Glioma on Chips Analysis of glioma cell guidance and interaction in microfluidic-controlled microenvironment enabled by machine learning

In biosystems, chemical and physical fields established by gradients guide cell migration, which is a fundamental phenomenon underlying physiological and pathophysiological processes such as development, morphogenesis, wound healing, and cancer metastasis. Cells in the supportive tissue of the brain, glia, are electrically stimulated by the local field potentials from neuronal activities. How the electric field may influence glial cells is yet fully understood. Furthermore, the cancer of glia, glioma, is not only the most common type of brain cancer, but the high-grade form of it (glioblastoma) is particularly aggressive with cells migrating into the surrounding tissues (infiltration) and contribute to poor prognosis. In this thesis, I investigate how electric fields in the microenvironment can affect the migration of glioblastoma cells using a versatile microsystem I have developed. I employ a hybrid microfluidic design to combine poly(methylmethacrylate) (PMMA) and poly(dimethylsiloxane) (PDMS), two of the most common materials for microfluidic fabrication. The advantages of the two materials can be complemented while disadvantages can be mitigated. The hybrid microfluidics have advantages such as versatile 3D layouts in PMMA, high dimensional accuracy in PDMS, and rapid prototype turnaround by facile bonding between PMMA and PDMS using a dual-energy double sided tape. To accurately analyze label-free cell migration, a machine learning software, Usiigaci, is developed to automatically segment, track, and analyze single cell movement and morphological changes under phase contrast microscopy. The hybrid microfluidic chip is then used to study the migration of glioblastoma cell models, T98G and U-251MG, in electric field (electrotaxis). The influence of extracellular matrix and chemical ligands on glioblastoma electrotaxis are investigated. I further test if voltage-gated calcium channels are involved in glioblastoma electrotaxis. The electrotaxes of glioblastoma cells are found to require optimal laminin extracellular matrices and depend on different types of voltage-gated calcium channels, voltage-gated potassium channels, and sodium transporters. A reversiblysealed hybrid microfluidic chip is developed to study how electric field and laminar shear can condition confluent endothelial cells and if the biomimetic conditions affect glioma cell adhesion to them. It is found that glioma/endothelial adhesion is mediated by the Ang1/Tie2 signaling axis and adhesion of glioma is slightly increased to endothelial cells conditioned with shear flow and moderate electric field. In conclusion, robust and versatile hybrid microsystems are employed for studying glioma biology with emphasis on cell migration. The hybrid microfluidic tools can enable us to elucidate fundamental mechanisms in the field of the tumor biology and regenerative medicine.Okinawa Institute of Science and Technology Graduate Universit

Population genetics in microchannels

Spatial constraints such as rigid barriers affect the dynamics of cell populations, potentially altering the course of natural evolution. In this paper, we study the population genetics of Escherichia coli proliferating in microchannels with open ends. Our experiments reveal that competition among two fluorescently labeled E. coli strains growing in a microchannel generates a self-organized stripe pattern aligned with the axial direction of the channel. To account for this observation, we employ a lattice population model in which reproducing cells push entire lanes of cells towards the open ends of the channel. By combining mathematical theory, numerical simulations, and experiments, we find that the fixation dynamics is extremely fast along the axial direction, with a logarithmic dependence on the number of cells per lane. In contrast, competition among lanes is a much slower process. We also demonstrate that random mutations appearing in the middle of the channel and close to its walls are much more likely to reach fixation than mutations occurring elsewhere.Comment: 21 pages, 14 figure

Anti-Inflammatory and Anticoagulative Effects of Paeonol on LPS-Induced Acute Lung Injury in Rats

Paeonol is an active component of Moutan Cortex Radicis and is widely used as an analgesic, antipyretic, and anti-inflammatory agent in traditional Chinese medicine. We wanted to determine the role of paeonol in treating adult respiratory distress syndrome (ARDS). We established an acute lung injury (ALI) model in Sprague-Dawley rats, which was similar to ARDS in humans, using intratracheal administration of lipopolysaccharide (LPS). The intraperitoneal administration of paeonol successfully reduced histopathological scores and attenuated myeloperoxidase-reactive cells as an index of polymorphonuclear neutrophils infiltration and also reduces inducible nitric oxide synthase expression in the lung tissue, at 16 h after LPS administration. In addition, paeonol reduced proinflammatory cytokines in bronchoalveolar lavage fluid, including tumor-necrosis factor-α, interleukin-1β, interleukin-6, and plasminogen-activated inhibition factor-1. These results indicated that paeonol successfully attenuates inflammatory and coagulation reactions to protect against ALI

Voltage-gated ion channels mediate the electrotaxis of glioblastoma cells in a hybrid PMMA/PDMS microdevice

Transformed astrocytes in the most aggressive form cause glioblastoma, the most common cancer in the central nervous system with high mortality. The physiological electric field by neuronal local field potentials and tissue polarity may guide the infiltration of glioblastoma cells through the electrotaxis process. However, microenvironments with multiplex gradients are difficult to create. In this work, we have developed a hybrid microfluidic platform to study glioblastoma electrotaxis in controlled microenvironments with high throughput quantitative analysis by machine learning-powered single cell tracking software. By equalizing the hydrostatic pressure difference between inlets and outlets of the microchannel, uniform single cells can be seeded reliably inside the microdevice. The electrotaxis of two glioblastoma models, T98G and U-251MG, requires an optimal laminin-containing extracellular matrix and exhibits opposite directional and electro-alignment tendencies. Calcium signaling is a key contributor in glioblastoma pathophysiology but its role in glioblastoma electrotaxis is still an open question. Anodal T98G electrotaxis and cathodal U-251MG electrotaxis require the presence of extracellular calcium cations. U-251MG electrotaxis is dependent on the P/Q-type voltage-gated calcium channel (VGCC) and T98G is dependent on the R-type VGCC. U-251MG electrotaxis and T98G electrotaxis are also mediated by A-type (rapidly inactivating) voltage-gated potassium channels and acid-sensing sodium channels. The involvement of multiple ion channels suggests that the glioblastoma electrotaxis is complex and patient-specific ion channel expression can be critical to develop personalized therapeutics to fight against cancer metastasis. The hybrid microfluidic design and machine learning-powered single cell analysis provide a simple and flexible platform for quantitative investigation of complicated biological systems

Glioblastoma adhesion in a quick-fit hybrid microdevice

Translational research requires reliable biomedical microdevices (BMMD) to mimic physiological conditions and answer biological questions. In this work, we introduce a reversibly sealed quick-fit hybrid BMMD that is operator-friendly and bubble-free, requires low reagent and cell consumption, enables robust and high throughput performance for biomedical experiments. Specifically, we fabricate a quick-fit poly(methyl methacrylate) and poly(dimethyl siloxane) (PMMA/PDMS) prototype to illustrate its utilities by probing the adhesion of glioblastoma cells (T98G and U251MG) to primary endothelial cells. In static condition, we confirm that angiopoietin-Tie2 signaling increases the adhesion of glioblastoma cells to endothelial cells. Next, to mimic the physiological hemodynamic flow and investigate the effect of physiological electric field, the endothelial cells are pre-conditioned with concurrent shear flow (with fixed 1 Pa shear stress) and direct current electric field (dcEF) in the quick-fit PMMA/PDMS BMMD. With shear flow alone, endothelial cells exhibit classical parallel alignment; while under a concurrent dcEF, the cells align perpendicularly to the electric current when the dcEF is greater than 154 V m(-1). Moreover, with fixed shear stress of 1 Pa, T98G glioblastoma cells demonstrate increased adhesion to endothelial cells conditioned in dcEF of 154 V m(-1), while U251MG glioblastoma cells show no difference. The quick-fit hybrid BMMD provides a simple and flexible platform to create multiplex systems, making it possible to investigate complicated biological conditions for translational research

Tumour-on-a-chip: microfluidic models of tumour morphology, growth and microenvironment

Cancer remains one of the leading causes of death, albeit enormous efforts to cure the disease. To overcome the major challenges in cancer therapy, we need to have a better understanding of the tumour microenvironment (TME), as well as a more effective means to screen anti-cancer drug leads; both can be achieved using advanced technologies, including the emerging tumour-on-a-chip technology. Here, we review the recent development of the tumour-on-a-chip technology, which integrates microfluidics, microfabrication, tissue engineering and biomaterials research, and offers new opportunities for building and applying functional three-dimensional in vitro human tumour models for oncology research, immunotherapy studies and drug screening. In particular, tumour-on-a-chip microdevices allow well-controlled microscopic studies of the interaction among tumour cells, immune cells and cells in the TME, of which simple tissue cultures and animal models are not amenable to do. The challenges in developing the next-generation tumour-on-a-chip technology are also discussed

Geographical heterogeneity and influenza infection within households

Although it has been suggested that schoolchildren vaccination reduces influenza morbidity and mortality in the community, it is unknown whether geographical heterogeneity would affect vaccine effectiveness

Relationships between serum HER2 ECD, TIMP-1 and clinical outcomes in Taiwanese breast cancer

<p>Abstract</p> <p>Background</p> <p>Serum levels of the extracellular domain of HER2/neu (HER2 ECD) have been demonstrated to be associated with clinical outcomes. A disintegrin and metalloproteinase-10, a sheddase of HER2/neu, can drive cancer progression and its activity is inhibited by tissue inhibitor of metalloproteinase-1 (TIMP-1). However, elevated TIMP-1 expression has been associated with a poor prognosis of breast cancer. Therefore, this study was performed to explore the relationships between serum HER2 ECD, TIMP-1 and clinical outcomes.</p> <p>Methods</p> <p>One hundred and eighty-five female breast cancer patients, who received curative mastectomy without neo-adjuvant chemotherapy at Chang-Gung Memorial Hospital, were recruited with informed consent for this study. Pre-operative serum levels of HER2 ECD and TIMP-1 were measured using an enzyme-linked immunosorbent assay.</p> <p>Results</p> <p>Twenty-three cases (12.4%) were classified HER2 ECD positive. HER2 ECD positivity was significantly associated with age, lymph node involvement, histological grade, estrogen receptor status, progesterone receptor status, tissue HER2/neu overexpression, and disease-free survival (DFS). In an age, stage, ER and HER2/neu status matched subgroup (N = 41), the serum level of TIMP-1 was significantly associated with HER2 ECD positivity and DFS.</p> <p>Conclusions</p> <p>A high serum TIMP-1 was significantly associated with HER2 ECD positivity and a poorer DFS among Taiwanese primary breast cancer patients with HER2 overexpression.</p