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. 2023 Feb 28;324(5):G341–G353. doi: 10.1152/ajpgi.00163.2022

Figure 4.

Figure 4.

ATF4 activation mediates ER stress-triggered CD36 upregulation in hepatocytes. A: AML-12 cells were treated with tunicamycin (5 µM), a chemical ER stress inducer, for 16 h. PPARγ transactivation was determined by detecting its downstream target genes using real-time qPCR. Data are expressed as means ± SD, n = 4 separate experiments. Student’s t test was used for statistical evaluation (*P < 0.05; **P < 0.01 vs. corresponding control). B–E: AML-12 cells were transfected with either scramble siRNA (si-Ctrl) or Pparg siRNA (si-Pparg) for 24 h before tunicamycin treatment (5 µM) for 16 h. PPARγ expression (B and C), CD36 gene expression (D), and ATF4 expression (C and E) were determined by real-time qPCR and Western blotting, respectively. Data were expressed as means ± SD, n = 4 separate experiments. Student’s t test was used for statistical evaluation (**P < 0.01; ***P < 0.001). F and G: AML-12 cells were transfected with either scramble siRNA (si-Ctrl) or Atf4 siRNA (si-Atf4) for 24 h before tunicamycin treatment (5 µM) for 16 h. Gene expression of Atf4 (F) and CD36 (G) was determined by real-time qPCR and data were expressed as means ± SD, n = 3 separate experiments. Student’s t test was used for statistical evaluation (**P < 0.01; ***P < 0.001). H and I: male C57 BL/6 mice were intraperitoneally injected with either vehicle or tunicamycin (2 mg/kg body wt). The liver samples were harvested 16 h later. Hepatic CD36 expression was determined by real-time qPCR and Western blotting, respectively. Data were expressed as means ± SD, n = 3 mice. Differences between the two groups were determined using Student’s t test (**P < 0.01 vs. control group). J: total RNA was extracted from the liver tissues of control and hepatocyte-specific Atf4 knockout (LsATF4-KO) mice and subjected to real-time qPCR for CD36 gene expression. Data were expressed as means ± SD, n = 4 mice/group. Differences between the two groups were determined using Student’s t test (*P < 0.05 vs. control group). ATF4, activating transcription factor 4; CD36, cluster of differentiation 36; ER, endoplasmic reticulum; PPARγ, peroxisome proliferator-activated receptor γ.