Overview on novel strategies and current guidelines for hematopoietic stem cell mobilisation and collection.

The success of the autologous stem cell transplantation is strictly related to an adequate hematopoietic stem cell mobilization and collection. The minimum threshold for a successful mobilization is currently defined as 2 × 106/kg CD34+ cells. However, the optimal stem cell mobilization strategy is still controversial. The availability of plerixafor, a selective and reversible CXCR4 inhibitor, has been associated with an higher use of chemo-free protocols by many centres. In the near future, it is conceivable that artificial intelligence may became more accurate and comprehensive, possibly guiding clinicians in choosing the optimal mobilisation treatment for the various patients undergoing hematopoietic stem cell transplantation. Machine learning-based scoring models may be the basis for the development of "intelligent" mobilisation algorithms.

Postremission Consolidation by Autologous Hematopoietic Cell Transplantation (HCT) for Acute Myeloid Leukemia in First Complete Remission (CR) and Negative Implications for Subsequent Allogeneic HCT in Second CR: A Study by the Acute Leukemia Working Party of the European Society for Blood and Marrow Transplantation (EBMT).

After autologous hematopoietic cell transplantation (HCT) in the first complete remission (CR1), patients with acute myeloid leukemia (AML) may relapse and undergo allogeneic HCT in the second complete remission (CR2). The aim of this study was to analyze the outcome of allogeneic HCT performed in CR2 comparing patients with prior consolidation by autologous HCT versus patients with chemotherapy consolidation. Included were 2619 adults with allogeneic HCT in CR2 from 2000 to 2017 with (n = 417) or without (n = 2202) prior autologous HCT. Patient groups were not entirely comparable; patients with prior autologous HCT were younger, had less often a favorable cytogenetic profile, had more commonly donors other than matched siblings, and more often received reduced-intensity conditioning. In multivariate analysis, nonrelapse mortality risks in patients with prior autologous HCT were 1.34 (1.07 to 1.67; P = .01) after adjustment for age, cytogenetic risk, transplant year, donor, conditioning intensity, sex matching, interval diagnosis-relapse, and relapse-allogeneic HCT as compared with chemotherapy consolidation. Similarly, risks of events in leukemia-free survival and graft-versus-host disease, relapse-free survival were higher with prior autologous HCT, 1.17 (1.01 to 1.35), P = .03 and 1.18 (1.03 to 1.35), P = .02, respectively. Risk of death was also higher, 1.13 (0.97 to 1.32), P = .1, but this was not significant. Postremission consolidation with autologous HCT for AML in CR1 increases toxicity of subsequent allogeneic HCT in CR2.

WITHDRAWN: Convalescent plasma, an apheresis research project targeting and motivating the fully recovered COVID 19 patients: A rousing message of clinical benefit to both donors and recipients alike.

The Publisher regrets that this article is an accidental duplication of an article that has already been published, https://doi.org/10.1016/j.transci.2020.102794. The duplicate article has therefore been withdrawn. The full Elsevier Policy on Article Withdrawal can be found at https://www.elsevier.com/about/our-business/policies/article-withdrawal.

NEW HORIZONS ON STEM CELL CRYOPRESERVATION THROUGH THE ARTIFICIAL EYES OF CD 34+, USING MODERN FLOW CYTOMETRY TOOLS.

Hematopoietic stem cell (HSC) cryopreservation is a critical step in autologous and cord blood transplantation (CBT). In most circumstances, cryopreservation is performed in a mixture containing dimethyl sulfoxide (DMSO), since DMSO is necessary to secure cell viability. Most centers use a controlled rate (slow) freezing before the long-term storage at vapor phase liquid nitrogen (LN2) temperatures (≤ -160 °C). The primary objectives for laboratories supporting HSCT programs are to provide secure storage for leukapheresis and cord blood products, and to adequately characterize the functional properties of the grafts before their infusion. In the autologous setting, the large majority of the published results dealt with the assessment of the graft before cryopreservation. On the contrary, in CBT, before a CB unit is released, a sample obtained from a contiguous segment of that CB unit needs to be tested to verify HLA type and cell viability. The effects of graft handling, cryopreservation, storage and thawing on the recovery of CD34+ cells needs to be carefully analyzed and standardized on a global level. Some technical unresolved issues still limit the application of the ISHAGE derived single platform flow cytometry protocol for the assessment of the thawed material; based on these considerations, an adaptation of both the acquisition setting and the gating strategyis necessary for reliable measurement of CD34-expressing HSC in cryopreserved grafts. Artificial intelligence applied to "big data" may provide a new tool for improving advanced processing procedures and quality management guidelines in this area of investigation.

Manipulation, and cryopreservation of autologous peripheral blood stem cell products in Italy: A survey by GITMO, SIDEM and GIIMA societies.

There is considerable heterogeneity in manipulation and cryopreservation of hematopoietic stem cells (HSC) for autologous HSC transplantation across Europe and Italy. To better address this point, three Italian Scientific Societies (GITMO- Gruppo Italiano per il Trapianto di Midollo Osseo; SIDEM- Società Italiana Emaferesi e Manipolazione Cellulare; and GIIMA- Gruppo Italiano Interdisciplinare Manipolazione e Aferesi per Terapie Cellulari), in collaboration with the Competent Authority "National Transplant Center" (CNT) sent to 85 Italian transplant centers (TC) a survey, which included 12 questions related to the most critical elements in graft processing. Fifty-nine centers (70 %) responded to the questionnaire. Overall, this survey demonstrates that the majority (>90 %) of responding TC used standardized procedures for HSC processing; however, an intercenter heterogeneity was clearly documented in several standard operating procedures adopted by different TC. These results seem to suggest that further standardization and efforts are needed to provide recommendations and guidelines on HSC manipulation, cryopreservation and functional assessment of cryopreserved material for autologous HSCT.

[Cultured platelets]. / Plaquettes sanguines de culture : état de l'art.

Blood platelets are anucleated elements of the blood. With a diameter of 2 to 3 µm, they are the smallest elements of blood. While their main role is to stop or prevent bleeding, they are also involved in other functions, such as immunity, inflammation or tumour progression. The development of biotechnology and the knowledge acquired about the mechanisms regulating the biogenesis of platelets makes the production of cultured platelets a viable option today. Consequently, this type of product could have its place in meeting a number of transfusion challenges such as alloimmunization or refractory states. However, culture yields remain low and many hurdles still need to be overcome before considering an application in transfusion. This article reviews the rationale for the production of cultured platelets for transfusion and summarizes the main advances in the field while highlighting its limitations.

Development of adaptive immune effector therapies in solid tumors.

State-of-the-art treatment strategies have drastically ameliorated the outcome of patients affected by cancer. However, resistant and recurrent solid tumors are generally nonresponsive to conventional therapies. A central factor in the sequence of events that lead to cancer is an alteration in antitumor immune surveillance, which results in failure to recognize and eliminate the transformed tumor cell. A greater understanding of the dysregulation and evasion of the immune system in the evolution and progression of cancer provides the basis for improved therapies. Targeted strategies, such as T-cell therapy, not only generally spare normal tissues, but also use alternative antineoplastic mechanisms that synergize with other therapeutics. Despite encouraging success in hematologic malignancies, adaptive cellular therapies for solid tumors face unique challenges because of the immunosuppressive tumor microenvironment, and the hurdle of T-cell trafficking within scarcely accessible tumor sites. This review provides a brief overview of current cellular therapeutic strategies for solid tumors, research carried out to increase efficacy and safety, and results from ongoing clinical trials.

Salvage high-dose chemotherapy in female patients with relapsed/refractory germ-cell tumors: a retrospective analysis of the European Group for Blood and Marrow Transplantation (EBMT).

BACKGROUND: High-dose chemotherapy (HDC) with hematopoietic progenitor cell transplantation is a standard option for relapsed/refractory testicular germ-cell tumor (GCT), but only few data have been reported in female patients with GCT. We conducted a retrospective analysis of female patients with GCT treated with HDC and registered with the European Society for Blood and Marrow Transplantation. PATIENTS AND METHODS: Between 1985 and 2013, 60 registered female patients with GCT, median age 27 years (range 15-48), were treated with salvage HDC. Forty patients (67%) had primary ovarian GCT, 8 (13%) mediastinal, 7 (12%) retroperitoneal and 5 (8%) other primary sites/unknown. Twenty-two patients (37%) received HDC as second-line therapy, 29 (48%) as third-line, and 9 (15%) as fourth- to sixth-line. Nine of 60 patients (15%) received HDC as late-intensification with no evidence of metastasis before HDC. The conditioning HDC regimens comprised carboplatin in 51 of 60 cases (85%), and consisted of a single HDC cycle in 31 cases (52%), a multi-cycle HDC regimen in 29 (48%). RESULTS: Nine cases who underwent late intensification HDC were not evaluable for response. Of the other 51 assessable patients, 17 (33%) achieved a complete response (CR), 8 (16%) a marker-negative partial remission (PRm-), 5 (10%) a marker-positive partial remission, 5 (10%) stable disease, and 13 (25%) progressive disease. There were 3 toxic deaths (6%). With an overall median follow-up of 14 months (range 1-219), 7 of 9 (78%) patients with late intensification and 18 of the 25 patients (72%) achieving a CR/PRm- following HDC were free of relapse/progression. In total, 25 of 60 patients (42%) were progression-free following HDC at a median follow-up of 87 months (range 3-219 months). CONCLUSIONS: Salvage HDC based on carboplatin represents a therapeutic option for female patients with relapsed/refractory GCT.

Repercussions of preterm birth on symptoms of asthma, allergic diseases and pulmonary function, 6-14 years later.

BACKGROUND: Prevalence of allergic diseases and impaired pulmonary function may be high in children born prematurely. This study aimed to assess pulmonary function and prevalence of asthma, atopic diseases and allergic sensitisation in these patients. METHODS: A cross-sectional study was conducted with children aged 6-14 years who were born prematurely with birth weight <2000g from January 2008 to May 2011. Exclusion criteria were: major malformations, or acute respiratory disorders. The International Study of Asthma and Allergies in Childhood questionnaire was applied followed by allergic skin prick test and spirometry. RESULTS: The study included 84 children aged 9.3±2.3 years born at mean gestational age of 31.8±2.4 weeks. The prevalence of current asthma was 25%, more severe asthma was 15.5%; rhinitis was 38.1%; flexural eczema was 8.3%; and a positive skin-prick test was 69.6%. Frequencies of children with values <80% of predicted were: FVC (8.3%), FEV1 (22.6%), and FEV1/FVC ratio (16.7%). Prevalence of children with FEF25-75% <70% of the predicted value was 32.4%, positive bronchodilator response was observed in 20.5% of cases, and altered pulmonary function in 42.9%. Factors associated with altered pulmonary function were oxygen dependency at 28 days of life (OR: 4.213, p=0.021), the presence of wheezing in childhood (OR: 5.979, p=0.014) and infant's height (OR: 0.945, p=0.005). CONCLUSIONS: There was a high prevalence of severe asthma, allergic sensitisation, and altered pulmonary function among children and adolescents born prematurely. Bronchopulmonary dysplasia and a history of wheezing were risk factors for altered pulmonary function.

Prevalence and associated factors for asthma in Brazilian and Japanese schoolchildren living in the city of São Paulo, Brazil.

OBJECTIVES: Ethnic background interferes on the prevalence of asthma among schoolchildren (4 to 9 years old, SC) born and living in São Paulo, Brazil. METHODS: International Study of Asthma and Allergy in Childhood (ISAAC)'s written standard and complementary questionnaires were applied to SC (similar socioeconomic status) living in the city of São Paulo: no-Japanese Brazilian (NJB, N = 306) and Japanese Brazilian (third generation, born in Brazil, from Japanese families with no miscegenation, JB, N = 258). RESULTS: The prevalence of current asthma was significantly higher among NJB in comparison to JB (22.2% vs 14.7%, respectively). To have rhinitis and to exercise less than once/week were risk factors for both groups of children. CONCLUSION: Although both groups were apparently exposed to the same environment, other cultural differences do not allow us to conclude about the ethnic component having greater influence than the environment in the development of asthma in these individuals.

Mutation frequencies of GNAQ, GNA11, BAP1, SF3B1, EIF1AX and TERT in uveal melanoma: detection of an activating mutation in the TERT gene promoter in a single case of uveal melanoma.

BACKGROUND: Uveal melanoma is the most frequent primary tumour of the eye. It is molecularly clearly distinct from cutaneous melanoma and shows a different pattern of driver mutations. The influence of sunlight ultraviolet (UV) exposure on the aetiology of uveal melanoma is a matter of debate. The recent identification of driver mutations in the promoter of the telomerase reverse transcriptase (TERT) gene with UV-induced cytidine-to-thymidine transitions in cutaneous melanoma prompted us to investigate whether these mutations also occur in uveal melanoma. METHODS: We analysed 50 cases of uveal melanoma obtained from enucleation surgery for mutations in the genes GNAQ, GNA11, BAP1, SF3B1, EIFAX1 and TERT, measured gene expression using microarrays and analysed gene copy numbers by SNP arrays. RESULTS: We detected a TERT mutation in only one case of a 57-year-old white male patient with clinical and histopathological features typical for uveal melanoma. The tumour showed mutations in GNA11 and EIF1AX that are typical for uveal melanoma and absent from cutaneous melanoma. No mutations were detected in GNAQ, BAP1 and SF3B1 that are frequently mutated in uveal melanoma. Both copies of chromosome 3 were retained. Several tumours among which the one carrying the TERT promoter mutation showed elevated TERT expression. Consistent with previous reports, GNAQ is inversely associated with chromosome 3 monosomy and metastasis. BAP1 mutations are significantly associated with chromosome 3 monosomy but not with relapse. CONCLUSION: These data indicate that TERT mutations are rare in uveal melanoma. No conclusion can be drawn on their potential influence on tumour progression.

Impact of an active lifestyle on heart rate variability and oxidative stress markers in offspring of hypertensives.

Familial history of hypertension is associated with autonomic dysfunction and increase in blood pressure (BP). However, an active lifestyle has been found to improve a number of health outcomes and reduce all-cause mortality. The aim of the present study was to investigate the effects of an active lifestyle on hemodynamics, heart rate variability (HRV) and oxidative stress markers in offspring of hypertensive parents. One hundred twenty-seven subjects were assigned into four groups: sedentary offspring of normotensives (S-ON) or hypertensives (S-OH); and physically active offspring of normotensives (A-ON) or hypertensives (A-OH). Diastolic BP and heart rate were reduced in the physically active groups when compared to S-OH group. A-ON and A-OH groups presented increased values of RR total variance when compared to the sedentary ones (A-ON: 4,912 ± 538 vs. S-ON: 2,354 ± 159; A-OH: 3,112 ± 236 vs. S-OH: 2,232 ± 241 ms2). Cardiac sympato-vagal balance (LF/HF), systemic hydrogen peroxide and superoxide anion were markedly increased in S-OH group when compared to all other studied groups. Additionally, important correlations were observed between LF/HF with diastolic BP (r = 0.30) and hydrogen peroxide (r = 0.41). Thus, our findings seem to confirm an early autonomic dysfunction in offspring of hypertensive parents, which was associated with a systemic increase in reactive oxygen species and blood pressure. However, our most important finding lies in the attenuation of such disorders in offspring of physically active hypertensives, thus emphasizing the importance of a physically active lifestyle in the prevention of early disorders that may be associated with onset of hypertension.

Functional properties of human platelets derived in vitro from CD34+ cells.

The in vitro production of blood platelets for transfusion purposes is an important goal in the context of a sustained demand for controlled products free of infectious, immune and inflammatory risks. The aim of this study was to characterize human platelets derived from CD34+ progenitors and to evaluate their hemostatic properties. These cultured platelets exhibited a typical discoid morphology despite an enlarged size and expressed normal levels of the major surface glycoproteins. They aggregated in response to ADP and a thrombin receptor agonist peptide (TRAP). After infusion into NSG mice, cultured and native platelets circulated with a similar 24 h half-life. Notably, the level of circulating cultured platelets remained constant during the first two hours following infusion. During this period of time their size decreased to reach normal values, probably due to their remodeling in the pulmonary circulation, as evidenced by the presence of numerous twisted platelet elements in the lungs. Finally, cultured platelets were capable of limiting blood loss in a bleeding assay performed in thrombocytopenic mice. In conclusion, we show here that cultured platelets derived from human CD34+ cells display the properties required for use in transfusion, opening the way to clinical trials.

Loss of Thy-1 (CD90) antigen expression on mesenchymal stromal cells from hematologic malignancies is induced by in vitro angiogenic stimuli and is associated with peculiar functional and phenotypic characteristics.

BACKGROUND: Little is known about human mesenchymal stromal cell (hMSC) phenotypic and functional subsets in response to environmental stimuli. The strategy used in this study focused on defining hMSC functional subpopulations based in particular on their Thy-1 (CD90) antigen (Ag) surface expression. METHODS: The effect of different in vitro microenvironmental conditions on the isolation and expansion of bone marrow-derived (BM) hMSC from hematologic malignancies (HM) and normal samples (NS) was assayed. hMSC clonogenic and differentiation potential, phenotypic profile and long-term capacity to sustain in vitro hemopoiesis were considered in relation to the different expansion protocols. RESULTS: The results showed that angiogenic supplements used in combination with low serum content gave rise to the appearance of Thy-1(-) HM-MSC with high proliferative potential, capable of restoring the typical HM stromal impairment. The expression of the CD271 was partially maintained. We further report an enhancement towards the osteogenic and adipogenic differentiation capacity by the Thy-1(-) HM-MSC subset. Despite the angiogenic treatment, the Thy-1(-) MSC stopped short of full endothelial differentiation. DISCUSSION: In this paper we provide evidence that in vitro angiogenic stimuli generate HM-MSC lacking CD90 Ag expression. The Thy-1(-) MSC subset is characterized by peculiar functional and phenotypic characteristics, thus supporting the role played by the microenvironment in selecting particular hMSC subsets maintaining normal tissue homeostasis or inducing pathologic processes.

A functional role for soluble HLA-G antigens in immune modulation mediated by mesenchymal stromal cells.

BACKGROUND: It has been suggested that soluble factors produced by bone marrow (BM) mesenchymal stromal cells (MSC) play a fundamental role in mediating immune modulation. HLA-G antigens (Ag) are major histocompatibility complex (MHC) class Ib molecules characterized by a limited polymorphism and a splicing mechanism that regulates the production of membrane-bound and soluble isoforms. Interleukin-10 (IL-10) cytokine is one of the main up-modulators of soluble HLA-G Ag (sHLA-G) production by CD14+ peripheral blood monocyte cells and increased IL-10 levels are reported to be associated with MSC immune modulation. METHODS: We investigated, by specific enzyme-linked immunosorbent assay (ELISA), the possible role of sHLA-G molecules in the inhibition of the peripheral blood mononuclear cell (PBMC) response to phytohemagglutinin (PHA) mediated by MSC from different sources. RESULTS: There was a significant correlation between the presence of increased levels of sHLA-G and IL-10 in the MSC/PBMC/PHA culture supernatants and lymphoproliferative inhibition. Neutralizing experiments performed with monoclonal Ab directed against HLA-G and IL-10 molecules confirmed the inhibitory ability of sHLA-G Ag. Furthermore, exogenous IL-10 induced sHLA-G molecule secretion by MSC alone in a polymorphic way, while a longitudinal analysis confirmed the loss of MSC inhibitory functions in relation to in vitro MSC aging. DISCUSSION: Overall the results obtained suggest a functional role for sHLA-G molecules in inhibiting the PBMC response mediated by MSC. Moreover, the ability of IL-10 to induce sHLA-G Ag production by MSC alone could be proposed as a marker of MSC functional ability.

Decreased thrombotic tendency in mouse models of the Bernard-Soulier syndrome.

OBJECTIVE: The platelet glycoprotein (GP)Ib-V-IX complex is a receptor required for normal hemostasis deficient in the Bernard-Soulier bleeding disorder. To evaluate the consequences of GPIb-V-IX deficiency in thrombosis we generated mouse models of the disease by targeting the GPIb beta subunit. METHODS AND RESULTS: Complete deletion (GPIb beta-/-) or an intracellular truncation (GPIb beta deltaIC-/-) reproduced typical and variant forms of Bernard-Soulier, with absent and partial (20%) expression of the complex on the platelet surface. Both strains exhibited thrombocytopenia and enlarged platelets with abnormal microtubular structures but normal granule composition. They exhibited prolonged tail bleeding times, which were less pronounced in GPIb beta deltaIC-/-. Decreased thrombus formation was observed after blood perfusion over a collagen coated surface at high shear. Resistance to vascular occlusion and an abnormal thrombus composition were observed in a model of FeCl3-induced lesion of carotid arteries. In a model of laser-induced lesion of mesenteric arterioles, thrombosis was strongly reduced in GPIb beta-/- mice, while a more modest effect was observed in GPIb beta deltaIC-/- animals. Finally, the two strains were protected against death in a model of systemic thromboembolism. CONCLUSIONS: This study provides in vivo evidence of a decreased thrombotic tendency linked to defective platelet GPIb-V-IX in mouse models of Bernard-Soulier syndrome.

Microtubule plus-end tracking Adenopolyposis Coli negatively regulates proplatelet formation.

Platelets are produced upon profound reorganization of mature megakaryocytes (MK) leading to proplatelet elongation and release into the blood stream, a process termed thrombopoiesis. This highly dynamic process requires microtubules (MT) reorganization by mechanisms that are still incompletely understood. Adenomatous polyposis coli (APC) is a microtubule plus-end tracking protein involved in the regulation of MT in a number of cell systems and its inactivation has been reported to alter hematopoiesis. The aim of our study was to investigate the role of APC in megakaryopoiesis and the final steps of platelet formation. Down-regulation of APC in cultured human MK by RNA interference increased endomitosis and the proportion of cells able to extend proplatelets (68.8% (shAPC1) and 52.5% (shAPC2) vs 28.1% in the control). Similarly an increased ploidy and amplification of the proplatelet network were observed in MK differentiated from Lin- cells of mice with APC-deficiency in the MK lineage. In accordance, these mice exhibited increased platelet counts when compared to wild type mice (1,323 ± 111 vs 919 ± 52 platelets/µL; n = 12 p 0.0033**). Their platelets had a normal size, ultrastructure and number of microtubules coils and their main functions were also preserved. Loss of APC resulted in lower levels of acetylated tubulin and decreased activation of the Wnt signaling pathway. Thus, APC appears as an important regulator of proplatelet formation and overall thrombopoiesis.

Pancreatic neuroendocrine tumors in MEN1 disease: a mono-centric longitudinal and prognostic study.

PURPOSE: Multiple endocrine neoplasia type 1 (MEN1) is an inherited endocrine neoplastic syndrome associated with a greater risk of endocrine tumor development like pancreatic neuroendocrine tumors (p-NET), with different clinical characteristics from sporadic ones. This paper aims to compare clinical, hystological and morphological aspects of p-NET in patients affected from MEN1 (MEN1+) and not-affected ones (MEN1-). METHODS: We performed a retrospective observational study. Data was collected between December 2010 and December 2015, including patients with a histological diagnosis of p-NET and radiological imaging. We compared clinical, histological, radiological, and prognostic aspects of MEN+ p-NET with MEN-1 p-NET. RESULTS: Of the 45 patients enrolled, 13 MEN1+ and 21 MEN1- cases were analyzed. Frequency of not secreting p-NETs and insulin secreting p-NETs, histopathological grades and Ki67 expression were superimposable between MEN1+ and MEN1- patients. MEN1+ pNETs are more often multicentric compared to MEN1- pNETs. Frequency of liver and nodes metastatic spread was higher in MEN1- p-NET compared to MEN1+ p-NET. Analyzing p-NET according to the disease outcome, we found that recovered and stable p-NETs in MEN1+ patients, compared to MEN1- cases, are diagnosed at lower age (p = 0.04/p = 0.002) and that are more frequently multifocal lesions (p = 0.009/p = 0.002). CONCLUSIONS: In our study pNETs in MEN1+ and pNETs in MEN1- don't significantly differ for prognosis but only for clinical features. p-NET stage disease and prognosis can be positively influenced by early diagnosis and screening in index patients' first-degree relatives.