Long-term remission after successful pregnancy in autologous peripheral blood stem cell transplanted system lupus erythematosus patients

Pregnancies in systemic lupus erythematosus (SLE) patients are high risk to both mother and fetus because of increased rates of complications. During the past years, we have treated many cases of SLE patients using autologous peripheral blood stem cell transplantation with good outcome after pregnancy. The rate of maternal hypertension and lupus nephritis was greatly reduced in autologous peripheral blood stem cell transplanted group (n = 11) when compared to non-transplant group (n = 39) (P < 0.05). In addition, the outcome of lupus flare activity of the mother after delivery is significantly better in transplanted group than that in non-transplanted group (P < 0.05). Here, we describe two typical cases of long duration (>6 years) of remission after successful pregnancy in refractory SLE patients post-autologous peripheral blood stem cell transplantation. Our report demonstrated that peripheral blood stem cell transplantation is safe and effective, thereby could be recommended as prior strategy in refractory SLE patients, especially for those women of child-bearing age who plan for pregnancy.     

Changes of immunological markers after autologous peripheral blood stem cell transplantation

Recently high-dose chemotherapy with peripheral blood stem cell transplantation (PBSCT) has become an important treatment for hematological and solid tumors. Methods: Immunological parameters were examined before and after PBSCT in 9 patients with lung cancer and 13 patients with malignant lymphoma. Findings were compared with those for bone marrow transplantation (BMT). Peripheral blood cells were analyzed for phenotype and the levels of cytokines and soluble factors were measured. Results: After PBSCT, activated T cells (CD3⁺HLA-DR⁺ cells, CD8⁺HLA-DR⁺ cells) and suppressor/cytotoxic T cells (CD8⁺CD11b⁻ cells) were significantly higher in the patients with lung cancer than in those with malignant lymphoma. Serum levels of interleukin-4 and soluble interleukin-2 receptor were also significantly higher in the patients with lung cancer than in those with lymphoma. On the other hand, the serum levels of interferon γ, tumor necrosis factor α, interleukin-6, soluble human leukocyte antigen class 1, and soluble thrombomodulin were significantly increased after bone marrow transplantation. The transfused peripheral stem cells of lung cancer and lymphoma patients had a similar number of granulocyte/macrophage-colony-forming units, but lung cancer patients had significantly more CD34-positive cells. Conclusion: By reinfusing large numbers of autologous immune cells, PBSCT may accelerate immune reconstitution, with T cells being likely to have a marked therapeutic potential. The changes after PBSCT were greater in patients with lung cancer than in lymphoma patients. These blood cells are potent mediators of anticancer activity and could play an important role in the elimination of autologous malignant cells. Received: 20 April 1998 / Accepted: 24 August 1998     

Allogeneic transplantation using CD34+ selected peripheral blood progenitor cells combined with non‐mobilized donor T cells for refractory severe aplastic anaemia

Allogeneic haematopoietic stem cell transplantation is curative for severe aplastic anaemia (SAA ) unresponsive to immunosuppressive therapy. To reduce chronic graft‐versus‐host disease (GVHD ), which occurs more frequently after peripheral blood stem cell (PBSC ) transplantation compared to bone‐marrow transplantation (BMT ), and to prevent graft rejection, we developed a novel partial T‐cell depleted transplant that infuses high numbers of granulocyte colony‐stimulating factor‐mobilized CD 34⁺ selected PBSC s combined with a BMT ‐equivalent dose of non‐mobilized donor T‐cells. Fifteen patients with refractory SAA received cyclophosphamide, anti‐thymocyte globulin and fludarabine conditioning, and were transplanted with a median 8 × 10⁶ CD 34⁺ cells/kg and 2 × 10⁷ non‐mobilized CD 3⁺ T‐cells/kg from human leucocyte antigen‐matched sibling donors. All achieved sustained engraftment with only two developing acute and two developing chronic GVHD . With a 3·5‐year median follow‐up, 86% of patients survived and were transfusion‐independent. When compared to a retrospective cohort of 56 bone‐marrow failure patients that received the identical transplant preparative regimen and GVHD prophylaxis with the exception that the allograft contained unmanipulated PBSC s, partial T‐cell depleted transplant recipients had delayed donor T‐cell chimerism and relative reduction of 75% in the incidence of acute grade II ‐IV GVHD (13% vs. 52%; P  =  0·010) and of 82% in chronic GVHD (13% vs. 72%; P  =  0·0004). In multivariate analysis, partial T‐cell depleted transplants remained significantly associated with a reduced risk of GVHD . In conclusion, for patients with refractory SAA , this novel transplant strategy achieves excellent engraftment and survival when compared to unmanipulated PBSC transplants and dramatically reduces the incidence of both acute and chronic GVHD .     

Treatment of Autoimmune Disease by Intense Immunosuppressive Conditioning and Autologous Hematopoietic Stem Cell Transplantation

Multiple sclerosis, systemic lupus erythematosus, and rheumatoidarthritis are immune-mediated diseases that are responsive tosuppression or modulation of the immune system. For patients withsevere disease, immunosuppression may be intensified to the point ofmyelosuppression or hematopoietic ablation. Hematopoiesis and immunitymay then be rapidly reconstituted by reinfusion of CD34⁺progenitor cells. In 10 patients with these autoimmune diseases, autologous hematopoietic stem cells were collected from bone marrow ormobilized from peripheral blood with either granulocytecolony-stimulating factor (G-CSF) or cyclophosphamide and G-CSF. Stemcells were enriched ex vivo using CD34⁺ selection andreinfused after either myelosuppressive conditioning withcyclophosphamide (200 mg/kg), methylprednisolone (4 g) and antithymocyte globulin (ATG; 90 mg/kg) or myeloablative conditioning with total body irradiation (1,200 cGy), methylprednisolone (4 g), andcyclophosphamide (120 mg/kg). Six patients with multiple sclerosis, 2 with systemic lupus erythematosus, and 2 with rheumatoid arthritis haveundergone hematopoietic stem cell transplantation. Mean time toengraftment of an absolute neutrophil count greater than 500/µL (0.5 × 10⁹/L) and a nontransfused platelet count greater than20,000/µL (20 × 10⁹/L) occurred on day 10 and 14, respectively. Regimen-related nonhematopoietic toxicity was minimal.All patients improved and/or had stabilization of disease witha follow-up of 5 to 17 months (median, 11 months). We conclude thatintense immunosuppressive conditioning and autologous T-cell-depletedhematopoietic transplantation was safely used to treat these 10 patients with severe autoimmune disease. Although durability ofresponse is as yet unknown, all patients have demonstrated stabilization or improvement.     

Use of CD34+ autologous stem cell transplantation in the treatment of children with refractory systemic lupus erythematosus

We report on the unique effects and benefits of autologous stem cell transplantation in childhood systemic lupus erythematosus (SLE) and describe this procedure in two young girls with severe and refractory disease. The patients stem cells were mobilized with granulocyte colony-stimulating factor (G-CSF) and collected by CS-3000 Blood Cell Separator (Baxter Healthcare, Round Lake, Ill., USA), and the CliniMACS CD34⁺ cell selection device (Miltenyi Biotech, Bergisch Gladbach, Germany) was used to obtain CD34⁺ cells. A total of 1.7×10⁶ and 1.0×10⁶/kg CD34⁺ cells were obtained, with 2.0×10⁵ and 1.0×10⁴/kg of CD3⁺ cells remaining, respectively. The conditioning regimen consisted of cyclophosphamide (50 mg/kg per day for 4 days) plus antithymocyte globulin (ATG-Fresenius, 5 mg/kg per day for 3 days). Neutrophil counts recovered within 9 days in both cases. Within 15 days, the platelet counts recovered and were sustained over 100×10⁹/l. Cushingoid features disappeared completely 3 months after transplantation because of the removal of corticosteroid medication. One 13-year-old child increased her height by 5 cm in 6 months after stopping steroids. She had not increased her height in her previous 7 years of disease. As of the time of this report, the first patient remains in clinical and laboratory remission for nearly 4 years, while the second suffered a relapse of thrombocytopenia 9 months post-transplantation. One residual effect of their treatment is that their CD4⁺ cell counts remained in the lower range after one year of transplant. The effect of this conditioning regimen plus CD34⁺ autologous stem cell transplantation on these two children with refractory SLE was beneficial, but long-term follow-up data and additional experience with this procedure are required. Autologous stem cell transplantation may limit the long-term toxicity of therapy in childhood SLE.     

Successful treatment of cytomegalovirus enteritis after unrelated allogeneic stem cell transplantation by the infusion of ex vivo‐expanded CD4+ lymphocytes derived from the recipient's peripheral blood donor cells

Adoptive immunotherapies have been developed for antiviral agent‐refractory cytomegalovirus (CMV) disease after stem cell transplantation (SCT). However, the application of such strategies is limited, particularly in terms of need for donor cooperation regarding blood sampling and inaccessibility in the setting of cord blood transplantation. Herein, we describe the first successful treatment of antiviral agent‐refractory CMV enteritis after allogeneic SCT by the infusion of ex vivo‐expanded donor‐derived CD4⁺ lymphocytes obtained from the recipient's peripheral blood.     

Loss of quiescence and impaired function of CD34+/CD38low cells one year following autologous stem cell transplantation

Patients who have undergone autologous stem cell transplantation are subsequently more susceptible to chemotherapy-induced bone marrow toxicity. In the present study, bone marrow primitive progenitor cells were examined one year after autologous stem cell transplantation and compared with normal bone marrow and mobilized peripheral blood stem cells. Post-transplantation bone marrow contained a significantly lower percentage of quiescent cells in the CD34⁺/CD38^low fraction compared to normal bone marrow. In addition, we observed a strong decrease in stem cell/primitive progenitor frequency in post-transplantation CD34⁺ cells as defined by long-term culture assays. Measurement of the levels of reactive oxygen species by flow cytometry revealed comparable levels in post-transplantation and normal bone marrow CD34⁺/CD38^low cells, while significantly higher levels of reactive oxygen species were observed in CD34⁺/CD38^high cells following autologous stem cell transplantation compared to normal bone marrow. Moreover, post-transplantation CD34⁺ bone marrow cells demonstrated an increased sensitivity to buthionine sulfoximine, a trigger for endogenous production of reactive oxygen species. Gene expression analysis on CD34⁺ cells revealed a set of 195 genes, including HMOX1, EGR1, FOS and SIRPA that are persistently down-regulated in mobilized peripheral blood cells and post-transplantation bone marrow compared to normal bone marrow. In conclusion, our data indicate that the diminished regenerative capacity of bone marrow following autologous stem cell transplantation is possibly related to a loss of quiescence and a reduced tolerability to oxidative stress.     

A phase I-II trial of autologous peripheral blood stem cell transplantation in the treatment of refractory autoimmune disease

OBJECTIVES: To carry out a phase I-II trial to elucidate the feasibility and efficacy of high dose cyclophosphamide (CY) supported by autologous peripheral blood stem cell transplantation (PBSCT) in the treatment of severe and refractory autoimmune disease (AD). METHODS: Peripheral blood stem cells (PBSCs) were mobilised during haematological recovery after relatively high dose CY (2 g/m2) for 2 days, followed by administration of granulocyte colony stimulating factor. After collecting PBSCs--more than 2x10(6) CD34+ cells/kg--by apheresis, CD34+ cells were immunologically selected and cryopreserved. Eight patients were enrolled--five had systemic sclerosis (SSc) alone, one had SSc with systemic lupus erythematosus, one amyopathic dermatomyositis (ADM), and one Wegener's granulomatosis (WG). All of the patients were treated with high dose CY (50 mg/kg) for 4 days and autologous PBSCT. RESULTS: Haematopoietic reconstitution was rapid and sustained. Toxicity due to the regimen included various infections such as pneumonia, sepsis, cystitis, herpes zoster, and acute heart failure. However, there was no treatment related mortality. Encouraging results were obtained after autologous PBSCT. Sclerosis of the skin was markedly improved in all of the patients with SSc. Interstitial pneumonia (IP), evaluated by PaO2, serum KL-6 levels, and pulmonary high resolution computed tomography, improved significantly. In a patient with ADM, severe and progressive IP also improved markedly. In a patient with WG, the size of the left orbital granuloma decreased substantially, resulting in reduction of the exophthalmos. CONCLUSIONS: These observations suggest that high dose CY with autologous PBSCT is feasible and may be effective in the treatment of severe and refractory AD.     

CD34+ selected stem cell boosts can improve poor graft function after paediatric allogeneic stem cell transplantation

Poor graft function (PGF) is a severe complication of haematopoietic stem cell transplantation (HSCT) and administration of donor stem cell boosts (SCBs) represents a therapeutic option. We report 50 paediatric patients with PGF who received 61 boosts with CD34⁺ selected peripheral blood stem cells (PBSC) after transplantation from matched unrelated (n = 25) or mismatched related (n = 25) donors. Within 8 weeks, a significant increase in median neutrophil counts (0·6 vs. 1·516 × 10⁹/l, P < 0·05) and a decrease in red blood cell and platelet transfusion requirement (median frequencies 1 and 7 vs. 0, P < 0·0001 and <0·001), were observed, and 78·8% of patients resolved one or two of their cytopenias. 36·5% had a complete haematological response. Median lymphocyte counts for CD3⁺, CD3⁺CD4⁺, CD19⁺ and CD56⁺ increased 8·3‐, 14·2‐, 22.‐ and 1·6‐fold. The rate of de novo acute graft‐versus‐host disease (GvHD) grade I–III was only 6% and resolved completely. No GvHD grade IV or chronic GvHD occurred. Patients who responded to SCB displayed a trend toward better overall survival (OS) (P = 0·07). Thus, administration of CD34⁺ selected SCBs from alternative donors is safe and effective. Further studies are warranted to clarify the impact on immune reconstitution and survival.     

Just‐in‐time rescue plerixafor in combination with chemotherapy and granulocyte‐colony stimulating factor for peripheral blood progenitor cell mobilization

Plerixafor, a recently approved peripheral blood progenitor cell mobilizing agent, is often added to granulocyte‐colony stimulating factor (G‐CSF) to mobilize peripheral blood progenitor cells in patients with lymphoma or myeloma who cannot mobilize enough CD34+ cells with G‐CSF alone to undergo autologous stem cell transplantation. However, data are lacking regarding the feasibility and efficacy of just‐in‐time plerixafor in combination with chemotherapy and G‐CSF. We reviewed the peripheral blood stem cell collection data of 38 consecutive patients with lymphoma (Hodgkin's and non‐Hodgkin's) and multiple myeloma who underwent chemomobilization and high‐dose G‐CSF and just‐in‐time plerixafor to evaluate the efficacy of this treatment combination. All patients with multiple myeloma and all but one patient with lymphoma collected the minimum required number of CD34+ cells to proceed with autologous stem cell transplantation (>2 × 10⁶/kg of body weight). The median CD34+ cell dose collected in patients with non‐Hodgkin lymphoma was 4.93 × 10⁶/kg of body weight. The median CD34+ cell dose collected for patients with multiple myeloma was 8.81 × 10⁶/kg of body weight. Plerixafor was well tolerated; no grade 2 or higher non‐hematologic toxic effects were observed. Am. J. Hematol. 88:754–757, 2013. © 2013 Wiley Periodicals, Inc.     

Immune recovery after autologous or rhG-CSF primed PBSC transplantation

Abstract: We performed a prospective study in 17 consecutive patients following autologous bone marrow (BM) or rhG-CSF primed peripheral blood stem cell (PBSC) transplantation, with the objective of comparing immune recovery between both procedures and to evaluate results in rhG-CSF mobilized peripheral blood stem cell transplantation (PBSCT). Kinetics of immune reconstitution showed differences, with a faster recovery of CD3+ and CD8+ T cells, and a more rapid and sustained recovery of CD8^±/CD56+ natural killer (NK) cells in the PBCSCT group. Autologous bone marrow transplantation (ABMT) was associated with a improved reconstitution of the CD19+/CD5^± subpopulation. Moreover, rhG-CSF mobilized PBSCT generated a greater recovery of CD8^±/CD56+ cells than previous data concerning transplantation with peripheral blood (PB) progenitors collected after myelosuppressive chemotherapy or myelosuppressive therapy plus rhG-CSF. Our results show differences in the rate and pattern of B and T lymphocytes reconstitution after ABMT and PBSCT. Additionally, we state an enhancement of CD56+ cells in patients undergoing PBSCT mobilized solely using rhG-CSF.     

Prediction model for CD34 positive cell yield in peripheral blood stem cell collection on the fourth day after G-CSF administration in healthy donors

Allogeneic peripheral blood stem cell transplantation (PBSCT) is an indispensable treatment option for hematological malignancy. The optimal collection day after granulocyte colony-stimulating factor (G-CSF) administration should be determined by peripheral blood pre-apheresis CD34 positive (CD34⁺) cell percentage. However, pre-apheresis CD34⁺ cell analysis is not available for most institutions in Japan. Prediction of the optimal collection day based on objective parameters, other than direct CD34⁺ cell count, is thus an important matter for investigation. To identify potential predictive factors, clinical parameters in 79 related donors who received allogeneic peripheral blood stem cell (PBSC) collection were analyzed. Eight factors were significantly correlated with the number of CD34⁺ cells per donor body weight on the fourth day (day 4) after G-CSF administration in univariate analysis. Using multi-regression analysis, we made a simple scoring system comprising age, sex, LDH on day 4 and RBC count at the baseline, which significantly predicted CD34⁺ cell yield (P = 0.048). This system allows us to determine the optimal PBSC collection day. When the score is 0 or 1 on day 4, starting apheresis on day 5 potentially helps avoiding the need for multiple harvests. Score 3 or 4 on day 4 is indicative of better performance if apheresis is started on day 4.     

T cell depletion utilizing CD34(+) stem cell selection and CD3(+) addback from unrelated adult donors in paediatric allogeneic stem cell transplantation recipients

CD34‐selected haploidentical and unrelated donor allogeneic stem cell transplantation (AlloSCT) in paediatric recipients is associated with sustained engraftment and low risk of acute graft‐versus‐host disease (aGVHD), but limited by delayed immune reconstitution and increased risk of viral and fungal infection. The optimal dose of donor T cells to prevent graft failure and minimize risk of early opportunistic infection and post‐transplant lymphoproliferative disorder (PTLD), while avoiding severe aGVHD, remains unknown. We prospectively studied CD34‐selected 8–10/10 human leucocyte antigen (HLA)‐matched unrelated donor (MUD) peripheral blood stem cell transplantation (PBSCT) in a cohort of 19 paediatric AlloSCT recipients with malignant (n = 13) or non‐malignant (n = 6) diseases. T cells were added back to achieve total dose 1·0–2·5 × 10⁵ CD3⁺/kg. GVHD pharmacoprophylaxis consisted only of tacrolimus. All patients engrafted neutrophils. Probabilities of grade II–IV aGVHD, limited chronic GVHD (cGVHD), and extensive cGVHD were 15·8%, 23·3%, and 0%, respectively. One patient developed PTLD. One‐year infection‐related mortality was 5·6%. T cell immune reconstitution was delayed. One‐year overall survival was 82·3%. Five patients with malignant disease ultimately died from progressive disease. CD34‐selected MUD PBSCT using a defined dose of T cell add‐back resulted in high rates of engraftment and low risk of grade II–IV aGVHD, early transplantation‐related mortality, and extensive cGVHD.     

Evans' syndrome following autologous peripheral blood stem cell transplantation for non‐Hodgkin's lymphoma

A 34‐year‐old woman with non‐Hodgkin's lymphoma received high‐dose chemotherapy followed by autologous peripheral blood stem cell transplantation (PBSCT). She developed Evans’ syndrome, the association of immune thrombocytopenia and autoimmune hemolytic anemia, 49 days after transplantation. Multiple autoimmune disorders may occur concurrently after autologous PBSCT.     

Serum levels of soluble CD30 in autologous peripheral blood stem cell transplantation

High-dose chemotherapy with peripheral blood stem cell transplantation (PBSCT) has become an important method of treatment for hematological and solid tumors. We examined levels of interleukin(IL)-4, interferon (IFN)γ, soluble (s) CD30, and sIL-2 receptor (R) before and after autologous PBSCT, and compared findings for PBSCT with those for bone marrow transplantation (BMT). We found significantly higher IL-4 levels 1 and 3 weeks after PBSCT than before PBSCT, while IFNγ levels remained almost unchanged after PBSCT. IFNγ levels were increased 3 weeks after BMT, although no increase in IL-4 level was observed. The serum sCD30 level was significantly higher 3 weeks after PBSCT, but not following BMT. For 34 samples on days 0 and 21 from 17 patients undergoing PBSCT, a strong correlation was observed between sCD30 and sIL-2R levels (r = 0.43, P < 0.01). However, no significant correlation between sCD30 and sIL-2R levels was found for BMT patients. These findings suggest that sCD30 is a useful marker for evaluating immunological activity following autologous PBSCT, and that the immunological conditions after autologous PBSCT may be associated with helper-T-cell-2-type immune responses. Received: 31 May 1999 / Accepted: 20 September 1999     

Acute but not chronic graft-versus-host disease is associated with a reduction of circulating CD4<Superscript>+</Superscript>CD25<Superscript>high</Superscript>CD127<Superscript>low/−</Superscript> regulatory T cells

Defects in central and peripheral tolerance are thought to contribute to life-threatening graft-versus-host disease (GvHD), a severe complication following allogeneic stem cell transplantation (SCT). Recent investigations have demonstrated regulatory T cells (Tregs) to suppress allogeneic immune reactions. Therefore, SCT patients with no or critically low numbers of Tregs may have an increased risk of GvHD. To address this hypothesis, we analyzed the recovery of CD4⁺CD25^highCD127^low/− Tregs in the peripheral blood of patients who have never developed GvHD (n = 6), patients who developed acute/chronic GvHD (n = 18), and patients who developed chronic GvHD without an earlier acute manifestation (n = 5) every 30 days for the first 6 months after peripheral blood SCT (PBSCT). The number of Tregs continuously improved in acute/chronic GvHD patients, but always remained lower than Tregs quantified in patients who never developed a GvHD. In contrast, chronic GvHD patients who did not develop acute GvHD earlier displayed significantly increased Treg cell numbers at the timepoint of chronic inflammation. These results indicate that numerically deficient Tregs following PBSCT are associated with the development of acute but not chronic GvHD.     

Enhancement of cyclosporin A-induced autologous graft-versus-host disease after peripheral blood stem cell transplantation by utilizing selected CD34(+) cells

Although autologous graft-versus-host disease (GVHD) can be induced by administration of cyclosporin A (CsA) after peripheral blood stem cell transplantation (PBSCT), the incidence appears to be remarkably lower compared to the incidence after bone marrow transplantation. The reduced incidence of autologous GVHD after PBSCT may be attributed to peripheral regulatory cells that are transferred with the stem cell inoculum. To determine whether transplantation of CD34-selected peripheral blood stem cells (PBSCs) leads to potentiation of autologous GVHD, five patients with malignant lymphoma were transplanted with CD34-selected PBSCs, followed by administration of CsA and interferon (IFN)-gamma. Inducibility of autologous GVHD and autocytotoxic activities of peripheral blood mononuclear cells (PBMCs) after transplantation were assessed. All patients demonstrated prompt hematologic recovery. Cytotoxic activity of PBMCs against autologous lymphocytes was detectable in four of four patients analyzed during a limited period from days 14 to 34 post-transplant. An erythematous rash compatible with GVHD, confirmed by skin biopsy, developed in three of five patients. One of the three patients developed not only skin, but also gut and liver GVHD. Transplantation of the CD34-selected stem cell graft that does not accompany transfusion of regulatory cells may potentiate the inducibility of autologous GVHD by the administration of CsA and IFN-gamma.     

Influence of ex vivo purging with CliniMACS CD34+ selection on outcome after autologous stem cell transplantation in non‐Hodgkin lymphoma

The major limitation of autologous stem cell transplantation (auto‐SCT) in non‐Hodgkin lymphoma (NHL) is relapse. Although autologous graft contamination may be a potential cause, prior purging of the autograft remains controversial. Therefore, we retrospectively analysed 56 consecutive patients with NHL receiving auto‐SCT at complete (n = 41) or partial remission (n = 15). Among them, 24 patients underwent autograft manipulation with positive selection of CD34⁺ cells using a CliniMACS device (purged group). Twenty‐five patients had received ≥2 previous chemotherapy regimens before auto‐SCT. After a median follow‐up of 41·4 months, transplant‐related mortality was observed only in unpurged group (n = 2; 3·6%). The 3‐year overall survival (91·7% vs. 56·1%, P = 0·009) and progression‐free survival (78·7% vs. 53·1%, P = 0·034) favoured CD34⁺ purification. While neutrophil recovery was similar, platelet recovery was delayed in the purged group. Cytomegalovirus reactivation was predominantly observed in the purged group, although no other clinically unmanageable infectious complications occurred. Although this study has the inevitable limitations of heterogeneity in previous treatment and NHL subtypes, and a small number of patients analysed, the high survival rate in the purged group may suggest the need for prospective randomized trials to determine the role of CD34⁺ purification in auto‐SCT for NHL.     

Long term cryopreservation in 5% DMSO maintains unchanged CD34+ cells viability and allows satisfactory hematological engraftment after peripheral blood stem cell transplantation

Peripheral blood stem cell cryopreservation is associated with cell damage and decreased viability. We evaluated the impact of up to 10 years of cryopreservation (5% DMSO) on viability of CD34⁺ cells utilizing graft samples of consecutive patients (2002–2012) with different malignancies who underwent stem cell collection and transplantation. Viability of CD34⁺ cells from oncohaematological patients measured after 5 weeks (97·2 ± 0·6%) or after 9–10 years of cryopreservation (95·9 ± 0·5%) was unaffected. Haemoglobin, granulocyte and platelet recovery after transplantation of long‐term cryopreserved grafts occurred within 8–13 days. CD34⁺ stem cells can be safely stored up to 9–10 years, without affecting cell viability and clinical effectiveness.     

Platelet-released supernatants enhance hematopoietic stem cell proliferation in vitro

Peripheral blood stem cell transplantation (PBSCT) is used to reconstitute normal hematopoiesis after myeloablative chemotherapy. The hematopoietic stem cells are collected from the blood by apheresis machines using density gradient centrifugation. Because of density similarities the grafts contain high levels of leukocytes and platelets that release various mediators into the grafts. The collected hematopoietic stem cells are therefore exposed to platelet released mediators including platelet-derived growth factor, transforming growth factor-β, vascular endothelial growth factor and platelet factor-4. To investigate whether platelet activation and secretion can affect hematopoietic stem cells during PBSCT, we cultured (i) normal cord blood stem cells and (ii) mobilized peripheral blood stem cells from autografts together with the total secretion product of thrombin activated platelets (i.e. platelet released supernatants). Platelet released supernatants enhanced the cell proliferation of both peripheral blood stem cell (PBSC) autograft and normal cord blood CD34⁺ cells. Our study shows that platelet secretion in PBSCT autografts affect the hematopoietic stem cell function and possibly thereby the hematopoietic reconstitution after PBSCT.