Mechanical Manipulation and Characterization of Biological Cells

Mechanical manipulation and characterization of an individual biological cell is currently one of the most exciting research areas in the field of medical robotics. Single cell manipulation is an important process in intracytoplasmic sperm injection (ICSI), pro-nuclei DNA injection, gene therapy, and other biomedical areas. However, conventional cell manipulation requires long training and the success rate depends on the experience of the operator. The goal of this research is to address the drawbacks of conventional cell manipulation by using force and vision feedback for cell manipulation tasks. We hypothesize that force feedback plays an important role in cell manipulation and possibly helps in cell characterization. This dissertation will summarize our research on: 1) the development of force and vision feedback interface for cell manipulation, 2) human subject studies to evaluate the addition of force feedback for cell injection tasks, 3) the development of haptics-enabled atomic force microscope system for cell indentation tasks, 4) appropriate analytical model for characterizing the mechanical property of mouse embryonic stem cells (mESC) and 5) several indentation studies on mESC to determine the mechanical property of undifferentiated and early differentiating (6 days under differentiation conditions) mESC. Our experimental results on zebrafish egg cells show that a system with force feedback capability when combined with vision feedback can lead to potentially higher success rates in cell injection tasks. Using this information, we performed experiments on mESC using the AFM to understand their characteristics in the undifferentiated pluripotent state as well as early differentiating state. These experiments were done on both live as well as fixed cells to understand the correlation between the two during cell indentation studies. Our results show that the mechanical property of undifferentiated mESC differs from early differentiating (6th day) mESC in both live and fixed cells. Thus, we hypothesize that mechanical characterization studies will potentially pave the way for developing a high throughput system with force feedback capability, to understand and predict the differentiation path a particular pluripotent cell will follow. This finding could also be used to develop improved methods of targeted cellular differentiation of stem cells for therapeutic and regenerative medicine

In Vitro, Non-Invasive Imaging and Detection of Single Living Mammalian Cells Interacting with Bio-Nano-Interfaces

Understanding of bio-nano-interfaces of living mammalian cells will benefit the identification of cellular alterations (e.g. nucleic acids, amino acids, biomechanics, etc.) due to external stimuli, the design of biomaterials (e.g. nanoparticles, nanotubes) and the investigation of the interaction between cells and bio-nano-interfaces (e.g. cell differentiation on 3D nanostructured materials). Analytical techniques can be applied to evaluate the chemical, physical, and mechanical properties of mammalian cells when exposed to such bio-nano-interfaces. In this study, non-invasive advanced spectroscopy techniques including atomic force microscopy (AFM) and Raman microscopy (RM), in conjunction with traditional biological methods are utilized to elucidate specific characteristic information for biological samples and how these property changes reflect the interaction with external stimuli. The focus of this dissertation is on the biophysical, biochemical and cytotoxic detection of mammalian cells interacting with bio-nano-interfaces, and this dissertation can be classified into three topics: biomechanics/cellular biopolymers measurement, bio-interfaces and nano-interfaces studies. For the topic of biomechanics/cellular biopolymers measurement, cellular biophysical and biomechanical properties could be used as differentiation markers to classify cellular differentiation. For the bio-interfaces part, it was observed that BRMS1 expression changed cellular biochemical and biomechanical properties, and the expressions of reactive oxidative species (ROS), apoptosis and cell viability of five types of cells displayed similar patterns over doxorubicin (DOX) incubation time. Secondly, A549 cells were treated with diesel exhaust particles (DEP) and resveratrol (RES) to study the effect of RES on the DEP-induced cells, and it was found that RES can alleviate DEP intervention on cellular structure and increase DEP-induced biomechanical and inflammatory changes. For the nano-interfaces topic, first we synthesized a hybrid nanoparticle with the multimodal properties of fluorescence imaging, Surface-enhanced Raman spectroscopy (SERS) detection and photothermal therapy (PTT) for single living cell analysis of epidermal growth factor receptor (EGFR) and specifically killed cancer cells with high EGFR expression. Additionally, to increase surface area, light-heat conversion efficiency and biocompatibility, we developed a silica coated nanoparticle conjugated with anti-human epidermal growth factor receptor 2 (HER2) antibody. Finally, three-dimensional TiO2 nanotubes with Au nanoparticles coating were synthesized and used to study trophoblast-derived stem-like cells growth on such 3D nanostructures

Microdevices and Microsystems for Cell Manipulation

Microfabricated devices and systems capable of micromanipulation are well-suited for the manipulation of cells. These technologies are capable of a variety of functions, including cell trapping, cell sorting, cell culturing, and cell surgery, often at single-cell or sub-cellular resolution. These functionalities are achieved through a variety of mechanisms, including mechanical, electrical, magnetic, optical, and thermal forces. The operations that these microdevices and microsystems enable are relevant to many areas of biomedical research, including tissue engineering, cellular therapeutics, drug discovery, and diagnostics. This Special Issue will highlight recent advances in the field of cellular manipulation. Technologies capable of parallel single-cell manipulation are of special interest

Mechanical characterisation and FEM modelling of biological deformation for surgical simulation

This thesis sought to explore the use of minimally invasive surgery via biomechanical simulation of soft tissue deformation and needle path planning insertion. When surgeons are placed under mechanical stress, human brain cells exhibit the viscoelastic behaviour of solid structures. However, the behavioural mechanisms of tissues/cells are not yet fully understood, and more information is needed to reliably calculate tissue/cell deformation. The research objectives and methodologies were: First, to objectively investigate and characterise the mechanical properties of biological tissues/cells by using experimental atomic force microscopy (AFM) data (see CHAPTER 3). This method was used to analyse the cell's mechanical behaviours with a developed numerical algorithm. The difference between two human brain cells (normal HNC-2 and U87 cancer cells) was studied to determine their mechanical properties so that these could then be applied to our proposed 3D model (see CHAPTER 5). Second, using the measured experimental AFM data, a system identification of AFM characterisation was implemented in another chapter (CHAPTER 4), which for comparison, was based on a MATLAB algorithm. The results showed that the model that was identified for AFM matched the measured experimental AFM data. Third, to establish a finite element method (FEM) for real-time modelling of nonlinear soft tissue deformation behaviours using a three-dimensional (3D) dynamic nonlinear FEM; this method was developed to establish the large-range deformation of tissue/cells with second- order Piola-Kirchhoff stress (CHAPTER 5). A Newmark numerical process was implemented to solve the partial differential equations (PDEs) that resulted from the FEM. Experimental analysis of biological human brain cells was conducted to verify and validate the nonlinear FEM for simulating deformation. Fourth, to establish a method for real-time motion plan modelling of nonlinear needle deflection during needle insertion using the third objective to implement the nonlinear FEM for needle path planning. Last, to use an application of bio-heat transfer of potential needle tip path planning by applying a bioheat transfer-based method (CHAPTER 6); this method was established for optimal path planning for needle insertion in the presence of soft tissue deformation. A bio- heat transfer was used to develop a temperature distribution for path planning to reach the target and avoid obstacles in cubic, liver and brain cell models. The algorithm defines the optimal path for needle tip placement; the needle tip placement is determined by the temperature distribution, which in turn, is based on soft tissue deformation that occurs in the process of needle insertion. When force was applied during the needle penetration process, the deflection accrued was based on the geometry of nonlinear material. Based on our simulation of 3D FEM discretisation of the Pennes' Bio-heat Transfer Equation, the distribution of the temperature from single point temperature sources was performed to determine the degree of transient thermal. Furthermore, the distribution was used to model thermal stresses and strains within the cell/tissue, which result from the heat source. The main contribution to this field is building a new conceptual design methodology for characterisation of the mechanical properties of biological cells by extracts of the mechanical properties of two biological human brain cells (normal HNC-2 and cancer U87 MG cells), and the experimental use of AFM for the first time. Also, linear FEM for soft tissue/needle insertion with large deformation is developed and adapted to our three-dimensional dynamic FEM soft tissue/cell modelling using numerical integration methods. Verification of the experimental work and the proposed method is examined mathematically and systematically using a system identification schema. Moreover, bio-heat transfer for needle insertion is implemented based on the proposed FEM soft tissue deformation modelling to represent path planning. The investigation of needle insertion into soft tissue/cell deformation using bioheat transfer FEM has not been done before

Professional English for biomedical engineering students

Навчальний посібник забезпечує аудиторну та самостійну роботу студентів третього курсу факультету біомедичної інженерії. Видання складається з восьми розділів (Units), які охоплюють професійно орієнтовані теми (Topics): “Introduction to biomedical engineering”, “Robotics in biomedical and healthcare engineering”, “Tissue engineering”, “Medical Imaging”, “Nanotechnology in biomedical engineering”, “Rehabilitation engineering”, “Biomaterials”, “Genetic engineering”. Розроблені вправи спрямовані на забезпечення знань, розвиток і удосконалення навичок і вмінь у читанні, говорнні, аудіюванні, письмі та перекладі, а також покращення лексичних та граматичних знань, навичок і умінь студентів. Завданням посібника є сприяння розширенню професійного тезаурусу студентів та підвищення мотивації студентів до автономного навчання

Intraocular lenses and their potential to prohibit posterior capsule opacification

Cataracts are the commonest cause of preventable blindness in the world. During surgery the natural lens is replaced with a polymeric intraocular lens (IOL), leaving the capsular bag in situ. The most common postoperative complication is scarring which is known as posterior capsule opacification (PCO). PCO occurs when residual lens epithelial cells (LECs) dedifferentiate and migrate onto the previously cell free posterior capsule. By modifying the IOL surface properties we can manipulate the cellular response. BioInteractions Ltd. is an innovative supplier of biomaterials, which aim to minimise, the host response, and provided the materials for this study. The aim of this study was to evaluate potential IOL coatings to reduce PCO. This can either be achieved by enabling a monolayer of LECs to attach to the posterior surface of the IOL, thus sandwiching the IOL to the capsular bag, or prohibiting cell attachment to the IOL entirely. Materials and Methods Various coatings were investigated incorporating: functional groups of poly ethylene glycol (PEG), sulphates, sulfonates, glycosaminoglycans (Heparin, (HEP) hyaluronic aid, (HA) and chondroitin sulphate (CS)) and zwitterionic monomers (10-30%). Ways to prevent dedifferentiation was also evaluated. LECs were seeded onto all coatings and monitored for a period of 7 – 14 days in cell culture. LECs were examined morphologically, cell nuclei were counted and growth curves were plotted. Water contact angle (CA) measurements were taken to measure the wettability of the coatings. Scanning electron microscope (SEM) analysis was performed to examine the topography of the coating. White light interferometry (WLI) analysis was conducted to analysis the surface roughness. Dedifferentiation of LECs and the use of TGFβ3 to neutralise or prevent dedifferentiation were also investigated. Results and Discussions Coatings with a greater number of water-based layers were the most hydrophilic, and did not offer the appropriate cell binding sites required to promote cell attachment. In general, little cell attachment was observed on HEP and HA coatings provided by BioInteractions Ltd., cell attachment varied on CS coatings provided by BioInteractions Ltd. When HA and CS were covalently bound onto amine coated coverslips a reduction in cell attachment was observed. The LEC response varied across different ratios of zwitterionic monomer within the coatings. Zwitterionic coatings were not cytotoxic to LECs and surface analysis demonstrated no clear link between wettability and roughness compared to cell attachment. Addition of transforming growth factor beta 2 (TGFβ2) was chosen as a successful dedifferentiation model. Addition of TGFβ3 had little influence at reversing dedifferentiation however it may offer some protection against differentiation. PCR analysis showed a change in regulation of collagens, integrins, matrix metallopeptidase and fibronectin 1 genes, when LECs were incubated with TGFβ2, TGFβ3 or untreated (control LECs). These genes may play important roles in PCO. Conclusions Incorporation of functional groups influenced the cellular response, however the coatings with more water-based layers prohibit cell attachment. The cellular response varied depending on GAG type and the conformation of GAG on the surface coating. HA and CS bound to amine-coated coverslips prohibited cell attachment at higher concentrations, indicating their potential to prohibit LEC attachment. There was no clear link between wettability and cell attachment on the novel zwitterionic coatings. The ratio of zwitterionic-component:arylic-based monomer(s) influenced cell attachment. TGFβ2 successfully dedifferentiated LECs. Further work is required to understand the influence of TGFβ3 on dedifferentiation

Life Sciences Program Tasks and Bibliography for FY 1996

This document includes information on all peer reviewed projects funded by the Office of Life and Microgravity Sciences and Applications, Life Sciences Division during fiscal year 1996. This document will be published annually and made available to scientists in the space life sciences field both as a hard copy and as an interactive Internet web page

Microplasma technology for influencing cell-surface interactions

Cataracts are the most common cause of preventable blindness worldwide. During cataract surgery a polymeric intraocular lens (IOL) is used to replace the cloudy natural lens. The most common post-operative complication is posterior capsule opacification (PCO). PCO is a wound healing response related to scarring, in which cellular changes disrupt the light path to the back of the eye through various processes, requiring a costly surgery to restore vision. The material of the IOL has been shown to affect PCO and it is hypothesised that the surface modification of IOL materials may be able to reduce the incidence of PCO. The use of plasmas established in the field of biomaterials modification and atmospheric pressure processes have significant benefits over the previous low pressure systems. In this work investigates the use of an atmospheric pressure plasma jet to modify the surface properties of polymeric materials, with the aim of developing a surface treatment method for use on IOLs. Materials and Methods The centre of polystyrene (PS) and poly(methyl methacrylate)(PMMA) surfaces were treated with an atmospheric pressure microplasma jet. The modification of surfaces was characterised by spatially resolved water contact angle, x-ray photoelectron spectroscopy (XPS) and atomic force microscopy (AFM). LECs were seeded onto surfaces and grown for 1-7 days. Cell attachment, growth and morphology were examined microscopically. The concentrations of some cytokines implicated in PCO (transforming growth factor-β2, basic fibroblast growth factor, interleukin-1, interleukin-6, and tumour necrosis factor-α) in culture medium were examined at specific time points. Tissue culture polystyrene and untreated materials served as controls. Atmospheric pressure plasma polymerisation of amine containing monomers using a plasma jet was also investigated. Results and Discussions The size of surface treatment could be tailored by altering flow rate and sample-nozzle distance. Surface treatment was due to an increase in surface oxygen content and plasma treatment did not cause a significant change in surface roughness. Plasma treatment increased the LEC adhesion to substrates. LECs were densely populated in the centre of treated materials and cells lacked the cobblestone morphology typical of epithelial cells. The secretion of inflammatory cytokines by LECs grown on plasma treated surfaces did not appear to be up-regulated in comparison to tissue culture polystyrene, however these results are preliminary. This work demonstrated that atmospheric pressure plasma polymerisation can be achieved using the plasma jet system to incorporate nitrogen functionalisation onto PS surfaces; however oxygen was also incorporated onto surfaces. Conclusions This work demonstrates that an atmospheric pressure microplasma jet can be used to modify surfaces in a spatially defined manner, without damaging the polymer surfaces. The increase in surface oxygen promotes cell adhesion which can be confined to an are