The loading plot (Figure 1B) revealed that signals at 3.04 ppm and 3.94 ppm dominates the discrimination, and this can be ascribed to a higher content of creatine in the treated cells, confirming the expected increased incorporation of creatine into the myotubes. The myotube protein expression in response

to creatine was analyzed by proteomics using selleck products 2-DGE. An obtained proteomic profile of myotube extracts is shown in Figure 2. Figure 1 PLS-DA scores plot of NMR-based metabonomic data. (A) PLS-DA scores plot from analysis of NMR-based metabonomic data obtained on extracts of control (open circles) and creatine monohydrate (CMH) treated C2C12 muscle cells (closed circles), (B) the X-loadings of the PLS-DA. The dominating signals at 3.04 and 3.94 ppm are ascribed to CH3 and CH2 in creatine, respectively. Vadimezan purchase The arrow shows a signal at 2.40 ppm, which was also found to

be significant in the discrimination of control and CMH-treated cells. The 2.40 ppm signal is tentatively assigned to malate. Figure 2 Proteomic profile of myotubes. Proteomic profile of myotubes as analyzed by 2-DGE visualized by silver staining. The positions of protein spots identified to be significantly different in controls and in creatine monohydrate-treated myotubes by PLS-DA of 2-DGE proteomics data are indicated. After the manual check of the automatically assigned number of spots, a total of 584 protein spots were annotated by the image analysis and used in the why further statistical

analysis. By PLS-DA, 28 proteins were found to be differentially expressed when comparing CMH-treated myotubes with the control myotubes (results not shown). The significance of the spots identified by the PLS-DA was further tested by statistical see more t-test (Table 1). Of the 28 protein spots in the PLS-DA model, 20 of these were found to be either significantly different (P < 0.05) or exhibited tendency to be significantly different (P < 0.1) by the t-test. Accordingly, the t-test confirms that the intensities of the majority of the spots identified by PLS-DA are considerably affected by CMH treatment. Of these, 13 were up-regulated by CMH treatment, while 7 were down-regulated. This shows, as probably expected, that CMH stimulates the expression of more proteins than it down-regulates. The spots which were identified by the t-test to be differentially expressed in the myotubes in response to CMH treatment were cut out from the gels, and subjected to MALDI-TOF MS analysis using peptide mass fingerprinting. Those protein spots which were identified by MS are listed in Table 2. The identified proteins include vimentin, malate dehydrogenase, peroxiredoxin, thioredoxin dependent peroxide reductase, 75 kDa and 78 kDa glucose regulated protein precursors.