Figure 4. Figure 4. Proposed Derivation of Various Tissues in the Patient. The findings were based on the results of polymerase-chain-reaction analysis. Both cell lines are represented to some extent in all tissues except blood.

This case represents an unusual example of tetragametic chimerism in a phenotypically normal, fertile XX/XX female who had no evidence of chimerism in peripheral blood. Figure 4 outlines the probable cause of this chimerism: separately fertilized XX zygotes, one with HLA haplotypes 1 and 3 and the other with haplotypes 2 and 4, are thought to have fused early in development. The distribution of cell lines varied in individual tissues, except in blood, which appeared to be derived from only one cell line, bearing HLA haplotypes 1 and 3. It is highly unlikely that the levels of the second cell line were below the limits of detection of our assays; we used sensitive techniques and multiple informative probes, which we have shown can identify as few as 1 in 100,000 cells in experimental mixes of two cell populations (unpublished data). Because of the single cell line in our patient's blood, blood-based studies of blood groups,14 molecular HLA typing,34 and DNA polymorphism analysis,10 which have all been used to identify chimeras, were not informative.

We are aware of only two other possible cases of human tetragametic chimeras with single cell lines in blood. In one case, discrepancies in the blood type between a woman and her children suggested that she was not the biologic mother.35 As in our patient, the children's nonpaternal haplotype was identified in maternal grandparents. However, the authors estimated that they would not have been able to detect a population of cells that was less than 0.5 percent of peripheral-blood cells.36 In another patient, a phenotypically normal man whose red cells were blood group B, chimerism was detected because of a surprisingly weak titer of antibody against group A and small amounts of group A substance on his red cells. The patient proved to be an XX/XY chimera with only XY lymphocytes in his blood. The XY line produced group B red cells; the XX line encoded a group A glycosyl transferase. The activity of this enzyme in nonhematopoietic XX tissues resulted in group A substance that was passively adsorbed by the patient's XY group B red cells.9

In a mouse model of tetragametic chimerism, in which blastomeres from two embryos were cocultured to form a chimera, 12 of 34 such mice had only one red-cell population in the blood even though they had two cell lines in other tissues.4 This finding could be consistent with the presence of a single cell line of clonal origin beginning early in development.37 Alternatively, a selective advantage could have caused one clone to be selected early in life. The latter possibility is supported by the finding, in a study of tetragametic rams, that one of the two red-cell lines completely disappeared over a period of five years in two of four chimeric animals.5

Because of the apparent rarity of tetragametic chimerism and the importance of the use of molecular techniques to confirm its presence, this condition may be underdiagnosed. Furthermore, if a single cell line predominates in the blood, the chimeric state may not be detected unless family studies are undertaken. Even then, the findings may be misinterpreted as ruling out maternity or paternity. Molecular studies of other tissues for chimerism should be considered in such cases. Furthermore, the need to consider this diagnosis may be increasingly relevant: in vitro fertilization is associated with a 33-fold increase in twinning16 and an increased incidence of tetragametic chimerism, possibly because the embryos are in close contact and fuse before they are implanted16 or because of double fertilization of an ovum with two nuclei.38,39

Finally, the tetragametic state has important implications for organ or stem-cell transplantation. Chimeras typically have immunologic tolerance to both cell lines. Even though our patient had only one cell line in her blood, her T lymphocytes did not respond to cells from family members with any combination of the four familial HLA haplotypes. These results are consistent with those of studies of tetragametic mice with single red-cell populations, which also demonstrated tolerance to skin grafts from parental strains.4 Thus, for a tetragametic human, a wider array of relatives (including, in our patient, all her children) and other persons may be eligible to be organ donors.