Objective To investigate the relationship between celastrol inhibition against multiple myeloma cell growth and unfolded protein response (UPR) and the related molecular mechanism, so as to provide new drug targets for multiple myeloma treatment. Methods Four multiple myeloma cell lines RPMI 8226, U266, SKO and KMS-11 were treated with different concentrations (proliferation:0.0-10.0 μmol/L; apoptosis:0.0-4.0 μmol/L; cell cycle:0.0-1.5 μmol/L) of celastrol for different periods (proliferation:1-3 d; apoptosis:1 d; cell cycle:1 d), and cell proliferation, apoptosis and cell cycle were examined. Western blotting analysis was used to detect the main molecules in the inositol-requiring enzyme 1 (IRE1), PRKR-like endoplasmic reticulum kinase (PERK), and activating transcription factor 6 (ATF6) signaling pathways of UPR, which included glucose-regulated protein 78 (GRP78), ATF6, PERK, eukaryotic initiation factor 2α (eIF2α), phosphorylated-eIF2α (p-eIF2α), C/EBP homologous protein (CHOP), IRE1 and phosphorylated-IRE1 (p-IRE1). After the lentivirus vector containing short hairpin RNA (shRNA) was used to interfere with eIF2α expression in RPMI 8226 cells, the effects of celastrol were detected on signaling molecule expression, apoptosis and cell cycle. Results Celastrol inhibited the proliferation of 4 multiple myeloma cells, induced apoptosis, and arrested the cell cycle at G₀/G₁ phase in dose- and time-dependent manners. RPMI 8226 cells were most sensitive to celastrol. For RPMI 8226 cells, when 0.5-2.0 μmol/L of celastrol was used for 30 min to 24 h, the p-eIF2α levels in the PERK signaling pathway of UPR were significantly increased (P<0.05 or P<0.01), and the downstream CHOP expression was risen correspondingly (P<0.05 or P<0.01), but the other two pathways ATF6 and IRE1 were not affected obviously. After interference of eIF2α expression with lentivirus vector containing shRNA, the effects of celastrol to increase CHOP expression, induce apoptosis and arrest cell cycle were significantly attenuated or disappeared. Conclusion Celastrol can inhibit the growth of a variety of multiple myeloma cells, and the activated eIF2α molecule in the UPR PERK signaling pathway may be one of the mechanisms.