31P NMR studies of cultured human tumor cells. Influence of pH on phospholipid metabolite levels and the detection of cytidine 5'-diphosphate choline.
202 MHz 31P NMR (11.7 T) was used to study the effects of culture medium pH on the levels of phosphate metabolites in three human tumor cell lines (XP29MAmal, a malignant xeroderma pigmentosum; CX-1, a colon carcinoma; KB, a squamous cell carcinoma of the oral cavity). Cells were cultured in Roux flasks in HAM's F-12 medium, and the pH was varied with the final medium change. After harvesting, 1-5 x 10(8) cells were suspended in Ringer/HEPES buffer at pH 7.4 and 4 degrees C for 31P NMR studies. Cell adhesion and growth rate decreased with decreasing pH, but, down to ca pH 6.1, trypan blue exclusion and the observed levels of nucleoside di- and triphosphates (range: 22-37% of total phosphates detected), phosphocreatine (PCr, 2-5%) and Pi (5-11%) did not vary significantly with pH. For XP29MAmal cells in exponential growth phosphocholine levels decreased from 18-28% at pH 7.0 to ca 5% at pH 6.0, while phosphoethanolamine levels increased from 2-7% to 15%. Glycerophosphocholine (GPC) levels increased from ca 7% at pH 7.2 to 13% at pH 6.3. At pH less than 6.3 cytidine 5'-diphosphate (CDP) choline became detectable (8-16%, delta p:P alpha = -8.13 ppm, P beta = 8.93 ppm, for PCr = 0 ppm). However, confluent cells did not accumulate CDP-choline when the pH was lowered. The cell lines CX-1 and KB also showed the pH effects described herein.
Kuesel
A C
AC
Central Spectroscopy Department, German Cancer Research Center, Heidelberg.
Graschew
G
G
Hull
W E
WE
Lorenz
W
W
Thielmann
H W
HW
D002794:Choline / Q000031:analogs & derivatives*; D003470:Culture Media; D003566:Cytidine Diphosphate Choline / Q000378:metabolism*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D009682:Magnetic Resonance Spectroscopy; D010743:Phospholipids / Q000378:metabolism*; D014407:Tumor Cells, Cultured / Q000378:metabolism*
D003470:Culture Media; D010743:Phospholipids; D003566:Cytidine Diphosphate Choline; D002794:Choline