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Resume Introduction L’urticaire chronique (UC) est une maladie invalidante. Les patients et leurs medecins sont souvent demandeurs d’une explication etiologique. Des etudes ont suggere que l’UC etait liee a la presence d’auto-anticorps seriques qui peuvent etre mis en evidence par un test intradermique au serum autologue (TSA). Objectifs Confirmer la frequence de positivite du TSA dans l’UC et evaluer l’utilite pratique de ce test en recherchant une correlation entre son resultat et la severite de l’urticaire (resistance aux anti-H1, score d’activite, duree d’evolution). Patients et methodes Les patients adresses pour UC entre le 1 octobre 2001 et le 31 mars 2005 ont fait l’objet d’explorations standardisees incluant interrogatoire, examen clinique, tests physiques, NFS, VS, CRP et recherche d’anti-thyroperoxydase (anti-TPO). Il leur a ete propose en outre de realiser un TSA. Les criteres d’inclusion etaient l’absence de cause trouvee a l’urticaire, l’acceptation du protocole incluant le prelevement sanguin puis l’injection du TSA trois semaines plus tard, la realisation des serodiagnostics pour les hepatites B et C et le VIH, l’arret des anti-H1 et des corticoides. Les criteres d’exclusion etaient l’existence d’un dermographisme, d’une urticaire physique, d’une vascularite urticarienne, l’absence d’arret des anti-H1 ou des corticoides. En avril 2006, nous avons contacte par courrier les patients pour preciser l’evolution de leur urticaire et de son traitement. Resultats Soixante-quatorze patients (67 femmes et sept hommes) âges en moyenne de 43 ans ont ete inclus. Le TSA etait positif chez 43 patients (58 %,) et negatif chez 31 (42 %). La seule difference notable, bien que statistiquement non significative (p = 0,23), entre les patients a TSA positif et ceux a TSA negatif, etait la presence d’anti-TPO chez 24 % des premiers contre 12 % des seconds. La survenue d’un angio-œdeme, la duree de l’UC, son score de severite, l’inefficacite relative des anti-H1, le recours aux corticoides ou a la ciclosporine etaient comparables dans les deux groupes. Discussion Le TSA est positif chez les patients ayant une UC, dans 58 % des cas de notre etude et dans la litterature avec une sensibilite estimee autour de 70 % et une specificite proche de 80 %. Mais si ce test peut aider a faire mieux comprendre la maladie au patient et ainsi a eviter de chercher vainement une autre cause ou de prescrire inutilement l’eviction d’allergenes alimentaires, notre etude montre que le resultat positif ou negatif du TSA n’a aucune repercussion sur le traitement et n’est pas associe a une plus grande severite de l’urticaire ni a une plus grande resistance au traitement. De ce fait, nous ne recommandons donc pas la pratique de cet examen en routine.

Les decouvertes de cibles therapeutiques et d'outils diagnostiques nouveaux sont aujourd'hui le fruit du developpement considerable de la biologie cellulaire et moleculaire et des biotechnologies. Apres le sequencage du genome humain, la pertinence des donnees obtenues par les techniques a haut debit, notamment celles issues de la genomique et de la proteomique, depend en premier lieu de l'assurance-qualite et de la tracabilite du materiel biologique a partir duquel sont effectuees les recherches, ainsi que des annotations qui qualifient les echantillons. Les Centres de Ressources Biologiques, qui incluent les biobanques, les cryobanques et les tumorotheques sont donc des infrastructures essentielles pour la decouverte d'outils innovants pour la prevention, le diagnostic et le traitement des maladies, et la mise au point de nouveaux medicaments. La constitution d'un reseau de Centres de Ressources Biologiques Humains sur des bases ethiques et juridiques harmonisees constitue aujourd'hui un enjeu majeur pour le developpement de la recherche medicale en Europe.

Recognition of the utility of skin allografts in the treatment of critical burn injuries goes back six decades. Since World War II, the use of allografts in burn treatment has been accepted practice. The material has proved to be highly adaptable as burn care has evolved through the years. However, a number of difficulties have emerged which limit utilisation of these grafts, including the possibility of disease transmission, absence of permanent take and shortage of supply [1]. Nevertheless, as has been clearly shown in two French surveys [2,3], the use of allografts remains unchallenged, despite improvements in local and general burn care. With the aims of improving availability and of reducing delays and administrative charges, a skin bank was planned and installed in Tours in 1986. Allograft procurement, as a component of multi-organ donations, was assigned to the burn surgeon on call; responsibility for skin preparation and preservation was assumed by the Tissue Bank Department of the Blood Centre. This arrangement proved to be effective and remains the same today. During the first 4 years of operation, supplies of allograft were generally adequate. Agreements were made with the other skin banks in France, especially Paris, to transfer supplies when appropriate to alleviate shortages and to distribute surfeits (Fig. 1). Recently, the number of procurements has declined markedly; in fact no allograft skin has been acquired locally for the last 3 years. It must be stressed that this development is not linked to disinterest, increased load of work of the teams involved, nor failure of the organisation. There has been a general decline in donors in France, which is especially obvious in Tours. Confronted by this situation, the Burn Centre in association with the Tissue Bank in Tours decided in 1998 to obtain glycerolised skin allografts from the Euro Skin Bank (GPA) in The Netherlands. This was considered appropriate in view

Molecular mechanisms leading to mobilization of hematopoietic cells from bone marrow (BM) to peripheral blood (PB) involve modulation of adhesion molecule expression on these cells that probably result in changes in adhesion capacity to the microenvironment. However, it is not clear whether these changes involve different stages or lineages of progenitor cells. In this study, we compared the capacity of mature and immature clonogenic progenitor cells from granulocyte colony-stimulating factor (G-CSF)-mobilized PB and normal BM CD34+ cells to adhere to complete marrow stroma. This functional capacity was assessed concurrently with molecular expression on CD34+ cells of integrins VLA-4 (alpha 4/beta 1), VLA-5 (alpha 5/beta 1), and LFA-1 (alpha L/beta 2) by interindividual (between mobilized PB and normal BM) and intraindividual (between mobilized PB and steady-state BM and PB in the same patient) analysis. The proportion of adherent clonogenic progenitor cells was significantly lower in PB than in BM, not only for total progenitor cells but also for mature and immature progenitor cells, and the difference was found for granulocytic and particularly for erythroid lineages. The lower adhesion capacity of PB CD34+ cells to stroma was associated with decreased expression (signal/noise MFI ratio) of integrin alpha 4, beta 1, alpha L, and beta 2 chains whereas that of alpha 5 chain did not differ from BM cells with the lowest expression level. Similar differences in integrin expression levels were also found between mobilized PB and steady-state BM CD34+ cells in the same patient except for the alpha L chain. Moreover, we demonstrated for the first time a strong positive correlation between mobilizing capacity and expression levels on mobilized CD34+ cells for the LFA-1 alpha L chain but not for VLA-4 or VLA-5. In conclusion, the decreased adhesion capacity of mobilized PB progenitor cells to stroma involves different maturation stages and different lineages. This is associated with down-regulation of integrins VLA-4 and LFA-1, but mobilizing capacity appears positively correlated with LFA-1 levels.

Post-graft hematopoiesis is characterized by long-term quantitative deficiency in marrow progenitor cells in both autologous and allogenic settings. In order to evaluate the function of post-graft progenitor cells, the proliferative capacity of marrow CD34(+) cells was evaluated in 10 patients 6 months after autologous bone marrow transplantation (ABMT) for non-Hodgkin's lymphoma and compared to that of 10 patients before ABMT and 10 normal controls. Immuno-selected CD34(+) cells were cultured for 7 days in liquid serum-free medium with a combination of early-acting GF consisting of stem cell factor, IL-3 and IL-1beta. Clonogenic efficiency of unselected cells for CFU-GM and BFU-E was decreased in post-graft patients compared to pre-graft and control patients. However, clonogenic efficiency of selected CD34(+) cells for CFU-GM was not different in post-graft, pre-graft and control patients but BFU-E values of post-graft patients remained lower than those of control patients. Decreased percentages of CD34(+) CD38(-) cells were observed in both post-graft and pre-graft patients while those of CD34(+) c-kit(+) cells were similar in all three patient groups. After 7-day liquid culture, expansion yields of total progenitor cells were significantly lower in post-graft patients (147 +/- 28%) than in pre-graft (255 +/- 27%) and control patients (246 +/- 23%). Post-graft deficiency in progenitor cell expansion was particularly marked for BFU-E (61 +/- 24%) compared to pre-graft patients (220 +/- 82%) and to controls (349 +/- 82%). These results indicate impaired proliferative potential of marrow CD34(+) cells several months after ABMT involving erythroid progenitor cells and/or commitment towards erythroid lineage from a more immature stage (pre-CFU).

Objective The present study was designed to compare directly the frequency of circulating LTC-IC and E-LTC-IC mobilized in peripheral blood (PB) after chemotherapy supported by either G-CSF (PB-G) or GM-CSF (PB-GM) in comparison to steady-state bone marrow (BM) and PB (PB-ST) values in the same patients. Materials and Methods Long-term cultures (LTC) were performed from 20 patients with malignant lymphoma at saturating cell concentrations to assess bulk progenitor cell production and by limiting dilution assay (LDA) to measure both frequency of LTC-IC and their proliferative and differentiation capacities. Results While CFC production in bulk LTC was higher at weeks 3–5 with PB-G than with PB-GM samples, week-5 LTC-IC and week-10 LTC-IC (E-LTC-IC) frequencies were not different using a LDA. However, the number of CFC derived from a single LTC-IC was higher in PB-G patients than in PB-GM patients ( p = 0.01). Interestingly, the frequency of LTC-IC per 1 × 10 5 MNC in mobilized PB positively correlated with one-year marrow progenitor cell recovery, in contrast to the number of autografted CD34 + cells and CFU-GM per kg. Conclusion Both G-CSF and GM-CSF resulted in similar increases in LTC-IC and E-LTC-IC in PB at comparable levels to those present in BM. However, the differentiation capacity of LTC-IC was higher after mobilization with G-CSF than with GM-CSF, suggesting qualitative differences in LTC-IC mobilized with these growth factors.

Some patients unexpectedly fail to mobilize sufficient numbers of haematopoietic progenitor cells (HPCs) into the peripheral blood for autologous transplantation. Considering the important role of the chemokine stromal cell-derived factor 1 (SDF-1) in HPC homing, we investigated a possible relationship between SDF1 gene polymorphism and HPC mobilization capacity in 63 patients with malignancy. Some 67% of the good mobilizers (or = 50 CD34(+) cells/microl) and only 36% of the intermediate/poor mobilizers were SDF1-3'A allele carriers (P = 0.032). In multivariate analysis, the presence of the SDF1-3'A allele was the only factor predictive of good CD34(+) cell mobilization (P = 0.025). This is the first report showing the involvement of genetic factors for HPC mobilization in humans and suggests a significant role for SDF-1 in this process.

Resume Propos Une etude multicentrique (12 centres) a ete realisee a partir des prelevements deellules souches peripheriques effectues en 1996 et 1997, sur 655 patients porteurs d'hemopathies ou de tumeurs malignes, afin de determiner les facteurs predictifs de qualite du greffon dans le but d'optimiser les procedures de recueil. Methodes Des fiches de recueil d'information ont ete completees pour 1 346 seances de cytapherese, soit 655 greffons. Les prelevements ont ete realises apres chimiotherapie d'induction et mise sous facteurs de croissance hematopoietiques (sauf pour les leucemies myeloides chroniques [LMC]). Chaque greffon a ete defini comme de ≪ bonne ≫ ou de ≪ mauvaise ≫ qualite, en fonction du nombre de CD34 + contenu. Un taux de CD34 + ≥2.10 6 /Kg de poids du receveur a ete defini de maniere consensuelle par le groupe de travail, d'apres les donnees de la litterature, comme critere de bon greffon. Les deux sous-groupes (bon greffon contre greffon de moins bonne qualite) ont ete compares en univarie puis grâce a un modele multivarie d'analyse discriminante. Resultats De nombreux parametres sont significativement lies au critere de qualite de la collection ; mais en analyse multivariee, trois facteurs predictifs se degagent: le taux de CD34 + circulantes, le nombre de cytaphereses et le nombre de masses sanguines traitees. Conclusion Le taux de CD34 + circulantes semble un outil decisionnel essentiel presageant de la qualite du greffon et du nombre de cytaphereses necessaires pour obtenir une collection suffisamment riche. Et il apparait necessaire de traiter au moins deux masses sanguines.

Purpose We assessed the usefulness of color Doppler imaging and spectral analysis in monitoring the effect of isovolemic hemodilution on retinal vein occlusion. Methods Color Doppler imaging was used to measure the systolic and diastolic blood flow velocities and the resistance index (RI) in the central retinal artery and the maximum and minimum blood flow velocities in the central retinal vein of affected eyes and contralateral unaffected eyes before and 1 day after isovolemic hemodilution in 70 adults (40 men and 30 women; mean age, 62.4 ± 13.7 years) who presented with retinal vein occlusion. Results With hemodilution, the hematocrit value was decreased from 41.8 ± 3.0 to 33.0 ± 3.0. In central retinal vein occlusion, the pretreatment arterial and venous flow velocities of affected eyes were significantly lower than those of unaffected eyes. After hemodilution in eyes affected by ischemic occlusion, a significant improvement in the RI in the central retinal artery was observed without changes in venous flow velocity. After hemodilution in eyes affected by nonischemic occlusion, a significant increase in maximal venous flow velocity only was noted. In branch retinal vein occlusion, no significant difference was observed in the RI between affected and unaffected eyes. After hemodilution, an improvement in RI was noted only for affected eyes. Conclusions The present results confirm the potential value of Doppler analysis in monitoring the impact of hemodilution on arterial and venous velocity in patients with central retinal vein occlusion. © 1998 John Wiley & Sons, Inc. J Clin Ultrasound26:119–124, 1998.

Continuing advances in life sciences and medical research are the result of the remarkable achievements of cell and molecular biology. In the post-sequencing era, the quality of the huge amount of data continuously generated by biotechnology, i.e. genomics, proteomics and high-throughput screening, depends on the quality assurance and the traceability of the original biological materials, including the annotations linked to these materials. Thus, biological resource centres are key infrastructures supporting biotechnology, bioprocessing and the development of new approaches in the prevention, diagnosis and treatment of diseases.

To evaluate the modification of pharmacodynamic parameters induced by the administration of L-asparaginase loaded into red blood cells, 13 patients received a single dose of L-asparaginase internalised into the carrier. The enzyme was loaded using a reversible lysis-resealing process. The dose per patient ranged from 30 to 200 i.u. kg-1. Considerable heterogeneity occurred between patients. the level of L-asparaginase circulating after 24 h represented 47% of the total injected dose as compared to 74.8% for red blood cells (RBCs). However, the half-life of the enzyme remaining in the circulation was very similar to that of the RBC carrier, i.e. 29 days and 27 days, respectively, compared with 8-24 h for the free enzyme. Sustained elimination of plasma L-asparagine occurred, the duration of which was dependent on the injected dose. A single injection of 30.i.u.kg-1 was sufficient to eliminate plasma L-asparagine over 10 days. With 150-200 IU.kg-1 the elimination period was extended to 50 days. These data show that the use of RBCs as carriers of L-asparaginase greatly improves the pharmacodynamic parameters of the drug.

Resume Les ulceres cutanes rebelles sont souvent refractaires a l'ensemble des therapeutiques classiques. L'Hemodilution Normovolemique a ete proposee des 1984 par Guenneguez et Ouvry, qui ont obtenu de bons resultats sur une petite serie de patients. Mais il s'agissait d'un protocole unicentrique utilisant une technique manuelle d'Hemodilution. Grâce aux Separateurs de cellules automatiques, il est maintenant possible d'effectuer ce geste, facilement et en toute securite chez des patients ambulatoires. Les donnees recentes concernant la physiopathologie des ulceres cutanes semblent confirmer le benefice que l'Hemodilution Normovolemique devrait apporter au traitement de cette pathologie. C'est pourquoi, un groupe de reflexion multidisciplinaire s'est cree afin de proposer une etude multicentrique evaluant le traitement des ulceres cutanes rebelles par Hemodilution Normovolemique durable.

Twenty six adult patients with low grade nodular non Hodgkin's lymphoma (NHL) were treated with autologous bone marrow transplantation. Conditioning regimen was BEAM-BEAC in 15 patients and TBI + Cyclophosphamide in 11 patients. Twenty one patients were grafted with haematopoietic stem cells, 12 after bone marrow purging and five with peripheral blood stem cells (PBSC). Two patients were treated in CR1 of leukemic phase, six in PR 1 and eighteen in sensitive relapse. With a median follow-up of 30 months, the actuarial survival is 91% and actuarial event free survival 67%. These data confirm some interest of ABMT in the treatment of low grade follicular NHL.

Objectifs Mettre en place une solution de partage des informations utiles aux acteurs de sante qui restent, apres contractualisation avec l’EFS, garant de la confidentialite et de la qualite et qui beneficient d’une identification securisee possible via le repertoire des professionnels de sante de l’ARS ; augmenter la securite transfusionnelle des patients ; s’integrer a l’environnement numerique du systeme de soins (DIP, ABM…) ; limiter le nombre d’examens redondants inutiles ; participer a l’amelioration de la gestion de l’identite des patients. Contenus et fonctionnalites en phase 1 Mise a disposition, en consultation et edition, du socle transfusionnel comportant les principales donnees IH (groupe ABOD RHK ± phenotypage etendu ± phenotypage elargi, RAI et identifications) et HLA/HPA/HNA, les protocoles et consignes transfusionnelles, les PSL transfuses, le signalement du risque de non disponibilite de PSL du fait d’une association rare d’antigenes erythrocytaires (par ex. D- C- e-) ou d’un phenotype erythrocytaire rare, d’un Allo-Ac d’interet transfusionnel, d’une greffe de CSH. En phase 2 Edition des comptes-rendus de resultats d’examens et envoi de donnees IH et HLA ILP. En phase 3 Integration des prescriptions connectees de PSL et analyses, lien avec les declarations d’effets indesirables receveurs d’interet ulterieur transfusionnel et ouverture possible a des donnees externes. Mise en œuvre Projet de l’EFS deploye par iterations regionales successives en commencant par l’EFS IDF avec lancement du marche IDF en mars 2015 pour un deploiement operationnel en 2015–2016 des 3 phases ; serveur d’identite national permettant l’interoperabilite des DPCT regionaux en 2017.

Summary. The roles of the chemokine stromal-derived factor 1 (SDF-1) and the matrix metalloproteinase 9 (MMP-9) in haematopoietic progenitor cell (HPC) mobilization are still unclear, particularly when patients are mobilized by granulocyte colony-stimulating factor (G-CSF) plus chemotherapy. We determined bone marrow (BM) and peripheral blood (PB) plasma levels of SDF-1, together with CXC-chemokine receptor 4 (CXCR-4) expression on CD34+ cells, and interleukin 8 (IL-8) and MMP-9 in 55 patients mobilized for autologous PB transplantation compared with 10 normal BM and PB samples. Plasma samples were tested at steady state (SS-) and after mobilization by cyclophosphamide and G-CSF administration (M-). SDF-1, CXCR-4, IL-8 and MMP-9 levels were significantly lower in SS- and M-PB than in SS-BM. Differences in SDF-1 levels between SS-PB and SS-BM were also observed after mobilization. We showed for the first time a clear relationship between the levels of circulating HPC, both at steady state and after mobilization, and those of secreted MMP-9 but not of SDF-1 or IL-8. However, a negative correlation was observed between mobilizing capacity and CXCR-4 expression on CD34+ cells. These findings suggest that G-CSF-induced mobilization of HPC from BM involves MMP-9, without reversing the positive gradient of SDF-1 between BM and PB.

Objective The aim of this study was designed to compare the in vivo long-term hematopoietic potential of bone marrow and peripheral blood grafts. Materials and Methods Marrow progenitor cell recovery was assessed for up to 4 years in 227 patients. One hundred patients were treated for malignant lymphomas by autologous bone marrow transplantation (BMT) and 127 by peripheral blood progenitor cell transplantation (PBPCT). Results Marrow progenitor cell counts were decreased for several years with both bone marrow and peripheral blood grafts. They were not different according to the origin of the graft, despite the reduced duration of peripheral blood cell recovery observed after PBPCT. Granulocyte colony-stimulating factor (G-CSF) used for PB graft mobilization and after transplantation resulted in faster neutrophil recovery compared to granulocyte-macrophage colony-stimulating factor (GM-CSF) with no evidence of decreased marrow progenitor cell recoveries. On the other hand, postgraft administration of GM-CSF enhanced long-term colony-forming unit granulocyte-macrophage reconstitution only after BMT. Factors that influenced marrow progenitor cell reconstitution have been identified by univariate and multivariate analysis: age, gender, type of lymphoma, and postgraft administration of hematopoietic growth factors (HGF) for the whole patient group; gender, graft progenitor cell yields, and type of HGF (G-CSF vs GM-CSF) for the PBPCT group; and only type of HGF for the BMT group. Conclusion Despite faster peripheral blood cell recovery, persistent deficiency of marrow progenitor cells was found several years after PBPCT, as observed after BMT. G-CSF–mobilized PBPCT resulted in faster neutrophil recovery compared to GM-CSF mobilization, with no difference in long-term hematopoietic reconstitution.

The aim of this study was to examine whether the severe prolonged deficiency in marrow clonogenic progenitor cells reported after autologous stem cell transplantation (ASCT) is associated with impairment of the primitive progenitor cell compartment. We performed Dexter-type marrow cultures and limiting dilution assays with CD34(+) cells from patients 1 year and/or later after autografting with peripheral blood stem cells for non-Hodgkin's lymphoma (NHL). Flow cytometric analysis was used to assess the CD38 antigen expression and apoptotic state (7-ADD(-)/annexin-V(+) cells) of the CD34(+) cell population. We found a dramatic decrease in both clonogenic progenitor cell production and frequency of long term culture-initiating cells (LTC-IC) in all the patients tested at 1 year, even in those displaying normal progenitor cell frequency. Surprisingly, the clonogenic capacity of each LTC-IC was not increased. Flow cytometric analysis of the CD34(+) cell population confirmed this quantitative defect, with a reduction in the CD38(dim/neg) cell population but no increase in apoptosis. This defect did not improve over time up to 4 years after transplantation. In addition, qualitative abnormalities were revealed, demonstrated by decreased CD34 antigen expression, together with impaired differentiating properties of LTC-IC toward erythroid lineage at 1 year. This study indicates that both quantitative and qualitative abnormalities of the primitive progenitor cell compartment are a constant feature up to 4 years after autologous stem cell transplantation.

Hemotherapy in developed countries is safer than ever but the present perception of society seems different than in the past. As Isbister wrote, during recent years, as blood safety has been progressively improved, the legal risks have been increasing. The first task is to study and define the risks for each activity. For me the management of risks may be achieved through the management of quality. The confidence of society in blood transfusion can probably best be obtained through a kind of recognised external quality assurance. I intend to describe our targets, our difficulties and the factors that have made our certification a success. I shall focus on several points: • what quality represents; • what our haemapheresis service is; • what ISO 9002 standards are and why we choose them; • our difficulties and the factors of success.

High doses of carboplatin or cisplatin combined with cyclophosphamide and etoposide followed by autologous bone marrow transplantation (ABMT) rescue have been used in the treatment of testicular tumors that have had a bad prognosis. Unusual cutaneous complications, evoking radiation-induced dermatitis, have been seen in two of eight patients with the same regimen. This new type of toxicity seems to be related to high-dose combination chemotherapies. Good results in the treatment of patients with testicular tumors on relapse who continue to respond to chemotherapy lead to the extension of this type of schedule and cutaneous toxicity will probably develop.

Abnormal hematopoiesis, including a deficiency of marrow progenitors and particularly of erythroid progenitors, has been described after autologous stem cell transplantation (ASCT), persisting for several years. In order to explain this deficiency, a resistance of marrow progenitors to stem cell factor (SCF) after ASCT was investigated. Marrow samples were harvested from pregraft patients at graft collection prior to ASCT, transplanted patients 6-24 months after high-dose therapy and control patients. CD34+ cells were cultured in a serum-free clonogenic assay with increasing doses of SCF. The clonogenic efficiency without SCF was lower for BFU-E in treated groups than in controls, whereas it was not different for CFU-GM. With increasing doses of SCF a dose-dependent effect was found on the numbers of both CFU-GM and BFU-E in all groups, although the maximal number of BFU-E remained lower in treated groups. However, the SCF dose that induced 50% of maximal BFU-E growth (D50) was similar in all groups. Furthermore, a dose-dependent effect on the size of BFU-E was found in all groups, with no difference in the proportion of large colonies. Thus, clonogenic erythroid progenitors from patients who have received myelotoxic treatments remain sensitive to SCF, with no evidence for a chemotherapy-related resistance.

Marrow stromal cells were evaluated several months after autologous BMT for their capacity to support both normal hemopoiesis and secrete the main growth factors involved in its control, G-CSF, GM-CSF, IL-3 and SCF. Stromal layers (SL) were obtained by long-term marrow cultures (LTMC) established from 15 patients (9 with hematologic malignancies and 6 with solid tumors) 3 months after autologous BMT and were compared to pre-graft patients. After irradiation, both post-graft and pre-graft SL were recharged with the same inoculum of normal marrow cells. As compared to pre-graft values, CFU-GM production on post-graft SL was significantly increased during the first 2 weeks of culture whereas it was decreased from week 3 to week 8. These findings were only observed in patients with hematologic malignancies and not in patients with solid tumors. Growth factor secretion was evaluated by ELISA in the supernatants of unstimulated and IL-1-stimulated SL from 10 post-graft patients, 13 pre-graft patients and 5 normal controls. In any group of patients, IL-3 was undetectable either spontaneously or after IL-1-stimulation. As compared to controls, secretion by IL-1-stimulated SL was not different for GM-CSF in pre- and post-graft patients but tended to be decreased for G-CSF in post-graft patients. SCF secretion, which was not induced by IL-1, appeared dramatically decreased in both pre- and post-graft patients. The capacity of post-graft SL to support CFU-GM growth in LTMC was correlated at week 1 with G-CSF secretion and from week 3 to week 8 with SCF secretion. These results suggest that microenvironment remains qualitatively damaged several months after BMT involving a decreased capacity both to support early hemopoiesis and to secrete SCF, particularly in patients grafted for hemopoietic malignancies.

The use of recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) as an adjunct to autologous bone marrow transplantation (ABMT) or peripheral blood progenitor cell (PBPC) transplantation was evaluated in 59 lymphoma patients. Patients were divided into three groups. In group I (n = 21) patients received rhGM-CSF (5 micrograms/kg daily) at the time of PBPC collection and during the recovery phase post-infusion. In group II (n = 12) patients received rhGM-CSF as an adjunct to ABMT. In group III (n = 26) they were grafted with bone marrow without rhGM-CSF. Administration of rhGM-CSF (groups I and II) significantly reduced the time to myeloid engraftment, the number of febrile days and the median duration of antibiotics administration and of hospital stay when compared with the group in which patients did not receive rhGM-CSF. The only difference between ABMT and PBPC, given with rhGM-CSF support, was observed in the duration of hospitalization (group Igroup II, P0.05). These data show that rhGM-CSF is highly effective in reducing the duration of aplasia following BMT and PBPC transfusion, and there appears to be little difference in efficacy between these techniques, provided that patients also receive rhGM-CSF.

Objective: A pilot clinical study was conducted to evaluate the toxicity of a single dose of L-asparaginase loaded in red blood cells (RBCs). Methods: Thirteen patients received a single dose of L-asparaginase in the range 30–200 IU ⋅ kg−1. The enzyme was loaded in one autologous blood unit using a lysis-resealing process. A control population of 33 patients receiving L-asparaginase intravenously were tested in parallel. IgG, IgM and IgE class anti-L-asparaginase antibodies were detected using specific radioimmunoassays. Results: L-Asparaginase pharmacodynamic parameters may be greatly improved by administration of the drug after internalisation in RBCs as compared to intravenous injection of free drug. The drug elimination was prolonged and similar to that of circulating carrier. After one injection of 30 IU ⋅ kg−1, plasma L-asparagine was eliminated in 10 days and this was extended to 50 days for 150–200 IU ⋅ kg−1. The drug was well tolerated and only transient variations were observed for some of the biological parameters measured. We did not reach the maximum tolerable dose (MTD) of L-asparaginase loaded in RBCs. No significant clinical toxicity was detected. In particular, no immune adverse effects were observed. Conclusion: This study opens new perspectives for the clinical utilisation of L-asparaginase. This mode of administration of the drug is able to improve pharmacodynamic parameters and enzymic efficacy and to increase the general tolerance of the treatment.

Summary. We assessed the effect of antitumoural therapy at intensive doses on the haemopoietic system using long-term marrow cultures (LTMC) established from 33 patients (25 with haematological diseases and eight with solid tumours) after autologous bone marrow transplantation (ABMT). When compared to 42 pre-graft patients, a decreased CFU-GM Production and a defect in stromal layer (SL) confluence were found after ABMT on day 90 but also on day 365. However, these abnormalities were observed only in patients with haematological diseases and no differences between pre-graft and post-graft results were found in patients with solid tumours. Among the patients with haematological diseases, on day 90 those with acute lymphoid leukaemias showed lower CFU-GM production whereas patients with non-Hodgkin's lymphomas developed more frequently subconfluent or confluent SL. Other factors studied such as sex, patient age, disease status, marrow purging and post-graft administration of growth factors did not appear to influence post-graft LTMC results. Multivariate analysis including all the patients has shown (a) that solid tumours were associated with higher CFU-GM production, and (b) that conditioning regimens with total body irradiation (TBI) or busulfan led more frequently to non-confluent SL. In conclusion, high-dose therapy followed by ABMT can induce a persistent impairment of the stem cell and stromal cell compartments, particularly in patients with haematological diseases conditioned with TBI, despite the absence of any alloimmune reaction and post-graft immunosuppressive therapy.

A study of CFU-GM sensitivity to mafosfamide was carried out in 67 candidates for ABMT. A lethal dose 95 (LD95) was calculated from the dose-response curve. Despite standardized treatment conditions of marrow buffy-coat cells, LD95 values that were normally distributed (median 100 micrograms/ml; mean +/- SEM 95.6 +/- 4.0 micrograms/ml) varied considerably from patient to patient (range 30-160 micrograms/ml). Three independent factors appeared to confer greater sensitivity of CFU-GM: low patient age, low CFU-GM rate in treated cells and prolonged delay of CFU-GM sensitivity test from last chemotherapy course. In contrast, sex, pathology, disease status, number of previous chemotherapies and use of CY before the test did not influence CFU-GM sensitivity. Forty-six patients were autografted with mafosfamide-treated marrows according to their LD95. The mean percentage of CFU-GM effectively recovered after graft purging was 3.96 +/- 2.09%. All patients engrafted well and their peripheral blood cell recoveries were correlated with graft CFU-GM content evaluated before purging but not after purging or after freezing. In multivariate analysis, this parameter remained the only factor predicting hematopoietic recovery among other patient variables such as sex, age, pathology, disease status, previous chemotherapies or TBI in conditioning regimens. In a subgroup of 39 patients with lymphoid malignancies compared with 25 patients autografted for non-Hodgkin's lymphomas with unpurged marrows, the delay in days (medians) was similar for leukocytes1 x 10(9)/l (19 vs 17 days) and for neutrophils0.5 x 10(9)/l (18 vs 17 days) while it was longer for platelets50 x 10(9)/l (34 vs 128 days).(ABSTRACT TRUNCATED AT 250 WORDS)

For several years, various enzymes loaded into red blood cells have been tried as circulating bioreactors. However, due to the bacterial or fungal origin of these enzymes, the in vivo use of such a bioreactor is questionable as concerned with potential human immunological adverse consequences. L-Asparaginase (L-Asp) is a well known enzyme used as a chemotherapeutic agent, but its use leads to important immunological intolerance reactions after intraveinous administration. Therefore it was a good candidate for such a study. The aim of this paper is to present some results for the immunological consequences following the injection of free or RBC loaded L-Asp in human patients.

Hematopoietic growth factors (GF) have greatly facilitated the mobilization and collection of peripheral blood (PB) stem cells. However, some patients fail to mobilize sufficient numbers of hematopoietic progenitor cells (HPC) to ensure rapid and durable engraftment. Interactions between the chemokine SDF-1 and its receptor CXCR4 have been shown to play a central role in the homing of HPC. To investigate a possible additional role of SDF-1 in the mobilization of CD34+ progenitor cells into PB, we examined the association of SDF1 gene polymorphism with circulating CD34+ cell counts after GF-priming. 63 patients (pts) were studied (48 malignant lymphoma, 8 myeloma and 7 solid tumors), after mobilization with G-CSF alone (n=11), chemotherapy (CT) and G-CSF (n=36) or GM-CSF (n=16). PB CD34+ cell counts were assessed when leukocytes reached 1,000/μl and graft collection performed when CD34+ counts reached 20/μl. The SDF1 3′G/A genotyping was performed on PB mononuclear cell DNA by polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) which can detect the G to A mutation. Out of the 63 pts, 21 were able to mobilize 50 CD34+ cells/μl or more in PB. Their characteristics were not statistically different from others in terms of diagnosis, age, sex, number of previous CT courses, previous radiotherapy, marrow involvement or mobilizing regimens. Distribution of SDF1 genotypes in normal controls and in patients according to their mobilizing capacity was as follows: These results provide evidence for an association of the SDF1 3′A allele with more efficient mobilization of CD34+ progenitor cells into PB and suggest an involvement of SDF-1 in mobilization mechanisms.

Non seminomatous germ cell testicular tumors (NSGCTT) is a very chemosensitive cancer; vepeside, cyclophosphamide or ifosfamide and cisplatin are the most effective drugs. Carboplatin is also active with a lack of nephrotoxicity, but there is probably a cross-resistance to cisplatin. High dose chemotherapy with autologous bone marrow transplantation is able to cure poor prognosis testicular cancers in first partial remission or in sensitive relapse. We report one case of non seminomatous testicular cancer which was progressive after chemotherapy with cisplatin and vepeside or teniposide, and which is in prolonged complete remission after conventional chemotherapy (vepeside, ifosfamide, carboplatin) and high dose chemotherapy (carboplatin, vepeside, cyclophosphamide) with autologous bone marrow transplantation.

Background and aim of the study: Autologous stem cell transplant (ASCT) has been shown to be an effective treatment for first-line and relapsed follicular lymphomas (FL). But no long term molecular remission was described. The aim of this retrospective study was to determine the clinical and molecular outcome of patients with FL who received ASCT during a 12-year period. Method: All patients who underwent ASCT for first-line or relapsed FL between January 1992 and December 2004 were included. Overall survival (OS) and progression-free survival (PFS) were estimated by Kaplan-Meier method and cumulative incidence of nonrelapse mortality (NRM), with relapse as a competing event, by Fine and Gray method. Patient characteristics: Seventy-one patients with a median age of 45 years and a median follow-up of 108 months were analysed. The majority were of the subtype grade 1 (57 %), had a high tumour burden (50 %) and were treated in firstline (52 %). After an anthracyclin-based induction regimen, 12 patients were in first complete remission (CR), 25 in first very good partial response (VGPR), 8 in second CR and 26 in second VGPR. They received BCNU, Etoposide, Aracytine, Melphalan (BEAM in 58 %) or Cyclophosphamide, total body irradiation (42 %) as conditioning for the ASCT. The majority of them received an unpurged graft (58%). Results : Thirty-eight patients were alive, 24 without progression between 4 and 12 years; 31 patients had died, 7 without progression. A total of 38 patients (55 %) developed recurrent lymphoma. Median OS was estimated at 8 years and 4 months. The ten-year PFS and the ten-year OS were 33 % and 47 %, respectively and the tenyear molecular PFS was 37 %. There was an apparent plateau on the remission duration curve at 32 % at 72 months and on the molecular remission duration curve at 37 % at 80 months. A plateau on the OS curve seemed to emerge at 41 % from the tenth year. Patients who received a purged graft had better OS and better PFS (median OS not reached versus 50 months, p = 0.08; median PFS not reached versus 22 months, p = 0.035) Three patients developed a secondary neoplasm and two a secondary myelodysplastic syndrome. The 10-year non-relapse mortality (NRM) was 20 %. Conclusion : This long follow-up study showed a plateau on the PFS and on the molecular PFS curves, suggesting that a selected group of patients might be cured by ASCT.