Stamp wound assay for studying coupled cell migration and cell debris clearance.

A new method for studying wound healing under realistic conditions in vitro was developed. The method involves creating defined patterns of damaged cell debris with poly(dimethyl)siloxane (PDMS) stamping. This novel assay permitted the quantification of wound healing rates in the presence of cell debris. Experimental results with this assay suggest that cell migration in the presence of cell debris is a two step process requiring (1) non-muscle myosin II-dependent cell clearance followed by (2) cell migration into newly cleared wound areas. The novel stamp wound assay allows the study of coupled cell migration and debris clearance and is a more realistic wound healing assay in vitro.

[1]  Amy Brock,et al.  Geometric determinants of directional cell motility revealed using microcontact printing. , 2003, Langmuir : the ACS journal of surfaces and colloids.

[2]  J. Klarlund,et al.  Wounding Induces Motility in Sheets of Corneal Epithelial Cells through Loss of Spatial Constraints , 2004, Journal of Biological Chemistry.

[3]  T. Mitchison,et al.  A high-throughput cell migration assay using scratch wound healing, a comparison of image-based readout methods , 2004, BMC biotechnology.

[4]  K. Beningo,et al.  A contractile activity that closes phagosomes in macrophages. , 1999, Journal of cell science.

[5]  G. Whitesides,et al.  Soft lithography in biology and biochemistry. , 2001, Annual review of biomedical engineering.

[6]  J. Fullard,et al.  Clearance of apoptotic corpses , 2009, Apoptosis.

[7]  Wei He,et al.  α4β1 Integrin Regulates Lamellipodia Protrusion via a Focal Complex/Focal Adhesion-independent Mechanism , 2002 .

[8]  A. Barbul,et al.  General principles of wound healing. , 1997, The Surgical clinics of North America.

[9]  G. Whitesides,et al.  Soft Lithography. , 1998, Angewandte Chemie.

[10]  P. Hebda,et al.  Stimulatory effects of transforming growth factor-beta and epidermal growth factor on epidermal cell outgrowth from porcine skin explant cultures. , 1988, The Journal of investigative dermatology.

[11]  P. Chavrier,et al.  Collective migration of an epithelial monolayer in response to a model wound , 2007, Proceedings of the National Academy of Sciences.

[12]  Dennis E. Discher,et al.  Inhibition of “self” engulfment through deactivation of myosin-II at the phagocytic synapse between human cells , 2008, The Journal of cell biology.

[13]  R. Birge,et al.  Innate apoptotic immunity: the calming touch of death , 2008, Cell Death and Differentiation.

[14]  F. Ren,et al.  Randomly oriented, upright SiO2 coated nanorods for reduced adhesion of mammalian cells. , 2009, Biomaterials.

[15]  M. Chaplain,et al.  A novel 'sandwich' assay for quantifying chemo-regulated cell migration within 3-dimensional matrices: wound healing cytokines exhibit distinct motogenic activities compared to the transmembrane assay. , 2006, Cell motility and the cytoskeleton.

[16]  T. Shaw,et al.  Wound repair at a glance , 2009, Journal of Cell Science.

[17]  I. M. Neiman,et al.  [Inflammation and cancer]. , 1974, Patologicheskaia fiziologiia i eksperimental'naia terapiia.

[18]  G. Whitesides,et al.  Patterning proteins and cells using soft lithography. , 1999, Biomaterials.

[19]  J. Scholes,et al.  Differences between the clearance of apoptotic cells by professional and non-professional phagocytes , 2000, Current Biology.

[20]  A. Bresnick,et al.  Rap1 activation in collagen phagocytosis is dependent on nonmuscle myosin II-A. , 2008, Molecular biology of the cell.

[21]  J. Forrester,et al.  Electric fields and MAP kinase signaling can regulate early wound healing in lens epithelium. , 2003, Investigative ophthalmology & visual science.

[22]  J. Guan,et al.  Wound-healing assay. , 2005, Methods in molecular biology.

[23]  C. Lo,et al.  Monitoring motion of confluent cells in tissue culture. , 1993, Experimental cell research.

[24]  Jan Feijen,et al.  A microfluidic wound-healing assay for quantifying endothelial cell migration. , 2010, American journal of physiology. Heart and circulatory physiology.

[25]  A. Bresnick,et al.  Rap 1 Activation in Collagen Phagocytosis Is Dependent on Nonmuscle Myosin IIA , 2008 .

[26]  R. Chandawarkar,et al.  In vivo delivery of heat shock protein 70 accelerates wound healing by up‐regulating macrophage‐mediated phagocytosis , 2006, Wound repair and regeneration : official publication of the Wound Healing Society [and] the European Tissue Repair Society.

[27]  Alan Hall,et al.  Rho GTPases: biochemistry and biology. , 2005, Annual review of cell and developmental biology.

[28]  Wei He,et al.  alpha4beta1 integrin regulates lamellipodia protrusion via a focal complex/focal adhesion-independent mechanism. , 2002, Molecular biology of the cell.

[29]  Niels Galjart,et al.  Differential effects of matrix and growth factors on endothelial and fibroblast motility: Application of a modified cell migration assay , 2006, Journal of cellular biochemistry.

[30]  M. Sheetz,et al.  Two distinct modes of myosin assembly and dynamics during epithelial wound closure , 2007, The Journal of cell biology.

[31]  R. Dickinson,et al.  Sarcomere mechanics in capillary endothelial cells. , 2009, Biophysical journal.

[32]  Joachim Wegener,et al.  Electrical wound-healing assay for cells in vitro. , 2004, Proceedings of the National Academy of Sciences of the United States of America.