis. Previously, we demonstrated reversal of the calcium effect in a neuroblastoma cell line using amlodipine and other calcium channel antagonists. In the present studies, we developed a CLN3 siRNA-inhibited primary rat neuron model to further study etoposide-induced calcium changes and apoptosis in CLN3 disease followed by recovery experiments with amlodipine. Our results show that intracellular calcium is significantly elevated in siRNA-inhibited cortical neurons after potassium chloride-induced depolarization. We were also able to show that amlodipine, a predominantly L-type dihydropyrimidine calcium channel antagonist can reverse the aberrant calcium elevations in this model of the disease. We performed an in situ TUNEL assay following etoposide-exposure to siRNA inhibited primary neurons, and apoptotic nuclei were detected providing additional evidence that increased neuronal apoptosis is associated with increased calcium levels. Amlodipine also reduced the absolute number of apoptotic cells in this experimental model.

Highlights of the 6th World Congress in Paediatric Cardiology and Cardiac Surgery.

The 6th World Congress in Paediatric Cardiology and Cardiac Surgery took place in Cape Town, South Africa, in February 2013. The congress is the largest meeting in the field of congenital and paediatric heart disease and attracts a global audience of specialists with the aim of sharing the latest multidisciplinary developments in research and clinical practice. The congress was commended as a huge success and this article aims to give a general flavor of the diverse meeting through detailing a few specific highlights from the various tracks.

Fungal myocarditis in a preterm neonate.

A male infant born at 25 weeks gestation presented with abdominal distension, was transferred to our institution for surgical management following suspected bowel perforation with severe sepsis. Umbilical catheter cultures grew Candida parapsilosis. At laparotomy, there was a large ileal perforation with peritonitis, he was treated with amphotericin, antibiotics and had an ileostomy. He had persistent pulmonary hypertension, requiring nitric oxide and high-frequency oscillatory ventilation. Serial echocardiograms revealed a patent ductus arteriosus (PDA), but also demonstrated increasing left ventricular hypertrophy and the development of bright areas within the septal myocardium. Further bright areas developed over a course of 2 weeks in his right ventricular outflow tract. After treatment for candidal infection, there was improvement in left ventricular thickness and brightness of the echogenic lesions was reduced. Biopsy of the lesions was discounted due to the risk of the procedure, the size of the infant and his improving clinical status.

Diagnosis of a patient with a kinetic variant of medium and short-chain 3-hydroxyacyl-CoA dehydrogenase deficiency by newborn screening.

Medium and short-chain 3-hydroxyacyl-CoA dehydrogenase deficiency is a rare cause of impaired mitochondrial fatty acid oxidation. We present a case report of a patient with hyperinsulinism and homozygosity for a novel mutation causing a kinetic variant of the enzyme. The diagnosis was initially inferred by abnormal newborn screening acylcarnitine analysis with elevated C4-hydroxyacylcarnitine.