Study on the Relationship between Cisplatin-resistance of Human Ovarian Cancer Cell and TLR9 under Hypoxia / 中国药房

ABSTRACT

OBJECTIVE:To study the relationship between cisplatin-resistant of human ovarian cancer cells (SKOV3) and Toll-like receptor 9 (TLR9) under hypoxia. METHODS:Immunofluorescence method was used to detect the TLR9 expression in rupture and intact membranes SKOV3. SKOV3 was taken,adding cisplatin,then CCK-8 was used to detect cell inhibition rate af-ter 6,12,24 h of added into TLR9 specific agonists CpG-ODN 2006 (group A) and its isotype control CpG-ODN 2006 control (group B). SKOV3 or SKOV3 pretreated by 0(not added),1,10,102,103 μmol/L TLR9 specific antagonist chloroquine for 3 h were taken,adding cisplatin,cell inhibition rate was detected after 24 h of added into SKOV3 normoxia(21% O2),hypoxia(1%O2)incubating 6,12,24 h,HICR under hypoxia in pretreatment was calculated. Western blot method was used to detect the multi-drug resistance associated protein(MRP)expression in SKOV3 under the conditions of normoxia 24 h cell supernatant(group C), hypoxia 24 h cell supernatant(group D),hypoxia 24 h cell supernatant+10 μmol/L chloroquine(group E). RESULTS:TLR9 was expressed in both cell membrane and cytoplasm of SKOV3. Compared with group A,cell inhibition rate in group B was decreased (P<0.01). Compared with normoxia cell supernatant,cell inhibition rate was decreased after hypoxia cell supernatant (P<0.05). Compared with not added chloroquine,adding 10,102 μmol/L chloroquine can obviously decrease HICR(P<0.01),and the most significant decrease was showed when using hypoxia 24 h cell supernatant+10 μmol/L chloroquine. Compared with group C,MRP expression in group D was strengthened (P<0.01);compared with group D,MRP expression in group E was weakened (P<0.01). CONCLUSIONS:TLR9 receptors can be activated under hypoxia,and cause SKOV3 to cisplatin-resistance,the effect may be related with up-regulating MRP expression.