Fig 5. KRT17 upregulates SLC2A1 and glucose uptake.

(A) Representative images of western blot analysis of Ca9/K17+ (C-4), Ca9, HSC3/K17- (Kd-4) and HSC3 for detecting KRT17, SLC2A1, and beta1-tubulin (TUBB). Representative images of three assays. (B) Expression of SLC2A1 in Ca9/K17+ (C-1, C-2, C-3, and C-4) and HSC3/K17- (Kd-1, Kd-2, Kd-3, and Kd-4) compared with Ca9 and HSC3, respectively, as revealed by densitometric analysis of the western blots. Expression level of protein X in a clone was normalized against TUBB and then against the control cells using the formula (Value of protein X in the clone / Value of TUBB in the clone) / (Value of protein X in the control cells / Value of TUBB in the control cells). The normalized expression levels were plotted on a log scale. Representative results of three independent assays. (C) Glucose uptake assay of Ca9 and Ca9/K17+ (C-4) cells as measured by fluorescence microscopy. The cells were treated with a fluorescent glucose analogue for 2 h. The fluorescence image was digitally analyzed and the signal in individual cells was depicted as a brightness unit. The box plot illustrates the maximum, third quartile, median, first quartile and minimum signals of 100 cells. Representative results of three assays. (D) Glucose uptake assay as measured by flow cytometry. The cells were treated with the fluorescent glucose analogue for 2 h and were harvested for flow cytometric analysis (n = 1000). (E) cDNA microarray analysis of KRT17 and SLC2A1 in 43 oral squamous cell carcinomas (OSCCs) and 7 normal controls. SLC2A1 was significantly upregulated in OSCC (P < 0.001), with an approximate three-fold increase both in the mean and the median value. There was a positive correlation between SLC2A1 and KRT17 expression (r = 0.46; ***P < 0.001). The scales on the horizontal and vertical axes represent absolute signal values. Crosses denote each OSCC case and filled circles denote normal controls. (E) Immunohistochemical expression of SLC2A1 in normal tongue epithelium and tongue cancer. Expression was greatest in lymphocytes (arrows in left upper and lower panels). In the normal oral epithelium, SLC2A1 was weakly expressed in the basal and spinous cells (left upper panel). In OSCC, SLC2A1 was upregulated, showing a level of expression comparable with lymphocytes (left and right lower panels). Scale bar, 100 μm.