| 1. |
- Porras, Ana Maria, et al.
(författare)
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Brain microvasculature endothelial cell orientation on micropatterned hydrogels is affected by glucose level variations
- 2021
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Ingår i: Scientific Reports. - : Springer Nature. - 2045-2322. ; 11:1
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Tidskriftsartikel (refereegranskat)abstract
- This work reports on an effort to decipher the alignment of brain microvasculature endothelial cells to physical constrains generated via adhesion control on hydrogel surfaces and explore the corresponding responses upon glucose level variations emulating the hypo- and hyperglycaemic effects in diabetes. We prepared hydrogels of hyaluronic acid a natural biomaterial that does not naturally support endothelial cell adhesion, and specifically functionalised RGD peptides into lines using UV-mediated linkage. The width of the lines was varied from 10 to 100 µm. We evaluated cell alignment by measuring the nuclei, cell, and F-actin orientations, and the nuclei and cell eccentricity via immunofluorescent staining and image analysis. We found that the brain microvascular endothelial cells aligned and elongated to these physical constraints for all line widths. In addition, we also observed that varying the cell medium glucose levels affected the cell alignment along the patterns. We believe our results may provide a platform for further studies on the impact of altered glucose levels in cardiovascular disease.
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| 2. |
- Porras, Ana Maria, et al.
(författare)
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Cell chirality exhibition of brain microvascular endothelial cells is dependent on micropattern width
- 2022
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Ingår i: RSC Advances. - : Royal Society of Chemistry. - 2046-2069. ; 12:46, s. 30135-30144
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Tidskriftsartikel (refereegranskat)abstract
- Left-right asymmetry is a conserved property in nature and observed in the human body, a property known as cell chirality. Cell chirality is often studied using micropatterned in vitro models. However, micropattern geometry and size often varies across different studies, making it challenging to compare results. Here, we utilized micropatterned RGD-peptide lines on hyaluronic acid hydrogels to investigate the effect of the micropattern width on the exhibited cell chirality bias of brain microvascular endothelial cells. Overall, this cell type exhibited a negative chirality bias on micropatterned lines ranging from 10 mu m to 400 mu m in width, where the negative bias was most pronounced on the 100 mu m wide lines. We also observed that this exhibited chirality bias varied across the line width. This work serves as a guide to determine optimal micropattern width for further investigations on cell chirality bias and its prominence in e.g., disease states or upon exposure to toxic substances.
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| 3. |
- Porras Hernandez, Ana Maria, et al.
(författare)
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A simplified approach to control cell adherence on biologically derived in vitro cell culture scaffolds by direct UV-mediated RGD linkage
- 2020
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Ingår i: Journal of materials science. Materials in medicine. - : Springer Nature. - 0957-4530 .- 1573-4838. ; 31:10
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Tidskriftsartikel (refereegranskat)abstract
- In this work, we present a method to fabricate a hyaluronic acid hydrogel with spatially controlled cell-adhesion properties based on photo-polymerisation cross-linking and functionalisation. The approach utilises the same reaction pathway for both steps meaning that it is user-friendly and allows for adaptation at any stage during the fabrication process. Moreover, the process does not require any additional cross-linkers. The hydrogel is formed by UV initiated radical addition reaction between acrylamide groups on the hyaluronic acid backbone. Cell adhesion is modulated by functionalising the adhesion peptide sequence RGD (arginine-glycine-aspartate) onto the hydrogel surface via radical mediated thiol-ene reaction using the non-reacted acrylamide groups. We show that 10 x 10 µm2 squares could be patterned with sharp features and a good resolution. The smallest area that could be patterned resulting in good cell adhesion was 25 x 25 µm2 squares, showing single-cell adhesion. Mouse brain endothelial cells adhered and remained in culture for up to 7 days on 100 x 100 µm2 square patterns. We see potential for this material combination for future use in novel organ-on-chip models and tissue engineering where the location of the cells is of importance and to further study endothelial cell biology.
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| 4. |
- Porras Hernandez, Ana Maria, et al.
(författare)
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Alignment of brain endothelial cells on patterned hyaluronic acid hydrogels
- 2020
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- Endothelial cells (ECs) line the blood vessel walls and present an elongated characteristic morphology. While the effect of micropatterning on different endothelial cells have been extensively studied, the effects on brain endothelial cells, which are highly specialized cells, have been overlooked [1]. Moreover, it has been shown that brain ECs do not elongate and align in response to shear stress, as e.g. HUVECs do. [2]. Hence, we set out to conclude how brain endothelial cells would behave on micropatterned lines. I We fabricated an RGD micropatterned photocrosslinked hyaluronic acid (HA-am) hydrogel substrate, with lines of controlled dimensions. These substrates were used to study cell elongation and alignment of the cell nuclei when adhering to lines raging from 10 µm to 100 µm in width.
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| 5. |
- Porras Hernández, Ana Maria, et al.
(författare)
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Confocal imaging dataset to assess endothelial cell orientation during extreme glucose conditions
- 2022
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Ingår i: Scientific Data. - : Springer Nature. - 2052-4463. ; 9:1
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Tidskriftsartikel (refereegranskat)abstract
- Confocal microscopy offers a mean to extract quantitative data on spatially confined subcellular structures. Here, we provide an imaging dataset of confocal z-stacks on endothelial cells spatially confined on lines with different widths, visualizing the nucleus, F-actin, and zonula occludens-1 (ZO-1), as well as the lines. This dataset also includes confocal images of spatially confined endothelial cells challenged with different glucose conditions. We have validated the image quality by established analytical means using the MeasureImageQuality module of the CellProfilerTM software. We envision that this dataset could be used to extract data on both a population and a single cell level, as well as a learning set for the development of new image analysis tools.
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| 6. |
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| 7. |
- Porras Hernández, Ana María
(författare)
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Micropatterning of hyaluronic acid hydrogels for in vitro models
- 2022
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The human body consist of a vast number of cells, and jointly, the cells, form tissues and organs. The cells interact and respond to their local microenvironment. The cellular microenvironment consists of a highly hydrated and compliant extracellular matrix, neighboring cells and circulating biochemical factors; and jointly, provide chemical and physical cues that regulate cell behaviour However, these cues are often not present in traditional in vitro models, where cells experience a stiff and unstructured environment. An approach to better mimic the in vivo microenvironment in vitro is to use hydrogels. Hydrogels are soft and highly hydrated polymers based on materials naturally found in the extracellular matrix of various tissues. Furthermore, these materials can be chemically functionalized to control the physical, chemical, and mechanical properties of the hydrogels. These functionalities can also be used to prepare micrometre sized cell adhesive regions, or micropatterns, on the hydrogel substrate. The micropatterns guide the cell shape and permit the study of the cell response to these changes in shape and function, which has been observed in e.g., endothelial cells from various origins. Taken all together, the aim of this work was to develop a hydrogel-based cell culture substrate that permits the control of the spatial adhesion of brain endothelial cells in order to study the morphological effects on these cells and contribute to the understanding of the function of brain endothelial cells in health and disease. This thesis demonstrates the functionalization of hyaluronic acid, a naturally occurring extracellular matrix polymer, to prepare photocrosslinkable hydrogels. Then, through photolithography, micropatterns of cell adhesive peptides were prepared on these hydrogels. Brain microvascular endothelial cells, a highly specialized type of endothelial cells, adhered to the micropatterns, and the effect on their alignment and cell chirality depending on the micropatterned sized was studied. Furthermore, changes in their alignment were also observed when exposed to different glucose concentration.
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| 8. |
- Tenje, Maria, et al.
(författare)
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A practical guide to microfabrication and patterning of hydrogels for biomimetic cell culture scaffolds
- 2020
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Ingår i: Organs-on-a-Chip. - The Netherlands : Elsevier. - 2666-1020. ; 2
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Forskningsöversikt (refereegranskat)abstract
- This review article describes microfabrication techniques to define chemical, mechanical and structural patterns in hydrogels and how these can be used to prepare in vivo like, i.e. biomimetic, cell culture scaffolds. Hydrogels are attractive materials for 3D cell cultures as they provide ideal culture conditions and they are becoming more prominently used. Single material gels without any modifications do however have their limitation in use and much can be gained by in improving the in vivo resemblance of simple hydrogel cell culture scaffolds. This review article discusses the most commonly used cross-linking strategies used for hydrogel-based culture scaffolds and gives a brief introduction to microfabrication methods that can be used to define chemical, mechanical and structural patterns in hydrogels with micrometre resolution. The review article also describes a selection of literature references using these microfabrication techniques to prepare organ and disease models with controlled cell adhesion, proliferation and migration. It is intended to serve as an introduction to microfabrication of hydrogels and an inspiration for novel interdisciplinary research projects.
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| 9. |
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| 10. |
- Cantoni, Federico, et al.
(författare)
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A microfluidic chip carrier including temperature control and perfusion system for long-term cell imaging
- 2021
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Ingår i: HardwareX. - : Elsevier. - 2468-0672. ; 10, s. e00245-
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Tidskriftsartikel (refereegranskat)abstract
- Microfluidic devices are widely used for biomedical applications but there is still a lack of affordable, reliable and user-friendly systems for transferring microfluidic chips from an incubator to a microscope while maintaining physiological conditions when performing microscopy. The presented carrier represents a cost-effective option for sustaining environmental conditions of microfluidic chips in combination with minimizing the device manipulation required for reagent injection, media exchange or sample collection. The carrier, which has the outer dimension of a standard well plate size, contains an integrated perfusion system that can recirculate the media using piezo pumps, operated in either continuous or intermittent modes (50–1000 µl/min). Furthermore, a film resistive heater made from 37 µm-thick copper wires, including temperature feedback control, was used to maintain the microfluidic chip temperature at 37 °C when outside the incubator. The heater characterisation showed a uniform temperature distribution along the chip channel for perfusion flow rates up to 10 µl/min. To demonstrate the feasibility of our platform for long term cell culture monitoring, mouse brain endothelial cells (bEnd.3) were repeatedly monitored for a period of 10 days, demonstrating a system with both the versatility and the potential for long imaging in microphysiological system cell cultures.
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