Rescue of K562 cells from MDM2-modulated p53-dependent apoptosis by growth factor-induced differentiation.

The wild-type human MDM2 protooncogene was tested for its ability to modulate apoptotic activity of the de novo expressed p53 tumor suppressor gene in K562 cells. We also studied the role of some cytokines in this phenomenon. K562, a human myeloid leukemia cell line, does not express p53 at the mRNA or protein level. In this study, we stably transfected K562 with eukaryotic vectors containing either normal p53 cDNA (pC53-SN3) or mutated p53 (143Val-->Ala) cDNA (pC53-SCX3). Transfectants expressing WT p53 or those expressing mutant p53 are called K562 SN and K562 SM respectively. Many leukemic cell lines undergo apoptosis when de novo WT p53 is expressed alone. In contrast, while the resulting clones (K562 SN and K562 SM) expressed p53, they did not undergo apoptosis. However, when treated with MDM2 mRNA antisense (MDM2 AS) oligodeoxynucleotides (ODNs), K562 SN demonstrated apoptotic features at both molecular and morphological levels. No change was observed when the other clones (K562 and K562 SM) were treated with MDM2 AS. Apoptosis induced in this manner was associated with a relatively small increase in intracellular calcium [Ca2+]i. Cells cultured in medium previously supplemented with recombinant human (rh) interleukin (IL)-3 and rh-erythropoietin (Epo) did not undergo apoptosis. Moreover, K562 SN cells were induced to differentiate. This differentiation was evaluated by measuring hemoglobin (Hb) level in cellular extracted proteins and by analyzing erythroid colony number and morphology. High Hb synthesis was obtained when K562 SN cells were cultured with cytokines (IL-3 + Epo) combined with MDM2 AS. Our results are consistent with the hypothesis that the function of the proto-oncogene MDM2 is to provide a 'feedback' mechanism for the p53-dependent pathway of apoptosis that could be shunted toward differentiation.

Role of p53 and RB on in vitro growth of normal umbilical cord blood cells.

Human umbilical cord blood (UCB) is rich in hematopoietic stem cells and progenitors and recently has been used in the clinic as an alternative source for graft and marrow repopulation. We tried to determine in vitro the roles of wild-type (wt) p53 and wt RB tumor/growth suppressor genes in the regulation of proliferation and maturation of hematopoietic UCB cells. CD34+ cells, isolated from mononuclear cells of UCB, were cultured in semisolid medium under conditions that favor growth of hematopoietic cells. We studied the level of expression of p53 and RB mRNAs and proteins during cell culture by Northern blot and cytofluorometry analysis, respectively. Sense (S), antisense (AS), or scrambled (missense [MS]) p53 and RB oligodeoxynucleotides (ODNs) were used to study the behavior of these cells in the absence of expression of p53 and/or RB. Adequate doses of p53 or RB ODNs inducing maximal inhibitory effect were used to study the behavior of these cells in the absence of expression of p53 and/or RB. Adequate doses of p53 or RB ODNs inducing maximal inhibitory effect with minimal cellular toxicity were determined. Exposure of CD34+ cells to p53 or AS, RB AS, or both p53 and RB AS but not other ODNs (sense or missense) resulted in a significantly increased number of colony-forming units-granulocyte/macrophage (CFU-GM) induced by interleukin-3 (IL-3) and/or granulocyte-macrophage colony-stimulating factor (GM-CSF). The number of erythroid colonies (CFU-E) and burst-forming units (BFU-E) derived from CD34+ cells in the presence of erythropoietin (Epo) was not significantly increased, whereas the number of such colonies was markedly increased in the presence of IL-3 + EPO upon p53 AS and/or RB AS treatment with hypothesis that wt p53 and RB are proliferation suppressor genes that interfere with normal maturation of hematopoietic cells.

Hematopoietic progenitor content of fetal cord blood collected using citrate-phosphate-dextrose: influence of holding temperature and delays.

Storage of unseparated fetal cord blood collected into citrate-phosphate-dextrose (CPD) for 48 hours has been studied using progenitor cell assays [plasma clot technique with plasma-conditioned medium, erythropoietin (EPO), erythropoietin + IL-3 as growth factors]; collected fetal blood volume was low [61 +/- 4 ml, mean +/- 95% confidence interval (CI)]. The CFU-GM concentration in total cord blood was 2460 +/- 1476 CFU-GM/ml, comparable to normal bone marrow collected into medium RPMI + heparin (2350 +/- 1258); 2 of 12 samples contained fewer than 10(3) CFU-GM. Storage of unseparated cord blood CFU-GM in CPD was better at 4 degrees C (rapid decrease at day (D) 2 from 15 +/- 9 x 10(4) at D0; 14 +/- 11 x 10(4) at D1 to 6 +/- 4 x 10(4) at (D2) than at 22 degrees C (strong decrease since D1: 5 +/- 4 x 10(4) CFU-GM). The CFU-GEMM content remained steady at 4 degrees C: from 16 +/- 16 x 10(4) at D0 to 18 +/- 16 at D2. Cryopreservation and thawing of unfractionated cord blood at D0 did not affect the CFU-FM cloning rates. Whether mature progenitors are more affected than stem cells by storage in CPD remains to be determined.

HLA antigen-sharing in couples with repeated spontaneous abortions and the birthweight of babies in successful pregnancies.

Previously, several groups reported an increase in HLA antigen-sharing in couples suffering from unexplained repeated spontaneous abortions. It was felt necessary to find out if HLA sharing could have any effect on children born after a successful pregnancy. The birthweight figures of children of 76 couples with repeated spontaneous abortions were analyzed. The results show a significantly lower birthweight in babies born from those couples, presenting a high incidence of HLA antigen-sharing, particularly concerning class II antigens.

Treatment of progressive systemic sclerosis by plasma exchange: long-term results in 40 patients.

The efficacy of plasma exchanges (PE) during the course of scleroderma has only been investigated for short periods. The aim of this study was to follow patients over a long enough period to observe the course of the clinical and paraclinical symptoms in the short, medium, and long term. Forty patients, 24 women and 16 men, were treated by PE and observed for 1-3, 3-12 and over 12 months. Immunological, biological and clinical course and any undesirable side effects were evaluated using a detailed questionnaire. Concomitant therapies were reported and most frequently consisted of corticosteroids, colchicine, factor XIII or vasodilators (nifedipine, captopril). The therapeutic effectiveness of PE was assessed on the basis of improvements in cutaneous, digestive, joint, muscular, lung, cardiovascular and renal lesions. Our findings confirmed the effectiveness of short-term PE on scleroderma (52% of the patients improved during the first 3 months). However, this improvement was transient (5% improvement between 3 and 12 months and only 2.5% over 12 months) and limited to the cutaneous and muscular lesions. Thus, PE cannot be recommended for the treatment of progressive systemic sclerosis.

[Neonatal exchange transfusion with irradiated whole blood. Preliminary results]. / L'exsanguino-transfusion néonatale avec sang total irradié. Résultats préliminaires.

Some clinical manifestations following exchange transfusion (ET) could result from graft versus host disease secondary to the introduction of viable foreign T lymphocytes: skin rash, fever, acute and sometimes bloody diarrhea or enterocolitis. Between February 1985 and January 1989 the blood used for 31 ET was irradiated at 40 grays. We compared the manifestations occurring during the days following ET to those occurring after 44 previous ET with non irradiated blood during the period January 1981 to January 1985. From 1981 to 1985, 13 of 44 infants developed problems within 3 days following ET: an erythematous macular skin rash in 4; gastrointestinal manifestations (diarrhea, vomiting and rectal bleeding, necrotizing enterocolitis) in 7; both skin lesions and a gastrointestinal problem in 2. Since 1985, 27 infants had no problems whereas only 4 developed gastrointestinal or cutaneous manifestations: NEC in a preterm infant, abdominal distension with rectal bleeding, fever and petechial rash in 2 infected infants. These data show a dramatic decrease of complications since the irradiation of blood products has been started: 30% with non irradiated, 13% with irradiated blood.

[Cholinergic crisis in a patient with myasthenia treated by plasma exchange and anticholinesterase agents]. / Crises cholinergiques chez un myasthénique traité par échanges plasmatiques et anticholinestérasiques.

A patient with myasthenia receiving treatment with anticholinesterase agents and plasma exchanges for an acute episode, developed three successive periods of neurological deterioration during which plasma cholinesterase levels were determined. The risk of onset of a cholinergic crisis under these circumstances has been reported in the literature but not documented. The accidents in the present case were related to cumulative overdose effects of anticholinesterase agents and depletion of cholinesterase, suggesting caution in the use of anticholinesterase agents when frequent plasma exchanges are being carried out in a patient with myasthenia.

[Therapeutic leukapheresis in a leukostasis syndrome complicating chronic lymphoid leukemia]. / Leucaphérèse thérapeutique au cours d'un syndrome de leucostase compliquant une leucémie lymphoïde chronique.

A 50 year old man with chronic lymphocytic leukemia (CLL) and extreme hyperleukocytosis (600 x 10(9)/liter) presented with a respiratory distress syndrome, congestive heart failure with cardiomegaly, endotoxic shock and anuria. Examination revealed nodes in all areas and hepatosplenomegaly; laboratory studies showed hypoxemia and a chest X-ray diffuse bilateral alveolar infiltrates. He was treated twice by leukapheresis using a cell separator. This procedure removed 10.1 x 10(10) white blood cells with marked clinical improvement and resolution of air-space diseases over the subsequent 48 hours. Despite this unusual indication for leukapheresis in CLL, this procedure may be of value in treating the leukemic phase of CLL with pulmonary leukostasis.

[Major hypertriglyceridemia, associated with severe acute pancreatitis, successfully treated with plasma exchange]. / Hypertriglycéridémie majeure, associée à une pancréatite aiguë grave, traitée avec succès par échanges plasmatiques.

A 27 year-old alcoholic patient with severe acute pancreatitis (4 Ranson's objective prognostic signs), complained of abdominal pain and vomiting and presented fever and rigors. The plasma was of milky turbidity and the concentration of triglycerides (TG) very high (26 Mmol/l, Normal: 3-1.6). Serum pancreatic amylase was elevated (262 U/l, Normal: 10-200). The electrocardiogram (ECG) showed S-T depression. The diagnosis was confirmed by computed tomography which showed a pancreatic phlegmon and a collection in the left pararenal space (Ranson's grade D). After PE on days 1 and 2: the amylase became normal, plasma clear, TG decreased (7.8 Mmol/l), the clinical picture improved and the ECG normal. Assisted ventilation was necessary over 10 days. Pancreatic morphology remained unchanged. The patient was discharged to intensive care on day 18. There was no indication for surgery. The fast drop in TG levels, the precursors of free fatty acids, may have limited their toxicity the pancreas, allowing a difficult stage to be over come and the course of the illness to interrupted. Normalization of the ECG requires emphasis. The association of HG, increased plasma amylase and abdominal pain justifies early iterative PE.

Evidence for a new HLA class II determinant present on cells from HLA-DR1 and/or -DR4 individuals.

Evidence for a new HLA class II specificity is presented. It is recognized by LE serum, which reacts with most DR1 and/or DR4 individuals (r = 0.86). Its frequency in the French population is 0.33. Absorption-elution experiments showed that the serum reactivity was not due to a mixture of anti-DR1 and anti-DR4 antibodies, but to a single antibody population which could be absorbed on and eluted from both DR1(+) or DR4(+) cells. LE specificity seemed to be expressed on DR but not on DQ molecules since the serum reacted with and could be absorbed by DR+,DQw- cells; it did not react with a DR-,DQw+ mutant cell, but did react with the DR+,DQw+ parental cell. The relationship between LE specificity and MC1 and Te23 specificities remains to be determined.

[Study of a sub-population of T-lymphocytes with the HLA-DR molecule in rheumatoid polyarthritis]. / Etude d'une sous-population T lymphocytaire à molécules HLA-DR dans la polyarthrite rhumatoïde.

The sub-population of T-lymphocytes which express the DR antigen was studied in 48 patients with rheumatoid arthritis and in 42 controls. The sub-population of T-lymphocytes which express DR was found to be larger in the patients with rheumatoid arthritis. This increase is even more marked in cases of rheumatoid arthritis with high erythrocyte sedimentation rates and with a high level of immune complexes.