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Biomedical subjects

B Genetet

Publications and source records attributed to B Genetet.

At least 37 records · Page 2Linked to original sources
PubMed1995

TAP2 gene polymorphism contributes to genetic susceptibility to multiple sclerosis.

MS is an autoimmune demyelinating disease that has been known to be associated with the HLA-DRB1*1501-DQA1*0102-DQB1*0602 haplotype. TAP1 and TAP2, two genes encoded within the MHC class II region between HLA-DP and -DQ loci, display genetic variability and are involved in the transport of antigenic peptides from the cytoplasm to the endoplasmic reticulum. Comparison of 116 MS patients with Caucasoid controls did not reveal any significant correlation between the previously described alleles of the TAP1 and TAP2 genes and MS. We report here an additional TAP2 dimorphism at codon 386, called I and J, corresponding to a silent mutation. An increased frequency of the J variant was observed in the patient population. The J mutation was not found in linkage disequilibrium with the HLA-DRB1*1501 allele and can be considered an additional genetic susceptibility marker of the disease.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1995

[Development of a control method for sample distributors].

The use in routine of distribution automations in Blood Bank laboratories should imply regular testing of the equipment by users. This work was carried out on Microlabs as well as on an IEMS. We used a coloured solution (Alizarine Red in pH9 buffer). We applied the Beer-Lambert law, knowing that any volume modification induces a change in optical density. After testing the linearity of the photometers used in the laboratory, we set up 4 distribution programmes so as to simulate classic distribution of diluted or undiluted serum: 1) for large volume dispensing, 2) for small volume distribution in a small amount of buffer, 3) for distribution of small volumes in large buffer volumes, 4) a specific IEMS programme. Results, expressed as coefficients of variation (CV) for each volume, varied from 9.8 to 0.7% according to the programme. These results show that large volume distribution does not pose any problem (CV < 5%). When dilution occurs, the error observed in serum distribution adds up to that of the thinner, and the smaller the two volumes, the greater is the error. It therefore appears necessary to find the best compromise between serum volume and thinner volume to obtain a CV < 5%. This error may appear negligible in semi-quantitative tests, but it may be important in quantitative tests. Lastly, this programme can easily be adapted in routine to monitor the dispensers efficiently.

Blood Specimen Collection↗
PubMed1995

[Development of a validation method for readers of ELISA microplates].

The results, either positive or negative, of the ELISA tests performed on blood donors to detect viral markers, are always validated by reading microplates on photometers. In Rennes, these microplates can be read on MR 7000 (Dynatech) or IEMS (Labsystems) photometers. This step, the last one in the technical sequence, conditions the results. Regular checking of the photometers therefore may enhance the quality of the testing sequence. To check the quality of the photometers, we use a 96-well microplate, the "Universal Calibration Test Plate" marketed by BIO-TEK Instruments USA. The plate includes 8 blank wells, 6 wells equipped with glass filters (3 stained and 3 unstained). This plate comes with a table showing the optical density in relation to the wavelength. We first verify the repeatability of each photometer, and secondly, verify accuracy. The plate also permits verifying the linearity of each photometer. In view of the results, we were able to state that both photometers were accurate, since the results obtained were compatible with an optical density of +/- 0.024 +/- 1%. Do. The tests are repeatable, with a variation coefficient below 0.5%, linearity was verified (r: 0.999 for both photometers). Reading the plate only takes a few minutes and permits daily control of the photometers. It only requires that the test results be reported on a control card to follow-up possible deviations, and technical maintenance can thus be carried out immediately.

Calibration↗
PubMed1995

[Study of class I HLA polymorphism in the Tunisian population].

The HLA class I gene polymorphism (HLA-A, -B) was investigated in a population of 102 Tunisians. Allele and haplotype frequencies as well as linkage disequilibrium between HLA-A and HLA-B loci were calculated and compared with other populations. The most frequent alleles were A2 (23%), A30 (12.5%), A3 (10.2%), A1 (8.5%), A23 (7.4%) for the HLA-A locus and B21 (14.3%), B44 (11.4%), B35 (9.6%), B5 (8.5%) for the HLA-B locus. The most frequent haplotype was A3 B21 (2.6%) and a positive linkage disequilibrium was found for the following allelic associations: A11 B35 (X2 = 6.8), A28 B35 (X2 = 5.3), and A30 B35 (X2 = 5). In conclusion, a specific distribution of HLA class I components in terms of antigen and haplotype frequencies characterizes the Tunisian population. This specific pattern may reflect the great ethnical diversity of this community. All these informations may be helpful in the future for HLA and disease association studies.

Alleles↗
PubMed1995

Thyroid cell survival in coculture with autologous peripheral or intrathyroidal lymphocytes.

OBJECTIVE: We have studied lymphocyte induced cytotoxicity and the production of interferon gamma (IFN-gamma) and tumour necrosis factor alpha (TNF-alpha) during coculture of thyrocytes and autologous lymphocytes from patients with Graves' disease and from normal subjects. PATIENTS: Thyroid tissues and lymphocytes were obtained from 28 patients with Graves' disease and from 9 control subjects. MEASUREMENTS: Lymphocyte induced cytotoxicity was evaluated on autologous thyrocytes using 5 metabolic tests: the MTT assay, the neutral red uptake, lactate dehydrogenase measurement and glutathione assay. IFN-gamma and TNF-alpha measurements were performed after 1, 5 or 7 days' coculture. RESULTS: The lymphocytes isolated from peripheral blood (PB lymphocytes) altered the morphology and the metabolism of autologous thyrocytes. The intrathyroidal lymphocytes isolated after Dispase digestion were not toxic whereas mechanically isolated lymphocytes exerted a little toxicity. No difference was seen between Graves' disease and normal cells. The supernatants from cocultures had higher IFN-gamma levels than those from lymphocyte cultures. In coculture, PB lymphocytes secreted more IFN-gamma and TNF-alpha than intrathyroidal lymphocytes. The PB lymphocyte induced cytotoxicity was not due to IFN-gamma and TNF-alpha alone. CONCLUSION: Peripheral blood lymphocytes are cytotoxic in vitro to autologous thyrocytes whereas intrathyroidal lymphocytes exert little or no cytotoxicity according to their isolation method. The mechanisms of lymphocyte induced toxicity remain to be explained.

Adult↗
PubMed1995

HLA class II gene polymorphism in Tunisians.

The polymorphism of HLA class II genes (HLA-DRB, DQB, DPB) was investigated in 101 Tunisians using polymerase chain reaction. (PCR) amplification and reverse dot blot (RDB) hybridization. Allele and haplotype frequencies, as well as DRB1-DQB1 linkage disequilibria, were calculated. A total of 26 DRB1 alleles were detected and the most prevalent variant was DRB1*0301 with an allelic frequency at 21.87%. In the DR1 group, DRB1*0102 was most frequent than DRB1*0101. In the DR4 group, DRB1*0403 was the most common allele and was associated with DQB1*0402. Interestingly this DRB1-DQB1 association has not been observed in other populations. With regard to the DR8 group, DRB1*0804 was the unique variant detected, whereas with the DR13 specificity, the most common variant was DRB1*1303 in Algerians also. Although the DQB1 polymorphism analysis showed an allelic distribution very close to that observed in caucasoids, many DRB1-DQB1 associations which have not been reported in studies of other populations, were described. Finally at the DPB1 locus DPB1*1701 and *1301 allele frequencies distinguish clearly this Tunisian sample from a French caucasoïd panel of 83 subjects. In conclusion, a specific distribution of HLA components in terms of gene and haplotype frequencies characterizises this Tunisian population. This specific pattern may reflect the great ethnic diversity of this community. All these informations may be helpful in the future for HLA and disease association studies.

Gene Frequency↗
PubMed1995

Ex vivo studies of polymorphonuclear neutrophils from patients with early-onset periodontitis (III). CR3 and LFA-1 expression by peripheral blood and gingival crevicular polymorphonuclear neutrophils.

In this study, we assessed the LFA-1 (CD18/CD11a) and CR3 (CD18/CD11b) expression on peripheral polymorphonuclear leukocytes (PB-PMN) and crevicular fluid polymorphonuclear leukocytes (CF-PMN), by subjects with a healthy periodontium (n = 7), gingivitis (n = 8), early-onset periodontitis (n = 17) and adult periodontitis (n = 8). Using flow cytometry analysis, the %s of CD18, CD11a and CD11b positive cells and the absolute numbers of fluorescent molecules were determined. No significant difference could be found among the 4 groups, for these 2 kinds of parameters, in PB-PMN or CF-PMN. However, a great difference could be noted between the results obtained from PB-PMN and those obtained from CF-PMN. The %s of positive CF-PMN were significantly lower than those of PB-PMN for the 3 sub-units (p < 0.001). The levels of CD18 and CD11b expressed by CF-PMN were higher than those expressed by PB-PMN and the difference was significant for CD11b (p < 0.001). On the contrary, the level of CD11a expressed on CF-PMN was significantly lower than that expressed by PB-PMN (p < 0.001). Hence, our current results show that early-onset periodontitis PMN can be quite normal and this fact is not surprising insofar as, in our study, these cells were perfectly functional and all the subjects were in good health. We concluded that the analysis of the leukocyte adhesion receptors expression on PB-PMN does not appear useful for helping to establish a differential diagnosis between the different forms of periodontitis.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1995

Relevance of 10 Caucasian HLA haplotypes in searches for unrelated bone marrow donors for 100 patients from a single center.

Unrelated donor searches for 100 Caucasian patients were referred to France Greffe de Moëlle Registry (FGM) from September 1987 (24,600 donors) to December 1993 (71,500 donors, 61% DR typed). After DR typing of HLA-A,B matched donors, unsuccessful searches were extended to other European Registries for 36 patients. Twenty two patients had a donor (FGM: 19, other Registries: 3) selected on: (1) HLA-A,B and DRB,DQB1 split identity; and (2) unidirectional relative response < 5% in MLR performed twice. Estimated probability of finding a compatible donor at 9 months in FGM was 12% (s.e. +/- 4%) and 25% at 2 years (s.e. +/- 6%). This probability was stringently dependent on a phenoidentity to one very common HLA-A,B,DR or B,DR haplotype (25% at 9 months when present, representing 19 of 19 patients with a compatible donor). Without this phenoidentity, the probability was zero per cent (P = 0.0001) in FGM searches and < 4% (n = 1) in extended searches. The MLR test was shown to be insensitive for screening for DPB1 mismatches. Clinical status influenced the probability of finding a compatible donor at one year ranging from 9% +/- 9% for ALL to 23% +/- 8% for CML (NS). Disregarding DPB1 mismatches is the most efficient way of increasing search efficiency.

Bone Marrow Transplantation↗
PubMed1995

[Exploration of the various steps of polymorphonuclear neutrophil function in diabetic patients].

UNLABELLED: Vascular complications in diabetic patients is a complex, probably multifactorial phenomena involving cellular phagocytosis. The aim of this study was to evaluate polymorphonuclear performance in 61 infection free diabetic patients based on tests of the different cell functions: 1) adhesion:adhesion molecule expression CD11a, CD11b, CD11c; adhesion test on nylon fibers. 2) chemotaxis:chemotaxis under aragose to FMLP (bacteria oligopeptide) and complement fractions. 3) Phagocytosis:latex beads. 4) Bacteriocidal power:chemoluminescence photometric oxidative potential before and after stimulation with opsonized zymosan and PMA; reduction of tetrazolium nitroblue. RESULTS were analyzed according to type of diabetes, glucose control, duration of the disease, history of infection and presence of vascular complications. RESULTS: compared with a group of 30 controls, the diabetic patients had a significant impaired polynuclear chemotaxis function (p < 0.001) with both spontaneous adhesion and increased expression of adhesion molecules (CD 11b, CD 11c). The chemoluminescence test was increased at the baseline level due to spontaneous polynuclear adhesion and increased production of free radicals. This response decreased after stimulation compared with controls. The type of diabetes, Hb A1c level and history of infection did not appear to have an effect. Inversely, changes in chemotaxis and chemoluminescence were greater in patients with vascular complications. In summary, all the functions of polynuclear neutrophils tested were altered in diabetic patients and could favor vascular complications and infections episodes.

Adult↗
PubMed1995

Distinct pattern of IL-2 and IFN-gamma gene expression in CD4 and CD8 T cells: cytofluorometric analysis at a single cell level using non-radioactive probes.

IL-2 and IFN-gamma gene expression was analyzed using an original method for in situ hybridization (ISH) with non-isotopic probes and flow cytometric analysis (FC). This method permits rapid detection of mRNA at a single cell level among in vitro activated human peripheral blood mononuclear cells (PBMC) and purified CD4 and CD8 T cell subsets. After stimulation with PMA and ionomycin (PMA+Io), cells were fixed at different times and hybridized with digoxigenin (DIG)-labelled RNA antisense or sense probes specific for IL-2 and IFN-gamma. The level of cytokine gene expression in individual cells was visualized using FITC-conjugated anti-DIG antibodies and the fluorescent signal was analyzed by flow cytometry. Specific hybridization with IL-2 and IFN-gamma antisense probes was detected among activated PBMC within lymphoid cells identified by their light scattering properties. Kinetic analysis of the frequency of mRNA producing cells exhibited a biphasic pattern with an early peak at 6-8 hrs. when percentages of IL-2 and IFN-gamma expressing cells reached 35 +/- 7% and 18 +/- 4%, respectively. Similar data were obtained by enzymatic detection on cell smears using AP-conjugated anti-DIG. Combination of ISH with FC was applied to the comparison of the pattern of cytokine gene expression between CD4 and CD8 T cell subsets isolated by negative selection using immunobeads and magnetic separation. IL-2 was expressed by activated CD8 T cells (25-35%), but CD4 T cells were the major producers of IL-2 as assessed by the high frequency of mRNA expressing cells (60%) and the large amount of mRNA per cell relative to the mean fluorescence intensity. In contrast, IFN-gamma mRNA was preferentially expressed by CD8 T cells (27-37%) and a minority of CD4 T cells (17-23%). Despite quantitative differences, kinetic analysis of IL-2 gene expression in CD4 and CD8 T cells showed similar profiles with an early peak at 6-8 hrs. Upregulation of IL-2 gene expression in CD4 T cells by CD28 co-stimulation increases the amount of IL-2 mRNA per cell as visualized by mean fluorescence intensity. In addition the effect of CD28 co-stimulation on IL-2 mRNA stabilisation was demonstrated by the maintenance of a high frequency of IL-2 expressing CD4 T cells and an elevated level of mRNA per cell for prolonged period after PMA+Io stimulation. By contrast CD28 co-stimulation had no obvious effect on IFN-gamma expression.

CD28 Antigens↗
PubMed1994

Ex vivo studies of polymorphonuclear neutrophils from patients with early-onset-periodontitis (II). Chemiluminescence response analysis.

Previous studies have indicated that oxygen-free radicals may cause damage to the periodontal tissues. This study compared the luminol-dependent chemiluminescence response (after stimulation with either opsonized zymosan or phorbol myristate acetate (PMA) of peripheral blood polymorphonuclear leukocytes (PMN) isolated from human subjects with a healthy periodontium (n = 7), gingivitis (n = 8), adult periodontitis (n = 8), or early-onset periodontitis (n = 17). These results were also compared with those obtained in a larger reference group which consists of 50 subjects without infection or inflammation, selected on the basis of laboratory investigations. An enhanced response was defined as being 2 standard deviations above the reference group mean; a reduced response was defined as being 2 standard deviations below this mean. Although PMN from patients with either gingivitis or periodontitis were often functionally activated (when compared to the PMN from the reference group), no significant differences could be found between the 4 groups, with regard to the chemiluminescence response means obtained in a basal state or after stimulation.

Adult↗
PubMed1993

Evidence for a polymorphism of HLA-G gene.

HLA-G gene polymorphism was analyzed by RFLP using seven restriction enzymes and an HLA-G locus-specific probe. Hybridization of 55 DNAs digested with three enzymes (Taq I, Pst I, and Bgl II) revealed two polymorphic bands in each case. RFLP patterns obtained with Taq I and Pst I corresponded to the same allelic polymorphism and differed from the Bgl II polymorphism. Combining both polymorphisms enabled determination of four alleles. Allelic frequencies were calculated: 40% of the subjects tested had allele 1, 36% had allele 2, 22% had allele 3, and 2% had allele 4. Analyzing the complete HLA class I phenotype revealed strong linkage disequilibrium with the HLA-A locus. The polymorphism described is located in the 3' flanking region of the gene. Moreover, extended HLA-A haplotypes were constructed by combining the HLA-G polymorphism with other class-I-sequence polymorphisms.

Base Sequence↗
PubMed1993

HLA-DR and -DQ genotyping by PCR-SSO in Shanghai Chinese.

The polymorphism of HLA class II (HLA-DRB1, -DQA1 and -DQB1) were investigated in 89 Shanghai Han Chinese using polymerase-chain reaction amplification and oligonucleotide (PCR-SSO) typing. Of the 43 DRB1 alleles tested, 24 were observed. DRB1*0901 (16.85%), *0803 (9.55%), *1202 (9.55%) and *1501 (12.92%) were most frequent and account for the high frequencies of DR9, DR8, DR5 and DR2 specificities in this population. The alleles DRB1*0101, *0102 and *1001 had very low frequencies. Among the 8 DQA1 and 13 DQB1 alleles tested, only DQA1*0401 and DQB1*0402 were absent. DQA1*0301, and DQB1*0301 and *0303 were among the most common alleles of the two loci respectively. Unusual DRB1-DQA1-DQB1 haplotypes were observed and a DRB1 "new" allele was suggested.

Alleles↗
PubMed1992

Interferon production in severe hemophiliacs with and without HIV antibodies.

The interferon (IFN) system, both serum IFN levels and the in vitro IFN production, was investigated in 38 clinically asymptomatic multitransfused hemophiliacs, half positive and half negative for HIV antibodies. In most patients, no circulating IFN was detected; similar levels of IFN-alpha were obtained after peripheral blood mononuclear cell (PBMC) stimulation with Sendai virus both in hemophiliacs and controls, while production of IFN-gamma following stimulation with phytohemagglutin (PHA) was diminished in a large number of patients irrespective of their HIV serology. These data indicate that the deficiency in IFN-gamma generation is not only related to HIV contamination but may be a direct consequence of the chronic antigenic stimulation through Factor VIII concentrates.

HIV Seropositivity↗
PubMed1992

Is there a mechanical factor of haemolysis in patients with positive IgG-type direct antiglobulin test?

In autoimmune haemolytic anaemia, the presence of antibodies on the erythrocyte membrane results in haemolysis through an immune process, but does it not alter the rheological properties of red blood cells (RBC), thus adding a mechanical factor to haemolysis? This study was designed to examine the rheological properties of erythrocytes sensitized with IgG-type antibodies. The study involved 20 patients with anaemia and positive direct antiglobulin test, including 12 with straightforward haemolysis, 10 samples sensitized in vitro, and 20 controls. The following haemorheological parameters were studied: erythrocyte filtration, blood and plasma viscosities, titration of adenosine triphosphate (ATP) and 2-3-DPG, erythrocyte morphology under scanning electron microscopy. The results showed increased erythrocyte rigidity (P less than 0.025) as well as higher blood viscosity compared to controls with similar haematocrit values, and unaltered ATP and 2-3-DPG (consistently with scanning electron microscope observations). These haemorheological disorders were more noticeable in patients with clear-cut haemolysis, and there was a correlation between the increase in erythrocyte rigidity indices and the haemolytic parameters, especially haptoglobin (P less than 0.001). The in vitro study confirmed the results obtained ex vivo. To conclude, the mechanical properties of antibody-coated erythrocytes are impaired, which may promote the immunological mechanism favouring haemolysis in the spleen.

Aged↗
PubMed1992

No major monoclonal lymphocyte population in the thyroid of patients with Graves' disease: study of gene rearrangement by restriction fragment length polymorphism and polymerase chain reaction.

The present study aimed at determining the mono-, oligo-, or polyclonal nature of intrathyroid lymphocytes at the DNA level in patients with Graves' disease. Two techniques were used to seek monoclonal rearrangement in DNA derived from intrathyroidal lymphocytes obtained from six patients. The first was restriction fragment length polymorphism using two specific probes from the B-chain of T-cell receptor and the other from the heavy chain immunoglobulin gene; the second was polymerase chain reaction using a couple of specific primers from the variable and joining regions of heavy chain immunoglobulins. The results for the patients with Graves' disease were compared with those obtained for circulating T-and B-lymphocytes, granulocytes (negative controls), and T- and B-leukemic cells (positive controls). The results with restriction fragment length polymorphism favored a polyclonal origin for the lymphocytes in all cases, since no rearrangement was visualized. The results with polymerase chain reaction were analogous, and the technique was 10 times more sensitive in the detection of rearrangement.

Adult↗