Preparation and characterization of a hydrogel containing anti-VEGF (bevacizumab) liposomes to prevent bone tissue repair at injured growth plates.
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1.7.1.1Effects of Chitosan on Cartilage Tissue Repair20
1.7.2Bevacizumab Encapsulated Liposome dispersed in Chitosan / ß-Glycerophosphate Thermosensitive Hydrogel21
chapter 2. AIMS AND HYPOTHESES22
Chapter 3 MATERIALS AND METHODS 23
3.1Chitosan Based Thermosensitive Hydrogel23
3.1.1Thermosensitive Solutions Preparation24
3.1.2Determination of Gelation Time:24
3.2Rheology25
3.3Preparation of Bevacizumab-Encapsulated Liposomes25
3.3.1Determination of Encapsulation Efficiency26
3.4Incorporation of Bevacizumab-Encapsulated-Liposomes into a Hydrogel (liposomal gel) and In vitro Release Analysis:27
3.5Enzyme-Linked Immunosorbent Assay27
3.6In Rats a Drill-Hole Growth Plate Injury Model28
3.6.1Animal Trial Design28
3.6.2Surgical Procedure29
3.6.3Specimen Collection29
3.6.3.1Blood Collection29
3.6.3.2Tissue Collection29
3.7Bone Histology30
3.7.1Bone Preparation30
3.7.2Micro-Computed Tomography Scans (µ-CT) and Bone Parameters Analysis31
3.7.3Histomorphological and Immunohistochemcial Analysis of Tibial:31
3.8Total RNA Isolation and Real-Time Quantitative Polymerase Chain Reaction:32
3.8.1RNA Extraction and cDNA Synthesis33
3.8.2Analysis of the expression of genes by Quantitative Real Time Polymerase Chain Reaction (RT-PCR)33
3.9Statistical Analysis34
Chatper 4 RESULTS35
4.1Characteristics of Chitosan/ß-Gp Hydrogels35
4.2Characteristics of Liposome37
References40
List of figures
Figure 1: Structure and function of different histologic zones in the growth plate9
Figure 2: Salter-Harris Classification System11
Figure 4: Picture33
Figure 5: Characteristics of Chitosan/ß-Gp Hydrogels (n=3)34
Figure 6: Viscosities of chitosan/ß-Gp hydrogels (chitosan: 6% (w/v), ß-Gp Salt: 10% (w/v), and chitosan: 10% (w/v), ß-Gp salt: 10% (w/v)) as a function of time at 37°C.35
Abbreviations
ANOVA
one-way analysis of variance
SEM
Standard error of mean
MSCs
Mesenchymal stem cells
OCN
Osteocalcin
Runx2
Runt-related transcription factor-2
TNF-a
Tumour necrosis factor-alpha
Summary
For the bone healing, bone development, and establishment, and growth of bone tumours, Angiogenesis is considered to be an enormous supplier. Growth plates are mainly situated at the end of all long bones, responsible for the lengthening of a child's skeleton via a tightly regulated process called endochondral ossification. The bone growth and function of the growth plate comprises of three zones, the resting zone, the differentiation, and proliferative zone and provisional and hypertrophic calcification zone. The bone tumours, impaired fracture healing therapy and skeletal defects ...