[Research of reversal effect of PESV to multi-drug resistance of leukemia stem cell].

Using the BALB/c mouse multidrug resistance model of leukemia, the effect of peptide extract from scorpion venom (PESV) to the upstream signal factors of P-gp of MDR leukemia stem cells on the mouse tumor block was observed, and the mechanism of PESV to reverse the MDR of LSC was studied. At the same time, the expression of P-gp, MDR1 mRNA and PI3-K, NF-κB were respectively detected through flow cytometry, RT-PCR, Western blot and Elisa, and the mouse liver, spleen were examined via histopathological methods. The results of the experiment were as follows: mice of the control group didn't show any obvious changes, while mice of the other six groups all showed arched back, emaciation, liver swell, and inflammation was found in all liver tissue. The expression level of P-gp and PI3K on the LSC membrane of mouse tumor block was down-regulated; the expression of MDR1 mRNA in the cytoplasm was obviously down in the PESV low dose group, and which was inordinately up in the middle dose group and the high dose group. The expression level of NF-κB in the leukemia stem cell nucleus remarkably decreased. PESV had a outstanding role of down-regulating PI3K, NF-κb, MDR1 which were all upstream factors of P-gp, and to a certain degree enhanced the sensitivity of LSC to ADM. Therefore, this experiment explained one of the mighty mechanism of PESV to reverse MDR of LSC, and provided a foundation to further study of combinational anti-cancer effects of PESV.

[Reversion Mechanism Study of PESV to Multidrug Resistance at Leukemia Stem Cell Level].

OBJECTIVE: To explore the effect of peptide extract from scorpion venom (PESV) to multidrug resistance (MDR) of leukemic stem cell (LSC) in vivo. METHODS: K562/A02 cells were cultured and collected in the logarithmic phase. K562/A02 stem cells were screened using immunomagnetic beads for reserve. K562/A02 LSC was injected to 5 of 40 BABL/c nude mice for preparing subcutaneous tumor. The rest 35 nude mice were then randomly divided into 7 groups, i.e., the normal control group, the model group, the Adriamycin (ADM) group, the PESV group, the ADM +high dose PESV group, the ADM + middle dose PESV group, the ADM +low dose PESV group, 5 in each group. Tumor tissue was embedded in all groups except the normal control group. One milliliter normal saline was peritoneally injected to mice in the model group after modeling, once per day. ADM 0. 05 mg was peritoneally injected to mice in the ADM group, once per other day. PESV 2 µg was peritoneally injected to mice in the PESV group, once per day. Mice in 3 ADM + PESV groups were peritoneally injected with ADM 0. 05 mg (once per other day) plus PESV (5, 2, and 1 µg respectively, once per day). All medication lasted for 14 days. P-glycoprotein (P-gp) was detected using flow cytometry. Breast cancer resistance protein (BCRP) and mRNA expression of multidrug resistance 1 (MDR1) were measured using RT-PCR. Aldehyde dehydrogenase 1 (ALDH1) was detected using immunohistochemistry. Phosphoinositide 3-kinase (PI3K) was detected using Western blot. NF-κB content was detected using ELISA. RESULTS: CD34 + CD38-ratio was 31.5% and IC50 was (60.33 ± 10. 68) µg/mL before K562/A02 cells were screened with immunomagnetic beads, while they were 92. 8% and (58. 33 ±9. 72) µg/mL after screen. The tumor formation rate was 100% in modeling mice. Compared with the model group, no statistical difference of each index occurred in the ADM group (P <0. 05). There was statistical difference in BCRP, MDR1 mRNA, or NF-κB factor between the model group and the PESV group (P <0. 05). The expression level of P-gp obviously decreased and the protein expression of P13K was down-regulated in 3 ADM + PESV groups (P <0. 05); mRNA expression of BCRP decreased and mRNA ex- pression of MDR1 obviously increased in the ADM + high dose PESV group and the ADM + middle dose PESV group, with statistical difference (P <0. 05). Protein expression of P13K was down-regulated in the ADM+ high dose PESV group, with statistical difference (P <0. 05). P-gp value, BCRP mRNA expression, MDR1 mRNA expression, PI3K, and NF-κB factor were all obviously down-regulated in the ADM +high dose PESV group, as compared with the ADM group and the PESV group respectively (P <0. 05). There was no statistical difference in ALDH1 positive rate among all groups (P >0. 05). Conclusion PESV combined ADM could down-regulate expression levels of P-gp, BCRP, MDR1, P13K, and NF-κB, strengthen the sensitivity of K562/A02 LSC to ADM in vivo, and reverse MDR of LSC.

Effects of the total saponins from Dioscorea nipponica on immunoregulation in aplastic anemia mice.

Dioscorea nipponica Makino, a popular folk medicine, exerts anti-inflammation properties. The present study investigated the therapeutic effect of the total saponins from Dioscorea nipponica Makino (TSDN) on aplastic anemia (AA) and possible immune regulation mechanisms. Using a mouse model of AA, three different doses of TSDN were orally administrated for 14 consecutive days. We first demonstrated that TSDN was found to be effective in alleviating pancytopenia with a hypocellular bone marrow as compared with AA model group. Moreover, gastrogavage administration of a medium dose of TSDN was found to dramatically increase the percentage of CD4(+) cells in bone marrow nucleated cells (BMNC) and restore the CD4(+)/CD8(+) ratio. The pro-inflammatory cytokine concentrations of IL-2 and IFN-γ were significantly decreased, and anti-inflammatory cytokine IL-4 was significantly increased in culture supernatant of BMNC. Further investigations showed that TSDN obviously inhibited Fas-FasL-induced BMNC apoptosis as well as effectively suppressed intracellular apoptosis protein of caspase-3 and -8 expressions. Taken together, these findings suggested that TSDN could alleviate AA by elevating the CD4(+)/CD8(+) T-cell ratio, inhibiting inflammatory Th1-cytokines, and exerting anti-apoptosis effects.