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Rhein is an active component of rhubarb; a traditional Chinese medicine reported to induce apoptosis and cause liver toxicity. However, rhein's apoptotic-inducing effects, as well as its molecular mechanisms of action on hepatic cells need to be further explored. In the present study, rhein was found to trigger apoptosis in primary human hepatic HL-7702 cells as showed by annexin V/PI double staining assay and nuclear morphological changes demonstrated by Hoechst 33258 staining. Moreover, it was observed that the mechanism implicated in rhein-induced apoptosis was caspase-dependent, presumably via ER-stress associated pathways, as illustrated by up-regulation of glucose-regulated protein 78 (GRP 78), PKR-like ER kinase (PERK), C-Jun N-terminal kinase (JNK) and CCAAT/enhancer-binding protein homologous protein (CHOP). Meanwhile, caspase-4 as a hallmark of ER-stress, was also showed to be activated following by caspase-3 activation. Furthermore, rhein also promoted intracellular elevation of calcium that contributed in apoptosis induction. Interestingly, pre-treatment with calpain inhibitor I reduced the effects of rhein on apoptosis induction and JNK activation. These data suggested that rhein-induced apoptosis through ER-stress and elevated intracellular calcium level in HL-7702 cells.

Apoptosis can result from activation of three major pathways: the extrinsic, the intrinsic, and the most recently identified endoplasmic reticulum (ER) stress-mediated pathway. While the two former pathways are known to be operational in human polymorphonuclear neutrophils (PMNs), the existence of the ER stress-mediated pathway, generally involving caspase-4, has never been reported in these cells. Recently, we have documented that arsenic trioxide (ATO) induced apoptosis in human PMNs by a mechanism that needs to be further investigated. In this study, using immunofluorescence and electron microscopy, we present evidence of ER alterations in PMNs activated by the ER stress inducer arsenic trioxide (ATO). Several key players of the unfolded protein response, including GRP78, GADD153, ATF6, XBP1 and eIF2alpha are expressed and activated in PMNs treated with ATO or other ER stress inducers. Although caspase-4 is expressed and activated in neutrophils, treatment with a caspase-4 inhibitor did not attenuate the pro-apoptotic effect of ATO at a concentration that reverses caspase-4 processing and activation. Our results demonstrate for the first time that the ER stress-mediated apoptotic pathway operates in human neutrophils.

A subgroup of caspase family of inflammatory caspases (-1, -4, -5, -11, and -12) play important role during cytokine maturation and inflammation but their regulation is not well understood as much as the initiator and effector caspases. Here, the biochemical mechanism of caspase-4 activation is elucidated. With citrate, a well-known kosmotrope to enhance the monomer-dimer transition, caspase-4 was activated approximately 40 times that was comparable with that of caspase-9 ( approximately 75-fold increments). The activation reaction was mainly bimolecular (n=1.67+/-0.04) for monomeric caspase-4. In addition, the interdomain cleavage was also responsible to activate caspase-4 more than 100-fold, again comparable with that of effector caspases where the proteolytic processing is considered as the sole activation mechanism. Thus, caspase-4 shows a novel activation mechanism of the synergism between dimerization and proteolysis that sharply differs from the established activation mechanism of dimerization for initiators and interdomain cleavage for effector caspases.

Due to chemotherapy, the majority of breast cancer patients survive, but frequently complain of chemotherapy-associated cognitive impairment. This phenomenon is termed "chemobrain" or "chemofog" in the literature. However, its mechanisms are unclear. The objective of this study was to investigate the mechanisms of paclitaxel (Px)-induced neurotoxicity, with a focus on endoplasmic reticulum (ER) stress. To investigate Px-induced neurotoxicity and ER stress induction, SK-N-SH cells were treated with 1, 10, 50, and 100 μM Px for 24 h. Neurotoxicity was assessed using MTS viability assays, and ER stress was assessed by evaluating the expression of phosphorylated elF2α (phospho-eIF2α), C/EBP homologous protein (CHOP), and cleaved caspase 4 and caspase 3 (the active form of each caspase). Furthermore, to investigate whether immunoglobulin heavy-chain binding protein (BiP) inducer X (BIX), which induces the molecular chaperone BiP, could attenuate Px-induced neurotoxicity, SK-N-SH cells were pre-treated for 12 h with 3.5 μM BIX before Px treatment. Neurotoxicity was observed in SK-N-SH cells treated with Px in a dose-dependent manner compared with vehicle control. Furthermore, phospho-eIF2α, CHOP, and activated caspase 4 and caspase 3 were significantly induced in Px-treated cells. In addition, pre-treatment with BIX significantly attenuated the induction of CHOP and activated caspase 4 and caspase 3. The viability of BIX pre-treated cells prior to Px treatment was significantly increased compared with cells that were not treated with BIX. Our results suggest that Px induces neurotoxicity in part through activating the ER stress response. Our findings should contribute to novel approaches regarding the mechanism of Px-induced neurotoxicity, including chemobrain.