Figure 7.
(A) Kaplan-Meier survival curve for overall survival of patients with low and high expression of LDHA. mRNA data were obtained from TCGA Liver Hepatocellular Carcinoma dataset. Based on LDHA expression, patients are classified as LDHA high expression and LDHA low expression. The red curve indicates a statistically significant reduction in overall survival with higher LDHA mRNA expression (P < .0071). (B) Schematic representation of experimental protocol 2. (C) Macroscopic view of livers from Ldha WT (left) or KO mice (right), liver-weight-to-body-weight ratio of WT, percentage of the tumor area, and the average of the size of Ldha WT or KO tumors (t test). ∗∗P < .01. Number and tumor size were evaluated in H&E-stained sections as described in the Materials and Methods. The histogram represents the means ± SD of 5 tumors per group (t test). (D) Macroscopic view of livers from Ldha WT treated with tamoxifen or oil, percentage of the tumor area, and liver-weight-to-body-weight ratio. The histograms represent means ± SD of 6 samples/group (t test). ns, not significant. (E) qRT-PCR analysis of Ldha mRNA levels in laser-microdissected WT HCCs compared with Ldha KO tumors. Gene expression is reported as a fold-change of mRNA from WT tumors relative to those from Ldha KO livers. Relative quantification analysis for each gene was calculated by the 2ΔΔCt method. The histogram represents the means ± SD of 3 tumors from 3 mice per group (t test). ∗∗P < .01. (F) Representative microphotographs of immunohistochemical staining for Ki-67 in the tumors (T) or surrounding livers (S) of Ldha WT or Ldha KO mice (top: 5×; bottom: 10×; sections counterstained with hematoxylin). (G) Representative microphotographs showing immunostaining for the cleaved form of CASP3 in the tumors (T) or surrounding liver (S) of Ldha WT and KO mice (CASP3 counterstained with Hematoxylin, Top 20×, Bottom 40×). Arrows indicate single CASP3-positive cells. (H) Apoptotic index (AI). AI was calculated as the number of Casp3-positive cells per field at 40× magnification. From 20 to 40 fields per liver were scored. Results were expressed as means ± SD of 3–4 mice per group (1-way analysis of variance). ∗P < .05.