The Experimental Research On The Inhibition Of Human Soluble Vascular Endothelial Growth Factor-1 On Leukemic Cell Proliferation

Vascular endothelial growth factor (VEGF) is one of the mostpowerful angiogenesis factors, which can promote the activation ,migration,and proliferation differentiation of endothelial cells, then forming vessels.VEGF also can enhance the permeability of vessels. Hypoxia and activationof oncogene are most effective factor to induce the expression of VEGF andits receptor(VEGFR). In adult physiological state, VEGF scarcely express,effecting by hypoxia and ischemia, many differentiated cells can expressedVEGF. Tumor cells will express VEGF induced by oncogene, the effect ofVEGF on angiogenesis has been widely identified. It is a new focus in thebiological targeted therapy to cure tumor with the method of antiangiogenesis.We have achieved significant progress in the research on solid tumor therapyby antiangiogenesis. Some angiogenic inhibitors have been saled, whichgained anticipative effects on the solid tumor therapy. Base on the aboveachievement , these good effects stimulate people to explore leukemictherapy by antiangiogenesis. Leukemia is a hematopoietic malignancy,deriving from bone marrow, which is different from solid tumor. Internal andoverseas research have made much work on whether antiangiogenesis can beapplied on the leukemia therapy. Already research demonstrated thathematological malignant tumor cells also express VEGF and VEGFR, whichcan stimulus the proliferation of malignant tumor cells and endothelial cellsthrough endocrine and paracrine path.Soluble VEGFR-1, also called sFlt-1 is the natural antagon of VEGF. Asdeficient of transmembrane domain and intracellular protein tyrosine kinasedomain, sFlt-1 do not excite the signal conduction, therefore ,we can utilizethe competive of sFlt-1 to consumpt the VEGF secreted by tumor cells,accordingly suppress the biologic activity of VEGF. The content of sFlt-1 inliving bodies is tiny, so in the most research , we take the advantage oftransgene technique, transfecting the cDNA of sFlt-1D4 to targeted cells andgain the stable protein of sFlt-1. Combining antiangiogenesis and genetherapy has potential perspective in tumor therapy.Reports showed that some leukemic cell lines express VEGF andVEGFR,In our experiment ,we take RT-PCR, Flow cytometer andenzyme-linked immunosorbent assay(ELISA) to identify the expression ofVEGF and Flt-1 in K562,HL60,U937 and bone marrow long-termculture-initiating cells(LTC-IC), the outcome showed that K562,HL60 andU937 cell lines expressed VEGF as well as its receptor,K562 line owned thehighest expression rate either in VEGF or Flt-1, expression rate of HL60 linewas the second, U937 line had a lower level of VEGF and Flt-1,neverthelessLTC-IC scarcely expressed VEGF and Flt-1. We detect VEGF and Flt-1usingthree types method , the result is more reliable, the result of Flow cytometeris consistent with that of ELISA, more over these two method can make theanalysis in quantity .There is difference in the expression rate of VEGF andFlt-1 among these leukemic cell lines, reflecting the loading dose of tumor,so we should pay attention to the level of VEGF and VEGFR in body whencure leukemia by antiangiogenesis.In the next step , sFlt-1 was added to the culture system after plating thesame amount of these leukemic cell lines, the consequence demonstrated thatthe inhibition on K562 and HL60 cell lines was as bigger as increasing thedose of sFlt-1,there was a significant difference on inhibition ratio whenadding various concentration of sFlt-1,p<0.05.These data indicated theinhibition of sFlt-1 on leukemic cell lines was concentration dependent. Thisconsequence demonstrate that , in the course of anti-VEGF therapy, we mustmaintain a high lever of medicine to guarantee the continuity and efficacy ofsFlt-1. As control group , the proliferation of LTC-IC was scarcely influenced.Based on the above findings, proliferation of normal bone marrow cells cannot be affected when the activity of VEGF is suppressed, so, it is feasibilityto transfect sFlt-1cDNA to bone marrow stroma cells for leukemia therapy.The next research will be on the animal, to certify sFlt-1 can suppressprogression of disease in leukemic mice and prolong life span. Thisbiotherapy can be considered as adjunctive therapy of chemotherapy,providing a new curative strategy for incurable and relapsing leukem.