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fbtwitterlinkedinvimeoflicker grey 14rssslideshare1
Languages: English
Types: Doctoral thesis
Subjects: UOW2
As a result of the enormous clinical need, cardiac tissue engineering has become a prime\ud focus of research within the field of tissue engineering. In this project, Poly(3-\ud hydroxyoctanoate), P(3HO), a medium chain length (mcl-PHAs) biodegradable,\ud biocompatible and elastomeric polyhydroxyalkanoate, was studied as a potential material for\ud cardiac tissue engineering. Mcl-PHAs are an alternative source of polymers produced by\ud Pseudomonas sp. As Gram-negative bacteria, Pseudomonas sp. contains lipopolysaccharides\ud in the membrane, which are co-purified with PHAs and may cause immunogenic reactions.\ud This limits the biomedical applications of the mcl-PHAs in several cases. In this work, the\ud Pseudomonas mendocina PHA synthase gene (phaC1) was expressed in the LPS free, GRAS,\ud Gram-positive microorganism, Bacillus subtilis so as to produce LPS-free mcl-PHAs. Our\ud results showed that the recombinant Bacillus subtilis containing the phaC1 gene produced\ud poly(3-hydroxybutirate), P(3HB), with a maximum yield of 32.3 % DCW, an unexpected\ud result. This result thus revealed the unusually broad substrate specificity of the PHA synthase\ud from P. mendocina, which is able to catalyse both medium and short chain length PHAs\ud biosynthesis depending on the metabolic pool available in the host organism. Sequence\ud comparison of this PHA synthase with stringent mcl-PHA synthases revealed possible\ud residues influencing the substrate specificity of PHA synthases.\ud As studies on mcl-PHAs remain limited mainly because of the lack of availability of mcl-\ud PHAs in large quantities, the capacity to scale-up P(3HO) production from 2 L to 20 L and\ud 72 L pilot plant bioreactors, based on constant oxygen transference, was studied.\ud The interaction of freshly isolated rat cardiomyocytes with the P(3HO) polymer, during\ud contraction, was studied when cells were stimulated at a range of frequencies of electrical\ud pulses or calcium concentrations. These results showed that P(3HO) did not have any\ud deleterious effects on the contraction of adult cardiomyocytes. P(3HO) cardiac patches nonporous,\ud porous or with P(3HO) electrospun fibres deposited on their surface were developed.\ud Our results showed that the mechanical properties of the final constructs were close to that of\ud the cardiac structures, with a Young’s modulus value of 0.41±0.03 MPa. Myoblast (C2Cl2)\ud cell proliferation was studied on the different constructs showing an enhanced cell adhesion\ud and proliferation when both porous and fibrous structures were incorporated together.\ud Finally, for further enhancement of the cardiac patch function, VEGF and RGD peptide were\ud incorporated. Results obtained in this project showed that the P(3HO) multifunctional cardiac\ud patches were potentially promising constructs for efficient cardiac tissue engineering.
  • The results below are discovered through our pilot algorithms. Let us know how we are doing!

    • Figure 6.1. Static strain vs. stress profile of the P(3HO) porous films. The initial slope and the maximum elongation are indicated with a black line.......................................................
    • Figure 6.2. Surface roughness analysis of two representative samples of P(3HO) porous films created by the particle leaching method........................................................................
    • Figure 6.14. Surface roughness analysis of two representative samples of A) P(3HO) neat film modified with 750nm fibers and B) P(3HO) porous films modified with 750nm fibers.........................................................................................................................................   163 Figure 6.15. Concentration of proteins adsorbed on the surface of neat P(3HO) films, porous P(3HO) films, Neat P(3HO) films + 750 nm fibers and Porous P(3HO) films + 750 nm fibers...................................................................................................................................
    • Figure 6.16. The % cell proliferation of C2C12 cell line on neat P(3HO) films containing porous structures, fibrous structures and both fibrous and porous structures...........................   165 Figure 6.17. SEM images of C2C12 cells at 24 hr on A), B) and C) neat P(3HO) and C), D) and E) porous films modified with 750 nm fibers....................................................................   166 Figure 7.1. FTIR spectra of as synthesized P(3HO) polymer vs. aminated P(3HO) (P(3HO)- NH2).........................................................................................................................................
    • Figure 7.2. FTIR spectra of P(3HO) polymer vs. P(3HO)-RGD.............................................
    • Figure 7.3. Surface wettability properties of P(3HO) vs. P(3HO)-RGD cardiac patches.......
    • Figure 7.9. Thermal profile of (A) unloaded P(3HB) microspheres and (B) VEGF loaded P(3HB) microspheres.............................................................................................................................. 183   Table 1.1. Mechanical properties of materials proposed for myocardial tissue engineering........
    • Adams H, Irving G, Koeslag H, Lochner D, Sandell R and Wilkinson C (1987). Betaadrenergic blockade restores glucose's antiketogenic activity after exercise in carbohydratedepleted athletes. J. Physiol. Lond. 386, 439-454.
    • Akaraonye E, Keshavraz T, Roy I (2010). Production of polyhydroxyalkanoates: the future green materials of choice. J. Chem. Technol. Biotechnol. 85: 732-743.
    • Akaraonye E, Moreno C, Knowles J, Keshavarz T, Ipsita Roy I (2012). Poly(3- hydroxybutyrate) production by Bacillus cereus SPV using sugarcane molasses as the main carbon source. Biothecnol. J. 7(2): 293-303.
    • Alvarez H, Kalscheuer R, Steinbuchel A (2000). Accumulation and mobilization of storage lipids by Rhodococcus opacus PD630 and Rhodococcus ruber NCIMB 40126. Appl.
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    • Ikada Y (2006). Challenges in tissue engineering. J. R. Soc. Interface. 3: 589-601.
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  • Discovered through pilot similarity algorithms. Send us your feedback.

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