Absorption Characteristic And Photodynamic Effect Of Two Hypocrellin B Derivatives In BGC-823Cells

Two derivatives of hypocrellin B (HB),17-5-amino-1-pentane-sulfonicacid-hypocrellin B（PENSHB）and15-Deacetyl-13-3-amino-1-propane-sulfonic acid-hypocrellin B（DPROHB）, in the aspects of absorptive characteristics, subcellularlocalization, mitochondrion damage, photodynamic therapy(PDT) effects and darktoxicity were compared with their parent HB in this study.（1）To investigate absorptive characteristics of PENSHB and DPROHB. HumanGastric Adenocarcinoma Cell (BGC-823) were cultured. PENSHB or DPROHB wereadded into culture medium in4μM concentration. To get the relationship betweenintracellular content and incubated time of photosensitizers, they were incubated fordifferent time. Samples were measured by spectrofluorophotometer The BGC-823cellswere incubated with PENSHB、 DPROHB in various concentrations for4h, therelationship between intracellular content and incubating concentrations ofphotosensitizers was ploted. The absorption curves of PENSHB and DPROHB byBGC-823cells were similar. The absorption content increased rapidly in the first1h, andreached platform at4h. In this experiment concentration range, intracellularphotosensitizers concentration are directly proportional to the concentrations of PENSHBand DPROHB, The curves show that their rising trend was similar. So the content ofPENSHB and DPROHB in BGC-823cells was dose and time dependent.（2）To validate the intracellular distribution of the two novel photosensitizers intumor cells, the BGC-823cells The cells were loaded with PENSHB or DPROHB incomplete medium in1h or4h. Photosensitizers fluorescence was imaged using a laserscanning confocal microscopy. The result show that photosensitizers are distributed incytoplasm, but not in cell nuclei.To assess whether two new photosensitizers binds to themitochondria, cells were co-loaded with Mito-Tracker, a mitochondria-specific dye, Our results show that photosensitizers localises to mitochondria in BGC-823cells, especially inthose cells incubated photosensitizers in4h. More interestingly, we found that theconcentration of photosensitizers near nucleus is higher than other parts in cells.（3）To compare the PDT effects and dark cytotoxicity between two newphotosensitizers and HB. The GBC-823cells were incubated with three photosensitizersrespectively in4h with different concentrations. Light-dependent cytotoxicity: cells wereirradiated by KTP laser under wavelength532nm with power density of20mW/cm2for1000s. The cytotoxicity group: cells were incubated with same medium withoutirradiation. Cell survival rates were measured by MTT assay respectively. Cell survivalrates to concentrations curve of each photosensitiser was plotted. The dark and light-dependent cytotoxicity IC50(50%inhibition concentration) of each photosensitizer werederived. The safety coefficient of the photosensitizers were calculated based on the darkand light-dependent cytotoxicity IC50. Light-dependent cytotoxicity IC50of PENSHB、DPROHB and HB were26.47、27.33、47.30nM, respectively. The statistical differenceexisted between HB and its derivatives, whereas no such difference existed betweenPENSHB and DPROHB. The cytotoxicity group: IC50of three photosensitizers were3.32、4.03、4.06μM, respectively. The safety coefficients of the individual photosensitizerwere125.43、147.46and85.84.Two novel hypocrellin derivatives PENSHB and DPROHB, with improved watersolubility, were absorpted faster than hypocrellin by cell. Additionally, they got a betterPDT effect under the similar dark cytotoxicity. In a word, PENSHB and DPROHB,asnew photosensitizers, have a good prospect of clinical application.