Regulation of liver hepcidin expression by alcohol in vivo does not involve Kupffer cell activation or TNF-α signaling

Abstract

Alcohol downregulates hepcidin expression in the liver leading to an increase in intestinal iron transport and liver iron storage. We have previously demonstrated that alcoholmediated oxidative stress is involved in the inhibition of hepcidin transcription by alcohol in vivo. Kupffer cells and TNF-␣ play a key role in alcohol-induced liver injury. The aim of this study was to define their involvement in the regulation of hepcidin expression by alcohol. Kupffer cells were inactivated or depleted by employing gadolinium chloride and liposomes containing clodronate, respectively. Rats pair fed with the alcohol-Lieber-DeCarli diet for 6 wk and mice fed with 20% ethanol in the drinking water for 1 wk were used as experimental models. Interestingly, alcohol downregulated hepcidin expression in the livers of rats and mice independent of gadolinium chloride or clodronate treatment. One week of alcohol treatment was sufficient to induce a significant increase in TNF-␣ levels and phosphorylation of NF-B subunit p65. The neutralization of TNF-␣ by specific antibodies inhibited p65 phosphorylation. However, neither the neutralization of TNF-␣ nor the lack of TNF-␣ receptor expression reversed alcohol-induced suppression of liver hepcidin expression. The level of alcohol-induced ROS in the liver was also undiminished following Kupffer cell inactivation or depletion. Our results demonstrate that alcohol-induced Kupffer cell activation and TNF-␣ signaling are not involved in the suppression of liver hepcidin expression by alcohol-mediated oxidative stress in vivo. Therefore, these findings suggest that alcohol acts within hepatocytes to suppress hepcidin expression and thereby influences iron homeostasis. alcoholic liver disease; iron; liver injury; oxidative stress HEPCIDIN, A CIRCULATORY PEPTIDE synthesized by the liver, plays a central role in the regulation of iron metabolism by controlling the iron uptake from the intestine and iron release from macrophages (16,18,20). Both acute and chronic alcohol exposure downregulate hepcidin expression in the liver (3,5,(7)(8)(9)21). This results in an increase in the expression of iron transporters in the duodenum and the iron storage protein ferritin in the liver (8, 9).The activation of Kupffer cells and inflammatory responses prime the liver for alcoholic liver disease (ALD) (1,23,25). The proinflammatory cytokine TNF-␣, mainly synthesized by Kupffer cells, plays a critical role in ethanol-induced liver injury (10,14,30). TNF-␣ has been reported to downregulate hepcidin expression in human hepatoma cells (17). However, LPS and the cytokines IL-1 and IL-6 induce liver hepcidin expression (11,17). Lou et al. (13) Besides cytokines, Kupffer cells activated following alcohol exposure also release reactive oxygen species (2,23,28). We have demonstrated that alcohol-mediated oxidative stress suppresses liver hepcidin expression and promoter activity by inhibiting the DNA-binding activity of the transcription factor, CCAAT/enhancer-binding protein ␣ (C/EBP ␣) in vivo (9). We investigat...

Is the iron regulatory hormone hepcidin a risk factor for alcoholic liver disease?

“…The inactivation of Kupffer cells by gadolinium chloride in rats pairfed with alcohol Lieber DeCarli diets for 6 wk or mice fed with ethanol in the drinking water for 1 wk did not reverse the alcohol-induced suppression of hepcidin expression in the liver [100] . Moreover, similar results were obtained when Kupffer cells were depleted by liposomes containing clodronate [100] . When phagocytosed by the Kupffer cells, clodronate released from the liposomes induces apoptosis and thereby depletes the Kupffer cell [101] .…”

“…When co-cultured, Kupffer cells have been suggested to exert a negative effect on hepcidin synthesis in hepatocytes [99] . However, the depletion or inactivation of the Kupffer cells has been reported to not induce any significant changes in basal hepcidin expression levels in the livers of both control and alcohol-treated animals in vivo [100] . Interestingly, one week of ethanol treatment (in the drinking water) was sufficient to induce NF-κB activation and TNF-α and IL-6 release in mice, compared to control mice fed with plain water [100] .…”

“…Interestingly, one week of ethanol treatment (in the drinking water) was sufficient to induce NF-κB activation and TNF-α and IL-6 release in mice, compared to control mice fed with plain water [100] . The neutralization of TNF-α inhibited the activation of NF-κB [100] . However, neither the neutralization of TNF-α nor the absence of TNF-α receptorⅠand Ⅱ expression altered the effect of alcohol on hepcidin expression (i.e.…”

“…The role of Kupffer cell activation and TNF-α signaling in the regulation of hepcidin expression by alcohol in vivo has also been reported [100] . The inactivation of Kupffer cells by gadolinium chloride in rats pairfed with alcohol Lieber DeCarli diets for 6 wk or mice fed with ethanol in the drinking water for 1 wk did not reverse the alcohol-induced suppression of hepcidin expression in the liver [100] .…”

The Hypoxia-Inducible Factor–C/EBPα Axis Controls Ethanol-Mediated Hepcidin Repression

“…Conversely, hepcidin deficiency leads to increased iron absorption and mobilization of iron stores, which can cause iron overload (6,29). Previous publications have shown that in rodents and humans, hepcidin is downregulated in response to ethanol treatment (5,11,14,16,17,19,20,31). Moreover, it has been shown that the increase in liver iron following alcohol consumption in ALD patients is directly due to low hepcidin expression (5,11,14,16,17,19,20,31).…”