Effect Of RhBMP-2m On Hematopoietic Restoration In Acute Hematopoietic Injured Mice And Preliminary Study Of The Mechanism

Bone morphogenetic protein is the member of the superfamily of TGF- P (Transforming Growth Factor- P ). Originally recongnized as bone inductive protein , BMP was subsequently found to play an important role not only in the repair of large bone defects but also in the induction of embryo development and activation of neural and hematopoietic tissue differentiation. BMP is supposed to have a promising prospect of clinical application.The characteristics of acute hematopoietic injury caused by radiation or chemical drugs are the damage of hematopoietic stem cells in the marrow and themicro hematopoietic environment, which lead to a pathological process characterized by infection and bleeding. So far there are no preventive drugs specifically effective to treat such damage. In the radiotherapy and chemotherapy of malignant tumors, many patients died of damage of their hematopoietic system.Our previous studies showed that rhBMP-2m could relieve acute hematopoietic injury caused by radiation of y -rays. BMP could treat and repair hematopoietic injury, whose function of inducing hematopoiesis was obviously superior to CSF, EPO and TPO. The mechanism is that BMP can upregulate some genes related to hematopoiesis, thus promoted the proliferation of hematopoietic stem cells and marrow stromal stem cells.In clinical practice, when chemicals are used to treat malignant tumors, their dosage is usually limited because they may inhibit hematopoiesis, thus the therapeutic efficacy is reduced. Therefore it is very important to protect hematopoietic tissues from damage in the chemotherapy. Based on our previous experiments, we set up a murine model of acute hematopoietic injury caused by intraperitonealy(IP) injecting CTX of 200 mg/kg body weight. The mice were divided randomly into 3 groups, namely CTX injection group(CTX group), BMP therapy group(CTX+BMP group) and PBS control group(Control group). CTX of 200 mg/kg per mouse was IP injected at a time to CTX+BMP group and CTX group so as to establish the experimental model of mouse bone marrow injury. In CTX+BMP group, the therapy started from the first day after injection of CTX by using IP injection of 0.5 mg/0.5 mL BMP per mouse each day. In control group, same volume PBS was injected only. Changes of peripheral blood leukocyte numbers in 3 groups were observed . On the 5th and 8th day after CTX injection, DNA content in mouse bone marrow karyocytes and variation of cell cycle wereanalysed by flow cytometry(FCM). Colony forming unit granulocyte-macrophage (CFU-GM) was cultivated simultaneously.To study the source of hematopoietic cells when rhBMP-2m is used to promote hematopoietic restoration in mice with acute hematopoietic injury, female BALB/c mice irradiated by 8.5 Gy Co Y ray, used as donors, received marrow cells from male mice of the same breed. When the marrow was successfully transplanted, the mice were shielded except their left legs which received local irradiation with 9 Gy electronic rays. Then they were treated with rhBMP-2m . Corresponding special primers were designed for the mRNA sequence of AEF and Sry, 2 sex-determining genes from Y-chromosome of male mice. The RT-PCR was used to detect the expression of AEF and Sry in the marrow cells of the femur both in the therapeutic group and irradiated group. The existence of Y-chromosome gene in marrow cells of the female mice that were the acceptor in the transplantation could indirectly lead to our judgment of the source of the new hematopoietic cells in the radiated mice. In this way we provide evidence to the source of the new hematopoietic cells after acute hematopoietic injury.In conclusion, BMP has some therapeutic effect on mouse bone marrow injury caused by CTX. The source of hematopoietic cells in the marrow of a mouse is probably its own stromal cells which, after induced by rhBMP-2m, turn into hematopoietic cells.