Effects Of Recombinant Human Tumor Necrosis Factor Alpha On Hematopoiesis Of Bone Marrow In Mice

Objectives To study the effect of different doses of nrhTNFa and synergistic action of nrhTNFa combined with chemotherapeutic drug on the hematopoiesis of bone marrow in mice, and to explore its toxic mechanism on mice bone marrow cells. Methods Low (1.25×10~6 U/kg), median (5.0×10~6 U/kg) and high (2.0×10~7 U/kg) doses of nrhTNFa were injected intramuscularly. The number of nucleated cells in mouse bone marrow as well as the number of white blood cells (WBCs) and platelets (PLTs) was counted and the content of haemoglobin (HB) was measured in peripheral blood. The hyperplastic degree of nucleated cells in mouse bone marrow was observed under light microscope. Apoptosis of mouse bone marrow cells was detected by flow cytometry, agarose gel electrophoresis and electron microscopy. Cell proliferation cycle of bone marrow in mice was evaluated by flow cytometry. Results 1. The number of bone marrow nucleated cells and peripheral blood cells was not affected by low dose of nrhTNFa, but inhibited by high dose of nrhTNFa. Median dose of nrhTNFa had no effects on the number of peripheral blood cells, but had a slight effect on the count of bone marrow nucleated cells (BMNCs). Different doses of nrhTNFa showed no synergetic action with cyclophosphamide (CTX). Meanwhile, the count of platelets was not affected by any dose of nrhTNFa.2. The effects of nrhTNFa on bone marrow cell apoptosis: (1).Under electron microscopy, the earlier morphological characteristics of apoptosis was observed in bone marrow cells of mice treated by high dose of nrhTNFa. (2).DNA flow cytometric analysis showed that the apoptosis percentage of bone marrow cells in mice treated by high, median and low doses of nrhTNFa was 1.2%, 2.4% and 16.2%, respectively.(3). "DNA ladder" was detected by agarose gel electrophoresis in bone marrow cells of mice treated by high and median doses of nrhTNFa.3. The effects of nrhTNFa on cell cycle: The number of cells in G2/M phase was increased while that in G0/G1 phase and S phase was decreased. This indicated that high dose of nrhTNFa arrested the cell proliferation at G2/M phase and inhibited mitosis of bone marrow cells in mice.Conclusions 1. Low dose of nrhTNFa had no inhibitory effects on mouse hematopoietic function. 2. Median dose of nrhTNFa had a slight effect on mouse hematopoietic function. 3. High dose of nrhTNFa inhibited mouse hematopoiesis, but had no synergetic action with CTX. 4. The inhibitory effects of nrhTNFa on mouse hematopoietic function were possibly related with its action of apoptosis induction and inhibition of mitosis. It has reversibility to the inhibitory effects on mouse hematopoietic function.