The influence of medium formulation on phosphomonoester and UDP-hexose levels in cultured human colon tumor cells as observed by 31P NMR spectroscopy.

High-resolution 31P NMR spectroscopy at 11.7 T was used to examine the influence of medium formulation (medium and serum type, and concentrations of glucose and inositol) on the cellular phosphate metabolism of CX-1 cells, a human colon cancer cell line derived from HT-29 cells. Striking differences in the 31P spectra of harvested CX-1 cells were observed. The largest variation was seen in the phosphocholine and UDP-hexose levels (up to seven-fold changes), with smaller differences in the levels of other phosphate metabolites. The major UDP-hexose species were found to be UDP-N-acetylglucosamine and UDP-N-acetylgalactosamine (ca 2:1 ratio), which have been proposed in the literature to be markers of cell differentiation status. Medium-induced alterations in metabolite levels were much greater than the normal variations seen in CX-1 control samples grown under identical conditions. They even exceeded the characteristic differences observed between different human tumor cell lines grown under one set of culture conditions. The remarkable sensitivity of CX-1 cellular phosphate metabolism to the culture environment has implications for the comparison of in vitro vs in vivo spectra, and for the interpretation of effects due to growth and therapy.

Characterization of a microcarrier cell culture system for 23Na MR spectroscopy studies.

A MR spectroscopy method is described for the simultaneous discrimination and observation of sodium from the three compartments created by an intact cell monolayer. Results are reported for Madin Darby Canine Kidney (MDCK) cells, an epithelial-like continuous cell line, cultured on Cytodex 1 microcarrier beads and perfused with medium containing 6 mM dysprosium (III) tripolyphosphate [Dy(TPP)2(7-)] as shift reagent. The sodium spectrum shows three resonances which are assigned to the shifted intrabead (basolateral) and extrabead (apical) pools and the unshifted intracellular pool. Ouabain inhibition of the Na(+)-K(+)-ATPase cellular pump mechanism was used to demonstrate the sensitivity of the method for monitoring intracellular sodium. The supported MDCK cells in this system remained viable after exposure for 5 h to medium containing Dy(TPP)2(7-) at a concentration of 6 mM, as determined by trypan blue dye exclusion and by comparison of the log growth rate and ability to form domes in subsequent generations of exposed cells vs unexposed controls.

Use of gel filtration in the preparation of biological fluids for magnetic resonance spectroscopy.

Analysis of biological fluids by proton magnetic resonance spectroscopy is often complicated by dynamic range problems created from the large water resonance. Gel filtration chromatography is found to be a simple and nondestructive method for exchanging D2O for H2O and for removing low molecular weight molecules from both plasma and urine, significantly improving subsequent one- and two-dimensional MRS spectra.