The endoplasmic reticulum stress response is involved in apoptosis induced by aloe-emodin in HK-2 cells.
Aloe-emodin (AE; 1,8-dihydroxy-3-hydroxymethyl-9,10-anthracenedione) is one of the primary active compounds in total rhubarb anthraquinones (TRAs), which induce nephrotoxicity in rats. However, it is still not known whether AE has a similar effect on human kidney cells. In this study, 3-(4,5,-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays showed that AE decreases the viability of HK-2 cells (a human proximal tubular epithelial cell line) in a dose- and time-dependent manner. AE induced G2/M arrest of cell cycle in HK-2 cells, which was detected with propidium iodide (PI) staining. This apoptosis was further investigated by Hoechst staining, transmission electron microscopy (TEM), DNA fragmentation, and Annexin V/PI staining. Apoptosis of the cells was associated with caspase 3 activation, which was detected by Western blot analysis and a caspase activity assay. In addition, changes in the endoplasmic reticulum (ER) ultrastructure as observed by TEM showed the effects of AE on ER. Treatment with AE also resulted in an increase in eukaryotic initiation factor-2α (eIF-2α) phosphorylation, X-box binding protein 1 (XBP1) mRNA splicing, c-Jun N-terminal kinase (JNK) phosphorylation, glucose-regulated protein (GRP) 78 and CAAT/enhancer-binding protein-homologous protein (CHOP) accumulation. These results suggest that AE induces ER stress in HK-2 cells, which is involved in AE-induced apoptosis. In conclusion, AE induces apoptosis in HK-2 cells, and the ER stress is involved in AE-induced apoptosis in vitro. The implications of the toxic effects of AE for clinical use are unclear and these findings should be taken into account in the risk assessment for human exposure.
School of Pharmaceutical Sciences, Center of Laboratory Animals, Sun Yat-sen University, Guangzhou 510006, PR China.
Shaohua
S
Zhu
Jing
J
Jin
Yan
Y
Wang
Zizhang
Z
Ouyang
Chen
C
Xi
Jia
J
Li
Yuwen
Y
Qiu
Jinzhi
J
Wan
Min
M
Huang
Zhiying
Z
Huang
D000880:Anthraquinones / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D001483:Base Sequence; D015153:Blotting, Western; D002453:Cell Cycle / Q000187:drug effects; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D017931:DNA Primers; D004268:DNA-Binding Proteins / Q000235:genetics; D004721:Endoplasmic Reticulum / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D005455:Fluorescent Antibody Technique; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D046529:Microscopy, Electron, Transmission; D012326:RNA Splicing; D012333:RNA, Messenger / Q000235:genetics; D000071701:Regulatory Factor X Transcription Factors; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors / Q000235:genetics; D000071717:X-Box Binding Protein 1
D000880:Anthraquinones; D017931:DNA Primers; D004268:DNA-Binding Proteins; D012333:RNA, Messenger; D000071701:Regulatory Factor X Transcription Factors; D014157:Transcription Factors; D000071717:X-Box Binding Protein 1; C000605543:XBP1 protein, human; C000605545:Xbp1 protein, rat; C518327:aloe emodin