This study reports on the protective effects of an activated form of Stat3 against Fas-mediated liver injury and the underlying mechanisms of these effects (Figure ). One of the major protective mechanisms against Fas-induced apoptosis in mouse liver seems to be a caspase-dependent mechanism. Stat3-C suppressed hepatic apoptosis through inhibition of both caspase-8 and -3 activities via upregulation of FLIP protein and through upregulation of Bcl-2 and Bcl-X
L proteins. The protective effect of Stat3 against apoptosis has already been explained partly by upregulation of antiapoptotic proteins such as Bcl-X
L and Survivin (13
). IL-6, one of the major ligands for Stat3, is also reported to suppress liver injury of various causes (19
). Adenovirally overexpressed “hyper IL-6,” a fusion protein of IL-6 and its soluble receptor, reversed fulminant liver failure induced by D
-galactosamine and also accelerated liver regeneration (35
). IL-6 also inhibits Fas-mediated liver injury through upregulation of Bcl-2, Bcl-X
L, and FLIP proteins (19
). These reports may support our present data.
Figure 7 Schematic view of the protective mechanisms of Stat3 against Fas-mediated liver injury. Activation of Stat3 upregulates caspase-related proteins (such as FLIP, Bcl-XL, and Bcl-2) and redox-related protein Ref-1. These proteins collaboratively prevent (more ...)
There are accumulating data indicating that ROS are involved in Fas-mediated apoptosis in various cells (36
). In the present study, the antioxidant NAC also reduced hepatic generation of ROS and activity of caspase-3 and -9, and prevented the liver from injury induced by Fas agonist. Gulbins et al. have already reported the involvement of Ras-mediated ROS in Fas-mediated cell death (40
). Fas may activate caspase activities at least partially via ROS generation and inducing apoptosis, effects that are inhibited by NAC in this pathway.
Interestingly, Stat3 reduced the hepatic generation of ROS following Jo2 administration through upregulation of Ref-1. Ref-1 is primarily a nuclear protein with dual and mutually exclusive nuclear functions: it is an endonuclease in the base excision repair pathway (41
) and it is a reducing agent that facilitates the DNA-binding activities of many redox-sensitive transcription factors including NF-κB (26
). This is a bifunctional protein that is upregulated in response to a host of stimuli that result in intracellular ROS generation (45
). Adenovirally overexpressed Ref-1 in mouse liver successfully improves postischemic liver injury through suppression of ROS generation and apoptosis in hepatic tissue (26
). This may also explain its role in the Stat3-dependent protection against Fas-induced oxidative liver injury through Ref-1. Stat3-C did not induce other redox-related proteins such as thioredoxin and MnSOD in the present study, though MnSOD induced by Stat3 protects cardiomyocytes against hypoxia/reoxygenation-induced injury (46
). In the present study, activities of caspase-3 and caspase-8 were both reduced by Stat3-C. The underlying mechanisms of this effect may involve the direct upregulation by Stat3-C of caspase-associated antiapoptotic proteins such as FLIP, Bcl-2, Bcl-X
L, and also caspase-3 inactivation via upregulation of Ref-1.
In this way, Stat3 may play crucial roles in protection against Fas-mediated liver injury. However, very interestingly, hepatic ROS generation and apoptosis were almost the same in control and LS3-KO mice. There may also exist in the cell compensatory mechanisms other than those dependent on Stat3 that are not yet known.
PKB (Akt), a serine-threonine kinase, possesses mitogenic and antiapoptotic actions in endothelial cells when stimulated by HGF (47
). Also, we have recently reported that Akt phosphorylates and inactivates the small GTPase Rac1, which results in the reduction of oxidative stress and apoptosis induced by hypoxia/reoxygenation in hepatocytes (48
). Ping et al. have reported that cardiac-specific transgenic activation of PKC-ε confers cardioprotection to the ischemic heart (49
). These data suggest that some mitogenic signaling molecules may possess antioxidant and/or antiapoptotic properties, and may also support our present data.
Stat3 is one of the most important transcription factors and plays important roles in the initiation of liver regeneration. Though Stat3’s effects upon cell cycle–related genes have been extensively studied, antiapoptotic effects and mechanisms of Stat3 have not been well studied. This study is the first to report that Stat3 protects against Fas-mediated hepatic apoptosis by its direct effects on caspase activity with antioxidant effects through Ref-1.