Epilepsy is the most common recurrent neurological condition, affecting a little over 1\% of the world's population. It is often debilitating because of significant psychological, social and cognitive burdens apart from the seizures themselves. Advancing treatment options for generalised epilepsy syndromes has proven a challenging problem, since they are often refractory to treatment with anti-epileptic medication, and are currently not amenable to surgical treatment, so that it is rarely ethical to conduct invasive intracranial EEG studies in human patients with these conditions in order to study them further. Although there has been controversy over the origin and networks involved in absence seizures, previous work by our group has shown that alternative areas of somatosensory cortex are important parts of the thalamocortical circuit in the generation of seizures. The work of this thesis aims to extend those studies by showing that the coupling between cortex and thalamus is altered during seizures. It is established that there is a process of resonance ocurring prior to and during seizures in relevant deep layers of cortex, and the mechanisms behind this are studied. Novel aspects of the work presented in this thesis include the examination of the EEG in the GAERS rat, a whole animal model of absence epilepsy, at the scales of both individual cortical microcircuits and whole cortical areas using simultaneous depth electrodes, and also at both very fine and long time scales. The studies described in this thesis show that the somatosensory cortex leads the thalamus in the first second of seizure onset, in agreement with the results of previous work. To account for the mechanisms behind this increased coupling at seizure onset, an analysis of the properties of resonance in different parts of cortex is presented. This indicates altered phase coupling prior to seizures, in the form of increased phase entrainment, phase coherence, and cross-frequency coupling. These changes are most marked in the junction between primary and secondary somatosensory cortex, with earlier, larger increases prior to seizures, and earlier decoupling at seizure termination. Dynamical analysis points to a number of processes underlying such increases in cortical resonance. Alterations in scaling behaviour are observed in different cortical regions, and also prior to seizures. Analysis of the Lyapunov spectrum of cortical depth and tetrode EEG data shows evidence for a separation in timescales of dynamics at seizure onset, and changes consistent with critical slowing. Further, gradual changes in the statistics of the largest exponents are seen with approaching seizures, suggesting an alteration in dynamical properties giving rise to critical slowing. The results of this thesis support the recently proposed cortical focus theory of absence seizure generation in GAERS rats, and provide new insight into cortical network changes relevant to seizures. This work provides a basis to explore the mode of action of antiepileptic drugs, and more targeted therapy for generalised absence seizures.

Thirty percent of people with epilepsy have seizures refractory to all available medical and surgical treatment options. The uncertainty caused by the constant threat of seizures is the most significant factor impacting people's quality of life with refractory epilepsy. Seizure forecasters aim to alleviate this uncertainty by advising when seizures will occur. This thesis explores the design of seizure forecasters with the aim of improving seizure forecaster performance and clinical utility. Constraints on the seizure forecasting problem are considered, and algorithms for seizure forecasting are explored. Often after a seizure, the total energy in an EEG recording is suppressed, taking seconds to minutes to return to typical values. This period is associated with impaired cognition, drowsiness, and possibly Sudden Unexplained Death in Epilepsy (SUDEP). The postictal period and its relationship to seizure length is explored, motivated by the recent discovery of bimodal seizure length distributions within some patients. Postictal length is defined as the time from seizure end to when average EEG values pass average interictal levels. The findings showed that postictal suppression length is bimodal, with the short seizure population having noticeably shorter postictal periods. For seizure forecasting, this has important implications for how seizures should be labelled. Separately classifying different seizure populations may improve seizure forecasting performance. Forecasters may soon be effective enough to be clinically useful. To optimize clinical utility, the user requirements for a seizure forecasting device need to be determined. Requirements include forecasting performance and the preferred format and timing of the information provided to assist patients. User requirements of a seizure forecaster are explored, providing constraints to the seizure forecasting task. An app for use by the public was made to reproduce the uncertainty of having epilepsy to model the seizure forecaster experience. The app also provided forecasts for ‘events’ to model the seizure forecaster experience. Using this model had two potential benefits: a larger population could be asked about their views on forecaster requirements, and the people being asked had experience using a forecaster. The app asked questions relating to forecaster design, including how forecast information was portrayed, how valuable the forecasts were, and how it alerted the user. The insights gained from user feedback will be helpful to guide the design of future seizure forecasters. Many factors, both exogenous and endogenous, are considered seizure precipitants and may have utility in seizure forecasting. The use of weather, sleep, and temporal features to forecast seizures was explored in this thesis. These features were chosen based on the availability of data. The weather features considered were temperature, maximum daily temperature, minimum daily temperature, humidity, wind speed, rainfall, pressure, and pressure change. The sleep features were current sleep stage, hours asleep, hours awake, hours in stage 1/2/3, the number of sleep transitions, and the time since waking. The temporal features explored were the hour of the day, the time of the week, and the lunar phase. Forecasts from features were combined using a Bayesian approach. Weather, sleep, and temporal factors all performed better than chance in some but not all patients, with temporal features performing the best and weather feature performing the worst. A combination of all features produced forecasts that outperformed individual features on average. Finally, the use of Long short-term Memory (LSTM) neural networks to forecast seizures over long time scales was explored. LSTM networks have memory and, thus, an advantage over most other classifiers such as support vector machines or convolutional neural networks (CNNs). The network included CNN and LSTM components. The CNN component was able to produce short-term forecasts, and the LSTM component was able to use this information to forecast over minutes, hours, or days. The algorithm performed nearly as well as the best performing algorithm on the same dataset, surpassing the previous best performance for some patients. The LSTM networks were also able to perform better than chance when forecasting seizures more than a day in advance.

Brain-computer interfaces (BCIs) have great potential to improve the quality of life for people with severe motor disabilities. By measuring and interpreting neural activity, BCIs can predict and express intention through an external computerised device. This creates an alternative mechanism for communication and control for people with paralysis. Volitional changes in oscillatory activity near the sensorimotor cortex, known as sensorimotor rhythms (SMRs), can be measured with invasive techniques, such as electrocorticography (ECoG), or with non-invasive techniques, such as electroencephalography (EEG). This work explored novel approaches for decoding SMRs from EEG with the aim of utilising neurophysiological principles. This thesis also investigated the viability of vascular electrocorticography (vECoG) as a SMR-based BCI modality. The common spatial patterns (CSP) algorithm is used to linearly combine information from multiple EEG electrodes in order to accentuate SMR activity. Typical patterns of SMR activity derived by this method were characterised using publicly available EEG datasets. It was found that both neurophysiologically probable and improbable patterns of activity were commonly extracted and that selecting for, and adapting to, neurophysiologically probable activity could improve decoding performance. These findings highlight the importance of considering spatial filter adaptation in EEG BCI decoder design. The range of decoding algorithms available to BCI practitioners is extensive and diverse. A universal method for explaining the predictions of EEG decoders in terms of neurophysiologically relevant factors was investigated. The validity of the explanations was demonstrated using simulated EEG data. The method was also employed to compare the behaviour of four categories of decoders using real, pre-recorded EEG data. The results indicated that all decoders were able to harness neurophysiologically plausible electrodes and cortical sources to make accurate predictions. However, the influence of artifactual activity was also found to contribute to high decoder accuracy. These findings emphasise the need to understand the predictive behaviour of decoders and the proposed method may be useful a tool to help BCI researchers achieve this understanding. Vascular electrocorticography (vECoG) measures intracranial neural activity by chronically implanting a stent-electrode array within the brain vasculature. By omitting the need to penetrate the skull, this minimally invasive technique has the potential to safely record high fidelity neural information and be used as a long-term, clinically useful BCI. Data from the first-in-human clinical trial of the Stentrode was used to characterise vECoG signal quality. Participant-specific re-referencing was shown to improve signal quality and the maximum bandwidth of the signal was found to be consistent with previous animal studies. A multiclass, online SMR decoder was also implemented and tested with a single participant. Discriminatory activity from multiple motor imagery classes could be observed, however, signal non-stationarity affected online decoding performance. Together, the signal quality and multiclass decoding results suggest that vECoG has significant potential as a recording modality for a clinically useful BCI. Overall, the findings presented in this thesis contribute to two key facets of SMR BCI research. In terms of decoding SMRs from EEG, they demonstrate the need to harness neurophysiological phenomena, avoid contamination from artifactual activity and improve the interpretability of complex decoding algorithms. Furthermore, the vECoG results add to the growing body of evidence implicating this modality as a beneficial way for interfacing with the brain in a minimally invasive manner.

Biological self-assembly is the foundation of the strength and elasticity which characterizes nature-derived biomaterials, including the cell’s architecture. Thus, an in-depth understanding of the mechanics behind this process can open the doors to various biomedical applications. For this purpose, this research employed novel experimental and characterization techniques to study self-assembly in biopolymers and biosystems. Initially, a droplet dissolution technique in liquid crystalline stages was used to analyze the process of self-assembly in two important biopolymers, silk and cellulose. Here, we report an effective and simple approach based on droplet dissolution in a liquid binary phase for the formation of silk fibroin transparent spheres as well as cellulose microbeads, both of which can span several hundred micrometers in diameter. The microstructure of the spheres formed at different ethanol concentrations was characterized by electron microscopy. High concentrations of ethanol caused droplets to be encased in a thin shell which collapses once it is taken out of the liquid phase. Generally, low ethanol concentrations produce transparent silk spheres and solid cellulose microbeads. This work on biopolymers demonstrates that controlled droplet dissolution self-assembling may be explored as a novel and effective way to tailor the microstructures of nature-derived biomaterials. The spheres generated in this manner have several different characteristics which can have multiple potential uses, such as templates for scaffolds, microcarriers, as well as photonics and nano-technological applications. The second part of this thesis investigated self-assembling in biosystems. Cell aggregates are an important tool in studying tissue remodelling, extracellular matrix formation, cell-cell interaction, and last but not the least, tissue-like biomechanical properties. A medium-throughput method was designed to characterize the mechanical properties of mesenchymal cell aggregates. This study was the first to present a precise and fast method to determine the Young’s modulus of mesenchymal cell aggregates, utilizing a step-by step aspiration technique. We were also able to recreate conditions that very closely resemble the in vivo environment, where the cells were found to be stretched, and the spheroids are soft and elastic Finally, potential applications of the self-assembled cell aggregates were explored in lung disease study and drug screening, specifically for Idiopathic Pulmonary Fibrosis (IPF). We demonstrated that the cell aggregates from IPF patients show an increase in stiffness, therefore mechanical testing of spheroids is an effective technique to study this disease. It was also found that a novel compound, PF670462, modulates the effect of TGF-beta and inhibits the fibrotic response of IPF cell aggregates. That is, this drug softens IPF spheroids and downregulates fibrogenic gene expression, therefore providing basis for the potential use of PF670462 in IPF treatment.

Dyskinetic cerebral palsy is characterised by variable involuntary movements resulting in sustained twisting and jerky movements often causing functional limitations. Assessing dyskinesia using observation-based clinical tools with sufficient reliability and sensitivity can be challenging. However, accurate assessment of dyskinesia is necessary for effective management and development of novel therapies to treat dyskinesia in children with cerebral palsy. In this dissertation, techniques in motion analysis are investigated to explore objective methods that can detect, differentiate and grade the severity of dyskinesia in children with cerebral palsy. In addition, mobile tools such as inertial measurement units are investigated as a possible strategy to address concerns in clinical translation. Three upper limb activities were performed by six typically developing children and twelve children with cerebral palsy. Upper limb movements were recorded using a marker-based optoelectronic motion capture system, inertial measurement units and electromyography. Features of kinematic joint angles, joint centre trajectories and muscle activity were extracted and analysed against clinical assessments of dyskinesia. Kinematic studies using a marker-based optoelectronic system are recommended over inertial measurement units and electromyography for assessing dyskinesia in children with cerebral palsy. Features of volitional and non-volitional movement identified differences between typically developing upper limbs and upper limbs with dyskinesia, albeit more consistently across activities on the non-volitional side. In contrast, differentiating upper limbs with spasticity from dyskinesia in children with cerebral palsy was challenging with only a small number of features demonstrating significant differences. Finally, kinematic-based objective measures capturing smoothness of movement during the reach-and-hold activity provided the best prospect of assessing the severity of dyskinesia in children with cerebral palsy. This dissertation provides encouraging results that support the use of objective measures to assess dyskinesia in children with cerebral palsy. However, there are clear technology barriers that need to be addressed to translate these assessment techniques into clinical practice. With continued research into improving the accuracy of inertial measurement units and other technological advances in markerless video tracking, the future remains promising for the clinical use of objective measures to assess dyskinesia in children with cerebral palsy.

Normal wrist function is ensured by a delicate interplay between bony architecture and soft-tissue constraints. Disruptions of this balance may alter carpal kinematics and joint contact, impairing the normal wrist physiology and biomechanics. The scapholunate interosseous ligament (SLIL), whose integrity is pivotal to normal scapholunate motion and that of the entire wrist, has been recognised as the most commonly injured carpal ligament. If left untreated, prolonged exposure to the altered joint motion may induce attrition of the secondary soft-tissue stabilisers, setting the stage for degenerative osteoarthritis. However, the exact role of these ligamentous stabilisers in maintaining normal scapholunate motion remains poorly understood. At present, many surgical procedures have been developed to treat scapholunate instability, but very few of them have been experimentally validated for their efficacy. Hence the aim of this investigation was twofold. Firstly, to evaluate the effects of different stages of scapholunate instability on wrist kinematics, and establish the relative contribution of various secondary scapholunate stabilisers to scapholunate motion and; secondly, to biomechanically evaluate the outcome of two reconstructive surgeries in treating scapholunate instability. As these experiments were highly invasive, a novel cadaveric wrist testing platform (CWTP), consisting of a computer-controlled dynamic wrist cadaver simulation apparatus (DWSA) and a biplane X-ray fluoroscopy system, was developed. A comprehensive set of validation experiments were carried out to assess the performance of the CWTP in carrying out in-vitro wrist simulations. It was shown that the DWSA was capable of achieving highly accurate and repeatable wrist movements, with robust control performance maintained even in the presence of severe ligament injuries to the wrist. The measurement accuracy and precision of the custom-built biplane X-ray fluoroscopy system were also evaluated. And the results demonstrated that the system was capable of resolving rigid body motion to a sub-degree and sub-millimetre accuracy. Dynamic wrist simulations, including flexion-extension (FEM), radial-ulnar deviation (RUD) and the dart-thrower's motion (DTM), were then performed in seven cadaveric wrist specimens. A four-stage ligament division protocol was developed based on a previous study that correlated specific anatomic lesions of the scapholunate supporting ligaments with different Geissler grades of scapholunate instability, as well as the widely-accepted injury mechanism of hyperextension, ulnar deviation and intercarpal supination. This included sequentially dividing the volar (v-) and the membranous (m-) SLIL in Stage I; the dorsal (d-) SLIL in Stage II; the radioscaphocapitate (RSC) and the long radiolunate (LRL) ligaments in Stage III; and the dorsal intercarpal (DIC) ligament, the scaphotrapeziotrapezoid (STT) ligament, and about 40% of the dorsal radiocarpal (DRC) ligament in Stage IV. Reconstructive surgeries were then performed with the goal of re-stabilising the severely disrupted scapholunate joint. Two different techniques, including a novel dorsal transarticular loop tenodesis (DTLT) procedure and the widely-used three-ligament tenodesis (3LT) procedure, were assessed for their efficacy in restoring the normal scapholunate biomechanics. With incremental ligament division, the scaphoid became progressively more flexed, ulnarly deviated, pronated and radially translated. The largest and the second-largest changes in the scaphoid kinematics were observed after the Stage II and IV ligament divisions, respectively. The motion of the dissociated scaphoid is likely to be primarily dictated by the bony geometry. Dividing the v- and m-SLIL, or the RSC and LRL ligaments, however, did not induce appreciable alterations. The lunate was found to be progressively more extended, supinated, and translated towards the volar and ulnar aspect of the radius. The most considerable changes in the lunate kinematics were noted after the complete division of the SLIL in Stage II. Its dissociation from the scaphoid was immediately evident, as the lunate rotation in the sagittal plane was observed to be significantly attenuated. Dividing the volar extrinsic ligaments in Stage III caused the lunate to pronate instead of rotating into further supination, as well as to become ulnarly translated. In comparison, apart from causing the lunate to become statistically more supinated, dividing the dorsal carpal ligaments and the STT ligament in Stage IV induced no noticeable alterations in the lunate kinematics. The DTLT resulted in a slightly insufficient reduction of the scaphoid in both the sagittal and transverse planes. And the procedure was found to induce a substantial amount of unphysiological volar translation of the scaphoid during ulnar deviation of the wrist. In contrast, the 3LT resulted in a much better correction of the scaphoid flexion; however, the scaphoid still appeared malrotated in pronation in the transverse plane. Furthermore, the pathological scaphoid supination was still present during wrist extension after the 3LT procedure, suggesting that the reconstruction was not capable in preventing the volar opening of the scapholunate interval. For the lunate, poor results were obtained after the 3LT, as the lunate remained collapsed in significant extension and supination. And since the tendon graft was tightened towards the ulnar side of the wrist, the 3LT not only did not correct the ulnar translation of the carpus, the condition was exacerbated by the procedure. The DTLT, on the other hand, led to a successful reduction of the dorsally collapsed lunate, which in turn restored its dynamic extension. The abnormal lunate supination was also corrected. However, cases of overcorrections were observed in these two planes. The results suggested that apart from the SLIL, the dorsal carpal and the STT ligaments might have a vital contribution to the scaphoid stability, and the volar extrinsic ligaments are essential in preventing ulnar translation of the carpus. For the DTLT procedure, a complementary dorsal capsulodesis might be beneficial in correcting the residual scaphoid flexion. And the non-physiological volar translation of the scaphoid during wrist ulnar deviation might be due to the passive stretching of the radially situated ECR-B graft used by the procedure, pronating the scaphoid and lunate as a unit towards the volar aspect of the radius. For the 3LT procedure, its inefficacy in preventing the volar opening of the scapholunate interval might be attributed to the ulnar and dorsal directed pull of the tendon graft, which might not be adequate for counteracting the supination tendency experienced by the scaphoid during wrist extension. Therefore it may be beneficial for a volar reconstruction of the scapholunate joint to be carried out in conjunction with the 3LT procedure. Furthermore, given the unsuccessful reduction of the lunate after 3LT, a transosseous passage of the tendon graft might be considered, instead of simply attaching it to the dorsum of the lunate. In summary, this study advances the understanding of normal and pathological wrist function, and provide a foundation for improved clinical practice in the surgical treatment of scapholunate injuries of the wrist.

Cells are dynamic, living structures that remodel themselves in response to stimuli from environment or in relation to cellular processes such as cell growth, proliferation, differentiation, migration and death. The change of cell mechanical property can be a biophysical indicator in response to the abnormal alteration in cell functionality under pathological conditions. The advances in tool development for cell mechanical measurement have facilitated in-depth discussion of cell mechanics, but heavily limited by low throughput and high cost. The emerging lab-on-a-chip microfluidic methods provide a promising solution due to the miniaturisation, among which the acoustofluidic method (the fusion of acoustics and microfluidics) appears to be advantageous due to its tunability, biocompatibility and acousto-mechanical nature. In this dissertation, I explored the application of surface acoustic wave (SAW) microfluidics in the area of cell mechanics, including establishing SAW devices for cell mechanical measurement, comparing SAW-based measurement with the benchmark from a conventional method, investigating the impacts on cell mechanical characteristic, and extending the concept to a high-throughput cytometry comparable to the real-world need. The results show that the SAW microfluidic method can provide an effective measurement on cell mechanical characteristics and probe the impact of cellular interior structure or cellular phenotype. It is consistent with the conventional benchmark and can be a complement for some cellular structures of interest. At last, it can operate as a continuous-flow high-throughput cytometry, which could be exploited in future studies related to cell mechanics.

The knee is the most injured joint in the body with injuries often caused by a tear in the anterior cruciate ligament (ACL). Over 70% of ACL injuries are non-contact, meaning that an excessive amount of force or moment is generated by the individuals themselves, which induces the tear. Non-contact ACL injuries most commonly occur following a perturbation. Perturbations make the athlete unbalanced or at loss of control, which ultimately alters their normal neuromuscular control and can lead to injury. In order to understand the effect of this unconscious neuromuscular response, this study aimed to induce unanticipated perturbations during walking in 10 male and 10 female athletes. Moreover, the effect of prophylactic knee braces was examined during these perturbations. Musculoskeletal modelling in OpenSim was used to calculate kinematics, kinetics, and muscle forces during these perturbations. Females portrayed muscle force patterns during perturbations, that are likely to increase the risk of ACL injury. Furthermore, females exhibited a larger difference in muscle forces between their dominant and non-dominant limbs. Finally, prophylactic knee braces significantly reduced peak quadriceps and soleus muscle forces during perturbations. Unlike planned movements in laboratory studies, unexpected perturbations provide with deeper insight in the mechanism of ACL injuries. Although ACL injuries do not typically occur during walking, potentially injurious movement patterns during a disturbance to natural balance while walking, could provide insight on what may be reproduced, on a higher scale, during high impact and high-speed athletic tasks that can effectively tear the ACL.

The damage to osteochondral tissue can develop degenerative joint diseases such as subchondral bone necrosis, osteochondral fracture, and osteoarthritis. In the development of these joint diseases, the subchondral bone plays a crucial role due to several risk factors: obesity, hereditary, aging, gender, and intense mechanical loads. The intense mechanical loads include both trauma/impact and repetitive loads which occur during high-intensity training and have been identified as the primary risk factor in the development of joint diseases. In this dissertation, we utilized mechanical tests combined with digital image correlation and micro-computed tomography to study the mechanical behaviour (in particular, Elastic Modulus and Energy Dissipation) of subchondral bone under both trauma/impact and repetitive loads. Results suggest that the presence of severe cartilage lesion in osteochondral tissue reduces the stiffness of cartilage and increases the compressive strain in the subchondral bone under trauma/impact loads. Moreover, the application of simulated physiological repetitive loads on subchondral bone has demonstrated the importance of the highest speeds in the development of the joint disease, consistent with the observations made in equine athletes. The results also exhibit the influence of microstructural properties on the mechanical behaviour of subchondral bone under repetitive loads. The results and approach of simulating physiological loading may be useful in optimizing training protocol and prevent joint diseases.

Epilepsy is the second-most common neurological disorder, with an estimated prevalence of 1% worldwide. Antiepileptic drugs (AEDs) are the frontline treatment but, despite more than 25 agents available, there can be significant difficulties in finding a treatment plan that controls seizures without unacceptable side effects. Furthermore, approximately 30% of patients do not obtain full control of seizures with the available AEDs. Therefore, there exists a need for novel therapies. Therapeutic electrical stimulation of the brain and peripheral nervous system has shown great promise in treating those with refractory epilepsy, but still little is known about the mechanisms underlying this. Recent work looking at prolonged recordings of the brain has revealed a deep rhythmicity to seizures that may provide a method of developing patient-specific prediction algorithms that could be used to titrate electrical stimulation therapies. What is missing in research and in clinical practise is an implantable device which provides enough flexibility to explore new paradigms and adapt to suit the needs of the patient. Safety considerations of implantable devices necessarily precedes large-scale clinical evaluation, and this has prevented exploration of speculative higher density arrays, which come at a cost to power, complexity and mechanical reliability with no guarantee of improvement to efficacy. Rather than redesign the system from scratch, this PhD thesis asks, “how can existing implantable technology be leveraged to create flexible high-density brain recording and stimulation systems?” In response to this question, an integrated circuit called XPAND was designed to enable seamless expansions of electrode count for a wide variety of commercial stimulators. Comprising a 6 x 64 element high voltage crosspoint switch powered and configured entirely using biphasic stimulation pulses, XPAND was designed with clinical safety in mind. Fabricated in Austria Microsystems H35 high voltage CMOS process, key modules within the chip were characterised, and the crosspoint switch shown to simultaneously route high voltage stimulation and ECG signals.

Simple epithelial tissue is formed by individual cells connecting together to form a monolayer sheet. This sheet generates the complex structures of organs by undergoing both large and small scale morphogenic events. Columnar epithelial cells have a height greater than their width and are polarised with a ring of circumferential contractile fibres localised to their apical cell-cell junctions. There has been considerable investigation into the movement of this apical network, especially using lasers to sever individual apical cell edges. From these apical investigations, a variety of mechanical mechanisms have been suggested to control epithelium movement. However, these mechanisms are contrasting, and are often derived for specific experiments, rather than general apical epithelium movement. Recently, it has been recognised that in additional to the apical network, the entire three-dimensional cell shape impacts epithelium movement. Additionally, there has been growing interest in understanding how epithelium scale movement is controlled by smaller scaler structures, such as contractile fibres. This thesis utilises both computational and experimental approaches to investigate how the mechanics of underlying structures can drive epithelium scale movement, with a focus on the contribution of individual cells and contractile fibres to apical epithelium movement upon laser severing. These investigations demonstrate that studying the mechanisms of underlying cell and sub cell structures can provide new insights into the causal mechanics of general epithelium movement. It is also shown that inclusion of these mechanisms can lead to greater prediction of epithelium movement.

Red blood cells (RBCs) squeeze through narrow capillaries as they transport oxygen to tissues and carbon dioxide to the lungs. The deformability of RBCs has been shown to depend on the viscoelasticity of the cell membrane and cytoplasm as well as the surface area to volume ratio (SA:V ratio) of the cell. In certain pathological diseases such as malaria, RBCs undergo restructuring of the membrane structure and modifications to the cell shape, which significantly reduce their deformability. Nonetheless, it is still unclear which factor has the greatest impact on the passage of RBCs through small capillaries. Here, we present a systematic analysis designed to identify the individual contributions of cell stiffness and SA:V ratio to the ability of RBCs to traverse narrow capillaries in a microfluidic device. We modified cellular rigidity using glutaraldehyde fixation, changed SA:V ratio by altering the buffer osmolarity and probed RBCs passage through microchannels. Our results showed that dramatic stiffening (~8 fold) had little effect (~6% retardation) on the ability of RBCs of the same geometry to traverse the channels. On the other hand, a moderate decrease (~13%) in the SA:V ratio affected the traversal of RBCs of similar stiffness more markedly (~19% decrease). We further studied RBCs infected by two different species of malaria parasites known to affect humans, Plasmodium falciparum and knowlesi. We found that P. falciparum rigidified the host RBC, but infected RBCs penetrated into microchannels with similar efficiency to uninfected RBCs. By contrast, P. knowlesi reduced the SA:V ratio of the host RBC resulting in restricted passage. We found that the earliest stage immature RBCs (reticulocytes) exhibited a similar SA:V ratio to mature RBCs and, despite being 30% larger, travelled into microchannels as efficiently as mature cells. Our finite element (FE) model provides a coherent rationale for our experimental observations, indicating that cell stiffness changes do not significantly affect RBC traversal in small capillaries due to the highly nonlinear mechanical behaviour of the cell membrane. Our numerical simulations predict that RBCs with low SA:V ratios are more prone to trapping in small capillaries (within the physiological size range) than RBCs with high membrane stiffness. Therefore, therapies targetting surface area to volume ratio of RBCs may be more effective than those that target cell stiffness.