Biomedical subjects
Publications and source records attributed to A Májský.
The author describes a peculiar case of a paternity suite where the accused died and his parents were also dead. Possible paternity was assessed by examination of HLA antigen of his ex-wife and daughter from his former marriage and in his sisters.
The author described an uncommon case when neither of two men (former husband and another man) who were considered possible fathers of the child, could be ruled out as potential fathers based on examination of signs of 19 erythrocyte, serum and enzyme systems and 59 HLA-A, B, C antigens and their subtypes. The eliminating capacity of the examination was 99.86%; paternity was, based on probability calculation, practically proved (p = 99.9925% and 99.90%). According to statements of parties in the dispute the other man is the child's father. The author discusses perspectives of DNA examinations.
The authors describe another subtype of antigen HLA-B27, described as HLA-B27P3. It was assessed, using 5 anti-HLA-B27 sera. It is identical with the weak antigen HLA-B27(HLA-B27w) described by the author in 1976.
In 79 leukaemic subjects (42 with acute leukaemia, 37 with chronic myelosis) the authors assessed HLA-DR antigens 1-5.7 and their frequency. As compared with a group of 130 healthy subjects, no significant difference was found. The results do not support a possible relationship of HLA-DR antigens and leukaemia.
For evaluation of the results of HLA examinations in paternity disputes the objective finding that the man has HLA antigens which the child inherited from the father, is decisive. The controversy as regards heredity ensuing from assessment of HLA haplotypes of the man by family studies is caused by recombination. The author presents a case of this type.
Analysis of the results of check-up examinations of HLA-A,B,C antigens in 62 cases of paternity demonstrated complete agreement of the first and second expert in 58.06% cases. Differences in 26 cases (= 41.94%) consisted in 10 cases in the detection of different HLA antigens from the first and second expert (owing in 6 cases to the different conclusion of the experts' account) and in 16 cases in the evidence of some HLA antigen only from one expert (usually by the revision). The most frequent errors pertained to the evidence of HLA antigens A28, A29, A30/31, A33, B16, B17, B21, B18, Bw22 and of the antigens of HLA-C locus; however, the well demonstrable HLA antigens A9, A10, B8, B13 were also not proved. The causes of the mistakes were analysed. It is necessary to avoid the errors of the administrative character and by the evaluation of the results in doubtful cases.
In 1974-1992 in the HLA reference laboratory of the Institute of Haematology and Blood Transfusion in Prague 15,731 sera sent by transfusion departments were examined; 4368 sera met the standard required for HLA typing, detecting 9 HLA-A, 18 HLA-B and 3 HLA-C antigens. Against these antigens with the exception of HLA-B41 and public antigens HLA-Bw4 and HLA-Bw6 monospecific sera were found: against 15 HLA antigens in a large number and against 12 HLA antigens--2 to 21 for the given antigen. Sera against subtypes of HLA antigens were sought only in rare instances, as in common practice it is sufficient (and sometimes essential) to type only basic HLA antigens.
Lymphocytes treated with a 6% and 4% AET solution after 60 or 120 minutes at 20 degrees or 37 degrees C display polyreactivity. Polyreactivity does not develop after short-term treatment (1-10 mins.). After treatment of lymphocytes with a 3% or 2% AET solution for a period of 20 or 30 mins. at 20 degrees or 37 degrees C the specific expresivity of HLA--A, B antigens rises; they are manifested, contrary to untreated lymphocytes, also by weak HLA sera. Addition of AET to the mixture of lymphocytes--HLA serum causes in the first stage of the microlymphocytotoxic test negativity of the test due to a block of the haemolytic activity of rabbit complement.
Forty-five specimens of lymphocytes from HLA-B27 positive subjects were examined by the micro-lymphocytotoxic test with 48 anti-HLA-B27 sera and the results were evaluated statistically by calculation of the correlation coefficient r. Based on the results of the statistical evaluation two groups of sera were determined which determined two groups of lymphocytes, carriers of different HLA-B27 subtypes. Six sera determined subtype B27P1, detected in four lymphocyte samples, another six sera subtype B27P2 found in three lymphocytes. The results of the statistical evaluation correspond to the serological reactions. The results indicate that HLA antigens can be reliably differentiated by simple methods.
The first allogenic bone marrow transplantation (TKD), when for the preparation whole body irradiation was used, was implemented in the Institute of Haematology and Blood Transfusion (UHKT) in Prague in 1986. Before June 1992 36 TKD were performed incl. 28 allogenic, 2 syngenic and 6 autologous. For the first time bone marrow from a non-related donor was transplanted. Of 30 allogenic and syngenic TKD to the present time 17 patients survive, i.e. 56.6% of the whole group. According to individual diagnoses 8 patients with the diagnosis of chronic myeloid leukaemia (CML) survive, 5 of 10 patients with the diagnosis of acute leukaemia (AL) and 3 of 4 patients with the diagnosis of severe aplastic anaemia (SAA) or with Fancon's anaemia (FA) resp. The survival period of the whole group is from 1-62 months since the transplantation. The main cause of death of 8 from 13 patients who died were infections associated with acute or chronic disease of the graft against the host (GVHD). In autologous TKD the bone marrow was treated with etoposide. Of the six transplanted patients with AL five survive 1.5-30 months after transplantation. The authors present some general information of pretransplantation preparation, prevention of GVHD, its incidence and results of TKD.
The author analyzed possibilities as regards the solution of paternity suits where paternity depends whether on the HLA 1 locus he is a homozygote or not. The authors presents a case of solution of a suit where the result depended on whether the man is a HLA-A2 homozygote or whether he has on the locus along with HLA-A2 also HLA-28 antigen.
The author investigated the effect of 20%, 15%, 10%, 5%, 2.5%, 1.25% chloroquine on HLA-A,B antigens on lymphocytes and on the reaction HLA-A,B antigen-antibody. HLA-A,B antigens are not released by chloroquine from the lymphocyte surface and their expressivity does not change, as was revealed by examination of the micro-lymphocytotoxic and absorption test. Higher concentrations of chloroquine cause poly-reactivity of lymphocytes. A 20%, 15% and sometimes also 10% and 5% of chloroquine solution block the reaction HLA-A, B antigen--HLA antibodies. The reason for this is the destructive action of chloroquine on rabbit complement.
Anti-idiotype antibodies against HLA specificities anti-HLA-A1, -A2, -A9, -A10, -B7, -B8, -B12 were detected in sera of 13 patients after transplantation of bone marrow and in five patients who had no transplantation but many blood transfusions. They had a multispecific character, inhibited concurrently 4-5 of the mentioned HLA antibodies. HLA antibodies were not present in sera of these patients and did not develop even after treatment with dithiothreitol. It seems therefore that anti-idiotypic antibodies have also a IgG structure. The bond of anti-idiotype antibodies with HLA specificities was firm and maximal: they were not released after 24 hours incubation at room temperature and did not form bonds with other HLA antibodies with the same specificity. The authors discuss the importance of anti-idiotype antibodies against HLA specificities for the outcome of bone marrow transplantation and therapeutically successful thrombocyte transfusion.
As part of research of juvenile hypertension the authors assessed the types of HLA antigens loci A and B in 242 children, adolescents and young adults with asymptomatic primary hypertension. The control group was formed by 300 normotonic blood donors. In the group of hypertonic subjects a significantly higher frequency of HLA-B5 antigen was found and a lower frequency of HLA-B8 and B35. Classification of the group of patients by the type of raised BP revealed the following: in subjects with a variable BP, as compared with the control group, antigens HLA-A1, B8, B35 and B40 were less frequent, in subjects with a permanently elevated blood pressure antigen HLA-B5 was more frequent and conversely antigen HLA-B35 was less frequent than in healthy subjects. From the investigation ensues a probable relationship between the presence of antigen HLA-B5 and the greater risk of the subsequent manifestation of the disease and conversely between the presence of antigen HLA-B35 in subjects at risk and probably a favourable prognosis of blood pressure readings. Contemporary findings cannot yet be considered unequivocal and it will be necessary to test their validity by extending the number of examined subjects.
HLA-DR antigens were examined in peripheral lymphocytes by means of a one-colour fluorescent microcytotoxic test using ethidium-bromide. Evidence of HLA-DR antigens was equally reliable as the two-colour fluorescent test. Contrary to the latter, however, faint non-specific positive results were obtained in sera not containing HLA-DR antibodies. This is caused probably by the fact that ethidium-bromide penetrates into cells with a very slightly damaged membrane. This reduces substantially the possibility of introduction of the method into common practice.
HLA sera were absorbed with erythrocytes with proved HLA antigens and blood cells with HLA-A, B antigens absorbed in vitro from serum. Both types of red cells were 5x to 15x washed in saline and exposed for two hours to the action of 15% chloroquine. The degree of absorption of HLA antibodies and the decline of the score were equal or slightly different after absorption with erythrocytes with own or quinked HLA antigens. Washing in saline or processing of blood cells with chloroquine did not affect the resulting absorption. From the results of the experiment ensues that the link of HLA-A, B antigens is equally firm blood cells isolated from the peripheral blood stream and blood cells where the antigens were linked to them in vitro from serum. This suggests that the view is justified that the majority of HLA-A, B antigens detected in blood cells is secondarily linked from plasma.
Typing of 38-43 HLA antigens ruled out paternity in 33.75% of 726 men suspected, during 198 revisions in 71.72%. In 63.57% the man was ruled out concurrently in loci HLA-A and B incl. in 79.33% where the man and child had all HLA-A, B antigens different. Locus HLA-A alone was ruled out in 8.01%, locus HLA-B in 28.17%; in a single case the man was ruled out in locus HLA-C only. By family investigations the man was ruled out in three of 11 cases. In two cases of paternity of twins the man was ruled out from paternity only of one child. In one case superfecundation was involved, confirmed in systems Duffy, Inv, Hp, AP, in the second case the man was ruled out from paternity of the second child in the Hp system. Male carriers of antigens of haplotypes frequent in the population were not ruled out from paternity in high percentages: with HLA antigens A1, B8 in 37.19%, A2, B7 in 19.54%, A2, B12 in 24.72%, A3, B7 in 37.14%.
The author describes his own modification of a two-colour fluorescent technique for typing of HLA-DR antigens. Concurrent examination of HLA-DR antigens in 34 subjects by this method and the monocytotoxic or B-lymphocytotoxic test revealed a greater sensitivity of the fluorescent technique; there was however also a tendency of non-specific positivity of results. More reliable results were obtained when the fluorescent technique was performed on Hammat plates, as compared with examinations using Terasaki plates. To eliminate erroneous results, the author recommends to use for HLA-DR typing two methods concurrently.