An HLA-A*2607 variant (A*260702) independently identified in two unrelated French families.

Abstract In this report, we describe the identification of a new variant of the A*2607 allele, named A*260702. This variant was detected independently in two unrelated potential bone marrow donors living in two different regions of France. Molecular typing and mono-allelic sequencing revealed that this allele differs from the original A*2607 allele by a single-nucleotide substitution in codon 56 (GGG --> GGT). This nucleotide change does not affect the amino acid sequence, both triplets coding for a glycine.

Xenogeneic ossicular implants: an experimental study of heterotopic, demineralized, lyophilized, porcine implants in the guinea-pig.

This study was done to compare the outcome of porcine ossicular implants in the middle ear and the subcutaneous dorsal region of the guinea-pig to those of allo-implants implanted in parallel in the dorsal region. The implants were heteropic, xenogeneic, demineralized (HCl), lyophilized and sterilized. The evaluation was histological (light microscopy and scanning electron microscopy) and immunological (immunofluorescence staining). Fifty-four guinea-pigs were implanted in the middle ear and 14 of them were also implanted subcutaneously in the dorsal region with xeno-implants and allo-implants. The middle ear implants were found to be constantly reossified and coated with normal mucosa with only a minimal immune reaction. In contrast, the dorsal xeno-implants were found to be the target of mononucleic infiltration, fibrous encapsulation and an influx of immunoglobulins resulting in segregation. The corresponding allo-implants were found to be partially reoccupied and reossified. These findings highlight the value of HCl demineralization in the induction of non-species-specific Bone Morphogenetic Protein and the failure of attempts at immuno-despecification. It appears that the fate of the implant depends less on its antigenic load than on the site of implantation. In this regard the middle ear is apparently very advantageous. The very good short-term tolerance and recovery observed in the middle ear xeno-implant suggest that these implants offer sufficiently good results to warrant clinical testing.

Factors influencing haemopoietic recovery following chemotherapy-mobilised autologous peripheral blood progenitor cell transplantation for haematological malignancies: a retrospective analysis of a 10-year single institution experience.

We retrospectively analysed the factors that influenced rate of haemopoietic recovery (HR) in 243 patients after transplantation with chemotherapy-mobilised autologous peripheral blood progenitor cells (PBPC). Approximately half the patients also received haemopoietic growth factors (HGF) for mobilisation. Conditioning for transplantation was with either chemotherapy alone or chemotherapy plus total body irradiation (TBI). Median time to recovery of granulocytes > or = 0.5 x 10(9)/l was 13 days (range 7-93 days) and of platelets > or = 50 x 10(9)/l 14 days (7-440). Speed of HR was greater, both for neutrophils and platelets for patients who received more rather than less CFU-GM than our median value of 18.9 x 10(4)/kg (P < 0.0001 in both instances) and more rather than less CD34-positive cells than our median value of 8.8 x 10(6)/kg (P < 0.0001 and P < 0.0005, respectively). For granulocyte recovery, in the multivariate analysis the dose of infused CFU-GM (P = 0.05) and the use of HGF for both mobilisation and post-transplantation (P < 0.0014) were significant positive factors. For platelet recovery in the multivariate analysis the dose of infused CFU-GM (P < 0.0016) was a positive factor. The use of busulphan and of TBI were significant adverse factors for rate of platelet recovery (P = 0.005 and 0.0004, respectively). When compared with non-HGF-mobilised PBPC, HGF-mobilised PBPC reduced the number of days of hospitalisation (28 vs 24, P = 0.0001) and of treatment with intravenous antibiotics (15 vs 11, P = 0.0004). These findings emphasise the importance of cell dose in accelerating haemopoietic recovery after autologous blood stem cell transplantation.

[Decalcified, lyophilized, sterile heterotopic porcine ossicular xenografts. Experimental evaluation in the guinea pig]. / Les xéno-implants ossiculaires hétérotopiques porcins décalcifiés, lyophilisés et stérilisés. Evaluation expérimentale chez le cobaye.

Using the guinea pig middle ear model, we assessed decalcified, lyophylized, sterile heterotopic porcine ossicular xeno-implants based on a histology (optic and electron scan microscope) and immunologic (immunofluorscence) methods. Implants were placed in the middle ear and others in the dorsal subcutaneous area. Allo-implants were compared as controls. Implants were placed in the middle ear in 54 animals and skin implants in 14. Under the influence of BMP, the implant ossified in all cases in the middle ear. Intense immune recruitment was not observed. Inversely, there was a mononuclear infiltration reaction to the skin implants with formation of a fibrous capsule, immunoglobulin and complement influx and consequently sequestration. The allo-implants were partially reossified. These findings confirm the value of decalcification with hydrochloric acid for BMP induction, independent of species and the failure of attempted immune despecification. Implant outcome is not dependent on its antigen load, which is high compared with its weight, but on the site of implantation. The middle ear appears to be a privileged site of implantation.

Immunosuppressive activity of sera of pregnant women on cytotoxic T-lymphocyte-mediated cytolysis. I. Characterization of the active fraction.

Sera of healthy pregnant women have a nonspecific inhibitory activity on the specific cytolytic activity of activated cytotoxic T lymphocytes in vitro. The serum fraction of pregnant females which is responsible for this activity has been characterized. The equivalent fraction derived from sera of healthy men has a similar but reduced activity compared with the female fraction. This suppressive factor seems to be activated during pregnancy. Moreover, this study sheds some light on the reasons behind the attenuation of the maternal immune system during pregnancy.

Measurement of residual leukocytes in platelet concentrate units.

Leukocyte contamination of various blood products is implicated in febrile transfusion reactions, in alloimmunization phenomena and immunosuppression, and in the transmission of viral infections. The biggest difficulty in interpreting published reports lies in the methods used to quantify residual leukocytes. As automated cell counters are not precise and sensitive enough to detect low cell counts, we compared two techniques: i) the classical method using the Nageotte haemocytometer; and ii) fluorescent staining with propidium iodide (PI). We performed these two techniques to evaluate the residual white cell count in 41 platelet units collected by cytapheresis. The difference in results obtained with the two techniques was borderline significant (P = 0.05). To evaluate the accuracy of both techniques, we added a known number of lymphocytes to platelet concentrates that had been filtered three times. There was a good correlation between the values obtained using the two techniques (r > 0.99). However, though the difference between the known value and PI results was not significant, the difference for Nageotte cell counter results was significant (P < 0.0001). From a practical point of view, the PI fluorescent technique is simple, rapid, and easy to carry out. Our results demonstrate the reliability of the PI technique for the evaluation of residual leukocytes.

[Technique for automating HLA-DR typing]. / Essai technique d'automatisation des groupages HLA DR.

Two types of anti-class II coated magnetic beads were compared for the use in isolation of B lymphocytes from whole blood. The HLA DR typing was performed in parallel after separation of the cells by the rosette technique. The depletion was monitored by Flow cytometry using CD2 and CD20 antibodies. Typing was performed using classic lymphocytotoxicity test of Propidium-Iodide-Carboxyfluorescein diacetate fluorescence. The results were scored with an ordinary microscope or by photometric evaluation. These techniques lead to simplification in cell isolation from small amounts of blood with an important gain of time and a less subjective interpretation of DR phenotyping was until now long and difficult.

The influence of cryopreservation on activity and surface markers of lymphokine-activated killer cells.

Ficoll-separated and monocyte-depleted mononuclear cells isolated from normal leukapheresis products were cryopreserved. These cells were incubated with or without 1,000 U/ml of recombinant interleukin-2 (rIL-2) for 4 days, and their lymphokine-activated killer (LAK) and natural killer (NK) activities were measured. IL-2 activation induced a significant increase in the expression of the CD25 antigen. There was no change in CD2, CD3, CD4, CD8, CD16, CD56 and CD57 cell marker expression. Cryopreservation did not induce any change in the membrane antigen expression and in the lymphocyte subsets. The NK activity was well preserved and the decrease of LAK activity of IL-2-activated cells after cryopreservation was not significant. In contrast, cells activated before cryopreservation had a significantly lower cytotoxic activity and the number of cells expressing the IL-2 receptor was also significantly reduced. However, the decrease of CD56 expression was not significant. CD25 expression seemed to be proportional to the LAK activity of the cells. This study demonstrated that cryopreserved lymphocytes, after 4 days of culture with rIL-2, could be as active and could express the CD25 and CD56 cell surface markers in the same manner as fresh LAK cells.

Lymphocyte immunization generates immunosuppressive factors in women with recurrent abortions.

Women with recurrent spontaneous abortions of unknown etiology were treated with injection of their partner's lymphocytes. The effect of lymphocyte immunization is evaluated by various immunological tests. The results demonstrate the appearance of immunosuppressive factors, especially P4 protein, present in normal pregnancy, which inhibits cellular cytotoxicity.

[Identification of HLA B27 antigen by flow cytometry]. / Identification de l'antigène HLA B27 par cytométrie de flux.

Cytofluorograph analysis after immunofluorescence with monoclonal antibodies can be useful for various antigens identification on the surface of cell membrane. This technic is described for antigen HLA B27 identification. This HLA antigen is known indeed to be linked to rhumatismal diseases. The fluorescence histograms obtained, allow to differentiate easily subjects which have HLA B27 phenotype and subjects which are HLA cross linked.

An inhibitor of cellular cytotoxicity is present in multiple sclerosis.

A blocking factor is present in the serum of patients with multiple sclerosis (MS). It is able to inhibit a cellular cytotoxicity reaction in vitro. This substance has characteristics similar to those of the 'P4' protein described in pregnant women and patients bearing tumors. Its possible contribution to MS pathogenesis is discussed.

Lupus erythematosus panniculitis with partial genetic deficiency of C2 and C4 in a child.

A 7-year-old girl developed recurrent febrile nodules and subcutaneous plaques on her arm, shoulder, and face. Primary acquired toxoplasmosis was diagnosed at onset, because of associated lymphadenopathy and detection of specific IgM. Skin lesions were responsive to therapy with systemic steroids but progressed to atrophy and poikiloderma within a few months. Subsequently, chloroquine therapy has been instituted and no relapse has occurred. Histopathologic findings and direct immunofluorescence studies were diagnostic of lupus erythematosus panniculitis. Electron microscopy showed lamellar inclusions within mast cells. Results of laboratory investigations disclosed leukopenia and hypocomplementemia with low C2 serum levels. A family study of the complement system, including C4 and Bf phenotyping and HLA A, B, and DR haplotyping, revealed the carriage of both C2 and C4A null alleles in the propositus. This observation suggests an additional link between lupus erythematosus panniculitis and the remainder of the lupus erythematosus clinical spectrum.

Characterization of P4, a natural inhibitor of cellular cytotoxicity reaction in pregnant women.

A factor present in the serum of pregnant women is able to inhibit nonspecifically a cellular cytotoxicity reaction. This glycoprotein called P4, is an alpha-globulin whose molecular weight is 200,000 daltons bound to albumin. Its isoelectric point is 4.1. By its various characteristics, P4 seems different from other proteins so far described.

Influence of HLA-A, B, and DR matching on the outcome of kidney transplant survival in preimmunized patients.

The outcome of 893 prospectively typed (HLA-A, B, and DR) and matched cadaveric kidney transplants--all first grafts, with patients being transfused before transplantation--was studied using actuarial survival methods. The effect of HLA-A, B and DR matching was only found to be significantly beneficial to graft survival in the group of 289 presensitized recipients: 70% and 43% graft survival at two years in the case of best-matched (4-6 HLA-A, B, and DR) identities versus mismatched (0 and 1 HLA-A, B, and DR) identities, respectively (P = 0.05). Although a cumulative effect of matching for antigens belonging to the 3 HLA-A, B, and DR series was observed among the group of preimmunized recipients, a trend arose in favor of the prominent role of the HLA-B alleles. No significant difference related to HLA matching was observed in the group of nonsensitized recipients. These results confirm previous observations and support efforts to give priority for matched kidneys to preimmunized patients.

Immunological analysis of women with repeated spontaneous abortions.

The study deals with 9 women with recurrent abortions who do not possess the blocking factor which is present during normal pregnancy and which inhibits, nonspecifically, a cellular cytotoxicity reaction in vitro. Electrophoresis of the women's serum revealed an abnormal glycoprotein, migrating in the alpha-globulin zone, which gives a precipitate with an anti-gamma-globulin serum after mercaptoethanol treatment. These last two findings may indicate a possible abortion.

Evidence of immunosuppressor factor in the serum of women taking oral contraceptives.

The serum of women taking oral contraceptives is able to inhibit an in vitro cytotoxicity reaction at the cell level. This effect is nonspecific. It seems to be due to the presence of an inhibitor of the lymphocyte activity and acting at the membrane level. This factor looks like that described in expectant mothers except that it exhibits a lower activity. This difference makes it possible to consider that its emergence depends on a specific hormonal impregnation.

Identification of a factor blocking a cellular cytotoxicity reaction in pregnant serum.

A blocking factor present in the serum of pregnant women is able to inhibit non-specifically a cellular cytotoxicity reaction. It is a glycoprotein, migrating in the alpha-globulin region with a molecular weight of about 210,000 daltons. Its absence from the serum of women with recurrent abortions could predict the outcome of pregnancy.