Biomedical subjects

Jian-Pei Fang

Publications and source records attributed to Jian-Pei Fang.

9 recordsLinked to original sources

PubMed2006

[Ex vivo expansion of megakaryocytic progenitors from mobilized human peripheral blood].

This study was aimed to investigate the effect of some culture system composed of various cytokine combinations (TPO, SCF, FL, IL-1, IL-3, IL-6) on ex vivo expansion of megakaryocytic progenitors induced from CD34+ cells of peripheral blood and to seek a optimal cytokine combination and culture time. Mononuclear cells were isolated from mobilized peripheral blood (MPB) by density gradient centrifugation over Ficoll. CD34+ cells were purified by using an immunomagnetic bead separation system. The selected CD34+ cells were seeded in Iscove's modified Kulbecco's medium (IMDM) supplemented with fetal calf serum (FCS) and various kinds of cytokines. After 15 days of culture, the content of CD41+ cells in culture system were determined by flow cytometry, and the number of megakaryocyte colony-forming unit (CFU-MK) was measured simultaneously. The results showed that after definited days of culture, the cytokine combination TPO/FL/IL-6/IL-3 was the most suitable for MPB to obtain high number of MK, and better than any other three groups (P < 0.05). The increase multiple of CD41+ cells was 93.97 +/- 17.27 on day 5 and 131.23 +/- 18.26 on day 10. On day 15, the proportion and the increase multiple of CD41+ cells decreased obviously. The expansion multiples of CFU-MK were 93.33 +/- 10.02 on day 5 and 120.67 +/- 13.01 on day 10, higher than any other groups. It is concluded that TPO/FL/IL-6/IL-3 combination was the best optimal for expansion ex vivo of megakaryocytic progenitors from MPB, and its suitable duration of culture was 10 days; a culture system for expansion ex vivo of megakaryocytic progenitors have been established in this study.

Antigens, CD34↗

PubMed2005

[Construction of pcDNA3.1(+)/A2E eukaryotic expression vector and its expression on K562 cell].

To construct pcDNA3.1(+)/A2E eukaryotic expression vector and obtain a stable expression on HLA-I negative human K562 cell, PCR technique was employed to amplify A2E cDNA from the multi-cistron expression vector pG/A2E carrying HLA-E and HLA-A2 cDNA through internal ribozyme entry site (IRES), the cDNA was subcloned into vector pcDNA3.1(+), thus a eukaryotic expression was constructed and named pcDNA3.1(+)/A2E; then, the recombinant plasmid was transferred into the target cells, followed by screening with G418 and limiting dilution; finally, flow cytometry was adopted to detect HLA-E expression on the target cells. The results showed that HLA-E molecules were successfully expressed on K562 cells transfected with pcDNA3.1(+)/A2E (27.76%) and the expression of HLA-E molecules was not detected on K562 cells transfected with pcDNA3.1(+). It is concluded that the pcDNA 3.1(+)/A2E eukaryotic expression vector was successfully constructed and the HLA-E molecules were expressed on K562 cells. The data presented here would be expected to lay a good basis for the research of the molecular mechanism of HLA-E function and the interaction between HLA-E and the receptor on NK cells, as well as the influence of the expression of HLA-E in vitro on NK cells.

Cloning, Molecular↗

PubMed2005

[Study on the expansion of megakaryocyte progenitors in vitro from cord blood].

This study was aimed to investigate the effect of various cytokines on megakeryocytes expansion in vitro from human cord blood CD34(+) cells in order to establish an optimal culture system for MK expansion. Mononuclear cells were obtained by Ficoll-Hapaque density gradient separation. CD34(+) cells were positively isolated using a CD34 progenitor cell isolation kit. CD34(+) cells were placed into 24 well plates at a concentration of 2 x 10(4) per well. Each well contained 1 ml of IMDM with the present of effective MK cells growth cytokines. Clonogenic potentials of MK progenitor were assayed using a methylcellulose cultures system. The results suggested that four cytokines (IL-3 + IL-6 + TPO + FLT3L) culture system could effectively induce and expand cord blood CD41(+) MK cells. The number of CD41(+) cells expanded 154.67 +/- 32.21-fold on day 7, and 193.23 +/- 25.24-fold on day 14. In conclusion, established expansion system in vitro for MK cells provides experimental foundation for recovery of platelets after cord blood transplantation.

Antigens, CD34↗

PubMed2004

[Analysis on HLA-E polymorphism in Guangdong Han population].

To investigate the distribution of HLA-E alleles and linkage between HLA-E and HLA-A or -B loci in Chinese Han in Guangdong area, HLA-E alleles were detected by using PCR-SSP in 150 unrelated healthy individuals from Guangzhou area; HLA-A, -B antigens typing in 106 individuals was carried out with NIH standard microlymphocytoxic method. Analysis of linkage was performed between HLA-E and HLA-A, -B. The results showed that three alleles of HLA-E could be detected in this population. They are E * 0101, E * 01031, E * 01032, with the frequency of 45.33%, 32.33%, 22.33% respectively. No E * 0102 and E * 0104 could be detected in all of these individuals. The analysis of linkage on two loci between HLA-E and HLA-A or -B showed that no significant difference could be found between expected frequencies and observed frequencies except B15/E * 01032 and A2/E * 01032. In conclusion, the high conservative polymorphism of HLA-E suggests that it's biological characteristic is different from that of classical HLA class Ia molecules.

Alleles↗

PubMed2003

Expression of killer cell inhibitor receptors on immunocompetent cells with relation to graft-versus-host disease after hematopoietic stem cell transplantation.

The study was aimed at the exploration of relationship between T cells expressing killer cell inhibitor receptors (KIR, CD158 and CD94) and graft-versus-host disease (GVHD) after hematopoietic stem cell transplantation. The expression rates of CD158a, CD158b and CD94 on T cells and NK cell were detected by flow cytometry and donor/recipient HLA-Cw was analyzed using PCR after peripheral blood stem cell transplantation (PBSCT) and umbilical cord blood transplantation (UCBT). After both PBSCT and UCBT, the rates of CD3(+)CD158a(+) and CD3(+)CD158b(+) T cells increased, especially the rate of CD8(+)CD158b(+) T cells. In both acute and chronic GVHD groups, the rate of CD3(+)CD158b(+) T cells increased, especially in acute GVHD. The CD94 mainly expressed on CD3(+)CD8(+) T cells. The percentage of the expression of CD94 on CD4(+) and CD8(+) cells after UCBT and PBSCT increased significantly. The expression of KIR in GVHD (early stage of transplantation) increased but the expression of KIR in chronic GVHD (advanced stage of transplantation) decreased. Five patients who HLA-Cw matched had no severe GVHD. In four patients who underwent allo-PBSCT and UCBT from related HLA-matched donors, only 2 patients had no aGVHD. Four patients underwent transplantation from unrelated HLA-matched donors had GVHD. These observations suggested that there is some relationship between GVHD and KIR expression on T cells. CD158b might be an inhibitory molecule of T cell activated at early stage after transplantation. Understanding the mechanism of GVHD with the expression of KIR on T cells, especially those binding the HLA-Cw might shed light on the establishment of the specific immunotolerance for the prevention of GVHD. To pay attention to HLA-Cw typing is very important to reduce GVHD and increase GVL effect in related or unrelated HLA-matched transplantation.

Antigens, CD↗

PubMed2002

[Establishment of a murine model for allogeneic umbilical cord blood transplantation].

This study was undertaken to establish a murine model for unrelated allogeneic umbilical cord blood transplantation (UCBT). The characteristics and percentage of hematopoietic stem/progenitor cells between near-term fetal and neonatal murine peripheral blood (FNPB) and bone marrow (BM) were evaluated by flow cytometry and semisolid methylcellulose culture. BABL/c (H-2(d)) recipient mice conditioned with high dose CTX were transplanted with FNPB form C57BL/6 (H-2(b)) mice and the survival rate, hematopoietic and immunological reconstruction, graft versus host disease (GVHD) and engraftment level were observed. The results showed that the numbers of day 14 CFU-GM and CFU-GEMM in FNPB (176.40 +/- 78.39)% and (141.40 +/- 56.57)%, respectively were much higher than those in BM (75.20 +/- 26.41)% and (68.80 +/- 23.95)%, respectively. Moreover the percentage of Sca-1(+) CD34(+) cell subsets in FNPB (3.63 +/- 1.13)% was also higher than that in BM (1.41 +/- 0.8 7)%. FNPB transplantation improved survival rate and reconstituted hematopoietic and immune function in recipients. There was no evidence of GVHD. Chimeric analysis showed that the proportion of donor cells in BM of recipients was 27.94% at 21 days after transplantation. It was concluded that FNPB contains more hematopoietic stem and progenitor cells with high expansion ability and weak allogeneic immunity, which was similar to human UCB. The murine model for allogeneic UCBT (C57BL/6-->BALB/c) was established successfully.

Animals↗

PubMed2002

[Association of the relationship between HLA-DQB1 alleles and major beta-thalassemia in 42 guangdong Chinese].

To investigate the relationship between HLA-DQB1* alleles and major beta-thalassemia, the HLA-DQB1* loci typing was performed with polymerase chain reaction-sequence specific primer (PCR-SSP) in 42 unrelated (unconsanguineous) patients with major beta-thalassemia and 45 normal control individuals in Guangdong Province. Results showed that the frequency of HLA-DQB1*06 allele in patient group (19.0%) was higher than that in the control group (4.4%) kappa(2) = 8.961, p < 0.01). Our data suggests that HLA-DQB1*06 allele is associated with pathogenesis of the major beta-thalassemia in Guangdong area.

Alleles↗

PubMed2001

[Clinical Study on A Patient with Intestinal Acute Graft-Versus-Host Disease Grade IV after Allo-Peripheral Blood Stem Cell Transplantation]

The purpose of this study was to designed to analyze and explore the diagnosis and treatment on a patient with beta-thalassemia major accompanied with intestinal acute graft-versus-host disease (aGVHD) grade IV after allo-peripheral blood stem cell transplantation (allo-PBSCT). The conditioning regimen was consisted of busulfan (18 mg/kg), cyclophosphamide (200 mg/kg), thiotepa (6 mg/kg) and horse anti-thymocyte globulin (110 mg/kg). The nucleated cells (10.6 x 10(9)/kg) and CD34(+) CD38(-) cells (11.7 x 10(6)/kg) were infused. At +14 days PBSCT, ANC and WBC achieved 0.66 x 10(9)/L and 1.4 x 10(9)/L respectively. The engraftment was successful but the patient depended on transfusion. The range of blood WBC was from 1.4 x 10(9)/L to 2.4 x 10(9)/L. The myelogram showed aplastic anemia. Hematopoiesis was restored continually after the second transfusion of the donor's cells. At +11 days the patient was suffered from cutaneous aGVHD grade II. Diarrhea occurred at +33 day when cutaneous aGVHD had been improved, and exaggerated obviously after the second transfusion at +39 days. The serious diarrhea continued for 2 months and intestinal aGVHD grade IV was finally diagnosed. Our conclusion was that intestinal aGVHD possibly occurred even when cutaneous aGVHD was improved. For a patient with refractory diarrhea after allo-PBSCT, intestinal aGVHD was considered first when the engraftment evidence was definited but peripheral blood cells and bone marrow were abnormal.

Journal Article↗

PubMedDate not supplied

Immunosuppressive treatment of aplastic anemia in Chinese children with antithymocyte globulin and cyclosporine.

Fifty-one children with aplastic anemia (AA) from 1993 to 2004 in the authors' institution were treated by 3 therapies: 11 patients in group 1 received the SSL-6 protocol; 16 patients in group 2 had CsA alone, where the dose of CsA began from 9-12 mg/kg in the initial 2 weeks and tapered off to 5 mg/kg later; 24 patients in group 3 were treated combining rabbit ATG (Pasteur, Merieux) 2.5 mg/kg for 5 days with CsA, which was the same dose as in group 2. The response was 27, 50, and 79%, respectively. The statistical analysis showed that the protocol of intensive immunosuppressive treatment (IST) was the most effective one and CsA was better than that of SSL-6. None of our patients developed clone diseases although the follow-up was as long as more than to 9 years. The data suggest that children with AA should receive IST as first-line therapy in developing countries. Hematopoietic stem cell transplantation (HSCT) is effective treatment for patients with aplastic anemia (AA). However, HSCT is not widely used in China for economic reasons and lack of donors. Immunosuppressive therapy (IST) is now the mainstay treatment for AA. To evaluate the effect of immunosuppressive therapy, combining antithymocyte globulin (ATG) with cyclosporine (CsA), a retrospective study on 51 children with AA from January 1993 to December 2004 treated in the authors' department was performed.

Adolescent↗