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Dive into the research topics where Mike A. Horton is active.

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Featured researches published by Mike A. Horton.


Biophysical Journal | 2002

Single Cell Mechanotransduction and Its Modulation Analyzed by Atomic Force Microscope Indentation

Guillaume Charras; Mike A. Horton

The skeleton adapts to its mechanical usage, although at the cellular level, the distribution and magnitude of strains generated and their detection are ill-understood. The magnitude and nature of the strains to which cells respond were investigated using an atomic force microscope (AFM) as a microindentor. A confocal microscope linked to the setup enabled analysis of cellular responses. Two different cell response pathways were identified: one, consequent upon contact, depended on activation of stretch-activated ion channels; the second, following stress relaxation, required an intact microtubular cytoskeleton. The cellular responses could be modulated by selectively disrupting cytoskeletal components thought to be involved in the transduction of mechanical stimuli. The F-actin cytoskeleton was not required for responses to mechanical strain, whereas the microtubular and vimentin networks were. Treatments that reduced membrane tension, or its transmission, selectively reduced contact reactions. Immunostaining of the cell cytoskeleton was used to interpret the results of the cytoskeletal disruption studies. We provide an estimate of the cellular strain magnitude needed to elicit intracellular calcium responses and propose a model that links single cell responses to whole bone adaptation. This technique may help to understand adaptation to mechanical usage in other organs.


Biophysical Journal | 2002

Determination of cellular strains by combined atomic force microscopy and finite element modeling.

Guillaume Charras; Mike A. Horton

Many organs adapt to their mechanical environment as a result of physiological change or disease. Cells are both the detectors and effectors of this process. Though many studies have been performed in vitro to investigate the mechanisms of detection and adaptation to mechanical strains, the cellular strains remain unknown and results from different stimulation techniques cannot be compared. By combining experimental determination of cell profiles and elasticities by atomic force microscopy with finite element modeling and computational fluid dynamics, we report the cellular strain distributions exerted by common whole-cell straining techniques and from micromanipulation techniques, hence enabling their comparison. Using data from our own analyses and experiments performed by others, we examine the threshold of activation for different signal transduction processes and the strain components that they may detect. We show that modulating cell elasticity, by increasing the F-actin content of the cytoskeleton, or cellular Poisson ratio are good strategies to resist fluid shear or hydrostatic pressure. We report that stray fluid flow in some substrate-stretch systems elicits significant cellular strains. In conclusion, this technique shows promise in furthering our understanding of the interplay among mechanical forces, strain detection, gene expression, and cellular adaptation in physiology and disease.


Ultramicroscopy | 2000

Adapting atomic force microscopy for cell biology

Petri Lehenkari; Guillaume Charras; Mike A. Horton

We present details of our AFM modifications to produce an adaptable imaging system for the cell biologist. We have designed and validated a new inverted microscope interface and a scan head with increased Z-range, based upon the TopoMetrix Explorer AFM. We have utilised these changes, together with home-made glass ball cantilevers, to obtain topographical information over cells with large Z-dimension (over 15 microm high), and mapped calcitonin-calcitonin receptor binding forces in living bone cells. We conclude that modified AFM can be used to evaluate intermolecular events in living cells and that this approach will ensure general application to the study of receptor-ligand interactions under truly physiological conditions.


Ultramicroscopy | 2001

Atomic force microscopy can be used to mechanically stimulate osteoblasts and evaluate cellular strain distributions

Guillaume Charras; Petri Lehenkari; Mike A. Horton

In this study, atomic force microscopy (AFM) was used to mechanically stimulate primary osteoblasts. In response to mechanical force applied by the AFM, the indented cell increased its intracellular calcium concentration. The material properties of the cell could be estimated and the membrane strains calculated. We proceeded to validate this technique experimentally and a 20% error was found between the predicted and the measured diameter of indentation. We also determined the strain distributions within the cell that result from AFM indentation using a simple finite element model. This enabled us to formulate hypotheses as to the mechanism through which cells may sense the applied mechanical strains. Finally, we report the effect of the Poisson ratio and the cell thickness on the strain distributions. Varying the Poisson ratio did not change the order of magnitude of the strains; whereas the cellular thickness dramatically changed the order of magnitude of the cellular strains. We conclude that AFM can be used for controlled mechanical stimulation of osteoblasts and that cellular strain distributions can be computed with a good accuracy when the cell is indented in its highest part.


Journal of Bone and Mineral Research | 2002

Nuclear Localization of Type I Parathyroid Hormone/Parathyroid Hormone‐Related Protein Receptors in Deer Antler Osteoclasts: Evidence for Parathyroid Hormone‐Related Protein and Receptor Activator of NF‐κB‐Dependent Effects on Osteoclast Formation in Regenerating Mammalian Bone

C. Faucheux; Mike A. Horton; J. S. Price

Parathyroid hormone‐related protein (PTHrP) is not required for osteoclastogenesis during embryonic development; however, after birth it has been shown to regulate osteoclast formation during tooth eruption. Our study explores the hypothesis that PTHrP also may regulate osteoclast differentiation in the regenerating skeletal tissues of deer antlers, bones capable of complete regeneration. Osteoclast‐like multinucleated cells (MNCs) formed spontaneously in micromass cultures derived from antler cartilage and these cells had the phenotypic characteristics of osteoclasts. PTHrP and receptor activator of NF‐κB ligand (RANKL) stimulated antler osteoclast formation although the effect of RANKL was less marked than that of PTHrP. The addition of osteoprotegerin (OPG) only partially decreased (by ∼65%) the number of osteoclasts in PTHrP‐treated cultures. To determine whether PTHrP also potentially could have direct effects on antler osteoclasts, we studied, by confocal microscopy, the expression of the type I PTH/PTHrP receptor (PTH1R) in MNCs cultured on glass and found the receptor protein to have a nuclear localization. In situ hybridization showed that antler MNCs also expressed PTH1R and PTHrP messenger RNAs (mRNAs). PTHrP was immunolocalized in MNCs cultured on glass but was undetectable in cells resorbing a dentine substrate. In tissue sections of antler cartilage, PTHrP and PTH1R were expressed in vitronectin receptor‐positive (VNR+) osteoclast‐like cells localized in the perivascular stroma. Thus, these data show that PTHrP plays a role in the regulation of osteoclast differentiation in regenerating skeletal tissues and that PTHrP can have effects on osteoclastogenesis that are independent of RANKL synthesis. Ours is the first study to describe the expression of the type I PTH/PTHrP receptor in mammalian osteoclasts at a protein and mRNA level, which indicates that PTHrP also may have a direct effect on osteoclasts. This also is the first study to show a nuclear localization of the PTHIR in cells of the osteoclast lineage, although the functional significance of this observation has yet to be established.


Expert Reviews in Molecular Medicine | 2000

New technologies in scanning probe microscopy for studying molecular interactions in cells

Petri Lehenkari; Guillaume Charras; Stephen A. Nesbitt; Mike A. Horton

Atomic force microscopy (AFM) is a specialised form of scanning probe microscopy, which was invented by Binnig and colleagues in 1986. Since then, AFM has been increasingly used to study biomedical problems. Because of its high resolution, AFM has been used to examine the topography or shape of surfaces, such as during the molecular imaging of proteins. This, combined with the ability to operate under known force regimes, makes AFM technology particularly useful for measuring intermolecular bond forces and assessing the mechanical properties of biological materials. Many of the constraints (e.g. complex instrumentation, slow acquisition speeds and poor vertical range) that previously limited the use of AFM in cell biology are now beginning to be resolved. Technological advances will enable AFM to challenge both confocal laser scanning microscopy and scanning electron microscopy as a method for carrying out three-dimensional imaging. Its use as both a precise micro-manipulator and a measurement tool will probably result in many novel and exciting applications in the future. In this article, we have reviewed some of the current biological applications of AFM, and illustrated these applications using studies of the cell biology of bone and integrin-mediated adhesion.


Biophysical Journal | 2004

Estimating the sensitivity of mechanosensitive ion channels to membrane strain and tension

Guillaume Charras; Beatrice A. Williams; Stephen M. Sims; Mike A. Horton


Single Molecules | 2000

Integration of Atomic Force and Confocal Microscopy

Mike A. Horton; Guillaume Charras; Christoph Ballestrem; Petri Lehenkari


Methods in Cell Biology | 2002

Biotechnological applications of atomic force microscopy

Guillaume Charras; Petri Lehenkari; Mike A. Horton


Methods in Cell Biology | 2002

Chapter 8 – Biotechnological Applications of Atomic Force Microscopy

Guillaume Charras; Petri Lehenkari; Mike A. Horton

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C. Faucheux

University College London

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Christoph Ballestrem

Wellcome Trust Centre for Cell-Matrix Research

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J. S. Price

University College London

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Beatrice A. Williams

Canadian Institutes of Health Research

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Stephen M. Sims

Canadian Institutes of Health Research

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