Ground Hogs' Day Lecture, Feb 2, 2005; Biology 2005 Albert HarrisH. V. Wilson made the single most important scientific discovery ever made here at UNC. He dissociated living sponges into their individual cells (random mixtures of differentiated cells), and observed that these reformed functioning sponges within 3 days. A few years later, he dissociated tissues of a kind of coral, and they also reformed normal anatomy. Since then, others discovered equivalent phenomena in some flat-worms, in some sea squirts, in the embryos of sea urchins up to the gastrula stage, and to some extent in frog embryos up to neurulation. Just about any organ can be dissected out of mouse and chicken embryos, and dissociated into a random mixture of individual cells (using trypsin & EDTA), mixed randomly with cells of a different organ, with the result that these differentiated cells will sort out from one another, and can often form tissues that look almost normal in histological sections. (They can't re-form functioning mice or chickens, however!!) A random mixture of liver cells and heart cells will sort out to form blobs of heart cells, in the interior of a mass of liver cells. What does it tell us about normal mechanisms of embryonic development that randomly mixed differentiated cells can sort out, sometimes to the extend of re-forming normal anatomy, and other times so that heart cells find & adhere to heart cells, and liver cells find & stick to liver cells? Our textbook supports 1 of several theories on this subject. (Malcolm Steinberg's "Differential Adhesion Hypothesis" sometimes called the "Thermodynamic Hypothesis" The majority believe that cell sorting results from differences in cell-cell adhesion molecules like the various cadherins, N-CAM etc. A few embryologists, including Steinberg (and also me) believe that gastrulation and neurulation are caused by the same mechanism as cell sorting. In a sense, we think of gastrulation as being the "sorting out" of the future mesoderm and endoderm, relative to the ectoderm. And we think of neurulation as being the sorting out of the future brain relative to the future skin. (On the other hand, most people think Steinberg and I are crazy to believe that sorting has the same cause as gastrulation, etc. Steinberg & I have vigorously disagreed for 30 years about what mechanical forces cause cell sorting. He thinks it's all differences in cell-cell adhesiveness, and I think it's a mixture of cell-cell adhesion molecules (to get liver cells with other liver, & heart with heart cells) combined with differences in strengths of contractility Some of the historical development of this subject: H. V. Wilson tried to create economically useful chimeric sponges, by dissociating cells of two different species and mixing them together. This didn't work, because cells of the two species sorted out according to species, before sorting out according to differentiated cell type. Different species wouldn't stay mixed. Similarly if you dissociate sea urchin blastula-stage embryos of two different species, and mix them randomly, they will only aggregate with cells of the same species. In contrast, cells of frogs and salamanders will stay mixed, and cells of birds and mammals stay mixed. In other words, amphibian, bird and mammal cells sort out by differentiated cell type, but do not sort out (well, maybe a little bit!) by species. Wilson rejected the idea that differentiated cells can sort out according to cell type. That idea was proposed by Julian Huxley, on the basis of experiments that repeated Wilson's, using different species of sponges. Wilson first believed that the randomly-mixed cells must be switching from one differentiated cell type to another, according to what location they were in. Later, Wilson adopted an even more extreme theory, that the new sponges were formed entirely by "archeocytes" that had previously not been differentiated at all! Some textbooks state this archeocyte theory as fact. Actually, living sponges undergo constant rearrangement of all their cells. This was also discovered here at UNC, by Cal Bond a former graduate student who was a TA in this course, and is now Biology Dept Chairman at Greensboro College. He and I also discovered that sponges crawl slowly, but the textbooks continue to say they are sessile. Johannes Holtfreter German, but anti-nazi, and then a refugee in Canada and then a Professor in the University of Rochester, New York. Dissociated amphibian cells of embryos of different species that have somewhat different colors. The colors of the cells allowed him to distinguish cells, and to find out whether they were switching from one cell type to another. Sort out by germ layer, and by germ layer subdivision. Mesoderm will sort out from ectoderm. Neural tube ectoderm will sort out from somatic (skin) ectoderm. Wrote a long and very stimulating paper with an MD-PhD student named Townes J. Experimental Zoology Vol 128 pages 53-120 Invented the concept of tissue affinity Gewebaffitat, in German Most readers took this to mean selective adhesiveness But Holtfreter also had ideas about negative affinity, and included ANY and all cell properties that would either get like cells together, or cause unlike cells to move apart. And he thought that gastrulation, neurulation etc. are caused by changes in Gewebaffitat. Because they have developed different Gewebaffitat properties, therefore cells rearrange, and we call these rearrangements gastrulation, neurulation etc. What do you think? Proof that cells rearrange, instead of re-differentiate; 1950s & 60s Aaron Moscona Studied sorting out of chicken heart cells from mouse liver cells, etc. using morphological differences between chicken and mouse nuclei as his marker for cell origin. John P. Trinkaus Labeled cells' DNA with tritiated thymidine, as a marker, and used this to prove that cells actually sort out by differentiated cell type, and don't switch from one cell type to another. Made time lapse films of pigmented retinal epithelial cells of embryonic eyes, sorting out from heart cells, liver cells etc. Because you could see the cells sorting out in these films, this also helped convince people that sorting really occurs . But sorting-out by cell types that never normally come into contact with each other persuaded Prof. Trinkaus that sorting-out can't be caused by normal embryological mechanisms. What do you think? Malcolm Steinberg (now at Princeton) invented a different kind of theory to explain cell sorting. He proposed that differentiated cells differ in the amount of adhesiveness to each other. Instead of liver and heart (for example) having different kinds of cell-cell adhesion proteins he hypothesized that they have (very) different amounts of the same adhesion molecules. He, Phillips and others proposed that thermodynamics will then cause cells to maximize their total adhesion by the cell type with the stronger adhesions bunching together into clumps surrounded by the cell type will the weaker adhesions. This is called the Differential Adhesion Hypothesis, or sometimes called the "thermodynamic hypothesis". Our textbook seems to believe in this theory, to judge from page 74. Steinberg and Phillips believed that they could measure "the work of adhesion" by measuring how strongly balls of cells resist flattening. Later, Edelman, Takeichi and others discovered specific cell-cell adhesion proteins, which have been named N-CAM, E-cadherin, P-cadherin etc. Nearly everyone now believes that cell-sorting is just a side-effect of differentiated cells having different kinds and combinations of cadherins. H. V. Wilson and Johannes Holtfreter are honored for guiding researchers to the discovery of cadherins. Steinberg collaborated with Takeichi in experiments in which different numbers of copies of genes for the same cadherin were transformed into cancerous cells, which caused them to sort out, with the cells having more gene copies going to the interior The person who has been most critical of Steinberg's theories happens to be me. J. Theoretical Biology, Vol 61, pages 267-285 (1976) My claim is that cells bunch together with others of the same differentiated cell type because they have different kinds of adhesion molecules,... and that their relative position (inside vs. outside) is caused by quantitative differences (differences in amount) of strengths of active contractions of cell surfaces, and... that Steinberg and Phillips were really measuring strengths of cell surface contractility, not amounts of cell-cell adhesion. (the "surface tensions" listed in fig 3.27 of the textbook) Notice the more general question: "What force(s) make(s) aggregations of cells tend to become spherical?"
Is it their maximization of cell-cell adhesions?
What do you think? Are you convinced by the Takiechi & Steinberg experiments in sorting out by cells with different numbers of genes for the same cadherin?
|