Conventional echocardiographic parameters are currently used in determining the timing for surgery in patients with mitral regurgitation. Since brain natriuretic peptide (BNP) rises in response to ventricular muscle stretch, and is to detect early heart failure, we hypothesized that BNP would be activated in patients with regurgitant valvular heart disease and concomitant left ventricular dilatation. Aim/Objectives: We therefore studied the pattern of changes in NT-pro BNP in patients with chronic severe rheumatic mitral regurgitation who were undergoing mitral valve replacement and compared this with the newer modality of tissue Doppler imaging (TDI). Setting: Patients submitted to surgery were prospectively evaluated over 8 months at Inkosi Albert Luthuli Central Hospital, Department of Cardiology. Controls were obtained from the outpatients' follow-up clinic. Methods: Simultaneous quantification of the severity of mitral regurgitation (MR), left ventricular (LV) end systolic volume (ESV), left atrial (LA) volume and Doppler filling ratios (mitral (E)/annulus (Ea)) were performed at baseline in all patients and was repeated at 1-week and at the six-week follow-up visit in surgical patients. Results: Both groups were similar for age and gender and echo-Doppler parameters in all patients preoperatively except LA size (p< 0.01) and volume (p<0.004) which were more elevated in the surgical group. Mean NT-pro BNP levels were markedly elevated preoperatively (262 pmolll) in all surgical cases compared to controls (57 pmol/l; p=0.0001). NT-pro BNP levels increased further at one week post surgery (395 pmol/l) and subsided at the six week follow-up visit (94 pmol/I). These changes were accompanied by significant reduction in LA (p= 0.003) and LV chamber dimensions (EDD = 0.004) with an increase in the ejection fraction from 42% at one week to 52 % at six weeks. Four patients had abnormally elevated NT-pro BNP levels (>53pmol/l) at the 6-week follow-up visit. A ROC curve was constructed for all variables to separate surgical cases from controls. The area under the curve was highest for NT-pro BNP (sensitivity= 96%, specificity 45 %). Conclusion: 1. There was a significant difference in the left atrial chamber size and volume, as well as Em/Ea (TDI) and NT-proBNP levels preoperatively between the two groups. The lack of a significant difference in the LV parameters between surgical and control groups suggest an almost total reliance on symptoms in deciding the timing of surgery which was reflected by markedly elevated NT-pro BNP in all surgical patients. 2. Postoperatively, there was a significant reduction in LA and LV dimensions. 3. The high false positivity rate for NT-pro BNP suggests that the test is most likely reflecting early LV decompensation in the less symptomatic control patients who rightly need surgery. 4. Tissue Doppler indices had similar sensitivity but low specificity compared to NT-proBNP. 5. Serial estimations of NT-pro BNP may prove useful in selecting patients for surgery. / Thesis (M.Med.Sc.)-University of KwaZulu-Natal, 2007.

Cardiac valves are dynamic, sophisticated structures which interact closely with the surrounding hemodynamic environment. Altered mechanical stresses, including pressure, shear and bending stresses, are believed to cause changes in valve biology, but the cellular and molecular events involved in these processes are not well characterized. Therefore, the overall goal of this project is to determine the effects of pressure and shear stress on porcine aortic valve leaflets biology. Results from the pressure study showed that elevated constant pressure (140 and 170 mmHg) causes significant increases in collagen synthesis. The increases were 37.5% and 90% for 140 and 170 mmHg, respectively. No significant differences in DNA and sGAG synthesis were observed under constant pressure. In the cyclic pressure study, the effects of both pressure magnitude and pulse frequency were studied. With the frequency fixed at 1.167 Hz, collagen and sGAG synthesis increased proportionally with mean pressure level. At a fixed pressure level (80-120 mmHg), collagen and sGAG synthesis were slightly increased by 25% and 14% at 0.5 Hz, respectively. DNA synthesis was significantly increased by 72% at 2 Hz. An experiment combining high magnitude (150-190 mmHg) and high frequency (2 Hz) demonstrated significant increases in collagen and sGAG synthesis (collagen: 74%, sGAG: 56%), but no significant changes in cell proliferation. Shear levels ranging from 1 to 80 dyne/cm2 were studied. Scanning electron microscopy results indicated that 48 hrs exposure to shear stress did not alter the circumferential alignment of endothelial cells. Collagen synthesis was significantly enhanced at 9 and 25 dyne/cm2, but not different from static controls under other shear conditions. Leaflets denuded of the endothelium were exposed to identical shear stress and showed very different responses. Collagen synthesis was not affected at any shear levels, but sGAG content was increased at shear of 9, 25 and 40 dyne/cm2. Further studies showed that the increases in collagen synthesis under pressure or shear stress was concurrent with a decline in the expression and activities of cathepsins L and S. This converse relationship between collagen synthesis and cathepsin activity indicated that cathepsins might be involved in valvular ECM remodeling.

Thesis (M.S.)--University of Melbourne, Dept. of Surgery (St.Vincent's Hospital),Faculty of Medicine, Dentistry and Health Sciences, 2006. / Typescript. Includes bibliographical references (leaves 118-141).

The gold standard treatment for patients with AVD is surgical replacement of the aortic valve with either mechanical or fixed tissue prostheses. These implants have a limited lifespan and are associated with serious adverse events. Patient autologous tissue engineered heart valves (TEHVs) offer a solution. Vital to the development of a TEHV is determining a source of donor tissue(s) that most closely mimics the native valve tissue. In pursuit of determining an alternative cell source for patient autologous TEHVs we compared a number of phenotypic and genotypic characteristics of atrial fibroblasts, dermal fibroblasts and differentiated bone marrow-derived progenitor cells (BMCs) and made a comparison to valvular interstitial cells (VICS). We demonstrate that while VICs share some phenotypic similarities with fibroblasts and BMCs, they also possess unique characteristics and demonstrate differential mRNA expression of key regulatory pathways that may influence their phenotype. / October 2016

According to statistics from the American Heart Association, valvular heart disease directly leads to about 20,000 deaths a year and contributes to an additional 50,000. While significant advancements have been made in the treatment options available for valvular heart disease, complications still occur. For this reason, the future of valvular heart disease treatment lies in understanding the physiology of the heart valve, and subsequently bioengineering a valve from one's own tissue to mimic native valve processes. Valvular interstitial cells (VICs) are the major cell type populating the valve matrix. In the inactive fibroblast-like state, these cells are responsible for extracellular matrix deposition. Activated VICs display a myofibroblast morphology characterized by the expression of alpha smooth muscle actin and are responsible for valve maintenance and repair. The activation of VICs is hypothesized to be stimulated by mechanical tension, which, in the presence of TGF-â1 allows the complete differentiation of VICs from the inactive to the active form. However, little is known about the potential for reversal or dedifferentiation from the active to inactive state. The purpose of this study was to determine whether substrate stiffness, the mechanical tension hypothesized to initiate VIC activation, modulates alpha smooth muscle actin expression in the presence and absence of TGF-â1. To mimic conditions found in vivo, substrates were varied from physiologic to pathological stiffness levels. Results showed that when freshly isolated VICs are cultured in the presence of serum, alpha smooth muscle actin expression increased on all substrate stiffnesses. In TGF-â-free medium, there was an apparent increase on all stiffness levels as well, but a statistical significance between groups could not be demonstrated. Immunoblots used to detect TGF-â1 showed that intracellular TGF-â1 was upregulated in VICs cultured in the presence of serum compared to those cultured in TGF-â-free medium. Taken together, these results suggest that freshly isolated VICs become activated, as indicated by increased expression of alpha smooth muscle actin, on all substrate levels in the presence of serum. It also appears as though unknown factors which are present in serum are required to stimulate significant autocrine production of TGF-â1. To determine whether VICs which had transitioned to the myofibroblast phenotype had the ability to dedifferentiate, cells were cultured on polystyrene for a minimum of four days then replated on substrates of varying stiffness. Analysis of alpha smooth muscle actin expression showed that, in the presence of serum and when replated on all of substrates used, alpha smooth muscle actin expression decreased, suggesting that these cells indeed have the potential to dedifferentiate. A change in cell morphology to a more rounded phenotype as well as the loss of visible stress fibers further supported this possibility. These studies represent a unique approach to studying phenotypic differentiation of valvular interstitial cells. Using acrylamide substrates of varying stiffness, and growth factor free media, we have shown that by altering substrate stiffness, changes in alpha smooth muscle actin expression consistent with differentiation and dedifferentiation can be induced. This potential for dedifferentiation suggests that in engineering the next generation of bioartificial valves, it may be possible to use the patient's own cells to seed the manufactured scaffold. This would avoid complications associated with current treatments, including immune rejections.

A Tissue Engineering (TE) approach to heart valve replacement has the aim of producing an implant that is identical to healthy tissue in morphology, function and immune recognition. The aim is to harvest tissue from a patient, establish cells in culture from this tissue and then use these cells to grow a new tissue in a desired shape for the implant. The scaffold material that supports the growth of cells into a desired shape may be composed of a biodegradable polymer that degrades over time, so that the final engineered implant is composed entirely of living tissue. The approach used at Swinburne University was to induce the desired mechanical and functional properties of tissue and is to be developed in an environment subjected to flow stresses that mimicked the haemodynamic forces that natural tissue experiences. The full attainment of natural biomechanical and morphological properties of a TE structure has not as yet been demonstrated. In this thesis a review of Tissue Engineering of Heart Valves (TEHVs) is presented followed by an assessment of biocompatible materials currently used for TEHVs. The thrust of the work was the design and development of a Bioreactor (BR) system, capable of simulating the corresponding haemodynamic forces in vitro so that long-term cultivation of TEHVs and/or other structures can be mimicked. A full description of the developed BR and the verification of its functionality under various physiological conditions using Laser Doppler Anemometry (LDA) are given. An analysis of the fluid flow and shear stress forces in and around a heart valve scaffold is also provided. Finally, preliminary results related to a fabricated aortic TEHV-scaffold and the developed cell culture systems are presented and discussed. Attempts to establish viable cell lines from ovine cardiac tissue are also reported.

Heart valve structures open and close during the cardiac cycle to provide unidirectional blood flow through the heart, critical for efficient cardiovascular function. Valve dysfunction results in either incomplete opening or incomplete closure of the valve. Both types of valve dysfunction decrease efficiency of blood flow, increasing the load on the myocardium and leading to secondary heart disease such as pathological hypertrophy and heart failure. There are currently no effective treatments to prevent or slow the progression of valve disease, and there are no pharmacological treatments for advanced valve disease. Although most valve disease is associated with aging, increasing evidence suggests that valve disease often has origins in development. Congenital valvuloseptal defects affect many newborns, ranging from life-threatening malformations requiring immediate repair to more subtle, often undiagnosed defects that increase susceptibility to valve disease later in life. Therefore, an improved understanding of the mechanisms of heart valve formation and maintenance of adult valves may serve as an important step in improving valve disease treatment options. In this work, the mechanisms of normal valve development and the role of Sox9 in developing and mature valves are further studied. The temporal and spatial expression of extracellular matrix genes and proteins are examined throughout normal murine valve development. Sox9 function in the processes of valve development and valve maintenance is examined using mouse models of conditional Sox9 loss-of-function. Heart valve phenotypes in mice with reduced Sox9 function are examined throughout development and in adult mice with resultant calcific valve disease. The possible causative mechanisms of calcific valve disease in mice with reduced Sox9 function are further investigated by identification of novel possible targets of Sox9 transcriptional regulation. Together these studies improve our understanding of heart valve development, characterize a model of heart valve calcification with genetic etiology, and identify and characterize novel targets of Sox9.

Bileaflet mechanical heart valves (BMHVs) are prone to thrombus formation in the hinge region due to high shear stress combined with stagnation regions. This thesis research addresses the hypothesis that models that isolate and mimic BMHV hinge geometries can be used to quantitatively characterize procoagulant potential using a novel in vitro blood flow system. Furthermore, these results can be correlated with digital particle image velocimetry (DPIV) measurements detailing flow fields for the same models. The significant findings were that: 1) recalcification of recirculating citrated blood markedly increases the magnitude of thrombus forming reactions and the sensitivity for their detection; 2) platelet activation, and the presence of adequate platelet numbers are essential for the activation of coagulation under conditions of high shear; and 3) thrombin formation can be inhibited by blocking the platelet receptors that facilitate platelet aggregation. The DPIV studies give some insight into why different channel geometries resulted in varying propensities for coagulation. The channel geometries with abrupt changes in diameter induced significantly higher levels of TAT and also formed jets that were subject to increased entrainment of the stagnant fluid in the chamber. This entrainment enables more mixing of the shear-activated platelets with the surrounding flow, which can propagate the coagulation cascade, thus increasing the chance for thrombus formation. The influence of abrupt changes in diameter was also evident in the BMHV human blood studies. The MP valve, which has a tortuous hinge pathway, induced significantly more TAT formation than the SJM Standard valve with a smoother hinge channel. Thus, BMHV hinge geometry should be as smooth and free of diameter changes as possible to eliminate stagnation regions that enable activated platelets to congregate and propagate the coagulation cascade. Leakage gap width also had a significant effect not only on procoagulant potential but also on platelet activation. Both the low and high leaker prototype valves had significantly higher levels of platelet activation compared to the SJM Standard valve, but only the low leaker valve demonstrated a higher propensity for coagulation. Thus, to minimize both platelet activation and thromboemboli formation, an optimal gap width should be maintained for BMHVs.

Thesis (M.S.)--Chemical Engineering, Georgia Institute of Technology, 2003. / Sambanis Athanassios, Committee Member ; Sotiropoulos Fotis, Committee Member ; Yoganathan Ajit, Committee Chair. Includes bibliographical references (leaves 239-243).

Thesis (M.S.)--Chemical Engineering, Georgia Institute of Technology, 2004. / Sambanis Athanassios, Committee Member ; Sotiropoulos Fotis, Committee Member ; Yoganathan Ajit, Committee Chair. Includes bibliographical references (leaves 239-243).