Stuart T. Fraser

The growth and study of living cells outside their native organisms forms the foundation of modern biology and underpin medicine. It has led to the identification of stem cells and the development of methods that can reprogram mature cells into pluripotent states, creating enormous potential for new therapies that can cure previously untreatable conditions and enable the repair of patient-specific tissues and organs. Accessing these advances, however, will require the development of sophisticated new cell culture materials and technologies. This Perspective article reviews the development of cell culture and current cell culture capabilities, with particular attention to the influence of spatial and temporal factors. We discuss traditional 2D culture, the complexities of 3D systems, and the emergence of 2.5D approaches as an alternative to high throughput 2D systems. Untapped potential and barriers to progress are identified while the new materials and technologies needed to drive the field forward are discussed.

This paper provides first-time evidence of covalent attachment of fluorescent molecules to plasma polymerised nanoparticles for diagnostic and therapeutic applications.

Background: Accumulation of lipid in the liver is the first hallmark of both alcohol-related liver disease (ALD) and non-alcohol-related fatty liver disease (NAFLD). Recent studies indicate that specific mutations in lipid genes confer risk and might influence disease progression to irreversible liver cirrhosis. This study aimed to understand the function/s of lipid risk genes driving disease development in zebrafish genetic models of alcohol- and non-alcohol related fatty liver. Methods: We used zebrafish larvae to investigate the effect of alcohol and high fat to model fatty liver and tested the utility of this model to study lipid risk gene functions. CRISPR/Cas9 gene editing was used to create knockdowns in 5 days post-fertilization zebrafish larvae for the available orthologs of human cirrhosis risk genes ( pnpla3, faf2, tm6sf2 ). To establish fatty liver models, larvae were exposed to ethanol and a high fat diet (HFD) consisting of chicken egg yolk. Changes in morphology (imaging), survival, liver injury (biochemical tests, histopathology), gene expression (qPCR) and lipid accumulation (dye specific live imaging) were analysed across treatment groups to test the functions of these genes. Results: Exposure of 5-day post-fertilization (dpf) WT larvae to 2% ethanol or HFD for 48 hours developed measurable hepatic steatosis. CRISPR-Cas9 genome editing depleted pnpla3, faf2 and tm6sf2 gene expression in these CRISPR knockdown larvae (crispants). Knockdown significantly increased effects of ethanol and HFD toxicity by increasing hepatic steatosis and hepatic neutrophil recruitment ≥2-fold in all three crispants. Furthermore, ethanol or HFD exposure significantly altered the expression of genes associated with ethanol metabolism ( cyp2y3 ) and lipid metabolism-related gene expression, including atgl (triglyceride hydrolysis), axox1, echs1 (fatty acid β-oxidation), fabp10a (transport), hmgcra (metabolism), notch1 (signaling) and srebp1 (lipid synthesis), in all three pnpla3, faf2 and tm6sf2 crispants. Nile Red staining in all three crispants revealed significantly increased lipid droplet size and triglyceride accumulation in the livers following exposure to ethanol or HFD. Conclusions: We identified roles for pnpla3, faf2 and tm6sf2 genes in triglyceride accumulation and fatty acid oxidation pathways in a zebrafish larvae model of fatty liver.

The lipid content of mammalian cells varies greatly between cell type. Current methods for analysing lipid components of cells are technically challenging and destructive. Here, we report a facile, inexpensive method to identify lipid content: intracellular flow cytometric lipid analysis (IFCLA). Distinct lipid classes can be distinguished by Nile Blue, Nile Red fluorescence or violet autofluorescence. Nile Blue is fluorescent in the presence of unsaturated fatty acids with a carbon chain length greater than 16. Cis-configured fatty acids induce greater Nile Blue fluorescence than their trans-configured counterparts. In contrast, Nile Red exhibits greatest fluorescence in the presence of cholesterol, cholesteryl esters, some triglycerides and phospholipids. Multiparametric SPADE analysis of hepatic cellular lipid distribution including Vitamin A autofluorescence is presented. This flow cytometric system allows for the rapid, inexpensive, non-destructive identification of lipid content and highlights the differences in lipid biology between cell types by imaging and flow cytometry.

There is a growing demand for polymer fiber scaffolds for biomedical applications and tissue engineering. Biodegradable polymers such as polycaprolactone have attracted particular attention due to their applicability to tissue engineering and optical neural interfacing. Here we report on a scalable and inexpensive fiber fabrication technique, which enables the drawing of PCL fibers in a single process without the use of auxiliary cladding. We demonstrate the possibility of drawing PCL fibers of different geometries and cross‐sections, including solid‐core, hollow‐core, and grooved fibers. The solid‐core fibers of different geometries are shown to support cell growth, through successful MCF‐7 breast cancer cell attachment and proliferation. We also show that the hollow‐core fibers exhibit a relatively stable optical propagation loss after submersion into a biological fluid for up to 21 days with potential to be used as waveguides in optical neural interfacing. The capacity to tailor the surface morphology of biodegradable PCL fibers and their non‐cytotoxicity make the proposed approach an attractive platform for biomedical applications and tissue engineering.

The representation of scientific knowledge and information on Wikipedia is controversial. While some entries present established scientific fact, other entries are devoid of information or present information of questionable rigour. Taking this and the problems associated with essay topics as assessment tasks into consideration, an assessment task has been developed in which senior physiology or histology students edit a Wikipedia entry which is relevant to the themes in the unit of study. Students develop digital literacy skills while improving the quality of scientific information on Wikipedia. Teaching and learning resources have been developed to aid the students transition from writing in a typical ‘essay style’ to writing in a style appropriate to Wikipedia. The assessment task and design presented here can be readily adapted to any field of study and is designed to help academics introduce editing of Wikipedia articles into existing curricula in an efficient and effective manner.

Red blood cells, or erythrocytes, make up approximately a quarter of all cells in the human body with over 2 billion new erythrocytes made each day in a healthy adult human. This massive cellular production system is coupled with a set of cell biological processes unique to mammals, in particular, the elimination of all organelles, and the expulsion and destruction of the condensed erythroid nucleus. Erythrocytes from birds, reptiles, amphibians and fish possess nuclei, mitochondria and other organelles: erythrocytes from mammals lack all of these intracellular components. This review will focus on the dynamic changes that take place in developing erythroid cells that are interacting with specialized macrophages in multicellular clusters termed erythroblastic islands. Proerythroblasts enter the erythroblastic niche as large cells with active nuclei, mitochondria producing heme and energy, and attach to the central macrophage via a range of adhesion molecules. Proerythroblasts then mature into erythroblasts and, following enucleation, in reticulocytes. When reticulocytes exit the erythroblastic island, they are smaller cells, without nuclei and with few mitochondria, possess some polyribosomes and have a profoundly different surface molecule phenotype. Here, we will review, step-by-step, the biophysical mechanisms that regulate the remarkable process of erythropoiesis with a particular focus on the events taking place in the erythroblastic island niche. This is presented from the biological perspective to offer insight into the elements of red blood cell development in the erythroblastic island niche which could be further explored with biophysical modelling systems.

The most common cell type in the human body, the red blood cell or erythrocyte, has a life span of approximately 3 months. To compensate for this massive cellular requirement and short life span, the major blood producing tissues contain vast numbers of erythroid progenitor cells. Erythroid progenitors differentiate progressively from hematopoietic stem cells to committed erythroid progenitors to reticulocytes lacking a nucleus and finally to functionally mature erythrocytes in the circulation. Different erythroid progenitor activity, representative of distinct stages of erythropoiesis, can be observed using semisolid colony assays. Distinct stages of erythroid maturation can also be monitored by flow cytometry. Here, we discuss the range of different technical approaches that are used to identify and quantify erythroid progenitors, with particular focus on the mouse as a model system.