Interestingly, RA fibroblast-like synoviocytes (FLS) express death receptors; yet, they are relatively resistant to FasL, TNF, and tumor necrosis (TNF)-related apoptosis-inducing ligand (TRAIL)-induced apoptosis [
3‐
5]. This resistance has been related to high expression of anti-apoptotic molecules such as Fas-associated death domain-like IL1 beta-converting enzyme-inhibitory protein (FLIP) [
6,
7], sentrin-1 [
8,
9], Bcl-2 [
10], Mcl-1 [
11], and constitutive activation of Akt [
12‐
14].
Apoptosis is a process highly regulated and crucial in many physiological situations, and could involve two main pathways; the extrinsic, by activation of death receptors (Fas, TNF-RI), and the intrinsic or mitochondrial pathway. In the extrinsic pathway, FasL, TNF, and TRAIL ligation leads to recruitment of Fas-associated via death domain (FADD) and procaspase-8, which form the death inducing signaling complex (DISC), where caspase-8 is activated. In turn, caspase-8 activates caspase-3, which causes DNA fragmentation and cell death. The mitochondrial pathway is induced by hypoxia, cytotoxic drugs and growth factor deprivation leading to liberation of cytochrome c (cyt c) and Apaf-1-mediated activation of the caspase-9 [
15‐
17]. This pathway is tightly regulated by members of the Bcl-2 family with anti-apoptotic function, such as Bcl-2, Bcl-xL, Bcl-w, Mcl-1, and A1, which prevent mitochondrial membrane permeability and release of cyt c. In contrast, other Bcl-family members, such as Bax, Bak, Bok, BH3 interacting domain death agonist (Bid), Bad, Bim, and Puma, are pro-apoptotic and promote mitochondrial membrane permeability [
18]. In some cell types, named type II cells, the two apoptotic pathways are connected through the cleavage of Bid by activated caspase-8. Truncated Bid translocates to the mitochondria causing release of cyt c and cell death [
19]. In contrast, in type I cells, death-receptor induced apoptosis is independent of Bid [
19]. It seems that both the intrinsic and extrinsic apoptotic pathways are involved in arthritis development. There is much evidence implicating the extrinsic pathway [review in ref [
20] and [
21]]. However, support for the role of the intrinsic pathway is scant, although very convincing. For example, mice lacking Bim [
22] or Bid [
23] develop a severe synovial inflammation and bone destruction in an arthritis model. Also, evidence suggests that RA FLS are type II cells [
24]. Therefore, it is necessary to investigate the relevance of the intrinsic pathway and its connection with the extrinsic pathway in the FLS resistance to apoptosis.
RA FLS typically show Akt activation that could contribute to the relative resistance to apoptosis by unknown mechanisms. Akt/PKB is a Ser/Thr protein kinase implicated in inhibition of apoptosis and stimulation of cellular growth in several tissues by mechanisms including phosphorylation of the pro-apoptotic proteins Bad [
25] and Bax [
26], and suppression of pro-apoptotic proteins such as Bim and PUMA, through phosphorylation of the forkhead pathway [
27]; favouring the anti-apoptotic effect of Mdm2 on p53 [
28]; and inhibition of cleavage of Bid protein [
29,
30].
The aim of this study was to investigate the connection of the death receptor stimulation with the intrinsic pathway in the apoptosis of the type II cells RA FLS, and to analyse the possible relation between constitutively activated phosphoinositol-3 (PI3) kinase/Akt and the mechanisms of resistance to Fas-mediated apoptosis.