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Modeling and Data Analysis in Cellular Biophysics
KTH, School of Engineering Sciences (SCI), Applied Physics, Cell Physics. (Cellens fysik)
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Cellular biophysics deals with the physical aspects of cell biology. This thesis presents a number of studies where mathematical models and data analysis can increase our understanding of this field.

During recent years development in experimental methods and mathematical modeling have driven the amount of data and complexity in our understanding of cellular biology to a new level. This development has made it possible to describe cellular systems quantitatively where only qualitative descriptions were previously possible. To deal with the complex data and models that arise in this kind of research a combination of tools from physics and cell biology has to be applied; this constitutes a field we call cellular biophysics. The aim of this doctoral thesis is to develop novel approaches in this field. I present eight studies where quantitative modeling and analysis are involved.

The first two studies concern cells interacting with their surrounding environment in the kidney. These cells sense fluid flow and respond with calcium (Ca2+) signals. The interaction between fluid and cells in renal tubular epithelium can be described by biomechanical models. This thesis describes a mathematical model of flow sensing by cilia with focus on the flow frequency response and time delay between the mechanical stress and the Ca2+ signaling response.

Intracellular Ca2+ is kept at a very low level compared to the extracellular environment, while several intracellular compartments have higher Ca2+ concentration than the cytoplasm. This makes Ca2+ an efficient messenger for intra­cellular signaling, the process whereby signals are transduced from an extracellular stimulus to an intracellular activity such as gene expression. An important type of Ca2+ signaling is oscillations in intracellular Ca2+ concentration which occur due to the concerted interplay between different transport mechanisms within a cell. A study in this thesis examines ways to explain these mechanisms in terms of a mathematical model. Another study in the thesis reports that erythropoietin can regulate the water permeability of astrocytes and that it alters the pattern of Ca2+ oscillations in astrocytes. In this thesis the analysis of this Ca2+ signaling is described.

Simulations described in one of the studies show how different geometries can affect the fluorescence recovery and that geometrically constrained reactions can trap diffusing receptors in dendritic spines. When separate time scales are present in a fluorescence revovery after photobleaching (FRAP) experiment the reaction and diffusion components can be studied separately.

Applying single particle tracking methods to the migration trajectories of natural killer cells shows that there is a correlation between the formation of conjugates and transient confinement zones (TCZs) in these trajectories in vitro. TCZs are also present in in vivo experiments where they show strong similarities with the in vitro situation. This approach is a novel concept in data analysis methods for tracking immune cells.

Abstract [sv]

Cellens biologiska fysik behandlar de fysikaliska aspekterna av cellbiologi. Denna avhandling presenterar ett antal studier där matematiska modeller och dataanalys kan öka vår förståelse av detta område.

Under senare år har utvecklingen av experimentella metoder och matematisk modellering drivit mängden data och komplexiteten i vår förståelse av cellbiologi till en ny nivå. Denna utveckling har gjort det möjligt att beskriva cellulära system kvantitativt där endast kvalitativa beskrivningar tidigare var möjliga. För att hantera de komplexa data och modeller som uppstår i denna typ av forskning krävs en kombination av verktyg från fysik och cellbiologi; detta utgör ett område vi kallar cellens biologiska fysik. Syftet med denna avhandling är att utveckla nya metoder inom detta område. Jag presenterar åtta studier där kvantitativ modellering och analys ingår.

De första två studierna behandlar hur celler interagerar med sin omgivning i njurarna. Dessa celler känner av ett vätskeflöde och svarar med kalcium (Ca2+)-signaler. Samspelet mellan vätska och celler i tubulärt njurepitel kan beskrivas med biomekaniska modeller. Denna avhandling beskriver en matematisk modell för flödeskänslighet hos cilier med fokus på flödesfrekvenssvar och tidsfördröjningen mellan den mekaniska påverkan och Ca2+-signaleringssvaret.

Intracellulärt Ca2+ hålls på en mycket låg nivå jämfört med den extracellulära miljön, samtidigt som flera intracellulära delar har högre Ca2+-koncentrationen än cytoplasman. Detta gör Ca2+ till en effektiv bärare för intracellulär signalering, den process där signaler överförs från ett extracellulärt stimuli till en intracellulär händelse, exempelvis genuttryck. En viktig typ av Ca2+-signalering är de oscillationer i intracellulär Ca2+-koncentration som uppstår på grund av det ordnade samspelet mellan olika transportmekanismer i en cell. En studie  i denna avhandling undersöker olika sätt att förklara dessa mekanismer i form av en matematisk modell. En annan studie i avhandlingen rapporterar att erytropoietin kan reglera vattenpermeabilitet av astrocyter och att det ändrar mönstret av Ca2+-oscillationer i astrocyter. I denna avhandling beskrivs analysen av denna Ca2+-signalering.

Simuleringar som beskrivs i en av studierna visar hur olika geometrier kan påverka fluorescensåterhämtning och att geometriskt begränsade reaktioner kan fånga in receptorer in i dendrittaggar. När separata tidsskalor förekommer i ett fluorescence revovery after photobleaching (FRAP)-experiment kan reaktions- och diffusionskomponenter studeras separat.

Tillämpande av single particle tracking-metoder på naturliga mördarceller visar att det finns ett samband mellan bildandet av konjugat och transient confinement zones (TCZs) i dessa trajektorier in vitro. TCZs förekommer också i in vivo-experiment där de visar stora likheter med in vitro-situationen. Denna strategi är ett nytt grepp inom dataanalys-metoder för att spåra immunceller.

Place, publisher, year, edition, pages
Stockholm: KTH , 2009. , viii, 72 p.
Series
Trita-FYS, ISSN 0280-316X ; 2009:60
National Category
Biophysics
Identifiers
URN: urn:nbn:se:kth:diva-11368ISBN: 978-91-7415-492-4 (print)OAI: oai:DiVA.org:kth-11368DiVA: diva2:274728
Public defence
2009-11-20, FD5, AlbaNova, Roslagstullsbacken 21, Stockholm, 13:00 (English)
Opponent
Supervisors
Note
QC 20100726Available from: 2009-11-03 Created: 2009-10-30 Last updated: 2010-07-26Bibliographically approved
List of papers
1. Microfluidic devices for studies of primary cilium mediated cellular response to dynamic flow conditions
Open this publication in new window or tab >>Microfluidic devices for studies of primary cilium mediated cellular response to dynamic flow conditions
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2008 (English)In: Biomedical microdevices (Print), ISSN 1387-2176, E-ISSN 1572-8781, Vol. 10, no 4, 555-560 p.Article in journal (Refereed) Published
Abstract [en]

We present the first microfabricated microfluidic devices designed specifically for studies of primary cilium mediated cellular response to dynamic flow. The primary cilium functions as a mechano-sensor in renal tubular epithelium, sensing the extracellular fluid flow. Malfunction of cilia has been implicated in e.g. polycystic kidney disease and other pathological conditions. Bending of the primary cilium by fluid flow has been shown to give rise to an intracellular calcium signal, however little is known about the sensitivity to flow duration, magnitude and direction. This paper presents a novel method for studying cilia forming cells in asymmetric microfluidic environments. The microfluidic devices presented here were designed for a dynamic control of the local fluid flow on a cellular level, and thus, enables studies of cellular responses to an amplitude, frequency and direction controlled cilium movement.

Keyword
cilia; primary cilium; microfluidic; flow sensitivity
National Category
Engineering and Technology
Identifiers
urn:nbn:se:kth:diva-11359 (URN)10.1007/s10544-008-9165-8 (DOI)000257546800010 ()18236160 (PubMedID)2-s2.0-47749089423 (Scopus ID)
Note
QC 20100723Available from: 2009-10-30 Created: 2009-10-30 Last updated: 2017-12-12Bibliographically approved
2. Mechanical Properties of Primary Cilia Regulate the Response to Fluid flow
Open this publication in new window or tab >>Mechanical Properties of Primary Cilia Regulate the Response to Fluid flow
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2010 (English)In: American Journal of Physiology - Renal Physiology, ISSN 0363-6127, E-ISSN 1522-1466, Vol. 298, no 5, 1096-1102 p.Article in journal (Refereed) Published
Abstract [en]

The primary cilium is a ubiquitous organelle present on most mammalian cells. Malfunction of the organelle has been associated with various pathological disorders, many of which lead to cystic disorders in liver, pancreas, and kidney. Primary cilia have in kidney epithelial cells been observed to generate intracellular calcium in response to fluid flow, and disruption of proteins involved in this calcium signaling lead to autosomal dominant polycystic kidney disease, implying a direct connection between calcium signaling and cyst formation. It has also been shown that there is a significant lag between the onset of flow and initiation of the calcium signal. The present study focuses on the mechanics of cilium bending and the resulting calcium signal. Visualization of real-time cilium movements in response to different types of applied flow showed that the bending is fast compared with the initiation of calcium increase. Mathematical modeling of cilium and surrounding membrane was performed to deduce the relation between bending and membrane stress. The results showed a delay in stress buildup that was similar to the delay in calcium signal. Our results thus indicate that the delay in calcium response upon cilia bending is caused by mechanical properties of the cell membrane.

Keyword
calcium; frequency; delayed membrane stress; modeling
National Category
Condensed Matter Physics
Identifiers
urn:nbn:se:kth:diva-11367 (URN)10.1152/ajprenal.00657.2009 (DOI)000276870800004 ()2-s2.0-77951616289 (Scopus ID)
Note
QC 20100726Available from: 2009-10-30 Created: 2009-10-30 Last updated: 2017-12-12Bibliographically approved
3. Modeling the impact of store-operated Ca2+ entry on intracellular Ca2+ oscillations
Open this publication in new window or tab >>Modeling the impact of store-operated Ca2+ entry on intracellular Ca2+ oscillations
2006 (English)In: Mathematical biosciences, ISSN 0025-5564, Vol. 204, no 2, 232-249 p.Article in journal (Refereed) Published
Abstract [en]

Calcium (Ca2+) oscillations play fundamental roles in various cell signaling processes and have been the subject of numerous modeling studies. Here we have implemented a general mathematical model to simulate the impact of store-operated Ca2+ entry on intracellular Ca2+ oscillations. In addition, we have compared two different models of the inositol 1,4,5-trisphosphate (IP3) receptor (IP3R) and their influences on intracellular Ca2+ oscillations. Store-operated Ca2+ entry following Ca2+ depletion of endoplasmic reticulum (ER) is an important component of Ca2+ signaling. We have developed a phenomenological model of store-operated Ca2+ entry via store-operated Ca2+ (SOC) channels, which are activated upon ER Ca2+ depletion. The depletion evokes a bi-phasic Ca2+ signal, which is also produced in our mathematical model. The IP3R is an important regulator of intracellular Ca2+ signals. This IP3 sensitive Ca2+ channel is also regulated by Ca2+. We apply two IP3R models, the Mak-McBride-Foskett model and the De Young and Keizer model, with significantly different channel characteristics. Our results show that the two separate IP3R models evoke intracellular Ca2+ oscillations with different frequencies and amplitudes. Store-operated Ca2+ entry affects the oscillatory behavior of these intracellular Ca2+ oscillations. The IP3 threshold is altered when store-operated Ca2+ entry is excluded from the model. Frequencies and amplitudes of intracellular Ca2+ oscillations are also altered without store-operated Ca2+ entry. Under certain conditions, when intracellular Ca2+ oscillations are absent, excluding store-operated Ca2+ entry induces an oscillatory response. These findings increase knowledge concerning store-operated Ca2+ entry and its impact on intracellular Ca2+ oscillations.

Keyword
calcium; oscillation; SOC
National Category
Condensed Matter Physics
Identifiers
urn:nbn:se:kth:diva-7064 (URN)10.1016/j.mbs.2006.03.001 (DOI)000242902200005 ()2-s2.0-33750721337 (Scopus ID)
Note
QC 20100726Available from: 2007-05-08 Created: 2007-05-08 Last updated: 2010-12-06Bibliographically approved
4. Erythropoietin modulation of astrocyte water permeability as a component of neuroprotection.
Open this publication in new window or tab >>Erythropoietin modulation of astrocyte water permeability as a component of neuroprotection.
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2009 (English)In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 1091-6490, Vol. 106, no 5, 1602-7 p.Article in journal (Refereed) Published
Abstract [en]

Disturbed brain water homeostasis with swelling of astroglial cells is a common complication in stroke, trauma, and meningitis and is considered to be a major cause of permanent brain damage. Astroglial cells possess the water channel aquaporin 4 (AQP4). Recent studies from our laboratory have shown that glutamate, acting on group I metabotropic glutamate receptors (mGluRs), increases the permeability of astrocyte AQP4, which, in situations of hypoxia-ischemia, will increase astrocyte water uptake. Here we report that erythropoietin (EPO), which in recent years has emerged as a potent neuro-protective agent, antagonizes the effect of a group I mGluR agonist on astrocyte water permeability. Activation of group I mGluRs triggers fast and highly regular intracellular calcium oscillations and we show that EPO interferes with this signaling event by altering the frequency of the oscillations. These effects of EPO are immediate, in contrast to the neuroprotective effects of EPO that are known to depend upon gene activation. Our findings indicate that EPO may directly reduce the risk of astrocyte swelling in stroke and other brain insults. In support of this conclusion we found that EPO reduced the neurological symptoms in a mouse model of primary brain edema known to depend upon AQP4 water transport.

Keyword
Aquaporin 4; Brain edema; Glutamate
National Category
Cell and Molecular Biology Neurosciences Cell Biology
Identifiers
urn:nbn:se:kth:diva-11360 (URN)10.1073/pnas.0812708106 (DOI)000263074600056 ()19164545 (PubMedID)2-s2.0-60849093177 (Scopus ID)
Note

QC 20150727

Available from: 2009-10-30 Created: 2009-10-30 Last updated: 2015-07-27Bibliographically approved
5. Role of diffusion limited space on water and salt homeostasis
Open this publication in new window or tab >>Role of diffusion limited space on water and salt homeostasis
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(English)Manuscript (preprint) (Other academic)
National Category
Condensed Matter Physics
Identifiers
urn:nbn:se:kth:diva-11365 (URN)
Note
QC 20100726Available from: 2009-10-30 Created: 2009-10-30 Last updated: 2010-07-27Bibliographically approved
6. Allosteric changes of the NMDA receptor trap diffusible dopamine 1 receptors in spines
Open this publication in new window or tab >>Allosteric changes of the NMDA receptor trap diffusible dopamine 1 receptors in spines
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2006 (English)In: Proceedings of the National Academy of Sciences of the United States of America, ISSN 0027-8424, E-ISSN 1091-6490, Vol. 103, no 3, 762-767 p.Article in journal (Refereed) Published
Abstract [en]

The dopaminergic and glutamatergic systems interact to initiate and organize normal behavior, a communication that may be perturbed in many neuropsychiatric diseases, including schizophrenia. We show here that NMDA, by allosterically modifying NMDA receptors, can act as a scaffold to recruit laterally diffusing dopamine D1 receptors (D1R) to neuronal spines. Using organotypic culture from rat striatum transfected with D1R fused to a fluorescent protein, we show that the majority of dendritic D1R are in lateral diffusion and that their mobility is confined by interaction with NMDA receptors. Exposure to NMDA reduces the diffusion coefficient for D1R and causes an increase in the number of D1R-positive spines. Unexpectedly, the action of NMDA in potentiating D1R recruitment was independent of calcium flow via the NMDA receptor channel. Thus, a highly energy-efficient, diffusion-trap mechanism can account for intraneuronal interaction between the glutamatergic and dopaminergic systems and for regulation of the number of D1R-positive spines. This diffusion trap system represents a molecular mechanism for brain plasticity and offers a promising target for development of antipsychotic therapy

Keyword
Fluorescence recovery after photo-bleaching; Lateral diffusion; Organotypic cultures; Receptor movement
National Category
Condensed Matter Physics
Identifiers
urn:nbn:se:kth:diva-7063 (URN)10.1073/pnas.0505557103 (DOI)000234727800047 ()2-s2.0-31444439409 (Scopus ID)
Note
QC 20100726Available from: 2007-05-08 Created: 2007-05-08 Last updated: 2010-11-11Bibliographically approved
7. Integrin-mediated cell attachment induces a PAK4-dependent feedback loop regulating cell adhesion through modified integrin alpha v beta 5 clustering and turnover
Open this publication in new window or tab >>Integrin-mediated cell attachment induces a PAK4-dependent feedback loop regulating cell adhesion through modified integrin alpha v beta 5 clustering and turnover
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2010 (English)In: Molecular Biology of the Cell, ISSN 1059-1524, E-ISSN 1939-4586, Vol. 21, no 19, 3317-3329 p.Article in journal (Refereed) Published
Abstract [en]

Cell-to-extracellular matrix adhesion is regulated by a multitude of pathways initiated distally to the core cell-matrix adhesion machinery, such as via growth factor signaling. In contrast to these extrinsically sourced pathways, we now identify a regulatory pathway that is intrinsic to the core adhesion machinery, providing an internal regulatory feedback loop to fine tune adhesion levels. This autoinhibitory negative feedback loop is initiated by cell adhesion to vitronectin, leading to PAK4 activation, which in turn limits total cell-vitronectin adhesion strength. Specifically, we show that PAK4 is activated by cell attachment to vitronectin as mediated by PAK4 binding partner integrin alpha v beta 5, and that active PAK4 induces accelerated integrin alpha v beta 5 turnover within adhesion complexes. Accelerated integrin turnover is associated with additional PAK4-mediated effects, including inhibited integrin alpha v beta 5 clustering, reduced integrin to F-actin connectivity and perturbed adhesion complex maturation. These specific outcomes are ultimately associated with reduced cell adhesion strength and increased cell motility. We thus demonstrate a novel mechanism deployed by cells to tune cell adhesion levels through the autoinhibitory regulation of integrin adhesion.

National Category
Cell Biology
Identifiers
urn:nbn:se:kth:diva-11366 (URN)10.1091/mbc.E10-03-0245 (DOI)000282275600004 ()2-s2.0-77958031728 (Scopus ID)
Funder
Swedish Research CouncilKnut and Alice Wallenberg Foundation
Note

QC 20100726

Available from: 2009-10-30 Created: 2009-10-30 Last updated: 2017-12-12Bibliographically approved
8. Single Cell Tracking of Natural Killer CellMigration in vivo and in vitro reveals Transient Migration Arrest Periods
Open this publication in new window or tab >>Single Cell Tracking of Natural Killer CellMigration in vivo and in vitro reveals Transient Migration Arrest Periods
Show others...
(English)Manuscript (preprint) (Other (popular science, discussion, etc.))
National Category
Biophysics
Identifiers
urn:nbn:se:kth:diva-11362 (URN)
Note
QC 20100726Available from: 2009-10-30 Created: 2009-10-30 Last updated: 2010-07-26Bibliographically approved

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