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Title Page
Abstract
Contents
I. Introduction 11
1.1. Overview of 3D cell printing in bone tissue engineering 11
II. PART 1. Evaluation of varying alginate bio-inks for suitable 3D bio platform 13
2.1. Introduction 13
2.2. Materials and methods 15
2.2.1. Preparation of alginate based bio-inks 15
2.2.2. Rheological measurements of the alginate bio-inks 15
2.2.3. Fabrication of 3D porous hydrogel matrices with various alginate bio-inks 15
2.2.4. In vitro cell culture and cell printing 15
2.2.5. Cell proliferation assay 16
2.2.6. Live and dead cell assay 16
2.2.7. Statistical analysis 16
2.3. Result and discussions 18
2.3.1. Printing of various alginate bio-inks 18
2.3.2. In vitro cell culture studies using alginate bio-inks 26
III. PART 2. The effects of 3D hydrogel complex with sulfated alginate on affinity based growth factor delivery and interactions for bone tissue engineering 30
3.1. Introduction 30
3.2. Materials and methods 32
3.2.1. Synthesis of alginate sulfate and novel bio-inks 32
3.2.2. Preparation of novel bio-inks 32
3.2.3. Rheological measurements of the alginate and alginate sulfate bio-inks 32
3.2.4. Fabrication of 3D hydrogel complex with alginate sulfate 32
3.2.5. In vitro cell culture and cell printing 33
3.2.6. Effect of the BMP-2 in hydrogel complex 33
3.2.7. In vitro cytotoxicity of alginate sulfate and cell viability & live and dead assay 34
3.2.8. ALP activity analysis and vonkossa staining 34
3.2.9. Analysis equipment 35
3.3. Results and discussions 36
3.3.1. Structural characterization 36
3.3.2. Printability of novel bio-inks with alginate sulfate 40
3.3.3. Effect of BMP-2 into 3D hydrogel complex 43
3.3.4. In vitro behaviors of alginate sulfate based bio-inks with BMP-2 in 3D hydrogel 46
IV. Conclusions 50
References 51
Table II.1. Properties of alginate bio-inks with different compositions and varying concentrations 20
Table III.1. The varying composition of alginate and alginate sulfate with BMP-2. 37
Table III.2. The calculated sulfur contents of synthesis product. 37
Figure II.1. Schematic diagram of the three-dimensional (3D) bio-printing process using different bio-inks... 21
Figure II.2. Photographs of alginate bio-inks of various compositions under different conditions. 22
Figure II.3. Mechanical properties of various alginate bio-inks 23
Figure II.4. Printability of alginate bio-inks. 24
Figure II.5. Photographs of the 3D porous hydrogel matrices with Ink 2, Ink 5, and Ink 8. 25
Figure II.6. Viability of the cells printed using the various alginate bio-inks (Ink 2, Ink 5, and Ink 8) up to... 28
Figure II.7. Fluorescence live/dead images of cells printed in the alginate bio-inks (Ink 2, Ink 5, and Ink 8)... 29
Figure III.1. Schematics of the 3D novel hydrogel complex. 38
Figure III.2. Preparation schematics of the sulfation of uronic acids in alginate. 39
Figure III.3. Mechanical properties of various novel bio-inks 41
Figure III.4. Printability and fabrication of the 3D hydrogel constructs using novel bio-inks. 42
Figure III.5. The effect of different concentrations of BMP-2 in 2D and cumulative amount of BMP-2 from... 45
Figure III.6. The cell proliferation of different ratios of novel bio-inks and live and dead staining assay up... 48
Figure III.7. The level of ALP activities and mineralization of the varying 3D novel hydrogel complex... 49
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