Lillie's Freemartin Research

Lillie's research on the freemartin, described earlier, was perhaps the most famous piece of reproductive science to come out of the Department of Zoology at the University of Chicago. According to Lillie, the bovine freemartin was a "natural experiment." Lillie's research (1916b, 1917a,b) ultimately revealed that a freemartin is a sterile female co-twin to a male, from a separate egg, but whose chorionic vessels (placentas) have merged with the male's in utero, allowing crossing of blood systems. That blood-borne chemicals—hormones—were consequential in embryological development was the major finding of this work.

The research began in 1914 when the manager of Lillie's private farm sent him a pair of twin calf fetuses with their membranes intact.^[44] Leon J. Cole, of the Department of Experimental Breeding in the College of Agriculture, University of Wisconsin, heard of Lillie's work on this problem and contacted him in hopes of correlating their efforts. Cole's department was working on multiple births in cattle: twins, double monsters, and freemartins.^[45] There was considerable sharing of materials between the two emerging centers of reproductive science because the freemartin problem was important to both.^[46] Lillie and his group relied primarily on the Chicago stockyards for freemartin material, acquired through a foreman at a Swift and Company abattoir and through the special efforts of the depart-

ment's collector. Lillie noted that "every uterus containing twins below a certain size from a certain slaughter house is sent to me for examination without being opened."^[47]

Lillie first investigated whether the twins came from the same or separate eggs. Prior research had argued for male twins from the same egg, largely because the twins were monochorial (attached to a single placenta), a phenomenon usually associated with one-egg twins. However, it did not make sense to Lillie that only one of a pair of male twins would be affected in utero. Lillie, who began to examine corpora lutea (sites of recently released eggs) in the ovaries, wrote Cole: "I am faced with the irritating difficulty that most of the uteri are received with one or both ovaries missing," making it impossible to determine whether one or two eggs had been released.^[48] Sufficient material gradually was amassed to demonstrate that there had consistently been two corpora lutea (one in each ovary), and two originally separate chorionic vessels (placentas) had fused in utero. Since the freemartin usually possessed mammary glands and both female and male external genitalia, and since if it were male, sex ratios would be strangely skewed, Lillie (1917a,b) concluded that it began as a female.

In 1916 both Cole and Lillie published early abstracts of their work in Science . Cole, reflecting his more genetic concerns, focused on sex ratios. Lillie (1916b:612, emphasis added) focused on sex differentiation through the exchange of blood between fetuses: "If one is male and the other female, the reproductive system of the female is largely suppressed, and certain male organs even develop in the female. This is unquestionably to be interpreted as a case of hormone action. It is not yet determined whether the invariable result of sterilization of the female at the expense of the male is due to more precocious development of the male hormones, or to a certain natural dominance of male over female hormones ." In his classic paper, published in 1917, Lillie emphasized that a vascular connection between the fetuses is requisite for development of a freemartin, and that influences of blood-borne hormones were acting on extant rudiments in the bisexual embryo stage. He concluded (1917a:415): "The course of embryonic sex-differentiation is largely determined by sex-hormones circulating in the blood." Figure 1 is one of the images that appeared in Lillie's article.

Such research continued at Chicago for some years.^[49] Lillie noted that the work had "wider application than we expected ... embryonic and astomoses blood vessels may have quite different results in different animals."^[50] Lillie also found that intersexes in goats and swine may be genetic rather than hormonal in origin.^[51] In 1917 he had thought that sex hormones were intensifiers of gene action, but by 1932 subsequent research had demonstrated the complete absence of sex differentiation in the absence of sex hormones (Lillie 1932:1–11, esp. 6; Danforth 1932). The freemartin work generated considerable interest in agricultural, popular sci-

Figure 1.
Lillie's freemartin research materials, Lillie (1917a).

ence, and medical circles. One popular article noted: "Twins in cattle may be about two percent of all births in some breeds and the two sexed twins form about half of all the twin births, making the matter of sterile cows that produce no milk of economic importance to the dairy industry."^[52] This work deepened connections among Lillie's group and agriculturally oriented centers of reproductive science headed by Cole at Wisconsin and by F. A. E. Crew at Edinburgh.^[53] Lillie's work is also cited as pathbreaking in terms of developing theories of immunological tolerance important in medicine (Billingham and Beer 1984). Such concerns were indicated by a popular science service's request to Lillie for a simple account of it in 1922: "There is a wide public interest just now in the subject of endocrinology. In fact the public seems ... to take it up as a fad in succession to the Freudian complexes now going out of fashion."^[54]

The importance of the freemartin work was multifold. First, the work clearly demonstrated hormonal influence on sex differentiation in utero. Thus the production of sex, a classic turn-of-the-century biological problem, involved not only genetic but also physiological processes. Second, the freemartin research "introduced biologists to the problems of the nature, origin, and action of sex hormones at a time when almost nothing was known about the subject" (Willier 1957:219). One might even say that Lillie imported endocrinology into the embryology of his day.^[55] Third, problems of sexual differentiation that the work posed were central to several other major reproductive investigations, including work by Emil Witschi, Carl Moore, and Dorothy Price.^[56]

The thorny problems of what "determines" sex versus what "differenti-

ates" organisms by sex during development that so absorbed Frank Lillie are classic examples of "boundary line" research. Star and Griesemer (1989) have found that such problems often become "borderline sinkholes," unresolved and commonly ignored. In fact, some years after his original work, Lillie (1932:5) reasserted that "we must make a radical distinction between" the two, with sex determination as chromosomal (genetic) and sex differentiation, in higher animals, as hormonal. That distinction has not held. Over seventy years after the freemartin research, many aspects of these problem have yet to be clarified and remain quite challenging.^[57] The biologist Anne Fausto-Sterling (1989, 1993, 1998) asserts that these problems may be particularly complicated by the sustained view among scientists that sex is dimorphic (divisible into two categories) rather than continuous, despite much empirical evidence to the contrary. The confounding of sex and gender by many reproductive scientists, and their commitments to dimorphic sex and to gender differences, may be contributing to the scientific problem.