Protective Effect Of RhPTH On Mice Hematopietic Injury

Objective:To study the improvment of recombinant human parathyroid hormone (rhPTH) on hematopoietic injury, four separated goals were explored. (1) To study the effect of rhPTH on mobilization of hematopoietic stem cells. (2)To investigate the protective effect of rhPTH on mice hematopietic injury induced by chronic benzene poisoning. (3)To examine the protective effect of rhPTH on acute aplastic anemia. (4)To explore the improving effect of rhPTH on chronic aplastic anemia.Methods:(1)Normal C57BL/6J mice were divided into two groups : rhPTH Group and Normal Saline(NS) Group. rhPTH Group received 80μg/kg rhPTH administration intraperitoneally for 7 days consecutively. NS Group received equal volume saline. The number of peripheral blood was counted by Hematology Analyzer; the number of hematopoietic progenitor cells in peripheral blood was measured by colony culture; CD34+ cells in peripheral blood were measured by Flow Cytometry.;the level of Granulocyte-Colony Stimulating Factor(G-CSF)and Stromal Cell Derived Factor 1(SDF-1)in serum were measured by ELISA. (2) Subcutaneous injection of 2 ml/kg benzene for 26 treatments in Kunming mice was used to establish chronic benzene poisoning model. Mice were divided into 3 groups:NS control treated Group ,40μg/kg rhPTH treated Group, 80μg/kg rhPTH treated Group. The model mice recieved the corresponding dose of rhPTH or saline intraperitoneally 5 times a week for 3 weeks. Bone marrow hematopoietic stem cells were measured by colony forming unit in the spleen (CFU-S). The number of bone marrow hematopoietic progenitor cells was measured by colony culture. The number of peripheral blood cells was counted by Hematology Analyzer. Spleen index was measured by weighting method. (3) To establish the acute aplastic anemia model, the cells from lymph nodes and thymus of DBA/2J mice were injected intravenously to Balb/c mice previously irradiated with a 6.0 Gy dose ofγ-ray. rhPTH treated group was administrated 80μg/kg rhPTH 5 times per week intraperitoneally. NS control treated group was injected with saline. Administration began one week before the model was established and lasted for 3 weeks. The level of CD34+ cells reflects hematopoietic stem cells;The number of bone marrow hematopoietic progenitor cells was measured by colony culture; the number of peripheral blood cells was counted by Hematology Analyzer; Spleen index was measured by weighting method.(4)The lymph nodes cells of DBA/2J mice were injected intravenously to Balb/c mice previously irradiated with a full body 5.0 Gy dose ofγ-ray. A model of chronic aplastic anemia was established. rhPTH treated group was administrated 80μg/kg rhPTH 5 times per week intraperitoneally. NS control treated group was injected with saline. The treatments were carried out one week before the model was made and lasted for 6 weeks. The level of CD34+ cells represents hematopoietic stem cells;the number of bone marrow hematopoietic progenitor cells was measured by colony culture; the number of peripheral blood was counted by Hematology Analyzer; Spleen index was measured by weighting method, the result of cell cycle reflected bone marrow cell proliferations.Results:(1)rhPTH significantly increased the number of leucocyte, hematopoietic progenitor cells and CD34+ cells in peripheral blood(p<0.05). However, it had no effect on serum G-CSF and SDF-1.(2)The CFU-S counts of mice exposed chronically to benzene in the present of either 40μg/kg rhPTH or 80μg/kg rhPTH were higher than those in NS control treated group(p<0.05);The spleen index in 2 rhPTH treated groups was notablly lower than that in NS control treated group(p<0.05); The number of bone marrow hematopoietic progenitor cells and peripheral blood counts showed no significant difference among the three groups.. (3) The number of bone marrow hematopoietic stem cells,bone marrow hematopoietic progenitor cells,peripheral blood counts and spleen index in model mice were notably lower than those in normal mice(p<0.05),reflecting the acute aplastic anemia mice model was successfully generated. The number of bone marrow hematopoietic stem cells in rhPTH treated group was significantly higher than that in NS control treated group(p<0.05). The number of bone marrow hematopoietic progenitor cells,peripheral blood counts and spleen index showed no significant difference between rhPTH treated group and NS control treated group. (4) The chronic aplastic anemia mice model was thus successfully established. The number of bone marrow hematopoietic stem cells and spleen index in rhPTH treated group was markly increased than that in NS control treated group(p<0.05). The number of bone marrow hematopoietic progenitor cells and peripheral blood counts in rhPTH treated group were insignificantly different from those in NS control treated group. The rate of cells in S-G2-M-phase in rhPTH treated group was significantly increased compared to the NS control treated group(p<0.05), reflecting an enhanced proliferating activity through rhPTH.Conclusions:1. rhPTH effectively induces mobilization of stem/progenitor cells.2. In three hematopoietic injury models, rhPTH has a protective effect in the recovery of hematopoietic stem cells but has no significant effect on the bone marrow hematopoietic progenitor cells and peripheral blood cells.3. rhPTH has a protective effect on hematopoietic system. Future studies are needed to determine dosing,treatment procedure and potential clinical application of our findings.