Freyer J P, Schor P L, Jarrett K A, Neeman M, Sillerud L O
Cellular and Molecular Biology Group, Los Alamos National Laboratory, New Mexico 87545.
Cancer Res. 1991 Aug 1;51(15):3831-7.
We have measured the 31P nuclear magnetic resonance spectra of EMT6/Ro multicellular tumor spheroids over a wide range of sizes under constant nutrient conditions which matched those used for culturing the spheroids. The amount of nucleotide triphosphate per cell decreased with spheroid growth, roughly in proportion to the decrease in cell volume. There was no correlation between the intracellular pH, the nucleotide triphosphate:Pi ratio, or the phosphocreatine:Pi ratio and either the spheroid cellularity, the mean cell volume, the S-phase fraction, the clonogenic capacity, or the amount of central necrosis. The phosphoryethanolamine:phosphorylcholine ratio also increased with increasing spheroid size. There was a negative correlation between the phosphoryethanolamine:phosphorylcholine ratio and the S-phase cell fraction or the mean cell volume; this ratio was positively correlated with the extent of central necrosis. The membrane degradation components glycerophosphorylcholine and glycerophosphorylethanolamine showed no significant changes with increasing spheroid size. These results imply that spheroid necrotic areas induced by chronic nutrient deficiencies are "invisible" to 31P nuclear magnetic resonance and that the development of cellular quiescence in spheroids is not caused by a decrease in the steady-state level of high-energy phosphates or a reduced intracellular pH. Together, these data support a model in which cells maintain normal steady-state levels of high energy phosphates until they are very close to necrotic cell death. This implies that the deterioration of 31P nuclear magnetic resonance spectra of tumors with increasing size is not caused by chronic nutrient deficiencies resulting from cells outgrowing the capillary supply, but rather is more related to transient nutrient deprivation phenomena.
我们在与培养球体所用条件相匹配的恒定营养条件下，测量了不同大小的EMT6/Ro多细胞肿瘤球体的31P核磁共振谱。每个细胞的三磷酸核苷酸量随球体生长而减少，大致与细胞体积的减少成比例。细胞内pH值、三磷酸核苷酸与无机磷酸（Pi）的比率或磷酸肌酸与Pi的比率，与球体细胞密度、平均细胞体积、S期细胞比例、克隆形成能力或中央坏死量之间均无相关性。磷酸乙醇胺与磷酸胆碱的比率也随球体大小的增加而升高。磷酸乙醇胺与磷酸胆碱的比率与S期细胞比例或平均细胞体积呈负相关；该比率与中央坏死程度呈正相关。膜降解成分甘油磷酰胆碱和甘油磷酰乙醇胺随球体大小增加无显著变化。这些结果表明，慢性营养缺乏诱导的球体坏死区域在31P核磁共振中是“不可见的”，并且球体中细胞静止状态的发展不是由高能磷酸盐稳态水平的降低或细胞内pH值的降低引起的。总之，这些数据支持一种模型，即细胞在非常接近坏死性细胞死亡之前维持高能磷酸盐的正常稳态水平。这意味着肿瘤31P核磁共振谱随大小增加而恶化不是由细胞生长超过毛细血管供应导致的慢性营养缺乏引起的，而是更与短暂的营养剥夺现象有关。
