Growth differentiation factor 11 (GDF11) - a promising anti-ageing factor - is highly concentrated in platelets.

Recent research suggests that growth differentiation factor 11 (GDF11) could reverse age-related diseases and that its blood concentration decreases with age. This poses plasma from young donors as a therapeutic GDF11 source to treat age-related diseases. In addition, the tissue source of circulating GDF11 remains unknown. We analysed GDF11 levels in paired samples of serum, plasma and platelet lysate (PL) from 23 volunteers. Plasma and PL were collected by plateletpheresis. Here, we show that GDF11 is highly concentrated in platelets and that the circulating levels reported in previous studies could be biased as a result of serum sample manipulation.

Severe infections after single umbilical cord blood transplantation in adults with or without the co-infusion of CD34+ cells from a third-party donor: results of a multicenter study from the Grupo Español de Trasplante Hematopoyético (GETH).

BACKGROUND: Umbilical cord blood transplantation (CBT) is an established alternative source of stem cells in the setting of unrelated transplantation. When compared with other sources, single-unit CBT (sCBT) is associated with a delayed hematologic recovery, which may lead to a higher infection-related mortality (IRM). Co-infusion with the sCBT of CD34+ peripheral blood stem cells from a third-party donor (TPD) (sCBT + TPDCD34+) has been shown to markedly accelerate leukocyte recovery, potentially reducing the IRM. However, to our knowledge, no comparative studies have focused on severe infections and IRM with these 2 sCBT strategies. METHODS: A total of 148 consecutive sCBT (2000-2010, median follow-up 4.5 years) were included in a multicenter retrospective study to analyze the incidence and risk factors of IRM and severe viral and invasive fungal infections (IFIs). Neutrophil engraftment occurred in 90% of sCBT (n = 77) and 94% sCBT + TPDCD34+ (n = 71) recipients at a median of 23 and 12 days post transplantation, respectively (P < 0.01). RESULTS: The 4-year IRM was 24% and 20%, respectively (P = 0.7), with no differences at day +30 (5% and 4%, respectively) and day +100 (10% and 8%, respectively). In multivariate analysis early status of the underlying malignancy, cytomegalovirus (CMV)-seronegative recipient and high CD34+ cell content in the cord blood unit before cryostorage (≥1.4 × 10(5) /kg) were protective of IRM. Among the causes of IRM, bacterial infections and IFIs were more common in sCBT (15% vs. 4%), while CMV disease and parasitic infections were more common in the sCBT + TPDCD34+ cohort (5% vs. 16%). CONCLUSION: These data show that sCBT supported with TPDCD34(+) cells results in much shorter periods of post-transplant leukopenia, but the short- and long-term rates of IRM were comparable to those of sCBT, presumably because immune recovery is equally delayed in both graft types.

Toxoplasmosis in cord blood transplantation recipients.

Toxoplasmosis is a devastating opportunistic infection that can affect immunocompromised patients such as cord blood transplantation (CBT) recipients. The clinical characteristics of 4 toxoplasmosis CBT patients treated at our institution are reviewed, together with 5 cases collected from the literature. The rate of toxoplasmosis in our hospital was 6% in CBT recipients and 0.2% in other types of allogeneic hematopoietic stem cell transplantation (P < 0.001). Five patients (56%) presented disseminated toxoplasmosis and 4 patients (44%) had localized infection in the central nervous system. In 5 of the 9 patients considered (56%), cytomegalovirus viral replication had been detected before the clinical onset of toxoplasmosis. Seven patients (78%) had previously developed graft-versus-host disease. All patients who exhibited disseminated disease died due to Toxoplasma infection. Pre-transplant serology was positive in 1 patient, negative in 3 patients, and not performed in another. Only 1 of these 5 patients with disseminated disease had received Toxoplasma prophylaxis with cotrimoxazole. It could be concluded that mortality in CBT patients with disseminated toxoplasmosis is unacceptably high. The negative results of serology in the majority of these cases, and its unspecific clinical presentation, makes diagnosis exceedingly difficult. Better diagnostic tests and prophylaxis strategy are needed in CBT recipients.

[Umbilical cord blood cell transplantation from an unrelated donor: dual transplantation]. / Trasplante de sangre de cordón umbilical más donante auxiliar: trasplante dual.

Our team conducted an original procedure of hematopoietic transplantation of umbilical cord blood (UCB) from an unrelated donor. The procedure consists of co-infusing hematopoietic stem cells selected from the blood of a third-party donor; it is conceived as a tool to shorten the engraftment period without preventing the engraftment of the UCB, even when using units with relatively low cell content and a low HLA compatibility. Between 1999 and 2008 we performed 64 transplantations in 60 adult patients (35 men and 25 women) with a median age of 34 years (range: 76-60) and a median weight of 70 kg (range: 43-95), all of whom were diagnosed with a high risk hematologic neoplasm (leukemia in most cases). Fludarabine, cyclophosphamide, ATG, and whole body irradiation or busulfan were used as conditioners. UCB was infused at medians of 2.4 x 107 CNT/kg (range: 1.14-4.30 x 107), 0.11 x 106 CD34+/kg (range: 0.035-0.37 x 106). Then, hematopoietic stem cells selected from the third-party donor were infused (2.43 x 106/kg [range: 1.05-3.34 x 106], with 0.3 x 104 CD3+/kg [range: 0.05-1.56 x 104]). Granulocyte engraftment occurred (ANC > 0.5 x 109/L) at a median of 10 days (range: 9-34 days), and the granulocyte engraftment of the UCB occurred in 21 days (range: 13-57 days). Complete UCB chimerism was observed in 37 days (range: 11-186 days) (previously double complete chimerism, presence of third-party donor and of cord) and platelet engraftment > 20 x 109/L in 33 days (range: 13 98 days) and > 50 x 109/L in 58 days (range: 14-106 days). Overall 3-year survival reached 51%, and 5 10 year-survival was 47% (plateau). Disease-free survival was 48% at three years, and 45% at 5 to 10 years; the mean follow-up of survivors was 48 months (range: 13-123 months). (Kaplan-Meier). In conclusion, early granulocyte recovery occurred thanks to a foster engraftment of hematopoietic stem from the third-party donor, which are not HLA-restricted; this is associated with a lower morbidity and mortality from infections secondary to neutropenia. There was also a high rate of engraftment and final full UCB chimerism, even with non-histocompatible UCB units (2/6 HLA mismatches) and with relatively low cell counts. In most cases, a single unit of UCB was sufficient. The incidence of severe GVHD and the percentage of relapses have been low. Opportunistic infections have occurred over a long period of time. This procedures makes allogeneic hematopoietic transplantation accessible to almost all patients.

Haplo-Cord transplantation compared to haploidentical transplantation with post-transplant cyclophosphamide in patients with AML.

For patients with AML, the best alternative donor remains to be defined. We analyze outcomes of patients who underwent myeloablative umbilical cord blood or haploidentical hemopoietic stem cell transplantation (HSCT) in Spain. Fifty-one patients underwent single umbilical cord blood transplantation supported by a third party donor (Haplo-Cord) between 1999 and 2012, and 36 patients received an haploidentical HSCT with post-transplant cyclophosphamide (PTCY-haplo) between 2012 and 2014 in GETH centers. The Haplo-Cord cohort included a higher proportion of patients with high disease risk index and use of TBI in the conditioning regimen, and hematopoietic cell transplantation-age Comorbidity Age Index was higher in PTCY-haplo patients. Cumulative incidence of neutrophil engraftment was 97% in the Haplo-Cord and 100% in the PTCY-haplo group, achieved in a median of 12 and 17 days, respectively (P=0.01). Grade II-IV acute GvHD rate was significantly higher in the PTCY-haplo group (9.8% vs 29%, P=0.02) as well as chronic GvHD rates (20% vs 38%, P=0.03). With a median follow-up of 61 months for the Haplo-Cord group and 26 months for the PTCY-haplo cohort, overall survival at 2 years was 55% and 59% (P=0.66), event-free survival was 45% vs 56% (P=0.46), relapse rate was 27% vs 21% (P=0.79), and non-relapse mortality was 17% vs 23% (P=0.54), respectively. In this multicenter experience, Haplo-Cord and PTCY-haplo HSCT offer valid alternatives for patients with AML. Neutrophil engraftment was faster in the Haplo-Cord cohort, with similar survival rates, with higher GvHD rates after haploidentical HSCT.

[Psychiatric patients in the pediatric emergency department of a tertiary care center: review of a 6-month period]. / Pacientes psiquiátricos en el servicio de urgencias de pediatría de un hospital terciario: revisión de un período de 6 meses.

INTRODUCTION: In the last few years, a marked increase in the number of psychiatric emergencies treated at pediatric emergency departments has been observed. The aim of the present study was to characterize these patients. PATIENTS AND METHODS: We performed a descriptive, prospective, cross sectional study, based on all psychiatric emergencies treated at the pediatric emergency department of Hospital General Universitario Gregorio Marañón, Madrid, from 1-10-04 to 31-3-05. The following variables were analyzed: age, sex, time of consultation, day of the week, day of the month, month of the year, psychiatric antecedents, previous psychiatric pharmacologic treatment, the person or service who took the child to hospital, diagnosis, and whether the patient was admitted to the hospital. RESULTS: Of a total of 36,449 emergencies, 79 were psychiatric (0.21 %). Sex rates were 48.1 % boys and 51.9 % girls. The mean (+/-2 SD) age was 13.73 +/- 2.5 years. Visits were most frequent on Mondays (19 %), in the evening, and in January and February. A total of 13.23 % of the patients were brought by extrahospital services. The main diagnoses were: behavioral disorders (36.76 %), anxiety disorders (20.58 %) and suicidal ideation or suicide attempt (13.23 %). There was a clear male predominance in behavioral disorders (67.85 %) and a female predominance in anxiety disorders (71.42 %) and suicidal ideation or suicide attempt (76.92 %). The hospitalization rate among these patients was 32.35 %. CONCLUSIONS: The incidence of psychiatric disorders in our pediatric emergency department was low. The patients were aged 11-15 years old, without differences between the sexes. Peak demand was reached on Mondays in January and February, at the end of the evening and beginning of the night. The most common diagnosis was behavioral disorder. The hospitalization rate was exceptionally high, more than 6 times higher than the average in our hospital.

Engraftment kinetics of human CD34+ cells from cord blood and mobilized peripheral blood co-transplanted into NOD/SCID mice.

We have reported short periods of post transplant neutropenia in human patients co-transplanted with cord blood (CB) and low numbers of haploidentical mobilized peripheral blood (MPB) CD34+ cells. To investigate the effect that the proportion of MPB to CB cells may have on engraftment kinetics, we have co-transplanted fixed numbers of human CB CD34+ cells mixed with different numbers of MPB CD34+ cells into NOD/SCID mice. We periodically quantified the proportion of human cells and the relative contribution of MPB and CB cells to the human engraftment on marrow aspirates. At the lowest MPB/CB ratios (5 : 1, 10 : 1), the contribution of CB cells predominated at all time points analyzed, and in three out of four experiments MPB cell contributions progressively decreased from day +15. At higher MPB/CB ratios, MPB cells had a more important contribution to both early and late engraftment, with the highest cell ratio resulting in only marginal CB cell engraftment. Therefore, our results showed greater potential, on a per cell basis, of human CB vs MPB cells for competitive sustained engraftment in the xenogeneic model used, which was only abrogated by the co-infusion of very high numbers of MPB cells.

Umbilical cord blood banking for unrelated transplantation: evaluation of cell separation and storage methods.

Cost-efficient umbilical cord blood (UCB) banking requires well-standardized methods of volume reduction and storage. To compare UCB fractionation using a technique of hydroxyethyl starch (HES) sedimentation with the Ficoll (double) and Percoll methods, 50 whole units was allocated randomly to each procedure. HES resulted in a significantly better recovery of mononuclear cells (87.5%), granulocyte/macrophage colony-forming units (CFU-GM) (88.4%), and CD34- cells (87.4%) and lesser volume reduction (85.5%). HES was the least laborious, time consuming, and expensive of the three procedures, costing 3.4- and 4.4-fold less than the Ficoll and Percoll methods, respectively. Five units processed by each method was frozen in 4.5-mL cryotubes under optimal conditions. After thawing, the greatest degree of recovery of viable nucleated cells and number of CFU-GM per unit were obtained using the HES procedure. Using 4.5-mL cryotubes, the calculated number of units that could be stored in 600-L containers was 3.8- and 2.2-fold higher for Ficoll- and Percoll-separated than for HES-separated units, respectively. Nevertheless, the higher direct costs of the density gradient separation procedures outweighed their lower storage cost. For long-term cryopreservation, we assessed the freezing of HES-processed units in 50-mL cryobags and their specifically designed canisters. We found cell recoveries similar to those obtained with cryotubes, but storage capacity was decreased. Special racks designed for these canisters resulted in a 5-fold increase over the number of units stored in standard cryobags. This system also is feasible for Percoll- and Ficoll-separated units, resulting in comparable storage costs for the three separation methods. We conclude that this HES procedure and the 50-mL cryobags constitute a cost-efficient system for large-scale UCB banking.

Conclusions of a national multicenter intercomparative study of in vitro cultures of human hematopoietic progenitors.

With the aim of developing a standardized program of clonogenic cultures, a multicenter intercomparative study of human CFU-GM, BFU-E and CFU-GEMM cultures was conducted. Aliquots of fresh mononuclear cord blood cells, as well as uniform culture materials and instructions for cell culture and for colony scoring were distributed to 28 national laboratories involved in hematopoietic research and transplantation. High interlaboratory coefficients of variation (CV) in the reported number of progenitors were found in our first intercomparative study (range 67-231%). To investigate the relevance of colony scoring in variations of the reported colony numbers, participants were invited to attend a meeting where a single culture dish was scored. In this case, the CVs ranged from 31% to 81%. A subsequent intercomparative assay was then conducted, and significant reductions in the inter-laboratory CVs were obtained with respect to the first study (CVs for colonies grown with two different media: CFU-GMs, 48% and 55%; BFU-Es, 70% and 62%; CFU-GEMMs, 70% and 51%; respectively). In most instances CVs were not significantly different from those obtained in the single plate scoring study, suggesting that the scoring process was the most relevant parameter accounting for variations in the reported colony numbers.

Evaluation of engraftment of ex vivo expanded cord blood cells in humans.

Cord blood transplants (CBT) result in high rates of engraftment in patients transplanted because of inherited diseases even across marked HLA disparities, mostly in children, with less severe manifestations of GVHD than BM and PBSC transplants. Evaluation of engraftment potential of CBT based on early progenitor content is difficult due to their inaccurate quantification. Instead, post-thaw nucleated cell counts (Pt-NCC) are commonly used for this purpose. We have analyzed engraftment as a function of pre-freeze nucleated cell counts (Pf-NCC) in patients receiving CBT because of inherited diseases. We have observed median times to engraftment of 26 days or less, shortest times ranging 8 to 13 days, late engraftment or graft failures tending to be associated with age >15 years and infusions of <3.7 x 10(7)/Pf-NCC/kg. These data may be appropriate references to evaluate engraftment of CBT performed with previously ex vivo expanded cells. CBT performed with units of which one aliquot has been previously culture-expanded have resulted in times to engraftment similar to the ones observed in the above-mentioned analysis. In these trials it is not possible to trace the actual origin of the early engrafting cells because the pre-cultured cells lack differentiating markers. To better evaluate the engraftment dynamics of culture-expanded CB cells in humans, we have used a model of simultaneously transplanting cells from two different donors to the same patient. Preliminary results of patients that have simultaneously received one uncultured CB unit and culture-expanded purified CB CD34+ cells obtained from a second one show no significant contribution of cultured cells to early engraftment, and no prohibitive unfavorable immunological problems have been observed.

HLA haploidentical cord blood cell transplant in a 15-year-old, 50 kg weight patient: successful treatment for chronic myeloid leukemia after myeloid blastic transformation.

A 15-year-old, 50 kg weight patient with CML had a myeloblastic transformation which reverted to Ph negative remission with intensive chemotherapy 5 years after diagnosis. Umbilical cord blood (UCB) from an HLA-haploidentical sister had been frozen 2 years and 9 months before, as she had no HLA-identical sibling and no suitable unrelated donor had been found. UCB transplant was selected on the basis of previous general experience with this kind of transplant, lack of a better choice of donor, and likelihood of a prompt relapse of the disease without delay and the patient developed grade II aGVHD as well as severe CsA toxicity which required discontinuation of the drug, anti-IL2r being given instead. Subsequently she only had histologic evidence of cGVHD and 1.5 years after the transplant she remains in complete hematologic remission with full chimerism and without evidence of the bcr/abl fusion gene. This case illustrates further possibilities of allo-transplantation using UCB.

Cord blood transplants: early recovery of neutrophils from co-transplanted sibling haploidentical progenitor cells and lack of engraftment of cultured cord blood cells, as ascertained by analysis of DNA polymorphisms.

The number of infused cells is a very important factor in cord blood transplant (CBT) engraftment. Prior ex vivo expansion of aliquots of transplanted cord blood (CB) units is being investigated as a procedure to increase engraftment potential, but results are difficult to evaluate due to a lack of markers for assessing the contribution of expanded cells. We transplanted five patients, infusing the best available CB unit and cells from a second donor simultaneously. In two patients, these cells were obtained from another frozen CB unit by CD34(+)positive selection and culture expansion; the other three patients received uncultured highly purified haploidentical CD34(+) cells. The first two patients had DNA from the culture expanded CB cells detected only for a few days around day +11 when the absolute neutrophil count (ANC) was >200/microl; thereafter and when the ANC was <500/microl, only donor DNA from the uncultured CB was detected. For the other three patients, DNA analysis showed early and transient granulocyte engraftment of haploidentical cells, progressively replaced by the CB-derived granulocytes. We concluded that: (1) simultaneous infusion of lymphocyte-depleted HLA highly mismatched haematopoietic progenitor cells has not produced unfavourable effects for CBT; (2) the double transplant model is suitable for evaluating the engraftment potential of ex vivocultured CB cells in the clinical setting; (3) the culture conditions used did not result in early recovery of ANC; and (4) co-transplantation of purified uncultured HLA haploidentical CD34(+) cells may reduce the time of neutropenia following CBT.

Infusion of lymphocytes obtained from a donor immunised with the paraprotein idiotype as a treatment in a relapsed myeloma.

A 48-year-old patient with IgA k multiple myeloma received a BMT from his HLA-matched sibling. After transplantation, the disease relapsed. Melphalan therapy followed by reinfusion of haemopoietic blood stem cells collected from the patient led to the improvement of the clinical status, although mixed chimerism and an elevated serum IgA persisted. Successful donor immunisation against an immunogenic preparation of the recipient monoclonal protein was performed before the infusion of donor T lymphocytes (DLI) into the patient. Ten weeks after the lymphocyte infusions, no monoclonal band was evidenced and donor complete chimerism was detected. The patient did not develop GVHD. Once complete remission was achieved, the idiotype vaccine was administered to the patient. Nineteen months after DLI, the patient remains in remission. Bone Marrow Transplantation (2000).

Donor leukocyte infusions for treatment of relapsed acute myeloid leukemia after allogeneic bone marrow transplantation.

A 24-year-old man with acute myelomonocytic leukemia (AML-M4) who relapsed 6 months after an allogeneic BMT was treated with chemotherapy followed by donor leukocyte infusions (4.19 x 10(8) mononuclear cells/kg). The patient developed grade II acute GVHD that responded to therapy with CsA and prednisone. Chimerism was assessed by PCR amplification of the MCT 118 hypervariable region. Fourteen months after donor leukocyte infusions the patient remains in complete remission, without any morphologic and cytogenetic evidence of leukemia, and with a complete donor chimerism. This case shows that donor leukocyte infusions are an effective therapy for some acute myeloid leukemia patients who relapse after allogeneic BMT.

Detection of maternal DNA in umbilical cord blood by polymerase chain reaction amplification of minisatellite sequences.

One of the concerns about the use of cord blood as a source of hematopoietic stem cells for allogeneic transplantation is the possibility of contamination by maternal cells which could cause life-threatening GVHD. We have assessed cord blood contamination using PCR analysis of several minisatellite regions to detect maternal DNA. Eighty mother-cord pairs were obtained for this study. In one case there were no specific maternal alleles at any loci and, therefore, cord blood could not be evaluated. Thus, there was a total of 79 informative cases for the detection of maternal cells in the fetal circulation. In most cases, the level of detection was between 0.5 and 1%. We detected maternal DNA in the cord blood sample in only one case (1.26%), and the analysis of dilution experiments led to an estimate of 0.5-1% maternal cells. In conclusion, using PCR amplification of hypervariable regions, maternal DNA is very rarely detected in the cord blood collected at birth, although this approach has a relatively low level of sensitivity.

Rhinocerebral mucormycosis following donor leukocyte infusion: successful treatment with liposomal amphotericin B and surgical debridement.

A 24-year-old male developed cytogenetic relapse of chronic myeloid leukemia (CML) four years after allogeneic BMT. After a year of treatment with IFN-alpha, he achieved a partial cytogenetic response. Treatment with donor leukocyte infusions (DLI) was given (total dose 1 x 10(8) T lymphocytes/kg). Two months later, he developed acute GVHD (skin and liver), that improved with CsA and methylprednisolone and resulted in cytogenetic remission with complete donor chimerism. One month later he developed rhinocerebral mucormycosis and was successfully treated with surgical debridement and liposomal amphotericin B (total dose 12 g). This is the first case of mucormycosis described after DLI.

Histological changes in glioblastoma after intratumoral injection of autologous lymphocytes and human lymphoblastoid interferon.

Seven glioblastomas were studied between 1 and 6 months after intratumoral injection of autologous lymphocytes and human lymphoblastoid interferon. Morphological study showed a great number of lymphocytes within the tumor tissue, and interactions between lymphocytes and glioblastoma cells, suggesting a killing phenomenon. These data support the potential usefulness of adoptive immunotherapy in patients with glioblastoma by means of intratumoral administration of activated lymphoid cells.

Cord blood transplants supported by co-infusion of mobilized hematopoietic stem cells from a third-party donor.

This open label clinical study provides updated evaluation of the strategy of single unit cord blood transplants (CBTs) with co-infusion of third-party donor (TPD) mobilized hematopoietic stem cells (MHSC). Fifty-five adults with high-risk hematological malignancies, median age 34 years (16-60 years) and weight 70 kg (43-95 kg), received CBTs (median 2.39 x 10(7) total nucleated cell (TNC) per kg and 0.11 x 10(6) CD34+ per kg) and TPD-MHSC (median 2.4 x 10(6) CD34+ per kg and 3.2 x 10(3) CD3+ per kg). Median time to ANC and to CB-ANC >0.5 x 10(9)/l as well as to full CB-chimerism was 10, 21 and 44 days, with maximum cumulative incidences (MCI) of 0.96, 0.95 and 0.91. Median time to unsupported platelets >20 x 10(9)/l was 32 days (MCI 0.78). MCI for grades I-IV and III-IV acute GVHD (aGVHD) were 0.62 and 0.11; 12 of 41 patients (29%) who are at risk developed chronic GVHD, becoming severely extensive in three patients. Relapses occurred in seven patients (MCI=0.17). The main causes of morbi-mortality were post-engraftment infections. CMV reactivations were the most frequent, their incidence declining after the fourth month. Five-year overall survival and disease-free survival (Kaplan-Meier) were 56 % and 47% (63% and 54% for patients