Table 1.
Primers used for qPCR gene expression analysis.
Fig 1.
Gene expression dynamics during liver development.
(A) Gene expression pattern of albumin, G6PC, and CYP3A4 in GSE132060, GSE25417, and GSE67848 datasets. (B) Gene expression pattern of NANOG, SOX2, and POU5F1 in GSE132060, GSE25417, and GSE67848 datasets. (C) Gene expression pattern of SLC2A2 in GSE132060, GSE25417, and GSE67848 datasets.
Fig 2.
Changes in SLC2A2 gene expression in hepatocellular carcinoma (HCC) based on its stage and correlation with oncofetal and stemness genes.
(A) The bar plot illustrates the expression levels of SLC2A2, SOX2, and POU5F1 genes in HCC samples at different stages (Stages I, II, and III). (B) The scatter plot depicts the correlation between the expression levels of the SLC2A2, stemness (SOX2 and POU5F1), and (C) oncofetal genes (AFP, SALL4, FOXM1, and IGF2BP1). Each data point represents an individual sample, and the correlation coefficients and p-values are indicated.
Fig 3.
SLC2A2 is associated with IGF1R pathways differentiation.
(A) SLC2A2 expression levels were determined using quantitative real-time polymerase chain reaction (qPCR) in or GLUT2 overexpression cells compared with control cells (n = 4). (B) GLUT2 protein levels were measured by western blotting (n = 3). (C) We confirmed whether the difference in SLC2A2 expression level was related to stemness (n = 3) or (D) the IGF1R pathways (n = 3) via qPCR. * p <0.05, ** p <0.01, and *** p <0.001.
Fig 4.
SLC2A2 is essential for liver differentiation in developing vertebrates
(A) RT‐PCR analysis of SLC2A2 and β‐actin using 4 dpf zebrafish embryos. Total RNA was isolated from uninjected control and SLC2A2 MO‐injected embryos (2.5, 5, and 10 ng). (B) Lateral view of zebrafish embryos after SLC2A2 MO-injection (2.5, 5, and 10 ng). The black arrow indicates heart edema in zebrafish embryos injected with 10 ng of SLC2A2-MO. (C) WISH images of uninjected embryos and SLC2A2-targeting morpholino-injected embryos at 5 dpf using fabp10a. The yellow line indicates fabp10a signal at the liver. (D) Using hepatoblast marker hhex, we captured WISH images of uninjected embryos and SLC2A2-targeting morpholino-injected embryos at 2 dpf. The white arrow indicates the hhex signal in the hepatoblast. (E) qRT-PCR analysis of fabp10a and hhex expression in uninjected embryos and SLC2A2-targeting morpholino-injected embryos at 4 dpf. mRNA expression is normalized to that of β-actin mRNA levels (*** indicates significance at p-value <0.001). Scale bars indicates 200 um. (F) Quantitative RT-PCR was used to measure hhex expression, normalized to β-actin mRNA levels. There was no significant difference (n.s.) observed in hhex expression between control and SLC2A2 MO-injected embryos. Data are represented as mean ± standard error of the mean (SEM). (G) Quantitative RT-PCR analysis was performed to evaluate the expression levels of igf1r. The mRNA expression levels were normalized to β-actin. A significant increase (p-value < 0.001, ***) in igf1r expression was observed in SLC2A2 MO-injected embryos compared to the control group. Data are presented as mean ± standard error of the mean (SEM). (G) Reduction in liver fluorescence intensity in zebrafish embryos following slc2a2 morpholino (MO) injection. Representative fluorescence microscopy images and quantitative analysis of liver fluorescence intensity in zebrafish embryos at 72, 96, and 120 hours post-fertilization (hpf). Uninjected embryos show consistent fluorescence in the liver across all time points, while embryos injected with slc2a2 MO exhibit a progressive reduction in fluorescence intensity. The fluorescence signal in slc2a2 MO-injected embryos decreased by approximately 60% at 96 hpf and 80% at 120 hpf compared to the uninjected controls.
Fig 5.
The IGF1R pathway is upregulated following slc2a2 knockdown in HepG2 cells and zebrafish embryos.
(A) qPCR analysis of IGF1R and IGF2 gene expression in HepG2 cells. Knockdown of slc2a2 using shGLUT2 significantly increased IGF1R and IGF2 expression compared to the pLKO control group (p < 0.05). Overexpression of slc2a2 (GLUT2 over) significantly decreased IGF1R and IGF2 expression compared to the pMSCV control group (p < 0.01 for IGF1R, p < 0.05 for IGF2). (B) qPCR analysis of igf1r mRNA expression in zebrafish embryos at 2 dpf. slc2a2 knockdown significantly increased igf1r expression compared to the control group (p < 0.001). (C) Immunohistochemistry (IHC) staining for IGF1R protein in zebrafish embryos at 2 dpf. slc2a2-MO treated embryos showed stronger IGF1R fluorescence intensity compared to control embryos, indicating increased IGF1R protein expression. All analyzed embryos (11/11) showed consistent results in both groups.