H-Y antigen and sexual dysgenesis in man

H-Y antigen is a surface component associated with the heterogametic sex of various species and supposed to induce testicular differentiation. Genes controlling directly or not the expression of H-Y antigen and testicular differentiation have been localized on Y as well as on X chromosome and even autosomal chromosome. However the genetical localization of the H-Y structural gene remains unknown. We analysed the expression of H-Y antigen in three types of sexual dysgenesis (males bearing XX caryotype, testicular feminization syndrome and one case of hermaphroditism) to clarify the function and the genetics of this antigen.     

Genetic aspects of H-Y antigen

Summary While it remains to be clarified what detection of H-Y antigen by current methods means, the existence of a factor governing testicular differentiation of the indifferent gonadal anlage seems to be well established. There are various kinds of evidence that H-Y antigen as a biologically meaningful factor has a complex genetical basis. There is the contribution of the Y chromosome which, independent of the number of other chromosomes, especially of X chromosomes, leads to a male phenotype. The X chromosome must be involved also because structural aberrations of its distal short arm influence the expression of the H-Y structural gene. Due to examples of autosomal inheritance of various forms of sex reversal, an autosomal gene is assumed to be involved as well. Arguments are presented favoring the assumption that the structural H-Y gene is autosomal, while genes on the X and Y chromosomes have a controlling function.This genetic control mechanism for H-Y antigen seems to have evolved secondary to placentation in mammals. In non-mammalian vertebrates, H-Y antigen is controlled by other factors, e.g. steroid hormones. While the functional role of H-Y antigen in directing differentiation of the heterogametic gonad appears to have been preserved during evolution, the mechanism of its control has changed. This latter mechanism is only poorly understood.     

Immunological and functional aspects of H-Y antigen

Summary Male-specific H-Y antigen may be defined by graft rejection, killer cell action or antibodies. Most commonly H-Y antigen is detected in assays using H-Y antisera. In these tests errors may arise from various causes: 1) Auto- and heteroantibodies cross-reacting with target cells. 2) Restriction phenomena. 3) MHC-dependent modification of the amount of H-Y antigen present on different tissues. 4) Modification of cell surface antigens by bacteria or viruses.Regarding the third definition of H-Y antigen, four different states can be distinguished in the mammalian male. H-Y occurs (1) as an integral part of the plasma membrane; (2) unspecifically attached to the membrane of human erythrocytes; (3) free in solution; (4) bound to its gonad-specific receptor.Redistribution experiments suggest that H-Y and ₂-m are associated on the cell membrane. Coredistribution is not found of H-Y and MHC antigens. An antibody blocking technique demonstrates association of H-Y and H-2D antigens on unfixed lymphoid, but not on testicular cells. Human erythrocytes lacking ₂-m do not integrate H-Y antigen into the cell membrane. Male erythrocytes, however, absorb H-Y antigen from the serum. The origin of H-Y antigen in the serum is not clear. It may be shed from cell membranes, derive from the testis which actively secretes H-Y antigen, or both.H-Y antigen is bound by a gonad-specific receptor. This receptor is present in the gonads of both sexes. H-Y antigen is supposed to mediate testis differentiation via this receptor. Reaggregation experiments in vitro using dissociated gonads of the newborn rat demonstrate that ovarian cells reorganize into testicular structures in the presence of H-Y antigen. The assumption cannot be confirmed that addition of H-Y antiserum to testicular cells results in ovarian structures. This finding, however, does not conflict with the view that H-Y antigen is involved in testis differentiation, e.g. by inducing testis cell-specific functions via the gonad-specific receptor.     

H-Y antigen: localization of the H-Y gene

Testicular development in a patient with deletion of the distal (fluorescent) segment of the Y chromosome is described. The presence of a normal dose of H-Y antigen was demonstrated by Goldberg's cytotoxicity test. It is concluded that the distal fluorescent segment of the Y chromosome is void of genes regulating H-Y antigen activity.     

H-Y antigen in the teleost

H-Y antigen, presumably the product of mammalian testis-determining genes, has been detected in three species of teleost fish, Xiphophorus maculatus. Haplochromis burtoni, and Oryzias latipes, and in hybrids of the genus Tilapia. In X. maculatus H-Y was most readily detected in YY males, suggesting that a genetic determinant of H-Y antigen expression may exist on the teleost Y-chromosome. Although H-Y was detected in males and not in feamles in each of the species that we studied, male heterogamety has not been firmly established for H. burtoni. Thus despite the extreme phlyogenetic conservation of H-Y genes and their association with the Y-chromosome, it remains open to question whether H-Y will always be found in the heterogametic sex, and whether serologically defined H-Y antigen plays any part in the differentiation of the teleost gonad.     

Serology of H-Y antigen

Summary Anti-H-Y antiserum is generally obtained from female inbred mice or rats that have been hyperimmunized with syngeneic male cells. The specificity of such antiserum is defined by its reactivity for male but not female cells. A number of conventional serological assays have been used to measure that reactivity. However, H-Y is a weak antigen, evidently represented sparingly on the surfaces of cells other than sperm, epidermal cells and brain cells; thus the srological assays for H-Y are technically difficult. Yet H-Y serology has enabled significant progress toward the understanding of primary sex differentiation.A recent advance in H-Y serology is the establishment of monoclonal anti-H-Y antisera which promise to facilitate analysis and clarification of the H-Y system.     

H-Y antigen in 46,XY gonadal dysgenesis

Summary Presence of H-Y antigen has been correlated with testicular differentiation, and absence of H-Y with failure of testicular differentiation, in a variety of mammalian species. To determine more precisely the relationship between expression of H-Y antigen and development of the testis, we studied the cells of phenotypic females with the 46,XY male karyotype. Blood leukocytes were typed H-Y⁺ in five XY females with gonadal dysgenesis, although in other studies blood leukocytes from XY females with gonadal dysgenesis were typed H-Y^-. Thus mere presence of H-Y antigen is not sufficient to guarantee normal differentiation of the testis. In the present paper we review evidence for an additional factor in gonadal organogenesis, the H-Y antigen receptor. We infer that testicular development requires engagement of H-Y and its receptor. It follows that XY gonadal dysgenesis is the consequence of functional absence of the H-Y testis inducer as in the following conditions: failure of synthesis of H-Y or failure of specific binding of H-Y.     

H-Y antigen in the study of sex determination and control of sex ratio

It has been proposed, on the basis of widespread phylogenetic conservation, that H-Y antigen is the inducer of primary sex, causing the undifferentiated XY gonad to become a testis in male heterogametic species such as the human and bovine. That proposition has withstood extensive testing in vivo and in vitro. Freemartin gonads are H-Y⁺, for example, and masculinization of the freemartin gonad has been attributed to soluble H-Y, borne and transmitted in the serum of the bull twin, and bound in ovarian receptors of the female. We have applied monoclonal H-Y antibodies to the identification of gender in embryos of the bovine. Our preliminary results imply presence of H-Y in bovine embryos of the morula and blastocyst stages recovered at about 6–12 days of gestation. Assignment of H-Y phenotype -- positive in males and negative in females -- allows selective implantation of male and female during embryo transfer. Thus in an early study, we correctly identified gender in 6 of 7 calves born healthy at term, after transfer of 8 blastocysts.     

On the secretion of H-Y antigen

It has been proposed that H-Y antigen secreted by cells of the Sertoli lineage is bound by receptors on these and other cells of the primordial gonad and thereby initiates formation of the testicular cords, and that H-Y is not an integral transmembrane component but a part of a ternary system with β₂-microglobulin and products of the MHC. It follows that cultured Daudi cells, which lack β₂-microglobulin and HLA, should secrete H-Y. This is consistent with evidence obtained with monoclonal H-Y antibody and an ELISA. By this method, free H-Y was demonstrable in the supernatant fluids of cultured Sertoli cells and Daudi cells. The assay provides a useful alternative to detection of H-Y in the complement-dependent cytotoxicity test.     

H-Y antigen in transsexuality,and how to explain testis differentiation in H-Y antigen-negative males and ovary differentiation in H-Y antigen-positive females

Summary H-Y antigen was determined in eight transsexual patients. Two of the four male-to-female transsexual patients typed as H-Y antigen-negative, while the other two typed as expected from their phenotypic and gonadal sex, namely H-Y antigen-positive. Of the four female-to-male transsexual patients, three typed as H-Y antigen-positive and one was H-Y antigen-negative, as expected. The presence of normal testes in H-Y antigen-negative males is assumed to result from a mutation of nucleotide sequences of the H-Y structural gene for antigenic determinants. Thus, an H-Y is produced with normal receptor-binding activity which can sustain the testis determination of the bipotent gonadal anlage. In the case of H-Y antigen-positive females with normal ovaries a deletion of the autosomally located H-Y structural gene is assumed. This deletion should affect sequences for repressor-binding (as was suggested for H-Y antigen-positive XX-males) and for receptor-binding activity of the H-Y antigen molecule. The resulting H-Y antigen is unable to bind to the gonadal receptor of the bipotent gonadal anlage. Thus an ovary is determined. The relevance of H-Y antigen for the aetiology of transsexualism is discussed.     

Studies on the H-Y antigen in rats

The H-Y antigen has been studied under a variety of experimental conditions in BN and Lewis rats. The results indicate that 1. graft size is crucially important in determining the fate of male skin isografts on females; 2. H-Y incompatible ear skin grafts survive significantly better than those of trunk origin; 3. prior exposure of females to male lymphoid cells greatly increases their capacity to reject male skin isografts; 4. neonatal castration has no influence on the expression of H-Y; 5. multiparity can induce unresponsiveness to H-Y; and 6. although BN females respond better than do Lewis females to H-Y, the antigen is stronger in Lewis males. These findings are compared with the results of similar experiments conducted with mice.Submitted in memory of Dr. Joy Palm, member of the Wistar Institute, who pioneered the genetic analysis of histocompatibility in rats.     

XY gonadal dysgenesis and the H-Y antigen

Summary H-Y antigen was determined in 12 patients affected by XY gonadal dysgenesis. Of these, three proved to be H-Y negative, and nine, including two sisters, were H-Y positive; two of the unrelated positive cases exhibited a reduced antigen titer. Therefore, this clinical condition must be genetically heterogeneous. It is assumed that in the negative cases and possibly in those with reduced antigen titer, the H-Y generating system is affected by mutation, while in the regular positive cases the target cells are unable to respond due to a defect of the gonad-specific H-Y antigen receptor.I dedicate this article to the memory of Ilse Aschmoneit