A Single Center 25-year Experience in Autologous Peripheral Blood Stem Cell Collection: A Focus on the Collection Efficiency

Background:Harvest of hematopoietic progenitor cells via leukapheresis is being used increasingly for autologous transplantation. Adequate yield of cells per kilogram body weight of recipient is required for a successful engraftment. Collection efficiency(CE)is a useful parameter to assess quality of peripheral blood stem cell (PBSC) collection program. In this study, we report a 25-year experience in a tertiary care Hospital in Italy.Patients and Methods:1,026 consecutives autologous PBSC collection procedure, performed in 763 patients, from January 1996 to December 2020 were retrospectively considered.Data regarding patients, Blood Cells Separators (BCS) , apheresisprocedures and PBSC products were collected in our database. In these 25 years different BCSwere adopted in our Apheresis Unit (AU). In the first period (1996-1999) we used Fresenius Com.Tec, in the central period (2000-2013) we used Cobe Spectra and inthe last period (2014-2020) Spectra Optia. Results:As regards the evaluation of patients before leukapheresis, the most significant data was the increasing number of CD34+ cells. Considering the PBSC collection procedure, there was a progressive increase in the processed blood volume with a shorter apheresis duration. Data related to the PBSC collection demonstrated an increasing CD34+ cell yield and efficiency a raise in CE that was 43% using Fresenius COM-TEC BCS, 49% using Cobe Spectra BCS and 53% using Spectra Optia BCS. . Conclusions:These results were observed considering a 25-year period, thus a great number of factors likely contributing to the observed results, including technological improvement of the instrumentation for leukapheresis,increased experience of the team operating in the Apheresis Unit, improved mobilization protocols, better criteria for patients’ selection. Focusing our attention on CE we observed quite satisfactory results with a median which rose from 43% to 53% with an increase of 10% in the observation period.

In vitro hematopoietic support of human umbilical cord blood cd34+ cells by human skeletal muscle-derived pericytes/perivascular cells / 中国组织工程研究

BACKGROUND: Perivascular cells have been shown to be the precursor cells of mesenchymal stem cells, which regulate the behavior of hematopoietic stem cells and support hematopoiesis through cell-to-cell contact or paracrine effects. Hematopoietic support of human skeletal muscle-derived pericytes/perivascular cells (hMD-PCs) remains to be studied. OBJECTIVE: To identify the biological characteristics of hMD-PCs isolated from human skeletal muscle and to study their supporting effect on umbilical cord blood CD34+ cells in vitro. METHODS: (1) hMD-PCs with phenotype CD146+ CD56-CD34-CD144-CD45- were sorted from human skeletal muscle by enzymatic digestion and multiparameter fluorescence-activated cell sorting, and their biological characteristics were identified. (2) The in vitro culture system of umbilical cord blood CD34+ cells co-cultured with human CD146+ hMD-PCs (experimental group) or with human bone marrow mesenchymal stem cells (positive control group) was established. After 1, 2 and 4 weeks of co-culture, the number of cells, the colony formation ability and immunophenotype were measured and statistically analyzed. RESULTS AND CONCLUSION: (1) CD146+ hMD-PCs were sorted by multiparameter fluorescence-activated cell sorting and the purity was (91.5±1.85)% (n=5). CD146+ hMD-PCs expressed mesenchymal surface markers CD73, CD90, CD105, CD44, and did not express hematopoietic cell and endothelial cell markers CD45, CD34, and CD31. After induced culture, CD146+ hMD-PCs could differentiate into osteoblasts, chondrogenesis, adipocytes and myoblasts. (2) There were no significant differences in the cell number, colony f ormation ability or immunophenotype (CD45+, CD34+ CD33-, CD14+, CD10+/CD19+) between experimental and positive control groups (P > 0.05, n=6). The number of cells in the blank control group without feeder was significantly decreased at 1 week of culture, and there was almost no cell survival at 2 weeks of culture. (3) In summary, CD146+ hMD-PCs, like human bone marrow mesenchymal stem cells, have hematopoietic support capacity in vitro.

Indomethacin enhances the sensitivity of CD34+ cells to imatinib by inhibiting β-catenin in blastic phase chronic myeloid leukemia / 解放军医学杂志

Objective To investigate the effect of indomethacin on cell cycle and apoptosis of CD34+ cells in blastic phase chronic myeloid leukemia, and explore the role of Wnt/β-catenin signal pathway in the molecular mechanism. Methods CD34+cells were sorted from bone marrow samples of blastic phase chronic myeloid leukemia, chronic phase chronic myeloid leukemia and normal cord blood by immune magnetic bead separation. The purity of CD34+ cells was detected by flow cytometry, and cellular morphology was observed by Wright's staining. The protein expression and location of β-catenin and BCR/ABL in CD34+ cells were analyzed by immunofluorescence assay. The changes of β-catenin protein expression in CD34+ cells treated with indomethacin and imatinib were detected by immunofluorescence, the cell apoptosis and cell cycle were observed by Wright's staining and flow cytometry, the mRNA level of c-myc and cyclin D1 was detected by real-time PCR, and the protein expression of BCR/ABL was detected by flow cytometry and immunofluorescence assay. Results CD34+ cells were successfully acquired with purity over 90%. β-catenin and BCR/ABL highly expressed in those CD34+ cells isolated from blastic phase chronic myeloid leukemia and mainly located in cytoplasm. Indomethacin combined with imatinib significantly suppressed the expression of β-catenin, increased the apoptotic rate and arrested the cell cycle in G0/G1 phase of blastic phase CD34+ cells (P<0.05), decreased the mRNA level of c-myc and cyclin D1 and the expression of BCR/ABL in blastic phase CD34+ cells (P<0.05), but threw no significant effect on normal CD34+ cells. Conclusions Indomethacin may significantly arrest the cell cycle and increase the apoptosis rate of CD34+ cells isolated from blastic phase chronic myeloid leukemia. It enhances the sensitivity of leukemia CD34+ cells to imatinib by inhibiting β-catenin expression, suppressing the transcription of c-myc and cyclin D1 and decreasing the expression of BCR/ABL protein.

Investigation on related factors of CD 34+ cells in peripheral blood mobilization effect by G-SCF stimulation / 国际检验医学杂志

Objective To investigate the influence factors of peripheral blood hematopoietic stem cell of healthy donor by granu‐locyte colony‐stimulating‐factors(G‐CSF) mobilization .Methods G‐CSF was subcutaneously injected in 24 cases of healthy donor for mobilizing hematopoietic stem cells .The T lymphocyte subgroup and blood routine data were detected .Results After G‐CSF stimulation ,peripheral blood CD3+ (% ) ,CD3+CD4+ (% ) ,white blood cell (WBC) count and platelet were significantly increased (P<0 .05) .And nucleated cells density of bone ,percentage of CD34+ cells on mobilized 4 ,5 ,6 d had no obvious difference .The correlation analysis showed that gender ,age and weight were negatively correlated with CD 34+cells percentage (P<0 .05) and pos‐itively correlated with WBC count (P<0 .01) .Conclusion Male donor is superior to female donor within a certain range ,the smal‐ler age ,the lighter weight ,the higher WBC count ,the higher the percentage of CD34+ cells in peripheral blood hematopoietic stem cells after G‐CSF mobilization .

Experimental study on repairing spinal cord injury by human umbilical cord blood CD34+cell transplantation at different time points / 天津医药

Objective To investigate the effect of transplantation of human umbilical cord blood CD34+cells on spinal cord injury. Methods CD34+cells were separated from fresh human umbilical cord blood by magnetic cell sorting. Ninety-six female Wistar rats were injured at T10 by IMPACTOR MODEL-Ⅱ, and then randomly assigned to three groups:Cyclo?sporin A (CsA)+Dexamethasone (Dex) treated group (Ⅰ, n=32), local transplantation of cells+CsA+Dex treated group (Ⅱ) at the first day after operation (DAO 1, n=32), local transplantation of cells+CsA+Dex treated group (Ⅲ) at DAO 6 (n=32). BBB locomotor scoring system was used to assess the recovery of the lower limbs. The survival and neural differentiation of transplanted cells at the injury site were observed by double immunofluorescence. The tissue vitality at the injury site was ob?served by 2,3,5-triphenyl-2H-tetrazolium chloride (TTC) staining, and the blood vessel density was observed by infusing mixture of Chinese ink and glutin followed by HE staining. Results BBB score at DAO 8-56 was significantly higher inⅡgroup than that of other two groups (P<0.05). TTC staining showed that the proportion of decreased vitality area was signifi?cantly smaller inⅡgroup than that of other two groups (P<0.01). The result of gelatin ink perfusion showed that the blood vessel density at the injury site was significantly bigger inⅡgroup than that of other two groups (P<0.01). There were more survival transplanted cells inⅡgroup than those of III group (per visual field, 7.51 ± 1.00 vs 5.51 ± 0.89,t=6.051, P<0.01). All the transplanted cells didn’t differentiate into neural cells. Conclusion Human umbilical cord blood CD34+cells can promote the recovery of the lower limbs after spinal cord injury by repairing blood vessels to increase tissue vitality at the in?jury site in rats.

ZNF330 promotes erythroid differentiation / 基础医学与临床

Objective To explore the effects of ZNF330 on erythroid differentiation of K562 cells and underlying mechanism .Methods Realtime PCR was performed to detect the expression of ZNF 330 in K562 cells induced by hemin .After CD34 +cells being infected by the recombination lentivirus ZNF 330-RNAi, Realtime PCR was applied to detect the expression of CD235a and γ-globin.The luciferase report assay was performed to examine if ZNF 330 could act as a trans-acting factor in 293T/17 cells.Co-Immunoprecipitation (Co-IP) was applied in 293T/17 cells to detect the interaction between ZNF 330 and ZNF408 which was involved in mRNA degradation .Results The ex-pression of ZNF330 was up-regulated after hemin treatment .The expression of CD235a andγ-globin decreased after inhibition expression of ZNF 330 had no effect on report gene .Co-Ip in two ways confirmed the direct binding be-tween ZNF330 and ZNF408 .Conclusions ZNF330 can promote erythroid differentiation , and a possible mecha-nism is that ZNF330 inhibits the function of ZNF 408 , a factor that is involved in mRNA degradation , through the interaction between the two proteins .

Recoleccón de células madre en sangre periférica mediante eféresis de grandes volúmenes / Peripheral blood stem cell collection by apheresis

Antecedentes: evaluar el rendimiento y calidad de las recolecciones de células madre en sangre periférica realizadas en el Hospital México entre junio del 2010 hasta junio del 2012, dado que existe una dosis mínima de células CD34+(células progenitoras hematopoyéticas) que definen la posibilidad de éxito del injerto en el trasplante de células madre. Métodos: estudio retrospectivo utilizando los registros de los pacientes y los procedimientos de recolecciones de células madre por aféresis efectuados en el servicio de hematología Hospital México. Para evaluar el rendimiento y calidad de las recolecciones y el efecto de estas en los tiempos de recuperación de los parámetros hematológicos de los pacientes trasplantados entre junio del 2010 y junio del 2012...

Mobilization of Peripheral Blood Stem Cells for Autologous Transplantation in Patients with Hematologic Malignancies

BACKGROUND: Autologous peripheral blood-stem cell transplantation (autoPBSCT) is the treatment of choice for hematologic malignancy, because the technique requires neither general anesthesia nor surgical intervention, amongst many other advantages. Despite these benefits, the risk of hematologic malignancy, as well as the effect of patient age and sex on the prediction of successful collection of autoPBSCT are still unclear. The purpose of this study was to examine whether the hematologic diagnosis of the disease, and age or sex affect the mobilization of CD34+ cells and mononuclear cells. METHODS: We retrospectively investigated 30 (6 multiple myeloma, 11 diffuse large B-cell lymphoma, 8 acute myeloid leukemia, 2 acute lymphoid leukemia, and 3 T-cell lymphoma) patients who underwent autoPBSCT between 2008 and 2011 at Daegu Catholic University Hospital. RESULTS: Patients with multiple myeloma had the highest average of both mononuclear cell (MNC) (2.07+/-0.67x10(8) cells/kg) and CD34+ cell (1.28+/-0.58x10(6) cells/kg) counts. Patients with T-cell lymphoma had both the lowest MNC (1.23+/-0.49x10(8) cells/kg) and CD34+ cell (0.20+/-0.6x10(6) cells/kg) counts. Male patients showed greater collected CD34+ cell counts (0.96+/-1.38x10(6) cells/kg) and MNC counts (1.71+/-0.76x10(8) cells/kg) than the female patients. Patients under the age of 44 had higher collected CD34+ cell counts (0.96+/-1.37x10(6) cells/kg) but lower counts of MNC (1.49+/-0.74x10(8) cells/kg). CONCLUSIONS: The collected MNC and CD34+ cell counts varied between the types of malignancies, and with respect to sex and age. However, only collected MNC counts were significantly different (P<0.05) among the different types of malignancies.

Biochemical and Parasitological Studies on the Effect of hUCB-Selected CD34+ Progenitor/Stem Cells in Mice Infected with Schistosoma mansoni

BACKGROUND AND OBJECTIVES: Placenta and blood that remained in the umbilical cord is routinely available as a discarded tissue after deliveries and it is free of any legal, moral, ethical or religious objections, providing a high number of multipotent CD34+ progenitor and stem cells. Using ex vivo isolated CD34+ cells from human umbilical cord blood (hUCB) have emerged as promising candidates to treat various diseases, including exogenous pathogenic infections. We have expanded to build a rational approach to study the effect of CD34+ cells after damaged liver tissues by the devastating human parasitic flatworm Schistosoma mansoni. METHODS AND RESULTS: Experimental studies were conducted in the Department of Zoology, Faculty of Science and Departments of Parasitology and Physiology, Faculty of Medicine, SCU, Egypt. We have studied the impact of ex vivo preparation of CD34+ cells from hUCB on S. mansoni-induced liver fibrosis de novo, and treated for shorter and longer periods in vivo. Ova count, ALT and albumin were measured at specific time interval and histopathological examination of liver was conducted to confirm the biochemical results. The data obtained were statistically analyzed by ANOVA between groups. It was found that the administration of CD34+ cells have modestly reduced liver damage; reduced the S. mansoni infection associated elevation in serum levels of ALT; significantly improved serum levels of albumin and reduced egg granuloma diameter in the livers. CONCLUSIONS: We demonstrated that CD34+ cells can markedly ameliorated liver fibrosis in vivo and may be beneficial for therapy to recover organ structure and/or function of S. mansoni-infected mice.

Hematologic Recovery after Tandem High-Dose Chemotherapy and Autologous Stem Cell Transplantation in Children with High-Risk Solid Tumors

Although the number of studies using tandem high-dose chemotherapy and autologous stem cell transplantation (HDCT/autoSCT) for the treatment of high-risk pediatric solid tumors has been increasing, documentation of hematologic recovery after tandem HDCT/autoSCT is very limited. For this reason, we retrospectively analyzed the hematologic recovery of 236 children with high-risk solid tumors who underwent tandem HDCT/autoSCT. The median numbers of CD34+ cells transplanted during the first and second HDCT/autoSCT were 4.3 x 10(6)/kg (range 0.6-220.2) and 4.1 x 10(6)/kg (range 0.9-157.6), respectively (P = 0.664). While there was no difference in neutrophil recovery between the first and second HDCT/autoSCT, platelet and RBC recoveries were significantly delayed in the second HDCT/autoSCT (P < 0.001 and P < 0.001, respectively). Delayed recovery in the second HDCT/autoSCT was more prominent when the number of transplanted CD34+ cells was lower, especially if it was < 2 x 10(6)/kg. A lower CD34+ cell count was also associated with increased RBC transfusion requirements and a higher serum ferritin level after tandem HDCT/autoSCT. More CD34+ cells need to be transplanted during the second HDCT/autoSCT in order to achieve the same hematologic recovery as the first HDCT/autoSCT.

Comparación de la actividad de 2 factores estimuladores de colonias de granulocitos de producción nacional: Hebervital y Leukocim / Comparison of the activity of 2 domestic- production factors for granulocyte colonies: Hebervital and Leukocim

En 35 pacientes que debían recibir terapia celular regenerativa se evaluó el efecto estimulante de 2 factores estimuladores de colonias de granulocitos de producción cubana: Hebervital y Leukocim, para la movilización de células madre hematopoyéticas hacia la sangre periférica. Los pacientes se seleccionaron de forma aleatoria en 2 grupos de tratamiento y se les suministró por vía subcutánea una dosis total de 40 mg/kg distribuidos en 4 subdosis de 10 mg/kg administrados cada 12 horas. Se realizó el conteo de las células mononucleares y células CD34+ en sangre periférica mediante citometría de flujo, antes y después de la estimulación. No se encontraron diferencias estadísticamente significativas en los conteos de las células CD34+ obtenidas posestimulación con el Hebervital y el Leukocim

In 35 patients who should have received regenerative cell therapy, it was evaluated the effect of 2 domestic production granulocyte colony stimulating factors: Leukocim and Hebervital for the mobilization of hematopoietic stem cells into peripheral blood. Patients were randomly selected into 2 treatment groups and were given a total dose of 40 mg/kg in 4 sub-doses of 10 mg/kg subcutaneously administered every 12 hours. Count was performed for mononuclear cells and CD34+ cells in peripheral blood by flow cytometry before and after stimulation. There were no statistically significant differences in the counts of CD34+ cells obtained after stimulation with Hebervital and Leukocim

Studies on mechanism of treatment of granulocyte colony-stimulating factor,recombinant human interleukin-11 and recombinant human interleukin-2 on hematopoietic injuries induced by 4.5 Gy γ-rays irradiation in beagles / 中华放射医学与防护杂志

Objective To investigate the mechanism of treatment of granulocyte colony-stimulating factor(rhG-CSF),recombinant human interleukin-11(rhIL-11)and recombinant human interleukin-2 (rhIL-2)on hematopoietic injuries induced by 4.5 Gy60 Coγ-ray irradiation in beagles,and to provide experimental evidence for the clinical treatment of extremely severe myeloid acute radiation sickness (ARS).Methods Sixteen beagle dogs were given 4.5 Gy60 Co γ-ray total body irradiation(TBI),then randomly assigned into irradiation control group,supportive care group or cytokines+supportive care (abbreviated as cytokines)group.In addition to supportive care,rhG-CSF,rhlL-11 and rhIL-2 were administered subcutaneously to treat dogs in cytokines group.The percentage of CD34+cells,cell cycle and apoptosis of nucleated cells in peripheral blood were examined by Flow cytometry.Results After 4.5 Gy 60 Co γ-ray irradiation,the CD34+cells in peripheral blood declined obviously(61.3%and 52.1% of baseline for irradiation control and supportive care group separately).The cell proportion of nucleated cells in Go/G1 phase was increased notably(99.27% and 99.49% respectively).The rate of apoptosis(26.93% and 21.29% separately)and necrosis(3.27% and 4.14%,respectively)of nucleated cells were elevated significantly when compared with values before irradiation(P<0.05) 1 d post irradiation.When beagles were treated with cytokines and supportive care,the CD34+cells in peripheral blood were markedly increased(135.6% of baseline).The effect of G0/G1 phase blockage of nucleated cells became more serious(99.71%).The rate of apoptosis(5.66%)and necrosis(1.60%)of nucleated cells were significantly lower than that of irradiation control and supportive care groups 1 d after exposure.Conclusions Cytokines maybe mobilize CD34+cells in bone marrow to peripheral blood,indce cell cycle block at G0/G1 phase and reduce apoptosis,and eventually cure hematopoieticinjuries induced by irradiation.

Effects of Mesenchymal Stem Cells Feeder Layer, Culture Sera and Freeze-thaw Lysates on Expansion and Differentiation of Cord Blood CD34~+ Cells in Vitro / 中山大学学报(医学科学版)

[Objective]To investigate the effects of mesenchymal stem cells (MSC) feeder layer, culture sere and freeze-thaw lysates on expansion and differentiation of cord blood CD34~+ cells in vitro. [Methods] MSC were isolated from human bone marrow and cultured until the third passage. Sera were obtained from the cultured MSC, and freeze-thaw lysates were obtained by repeated freeze-thaw procedures. Cord blood CD34~+ cells were isolated by magnetic cell separation system, and were co-cultured with the MSC feeder layer, culture sera, freeze-thaw lysates and hematopeietic growth factors (HGFs), respectively. The nucleated cells, CD34~+ cells, CD34~+CD38~- cells, CD41~+ cells and CD3~+ cells in the above culture system were detected by flow cytometry on day 6 and day 12. [Results] ①MSC feeder layer had a strong effect on nucleated cells, CD34~+,CD34~+CD38~- cells expansion. The MSC sera and freeze-thaw lysates had similar effect on cell expansion, but the effect was weaker than that of feeder layer (P<0.05). ② Both MSC sera and feeder layer inhibited cord blood CD34~+ cells differentiation toward CD3~+ cells or CD19~+ cells, and no significant differences were found between these two groups (P>0.05). ③ Both MSC sera and feeder layer promoted cord blood CD34~+ cells differentiation toward CD41~+ cells, and the effect was stronger in the feeder layer than that of the sera (P<0.05). ④ Freeze-thaw lysates had no effect on cell expansion and differentiation, and were similar with that of HGFs (P>0.05). [Conclusions] The MSC sera have positive effects on expansion of cord blood CD34~+ and CD34~+CD38~- cells, moreover they have the ability of promoting cord blood CD34~+ cells differentiation toward CD41~+ cells.

Factors influencing engraftment in hematological patients after human leukocyte antigen matched sibling allogeneic blood and marrow transplantation / 中华内科杂志

theengraftment of neutrophil and platelet after HLA-matched sibling allogeneic blood and marrow transplantation.Duration from diagnosis to trarmplantation was another factor influencing engraftment of platelet.

Study on optimized conditions for cord blood CD+34 cells amplification in vitro / 白血病·淋巴瘤

Objective To set up the optimized conditions,the amplification of eord blood CDCD+34 cells in vitro were analyzed by comparing the conditions such as different feeder-layers, stimulating-factors or purity/contents of those cells. Methods The cord blood CDCD+34 cells proliferation was analyzed by the methods of MTT, cell counting, and flow eytometer. The amplification and clone-forming ability of cord blood CDCD+34 cells were detected under optimized condition. Resulits The growth rates of cord blood CD+34 cell under optimized conditions(10 times) were significantly higher than that of the control(2.8 times) (P ＜0.01), and the cloneforming ability of cord blood CDCD+34 cells under optimized conditions(CFU-C 36.67±6.11) were also better than that of the control(CFU-C 16.33±1.53) (P ＜0.01). Conclusion The cord blood CD+34 cells proliferation can be promoted in the co-cultured system, and the character of the stem cells were kept well in that system.

Hemopoietic progenitor cell identification in fetal and adult blood / Célula progenitora hamatopoética: identificação em sangue fetal e de adulto

Hemopoietic progenitor cells give rise to all cellular elements of the blood and are of importance as a potential source of cells used for correction of various pathological conditions. The main objective of this study was to identify and quantitative hemopoietic progenitor cell in antenatal fetal blood, in cord blood at the time of delivery and in adult blood, using monoclonal antibodies to surface markers and flow cytometry. CD34+ cells, most of them probably representing progenitor cells, were detected in prenatal fetal blood as early as the 17th week of gestation. The proportion of these cells showed a tendency to decrease as the pregnancy progressed. Within the population of CD34+ cells, a relatively low proportion (less than 1 percent) were negative for the surface marker CD33 or HLA-Dr, indicating a population of primitive stem cells, i.e., progenitor cells no committed to a specific lineage. On the contrary, another group coexpressed CD33 or HLA-Dr, being more mature progenitor cells already committed to differentiate along a specific lineage. The percentage of CD34+ obtained in blood of adult patients after mobilization with chemotherapeutic agents and growth factors showed an average value of 2.7± 3.1 percent. The percentage of CD34+ in the apheresis products of various patients varied from 0.58 to 1.48. In some cases the cells were reinfused in the patient with good results. Our findings are in agreement with previous studies suggesting that CD34+ stem cells is a heterogeneous population, with each subset having variable degree o commitment to differentiate toward a specific cell lineage.

As células progenitoras hematopoéticas são as responsáveis pela produção de todos os elementos do sangue e são as potenciais fontes de células usadas para o tratamento de várias condições patológicas. O principal objetivo deste trabalho foi identificar e quantificar as células progenitoras hematopoiéticas no sangue fetal do período pré-natal, no sangue de cordão umbilical no momento do parto e no sangue do adulto, usando anticorpos monoclonais para marcadores de superfície e citometria de fluxo. As células CD34+ na maioria das vezes representam células progenitoras e foram detectadas no sangue fetal pré-natal tão precoce como na 17ª semana de gestação. A proporção destas células mostrou a tendência de diminuir durante a progressão da gestação. Dentro da população de células CD34+, uma proporção relativamente pequena (menos de 1 por cento) foi negativa para os marcadores de superfície CD33 ou HLA-Dr, indicando uma população de células primitivas, isto é, células progenitoras não comissionadas com uma linhagem específica. Ao contrário, outro grupo co-expressa CD33 ou HLA-Dr, sendo progenitores celulares mais maduros já comprometidos com linhagens específicas. A porcentagem de CD34+ obtida no sangue de adultos após mobilização com agentes quimioterápicos e fator de crescimento mostrou uma média de 2.7+/-3.1 por cento. 0 por cento de CD34+ no produto aferético de vários pacientes variou de 0.58 a 1.48. Em alguns casos as células foram infundidas nos pacientes com bons resultados. Nossos achados estão de acordo com estudos prévios sugerindo que células CD34+ sejam uma população heterogênea com cada subgrupo apresentando graus de comprometimento com diferentes linhagens específicas.

Expression of Peripheral Blood CD34++ Cells before and after Hyperbaric Oxygen Treatment in Patients with Brain Traumatic Injury / 中国康复理论与实践

@#Objective To investigate the change of peripheral blood CD34+ cells and white blood cells(WBC)count in the patients with brain traumatic injury in convalescence stage during hyperbaric oxygen(HBO)treatment.Methods The peripheral blood CD34+ cells and WBC count were measured in 11 patients with brain traumatic injury in convalescence stage who accepted HBO treatment before and 1,2,3,4,and 5 weeks during the treatment.Other 11 persons were measured once as controls.Results Compared with the controls,the number of CD34+ cells showed normal in the 1st and the 3rd week,but increased in the 2nd and the 5th week in the HBO group(P<0.05).It was increased in the 5th week compared with that in the 1st and the 3rd week(P<0.05)in the treatment group.The number of WBC showed no significance and the patients' symptoms improved during the HBO treatment.Conclusion HBO can mobilize bone marrow derived stem cells marked by CD34,which may participate in the restoration of brain traumatic injury in convalescence stage.

Immunal reconstitution after autologous purified CD+34 cells transplantation in patients with systemic lupus erythematosus / 中华内科杂志

Objective To investigate the variation of immune index in patients with systemic lupus erythematosus (SLE) treated with autologous purified CD+34 cells transplantation and to clarify the relationship with pathogenesis and prognosis. Methods Flow cytometry (FCM) and enzyme linked immunosorbent assay(ELISA) were used to test lymphocyte subsets, C3, C4, CH50, autoantibodies and immunoglobulin for 18 cases of SLE before and after transplantation. Results The results showed that the ratio of all the T cell subsets reduced obviously in early postgraft and recovered gradually in 1 to 3 months after transplantation except CD45 RO+CD+4 cells. The levels of serum C3, C4, CH50 increased significantly after transplantation. No case relapsed within one year after transplantation, but 2 patients relapsed one year after transplantation. The levels of the indexes in the patients with relapse were significantly lower than those in the patients with persistent remission, including C4 in the entire course, CH50 in the 3rd and 12th month after transplantation and CD45 RA+ CD+8 cells in the 6th month after transplantation. However, the ratio of CD45 RO+ CD+4 cells in the first month after transplantation in the patients with relapse was higher than that in the patients with persistent remission. Conclusion Autologous purified CD+34 cells transplantation is effective for treating SLE. Survey of immune indexes before and after transplantation is important to investigate the pathogenesis of SLE. Moreover, these immune indexes can be used to predict therapeutic efficacy of SLE.

Ex Vivo Generation of RBCs from CD34+ Cells in Human Umbilical Cord Blood and Expression Profile Analysis Using Microarray / 대한진단검사의학회지

BACKGROUND: In this study, we attempted to generate RBCs from CD34+ cells in cord blood using a 3-step culture protocol and also evaluated a change in immunophenotypic characteristics and expression profile according to erythropoietin (EPO) concentrations and culture duration. METHODS: Using mini-MACS columns, CD34+ cells were isolated from cord blood. The culture procedure comprised three steps. For each step, cells were cultured sequentially for 7 days in a serum free liquid medium with a specific combination of growth factors for 21 days. [1st step: Flt3-ligand (Flt3-L), thrombopoietin and stem cell factor (SCF); 2nd step: IGF-1, SCF and EPO; and 3rd step: IGF-1 and EPO] To evaluate the effect of EPO on proliferation and differentiation, cells were cultured with different EPO concentrations (0, 3, 10 & 20 U/mL). Cell count and morphology were monitored during the culture. For phenotyping, antibodies to CD34, CD38, CD45 and glycophorin A (GPA) were used. The expression profile of cultured cells was analyzed by 17, 000-gene microarray analysis. RESULTS: As EPO concentration increased, cell expansion was also increased, showing a maximum expansion at 20 U/mL. The cell population showed a gradual decrease in expression of CD34 and CD45, whereas the expression of GPA was not prominent in any conditions. However, we observed increased expression in some genes associated with erythropoiesis (e.g. glycophorin A, rhesus blood group CcEe antigens). CONCLUSIONS: This study shows that erythropoietin enhances the proliferation of hematopoietic progenitor cells. Our culture system did not achieve pure production of RBCs, but induced expression changes that indicated erythroid differentiation.

Ex vivo Expansion and Clonal Maintenance of CD34+ Selected Cells from Cord Blood and Peripheral Blood / 소아과

PURPOSE: Because of the unavailability of marrow transplantation, umbilical cord blood (CB) is increasingly being used. We evaluated the potential of ex vivo expansion and clonality in CD34+ cells separated from cord blood source and mobilized peripheral blood (PB) in a serum-free media. METHODS: The CD34+ cells, selected from CB and mobilized PB, were expanded with hematopoietic growth factors. They were then cultured for burst-forming units of erythrocytes (BFU-E), colony-forming units of granulocytes and monocytes (CFU-GM) and colony-forming units of megakaryocytes (CFU-Mk) at culture days 0, day 4, day 7, and day 14 with various growth factors. RESULTS: The CB-selected CD34+ cells showed significantly higher total cell expansion than those from the PB at day 7 (2 fold increase than PB). The CB-selected CD34+ cells produced more BFU-E colonies than did the PB on culture at days 7 and at day 14. Also, the CB-selected CD34+ cells produced more CFU-Mk colonies than did the PB on culture at day 4 and at day 7. CONCLUSION: The ex vivo expansion of the CB cells may be promising in producing total cellular expansion, CFU-Mk and BFU-E compared with PB for 7 to 14 days. The growth factors combination including megakaryocyte growth and development, flt3-ligand and interleukin-3 showed more expansion in the view of total cells and clonal maintenance compared with less combination.