Production of mature red blood cell by using peripheral blood mononuclear cells / 中国实验血液学杂志

Most protocols for in vitro producing red blood cells (RBC) use the CD34(+) cells or embryonic stem cells from cord blood, bone marrow or peripheral blood as the start materials. This study was purposed to produce the mature RBC in vitro by using peripheral blood mononuclear cells as start material. The peripheral blood mononuclear cells (PBMNC) were isolated from buffy coat after blood leukapheresis, the mature red blood cells (RBC) were prepared by a 4-step culture protocol. The results showed that after culture by inducing with the different sets of cytokines and supporting by mouse MS-5 cell line, the expansion of PBMNC reached about 1000 folds at the end of the culture. About 90% of cultured RBC were enucleated mature cells which had the comparable morphological characteristics with normal RBC. Colony-forming assays showed that this culture system could stimulate the proliferation of progenitors in PBMNC and differentiate into erythroid cells. The structure and function analysis indicated that the mean cell volume of in vitro cultured RBC was 118 ± 4 fl, which was slight larger than that of normal RBC (80-100 fl); the mean cell hemoglobin was 36 ± 1.2 pg, which was slight higher than that of normal RBC (27-31 pg); the maximal deformation index was 0.46, which approachs level of normal RBC; the glucose-6-phosphate dehydrogenase and pyrurvate kinase levels was consistant with young RBC. It is concluded that PBMNC are feasble, convenient and low-cost source for producing cultured RBC and this culture system is suitable to generate the RBC from PBMNC.

To identify a novel HLA-DRB1 allele in Chinese by sequencing / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To identify a novel HLA-DRB1 allele in Chinese.</p><p><b>METHODS</b>A novel HLA-DR allele was detected by PCR-SSP and SBT in a patient with leukemia.</p><p><b>RESULTS</b>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.</p><p><b>CONCLUSION</b>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.</p>

A recombination event occurring between HLA-B and DRB1 loci within a Chinese Han family / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To investigate a recombination event occurring between the HLA-B and DRB1 loci in a Chinese family with a leukemia patient.</p><p><b>METHODS</b>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.</p><p><b>RESULTS</b>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.</p><p><b>CONCLUSION</b>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.</p>

Identification of a novel HLA allele DRB1*1462 by sequence-based typing / 中国实验血液学杂志

This study was to identify a novel HLA-DRB1 allele in Chinese population by nucleotide sequence ana- lysis. The HLA typing of genes was performed by PCR-SSO and PCR-SSP, the ambiguous novel allele was identified by DNA sequence analysis. The results showed that the sequence of this new allele differed from DRB1*140101 by one nucleotide substitution at position 256 in exon 2 (G- > A), resulting in an amino acid change from Ala to Thr at codon 57. In conclusion, this allele is a novel one, which has been officially given the name DRB1*1462 by the WHO nomenclature committee in January 2006.

Investigation on induced expansion of erythroid cells from cord blood CD34(+) cells in vitro / 中国实验血液学杂志

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.