Activation of NADPH oxidase by transforming growth factor-β in hepatocytes mediates up-regulation of epidermal growth factor receptor ligands through a nuclear factor-κB-dependent mechanism

Abstract

The TGF-beta (transforming growth factor-beta) induces survival signals in foetal rat hepatocytes through transactivation of EGFR (epidermal growth factor receptor). The molecular mechanism is not completely understood, but both activation of the TACE (tumour necrosis factor alpha-converting enzyme)/ADAM17 (a disintegrin and metalloproteinase 17; one of the metalloproteases involved in shedding of the EGFR ligands) and up-regulation of TGF-alpha and HB-EGF (heparin-binding epidermal growth factor-like growth factor) appear to be involved. In the present study, we have analysed the molecular mechanisms that mediate up-regulation of the EGFR ligands by TGF-beta in foetal rat hepatocytes. The potential involvement of ROS (reactive oxygen species), an early signal induced by TGF-beta, and the existence of an amplification loop triggered by initial activation of the EGFR, have been studied. Results indicate that DPI (diphenyleneiodonium) and apocynin, two NOX (NADPH oxidase) inhibitors, and SB431542, an inhibitor of the TbetaR-I (TGF-beta receptor I), block up-regulation of EGFR ligands and Akt activation. Different members of the NOX family of genes are expressed in hepatocytes, included nox1, nox2 and nox4. TGF-beta up-regulates nox4 and increases the levels of Rac1 protein, a known regulator of both Nox1 and Nox2, in a TbetaR-I-dependent manner. TGF-beta mediates activation of the nuclear factor-kappaB pathway, which is inhibited by DPI and is required for up-regulation of TGF-alpha and HB-EGF. In contrast, EGFR activation is not required for TGF-beta-induced up-regulation of those ligands. Considering previous work that has established the role of ROS in apoptosis induced by TGF-beta in hepatocytes, the results of the present study indicate that ROS might mediate both pro- and anti-apoptotic signals in TGF-beta-treated cells.

The NADPH oxidase inhibitor VAS2870 impairs cell growth and enhances TGF-β-induced apoptosis of liver tumor cells

“…They were cloned by homology to the phagocytic NADPH oxidase gp91phox (NOX2) and have been described to function as signaling molecules in non-phagocytic cells [22]. In the liver, the isoforms NOX1, NOX2 and NOX4 are expressed by fetal and adult hepatocytes ( [5], Fig. 1C), although they might have opposite effects controlling cell death: NOX4 favoring apoptosis whilst NOX1 favors survival [6,8].…”

“…In fact, we have previously described that fetal rat hepatocytes show Nox1, Nox2, and Nox4 expression in basal conditions [5] showing apparent opposite roles in the control of liver cell death. Thus, NOX4 is necessary for the triggering of apoptosis induced by a physiological stimulus, such as the Transforming Growth Factor-beta (TGF-) [6] or antineoplastic drugs, such as doxorubicin [7].…”

“…Thus, NOX4 is necessary for the triggering of apoptosis induced by a physiological stimulus, such as the Transforming Growth Factor-beta (TGF-) [6] or antineoplastic drugs, such as doxorubicin [7]. On the contrary, NOX1 might be involved in protecting cells from TGF- proapoptotic signals in both fetal hepatocytes and hepatoma cells [5,8]. Importantly, we have recently reported an essential role for NOX1 in controlling autocrine growth through the Epidermal Growth Factor Receptor Page 4 of 27 A c c e p t e d M a n u s c r i p t 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 4 (EGFR), a mechanism which seems to be specific for liver tumor cells as compared to non tumoral hepatocytes [9].…”

Quinone compounds regulate the level of ROS production by the NADPH oxidase Nox4

“…Nox4 is unique among other Nox isoenzymes in that its activity is constitutive and may depend on a specific conformation of the dehydrogenase DH domain that should allow a spontaneous transfer of electrons from NADPH to FAD and to the hemes [24,25]. Data from various studies indicated clearly that Nox4 activity is regulated at the mRNA level, implying that an increase or decrease of ROS production by Nox4 is correlated to an up regulation [9,16,[26][27][28][29][30][31][32][33] or to a decline [34] of Nox4 transcripts. Although it has not been reported yet, post-translational regulation of Nox4 oxidase activity cannot be excluded.…”

High glucose activates PKC-ζ and NADPH oxidase through autocrine TGF-β1 signaling in mesangial cells

“…NADPH oxidase subunits include Nox1-5, p22 phox , p47 phox , p67 phox , and Rac (9). TGF-␤ 1 strongly enhances reactive oxygen species (ROS) production by NADPH oxidase in fibroblasts and hepatocytes (4,5,36). While the mechanisms are incompletely defined, this effect may be mediated in part through Smadinduced increases in the expression of Nox4 or other NADPH oxidase subunits (4).…”