Despite the accumulating evidence supporting an apoptotic role of caspase-4, negative results have been reported in tunicamycin- and thapsigargin-induced apoptosis in human multiple myeloma cell lines,
37 homoharringtonine-induced apoptosis in MUTZ-1 cells,
38 and Z α-1 antitrypsin-induced apoptosis in HEK293 cells.
39 Although apoptosis usually does not require de novo synthesis of caspases, ER stress may increase mRNA expression of certain caspases, including caspase-4.
40 Our study demonstrated that proinflammatory stimuli (IL-1β, TNF-α, LPS, IFN-γ, and monocyte coculture) induced both mRNA synthesis and activation of caspase-4 in hRPE cells. IL-1β–induced IL-8 protein production was reduced by the caspase-4 inhibitor Z-LEVD-fmk, suggesting that caspase-4 is involved in the proinflammatory responses of hRPE cells. These results are consistent with a previous study in which LPS-induced IL-8 mRNA synthesis and protein production are reduced by caspase-4 knockdown in human THP1 monocytic cell lines.
13 On the other hand, the ER-stress inducer tunicamycin induced expression of GRP78, a specific marker of ER stress. Although tunicamycin increased the expression of proapoptotic Bax, this study also found increased expression of the antiapoptotic Bcl-2 protein, resulting in an increased Bcl-2/Bax ratio. In most cases, the ER stress-induced apoptotic response is regulated by or dependent on the Bcl-2 family of proteins.
41 The Bcl-2/Bax ratio has been clinically used as an indicator of the susceptibility of tumor cells to the induction of apoptosis by chemotherapeutic agents.
42 However, the involvement of Bcl-2 and Bax in tunicamycin-induced apoptosis has not been well characterized. Increased Bcl-2 protein expression by tunicamycin has been reported in one previous report.
43 The increased Bcl-2 expression shown in our study, however, might not have affected caspase-4 activation because a previous study has shown that activation of caspase-4 by tunicamycin is only slightly affected by overexpression of Bcl-2.
9 The increased Bcl-2/Bax ratio by tunicamycin may suggest an early protective response of hRPE cells to apoptotic stimuli by enhanced expression of the antiapoptotic protein Bcl-2 to counteract the increase in proapoptotic protein Bax.