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Roles and regulations of starvation-induced hepatic steatosis in zebrafish

par Pozo Morales, Macarena
Président du jury Remmelink, Myriam
Promoteur Singh, Sumeet Pal
Publication Non publié, 2023-05-22

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Résumé :

During starvation, animals utilize the liver as lipid storage organ. For instance, patients suffering from anorexia nervosa, overnight fasted mice and starved zebrafish larvae develop fatty liver. However, the mechanisms underlying starvation-induced liver damage remain unknown. Further, it remains unclear if the resolution of hepatic steatosis is beneficial to the liver and the animal. Here, we investigated the role and regulation of starvation-induced hepatic steatosis using zebrafish (Danio rerio) as model system. By in vivo imaging of calcium and lipid droplets, we published a work showing negative correlation between the two. Using a genetically encoded calcium chelator (SpiCee), we showed that buffering of calcium signaling reduces the turnover of lipid droplets. On the other hand, we increased calcium transients in the liver by mobilization of endo-lysosomal Ca2+ stores, using selective two-pore complex 2 (TPC2) agonists. This led to a resolution of hepatic lipids. Thus, we established methods to manipulate starvation-induced hepatic steatosis by modulating Ca2+ signaling. Using this system, we investigated the impact of hepatic lipid droplet accumulation, and their turnover on the health of the organ and organism. Increasing Ca2+ flux by pharmacological activation of TPC2 channels efficiently reduced fat in the zebrafish liver. However, this process enhanced macrophage infiltration, hepatocyte phagocytosis and starvation-induced mortality. On the other hand, buffering cytoplasmic Ca2+ reduced lipid droplet turnover during starvation, leading to reduced liver inflammation and increased the lifespan of starved animals. With this, we propose that fatty liver helps with starvation resistance, while hepatic lipid flux (turnover) damages the liver. To understand how animals with enhanced starvation resistance deal with hepatic steatosis and liver damage, we investigated the starvation response in cavefish (Astyanax mexicanus) in collaboration with Dr. Nicolas Rohner. In the cavefish model, we found that surprisingly, animals do not accumulate fat in the liver upon starvation, protecting liver from damage, in contrast to surface fish and zebrafish. Using transcriptome analysis, we identified the long-chain fatty acid transporter, slc27a2a as one of the genes responsible for starvation-induced steatosis. Zebrafish and surface fish, but not cavefish, upregulated the expression of slc27a2a in the liver upon starvation. When a pharmacological inhibition of SLC27A2, we observed a decrease in the fat accumulation within the zebrafish hepatocytes during starvation. Therefore, our study demonstrates the turnover of lipid droplets in hepatocytes during starvation, which is regulated by Ca2+ dynamics, is detrimental to the liver and the animals. Further, we identify that SLC27A2 regulates the development of starvation-induced steatosis.In addition, we used the tools developed here to investigate the response of the liver to acute ethanol exposure. The liver sustains the earliest and the greatest degree of tissue injury from excessive drinking because it is the primary site of ethanol metabolism. Acute alcohol uptake leads to accumulation of fat in the liver; however, chronic alcohol abuse leads to alcoholic liver disease (ALD), which is characterized by hepatic inflammation, hepatocyte injury and ballooning. Using in vivo calcium imaging, we show that the hepatocytes initiate calcium flux upon alcohol stress. Further, we find that dampening of calcium signaling leads to hepatocyte ballooning in the context of acute alcohol stress, thereby developing an accelerated model of alcoholic liver damage. Using RNA-Sequencing, we identify PIKfyve / TRPML1 as potential regulators of calcium flux upon alcohol stress. TRPML1 is an endo-lysosomal calcium channel that is activated by the PI(3,5)P2, the product of PIKfyve activity. Notably, pharmacological inhibition of PIKfyve leads to hepatocyte ballooning. In this ongoing investigation, we hypothesize endo-lysosomal calcium stores as an important and new modulators of alcoholic liver disease.