[Gelatinous transformation of the bone marrow: A retrospective monocentric case series of 12 patients]. / Transformation gélatineuse de la moelle osseuse : une série rétrospective monocentrique de 12 cas.

PURPOSE: Gelatinous bone marrow transformation is a rare complication of unknown pathogenesis related to several underlying diseases. It is described as a focal loss of hematopoietic cells and a deposition of eosinophilic gelatinous substance rich in hyaluronic acid. We report a retrospective monocentric study followed by a literature review. METHODS: We have identified 12 patients with gelatinous transformation from 1999 to 2014. Clinical, biological and cytological data were collected. RESULTS: A bone marrow aspirate was performed for mild to severe pancytopenia (50%), less frequently for monocytopenia. Mean haemoglobin was 10.4g/dL, platelets 126.9G/L and leucocytes 3.82G/L. Gelatinous transformation was found in all patients. We have identified three hematologic malignancies, three cancers, three severe infections, two anorexia nervosa and one kidney injury. Ten patients had undernutrition, 9 out of the 12 patients died within a year due to advanced underlying disease and comorbidities. CONCLUSION: Gelatinous bone marrow transformation is found in various situations of severe undernutrition including anorexia nervosa, advanced cancers or severe sepsis, especially in HIV infection. The deposition of hyaluronic acid probably reduces haematopoiesis. Due to a complex inflammatory process, it alters the hematopoietic microenvironment and the bone marrow stroma. Severe undernutrition and other mechanisms are reviewed in this study. Gelatinous degeneration is still a rare disorder, indicative of an advanced underlying disease; its recognition could help guide investigations.

[Increased haemoglobin A2 levels in pseudoxanthoma elasticum]. / Augmentation de l'hémoglobine A2 au cours du pseudoxanthome élastique.

BACKGROUND: Pseudoxanthoma elasticum (PXE) is normally associated with mutations in the ABCC6 gene. A PXE phenotype without mutations in ABCC6 has been described in Greek and Italian patients presenting with beta thalassemia. We attempted to determine the incidence of beta thalassemia in a cohort of French patients with PXE. PATIENTS AND METHODS: Fifty patients with PXE were included in the study. Laboratory examinations comprised hemoglobin electrophoresis, ABCC6 gene study and in some studies: mutation analysis, beta-globin gene. RESULTS: No cases of beta thalassemia were diagnosed in this cohort of French patients with PXE. However, 20% of the latter exhibited a significant but isolated (i.e. without microcytic anemia) increase of hemoglobin A2 (HbA2). Statistical comparisons showed no difference in terms of geographical origin or severity of PXE between patients with high levels of HbA2 and those with normal levels of HbA2 other than the extent of cutaneous involvement. Study of the beta-globin gene displayed mutations only in the two patients with the highest recorded levels of HbA2. ABCC6 + beta-globin digenism was ruled out of the pathogenesis of PXE. DISCUSSION: The PXE phenotype seen in some patients with beta thalassemia appears to be associated with epigenetic modification of ABCC6 transcription and depends specifically on the beta globin locus. Isolated increase in HbA2 is probably a laboratory marker for PXE. Here again, a functional epigenetic reaction between ABCC6 and the beta-globin locus was suspected. However, these reciprocal interactions are clearly unequal since the change in ABCC6 transcription occurring during the course of beta thalassaemia is responsible for a PXE phenotype while increased HbA2 during the course of PXE has no clinical consequences.

[Vascular purpura in a patient with severe sarcoptic acariasis]. / Purpura vasculaire au cours d'une gale sévère.

INTRODUCTION: The common cutaneous manifestations of human sarcoptic acariasis are erythematous and pruriginous papules. Vascular purpura in patients with scabies has rarely been reported. We report a case of vascular purpura in the presence of a transient elevation of anticardiolipin antibodies. CASE REPORT: A 79 year-old woman was treated with prednisone and chlorambucil for a lymphoma. She was referred to our institution for diffuse sarcoptic acariasis. Clinical examination showed a petechial purpura on her legs. Histology of two biopsies of purpuric papules showed non inflammatory thrombosis of the upper dermis vessels. Investigations revealed a transient elevation of anticardiolipin antibodies, moderate hyperhomocysteinaemia and blood eosinophilia. Treatment consisted in ivermectin and repeated applications of benzyl benzoate lotion. The purpura healed within one month, without specific treatment. DISCUSSION: Histological findings of sarcoptic acariasis with vascular purpura correspond to leukocytoclasic vasculitis. Focal glomerulonephritis has been reported. In our case, histopathology showed a non inflammatory thrombosis of the dermal vessels. We hypothesize that the anticardiolipin antibodies, hyperhomocysteinaemia and blood eosinophilia may have been the cause of vascular thrombosis.

Primitive cytokines and cytokine receptors in invertebrates: the sea star Asterias rubens as a model of study.

It has been previously demonstrated that the sea star axial organ is a primitive immune organ. Phagocytic, lymphoid-like cells have been characterized with properties similar to those of vertebrates. There is also evidence for an invertebrate cytokine network because IL-1 and TNF-like activities are clearly demonstrable. In addition, the authors have previously described preliminary evidence for IL-2-like activity in the sea star. In the present report, the authors obtained evidence for the presence of IL-1- and IL-2-like molecules on axial organ cells. More interestingly, the results suggested that sea star cells express structures similar to human receptors for IL-1, IL-2, IL-6 and IFN-gamma.

Expression of V beta gene segments by Sezary cells.

The T-cell receptor V beta repertoire expressed by Sezary cells was determined in a series of 16 patients whose samples have been shown to contain a majority of tumor cells. By using anti-V beta monoclonal antibodies, polymerase chain reaction analysis of expressed V beta, and, in selected cases, nucleotide sequencing, we have shown that the expressed V beta segments belong to five V beta families (V beta 5, V beta 6, V beta 8, V beta 13, and V beta 18), which contain a large fraction of the T-cell receptor V beta repertoire and do not share significant similarities in complementary determining region 4. V beta segments from these five families were also found to be strongly expressed by CD4 + CD7- peripheral blood cells obtained by fluorescence-activated cell sorting from two healthy donors. The diversity of the V beta repertoire expressed by Sezary cells appears to be similar to that expressed by circulating non-neoplastic T cells. These data do not support the hypothesis that a common superantigen is involved in the initiation of this form of cutaneous T-cell lymphoma.

A Th0/Th2-like function of CD4+CD7- T helper cells from normal donors and HIV-infected patients.

We previously reported the expansion during HIV infection of a subset of CD4+ T cells characterized by a lack of CD7 cell surface expression. This CD4+CD7- subset showed in normal donors a lower cell proliferation than CD4+CD7+ autologous cells after CD3 triggering or CD28 costimulation. Our aim was to further characterize the Th function of this CD4+CD7- T cell subset, both in normal donors and in HIV-infected patients. Their CD4+CD7- cell proliferation and cytokine secretion were analyzed after cell-sorting and co-stimulation of the CD3 and CD28 pathways. In normal donors, the IL-2 produced by CD4+CD7- cells in response to a CD3 plus CD28 costimulus represented 50 +/- 28% of the autologous CD4+CD7+ cell IL-2 secretion. In addition, the CD4+CD7+ T cells produced higher amounts of IL-4 and IL-10 than the CD4+CD7+ cells with mean CD4+CD7-: CD4+CD7+ ratios of 5.4 +/- 4.5 and 26 +/- 25, for IL-4 and IL-10, respectively, whereas the IFN-gamma production was similar in both subsets. After a triggering of the CD3 complex alone, significant amounts of IL-2 were detectable in CD4+CD7+ cell supernatants only; conversely, IL-4 and IL-10 could be detected only in the culture medium from CD4+CD7- T cells. This profile of cytokine production was maintained both over time and at the clonal cell level in the CD4+CD7- T cell subset. In HIV-infected patients, the CD4+CD7- T cell expansion observed in relationship with disease progression was associated to an in vivo activation of the CD4+CD7- cells, as assessed by cell surface expression of the HLA-DR, CD25, CD71, and CD57 markers. The CD4+CD7- T cells from HIV-seropositive patients showed the same imbalance of cell proliferation and IL-2, IL-4, and IL-10 production as observed in normal donors despite low levels of proliferation and cytokine production. Together, our data indicate that the lack of CD7 expression defines a CD4+ Th cell subset with a Th0/Th2-like profile of cytokine secretion in normal individuals. The CD4+CD7- subset is expanded during HIV infection and characterized by the same although impaired profile of cytokine production. These CD4+CD7- T cells might play a role in the Th cell dysfunctions observed in HIV infection.

T lymphocyte subsets in primary antiphospholipid syndrome.

OBJECTIVE: To study T, natural killer (NK) and B blood cell subsets in primary antiphospholipid syndrome (APS). METHODS: We studied 10 patients with primary APS and 12 healthy subjects. Blood lymphocytes counts, proportions of B cells (CD19+), total CD5 and CD5 B cells, NK cells (CD16+CD56+), T cells (CD3+), CD4+ helper (naive CD4+CD45RA+, memory CD4+CD45RO+ and CD4+CD29+, activated CD4+CD25+), CD8 (immunoregulatory CD8+CD57+, activated CD8+HLA-DR+) T cell subsets were measured by flow cytometry analysis. RESULTS: In the primary APS group, we observed a lower total lymphocyte count (p = 0.009), an expansion of naive CD4 cells (p = 0.025), a lower proportion of memory CD4 cells (p = 0.04) and an increased ratio of naive/memory CD4 cells (p = 0.015). CONCLUSION: Blood T cells phenotypes in primary APS differ markedly from healthy controls, indicating that immunologic abnormalities in primary APS might extend beyond autoantibody production.

Identification of T-like and B-like lymphocyte subsets in sea star Asterias rubens by monoclonal antibodies to human leukocytes.

The axial organ of sea star Asterias rubens is a primitive immune organ. The total cell population was fractionated into two populations: adherent (B-like) and non-adherent cells (T-like) to nylon wool. These two cell subsets were previously defined as functionally acting as mammals T and B cells. In the present report, we pointed out that these T and B-like cells can be identified using mouse to human monoclonal antibodies. Reproducible results were obtained with anti-CD7 monoclonal antibody which detects the T-like cell subset and with anti-CD14 antibody that characterizes the B-like population.

Defective calcium response in B-chronic lymphocytic leukemia cells. Alteration of early protein tyrosine phosphorylation and of the mechanism responsible for cell calcium influx.

To study defective signal transduction via the Ag receptor of B-chronic lymphocytic leukemia cells (B-CLL), we examine in this report the Ca2+ response triggered by anti-mu antibody in 23 patients previously classified in three phenotypic groups. Altered Ca2+ changes are essentially found in CLL group II whose leukemic cells are characterized by a marked expression of the CD11b Ag. B-CLL cells from patients with a defective Ca2+ response present an altered pattern of protein tyrosine phosphorylation after anti-mu stimulation in comparison with normal human B cells and B-CLL cells from patients having a normal Ca2+ response. Most of the proteins usually tyrosine phosphorylated after the triggering of cell-surface IgM are concerned. This includes the gamma 1 isoform of phospholipase C, although the protein is normally present in B-CLL cells. These findings suggest that the interruption of the phosphoinositide pathway in B-CLL cells is very proximal, at the level in the signaling cascade between activated surface Ig receptors and protein tyrosine kinases. Simultaneously, we demonstrate that some low responding patients exhibit a decreased Ca2+ response to thapsigargin, an agent known to release intracellular Ca2+ without inositol 1,4,5-trisphosphate production. This suggests that an altered functioning of the mechanism leading to the cell Ca2+ influx in B cells can be also involved in the decreased Ca2+ response observed in B-CLL cells.

[Amplification of a circulating CD4+CD7- T lymphocytes subset in kidney transplantation]. / Amplification d'une sous-population lymphocytaire T circulante CD4+CD7- en transplantation rénale.

A minor subset of CD4+ T-cells (10 +/- 4 percent) that does not express the CD7 antigen is detectable in normal individuals. These CD4+CD7- T-cells have lower proliferative capacities than autologous CD4+CD7- T-cells. We observed an expansion of CD4+CD7- peripheral T-cells in 45 kidney transplant recipients. The CD4+CD7- TL expansion (23 +/- 20 percent was observed in both short term (< 6 months, n = 20) and long term transplanted patients (> 3 years, n = 25). These results significantly differed from those observed in patients on dialysis (10 +/- 4 percent, P < 0.002). The percentages of CD4+CD7- TL in kidney recipients are significantly correlated with the percentages of CD4+CD57+ and CD8+CD57+ T-cells subsets. A three-color immunofluorescence analysis indicated that 75 percent of the CD4+CD7- TL express the CD57 molecule contrasting with those in normal individuals (0.3 percent). Purified CD4+CD7- T-cells from kidney recipients have lower proliferative responses after PHA, CD2 and CD3 stimulations than autologous CD4+CD7+ TL. The proliferative response is partially restored by the addition of anti-CD28 moAb or IL2. We have highlighted, in kidney recipients with good graft function, an expansion of CD4+CD7- T lymphocytes which could be associated with CD4+ T-cell anergy and tolerance.

CD4+CD7-CD57+ T cells: a new T-lymphocyte subset expanded during human immunodeficiency virus infection.

CD7 and CD57 are two cell surface molecules related to the differentiation or functional stages of CD4+ T cells. The CD4+CD7- T cells represent a minor subset of CD4+ cells in normal individuals and are considered to contain the normal counterpart of Sézary T cells; the CD4+CD57+ peripheral blood lymphocytes (PBL) are detectable in long-term renal allograft recipients. We compared the cell surface expression of these CD7 and CD57 markers on CD4+ T lymphocytes in peripheral blood and lymphoid organs from normal individuals and human immunodeficiency virus (HIV)-infected patients. Our results indicate that CD4+CD7- T cells in normal PBL do not express CD57 and were poorly responsive to anti-CD3 monoclonal antibody (MoAb), the activation being restored by addition of anti-CD28 MoAb. This CD4+CD7- cell subset is increased in peripheral blood during HIV infection, and its progressive expansion mirrors both the absolute and relative decrease of CD4+ T cells. The lack of CD7 expression is correlated with CD57 acquisition on CD4+ T cells because CD4+CD7-CD57+ cells represent a major component of the CD4+CD7- subset in HIV-infected patients. Our results suggest that the presence and the expansion of CD4+CD7-CD57+ T lymphocytes, which do not behave as previously defined helper subsets, may participate to the immune dysfunction observed during HIV infection.

Characterization of normal human bone marrow by flow cytometry.

Fifteen bone marrow samples were obtained from consenting donors for allogenic bone marrow transplantation and were studied by flow cytometry. The simultaneous analysis, cell by cell, of size (forward angle light scattering), structure (right angle light scattering) and surface antigenic expression with 20 monoclonal antibodies allowed selection of ten reproducible cell subsets which were sorted and studied by microscopic examination in order to confirm their distinct characteristics. This very accurate analysis of a bone marrow cytogram should find applications in view of a better characterization of normal and pathological human bone marrow.

Proposals for a Phenotypic Classification of B-Chronic Lymphocytic Leukemia, Relationship with Prognostic Factors.

Immunophenotypic analysis was performed in 53 cases of B chronic lymphocytic leukemia using a large panel of monoclonal antibodies recognizing B, T, activation and myeloid antigens. Our results showed four patterns of reactivity: (a) several molecules were constantly expressed: CD19, CD20, CD24, CD37, HLA-DR, mu heavy chain, CD5, CD23, B5, CD32; (b) one antigen, CD11b, was found in 50 to 80% of the cases; (c) some markers were detected in less than 50% of the cases: CD25, CD38, CD71, CD11a, c, CD14b-c; (d) CD2 and CD16 were never detected. From these results, a phenotypic classification in three groups has been proposed and these groups were correlated with the progression of the disease, mainly with the lymphocyte doubling time of less than one year. We hypothesized that the leukemia cells could be at various stages of differentiation and/or activation according to their expression of activation and myeloid markers.

[Description and performance of a flow cytometry apparatus: the EPICS]. / Description et performances d'un appareil de cytométrie de flux: l'EPICS.

This is a report on our experience with the EPICS C (Coultronics) cytometric flux apparatus, a screening cell analyzer, employing a laser ray (2 or 5 watts); we obtained good results to analyze immunologically-tagged mononuclear blood cells with or without prior separation: for rhythm, repeatability, and contamination. The EPICS C machine proved to be effective to study the cell cycle using lymphoblastic cells, epithelial cells and cells from a breast cancer. Several screening trials were carried out with fluorescent ball bearings of various sizes; the quality of screening (purity and yield) appear optimal at a speed ranging from 500 to 1,000 bearings per second, using three parameters: the logarithm of green fluorescence, the integral function of green fluorescence, the diffraction of light to small angles. Thus, if results obtained for this analysis are entirely satisfactory, the screening function remains limited because it is slow.