Surface engineering of living myoblasts via selective periodate oxidation.
Cell surface molecules are vital for normal cell activity. To study the functions of these molecules or manipulate cell behavior, the ability to decorate cell surfaces with bioactive molecules of our choosing is a potentially powerful technique. Here, we describe the molecular engineering of living L6 myoblast monolayers via selective periodate oxidation of sialic acid residues and the application of this surface modification in the artificial aggregation of cells. The aldehyde groups generated by this reaction were used to selectively ligate a model molecule, biotin hydrazide, to the cell surfaces. Flow cytometry analysis after staining with fluorescently conjugated avidin revealed a concentration-dependent increase in fluorescence compared to untreated cells with a maximal shift of 345.1 +/- 27.4-fold and an EC(50) of 17.4 +/- 1.1 microM. This mild oxidation reaction did not affect cell number, viability, or morphology. We then compared this chemical technique with the metabolic incorporation of reactive cell surface ketone groups using N-levulinoylmannosamine (ManLev). In this cell line, only a 22.3-fold fluorescence shift was observed compared to untreated cells when myoblasts were incubated with a high concentration of ManLev for 48 hours. Periodate oxidation was then used to modify myoblast surfaces to induce cell aggregation. Crosslinking biotinylated myoblasts, which do not spontaneously aggregate in culture, with avidin resulted in the rapid formation of millimeter-sized, multicellular structures. These data indicate that sodium periodate treatment is an effective, noncytotoxic method for the in vitro molecular engineering of living cell surfaces with the potential for cell biology and tissue engineering applications.
De Bank
P A
PA
School of Pharmaceutical Sciences, University of Nottingham, University Park, Nottingham, NG7 2RD, United Kingdom.
Kellam
B
B
Kendall
D A
DA
Shakesheff
K M
KM
D000447:Aldehydes / Q000378:metabolism; D000818:Animals; D001710:Biotin / Q000031:analogs & derivatives* / Q000378:metabolism; D002449:Cell Aggregation / Q000187:drug effects* / Q000502:physiology; D002452:Cell Count; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D005240:Feasibility Studies; D005434:Flow Cytometry; D006595:Hexosamines / Q000494:pharmacology; D007659:Ketones / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D032446:Myoblasts / Q000166:cytology / Q000187:drug effects* / Q000502:physiology* / Q000648:ultrastructure; D010084:Oxidation-Reduction; D010504:Periodic Acid / Q000494:pharmacology*; D051381:Rats; D023822:Tissue Engineering / Q000379:methods
D000447:Aldehydes; D006595:Hexosamines; D007659:Ketones; D008565:Membrane Proteins; C105871:N-levulinoyl mannosamine; D010504:Periodic Acid; C053311:biotin hydrazide; D001710:Biotin; C009288:metaperiodate