Biochemical and Biophysical Research Communications
Augmenter of liver regeneration (ALR) protects human hepatocytes against apoptosis
Research highlights
► ALR decreases cytochrome c release from mitochondria. ► ALR protects hepatocytes against apoptosis induction by ethanol, TRAIL, anti-Apo, TGF-β and actinomycin D. ► ALR exerts a liver-specific anti-apoptotic effect. ► A possible medical usage of ALR regarding protection of liver cells during apoptosis inducing therapies.
Introduction
The liver has a regenerative potential, which permits recovery from functional disorders induced by hepatic injury [1]. During liver regeneration, regenerative factors released from parenchymal and non-parenchymal liver cells stimulate the proliferation of hepatocytes by induction of immediate early genes and stimulating the release of growth factors [2].
Besides well-known growth factors like hepatocyte growth factor (HGF) or epidermal growth factor (EGF), augmenter of liver regeneration (ALR) is another cytokine of vital importance. For both EGF and HGF, mitogenic effects on not only hepatic cells but also trophoblasts and myoblasts, respectively have been shown [3], [4]. ALR belongs to a novel group of so called cytozymes as it acts as a growth factor and a sulfhydryl oxidase enzyme, that binds FAD containing a redox-active CxxC disulfide proximal to a flavin ring [5]. ALR is known to support liver regeneration in experimental animals [6], [7]. Previous studies have shown that ALR activates the ras/Mek/Erk as well as the PI3K/Akt pathways [8]. The influence of ALR on signaling pathways differs from that of other growth factors, such as EGF. For instance, ALR causes a transient and EGF a permanent increase in ERK phosphorylation [8]. Although an activation of the PI3K/Akt signaling pathway has recently been shown, an anti-apoptotic effect of ALR on human hepatocytes has not been studied yet. This possible new feature of ALR was tested by examining ALR’s protective effects against various apoptotic inducers. Particularly in the field of cancer treatment, either of primary liver cancer or liver metastases, a therapy that protects the liver against apoptosis would be highly welcome [9].
For HGF, as one of the growth factors influencing the liver metabolism, anti-apoptotic and [10], [11], [12], [13] protective effects [14], [15] have already been found. However, cytokines or growth factors, such as EGF, HGF or IL-6 are problematic in this context, because they influence a diversity of organ systems [8]. For example, EGF activated ERK and stimulated proliferation not only in liver but also in cell lines of colonic, bronchial, pancreatic and gastric origin [8]. In contrast, ALR induced proliferation only in liver cell systems. However, to our knowledge the cell type specific anti-apoptotic effects of ALR have not yet been studied. Therefore the aim of this study was to investigate the promising characteristics of ALR regarding a possible liver-specific anti-apoptotic effect in vitro.
The aim of this study was to show a protection of hepatic cells by ALR administration from apoptosis induced by ethanol, tumor necrosis factor related apoptosis-inducing ligand (TRAIL), anti-Apo, transforming growth factor β (TGF-β) and actinomycin D (Act D) and that this protection occurs in primary human hepatocytes but not in pancreatic, colonic, bronchial and gastric cell lines. This would imply a possible medical usage of ALR regarding protection of liver cells during apoptosis inducing therapies.
Section snippets
Reagents
Recombinant human ALR (rhALR) was prepared as described previously [16]. Briefly, fractions containing rhALR protein were combined and dialyzed against dialysis buffer (25 mM Hepes, 0.1% Tween 20, and 1 mM EDTA, pH 8.2) at 4 °C, with three buffer changes. Afterwards, rhALR protein was concentrated using a 5 kDa cut-off ultrafree-15 centrifugal filter device (Millipore GmbH, Schwalbach, Germany) [17]. All antibodies used were purchased from Cell Signaling Technology (Beverly, MA, USA). TRAIL and
ALR decreases cytochrome c release from mitochondria
As an early sign of apoptosis, administration of ethanol resulted in an increase in cytochrome c release into the cytosol in primary human hepatocytes (Fig. 1). When treated with ethanol, cells cultured in the presence of rhALR exhibited a clear decrease in cytosolic cytochrome c compared to the cells incubated with ethanol alone (Fig. 1). The influence of ALR on the release of cytochrome c from mitochondria into the cytosol suggests that the anti-apoptotic effect of ALR is at least partially
Discussion
Therapeutic options stimulating anti-apoptotic mechanisms in hepatocytes would be highly welcome in several clinical settings, e.g. non-alcoholic steatohepatitis (NASH), alcohol mediated hepatitis and cholestatic liver diseases, to retard fibrotic progression and potentially prevent cirrhosis [23]. As hepatocyte cell death promotes hepatic fibrosis, therapeutic hepatoprotective strategies focusing on protection of hepatocytes and prevention of hepatocytic apoptosis are important. However,
Acknowledgments
We thank C. Putz for excellent technical assistance and Dr. S.M.L. Lee for great support in finishing this manuscript. We acknowledge the Human Tissue and Cell Research (HTCR) Foundation for supporting our research by making human liver tissue available as well as hepacult GmbH for supplying cell isolation and cultivation technology. This study was supported in part by a grant from the BMBF (Virtual Liver, 0315759) and a grant from the Bohnewand Foundation.
References (32)
- et al.
EGF stimulates proliferation in the bovine placental trophoblast cell line F3 via Ras and MAPK
Placenta
(2010) - et al.
Hepatocyte growth factor (HGF) signals through SHP2 to regulate primary mouse myoblast proliferation
Exp. Cell Res.
(2009) - et al.
Augmenter of liver regeneration causes different kinetics of ERK1/2 and Akt/PKB phosphorylation than EGF and induces hepatocyte proliferation in an EGF receptor independent and liver specific manner
Biochem. Biophys. Res. Commun.
(2010) - et al.
Hepatocyte growth factor suppresses tumor cell apoptosis in nasopharyngeal carcinoma by upregulating Bcl-2 protein expression
Pathol. Res. Pract.
(2009) - et al.
Anti-apoptotic effect of hepatocyte growth factor from actinomycin D in hepatocyte-derived HL7702 cells is associated with activation of PI3K/Akt signaling
Toxicol. Lett.
(2006) - et al.
Effects of hepatocyte growth factor on glutathione synthesis, growth, and apoptosis is cell density-dependent
Exp. Cell Res.
(2008) - et al.
Hepatocyte growth factor protects hepatocytes against oxidative injury induced by ethanol metabolism
Free Radic. Biol. Med.
(2009) - et al.
Hepatocyte growth factor has protective effects on crystal–cell interaction and crystal deposits
Urology
(2006) - et al.
Regulation of polyamine synthesis in human hepatocytes by hepatotrophic factor augmenter of liver regeneration
Biochem. Biophys. Res. Commun.
(2006) - et al.
Cellular damage to human hepatocytes through repeated application of 5-aminolevulinic acid
J. Hepatol.
(2003)
Characterization of immune responses in gastric cancer patients A possible impact of H. pylori to polarize a tumor-specific type 1 response?
Clin. Immunol.
A rapid and simple method for measuring thymocyte apoptosis by propidium iodide staining and flow cytometry
J. Immunol. Methods
Analysis of CD95 threshold signaling: triggering of CD95 (FAS/APO-1) at low concentrations primarily results in survival signaling
J. Biol. Chem.
Augmenter of liver regeneration: an important intracellular survival factor for hepatocytes
J. Hepatol.
Human augmenter of liver regeneration is important for hepatoma cell viability and resistance to radiation-induced oxidative stress
Free Radic. Biol. Med.
Liver regeneration is impaired in lipodystrophic fatty liver dystrophy mice
Hepatology
Cited by (46)
TMPRSS11D/ALR-mediated ER stress regulates the function of myeloid-derived suppressor cells in the cervical cancer microenvironment
2023, International ImmunopharmacologyAugmenter of liver regeneration: Mitochondrial function and steatohepatitis
2022, Journal of HepatologyDeceleration of Liver Regeneration by Knockdown of Heme Oxygenase-1 is Associated With Impairment of Liver Injury Recovery After Reduced-Size Liver Transplantation in Rats
2020, Transplantation ProceedingsCitation Excerpt :In agreement with them, the ALT and AST activity, a hallmark of hepatocellular damage, was also inhibited by SnPP pretreatment, were paralleled to HO-1 mRNA expression along with accelerated liver proliferation after HSLT, and this phenomenon was supported by histologic outcomes. Augmenter of liver regeneration significantly reduces apoptosis induced by ethanol, TGF-β, and actinomycin D [17], and HO-1 participates in protection of the organs by inhibiting cell apoptosis response [5]. So, we next examined whether HO-1 ameliorates hepatocyte apoptosis in HO-1 induced liver protection.
Augmenter of liver regeneration: Essential for growth and beyond
2019, Cytokine and Growth Factor ReviewsCitation Excerpt :It was shown that rALR reduces caspase 3/7 activity after free fatty acid- [64] and bile acid-induced injury (unpublished data), which may be due to reduced expression of Bax [64]. Furthermore, rALR reduces EtOH- and death ligand-induced apoptosis by reducing cytochrome C release [145]. This was further confirmed in a study showing that rALR reduces cell damage after partial hepatectomy by inducing the expression of anti-oxidative clusterin and anti-apoptotic Bcl2 and suggested that rALR boosts liver regeneration through apoptosis attenuation rather that inducing proliferation [66].
- 1
Both authors contributed equally to this study.