scholarly journals The Effect of Vascular Graft and Human Umbilical Cord Blood-Derived CD34+ Stem Cell on Peripheral Nerve Healing

2018 ◽  
Vol 6 (11) ◽  
pp. 1946-1952
Author(s):  
Ali Yilmaz ◽  
Abdullah Topcu ◽  
Cagdas Erdogan ◽  
Barbaros Sahin ◽  
Gulcin Abban ◽  
...  
Keyword(s):  
Stem Cell ◽  
Electron Microscope ◽  
Sciatic Nerve ◽  
Peripheral Nerve ◽  
Cord Blood ◽  
Vascular Graft ◽  
Nerve Damage ◽  
Cd34 Cells ◽  
The Right ◽  
Synthetic Vascular Graft

AIM: There are many trials concerning peripheral nerve damage causes and treatment options. Unfortunately, nerve damage is still a major problem regarding health, social and economic issues. On this study, we used vascular graft and human cord blood derived stem cells to find an alternative treatment solution to this problem. MATERIAL AND METHODS: We used 21 female Wistar rats on our study. They were anesthetized with ketamine and we studied right hind limbs. On Group 1, we did a full layer cut on the right sciatic nerve. On Group 2, we did a full layer cut on the right sciatic nerve, and we covered synthetic vascular graft on cut area. On Group 3, we did a full layer cut on right sciatic nerve, and we covered the area with stem cell applied vascular graft. RESULTS: At the end of postoperative 8. weeks, we performed EMG on the rats. When we compared healthy and degenerated areas as a result of EMG, we found significant amplitude differences between the groups on healthy areas whereas there was no significant difference on degenerated  areas between the groups. Then we re-opened the operated area again to reveal the sciatic nerve cut area, and we performed electron microscope evaluation. On the stem cell group, we observed that both the axon and the myelin sheet prevented degeneration. CONCLUSION: This study is a first on using synthetic vascular graft and cord blood derived CD34+ cells in peripheral nerve degeneration. On  the  tissues  that  were  examined  with  electron  microscope,  we  observed  that  CD34+  cells  prevented  both  axonal  and  myelin  sheath degeneration. Nerve tissue showed similar results to the control group, and the damage was minimal.

scholarly journals High Integrity and Fidelity of Long-Term Cryopreserved Umbilical Cord Blood for Transplantation

2021 ◽  
Vol 10 (2) ◽  
pp. 293
Author(s):  
Gee-Hye Kim ◽  
Jihye Kwak ◽  
Sung Hee Kim ◽  
Hee Jung Kim ◽  
Hye Kyung Hong ◽  
...  
Keyword(s):  
Stem Cell ◽  
Cord Blood ◽  
Umbilical Cord ◽  
Umbilical Cord Blood ◽  
Clinical Applications ◽  
Hematopoietic Stem ◽  
Cd34 Cells ◽  
Hsc Transplantation

Umbilical cord blood (UCB) is used as a source of donor cells for hematopoietic stem cell (HSC) transplantation. The success of transplantation is dependent on the quality of cord blood (CB) units for maximizing the chance of engraftment. Improved outcomes following transplantation are associated with certain factors of cryopreserved CB units: total volume and total nucleated cell (TNC) count, mononuclear cell (MNC) count, and CD34+ cell count. The role of the storage period of CB units in determining the viability and counts of cells is less clear and is related to the quality of cryopreserved CB units. Herein, we demonstrate the recovery of viable TNCs and CD34+ cells, as well as the MNC viability in 20-year-old cryopreserved CB units in a CB bank (MEDIPOST Co., Ltd., Seongnam-si, Gyeonggi-do, Korea). In addition, cell populations in CB units were evaluated for future clinical applications. The stable recovery rate of the viability of cryopreserved CB that had been stored for up to 20 years suggested the possibility of uses of the long-term cryopreservation of CB units. Similar relationships were observed in the recovery of TNCs and CD34+ cells in units of cryopreserved and fresh CB. The high-viability recovery of long-term cryopreserved CB suggests that successful hematopoietic stem cell (HSC) transplantation and other clinical applications, which are suitable for treating incurable diseases, may be performed regardless of long-term storage.

Clinical-Scale Cultures of Cord Blood CD34+Cells to Amplify Committed Progenitors and Maintain Stem Cell Activity

2011 ◽  
Vol 20 (9) ◽  
pp. 1453-1464 ◽  
Author(s):  
Zoran Ivanovic ◽  
Pascale Duchez ◽  
Jean Chevaleyre ◽  
Marija Vlaski ◽  
Xavier Lafarge ◽  
...  
Keyword(s):  
Stem Cell ◽  
Cord Blood ◽  
Cell Activity ◽  
Clinical Scale ◽  
Cd34 Cells ◽  
Stem Cell Activity ◽  
Cord Blood Cd34

scholarly journals Histopathological analysis of gangliosides use in peripheral nerve regeneration after axonotmesis in rats

2008 ◽  
Vol 23 (4) ◽  
pp. 364-371 ◽  
Author(s):  
Camila Maria Beder Ribeiro ◽  
Belmiro Cavalcanti do Egito Vasconcelos ◽  
Joaquim Celestino da Silva Neto ◽  
Valdemiro Amaro da Silva Júnior ◽  
Nancy Gurgel Figueiredo
Keyword(s):  
Sciatic Nerve ◽  
Peripheral Nerve ◽  
Nerve Regeneration ◽  
Schwann Cells ◽  
Peripheral Nerve Regeneration ◽  
Cell Activity ◽  
Control Group ◽  
Histopathological Analysis ◽  
Male Wistar Rats ◽  
The Right

PURPOSE: To analyze the action of gangliosides in peripheral nerve regeneration in the sciatic nerve of the rat. METHODS: The sample was composed of 96 male Wistar rats. The animals were anaesthetized and, after identification of the anaesthesic plane, an incision was made in the posterior region of the thigh, followed by skin and muscle divulsion. The right sciatic nerve was isolated and compressed for 2 minutes. Continuous suture of the skin was performed. The animals were randomly divided into two groups: the experimental group (EG), which received subcutaneous injection of gangliosides, and the control group (CG), which received saline solution (0.9%) to mimic the effects of drug administration. RESULTS: No differences were observed between the experimental and control groups evaluated on the eighth day of observation. At 15 and 30 days the EG showed an decrease in Schwann cell activity and an apparent improvement in fibre organization; at 60 days, there was a slight presence of Schwann cells in the endoneural space and the fibres were organized, indicating nerve regeneration. At 15 and 30 days, the level of cell reaction in the CG had diminished, but there were many cells with cytoplasm in activity and in mitosis; at 60 days, hyperplastic Schwann cells and mitotic activity were again observed, as well as nerve regeneration, but to a lesser extent than in the EG. CONCLUSION: The administration of exogenous gangliosides seems to improve nerve regeneration.

scholarly journals A Naive NK Cell Repertoire in the Circulation of Haploidentical Stem Cell Donors Pre Mobilization Predicts Rejection of Cord Myeloid Cells in Patients Undergoing Combined Haploidentical and Unrelated Cord Blood HSCT

Blood ◽  
2016 ◽  
Vol 128 (22) ◽  
pp. 2199-2199
Author(s):  
Mattias Carlsten ◽  
Robert N. Reger ◽  
Ritesh Kotecha ◽  
Enkhtsetseg Purev ◽  
Xin Tian ◽  
...  
Keyword(s):  
Stem Cell ◽  
Cord Blood ◽  
Nk Cells ◽  
Nk Cell ◽  
Stem Cell Mobilization ◽  
Cell Repertoire ◽  
Post Transplant ◽  
Cd34 Cells ◽  
Cell Mobilization ◽  
Unrelated Cord Blood

Abstract Background: For patients (pts) with severe aplastic anemia (SAA) lacking an HLA identical donor, outcomes of hematopoietic stem cell transplantation (HSCT) using unrelated cord blood (UCB) units or haplo-identical donors (HDs) have historically been associated with high graft failure rates and poor survival. In an ongoing clinical trial at the NHLBI, we have observed excellent engraftment (100%) and survival (91%) in SAA pts (n=27) receiving a transplant that co-infuses a single UCB unit with CD34-selectedCD3-depletedcells from a haplo-identical relative. Although cord myeloid engraftment(defined as cord ANC >500/μL) occurred at<day 100 in the majority of pts, a significant fraction of pts had delayed (>day 100) or no cord myeloid engraftment. In this analysis, we investigated factors that may have impeded cord myeloid engraftment following UCB/HD transplantation. Methods: Flow-based NK cell phenotyping using a BD Fortessa II instrument was performed on blood obtained pre-transplant from HDs used for the first 18 SAA pts undergoing UCB/HD transplantation. Lineage specific chimerism was measured by PCR of microsatellites (PowerPlex 16 HS Systemkit/Promega) using DNA from flow sorted cells (BD FACSAria) collected multiple time points post-transplant.KIR-ligand incompatibility in the HD vs UCB directionwas defined using high-resolution HLA typing. Results: 13/18 (72%) pts had cord myeloid engraftment before day 100 while 5/18 (28%) had delayed or no cord myeloid engraftment. Remarkably, delayed or no cord myeloid engraftment occurred exclusively in pts transplanted with KIR-ligand incompatibility in the HD vs UCB direction (n=9) (Figure 1A). In contrast, all 9 pts transplanted with KIR-ligand compatibility in the HD vs UCB direction achieved cord myeloid engraftment by ²day 48 (median day 35) post-transplant. Chimerism analysis performed on blood obtained 30+ days post-transplant revealed NK cell chimerism was ³ 90% cord in origin in all 9 pts transplanted with KIR-ligand compatible grafts. In contrast, amongst the 9 pts receiving a KIR-ligand incompatible transplant, NK cell chimerism was predominantly HD in origin with only a minor fraction of cord NK cells detected 30-200 days post-transplant (Figure 1B). Predominant HD NK cell chimerism in pts receiving a KIR-ligand incompatible transplant was associated with lower degrees of cord myeloid chimerism compared to KIR-ligand compatible recipients. Further analysis of the KIR-ligand incompatible cohort revealed distinct heterogeneity in the time to cord myeloid engraftment (Figure 1A). Although delayed or no cord myeloid engraftment was observed in 5/9 recipients of KIR-ligand incompatible transplants, 4/9 pts in this cohort had cord engraftment at a similar time as pts transplanted with KIR-ligand compatible grafts (median 35 vs. 35 days). This variability in time to cord myeloid engraftment was not associated with stem cell dose, degree of HD NK cell chimerism, type of KIR-ligand incompatibility or KIR haplotype. However, we observed a strong correlation between the proportion of naive NK cells in circulation of HDs before stem cell mobilization with delayed or no myeloid cord engraftment (Figure 1C). With the exception of one patient who had failed HD engraftment, only transplants of CD34+ cells from HDs with a predominantly naive NK cell repertoire, expressing high frequencies of the NKG2A receptor concomitant with low frequencies of NKG2C, Lir-1 and CD57 resulted in delayed or no cord myeloid engraftment (p<0.05). Conclusions: Our study provides the first evidence that NK cells from engrafting CD34+ cells from selected HDs can significantly delay or completely inhibit cord myeloid engraftment following UCB/HD transplantation. Suppression of cord hematopoiesis appears to be restricted to NK cells originating from HDs withHD vs UCB KIR-ligand incompatibility who have a large naive NK cell repertoire in their circulation prior to stem cell mobilization. The myelosuppressive effects of these NK cells are consistent with recentlypublished data showing a naive NK cell repertoire in stem cell donors predicts a reduced risk of AML relapse post-allogeneic HSCT.Further studies defining the mechanisms through which naive NK cells suppress cord hematopoiesis followingUCB/HDtransplantation could shed insights into methods to optimize NK cell mediated graft-vs-leukemia followingallogeneicHSCT of myeloid leukemias. Disclosures No relevant conflicts of interest to declare.

Ex-Vivo Generation and Expansion of Both Lymphoid and Myeloid Lineages from Human Cord Blood (CB) HSC Using a Serum-Free Human Mesenchymal Stem Cell Based Culture System.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 2888-2888
Author(s):  
Ana Frias ◽  
Christopher D. Porada ◽  
Kirsten B. Crapnell ◽  
Joaquim M.S. Cabral ◽  
Esmail D. Zanjani ◽  
...  
Keyword(s):  
Stem Cell ◽  
Cord Blood ◽  
Culture System ◽  
Ex Vivo ◽  
Cell Number ◽  
Human Cord Blood ◽  
Serum Free ◽  
Gm Csf ◽  
Cd34 Cells

Abstract The in vitro culture of a hematopoietic stem cell (HSC) graft with either media containing animal-derived components or a feeder layer with ill-defined pathogenic potential such as xenogeneic cell lines or cells modified by viral transformation poses risks that concern scientists and regulatory agencies. In the present studies, we avoided these risks by evaluating the ability of a human stromal-based serum free culture system (hu-ST) to support the ex-vivo expansion/maintenance of human CB HSC. CB CD34+ enriched cells were cultured in serum free medium in the presence of hu-ST with SCF, bFGF, LIF and Flt-3, and the cultures were analyzed for expansion, phenotype and clonogenic ability. We have previously reported the ability of this culture system to allow the successful expansion/maintenance of HSC along the myeloid pathway. In the present study, we investigated whether we could further develop this culture system to simultaneously expand myelopoiesis and lymphopoiesis in vitro. To this end, cord blood CD34+ cells were cultured for a total of 28 days and analyzed every 3 days for expansion and phenotype. There was a progressive increase in CD34 cell number with time in culture. The differentiative profile was primarily shifted towards the myeloid lineage with the presence of CD33, CD15, and CD14. However, a significant number of CD7+ cells were also generated. At week 2 of culture, we observed that 30% of the cells in the culture were CD7 positive. These CD7+CD2-CD3-CD5-CD56-CD16-CD34- cells were then sorted and either plated on top of new irradiated hu-ST layers in the presence of SCF, FLT-3, IL-7, IL-2, and IL-15, or cultured with IL-4, GM-CSF, and FLT-3 in the absence of stroma. Both of these cultures were maintained for an additional 2 weeks. In both sets of cultures, further expansion in the total cell number occurred with the time in culture, and by the end of the week 2, we observed that 25.3±4.18% of the cells had become CD56+ CD3-, a phenotype consistent with that of NK cells. Furthermore, cytotoxicity assays were performed and showed cytotoxic activity that increased in an E:T ratio-dependent fashion. 38.6% of the CD7+ cells grown in the presence of IL-4, GM-CSF, and FLT-3 became CD123+CD11c-, a phenotype characteristic of nonactivated dendritic cells, while 7.3–12.1% adopted an activitated dendritic cell phenotype CD83+CD1a+. In summary, we developed an in vitro culture system that reproducibly allows the effective ex vivo expansion of human cord blood HSCs while maintaining the capability of generating both myeloid and lymphoid hematopoiesis in vitro.

Impaired Alloantigen Presenting Activity of Cord Blood Nucleated Cells.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2203-2203
Author(s):  
Sandeep Chunduri ◽  
Dolores Mahmud ◽  
Javaneh Abbasian ◽  
Damiano Rondelli
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
T Cell ◽  
Cord Blood ◽  
Mononuclear Cells ◽  
Ex Vivo ◽  
Solid Organ Transplantation ◽  
T Cell Response ◽  
Solid Organ ◽  
Cd34 Cells

Abstract Transplantation of HLA-mismatched cord blood (CB) nucleated cells has limited risk of severe acute graft-versus-host disease and graft rejection. This may depend on naïve T cells not yet exposed to many antigens and on immature antigen-presenting cells (APC) not delivering appropriate signals to allogeneic T cells. In order to test the APC activity of human circulating CB cells in-vitro, we initially used irradiated CB mononuclear cells (MNC) or immunomagnetically selected CD34+ cells, CD133+ cells, or CD14+ monocytes to stimulate the proliferative response of incompatible blood T cells in mixed leukocyte culture (MLC). CB MNC failed to induce allogeneic T cell proliferation, while CD34+ and CD133+ progenitors or CD14+ monocytes induced potent T cell alloresponses. Nevertheless, since allogeneic T cell response was not restored after depletion of CD3+ cells in the CB, nor the add-back of irradiated CB MNC to CD34+ or CD14+ stimulators inhibited allo-T cells, a direct suppressive effect of CB MNC was excluded. Allogeneic peripheral blood cytotoxic T-lymphocyte (CTL) responses were not induced after 7 days of stimulation with irradiated CB MNC, although after 4 weekly rechallenges with CB MNC, on average a 23% lysis of antigen-specific CB PHA-blasts was observed at the highest effector:target ratio (50:1). To test the tolerogenic potential of CB MNC, T cells initially exposed to CB MNC were rechallenged in secondary MLC with CB MNC, or CD34+ cells, or monocyte-derived dendritic cells (Mo-DC) generated in liquid culture with GM-CSF and IL-4. Allogeneic T cells were still unresponsive upon rechallenge with CB MNC, but proliferated upon 3 days of restimulation with CD34+ cells or Mo-DC from the same CB. Surprisingly, the supernatant of these latter MLCs did inhibit completely a 3rd party MLC. Instead, the supernatant of blood T cells that had been activated by CB CD34+ cells or Mo-DC both in primary and secondary MLC did not. These results show an impaired allo-APC activity of CB MNC but not CB CD34+ cells, and suggest that T cells releasing immunosuppressive cytokines may be activated by CB MNC and then expanded by a second more potent stimulation with professional APC. This hypothesis could explain the sustained engraftment of HLA-mismatched CB stem cell transplants in humans. Based on these results, the in-vivo or ex-vivo downregulation of T cell alloreactivity induced by CB MNC will be tested in experimental models of stem cell, as well as solid organ transplantation.

Quantitative Monitoring of Lineage-Specific Chimerism after Dual Hematopoietic Stem Cell Transplantation (Umbilical Cord Blood with Co-Infusion of Third Party Donor CD34+ Cells).

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 5387-5387
Author(s):  
Noemi Sanchez-Hernandez ◽  
Cintia Manzano ◽  
Diana Barroso ◽  
Gustavo Iglesias ◽  
Pascual Balsalobre ◽  
...  
Keyword(s):  
T Cells ◽  
Stem Cell ◽  
Stem Cell Transplantation ◽  
Cord Blood ◽  
Umbilical Cord ◽  
Umbilical Cord Blood ◽  
Graft Rejection ◽  
Third Party ◽  
Qrt Pcr ◽  
Cd34 Cells

Abstract Background: Allogeneic umbilical cord blood (UCB) stem cell transplantation (SCT) shows some advantages (HLA matching requirements or availability) respect to SCT using other sources of matched unrelated donor (MUD) stem cells. However, it is correlated with slower engraftment, increasing risk of infections and early mortality. It has been recently shown that co-infusion of third party donor (TPD) CD34+ cells (dual SCT) is useful to speed up engraftment. Objective: To evaluate the usefulness of lineage-specific chimerism quantification in the management of this transplant setting. Patients and methods: 8 dual SCT (Tables 1, 2) in 7 patients (1 CML-BC, 2 AML-M2, 1 AML-M4, 1 ALL-Ph+, 1 biphen. ALL, 1 NHL). Chimerism was analyzed by STR-PCR (AmpFlSTR SGM Plus, Applied Biosystems; sensitivity 1%) and quantitative real-time PCR (qrt-PCR) of null alleles and insertion/deletion polymorphisms (Light Cycler, Roche; sensitivity 0,01%). Peripheral blood (PB) and leukocyte lineages (T cells, CD3+, and myeloid cells, CD15+), isolated by positive selection using automated immunomagnetic technology (AutoMACS, Miltenyi Biotec), were analyzed weekly. Bone marrow (BM) was analyzed at days +30, +100, +180 and +365). Results: 7/8 cases showed initially a high proportion of TPD cells in PB which were progressively replaced by UCB cells. UCB complete chimerism (UCB-CC, absence of recipient or TPD cells even in qrt-PCR assays) was acquired in a median of 22.5 days (range 18–39). In one patient, fully HLA-mismatched with the TPD, no TPD cells were observed after dual SCT. 4/8 cases showed recipient cells in PB after dual SCT during a median period of 12 days (range 4–18 days). In 3/8 cases, recipient cells were found after CC had been acquired, which allowed early diagnosis of 1 graft rejection and 2 relapses. T cells (CD3+) are mainly of UCB origin early after dual SCT and reach UCB-CC a median of 7 days (range 0–21) before PB. However, myeloid cells (CD15+) derive primarily from the TPD and reach CC together with PB. TPD cellularity favoured early engraftment (before UCB-CC took place) in 4 cases. In this context, only one important infectious complication (hepatosplenic tuberculosis) was observed, which resolved with the appropriate treatment. Conclusions: Lineage-specific chimerism quantification allowed a close monitoring of the dynamics of engraftment of cells of both donors which is of key importance in this SCT setting. Moreover, lineage-specific chimerism analysis was useful to diagnose one graft rejection and two relapses (the patient with NHL showed a ganglionar relapse in CC). Table 1. Transplantation characteristics. Table 2. Transplantation results. Median (range).

Potential Increase in Graft Failure Risk after Reduced Intensity Conditioning Stem Cell Transplant (RIT) Using Umbilical Cord Blood for Children with Primary Immunodeficiency.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 2006-2006
Author(s):  
Jessica Guarino ◽  
Morris Kletzel ◽  
Reggie Duerst ◽  
David Jacobsohn ◽  
Jennifer Schneiderman ◽  
...  
Keyword(s):  
Stem Cell ◽  
Cord Blood ◽  
Median Time ◽  
Primary Immunodeficiency ◽  
Graft Failure ◽  
Conditioning Regimen ◽  
Reduced Intensity Conditioning ◽  
Cd34 Cells ◽  
Unrelated Donors ◽  
Reduced Intensity

Abstract Allogeneic stem cell transplantation is curative for patients with primary immunodeficiency. A unique reduced intensity conditioning regimen has been developed to maximize cure rate and minimize transplant-related toxicity. Between 2000 and 2007, we performed 16 RIT in patients with hyper-IgM syndrome (n=2), severe combined immune deficiency (SCID) (n=10), immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome (IPEX) (n=2), Wiskott-Aldrich syndrome (n=1), and X-linked lymphoproliferative disease (n=1). There were 12 males and 4 females, with a median age at the time of RIT of 8.0 months (range 1.1 months to 8.9 years). Donor sources included peripheral blood stem cells (PBSCs) from matched related donors (n=7) or unrelated donors (n=2), and cord blood units (CBUs) (n=7). The median cell dose was 8.55 × 108 total nucleated cells (TNC)/kg and 6.7 × 106 CD34+cells/kg for the PBSC group, and 1.99 × 108 TNC/kg and 0.71 × 106 CD34+cells/kg for the CBU group. The conditioning regimen consisted of Fludarabine (Flu), days -10 through -5, intravenous Busulfan (Bu), days -5 and -4, and Anti-thymocyte globulin (ATG), days -4 through -1. GVHD prophylaxis included tacrolimus/prednisone (n=1), cyclosporine A (CSA) alone (n=2), and CSA/mycophenolate mofetil (n=13). All patients who received PBSCs from related and unrelated donors engrafted (9/9), whereas only 4/7 (57%) patients who received CBUs engrafted. The 3 patients who experienced primary graft failure had the following diagnoses; IPEX, T−B−NK+ SCID and T−B+NK+ SCID. Their cord blood doses were 0.78, 1.19 and 1.99 × 108 TNC/kg, and 0.11, 0.69, and 0.55 × 106 CD34+cells/kg, respectively. For the 13 patients who engrafted, median time to absolute neutrophil count (ANC) &gt;1000 was 19 days (range 4 to 53) and median time to platelets &gt;50K was 23 days (range 14 to 90). The ANC never dropped &lt;500 for 8/13 (62%) patients, and platelets never dropped &lt;20K for 9/13 (69%) patients who engrafted. VNTR analysis of donor cell contribution showed that full donor chimerism was achieved in 8/16 patients (50%; 6 received PBSCs and 2 received CBUs), and, partial donor chimerism was achieved in 5/16 patients (31%; 3 received PBSCs and 2 received CBUs). Toxicities within 100 days post-RIT included bacteremia (n=8), candidemia (n=1), and viral infection (n=6). All infections were effectively treated and patients fully recovered. No episodes of seizure or veno-occlusive disease were experienced. No mucositis or severe nausea/vomiting was reported. No grade III/IV acute graft-versus-host disease (GVHD) or chronic GVHD was seen. Three patients died within 100 days post-RIT from causes related to primary or secondary graft failure. The overall survival was 81.0% at 2 year post-RIT (95% CI 89.5–71.5). All deceased patients received CBUs as the donor source. One of these patients were considered high-risk with pre-RIT Lansky score =30%. If this patient was excluded from analysis, the 100 day RIT related mortality was 13.3%. This retrospective analysis revealed that RIT with Flu/Bu/ATG conditioning is well tolerated in children with primary immunodeficiency. The use of CBUs, however, appeared to increase the risk of graft failure. A larger study of the use of RIT in primary immunodeficiency could further examine this hypothesis.

The Polycomb Gene BMI1 Collaborates with BCR-ABL in Leukemic Transformation of Human Cord Blood CD34+ Cells.

Blood ◽  
2008 ◽  
Vol 112 (11) ◽  
pp. 1350-1350
Author(s):  
Aleksandra Rizo ◽  
Sandra Olthof ◽  
OS van Ronald ◽  
Bert HJ Dontje ◽  
Edo Vellenga ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Stem Cell ◽  
B Cells ◽  
Cord Blood ◽  
Bone Marrow Microenvironment ◽  
Leukemic Transformation ◽  
Human Cord Blood ◽  
Self Renewal ◽  
Cd34 Cells

Abstract Previously, we demonstrated that BMI1 acts as a stem cell maintenance factor for human stem/progenitor cells. Here, we report that BMI1 collaborates with BCR-ABL in inducing leukemogenic transformation of human cord blood (CB) CD34+ cells. BMI1 and BCR-ABL were co-expressed into CB CD34+ cells (further referred as B/B cells) using a retroviral approach and cells were transplanted into NOD-SCID mice. In two out of five mice we observed leukemia within 4 months after transplantation. Chimerism levels reached 80–90% in the bone marrow and peripheral blood and morphological analysis revealed the appearance of primitive blast-like human hematopoietic cells with features that recapitulate human lymphoid leukemia. The mice were lethargic, with splenomegaly and infiltration of leukemic cells in the spleen, liver and the bone marrow and immunophenotypical analyses revealed that the cells expressed CD34 and CD19. To further understand the mechanisms underlying the leukemic transformation we performed ex-vivo long-term cultures on bone marrow stroma. We observed that the double transduced B/B cells had a strong proliferative advantage and elevated self-renewal potential as compared to controls. Expanding cultures could be maintained for over 20 weeks and Cobblestone Area Forming Cells (CAFCs) could be harvested and replated to initiate new expanding cocultures. Stem cell frequencies were determined in Long-Term Culture-Initiating Cell (LTC-IC) assays and frequencies were enhanced over 100-fold as compared to controls. Depending on the MS5 co-culture conditions, both myeloid as well as lymphoid long-term cultures could be established, indicating that extrinsic factors might dictate the lineage fate of transformed cells. To determine the necessity of a bone marrow microenvironment, we performed stroma-free liquid cultures and observed that the B/B cells were capable of expanding over 23 weeks, BMI1 cells were able to grow for 16 weeks and, importantly, BCR-ABL cells were not able to propagate long-term in stromain-dependent cultures. Thus, these data suggest that BCR-ABL cells are still dependent on cues from the bone marrow microenvironment for long-term self-renewal, and that co-expression of the intrinsic stem cell regulator BMI1 might alleviate this necessity of BCR-ABL+ cells for a microenvironment. Experiments in which B/B-transduced cells were sorted into HSC, CMP, GMP and MEP populations indicated that long-term self-renewal and expansion could particularly be imposed on the HSC population, and much less efficiently on progenitor subpopulations. In order to study whether the B/B-leukemic stem cells could be targeted by Imatinib, we applied a short pulse of Imatinib to expanding MS5 cocultures for 7 days. While the vast majority of cells in all cultures did not survive, in the B/B-transduced group a population of immature cells remained that was capable of re-initiating proliferative cultures of self-renewing CAFCs with very high frequencies (1/96 as determined by LTC-IC assays). Finally, we asked whether retroviral introduction of BMI1 in BCR-ABL+ CD34+ cells isolated from CML patients in chronic phase that expressed low endogenous BMI1 levels would affect long-term growth and self-renewal. Upon overexpression of BMI1 we observed increased proliferation capacity of the BMI1 transduced CML cells, and cultures could be maintained for much longer periods than control-transduced cultures. In conclusion, our data indicate that BMI1 collaborates with BCR-ABL in leukemic transformation, and our human-based system should provide a useful model to study the pathology of leukemias and test new drug entities.

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