Noninvasive Evaluation Of Radiation-Enhanced Glioma Cell Invasiveness By Using 14.7-T High Resolution ~1H-MRS

Objective:To study the effect of a range of single X-rays on the activity of MMP-2 and water-soluble metabolites of U87 and C6 glioma multicellular spheroids and obtain the correlation between metabolite characteristics of glioma cells and invasion ability in vitro at the cellular level after irradiation.Methods:To obtain the activity of MMP-2 and metabolite ratios of glioma cells after irradiation with different dosages of X-ray.Irradiation of X-ray at Ogy, 1gy,5gy, 10gy, 15gy exposed U87 and C6 glioma cells.Perchloric acid extracting (PCA) method was used to gain water-soluble metabolites [Choline (Cho), Creatine(Cr), and N-acetylaspartate (NAA)]. The metabolites were observed by using 14.7T 1H-MRS in vitro. Matrigel invasion assay and gelatin zymography was performed to test the invasion ability of U87 and C6 glioma cells. After 20h, serum-free medium from U87 and C6 cells was collected for gelatin zymography. One part of cells from every irradiated sample was used to measure matrigel invasion assay and the other part of cells was used to get’H-MRS patterns by the PCA method.Results:Good MR spectra of PCA method from U87 and C6 glioma cells were obtained. Both radiation-induced MMP-2 activity and the Cho/Cr, Cho/NAA ratios increased after irradiation and their increase in the manner of dosage dependence. The MMP-2 activity and the Cho/Cr, Cho/NAA ratios increased up to 10Gy for glioma cells and decreased thereafter. The Cho/NAA ratio of U87 cells increased from 3.55±.06 (OGy) to 9.13±0.30 (10Gy), and then decline to 5.94±0.15 (15Gy), The Cho/NAA ratio of C6 cells increased from 1.07±0.24 (OGy) to 2.77±0.36 (10Gy), and then decline to 1.73±0.41 (15Gy). The Cho/Cr ratio of U87 cells increased from 2.10±0.29 (OGy) to 4.92±0.07 (10Gy), and then decline to 3.69±0.15 (15Gy), The Cho/Cr ratio of C6 cells increased from 0.38±0.08 (OGy) to 0.79±0.03 (10Gy), and then decline to 0.55±0.08 (15Gy).Further more, the invasion of glioma cells highly correlated positively with the Cho/Cr and Cho/NAA ratios. Both Cho/Cr ratio and Cho/NAA of U87 and C6 glioma cells were highly positive correlated to with the number of invaded cell in the matrigel invasion assay. The Pearson’s correlation coefficient (r) values of U87 cells were 0.89 (Cho/Cr ratio versus invasion ability) and 0.91 (Cho/NAA ratio versus invasion ability) (P<0.01). The Pearson’s correlation coefficient (r) values of C6 cells were 0.80 (Cho/Cr ratio versus invasion ability) and 0.73 (Cho/NAA ratio versus invasion ability) (P＜0.01).Conclusion:High-resolution H-MRS in vitro can detect the subtle biochemical changes at the cellular level. Sublethal irradiation promotes the activity of MMP-2 of U87 MCS and leads the Cho/Cr ratio increase. H-MRS may have the potential to aid in display the tumor invasion range.