METABOLIC CHANGES IN MESENCHYMAL STEM CELLS IN OSTEOGENIC MEDIUM MEASURED BY AUTOFLUORESCENCE SPECTROSCOPY

¹ Department of Opthalmology, Johns Hopkins University, Baltimore, Maryland
² Department of Biomedical Engineering, Johns Hopkins University, Baltimore, Maryland
³ Department of Cell Biology, Johns Hopkins University, Baltimore, Maryland

^* To whom correspondence should be addressed. E-mail: Rchuck1{at}jhmi.edu.

	Abstract

Purpose: To measure metabolic changes in mesenchymal stem cells (MSCs) placed in osteogenic medium by autofluorescence spectroscopy.

Methods: MSCs were plated in stem cell-supporting or osteogenic medium and imaged. Shift from the basic growth environment to the inductive osteogenic environment was confirmed by reverse transcriptase (RT)-PCR. Reduced pyridine nucleotides were detected by exciting near 366 nm and measuring fluorescence at 450nm, and oxidized flavoproteins were detected by exciting at 460 nm and measuring fluorescence at 540nm. The ratio of these fluorescence measurements, redox fluorometry, is a non-invasive measure of the cellular metabolic state.

Results: The detected pyridine nucleotide to flavoprotein ratio decreases upon transitioning from the stem cell to the differentiated state, as well as with increasing cell density and cell-cell contact.

Conclusion: MSC metabolism increases upon placement in differentiating medium, and with increasing cell density and contact. Redox fluorometry is a feasible, non-invasive technique for distinguishing MSCs from further differentiated cells.