Since about 15 years intensive chemotherapy followed by autologous bone marrow transplantation has been used on the basis of "dose-response" principle to treat certain children with tumours of sombre prognosis. At present, the main indications for this method are metastatif neuroblastoma in less than one-year old children, non-Hodgkin's malignant lymphomas in partial remission or relapse, refractory or recurrent Hodgkin's disease and some peculiar forms of Wilms' tumour. In other tumours, such as rhabdomyosarcoma, Ewing's sarcoma or brain tumours, the indications have not yet been clearly determined. The treatment must be administered as part of multicentre French or European trials conducted in specialized centres. The practice and application of autologous bone marrow transplantation are being revolutionized by the availability of haematopoietic growth factors and the development of the peripheral blood stem cells reinjection technique. Genic therapy will soon have major repercussions in this field.

The CD 34 antigen is a glycoprotein found on the surface of hematopoietic stem cells and early committed progenitors. The CE 34 stem cells from 14 samples of bone marrow, cord blood or leucapheresis were isolated using a positive selection procedure involving an anti CD 34 biotinylated monoclonal antibody and an avidin immunoabsorption device. Results showed that in 60 percent of samples, the positively-selected fractions contained more than 70 percent CD 34 cells. Concentration in CFU-GM and BFU-E progenitors increased 15 and 26 fold respectively in the CE 34 enriched samples. Long-term culture of two samples demonstrated that nearly all of the most immature progenitors were recovered in the procedure. These preliminary results showed that the positive selection technique of CD 34 hematopoietic stem cells is now available for use in autologous or allogeneic hematopoietic stem cell transplantation.

3'-azido-3'-deoxythymidine (zidovudine-AZT) was the first clinically approved reverse transcriptase inhibitor for the treatment of acquired immunodeficiency syndrome (AIDS) and recently, 2',3'-dideoxyinosine (didanosine-ddI) has also been approved in France. These nucleoside analogs have no intrinsic anti-HIV activity and must be metabolized to their respective 5'-triphosphates by means of kinases, nucleotidases, or other activating enzymes present naturally in cells. The presence and activity of the necessary intracellular enzymes for activation of nucleoside analogs is highly dependent on species, cell type, and cell cycle stage, illustrating the importance of cellular functions in the mechanism(s) of action or toxicity of nucleoside analogs. Although the apparent plasma elimination half-life of the parent drug varies between 1 and 2 hours, the active triphosphate derivatives have intracellular half-lives between 4 and 12 h with 2',3'-dideoxyadenosine-5'-triphosphate (ddATP), the active component of ddl being one of the most stable with an intracellular half-life of 8 to 12 hours which result in relatively infrequent dosing as compared to other classes of potential anti-HIV drugs under development. The multifactorial mechanism(s) of toxicity of this class of drugs likely explains the different spectrums of toxicity observed with the various nucleoside analogs, and demonstrates the uniqueness of each compound. Recently, AZT-resistant strains have been isolated from AZT-treated patients, probably reflecting a sequential acquisition of amino-acid mutations in the HIV-RT. Of importance, cross-resistance was demonstrated with other compounds with an 3'-azido group, but no cross-resistance was detected with either ddl of 2',3'-dideoxycytidine.(ABSTRACT TRUNCATED AT 250 WORDS)

Patients with recurrent lymphoma have a poor prognosis, even when a new remission can be obtained. In follicular lymphomas this remission is shorter. In aggressive lymphomas less than 10% of the patients survive for 5 years. Salvage treatments include drugs that were not administered in the induction of treatment. In follicular lymphomas new drugs which interfere with adenosine deaminase are very promising. In view of the bad results of rescue treatment and of the good sensitivity of lymphomas, even when recurrent, to radiotherapy and chemotherapy, intensive treatments combined with allogeneic or autologous graft of haematopoietic stem cells have been tested, and the results obtained seem to be better than those of conventional treatments. The position of these new treatments will be more clearly defined when the results of randomized studies are known. However, the prognosis of recurrent lymphoma can be further improved by reducing the toxicity of intensive treatments with the use of haematopoietic growth factors, by ridding the graft of malignant cells and by stimulating the post-graft immune defence to diminish the incidence of recurrences.

The term "aggressive lymphoma" applies to the intermediate severity and high malignancy lymphomas of the international classifications. Despite their heterogeneity, the lymphomas of this group should be cured by modern treatments. The therapeutic approach depends on the presence or absence of adverse prognostic factors. When these factors are absent cure can be obtained in 80 to 90% of the cases by chemotherapy or chemo-radiotherapy. The basic treatment rests on chemotherapy adjusted to the severity of the disease. In its least severe forms the different chemotherapeutic combinations available have about the same effectiveness, and it is rational to choose the least toxic of them. Intensive treatments are reserved to patients with several factors of poor prognosis. In such cases, complete remission rates may be close to 65%, but the percentage of long-term survival is only 40%, and new therapeutic approaches are required. At present, in addition to intensified conventional therapy made possible by growth factors, the possibility or early intensification with haematopoietic stem cells autografts is being studied. Intensive treatments can only be given to young subjects. Lymphoblastic and Burkitt's must be approached differently when medullary or meningeal tissues are invaded.

The control of granulocyte and macrophage formation and function offers many insights into the developmental processes involved in hemopoiesis. Regulation of granulocyte and macrophage populations is mainly controlled by the actions of four glycoprotein regulators--the colony stimulating factors. These factors exhibit some redundancy in their actions but each is capable of regulating multiple events in the production, differentiation commitment, maturation and functional activities of the cells. The design of this regulatory system is such that it can ensure precise cell production under basal conditions but is also able to achieve large variations in cell production in response to emergency situations.

The neural crest is part of a larger embryonic structure, the neural folds, belonging to the neural primordium of the Vertebrate embryo. The neural fold is formed by the anterior and lateral ridges of the neural anlage, which fuse mediodorsally when the neural tube closes. Anteriorly, the epithelium of the neural fold does not convert into mesenchymal cells and yields Rathke's pouch, the olfactory organ and the epithelium of the mouth roof, of the upper lip and of the frontal region of the head. From the level of the diencephalon (at the level of the epiphysis) downwards the neural fold epithelium undergoes the epitheliomesenchymal transition and yields the neural crest cells which become later on highly diversified and form various structures and tissues throughout the body. A large amount of data have shown that the environmental cues exerted on crest cells both during their migration and when they have reached their target sites are critical in determining their fate. In order to understand the mechanisms through which environmental factors influence crest cell differentiation, the developmental capacities of single neural crest cells were investigated at different time points of their ontogeny. Single cell cultures of crest cells have revealed that already at the migratory stage the neural crest is made up of cells at different states of determination. In particular, the analysis of clones obtained from single cell cultures of cephalic migratory crest cells has shown that, although many clonogenic cells are multipotent to varying degrees, others are committed to give rise to one single derivative. Totipotent progenitors able to generate representatives of virtually all the phenotypes (neuronal, glial, melanocytic and mesectodermal) encountered in cephalic neural crest derivatives were also found. We proposed that they represent stem cells analogous to those which in the hemopoietic system generate the various types of blood cells. The neural crest stem cell gives rise to diverse progenitors that become progressively restricted in their potentialities according to an essentially stochastic mechanism while dividing during and after completion of the migration process. Similar cloning experiments of crest cells that have already reached their target organs, i. e. sensory ganglia or enteric plexuses, showed that the phenotypic repertoire expressed by crest-derived cells decreases with increasing embryonic age. Efforts are made to elucidate the nature of the factors which influence either the survival and/or the differentiation of neural crest cells in the various types of environments in which they evolve.(ABSTRACT TRUNCATED AT 400 WORDS)

Human fetal blood tissues implanted in the SCID immuno-deficient mouse are tolerated and develop considerably. The bone marrow maintains its autonomous hematopoietic function for several months. Conversely, thymic lymphopoiesis can continue over the long term only in the presence of an appropriate source of stem cells. This in vivo replica of the human blood system can be used to test the hematogenic abilities of candidate populations of human hematopoietic stem cells, as suggested by the hematogenic reconstitution of human thymus and marrow in SCID mice obtained from purified CD34+ precursors. In the same model, hematogenic activity of CD34+ cells was limited to the sub-population of these precursors coexpressing the antigen Thy-1. These results, combined with those of in vitro analyses, suggest that the population of CD34+ Thy-1+ components of fetal liver and of the bone marrow is extremely rich in human hematopoietic stem cells.

Using avian chimeras, we have demonstrated earlier that the stem cells seeding the definitive hemopoietic organs form within the embryo rather than in the yolk sac. We now report experimental evidence indicating that hemopoietic progenitors appear in mouse embryos in the para-aortic splanchnopleura, a location similar to the one that produces stem cells in avian embryos. This structure obtained from E8.5 embryos was grafted under the kidney capsule of adult SCID mice. One compartment of the B lymphoid system became reconstituted with cells derived from the graft, that were identified with genetic and antigenic markers. In vitro data are also in favor of the production of progenitors from this structure. Finally a strategy designed to understand the developmental links between the endothelial network and hemopoietic cells is described. It is based on the expression patterns of two protooncogenes, c-ets1 and c-myb, activated during the amplification period of each of these lineages.

Fifty cases of chronic pure red cell aplasia in adults (idiopathic in 32 cases, associated with chronic lymphocytic leukaemia in 18) were followed up after studying their erythroblastic precursors in vitro. Analysis of response to immunomodulators confirmed the existence, in patients with idiopathic chronic pure red cell aplasia, of a relationship between in vitro behaviour and obtention of a remission: patients with normal differentiation of erythroblasts consistently responded to treatments, whereas treatments were usually ineffective when the erythroblasts did not differentiate in vitro. When in vitro differentiation was below normal, responses to treatments were varied. In patients with chronic lymphocytic leukaemia, the erythroblastic precursors were normal in 15/18 cases, and response to immunomodulators was observed in 13/18 cases. A second objective of this study was to determine, on a large series, the most adequate immunomodulator treatment. In idiopathic chronic pure red cell aplasia the most efficient therapy was corticosteroids and cyclophosphamide administered separately or together. The antilymphocyte serum gave disappointing results. In pure red cell aplasia associated with leukaemia cyclophosphamide was the most frequently effective drug, with corticosteroids taking second rank; however, infections were frequent. The treatments to be considered when both corticosteroids and cyclophosphamide fail are discussed.

The stochastic model of stem cell differentiation is in accord with experimental findings but does not explain hematopoietic homeostasis. We discuss how positive and negative controls by cytokines and inhibitors could maintain homeostasis, even though progenitor commitment towards the various hematopoietic lineages may be stochastic.

In our study, the rate of pregnancy by transfer and puncture was not significantly different in unexplained and in tubal infertility, but the mean number of transferred embryos was significantly higher in the first group. To explain these data, we compared the quality of embryos in 32 punctures realized among 29 women with unexplained infertility and in 171 punctures planned among 156 women with tubal infertility. The percentage of embryos with 4 or more blastomeres was significantly lower in the unexplained infertility group than in the pure tubal infertility group.

Hybrid materials for use as bone substitutes are composed of a synthetic substance to which autologous osteogenic cells are bound. Identification of osteogenic stem cells among bone marrow stromal cells was followed by the demonstration that a host of growth factors, hormones, and cytokines have effects on cultured osteoblasts. Porous ceramics have been developed as vectors for osteogenic cells previously grown in vitro. Porous hydroxyapatite beads bearing stromal cells were used to fill experimental ulnar defects in dogs. The cells were first grown on flat supports then on the three-dimensional hydroxyapatite network. Invasion by histologically identifiable bone tissue was seen in the stroma cell-bearing implants but not in control implants without stromal cells. Materials containing osteogenic cells may prove extremely useful since bone substitutes including hydroxyapatite ceramics are implanted in sites where osteogenic cells are scarce.

In 1987, Prather et al. have performed the first embryo cloning by nuclear transfer in the bovine species. Since, many researchers try to develop and to apply the technique. While the enucleation of the recipient oocyte, the injection of the donor blastomere and the fusion procedure are now well controlled, on the other hand, maturation and activation as the development and freezing of the cloned embryos need to be more investigated. The cloned embryo is more fragile. An increase in embryonic mortality is observed after transfer in a recipient cow.

Gene therapy is a new field of biomedical research just entering clinical practice. Recombinant retroviruses are at present the best vectors for gene transfer into a permanently dividing tissue like the hematopoietic tissue. The ex vivo strategy consists of an autologous transplantation of genetically modified cells. Although the hematopoietic tissue can be manipulated with comparative ease, its development and its structure are rather complex. Therefore, the different sources of hematopoietic tissue and the diverse cell types represent a wide array of targets for gene transfer. In vivo studies in mice have shown that the hematopoietic stem cells can be efficiently transduced by retroviral vectors carrying therapeutic genes. In contrast, preclinical studies in large outbred animals indicate that only a small proportion of stem cells can be transduced, although much better results can be obtained with primitive progenitors detectable in vitro. However, some clinical trials have already shown interesting and useful results.

This paper reviews the present status of the transplantation of autologous stem cells and the recent development of cytokines. Since 1977, autologous bone marrow transplantation has considerably expanded its scope. In France, more than 1,300 autologous bone marrow transplantations (ABMT) are performed per year, in parallel to 560 allogeneic bone marrow transplantation (BMT). ABMT has become a routine procedure for the treatment of patients with acute leukemias (results very similar to those of BMT), lymphomas, myelomas, as well as some selected tumors (breast and testicular cancers). In parallel to the development of ABMT and BMT, more than 30 cytokines have been individualized. Many of them are presently tested in clinical trials and some (GMCSF, GCSF, Erythropoietin, Interferons) are marketed. One of the most recent development which combines the two fields includes the selection and purification of immature normal stem cells recognized by the CD 34 monoclonal antibodies, their expansion in vitro by liquid culture in presence of multiple cytokines and gene transfer before autografting. These fields are reviewed.