SVP-B5 peptide from Buthus martensii Karsch scorpion venom exerts hyperproliferative effects on irradiated hematopoietic cells

Abstract

Previous studies have demonstrated the radioprotective efficacy of scorpion venom peptide, fraction II (SVPII) from the venom of Buthus martensii Karsch. In the present study, the SVP-B5 polypeptide, which is one of the active components of SVPII, was purified using a two-step chromatographic process. SVP-B5 significantly promoted the proliferation of irradiated M-NFS-60 mouse-derived myelocytic leukemia cells. In addition, SVP-B5 effectively and persistently promoted hematopoietic recovery and expansion of hematopoietic cells after irradiation as demonstrated by cobblestone area forming cell and long-term bone marrow culture assays. Treatment of M-NFS-60 cells with SVP-B5 upregulated the expression of interleukin 3 receptor and activated the Janus kinase-2/signal transducer and activator of transcription 5 signaling pathway. In conclusion, the present study demonstrated that SVP-B5 has growth factor-like properties and may be used as a therapeutic modality in the recovery of severe myelosuppression, which is a common side effect of radiotherapy.

Keywords: hematopoiesis; interleukin-3; irradiation; polypeptides; scorpion venom.

Figure 1.

Isolation, purification and identification of scorpion venom. (A) Three protein peaks were identified in the scorpion venom crude extract (peaks I, II, and III) obtained Sephadex G-50 gel chromatography. (B) Seven peaks (B1-B7) were identified within peak II separated by CM-Sepharose FF ion chromatography. Products were further characterized by reverse-phase high-performance liquid chromatography.

Figure 2.

Effects SVP-B4 and SVP-B5 on cell proliferation. Irradiated M-NFS-60 cells treated with SVP-B4 and SVP-B5 for (A) 24 and (B) 48 h. The cell proliferation rate is expressed as the fold increase compared with the control group. *P<0.05; **P<0.01; ***P<0.005 vs. untreated.

Figure 3.

Effect of SVP-B5 on CAFCs and CFU-GMs. CAFC results at (A) 14 days and (B) at 35 days. CFU-GM results at (C) day 7 and (D) day 14. PBS was used as a vehicle in the control cells. *P<0.05 and **P<0.01vs. vehicle. CAFCs, cobblestone area forming cells; CFU-GMs, granulocyte-monocyte colony forming units; IR-0 Gy, not irradiated; IR-2 Gy, irradiated with X-rays at 2 Gy; BMCs, bone marrow cells.

Figure 4.

Effect of SVP-B5 on the expression of IL-3R. (A and B) M-NFS-60 cells were treated with SVP-B5 for (A) 24 and (B) 48 h and expression of IL-3R was observed by immunohistochemistry (magnification, ×400). (C) Representative western blot showing the increased expression of IL-3R in M-NFS-60 cells treated with SVP-B5 for 24 h. GAPDH served as a loading control. Cells treated with IL-3 served as a positive control, and an untreated CTL group was also used. CTL, control; IL-3R, interleukin 3 receptor.

Figure 5.

Effect of SVP-B5 on the phosphorylation of JAK2 and STAT5. Representative western blots displaying increased levels of P-JAK2 and P-STAT5 after SVPB5 (1 µg/ml) treatment with SVP-B5 for different periods. Cell treated with IL-3 served as a positive control. P-JAK2, phosphorylated Janus kinase 2; STAT5, signal transducer and activator of transcription 5; IL, interleukin.