ABSTRACT
The pathogenesis of aplastic anemia (AA) was explored and the effects of AA serum on the expression of crucial cyclin D isoform (cyclin D3) in umbilical cord blood hematopoietic stem/progenitor cells were observed. The CD34+ cells were isolated from the cord blood with MIDI-MACS Semi-solid methylcellulose culture technique was used to measure the formation of CFU-GM; The expression level of cyclin D3 was assayed by semi-quantitative RT-PCR and Western-blot after the hematopoietic stem/progenitor cells were incubated in AA serum. The results showed that the AA serum could inhibit the formation of CFU-GM and down regulate the expression level of the cyclin D3 at the mRNA and protein level respectively. In conclusion, the AA serum could inhibit the proliferation of hematopoietic stem cells and down regulate level of cyclin D3, which might be one mechanism of hematopoiesis inhibition in AA.
Subject(s)
Anemia, Aplastic/blood , Antigens, CD34/metabolism , Cells, Cultured , Colony-Forming Units Assay , Cyclins/biosynthesis , Cyclins/genetics , Fetal Blood/cytology , Hematopoietic Stem Cells/cytology , Protein Isoforms/biosynthesis , Protein Isoforms/genetics , RNA, Messenger/biosynthesis , RNA, Messenger/genetics , SerumABSTRACT
The pathogenesis of aplastic anemia (AA) was explored and the effects of AA serum on the expression of crucial cyclin D isoform (cyclin D3) in umbilical cord blood hematopoietic stem/progenitor cells were observed. The CD34+ cells were isolated from the cord blood with MIDI-MACS Semi-solid methylcellulose culture technique was used to measure the formation of CFU-GM; The expression level of cyclin D3 was assayed by semi-quantitative RT-PCR and Western-blot after the hematopoietic stem/progenitor cells were incubated in AA serum. The results showed that the AA serum could inhibit the formation of CFU-GM and down regulate the expression level of the cyclin D3 at the mRNA and protein level respectively. In conclusion, the AA serum could inhibit the proliferation of hematopoietic stem cells and down regulate level of cyclin D3, which might be one mechanism of hematopoiesis inhibition in AA.
Subject(s)
Female , Humans , Male , Anemia, Aplastic , Blood , Antigens, CD34 , Metabolism , Cells, Cultured , Colony-Forming Units Assay , Cyclin D3 , Cyclins , Genetics , Fetal Blood , Cell Biology , Hematopoietic Stem Cells , Cell Biology , Protein Isoforms , Genetics , RNA, Messenger , Genetics , SerumABSTRACT
AIM: To explore the pathogenesis of aplastic anemia (AA), we identified the crucial isoform of cyclin D that determine the proliferation of the cord blood CD34~+ cells and observed effects of AA serum on the expression of crucial cyclin D isoform in umbilical cord blood CD34~+ cells. METHODS: The CD34~+ cells were isolated with MIDI-MACS system. The isoforms of cyclin D were detected by RT-PCR and Western blotting. Methylcellulose culture system was used to measure the formation of CFU-GM. The expression level of crucial cyclin D isoform was assayed by RT-PCR and Western blotting after the CD34~+ cells were incubated in AA serum. RESULTS: The crucial cyclin D isoform in CD34~+ cells was cyclin D3. The AA serum inhibited the formation of CFU-GM and down-regulated expression level of the cyclin D3 from the mRNA to protein level, respectively. CONCLUSION: The AA serum inhibits the proliferation of CD34~+ cells and down-regulates level of cyclin D3, this may be one of hematopoiesis inhibition mechanisms in AA.