| 1. |
- Gustavsson, Anna-Karin, 1986, et al.
(författare)
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Allosteric regulation of phosphofructokinase controls the emergence of glycolytic oscillations in isolated yeast cells
- 2014
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Ingår i: The FEBS Journal. - : Wiley. - 1742-464X .- 1742-4658. ; 281:12, s. 2784-2793
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Tidskriftsartikel (refereegranskat)abstract
- Oscillations are widely distributed in nature and synchronization of oscillators has been described at the cellular level (e.g. heart cells) and at the population level (e.g. fireflies). Yeast glycolysis is the best known oscillatory system, although it has been studied almost exclusively at the population level (i.e. limited to observations of average behaviour in synchronized cultures). We studied individual yeast cells that were positioned with optical tweezers in a microfluidic chamber to determine the precise conditions for autonomous glycolytic oscillations. Hopf bifurcation points were determined experimentally in individual cells as a function of glucose and cyanide concentrations. The experiments were analyzed in a detailed mathematical model and could be interpreted in terms of an oscillatory manifold in a three-dimensional state-space; crossing the boundaries of the manifold coincides with the onset of oscillations and positioning along the longitudinal axis of the volume sets the period. The oscillatory manifold could be approximated by allosteric control values of phosphofructokinase for ATP and AMP.
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| 2. |
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| 3. |
- Gustavsson, Anna-Karin, 1986, et al.
(författare)
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Effect of External Acetaldehyde on Glycolytic Oscillations in Individual Yeast Cells
- 2013
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Ingår i: Optical Molecular Probes, Imaging and Drug Delivery.
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- Yeast cells in dense cultures can synchronize their glycolytic oscillations via acetaldehyde. Combining optical tweezers with microfluidics, the effect of external acetaldehyde on glycolytic oscillations in individual cells has been investigated.
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| 4. |
- Gustavsson, Anna-Karin, 1986, et al.
(författare)
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Entrainment of heterogeneous glycolytic oscillations in single cells
- 2015
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 5
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Tidskriftsartikel (refereegranskat)abstract
- Cell signaling, gene expression, and metabolism are affected by cell-cell heterogeneity and random changes in the environment. The effects of such fluctuations on cell signaling and gene expression have recently been studied intensively using single-cell experiments. In metabolism heterogeneity may be particularly important because it may affect synchronisation of metabolic oscillations, an important example of cell-cell communication. This synchronisation is notoriously difficult to describe theoretically as the example of glycolytic oscillations shows: neither is the mechanism of glycolytic synchronisation understood nor the role of cell-cell heterogeneity. To pin down the mechanism and to assess its robustness and universality we have experimentally investigated the entrainment of glycolytic oscillations in individual yeast cells by periodic external perturbations. We find that oscillatory cells synchronise through phase shifts and that the mechanism is insensitive to cell heterogeneity (robustness) and similar for different types of external perturbations (universality).
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| 5. |
- Gustavsson, Anna-Karin, 1986, et al.
(författare)
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FEBS Journal Prize Lecture: Sustained glycolytic oscillations in individual isolated yeast cells
- 2013
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Ingår i: FEBS Journal. - : Wiley. - 1742-4658 .- 1742-464X. ; 280:Suppl. S1
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Konferensbidrag (övrigt vetenskapligt/konstnärligt)abstract
- Yeast glycolytic oscillations have been extensively studied since the 1950s in dense populations of cells and in cell-free extracts. Until recently, sustained oscillations had only been observed at the population level, i.e. for synchronized cultures at high biomass concentrations. One question that had not been satisfactorily addressed was whether individual cells display qualitatively different behaviour from the mean behaviour of a population of cells. We were able to observe sustained oscillations in individual isolated cells using a sophisticated experimental setup in which the concentration of metabolites in glycolysis was quantified by measuring the autofluorescence intensity from NADH molecules in the individual cells, the extracellular environment was controlled both spatially and temporally using microfluidics, and the cell density and position of the cell array within the microfluidic flow chamber was varied using optical tweezers. We thus showed that a high cell density is not a requirement for induction of oscillatory behaviour. A detailed kinetic model for the cellular reactions was adjusted to describe isolated cells in a microfluidic flow chamber. It was successfully used to simulate the heterogeneity in the oscillatory response of the individual cells, assuming small differences in a single internal parameter. In further studies we have investigated the precise conditions for autonomous oscillations at the single cell level. We have also investigated how the extracellular environment affects the characteristics of the oscillations and the heterogeneity between cells. This setup also enables studies of cell-to-cell distance and flowrate dependence on cell communication and synchronization.
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| 6. |
- Gustavsson, Anna-Karin, 1986
(författare)
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Glycolytic oscillations in individual yeast cells
- 2015
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Oscillations in the concentration of yeast glycolytic intermediates have been intensively studied since the 1950s, but these studies have so far been limited to observations of average oscillatory behavior in synchronized cultures. Hence, it has remained unknown whether the onset of oscillations is a collective property of the population which requires a high cell density, or if individual cells can oscillate also in isolation. To determine the mechanisms behind oscillations, cell-cell interactions and synchronization, and to investigate the role of cell-cell heterogeneity, oscillations have to be studied on the single-cell level. The aims of this project were to determine whether individual cells in isolation can oscillate and if there is large heterogeneity among individual cells, to determine if a fluid flow affects the oscillatory behavior, to identify the precise conditions required for oscillations to emerge in individual cells, to investigate the mechanism behind oscillations, and to elucidate the mechanism behind synchronization, its robustness to cell heterogeneity and its universality with respect to different chemical species. In this work it was shown that glycolytic oscillations can be induced and studied in individual, isolated yeast cells by combining optical tweezers for cell positioning, microfluidics for environmental control and fluorescence microscopy for detection. My single-cell data revealed large heterogeneity and four categories of cell behavior were identified. It was also verified that the oscillatory behavior was determined by the concentrations of glucose and cyanide in the extracellular environment rather than the flow rates used in the microfluidic flow chamber. Varying the concentrations of glucose and cyanide, the precise conditions for oscillations to emerge in individual cells were determined and it was shown that individual cells can oscillate also at conditions where no oscillations are detected in populations. This indicates that loss of oscillations in a population can be caused by desynchronization rather than by loss of oscillations in individual cells. Investigation of single-cell responses using a detailed kinetic model showed that the onset of oscillations could be described by allosteric regulation of the enzyme phosphofructokinase by AMP and ATP. To determine the mechanism behind synchronization and to assess its robustness and universality, entrainment of oscillations in individual yeast cells by periodic external perturbations was investigated. It was found that oscillatory cells synchronize through phase shifts and that the mechanism is insensitive to cell heterogeneity (robustness) and similar for different types of external perturbations (universality). The results presented in this work have advanced our understanding of the complex set of reactions in energy metabolism and the mechanisms through which cells oscillate, communicate, and synchronize. Pursuing these studies will hopefully not only give further information about glycolysis in yeast, but also about energy metabolism, oscillations, and communication in other biological systems, such as oscillatory insulin secretion from islets of beta-cells.
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| 7. |
- Gustavsson, Anna-Karin, 1986, et al.
(författare)
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Heterogeneity of glycolytic oscillatory behaviour in individual yeast cells
- 2014
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Ingår i: FEBS Letters. - : Wiley. - 0014-5793. ; 588:1, s. 3-7
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Forskningsöversikt (refereegranskat)abstract
- There are many examples of oscillations in biological systems and one of the most investigated is glycolytic oscillations in yeast. These oscillations have been studied since the 1950s in dense, synchronized populations and in cell-free extracts, but it has for long been unknown whether a high cell density is a requirement for oscillations to be induced, or if individual cells can oscillate also in isolation without synchronization. Here we present an experimental method and a detailed kinetic model for studying glycolytic oscillations in individual, isolated yeast cells and compare them to previously reported studies of single-cell oscillations. The importance of single-cell studies of this phenomenon and relevant future research questions are also discussed.
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| 8. |
- Gustavsson, Anna-Karin, 1986, et al.
(författare)
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Induction of sustained glycolytic oscillations in single yeast cells using microfluidics and optical tweezers
- 2012
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Ingår i: Proc. SPIE, 8458 s. 84580Y. - : SPIE. ; 8458
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Konferensbidrag (refereegranskat)abstract
- Yeast glycolytic oscillations have been studied since the 1950s in cell free extracts and in intact cells. Until recently, sustained oscillations have only been observed in intact cells at the population level. The aim of this study was to investigate sustained glycolytic oscillations in single cells. Optical tweezers were used to position yeast cells in arrays with variable cell density in the junction of a microfluidic flow chamber. The microfluidic flow chambers were fabricated using soft lithography and the flow rates in the different inlet channels were individually controlled by syringe pumps. Due to the low Reynolds number, the solutions mixed by diffusion only. The environment in the junction of the chamber could thus be controlled by changing the flow rates in the inlet channels, with a complete change of environment within 2 s. The optimum position of the cell array was determined by simulations, to ensure complete coverage of the intended solution without any concentration gradients over the cell array. Using a DAPI filter set, the NADH auto fluorescence could be monitored in up to 100 cells simultaneously. Sustained oscillations were successfully induced in individual, isolated cells within specific flow rates and concentrations of glucose and cyanide. By changing the flow rates without changing the surrounding solution, it was found that the cell behavior was dependent on the concentration of chemicals in the medium rather than the flow rates in the range tested. Furthermore, by packing cells tightly, cell-to-cell interaction and synchronization could be studied.
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| 9. |
- Gustavsson, Anna-Karin, 1986
(författare)
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Single Cell Studies of Glycolytic Oscillations Using Microfluidics and Optical Tweezers
- 2012
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Licentiatavhandling (övrigt vetenskapligt/konstnärligt)abstract
- The complex set of reactions in metabolism has been extensively studied in populations with millions of cells, but much information can still be gained by studying the heterogeneous metabolic behaviour in individual cells. The first part of energy metabolism is called glycolysis and is very similar in mammalian cells, such as human cells, and in yeast cells. If yeast cells are exposed to certain concentrations of glucose and cyanide, the concentration of metabolites in glycolysis starts to oscillate. These glycolytic oscillations have been studied since the 1950s in both intact cells and in yeast extracts. In dense cell cultures, glycolytic oscillations are synchronized via the metabolic intermediate acetaldehyde, which rapidly diffuses through the cell membrane. Synchronization is a requirement when oscillations from millions of cells in a population are to be investigated. Such studies will, however, only give information about the population average and the oscillatory behaviour on the single cell level will be unknown. Several questions have for long been unanswered, such as why, as reported, a population of cells loses its oscillations. This could be due to non-oscillatory cells but it might also be an effect of desynchronization of the oscillations. Another question is if there is a large heterogeneity in for instance amplitude and period time of the oscillations between the individual cells. If a population of cells will show complete, partial or no synchronization depends both on the heterogeneity on the single cell level and on the coupling strength between the cells. In this work, these questions have been investigated by combining optical tweezers for cell positioning in arrays with variable cell density, microfluidics for spatial and temporal environmental control and fluorescence microscopy for detection of the responses on the single cell level. It was found that sustained glycolytic oscillations can be induced in isolated yeast cells and hence that a high cell density is not a requirement for oscillations to occur. A large heterogeneity in the oscillatory behaviour could also be seen on the single cell level. This setup enables future studies where important information about metabolism can be gained and where cell-to-cell coupling and synchronization can be further investigated.
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| 10. |
- Gustavsson, Anna-Karin, 1986, et al.
(författare)
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Studying Glycolytic Oscillations in Individual Yeast Cells by Combining Fluorescence Microscopy with Microfluidics and Optical Tweezers.
- 2019
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Ingår i: Current protocols in cell biology. - : Wiley. - 1934-2616 .- 1934-2500. ; 82:1
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Tidskriftsartikel (refereegranskat)abstract
- In this unit, we provide a clear exposition of the methodology employed to study dynamic responses in individual cells, using microfluidics for controlling and adjusting the cell environment, optical tweezers for precise cell positioning, and fluorescence microscopy for detecting intracellular responses. This unit focuses on the induction and study of glycolytic oscillations in single yeast cells, but the methodology can easily be adjusted to examine other biological questions and cell types. We present a step-by-step guide for fabrication of the microfluidic device, for alignment of the optical tweezers, for cell preparation, and for time-lapse imaging of glycolytic oscillations in single cells, including a discussion of common pitfalls. A user who follows the protocols should be able to detect clear metabolite time traces over the course of up to an hour that are indicative of dynamics on the second scale in individual cells during fast and reversible environmental adjustments. © 2018 by John Wiley & Sons, Inc.
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| 11. |
- Gustavsson, Anna-Karin, 1986, et al.
(författare)
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Sustained glycolytic oscillations in individual isolated yeast cells
- 2012
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Ingår i: Febs Journal. - : Wiley. - 1742-464X. ; 279:16, s. 2837-2847
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Tidskriftsartikel (refereegranskat)abstract
- Yeast glycolytic oscillations have been studied since the 1950s in cell-free extracts and intact cells. For intact cells, sustained oscillations have so far only been observed at the population level, i.e. for synchronized cultures at high biomass concentrations. Using optical tweezers to position yeast cells in a microfluidic chamber, we were able to observe sustained oscillations in individual isolated cells. Using a detailed kinetic model for the cellular reactions, we simulated the heterogeneity in the response of the individual cells, assuming small differences in a single internal parameter. This is the first time that sustained limit-cycle oscillations have been demonstrated in isolated yeast cells. Database ?The mathematical model described here has been submitted to the JWS Online Cellular Systems Modelling Database and can be accessed at free of charge.
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| 12. |
- Mojica Benavides, Martin, 1983, et al.
(författare)
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An Optical Tweezers, Epi-Fluorescence/Spinning disk confocal- and microfluidic-setup for synchronization studies of glycolytic oscillations in living yeast cells
- 2016
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Ingår i: Proceedings SPIE 9922, Optical Trapping and Optical Micromanipulation XIII. San Diego; USA. 28 August -1 September 2016. - : SPIE. - 9781510602359
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Konferensbidrag (refereegranskat)abstract
- Due to the significant importance of glycolytic oscillations studies and the recent breakthroughs on single cell analysis, a further interest arrives with intracellular and intercellular responses. Understanding cell-cell communication can give insight to oscillatory behaviors in biological systems, such as insulin secretion from pancreatic beta-cells. The aim of this work consists on the manipulation of living yeast cells to study propagation and synchronization of induced glycolytic oscillations. A setup, consisting of an optical tweezers system and microfluidic devices coupled with fluorescence imaging was designed to perform a time dependent observation during artificially induced glycolytic oscillations. Multi-channel flow devices and diffusion chambers were fabricated using soft lithography. Automatized pumps controlled specific flow rates of infused glucose and cyanide solutions, used to induce the oscillations. Flow and diffusion in the microfluidic devices were simulated to assure experimentally the desired coverage of the solutions across the yeast cells, a requirement for time dependent measurements. Using near infrared optical tweezers, yeast cells were trapped and positioned in array configurations, ranging from a single cell to clusters of various symmetries, in order to obtain information about cell-cell communications during the metabolic cycles. Confocal illumination of an entire focal plane using a spinning disk, will allow acquirement of NADH periodic fluorescence signals during glycolytic oscillations. This method permits an improvement of the 2D projection images obtained with wide field microscopy to a tomographic description of the subcellular propagation of the oscillations.
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| 13. |
- Schüler, Emil, et al.
(författare)
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Solar photocatalytic and electrokinetic studies of TiO2/Ag nanoparticle suspensions
- 2013
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Ingår i: Solar Energy. - : Elsevier BV. - 0038-092X. ; 96, s. 220-226
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Tidskriftsartikel (refereegranskat)abstract
- Composites of TiO2 and Ag nanoparticles were used to study photodegradation of aqueous Rhodamine B under natural sunlight exposure. The addition of Ag powder resulted in a ∼45% increase of the photocatalytic activity of TiO2 after 30 min, which is attributed to the suppression of the photo-generated electron–hole pair recombination. Electrokinetic studies on TiO2, Ag, and TiO2/Ag aqueous solutions address the question of how the electron transfer occurs. It was found that the TiO2 and TiO2/Ag zeta potential versus pH behavior is virtually identical which suggests that the Ag particles are encapsulated by the TiO2 particles, allowing the electron transfer across the ‘static’ semiconductor–metal interface.
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| 14. |
- van Niekerk, David, et al.
(författare)
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Phosphofructokinase controls the acetaldehyde induced phase shift in isolated yeast glycolytic oscillators.
- 2019
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Ingår i: The Biochemical journal. - 1470-8728. ; 476:2, s. 353-363
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Tidskriftsartikel (refereegranskat)abstract
- The response of oscillatory systems to external perturbations is crucial for emergent properties such as synchronization and phase locking, and can be quantified in a phase response curve. In individual, oscillating yeast cells, we characterized experimentally the phase response of glycolytic oscillations for external acetaldehyde pulses, and followed the transduction of the perturbation through the system. Subsequently, we analyzed the control of the relevant system components in a detailed mechanistic model. The observed responses are interpreted in terms of the functional coupling and regulation in the reaction network. We find that our model quantitatively predicts the phase dependent phase shift observed in the experimental data. The phase shift is in agreement with an adaptation leading to synchronization with an external signal. Our model analysis establishes that phosphofructokinase plays a key role in the phase shift dynamics as shown in the phase response curve, and adaptation time to external perturbations. Specific mechanism-based interventions, made possible through such analyses of detailed models, can improve upon standard trial and error methods, e.g. melatonin supplementation to overcome jet-lag, which are error prone, specifically, since the effects are phase and dose dependent.
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| 15. |
- Wastesson, Karin, 1986-
(författare)
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Learning Managerial Work : First-line Managers’ Learning in Everyday Work within Swedish Elderly Care
- 2024
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- This study’s overall aim is to contribute knowledge about first-line managers’ learning in everyday work within the context of elderly care. The study used a qualitative research approach and was carried out within four Swedish elderly care organisations. A total of 40 first-line managers were interviewed, 10 of whom kept time-use diaries. The theoretical framework is based on a workplace learning perspective originating from theory of situated learning. Situated learning theory includes the concepts of community of practice, legitimate peripheral participation, and membership, which serve as analytical tools to illuminate characteristics of learning from various angles. In addition, the concept of gender has been used to gain a deeper understanding for managers’ workplace learning in the female-dominated elderly care context.The findings showed that managers’ learning happened in a stream of varied tasks and interactions shaped by conditions in the workplace. The managers’ work was characterised by unpredictability and changing circumstances, which meant they continuously had to learn how to handle new situations and expand their repertoire of managerial practices. One common perception among themanagers was the importance of being able to make quick decisions with limited information, and assess the results afterwards. The managers learned how to deal with work situations by either maintaining and modifying current practices or inventing new ones. In addition to these three practices, their learning was affected by different conditions, particularly professional experience, work relationships and organisational conditions. The findings further showed that the managers had to learn to deal with expectations of how they should act in the managerial role based on their gender, and learn to navigate between gender ideals that permeated the female-dominated elderly care environment.Three conclusions were drawn from this study. The first was that managers took great responsibility for their own learning, including what they needed to know and how they would learn it. Despite the fact that they all had access to resources provided by the employer, managers often chose alternative ways to learn, usually by relying on informal networks and close personal relationships. As a result of this self-directed learning, they were able to make decisions that suited their learning needs, and effectively proceed in practice without having to confirm their chosen methods.The second conclusion was that work relationships played a central part in managers’ learning, within both the care work community and the first-line manager community. Work relationships with other first-line managers provided support for learning through, for example, knowledge exchange and joint discussions, as well as emotional support. Work relationships with subordinates were significant for learning, and could result in solutions to complex issues, which could have a direct effect on the daily operation of care work services. Due to the diverse mix of professions, varying interests, and formal positions of authority in the care work community, managers were required to devote considerable time and effort to facilitate collaboration and a shared repertoire. As a result, learning was seen as a stimulating and enjoyable experience, but was also demanding and sometimes boring.The third conclusion was that in the context of the female-dominated elderly care gender operated differently in two communities of practices. Male privilege was still prominent in the care work community, as men were accepted and perceived as legitimate leaders among their subordinates. Female managers instead had to navigate and balance the expectations associated with femininity and the managerial role in this community. However, the female-dominated elderly care context provided female managers with more opportunities to connect with equal peers and establish influential positions, whereas male managers could encounter challenges in gaining access to learning and participating in the first-line manager community.
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