Effect of IL-33 on the proliferation and migration of hepatocellular carcinoma cells by regulating NF-κB signaling pathway

Abstract

Abstract: Objective To investigate the effect of IL-33 on the regulation of NF-κB signaling pathway and inflammatory factors on the proliferation and migration of HepG2 cells.Methods RT-PCR was used to detect the expression of IL-33 in different hepatocellular carcinoma cell lines (LO2, MHCC97H, LM3, HepG2, SMCC7721, Hep3B). Western Blot was used to assess the regulation of IL-33 on the activation of NF-κB signaling pathway-related proteins (NF-κB, p-NF-κB, IKB, p-IKB) in HepG2 cells. ELISA was used to measure the regulation of IL-33 on the release of inflammatory factors (IL-1α, IL-1β, IL-6, IL-18) in HepG2 cells. The EdU proliferation test was conducted to detect the effect of IL-33 on the proliferation of HepG2 cells. The CCK8 test was performed to assess the viability of HepG2 cells after 24 h and 48 h of treatment with IL-33. Transwell cell test and scratch test were conducted to examine the effect of IL-33 on HepG2 cell migration.Results The RT-PCR results showed a statistically significant difference in the relative expression of IL-33mRNA between LO2 (1.01 ± 0.01), MHCC97H (1.08 ± 0.05), LM3 (1.76 ± 0.21), HepG2 (3.88 ± 0.35), SMCC7721 (2.24 ± 0.43), and Hep3B (2.13 ± 0.30) (F=19.621, P=0.001). LO2 cells and MHCC97H cells had similar expression levels, while LM3, HepG2, SMCC7721, and Hep3B showed higher expression, with HepG2 cells having the highest expression. In the 24-hour group treated with IL-33, the levels of IL-1α (3185.25 ± 194.67), IL-1β(2103.69 ± 124.75), IL-6 (446.58 ± 36.82), and IL-18 (1492.60 ± 180.31) were higher than those in the control group (1001.58 ± 18.65, 1642.81 ± 45.66, 151.34 ± 20.13, 915.48 ± 36.15), with statistical significance (P<0.05).The relative expression levels of phosphorylated NF-κB protein (4.53 ± 0.36) and phosphorylated IKB protein (2.76 ± 0.28) were higher in the 24-hour group treated with IL-33 compared to the control group (1.25 ± 0.23, 0.49 ± 0.09), with statistical significance (P<0.05). The levels of IL-1α, IL-1β, IL-6 and IL-18 factors were higher in the 24-hour group treated with IL-33 than the control group, with a statistically significant difference (P<0.05). The IL-33 treated group showed higher cell proliferation (1.41 ± 0.08), 24-hour cell viability (135.69 ± 6.88)%, and 48-hour cell viability (176.34 ± 25.69)% compared to the control group's cell proliferation (0.86 ± 0.04), and cell viability at 24 hours (98.65 ± 1.05)%, and 48 hours (119.89 ± 10.86)%, with statistically significant differences (t=13.750, P=0.000; t=9.218, P=0.000; t=3.506, P=0.017). The results of the Transwell chamber and scratch experiments indicated that the number of cell invasions (256.38 ± 10.11) and migration relative distance (0.74 ± 0.05) in the IL-33 treated group for 24 hours were higher than those in the control group (185.63 ± 23.54 and 0.45 ± 0.12, respectively), with statistically significant differences (t=4.783, P=0.005; t=3.864, P=0.012).Conclusion IL-33 can affect the inflammatory microenvironment of hepatoma cells by regulating the NF-κB signaling pathway and promote the proliferation and migration of HepG2 cells.

Key words: Hepatocellular carcinoma, IL-33, NF-κB signaling pathway, Inflammation, Proliferation, Migration

ZHAO Ying, TIAN Ming. Effect of IL-33 on the proliferation and migration of hepatocellular carcinoma cells by regulating NF-κB signaling pathway[J]. Chinese Hepatolgy, 2023, 28(11): 1331-1334.

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