[Effect of anti-TGF-beta antibody on ex vivo expansion and expression of adhesion molecules of human cord blood CD34+ cells].
TGF-beta, as an inhibitor of hemopoiesis, excreted by hematopoietic stem and progenitor cells, down-regulates the expression of cytokines such as Flt-3 ligand, SCF, IL-3 etc on the stem and progenitor cells. The effect of anti-TGF-beta antibody on ex vivo expansion and expression of adhesive molecules on cord blood CD34(+) cells was studied in this research. The CD34(+) cells from six units of fresh umbilical cord blood were enriched by density gradient sedimentation and purified by miniMACS cell isolation system, and plated them into the SFEM serum free culture system which containing SCF, Flt-3L, TPO and IL-3 in the condition of 37 degrees C, 5% CO2, and saturated moisture. There were three groups in this experiment: (1) blank group: same as the culture system described above; (2) control group: added with normal rabbit IgG into the mentioned culture system; (3) test group: the same culture system with anti-TGF-beta1 antibo-dy. Cultured for 6 days, the number of mononuclear cells (MNC) was counted, the expression of CD34 antigen, CD117 (c-kit) antigen, CD11a antigen, CD49d antigen and CD33 antigen was tested with FCM. Meanwhile, cells of the three groups were plated in the methylcellulose culture system for 14 days, the number of CFU-GEMM, BFU-E, CFU-GM was counted. The results indicated that the expansion multiples of MNC, CD34(+) cells, CD34(+)c-kit(+) cells, CFU-GEMM in the test group (41.82 +/- 13.49, 15.62 +/- 6.95, 13.36 +/- 6.12, 11.07 +/- 4.05) were significantly higher than in the control group (28.86 +/- 9.03, 10.40 +/- 4.98, 9.04 +/- 4.40, 6.36 +/- 2.37) (P = 0.001, 0.002, 0.003, 0.002) respectively. The expansion multiple of more primitive CD34(+)c-kit(-) subpopulation in the test group (69.10 +/- 41.06) was even higher than in the control group (27.29 +/- 10.40) (P = 0.024). Adhesion molecule expression on the CD34(+) cells after short-term expansion: the expression of CD11a on the CD34(+) cells of the original cord blood was (61.73 +/- 4.13)%, and CD49d was (55.12 +/- 5.22)%. After expansion in each group the expression of CD11a on the CD34(+) cells did not change with statistical significance (P > 0.05), the expression of CD49d increased (P < 0.05). Compared with blank group and control group, anti-TGF-beta antibody did not impact on the expression of CD11a and CD49d (P > 0.05). It is concluded that anti-TGF-beta antibody can synergize other cytokines to effectively enhance the proliferation of cord blood NC, CD34(+) cells, progenitor subpopulation of CD34(+)c-kit(-) cells, and increase the output of more primitive progenitor colony, CFU-GEMM and BFU-E. At the same time, anti-TGF-beta antibody did not depresss the expression of adhesion molecules on CD34(+) cells.
Qian
Jun
J
Department of Hematology, Beijing Tongren Hospital Affliated to Capital University of Medical Sciences, Beijing 100730, China.
Liu
Fu-Qiang
FQ
Wu
Yi-Ping
YP
Wang
Yang
Y
Kang
Wen-Ting
WT
D000906:Antibodies / Q000494:pharmacology*; D018952:Antigens, CD34 / Q000032:analysis*; D040881:CD11a Antigen / Q000032:analysis; D015815:Cell Adhesion Molecules / Q000032:analysis*; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D005260:Female; D005312:Fetal Blood / Q000166:cytology / Q000276:immunology; D005434:Flow Cytometry; D006801:Humans; D039441:Integrin alpha4 / Q000032:analysis; D011247:Pregnancy; D019009:Proto-Oncogene Proteins c-kit / Q000032:analysis; D016212:Transforming Growth Factor beta / Q000276:immunology*
D000906:Antibodies; D018952:Antigens, CD34; D040881:CD11a Antigen; D015815:Cell Adhesion Molecules; D016212:Transforming Growth Factor beta; D039441:Integrin alpha4; D019009:Proto-Oncogene Proteins c-kit