Pannexin1 contributes to pathophysiological ATP release in lipoapoptosis induced by saturated free fatty acids in liver cells

Abstract

Hepatocyte lipoapoptosis induced by saturated free fatty acids (FFA) contributes to hepatic inflammation in lipotoxic liver injury, and the cellular mechanisms involved have not been defined. Recent studies have shown that apoptosis in nonhepatic cells stimulates ATP release via activation of pannexin1 (panx1), and extracellular ATP functions as a proinflammatory signal for recruitment and activation of the inflammatory cells. However, it is not known whether lipoapoptosis stimulates ATP release in liver cells. We found that lipoapoptosis induced by saturated FFA stimulated ATP release in liver cells that increased extracellular ATP concentration by more than fivefold above the values observed in healthy cells. This sustained pathophysiological ATP release was not dependent on caspase-3/7 activation. Inhibition of c-Jun NH(2)-terminal kinase (JNK), a key mediator of lipoapoptosis, with SP600125 blocked pathophysiological ATP release in a dose-dependent manner. RT-PCR analysis indicated that panx1 is expressed in hepatocytes and multiple liver cell lines. Notably, inhibition of panx1 expression with short hairpin (sh)RNA inhibited in part pathophysiological ATP release. Moreover, lipoapoptosis stimulated uptake of a membrane impermeable dye YoPro-1 (indicative of panx1 activation), which was inhibited by panx1 shRNA, probenecid, and mefloquine. These results suggest that panx1 contributes to pathophysiological ATP release in lipoapoptosis induced by saturated FFA. Thus panx1 may play an important role in hepatic inflammation by mediating an increase in extracellular ATP concentration in lipotoxic liver injury.

Nucleotides Released From Palmitate-Challenged Muscle Cells Through Pannexin-3 Attract Monocytes

“…The small dye YO-PRO-1 has a molecular weight (630 g/mol) close to that of ATP (507 g/mol), readily diffuses through open Cx/pannexin channels, and fluoresces upon binding to nucleic acids; hence, it has been used to ascertain the presence of open channels at the cell surface (15). YO-PRO was administered to myotubes treated with PA, PO, or BSA, along with a large Texas-Red-dextran (10 kDa) polysaccharide that cannot go through Cx/pannexin channels but enters cells with damaged membranes (Supplementary Fig.…”

“…Pannexins are recently discovered channel-forming proteins that allow the release of cytoplasmic molecules to the extracellular space (13,14). Whereas pannexin-1 and its role in the physiological release of small molecules have been widely studied (15)(16)(17), the functions of pannexin-2 and pannexin-3 remain elusive. Moreover, the contribution of pannexin channels and small molecule release during metabolic inflammation remains unexplored.…”

Evaluation of YO-PRO-1 as an early marker of apoptosis following radiofrequency ablation of colon cancer liver metastases

“…It is also possible that the transient activation of the plasma membrane can be a death trigger independent of proinflammatory caspase-1 and TLR signaling. Alternatively, liver cells may be highly sensitive to any manipulation and undergo early cellular changes (increased plasma membrane permeability) and will subsequently undergo cell necrosis possibly via different pathways (Emmett et al 2008;Xiao et al 2012). Small deviations from the norm cause fast responses, executed very efficiently by the hepatocytes.…”

“…This is most likely an indication of advanced injury and irreversible apoptosis due to the ablation treatment Xiao et al 2012). Coordinated activation of P2X receptors and Pannexin1 results in diverse arrays of membrane trafficking events, including opening of pores in the membrane, allowing large molecules, such as YP1 to enter cells (Iglesias et al 2008;Qu et al 2011;Xiao et al 2012). This observation may explain the discordance between cell death marker TUNEL and cell-permeability marker YP1 in the current study.…”

Purinergic signalling in the liver in health and disease

“…Hepatocyte lipoapoptosis induced by saturated free fatty acids contributes to hepatic inflammation in lipotoxic liver injury. It has been suggested that pannexin1 may play an important role in hepatic inflammation by mediating an increase in ATP release in lipotoxic liver injury [259]. APAP is often used to treat fever and pain, but it can cause damage to hepatocytes.…”