THE DISTRIBUTION OF THE ABO BLOOD GROUPS IN PERSONS WITH LEWIS ANTIBODIES

Transcription

1 THE AMEHICAN JOURNAL OF CLINICAL PATHOLOGY Copyright 1909 by The Williams & Wilkins Co. Vol.51, 1 Printed in U.S.A. THE DISTRIBUTION OF THE ABO BLOOD GROUPS IN PERSONS WITH LEWIS ANTIBODIES RICHARD I-I. WALKER, M.D., LARRY D. GRIFFIN, MT (ASCP) BB, AND MARK KASHGARIAN M.D., M.P.H. Department of Pathology, University of Tennessee, and the Blood Bank and Transfusion Service, City of Memphis Hospitals, Memphis, Tennessee The independence of the Lewis blood group system from the ABO system was first established by Mourant in and has been confirmed by Race and Sanger. 27 Many associations have been observed between these systems, however. Among these is the similarity of the biochemical characterization of ABH and Le" substances, 16,18 ' 5, 16,24 lo, 24,29 serologic cross-reactions, the proposed common genetic pathways for the production of these substances, 8,31 the variation in the frequency of the Le b red cell phenotype 3 and the strength of the expression of the Le" red cell phenotype 11 depending on the ABO phenotype, the suppression of the Le x character by the Ai gene, 2 the relationship between ABH secretion and the Lewis red cell phenotype, 13, u the Magard serum, 1 the close association of Le b and H, 5,35 and the possible correlation of the ABO and Lewis phenotypes on blood lipids. 6 Jordal in first recognized the high frequency of the Group A phenotype among 23 persons with anti-lc 11 and anti-le x in their sera. He further noted that the 22 anti-lewis sera previously reported came from persons with an excess of the Group A phenotype. Giblett and Mollison 22 noted a decrease in Group 0 persons among 66 donors of anti-lc". Similar findings were reported in 1965 by Kissmeyer-Nielsen in a large Scandinavian population. 17 The present retrospective study from our laboratoiy was conducted because of the apparent difference of ABO blood group distribution among a predominantly Negro population with Lewis antibodies, in contrast to the population from which they came. METHODS AND MATERIALS The subjects of this study were 3S6 persons with Lewis antibodies in their sera. Received February 2G, Blood specimens from these persons were processed by the Blood Bank and Transfusion Service of the City of Memphis Hospitals during the period January 1, 1963, to November 29, All persons in whom Lewis antibodies were identified during this period were included and their ABO groups were noted. The City of Memphis Hospitals is a large general hospital complex which serves primarily the indigent population of Memphis and surrounding areas, and its patients are predominantly Negro. The expected ABO phenotype frequencies were established by tallying the ABO groups of S903 inpatients from January 1, 1963, to July 1, These were determined from consecutive samples submitted to the blood bank for typing and crossmatching. The ABO groups of approximately 2000 consecutive prenatal outpatients and 2000 consecutive donors (selectively drawn for optimal inventory group distribution) were also tallied for comparison. It was from this same population (inpatients, prenatal outpatients, and donors) that the persons with atypical antibodies were derived. An additional comparison was made between the ABO phenotypes of persons with Lewis antibodies and the ABO phenotypes of persons with atj'pical antibodies which were other than Lewis in specificity (non-lewis). Antibody screens are conducted on all samples submitted, using saline-washed Group O reagent red cells, human.* A 30- min. incubation at room temperature and reading after light centrifugation were followed by a 15-min. incubation at 37 C. and the saline-indirect antiglobulin test.t Sera with macroscopically positive tests were * Hemantigen, Pfizer Diagnostics, Inc., New York, N. Y f Anti-human serum (7 + non-7), Ortho Diagnostics, Raritan, N. J

2 WALKER ET AL. Vol. 51 TABLE 1 FREQUENCY AND PERCENTAGES OF ABO PHBNOTYPES Jndi\ iduals with An ibodies in th eir Sera* Phenotype Anti-Le" Anti-Le* Non-Lewis antibodies Inpatient population A B 0 AB Total S S IS * Does not include 24 persons with anti-le 1 ' antibodies. This group was too small to evaluate. then tested against a commercial cell panel* in saline at room temperature and at 5 C. and in albumin at 37 C, and the albumin-indirect antiglobulin test (Ortho Diagnostics). Only sera in which the antibody specificity was established were included in this study. Frequently the specificity was confirmed by a second red cell panel* when the first study was not decisive. On a few occasions, the test cells were trypsinized for Lewis problems, but this was not performed routinely. The Lewis antibodies were not tested against cord cells or adult cells of phenotypes other than O. ABH secretor status was not determined in the donors of these Lewis antibodies. The anti-lc" sera agglutinated Le(a+b ) adult Group O red cells and showed no activity against Le(a b+) or Le(a b ) cells under the conditions tested. Anti-Lc 1 ' sera agglutinated Le(a b+) cells and failed to agglutinate Le(a+b ) or Le(a b-) cells. Anti-Lc x is defined in this study as an antibody which agglutinates adult Group O red cells of the phenotypes Le(a+b ) and Le(a b + ) but fails to agglutinate adult Group O red cells of the Le(a b ) phenotype (a modification of the proposal of Miller and associates 21 )-t Although the * Pfizer Diagnostics, Inc., or Hyland Laboratories, Los Angeles, Calif f The term anti-le* was first, proposed by Miller and associates 21 to indicate a Lewis antibody which cross-reacts with the Le" and Le b determinants. term anti-lc" + Lc 1 ' is frequently used to describe this antibody, anti-le x seems more appropriate because it denotes the crossreacting character of most of these antibodies. Absorption and elution tests to document this cross-reacting component were not performed on the anti-le x sera in this study. The anti-le x sera thus probably represent a heterogeneous group as suggested by Andersen. 2 RESULTS Table 1 gives the frequencies and proportions of the ABO groups of the subjects in the study. The ABO frequencies of the "blood bank" inpatients found in the last column of Table 1 were used as the expected frequencies. This was our largest source of patients with antibodies; the ABO frequencies and proportions of the other two source groups did not differ significantly from this group. The ABO frequencies of the inpatients also do not differ significantly from those published for the American Negro. 26 The phenotypes of a sampling of persons with non-lewis antibodies are found in the third column of Table 1. The 259 individuals of this group are given as a secondary control to compare with the phenotypes of persons with Lewis antibodies given in the first and second columns of Table 1. However, we have not used this control for statistical comparisons with the groups possessing Lewis antibodies. The frequencies of the phenotype Groups A, B, and AB are all increased and the fre-

3 Jan ABO GROUPS IN PERSONS WITH LEWIS ANTIBODIES quency of Group 0 is decreased in the groups with Lewis antibodies when compared with the expected percentages derived from our inpatients or with the group of 259 persons with non-lewis antibodies. These differences can perhaps be more readily seen in Table 2, in which the gene frequencies of the four groups are given. The method of calculation is that of Fisher and Taylor 27 in which the AB phenotype is not used to estimate the gene frequencies, inasmuch as the numbers in this group with antibodies are relatively small and the AB group is thus subject to errors. To speak of an increase in A and B or a decrease in 0 is to refer to fundamentally the same phenomenon in a distribution of this sort. The decision as to whether there is a primary increase or decrease in a specific phenotype must come from biologic evidence rather than from a statistical evaluation of the distribution of phenotypes. The statistical evaluation of the phenotypes is shown in Table 3. Six comparisons are given in this table. The groups with anti-le" and anti-le x are compared with Gene A B 0 TABLE 2 ESTIMATED GENE FREQUENCIES Anti-Lc" Study Groups Anti-Le" Non-Lewis antibodies Inpatient population expected percentages derived from the inpatient group previously mentioned and with each other. The first three comparisons are between the distribution of A, B, O, and AB phenotypes (4 X 2 x 2 tests) and the second three comparisons combine the frequencies of A, B, and AB phenotypes so that the O phenotype can be compared with the non-0 phenotype (2 X 2 x 2 tests). The group with anti-le x is highly significant when compared with the expected frequencies or with the anti-lc" group. The distribution of ABO phenotypes is also significantly different between the anti-le" group and the expected percentages. DISCUSSION As can be seen from an inspection of the three tables, the ABO phenotj'pe distribution of the anti-le" and the anti-le* groups is distinctly different from that of the general population. Because of the small numbers, no statistical evaluation of the anti-le b group can be made. However, the ABO phenotype distribution in this small anti-lc h group appeared similar to that observed in the anti-le x group. It should be emphasized that we have considered the groups anti- Le" and anti-le x as mutually exclusive, although this may not be correct. Wiener 35 ' 36 considers anti-le x to be a more potent anti-le" which is capable of detecting the Le" character in persons of the apparent erythrocyte phenotype Le(a b+). It has been reported that all persons with TABLE 3 STATISTICAL COMPARISONS OF PHENOTYPES Comparison X* df Probability ABO phenotype Anti-Lc" vs. expected* Anti-Lc 1 vs. expected* Anti-Le" vs. anti-le* O and non-0 phenotypes Anti-Lc" vs. expected* Anti-Lc* vs. expected* Anti-Lc" vs. anti-lc* * { 2t 2t <0.05 (significant) <0.02 (significant) * Expected value calculated from percentages of inpatient population. f One degree of freedom (df) subtracted for comparison with an expected frequency. t x 2 calculated using Yates' correction.

4 6 WALKER ET AL. Vol. 51 anti-le" in their sera are Le(a b ) and are secretors of ABH substances. 21,36 A possible mechanism for the unusual ABO group frequencies in persons with Lewis antibodies is suggested by the evidence that Le(a b ) nonsecretors of ABH apparently do not produce anti-lc". 21 This may be a result of the close chemical similarity between Le" substance and precursor substance which these persons have present in large amounts, according to the theory of Morgan, 24 Watkins, 31 and Ceppellini. 8 Biochemical studies have indicated a close similarity in the characterization of precursor substance, Le!V, and H substances. 16,24,29 Bird 5 has demonstrated a single substance common to all types of saliva, purified blood group substances, the "Fl" material, and Type XIV pneumococcal polysaccharide by the Ouchterlony technic, using the Ricinus communis precipitin. Titration studies by Bird 5 indicated that quantitative differences existed between these substances with the "Fl" material and Type XIV polysaccharide having the greatest activity, less activity in H and Le a substance, and least activity in A and B substances. Similar cross-reactivity has been described by Rabat 10 and Morgan 24 using Type XIV pneumococcal antibody. These similarities of autogenous macromolecules, precursor substance and Le a, may be responsible for the suppression of antibody response 9 to Le a substance in Le(a b ) ABH nonsecretors. Miller and associates 21 found that Le(a b ) nonsecretors of ABH always had "Le a " substance in their saliva at a 1:5 dilution and frequently in a 1:10 dilution, whereas Le(a+b ) ABH nonsecretors always had complete inhibition titers for Le" substance in 1:100 dilutions of saliva and generally in 1:1000 dilutions. The partial inhibition of anti-lc a serum by saliva from the Le(a b ) ABH nonsecretors has been confirmed by Andersen 2 and Bianco and colleagues. 4 This is most likely the result of a cross-reaction of anti-le" with precursor substance. Cross-reactions between Le a and Le b substances have also been postulated by Miller and associates. 21 The low frequency of Group O persons with Lewis antibodies could be explained by the similarities between H substance and Le" substance, inasmuch as Group O persons have the greatest amounts of H substance and may fail to recognize Le a substance as foreign or "not self." Similarities between the H and Le a specificities have been demonstrated by Watkins and Morgan 33 using inhibition tests with 1-fucose. Immunologic responsiveness of an individual is dependent in part on the structure of autogenous macromolecules. Thus, the immunogenicity of Lewis substances is indirectly dependent on the genes responsible for the synthesis of related autogenous macromolecules. The structure of the nonreducing terminal ends of the ABH and Lewis determinants has been studied extensively by Morgan, 24 Watkins, 32 Schiffman and Marcus, 29 and Lloyd and associates. 18, 19 The molecules differ only in the substitution of monosaccharide units at the nonreducing ends. These few substitutions account for the separate serologic specificities. Aside from these terminal substitutions, the molecules are remarkably similar. Weigle 34 has shown that tolerance to serum protein antigens is directed not only to the individual determinant groups but also to the over-all chemical and physical properties of the antigen. Group O persons may be more "tolerant" to Lewis substances than are persons of other phenotypes because of the close similarities between H and Lewis substances. 23,30 The presence of the Se gene seems to be essential for anti-le" production, but the amount of H substance appears to influence predisposition to anti-le" formation. The quantity of ABH produced by secretors has been suggested to be under genetic control, 10 which might also influence suppression of anti-le a production in persons with the greatest amounts of H substance. Lodge and colleagues 20 have recently presented evidence for cross-reactions of anti-le x with precursor substance and H substance. The Aagaard type sera (anti-le bl ) provide additional evidence for similarity between H and Le a substances. These sera have specificity for the product of the Se and Le genes (called Hi by Bird 6 and Wiener 36 ) but

5 Jan ABO GROUPS IN PERSONS WITH LEWIS ANTIBODIES they may weakly cross-react with separate H or Le" substances in the saliva of persons possessing either the Se or Le gene, respectively, but not both. 7 ' 8 Thus, we do not believe that an antibody with anti-le bl specificity will be found in a person of the phenotype Le(a+b ), ABH nonsecretor. Three persons have been reported who possess anti-lc 1 ' 11 in their sera who are Le(a+b ) ABH nonsecretors. 7,12 Persons with anti-le b in their sera are almost always Le(a b ) and are nonsecretors of ABH substances. 21 ' 30 Miller and associates 21 found that the majority of Le(a b ) secretors of ABH had significant amounts of substance in their saliva inhibiting anti-lc b (anti-le bh ) and trace amounts were often found in the remainder of this phenotypic group. Predisposition for anti-lc b production is thus also modified by the ABO group showing an even more pronounced reciprocal relationship with the quantity of H in the body. This accounts for the presence of anti-h in many anti-le b sera. Indeed, Race and Sanger 28 state that they have not observed an anti-lc b in the serum of a Group 0 person. It has been proposed 21 that the weakness of Le b reactions with blood of Groups Ai, B, and AiB is a result of their relative lack of H substance, rather than of epistasy. 3 SUMMARY The frequencies of the ABO groups in Negroes with Lewis antibodies were compared with their frequencies in the parent population and with a control group of persons with antibodies other than Lewis in specificity, derived from the same parent population. The statistical analysis of Negroes with Lewis antibodies revealed a significant decrease in the frequency of Group 0 individuals and an excess of Groups A, B, and AB persons, in contrast with the two control populations. This was most striking in persons with anti-lc x and to a lesser degree in persons with anti-lc". This is an additional association between the ABO and Lewis blood group systems. A mechanism is postulated based on genetic factors and on the dependence of antibody responses on the structure of autogenous macromolecules. The observation that anti-lc" is produced only by secretors of ABH suggests that precursor substance may suppress anti-lc" production. The pattern of ABO phenotype frequencies in persov.s with Lewis antibodies tends to be inversely proportional to the amounts of H substance present in persons of different ABO phenotypes. Thus, although an ABH secretor environment appears to be essential for anti-lc" production, the amount of H substance present appears to influence predisposition to the production of anti-lc". This theory is consistent with biochemical and serologic evidence of similarities between H and Le". REFERENCES 1. Andersen, J.: Modifying influence of the secretor gene on the development of the ABH substances: a contribution to the conception of the Lewis group system. Vox Sang., 8: 251-2G1, Andersen, J.: Serological studies on two Lewis sera. Acta path, et microbiol. scandinav., 48: 374-3S4, Andresen, P. H.: The blood group system L, a new blood group L 2, a case of epistasy within the blood Groups. Acta Path, et microbiol. scandinav., 25: , Bianco, I., Silvestroni, E., Lawler, S. D., Marshall, R., and Siniscaloo, M.: Further contributions to the study of Lewis and secretor characters. Vox Sang., 5: S, Bird, G. W. G.: llaemagglutinins in seeds. Brit. M. Bull., 15: 1G5-16S, Blankenhorn, D. H., Jensen, J., Chin, H. P., and Sturgeon, P.: Genetic influence upon blood lipids: an unexpected correlation with ABO and Lewis blood groups. J. Clin. Invest., 43: 1262, 1964 (abstract). 7. Brendemoen, O. J.: Further studies of agglutination and inhibition in the Le"-Le b system. J. Lab. & Clin. Med., 36: , Ceppellini, R.: Physiological genetics of human blood factors. In Ciba Foundation Symposium on Biochemistry of Human Genetics. Boston: Little, Brown and Company, 1959, p Cinader, B.: Dependence of antibody responses on structure and polymorphism of autologous macromolecules. Brit. M. Bull., 19: , Clarke, C. A., MeConnell, R. B., and Sheppard, P. M.: A genetical study of the variations in ABH secretion. Ann. Human Genet. 24: , Cutbush, M., Giblett, E. R. and Mollison, P. L.: Demonstration of the phenotype Le(a+b+) in infants and adults. Brit. J. Haemat., 2: , Garratty, G., and Kleinschmidt, G.: Two ex-

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