[Influence of cytokine combinations on proliferation and differentiation of umbilical cord blood CD34(+) cells into megakaryocytes/platelets in vitro].

In order to investigate the influence of cytokine combinations on proliferation and differentiation of human umbilical cord blood CD34(+) cells into megakaryocytes/platelets in vitro, the CD34(+) cells from human umbilical cord blood were amplified in serum-free medium StemSpan(SFEM) supplemented with several cytokine combinations by three-phase culture system. The effects of the cytokine combinations were compared. The results showed that at day 14 of the first culture phase, the CD34(+) cells cultured with cytokine combinations SCF + TPO + FL + IL-3 were amplified (11 000 ± 1 000) times, which were significantly higher than that of cells cultured with SCF + TPO + FL, but were not significantly different from that of cells cultured with SCF + TPO + IL-3 or SCF + TPO + FL + IL-3+ hydroxyl-corticosteroids. At day 7 of the second culture phase, the CD34(+) cells cultured with cytokine combination SCF + TPO + FL + IL-11 were amplified by (204666.7 ± 11718.9) times, which were significantly higher than that of cells cultured with SCF + TPO + FL + IL-3, but were not significantly different from that of cells cultured with SCF + TPO + FL + IL-11 + BMP4 + VEGF. At day 3 and day 6, the CD34(+) platelet-like cells accounted for about (39.8 ± 1.9)%, (39.7 ± 2.6)% and (25.5 ± 1.4)%, (23.1 ± 3.5)% cultured with SCF + TPO + FL + IL-11 and SCF + TPO + FL + IL-11 + BMP4 + VEGF, and significantly higher than that of the cells cultured with SCF + TPO + FL + IL-3. It is concluded that the cytokine combination of SCF + TPO + FL + IL-3 is most suitable cytokines combination for the amplification of CD34(+) hematopoietic progenitor cells. The cytokine combination of SCF + TPO + FL + IL-11 is preferred for the proliferation and differentiation of megakaryocytes, this study lays an experimental basis for investigating the proliferation and differentiation of CD34(+) into megakaryocytes/platelets in vitro.

[To identify a novel HLA-DRB1 allele in Chinese by sequencing].

OBJECTIVE: To identify a novel HLA-DRB1 allele in Chinese. METHODS: A novel HLA-DR allele was detected by PCR-SSP and SBT in a patient with leukemia. RESULTS: The sequence of the novel allele was different from all other known alleles. The novel allele differed from the closet matching allele HLA-DRB1*1404 by one nucleotide substitution in exon 2, at position 33 T>C, this resulted in an amino acid change from Tyr to His at codon 17. CONCLUSION: The novel allele is confirmed as a new HLA allele and it was officially named HLA-DRB1*1461 by WHO Nomenclature Committee in May, 2006.

[Investigation on induced expansion of erythroid cells from cord blood CD34(+) cells in vitro].

This study was aimed to investigate a beneficial approach for resolving the deficiency of blood source, preventing the infection resulting from blood transfusion and overcoming the knotty match of patients with rare blood group by using massive expansion of erythroid cells from cord blood CD34(+) cells in vitro. The CD34(+) cells from human cord blood were cultured in serum-free medium supplemented with stem cell factor (SCF), interleukin-3 (IL-3) and erythropoietin (EPO) for 1 week, then expansion and differentiation of CD34(+) cells into erythroid cells were supported by co-culture with human mesenchymal stem cells (MSCs) derived from bone marrow for 2 weeks. The results indicated that after culture for 23 days, the expansion multiple of total cell number reached 2.52 x 10(5), and over 95% of these cells were erythroid cells as compared with less than 1% of myelomonocytic (CD14(+) or CD15(+)) cells and megakaryocytic (CD41(+)) cells. However, the culture system without MSC support was significantly disadvantaged both in expansion ability and ratio of erythroid cells when compared with MSC supporting system. It is concluded that the erythroid cells can be produced from CD34(+) cells in large scale by culturing in the system comprised of cytokine sets and MSC feeders, in which MSCs can support the proliferation and differentiation of erythroid cells.

[A recombination event occurring between HLA-B and DRB1 loci within a Chinese Han family].

OBJECTIVE: To investigate a recombination event occurring between the HLA-B and DRB1 loci in a Chinese family with a leukemia patient. METHODS: HLA class I (-A and -B) low resolution typing was carried out by polymerase chain reaction-sequence specific oligonucleotide, PCR-SSO). HLA class II low resolution typing was performed by PCR-sequence specific primer (PCR-SSP). And HLA class I and II high resolution typing was done by sequencing-based typing (SBT). Then the recombination event was analyzed by family study. RESULTS: The 2 haplotypes of the patient were A*3101-B*1301-DRB1*0701 and A*3303-B*4403-DRB1*1302. His father's 2 haplotypes were A*3001-B*1302-DRB1*0701 and A*3101-B*1301-DRB1*1501. Family study demonstrated that the HLA-A*3101-B*1301 was from one of his father's chromosome and the DRB1*0701 was from the other chromosome of his father. So the result indicated that the recombination event occurred between the HLA-B and -DRB1 loci during meiosis of his father and resulted in a new HLA haplotype that was transferred to the son. CONCLUSION: A HLA-B/DR recombination event occurring between the HLA-B and -DRB1 loci has been found in a Chinese family, which may help further study of the mechanism of HLA recombination.