Engineering myocardial tissue patches with hierarchical structure-function.

Complex hierarchical organization is a hallmark of tissues and their subsequent integration into organs. A major challenge in tissue engineering is to generate arrays of cells with defined structural organization that display appropriate functional properties. Given what is known about cellular responses to physiochemical cues from the surrounding environment, we can build tissue structures that mimic these microenvironments and validate these platforms using both experimental and computational approaches. Tissue generation encompasses many methods and tissue types, but here we review layering cell sheets to create scaffold-less myocardial patches. We discuss surgical criteria that can drive the design of myocardial cell sheets and the methods used to fabricate, mechanically condition, and functionally test them. We also focus on how computational and experimental approaches could be integrated to optimize tissue mechanical properties by using measurements of biomechanical properties and tissue anisotropy to create predictive computational models. Tissue anisotropy and dynamic mechanical stimuli affect cell phenotype in terms of protein expression and secretion, which in turn, leads to compositional and structural changes that ultimately impact tissue function. Therefore, a combinatorial approach of design, fabrication, testing, and modeling can be carried out iteratively to optimize engineered tissue function.

Complications and outcomes of pandemic 2009 Influenza A (H1N1) virus infection in hospitalized adults: how do they differ from those in seasonal influenza?

BACKGROUND: It is unclear whether pandemic 2009 influenza A (pH1N1) infection caused more significant disease among hospitalized adults than seasonal influenza. METHODS: A prospective, observational study was conducted in adults hospitalized with polymerase chain reaction-confirmed pH1N1 infection in 2 acute-care general hospitals from June 2009 to May 2010 (n = 382). Complications and outcomes were described and compared with those in a seasonal influenza cohort (2007-2008, same hospitals; n = 754). RESULTS: Hospitalized patients with pH1N1 influenza were younger than those with seasonal influenza (mean age ± standard deviation, 47 ± 20 vs 70 ± 19 years) and fewer had comorbid conditions (48% vs 64%). The rate of positive immunofluorescence assay results was low (54% vs 84%), and antiviral use was frequent (96% vs 52%). Most patients in both cohorts developed complicated illnesses (67.8% vs 77.1%), but patients with pH1N1 influenza had higher rates of extrapulmonary complications (23% vs 16%; P = .004) and intensive care unit admission and/or death (patient age <35 years, 2.3% vs 0%; 35-65 years, 12.4% vs 3.2%; >65 years, 13.5% vs 8.5%; adjusted odds ratio [OR] 2.13; 95% confidence interval [CI], 1.25-3.62; P = .005). Patients who received antiviral treatment within 96 h after onset had better survival (log-rank test, P < .001). However, without timely treatment, the mortality risk was higher with pH1N1 infection (9.0% vs 5.8% for seasonal influenza; adjusted OR, 6.85; 95% CI, 1.64-28.65; P = .008]. Bacterial superinfection worsened outcomes. CONCLUSIONS: Adults hospitalized for pH1N1 influenza had significant complications and mortality despite being younger than patients with seasonal influenza. Antiviral treatment within 96 h may improve survival.

Cell culture platform with mechanical conditioning and nondamaging cellular detachment.

Cells implanted after injury may remodel undesirably with improper mechanical stimulation from surrounding tissue. Proper conditioning of tissue engineered constructs before implantation can lead to suitable tissue architectures, along with an extracellular matrix (ECM) environment that more closely mimics native tissue. Additionally, cell implantation without bulky polymeric scaffolding is often desirable. Previous researchers have created devices capable of applying mechanical forces to cells (e.g., stretch) but cellular removal from these devices, such as by trypsin, often results in irreversible damage. Conversely, devices are available that can detach intact cells, but these are inelastic, nonstretchable substrates. We have created a cell culture platform that allows for mechanical conditioning and then subsequent nondamaging detachment of those cells. We have modified silicone culture surfaces, to incorporate thermally responsive polymers of N-isopropylacrylamide (NIPAAm) to create an elastic substrate that can also change surface properties with temperature change. A copolymer of NIPAAm and 10percent w/w acrylic acid (AAc) was conjugated to an amine-bonded silicone surface through carbodiimide chemistry. Cells were able to attach to the resulting surfaces at 37 degreeC and showed detachment by rounded morphology at 25degreeC. Following mechanical stretching, cells were still able to spontaneously detach from these modified silicone surfaces with temperature change.

Flow visualization techniques in a mock ventricle supported by a nonpulsatile left ventricular assist device.

Little is known about flow patterns in ventricles supported by continuous flow left ventricular assist devices (LVADs), and valuable information can be obtained with simple flow visualization experiments. We describe the application of several experimental techniques for the in vitro study of ventricular flow patterns (e.g., unsteadiness, vortical motions, stagnation regions) in the presence of a continuous flow LVAD. We used dye streaks, particle paths, and hydrogen bubble techniques to visualize fluid flow in an idealized, static, transparent mock ventricle attached to a Jarvik 2000 continuous flow LVAD. We recorded ventricular flow behavior at various pump speeds while independently adjusting pump flow (by varying the afterload) to emulate in vivo pump flow at various phases of the cardiac cycle. Changes in ventricular flow behavior at different pump flow rates may be of clinical relevance, because continuous flow pumps are extremely sensitive to inflow and outflow pressures and instantaneous pump flow varies significantly at different points throughout the cardiac cycle. Further work is needed to quantitatively compare the flow behavior of different continuous flow devices in a pulsatile ventricular model.

Abundance and location of proteoglycans and hyaluronan within normal and myxomatous mitral valves.

INTRODUCTION: Extracellular matrix changes occur in many heart valve pathologies. For example, myxomatous mitral valves are reported to contain excess proteoglycans and hyaluronan. However, it is unknown which specific proteoglycans are altered in myxomatous valves. Because proteoglycans perform varied functions in connective tissues, this study was designed to identify and localize three matrix-associated proteoglycans, as well as hyaluronan and the hyaluronan receptor for endocytosis, within myxomatous and normal mitral valves. METHODS: Human mitral posterior leaflets (control, n=6-9; myxomatous, n=14-21; mean age, 61 years for all groups) were histochemically stained for proteoglycan core proteins, hyaluronan, and the hyaluronan receptor for endocytosis. Stain intensity was semiquantitatively graded to determine differences in marker abundance between normal and myxomatous valves. The proteoglycans were localized to different regions of the leaflet by correspondence to parallel Movat-stained sections. RESULTS: The proteoglycans decorin, biglycan, and versican were more abundant in myxomatous valves than in normal controls (P<.03). There was a gender effect on proteoglycan presence, but no age-related trends were observed. Hyaluronan and the hyaluronan receptor for endocytosis were distributed throughout all valves. There was no significant difference in hyaluronan between groups, but expression of the hyaluronan receptor for endocytosis was reduced in myxomatous valves compared to normal controls (P<.002). CONCLUSION: Excess decorin, biglycan, and versican may be associated with the remodeling of other matrix components in myxomatous mitral valves. Decreased expression of the hyaluronan receptor for endocytosis in myxomatous valves suggests that hyaluronan metabolism could be altered in myxomatous mitral valve disease. These findings contribute towards elucidating the pathogenesis of myxomatous mitral valve disease and developing potential new therapies.

Mitral valvular interstitial cells demonstrate regional, adhesional, and synthetic heterogeneity.

BACKGROUND/AIMS: Because various regions of the mitral valve contain distinctive extracellular matrix enabling the tissues to withstand diverse mechanical environments, we investigated phenotype and matrix production of porcine valvular interstitial cells (VICs) from different regions. METHODS: VICswere isolated from the chordae (MCh), the center of the anterior leaflet (AlCtr), and the posterior leaflet free edge (PlFree), then assayed for metabolic, growth, and adhesion rates; collagen and glycosaminoglycan (GAG) production, and phenotype using biochemical assays, flow cytometry, and immunocytochemistry. RESULTS: The AlCtr VICs exhibited the fastest metabolism but slowest growth. PlFree cells grew the fastest, but demonstrated the least smooth muscle alpha-actin, vimentin, and internal complexity. AlCtr VICs secreted less collagen into the culture medium but more 4-sulfated GAGs than other cells. Adhesion-based separation resulted in altered secretion of sulfated GAGs by MCh and AlCtr cells but not by the PlFree cells. CONCLUSIONS: VICs isolated from various regions of the mitral valve demonstrate phenotypic differences in culture, corresponding to the ability of the mitral valve to accommodate the physical stresses or altered hemodynamics that occur with injury or disease. Further understanding of VIC and valve mechanobiology could lead to novel medical or tissue engineering approaches to treat valve diseases.