A Study On The Effective Mechanisms Of Recombinant Human Erythropoietin On The Immune Function Of Bone Marrow Suppression Mice

Objective: Carcinomas are fatal diseases to the health of children and often have much sequel to affect their development, it's mortality stand second on the list of age from 5 to 14 in our country. The major treatment measure is chemotherapeutics at present, but chemical drugs can also kill normal cells when they kill tumor cells, the side effects are serious. Bone marrow suppression is one of the common side effects, it can cause anemia, lower immune function, these patients can be infected easily, the mortality is high. Chemotheraputics and tumors can cause severe anemia, patients need blood transfusion many times, but blood transfusion has a lot of side effects. It has great clinical significance to search for methods and drugs which can prevent and cure anemia and improve lower immune function. Recombinant human erythropoietin (rh-EPO) treatment has been used successfully for end-stage renal disease and cancer-related anemia. Recent studies have found that EPO and EPO-R also consist in non-erythrocyte cells, EPO can affects blood system, immune system, nerve system, etc. This research use mice model to study the effect of rh-EPO on hemoglobin (Hb), white blood cell (WBC), red blood cell (RBC), platelate(PLT), T lymphocyte proliferative responsiveness, red blood cell immune function, production of interlikin-2 (IL-2) and tumor necrosis factor α(TNF-α). Methods: 60 six-week-old mice were randomly divided into: (1) cyclophosphamide+saline group (CTX+NS): saline was administered every other day for six times at the back flank, on the 4, 8 days cyclophosphamide of 150mg/kg.t was administered respectively in the abdomen area. (2) Cyclophosphamide+lower dosage rh-EPO group (CTX+LDrh-EPO): rh-EPO of 250 IU/kg.t was administered every other day for six times, on the 4, 8 days cyclophosphamide of 150mg/kg.t was administered respectively. (3) Cyclophosphamide+higher dosage rh-EPO group (CTX +HDrh-EPO): rh-EPO of 500 IU/kg.t was administered every other day for six times, on the 4, 8 days cyclophosphamide of 150mg/kg.t was administered respectively. (4) saline normal control group (NS): only saline was administered. There were 15 mice in each group. Mice were killed at the directed time and blood, spleen were prepared. The changes of Hb, WBC, NE/LY, PLT, T lymphocyte proliferative responsiveness (measured with MTT methods), red blood cell immune function (C3b-R%) , and production of IL-2 and TNF-αby splenocytes (measured with radioimmunoassay) were observed. Results: During this experiment, 60 mice were all left in the research. (1) Levels of Hb, RBC, WBC and PLT in CTX+NS group were (87.300±12.783)g/L, (2.620±0.523)×1012/L, (3.553±0.276)×109/L , (108.500±9.637)×109/L respectively.They were (102.860±8.941) g/L, (3.107±0.276)×1012/L, (4.610±0.621)×109/L, (140.364±12.283)×109/L respectively in normal control group. These index in CTX+NS group were significantly lower than that in normal control group (p<0.05) , so the model were successfully made. (2) The changes of WBC and PLT: levels of WBC in CTX+LDrh-EPO group (3.671 ±0.322)×109/L and CTX+HDrh-EPO group (3.734±0.343)×109/L were little higher than that in CTX+NS group (p>0.05). Levels of PLT in CTX+LDrh-EPO group (110.146±8.453)×109/L and CTX+HDrh-EPO group (112.760±9.362)×109/L were little higher than that in CTX+NS group (p>0.05). (3) Changes of NE/LY: Levels of NE/LY in CTX+NS group (0.615±0.034) was significantly lower than that in normal control group (1.115±0.072), (p<0.01). While levels of NE/LY in CTX+LDrh-EPO group (0.624±0.048) and CTX+HDrh-EPO group (0.641±0.042) were little higher than that in CTX+NS group (p>0.05). (4) Changes of Hb and RBC: Levels of Hb in CTX+LDrh-EPO group (94.825±8.204)g/L and CTX+HDrh-EPO group (106.720±10.717) g/L were significantly higher than that in CTX+NS group (p<0.05). Levels of RBC in CTX+LDrh-EPO group (2.846±0.257)×1012/L and CTX+HDrh-EPO group (3.203±0.321)×1012/L were significantly higher than that in CTX+NS group (p<0.05). Compared between CTX+LDrh-EPO group and CTX+HDrh-EPO group, levels of Hb and RBC in CTX+HDrh-EPO group were significantly higher than that in CTX+LDrh-EPO group (p<0.05). (5) Changes of red blood cellimmune function (C3b-R%): Level of C3b-R% in CTX+NS group (12.813±1.114) was significantly lower than that in normal control group (16.633±1.417), (p<0.01). Levels of C3b-R% in CTX+LDrh-EPO group (14.257±1.058) and CTX+HDrh-EPO group (15.846±1.593) were significantly higher than that in CTX+NS group(p<0.05). Compared between CTX+LDrh-EPO group and CTX+HDrh-EPO group, level of C3b-R% in CTX+HDrh-EPO group was significantly higher than that in CTX+LDrh-EPO group (p<0.05). (6) Changes of T lymphocyte proliferative responsiveness OD index (A): Level of A in CTX+NS group (0.155±0.013) was significantly lower than that in normal control group (0.283±0.05), (P<0.01). Levels of A in CTX+LDrh-EPO Group (0.199±0.021) group and CTX+HDrh-EPO group (0.241±0.024) were significantly higher than that in CTX+NS group (p<0.05), level of A in CTX+HDrh-EPO group was significantly higher than that in CTX+LDrh-EPO group (p<0.05). (7) Changes of IL-2: Level of IL-2 in CTX+NS group (207.217±4.223) pg/ml was significantly lower than that in normal control group (320.630±10.442) pg/ml, (p<0.01). Levels of IL-2 in CTX+LDrh-EPO group (230.302±9.724) pg/ml and CTX+HDrh-EPO group (285.901±11.386) pg/ml were significantly higher than that in CTX+NS group (p<0.05), level of IL-2 in CTX+HDrh-EPO group was significantly higher than that in CTX+LDrh-EPO group (p<0.05). (8) Changes of TNF-α: Level of TNF-αin CTX+NS group (0.252±0.021) ng/ml was significantly lower than that in normal control group(0.297±0.016) ng/ml, (p<0.05). While levels of TNF-αin CTX+LDrh-EPO group (0.257±0.018) ng/ml and CTX+HDrh-EPO group (0.261±0.017) ng/ml were little higher than that in CTX+NS group (p>0.05). (9) Correlation between C3b-R% and A, C3b-R% and IL-2: There were positive correlation between C3b-R% and A, C3b-R% and IL-2. The correlation coefficient between C3b-R% and A in CTX+LDrh-EPO group was (r=0.812, p<0.01), in CTX+HDrh-EPO group was (r=0.765, p<0.01). The correlation coefficient between C3b-R% and IL-2 in CTX+LDrh-EPO group was (r=0.689, p<0.01), in CTX+HDrh-EPO group was (r=0.755, p<0.01). Conclusions: (1) In respect that the observation of Hb, RBC, WBC and PLT, we believe that we duplicated the bone marrow suppression model successfully. (2) Cyclophosphamide-treated mice had lower WBC and Hb, lower red blood cell immune function and T lymphocyte proliferative responsiveness, lower production of IL-2 and TNF-α. Cyclophosphamide-treated mice had an improvement in Hb, T lymphocyte proliferative responsiveness, red blood cell immune function and IL-2 after rh-EPO administration, while levels of WBC and TNF-αwere not changed. There were positive correlation between red blood cell immune function and T lymphocyte proliferative responsiveness, between red blood cell immune function and IL-2 level. (3) rh-EPO administration had an improvement in T lymphocyte proliferative responsiveness, red blood cell immune function and IL-2 level as well as improve anemia. (4) The...