1,721,074 research outputs found
Formation of miniaturized 3D tumour spheroids for drug screening
No full textCancer is one of the most life-threatening diseases and has affected many in the world. Currently, anti-cancer drugs are being tested by in vitro cell- based study and animal testing before drugs can move on to clinical trials. Traditional 2D cell culture is widely used for in vitro study due to the lower cost and simplicity. However, in recent years, 2D cell culture is increasingly reported to have inaccurate and misleading data. This led to an increase interest in developing 3D culture platform for the screening of anti-cancer drugs. Among the 3D cell culture models, multi-cellular spheroids are commonly used as they accurately mimic tumour architecture. There are many existing methods to fabricate multi-cellular spheroids but lacked high throughput and consistency. In this study, co-axial electrospray will be used to fabricate hydrogel spheroids with core-shell structure and its parameters will be optimised in order to obtain the desired size and consistent shape. Effect of voltage, hardening bath, needle size, total flow rate and flow rate ratio will be studied based on the morphology of spheroids and its consistency. With the optimised parameters, MCF-7 and L929 cells will be encapsulated in core and shell compartment respectively to form multi-cellular spheroids as proof of concepts. Cell viability and cellular spheroids formation will be characterized with fluorescence staining.Bachelor of Engineering (Materials Engineering
Modulation of human mesenchymal stem cells towards neuronal differentiation via physical cues from scaffold design
No full textThe influential role of physical cues from microenvironment in directing stem cell differentiation has been recognition. In spite of extensive studies reported in modulation of stem cell fate via various physical cues, little has been explored in neuronal differentiation event with either single or combination of these physical cues.Doctor of Philosophy (MSE
Optimization of drug release profile for 2D and 3D electrospun fibres for liver fibrosis
No full textThe increasing trend of liver-related diseases has resulted in numerous cases of death and is
currently highly ranked in terms of deadly diseases. Therefore, more focus has been put on
finding a solution for the various type of liver diseases. All liver diseases progress through
liver fibrosis defined by scar tissue formation. With the fast development of health-related
researches, many methods have been proposed to overcome the symptoms of liver fibrosis.
However, there are many limitations to the present methods. One of the methods that had
recently come into the spotlight is the use of electrospinning to fabricate fibre to prepare
scaffolds for drug delivery.
Our current work investigates the effects of electrospinning on the fabricated fibre and the
process of encapsulating Decorin in the electrospun fibre for release study and full
encapsulation study. There is also the use of BSA as the protein to optimise the encapsulation
percentage within the electrospun fibre. Characterization method like Micro BCA Assay,
ELISA Assay, spectrophotometry and scanning electron microscopy (SEM) will be
employed.Bachelor of Engineering (Materials Engineering
Synthesis and characterization of biodegradable elastomers for soft tissue engineering
No full textHerein, the synthesis and evaluation of a new group of biodegradable elastomeric polyesters,
poly(1, 12-dodecandiol-co-malic acid) is performed. The main objectives of this project are to
design a malic acid-based aliphatic polyester material with desirable modulus, elasticity and
degradation rate.
Two-dimensional poly(1, 12-dodecandiol-co-malic acid) scaffolds were prepared by reacting
malic acid with different diol ratios under nitrogen flow to form pre-polymers. Pre-polymers
were then post-polymerized at 140oC and 160oC under vacuum (30 kPa) for various days.
8 scaffolds with different acid to diol ratios were synthesized and post-polymerization under
different conditions and then characterized. All scaffolds demonstrated good thermal stability.
Seven out of eight materials passed the in vitro cytotoxicity test with the Young’s modulus
ranged from 1.13±0.65MPa to 4.04±0.78MPa. Elongation was as high as 1070.48±265.14%. The
mechanical properties and degradation rate could be controlled by choosing different acid to diol
ratios as well as by modulating the post-polymerization conditions.
In addition, this study would like to propose PDDM1-1,21,0 and PDDM1-2,36,0 for further
investigation as they showed desirable thermal and mechanical properties, degradation rate and
excellent results for cell growth.Bachelor of Engineering (Materials Engineering
Polycaprolactone scaffold fabricated via selective laser sintering for cardiac tissue engineering
No full textCardiac tissue engineering has been emerged as one of the promising area in the
biomedical engineering to repair or replace damaged tissue. It focused on growing
cells by using temporary three – dimensional biomaterial scaffold acting as a support
to guide cell proliferation. An advanced scaffold fabrication technique such as Rapid
Prototyping (RP) processed has been utilized to overcome the limitation of the
conventional base method. Selective Laser Sintering was utilized to fabricate tissue
engineering scaffolds due to its high reproducibility and good control of pore size.
Polycaprolactone (PCL) is a semicrystalline polymer that has been proven as a
biocompatible and bioresorbable material for tissue engineering applications. In this
report, it is aimed to show that by varying the sintering parameter of the SLS system,
mechanical properties in the tensile mode of the PCL scaffold can be changed to
obtain low stiffness. The SLS parameters named laser power, laser scan speed and
part bed temperature were varied. The laser power between 1 – 5 W, the laser scan
speed between 100 – 300 inch/s and constant part bed temperature of 50 °C were
tested. The Young’s modulus of PCL scaffold varied from 3 MPa to 42 MPa and the
maximum elongation varied from 12 % to 162 %. Fracture surface of the scaffold
after break was also investigated to know the failure mode and avoid catastrophic
failure. Although it was shown that SLS is feasible to fabricate PCL scaffold, the
mechanical properties has not reached the target in order of tens kPa. Thus, further
process need to be carried out to fabricate the scaffold that match with the tensile
stiffness of the native myocardium.Bachelor of Engineering (Materials Engineering
Bio-ink properties of gelatin methacrylamide
No full textIn 2012, Singapore had 456 patients on the kidney transplantation waiting list and the shortage of organ transplantation drives the need for new regeneration therapy. It is widely believed that high precision 3D Bioprinting will be a new technology to improve regeneration therapy. The key challenge of the 3D Bioprinting is subjected to the availability of appropriate printable and cell compatible materials as bio-ink. Gelatin methacrylamide (GelMA) is a hydrogel material that has excellent cell viability and cytocompatibility which can potentially be used as a bio-ink material. However, limited research have been done on the feasibility of using GelMA as a bio-ink material. The objective of this study is to investigate the feasibility of using GelMA as a printable bio-ink material. The feasibility in this study of bio-ink will only cover the printability. GelMA that used in this study was chemically modified from gelatin Type A and Type B and was carried out using different percentages of degree of substitution (% of DS) and different concentrations. This study involved rheology studies and 3D Bioprinting. Rheology studies covered flow behaviour of the GelMA solutions under various shear rates and various temperatures. Frequency sweep test was also conducted to study the viscoelasticity of GelMA. Results obtained from rheology studies showed GelMA are shear thinning materials and their viscosities are dependent on % of DS, concentration and temperature. 20% weight per volume (% w/v) GelMA 2.2 Type A and Type B were selected as the test materials for 3D Bioprinting. These two samples have higher storage modulus properties (G’>G”). From 3D Bioprinting, 20 % w/v GelMA 2.2 Type A shows better resolution and shape fidelity than Type B. In summary, GelMA is feasible to be used as a bio-ink material as it exhibits the component of printability such as shear thinning properties and higher storage modulus properties (G’>G”). Due to machine limitation, not all GelMA samples could be studied in this project. Future studies on GelMA could involve optimising other printing parameters to explore its printing possibility.Bachelor of Engineering (Materials Engineering
Synthesis and cell culture on 2D and 3D biodegradable elastomer
No full textThe objective of this project is to explore the optimum curing time and reactants ratio in the synthesis of a biodegradable elastomer for myocardial tissue engineering. In line with this objective, a 3D biodegradable elastomeric scaffold that supports cell proliferation and differentiation for use in myocardial tissue engineering.Bachelor of Engineering (Materials Engineering
Cell-materials interaction and its implications on stem cell fate
No full textHuman mesenchymal stem cells (hMSCs) continue to attract prominence in tissue
engineering due to their immunosuppressive property, self renewability, and
multi-lineage differentiation potential. While emphasis has always been placed on
inducing differentiation of hMSCs into the targeted cells of interest by biological
methods in the past, biophysical methods such as mechanical stimulation and
cellular morphology are now emerging at a rapid pace. In the physical methods,
the interaction between the cell and material is of utmost importance but
systematic study of this interaction is still limited to date. In this investigation,
cell-materials interaction represented by the development of focal adhesion (FA)
was studied systematically and the relationship between FA and stem cell
differentiation (especially myogenic differentiation) was investigated. Results
indicated that the FA development of hMSCs can be modulated by
micropatteming but there was a synergistic effect between FA regulation and
matrix stiffness. Elongated FA could be modulated on the substrates with
intermediate stiffness ((polydimethylsiloxane (PDMS) with a stiffness of 12.6 kPa
and polyacrylamide (PA) gel with a stiffness of 10.2 kl'a) and with collagen type I
(COLI) as the inking protein. hMSCs with elongated FA showed specific
myogenic differentiation at both transcription and translation levels compared with
cells with dense FA and those in control group. Mechanistic study showed that
elongated FA recruited integrin ~3 clusters, activated RhoA signaling pathway,
aligned stress fibers, and increased cellular tension via activation of RhoA
signaling pathway and up-regulation of myosin light chain kinase (MLCK).
Moreover, the interplay of cell shape and FA on hMSCs differentiation was investigated and the key role of elongated FA in driving hMSCs myogenic
differentiation was confirmed. Similar shaped hMSCs with different FA
morphology and distribution were induced using micropatterning technique and it
was found that the elongated FA was more supportive of myogenic differentiation.
Collectively, this study demonstrates a novel chemical/biological free and feasible
method of inducing myogenic differentiation of hMSCs and improves our
understanding of the micropatterning platform in affecting the differentiation.DOCTOR OF PHILOSOPHY (MSE
Engineering 3D scaffolds with iPSCs towards regeneration of cardiac tissues
No full textThis work focuses on engineering 3 dimensional (3D) fibrous hybrid scaffolds with human induced pluripotent stem cells (hiPSC) for cardiac tissue engineering (CTE). Natural materials are highly bioactive, yet they are limited by their high batch-to-batch variability, and their poorly understood bioactivity mechanisms, particularly, in affecting stem cell fate. Conversely, commonly used synthetic materials, while offering good control over different parameters, generally lack the suitable bioactivity for cellular interactions. In this work, we propose comparing and combining natural and synthetic scaffolds to enjoy the advantages of both platforms while circumventing their inherent limitations. The resulting 3D scaffolds will improve understanding of the cell material interactions and might produce a potential treatment for cardiac regeneration.
One ideal group of natural biomaterials can be obtained by decellularization, yielding tissue specific bioactive, and cell supportive scaffolds of either solid, or liquid forms. Our lab has isolated a porcine cardiac extra cellular matrix (pcECM), in both solid and liquid forms, which preserves the 3D architecture of the heart ECM, while maintaining the bioactivity for cellular interaction. Nevertheless, the exact contribution of such pcECM to stem cell and tissue specific lineage commitment, and the possible mechanisms governing such bioactivity remain largely unknown. The complexity of the pcECM composition and 3D architecture hinder our ability to discriminate between different effectors and their resulting individual or combined effects. Hence, it is necessary to generate synthetic 3D biomimetic scaffolds with controllable architecture and bioactivity profiles that would enable the study of various components in a modular way.
Electrospinning is an accessible and inexpensive means to fabricate fibrous matrices but the fundamental limitation with traditional electrospinning is that the scaffold produced is usually two dimensional (2D) dense mats rather than 3D porous structures. Here, we improvised the liquid-collector of electrospinning to fabricate 3D fibrous scaffolds with high porosity. Though this ECM mimicking synthetic polymer scaffolds offer high reproducibility, they generally lack the bioactivity inherent to natural ECM biomaterials. Thus, different approaches have been used to confer bioactivity to synthetic materials, such as natural materials (e.g., short bioactive peptides) either on the surface or within the bulk, pre-culturing the scaffold with ECM producing cells (e.g., fibroblasts, and mesenchymal stem cells), and physical and/or chemical surface modifications.
Our work aims to obtain 3D composite scaffolds (3DCS) with ECM mimicking synthetic ultra-structures, and tissue specific biochemical cues by fabricating 3D electrospun polymeric scaffolds and functionalizing them with liquidized pcECM. The 3DCS produced were studied in comparison to the natural pcECM highlighting the roles of architecture, biochemical composition, and various combinations thereof, in affecting the function and fate of seeded human multi and pluri-potent stem cells. hiPSCs were used in this study as an ideal model cell with clinical relevancy, given their possible autologous sourcing, and their ability to differentiate into all cardiac cell types, in particular, beating cardiomyocytes (hiPSC-CM). We hypothesized that, bioactive 3D scaffolds (pcECM and/or 3DCS) that maintain a balance and cooperation between architectural and biochemical signals, are needed to initialize differentiation of hiPSC towards cardiac lineages.
Our results show that the pcECM can be mimicked by wet electrospinning of poly lactide-co-glycolide (PLGA), and poly lactide-co-ε-caprolactone (PLCL). However, based on the evaluated properties, and reproducibility of the 3D synthetic scaffolds, only 3D PLGA exhibited adequate profile and was therefore used for further studies. After modification with pcECM gel, the 3DCS displayed similarities with pcECM in terms of morphology, chemistry, biochemical composition. The 3DCS also displayed cardiac relevant mechanical properties and did not elicit any immunogenicity in vitro. 3DCS also displayed the ability for cellular attachment and growth under static conditions when human mesenchymal stem cells (hMSCs) were used as model cells. hiPSC-CM seeded 3DCS maintained CM viability, beating functionality and phenotypic identity for two weeks as evaluated by protein expression. Moreover, the scaffolds’ microenvironment supported the calcium handling ability. Finally, hiPSCs seeded on these scaffolds, differentiated into cardiac lineage cells spontaneously without the addition of any external factors or molecules, asserting the role and importance of a tissue specific biochemical microenvironment for cardiac applications. Taken together, our results here contribute to the understanding of how the biology and architecture of the pcECM can affect and determine the fate of the seeded hiPSCs. This knowledge is relevant not only for basic research but also for possible CTE applications.Doctor of Philosoph
Development of biomimetic 3D cancer platform for drug testing
No full textCancer accounts for most of the deaths in the world and it is mainly due to metastasis that makes treatment of cancer difficult. Epithelial-to-Mesenchymal Transition plays an important role in tumour metastasis as well as in tumour recurrence, the conversion of epithelial cells to mesenchymal cells can result in the acquisition of migratory and invasive properties. There are numerous studies on how stiffness of extracellular matrix, signals from the environment, type of extracellular matrix and the presence of integrins can affect cell behavior and migration. However, there are limited studies looking at how the physical and material characteristics of the microenvironment affect the Epithelial-to-Mesenchymal Transition and Mesenchymal-to-Epithelial Transition of highly invasive tumor cell lines. Conventional 2D cell culture models have also been proven to be inadequate in mimicking the environment of 3D native tissues. In response, a 3D culture model has been gradually gaining popularity among research groups due to its better representation of the microenvironment of living tissues. Therefore in this study, we propose a 3D biomimetic culture model constructed with 5% PEG 5% Gelatin matrix and collagen I matrix to study the EMT and MET transition in the human body environment. MDA-MB-231 cancer spheroids were then placed in the PEG + Gelatin matrix in a polyethylene terephthalate plastic construct before the addition of a collagen matrix layer at the top. From this study, we had found out that the design of the construct was suitable to mimic the EMT and MET phenomenon. However, there were a few limitations of the culture model. First, spheroids were not fully immersed in PEG + Gelatin matrix as they sunk into the collagen matrix when added. Also, the non-invading cancer cells in the spheroids facing the PEG + Gelatin matrix were suspected to have poor nutrient gradient and the inadequate presence of integrins in the matrix. Hence, future work or improvement to the 3D biomimetic culture model have to be done before we could fully understand whether EMT and MET of highly invasive tumor cell lines are critically determined by the physical and material characteristics of the microenvironment.Bachelor of Engineering (Materials Engineering
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