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Engineering of fibrillar decorin matrices for a tissue-engineered trachea

Decorin is a structural and functional proteoglycan (PG) residing in the complex network of extracellular matrix (ECM) proteins in many connective tissues. Depending on the protein core and the glycosaminoglycan chain, PGs support cell adhesion, migration, proliferation, differentiation, ECM assembly and growth factor binding. For applications in tissue engineering, it is crucial to develop reliable, ECM-mimicking biomaterials. Electrospinning is a suitable method for creating three-dimensional (3D), fibrillar scaffolds. While there are numerous reports on the electrospinning of proteins including collagen, to date, there are no reports on the electrospinning of PGs. In the following study, we used electrospinning to generate decorin-containing matrices for tracheal tissue engineering applications. The electrospun scaffolds were analyzed using scanning electron microscopy, atomic force microscopy, contact angle measurements and dynamic mechanical analysis. Additionally, we confirmed PG functionality with immunostaining and 1,9-dimethylmethylene blue. To determine cell-matrix-interactions, tracheal cells (hPAECs) were seeded and analyzed using an FOXJ1-antibody. Moreover, interactions of the electrospun scaffolds with immune-mediated mechanisms were analyzed in detail. To conclude, we demonstrated the feasibility of electrospinning of decorin to generate functional 3D scaffolds with low immunogenicity for hPAEC expansion. Our data suggest that these hybrid materials may be suitable as a substrate for tracheal tissue engineering.

Percutaneous SAPIEN S3 Transcatheter Valve Implantation for Post-Left Ventricular Assist Device Aortic Regurgitation.

Aortic regurgitation was found to develop in a considerable share of patients supported with continuous flow left ventricular assist devices (LVADs). The resulting circulatory loop renders LVAD operation inefficient so that symptoms of heart failure develop in spite of high LVAD flows. In patients with a high reoperative risk, transcatheter aortic valve implantation may be considered as an alternative to reoperative valve surgical procedures. We report a case of percutaneous transcatheter aortic valve implantation using the SAPIEN S3 (Edwards Lifesciences, Inc, Irvine, CA) valve for post-LVAD aortic regurgitation.

Methylene blue modulates transendothelial migration of peripheral blood cells.

Vasoplegia is a severe complication after cardiac surgery. Within the last years the administration of nitric oxide synthase inhibitor methylene blue (MB) became a new therapeutic strategy. Our aim was to investigate the role of MB on transendothelial migration of circulating blood cells, the potential role of cyclic cGMP, eNOS and iNOS in this process, and the influence of MB on endothelial cell apoptosis. Human vascular endothelial cells (HuMEC-1) were treated for 30 minutes or 2 hours with different concentrations of MB. Inflammation was mimicked by LPS stimulation prior and after MB. Transmigration of PBMCs and T-Lymphocytes through the treated endothelial cells was investigated. The influence of MB upon the different subsets of PBMCs (Granulocytes, T- and B-Lymphocytes, and Monocytes) was assessed after transmigration by means of flow-cytometry. The effect of MB on cell apoptosis was evaluated using Annexin-V and Propidium Iodide stainings. Analyses of the expression of cyclic cGMP, eNOS and iNOS were performed by means of RT-PCR and Western Blot. Results were analyzed using unpaired Students T-test. Analysis of endothelial cell apoptosis by MB indicated a dose-dependent increase of apoptotic cells. We observed time- and dose-dependent effects of MB on transendothelial migration of PBMCs. The prophylactic administration of MB led to an increase of transendothelial migration of PBMCs but not Jurkat cells. Furthermore, HuMEC-1 secretion of cGMP correlated with iNOS expression after MB administration but not with eNOS expression. Expression of these molecules was reduced after MB administration at protein level. This study clearly reveals that endothelial response to MB is dose- and especially time-dependent. MB shows different effects on circulating blood cell-subtypes, and modifies the release patterns of eNOS, iNOS, and cGMP. The transendothelial migration is modulated after treatment with MB. Furthermore, MB provokes apoptosis of endothelial cells in a dose/time-dependent manner.

Xeno-immunogenicity of ice-free cryopreserved porcine leaflets

BACKGROUND Undesirable processes of inflammation, calcification, or immune-mediated reactions are limiting factors in long-term survival of heart valves in patients. In this study, we target the modulatory effects of ice-free cryopreservation (IFC) of xenogeneic heart valve leaflet matrices, without decellularization, on the adaptive human immune responses in vitro. METHODS We tested porcine leaflet matrices from fresh untreated, conventionally cryopreserved (CFC), and IFC pulmonary valves by culturing them with human blood mononuclear cells for 5 d in vitro. No other tissue treatment protocols to modify possible immune responses were used. Matrices alone or in addition with a low-dose second stimulus were analyzed for induction of proliferation and cytokine release by flow cytometry-based techniques. Evaluation of the α-Gal epitope expression was performed by immunohistochemistry with fluorochrome-labeled B4 isolectin. RESULTS None of the tested leaflet treatment groups directly triggered the proliferation of immune cells. But when tested in combination with a second trigger by anti-CD3, IFC valves showed significantly reduced proliferation of T cells, especially effector memory T cells, in comparison with fresh or CFC tissue. Moreover, the cytokine levels for interferon-γ (IFNγ), tumor necrosis factor α, and interleukin-10 were reduced for the IFC-treated group being significantly different compared with the CFC group. However, no difference between treatment groups in the expression of the α-Gal antigen was observed. CONCLUSIONS IFC of xenogeneic tissue might be an appropriate treatment method or processing step to prevent responses of the adaptive immune system.

Initiation of an Inter-hospital ECMO Transfer Program for Patients Suffering From Severe Acute Heart and/or Pulmonary Failure

1. Holmes Jr DR, Mack MJ, Kaul S, Agnihotri A, Alexander KP, Bailey SR, et al. 2012 ACCF/AATS/SCAI/STS expert consensus document on transcatheter aortic valve replacement. J Am Coll Cardiol. 2012;59:1200–54. 2. Mummert J, Sirois E, Sun W. Quantification of biomechanical interaction of transcatheter aortic valve stent deployed in porcine and ovine hearts. Ann Biomed Eng. 2013;41:577–86. 3. Tzamtzis S, Viquerat J, Yap J, Mullen MJ, Burriesci G. Numerical analysis of the radial force produced by the medtronic-corevalve and edwards-sapien after transcatheter aortic valve implantation (TAVI). Med Eng Phys. 2013;35:125–30. 4. Achenbach S, Delgado V, Hausleiter J, Schoenhagen P, Min JK, Leipsic JA. SCCT expert consensus document on computed tomography imaging before transcatheter aortic valve implantation (TAVI)/transcatheter aortic valve replacement (TAVI). J Cardiovasc Comput Tomogr. 2012;6:366–80. 5. Hayashida K, Bouvier E, Lefevre T, Chevalier B, Hovasse T, Romano M, et al. Transcatheter aortic valve implantation for patients with severe bicuspid aortic valve stenosis. Circ Cardiovasc Interv. 2013;6:284–91. 6. Sun W, Li K, Sirois E. Simulated elliptical bioprosthetic valve deformation: Implications forasymmetrictranscathetervalvedeployment. JBiomech.2010;43:3085–90.

Pre-filling of the extracorporeal circuit with autologous blood is safe, but not effective in optimizing biocompatibility in high-risk patients

Objectives: Haemodilution resulting from crystalloid priming of the cardiopulmonary bypass circuit represents a major risk factor for blood transfusions in high-risk cardiac surgery patients. We designed this study to evaluate the effects of antegrade autologous priming (AAP) on reducing perioperative blood transfusion and markers of the inflammatory response in older patients (>75 years). Methods: Seventy-two patients undergoing first-time coronary bypass and/or aortic valve replacement were prospectively randomised to a cardiopulmonary bypass (CPB) with or without AAP. AAP was performed by adding the patient’s own blood to the prime solution (mean 280ml). Perfusion and anaesthetic techniques were as usual. The haematocrit was maintained at a minimum of 21% during CPB. Patients were well matched for all preoperative variables, including established transfusion risk factors. The primary endpoint was the requirement of red cell transfusion. The surrogate endpoints were renal function, inflammatory response and ischaemic parameters. Blood samples were drawn pre- and intraoperatively and at intervals of 6 hours till POD 6. Results: Current analysis shows no differences in patients receiving homologous packed red cell transfusions. Also, markers of the inflammatory response (IL6, IL8), renal function (cystatin C, creatinine) and myocardial ischaemia (troponin T, CK-MB) were comparable in both groups (p>0.05). Clinical outcomes were similar with respect to pulmonary, renal and hepatic function, length of ICU stay and hospital stay. Conclusion: These data suggest that antegrade autologous priming is a safe procedure, but an ineffective way for improving biocompatibility and reducing the need for blood transfusion in older patients.

Effects on human heart valve immunogenicity in vitro by high concentration cryoprotectant treatment.

It has been shown previously that cryopreservation, using an ice‐free cryopreservation method with the cryoprotectant formulation VS83, beneficially modulated immune reactions in vivo and in vitro when compared with conventionally frozen tissues. In this study, we assessed the impact of a VS83 post‐treatment of previously conventionally frozen human tissue on responses of human immune cells in vitro. Tissue punches of treated and non‐treated (control) aortic heart valve tissue (leaflets and associated aortic root) were co‐cultured for 7 days with peripheral blood mononuclear cells or enriched CD14+ monocytes. Effects on cellular activation markers, cytokine secretion and immune cell proliferation were analysed by flow cytometry. Flow cytometry studies showed that VS83 treatment of aortic root tissue promoted activation and differentiation of CD14+ monocytes, inducing both up‐regulation of CD16 and down‐regulation of CD14. Significantly enhanced expression levels for the C‐C chemokine receptor (CCR)7 and the human leukocyte antigen (HLA)‐DR on monocytes co‐cultured with VS83‐treated aortic root tissue were measured, while the interleukin (IL)‐6 and monocyte chemoattractant protein (MCP)‐1 release was suppressed. However, the levels of interferon (IFN)γ and tumour necrosis factor (TNF)α remained undetectable, indicating that complete activation into pro‐inflammatory macrophages did not occur. Similar, but non‐significant, changes occurred with VS83‐treated leaflets. Additionally, in co‐cultures with T cells, proliferation and cytokine secretion responses were minimal. In conclusion, post‐treatment of conventionally cryopreserved human heart valve tissue with the VS83 formulation induces changes in the activation and differentiation characteristics of human monocytes, and thereby may influence long‐term performance following implantation. Copyright

Suspected involvement of EPTFE membrane in sterile intrathoracic abscess and pericardial empyema in a multi-allergic LVAD recipient: a case report

Device-related infections in recipients of left ventricular assist devices (LVAD) have been recognized as a major source of morbidity and mortality. They require a high level of diagnostic effort as part of the overall burden resulting from infectious complications in LVAD recipients. We present a multi-allergic patient who was treated for persistent sterile intrathoracic abscess formation and pericardial empyema following minimally invasive LVAD implantation including use of a sheet of e-polytetrafluoroethylene (ePTFE) membrane to restore pericardial integrity. Sterile abscess formation and pericardial empyema recurred after surgical removal until the ePTFE membrane was removed, suggesting that in disposed patients, ePTFE may be related to sterile abscess formation or sterile empyema.

Midterm follow up in patients with reduction ascending aortoplasty

BackgroundThe reduction ascending aortoplasty in patients with an aortic ectasia/dilatation is a common procedure during concomitant cardiac operations. Aim of the follow up study was the evaluation of possible re-dilatation and complications.MethodsFrom 1998 to 2010 124 patients (69% male; mean age 66.6 ± 12 ys) with ectasia of the ascending aortic who had no further indication for an aortic replacement, were included. The mean preoperative diameter of the ascending aorta was 4.2 ± 0.6 cm. The patients risk profile was moderate (mean EF 51% ± 11%, Euroscore 4.2 ± 2.1). To treat the dilatation of the ascending aorta, a longitudinal incision was performed and a strip of the aortic wall was resected. A reduction aortoplasty was carried out with a double-layered suture line using a 4/0 Prolene mattress suture with an additional 4/0 Prolene running suture. A follow up (rate 95%) was performed by echocardiography- and clinical examination.ResultsAll patients underwent reduction aortoplasty associated with a primary cardiac surgical procedure (AVR 63%, CABG 13%, other or combination 24%). The intrahospital mortality rate was 4%. Four aortic bleeding complications occurred. After a mean postoperative period of 57 ± 39 months, the ascending aortic diameter (3.6 ± 0.6 cm) was still significantly (P < 0.01) reduced. No postoperative aortic-related complications including aortic rupture, dissection and reoperation were observed. In 4 patients, the ascending aorta had re-dilated to the preoperative diameter.ConclusionReduction ascending aortoplasty without external wrapping is a safe procedure with acceptable midterm results in patients with asymptomatic dilatations and concomitant cardiac surgical procedures.

Left ventricular non-compaction cardiomyopathy and left ventricular assist device: a word of caution

BackgroundIn patients with left ventricular non-compaction (LVNC), implantation of a left ventricular assist device (LVAD) may be performed as a bridge to transplantation. In this respect, the particular characteristics of the left ventricular myocardium may represent a challenge.Case presentationWe report a patient with LVNC who required urgent heart transplantation for inflow cannula obstruction nine months after receiving a LVAD. LVAD parameters, echocardiography and examination of the explanted heart suggested changes of left ventricular configuration brought about by LVAD support as the most likely cause of inflow cannula obstruction.ConclusionsWe conclude that changes experienced by non-compacted myocardium during LVAD support may give rise to inflow cannula obstruction and flow reduction. Presence of LVNC mandates tight surveillance for changes in LV configuration and LVAD flow characteristics and may justify urgent transplantation listing status.