Hayflick, His Limit, and Cellular Ageing Clearly a Next Step Is Automation

PERSPECTIVES Finally, the most apparent drawback is the Whether or not nucleic acid computers 175–179 (2000). 3. Faulhammer, D., Cukras, A., Lipton, R. J. & Landweber, L. time required for each computation. ultimately prove feasible, they have already F. Molecular computation: RNA solutions to chess Whereas a simple desktop computer can contributed to multi-disciplinary science by problems. Proc. Natl Acad. Sci. USA 97, 1385–1389 (2000). solve the seven-city instance of the Travelling causing us to question the nature of comput- 4. Ouyang, Q., Kaplan, P. D., Liu, S. & Libchaber, A. DNA Salesman Problem in less than a second, ing and to forge new links between the biolog- solution of the maximal clique problem. Science 278, 1 446–449 (1997). Adleman took seven days . The use of DNA ical and computational sciences. For example, 5. Henegariu, O., Heerema, N. A., Dlouhy, S. R., Vance, G. H. chips2 or other approaches may eventually it has led us to focus on the nature of biologi- & Vogt, P. H. Multiplex PCR: Critical parameters and step- by-step protocol. Biotechniques 23, 504–511 (1997). lead to automation, which would save con- cal DNA computations, such as the assembly 6. Karp, G. Cell and Molecular Biology: Concepts and siderable amounts of time, but fundamental of modern genes from encrypted building- Experiments 2nd edn (John Wiley & Sons, New York, 1999). DNA computing technology needs to blocks in the genomes of some single-celled 7. Seife, C. RNA works out knight moves. Science 287, advance far beyond its current bounds before ciliates (FIG. 5)14. After all, our bodies already 1182–1183 (2000). 8. Condon, A. & Rozenberg, G. (eds) Prelim. Proc. 6th Int. it can be made practical. contain millions of complicated, efficient, Meet. DNA Based Computers (Leiden Univ., The DNA computing has its advantages, evolved molecular computers called cells. Netherlands, 2000). 9. Meller, A. et al. Rapid nanopore discrimination between though. One is its massive parallelism — that Adam J. Ruben and Laura F. Landweber are in the single polynucleotide molecules. Proc. Natl Acad. Sci. is, brute-force algorithms can search through Department of Ecology and Evolutionary Biology, USA 97, 1079–1084 (2000). 10. Sakamoto, K. et al. Molecular computation by DNA quadrillions of molecules at the same time Princeton University, Princeton, hairpin formation. Science 288, 1223–1226 (2000). and find a correct solution, akin to in vitro New Jersey 08544, USA. 11. Seeman, N. C. DNA engineering and its application to biotechnology. Trends Biotechnol. 17, 437–443 (2000). selection3. Another is miniaturization. And e-mail: [email protected] Links 12. Winfree, E. et al. in DNA Based Computers II. DIMACS once the procedures are under control, the Series in Discrete Mathematics and Theoretical Computer Science Vol. 44 (eds Landweber, L. F. & Baum, raw materials cost less too.“Here’s nature’s FURTHER INFORMATION DNA computing: a E. B.) (American Mathematical Soc., Providence, Rhode toolbox,”commented Adleman7,“a bunch of primer | Laura Landweber’s homepage Island, 1999). 13. Winfree, E., Liu, F., Wenzler, L. A. & Seeman, N. C. little tools that are dirt cheap; you can buy a Design and self-assembly of two-dimensional DNA DNA strand for 100 femtocents.” crystals. Nature 394, 539–544 (1998). 1. Adleman, L. Molecular computation of solutions to 14. Landweber, L. F., Kuo, T.-C. & Curtis, E. A. Evolution and combinatorial problems. Science 266, 1021–1023 (1994). assembly of an extremely scrambled gene. Proc. Natl The near future 2. Liu, Q. et al. DNA computing on surfaces. Nature 403, Acad. Sci. USA 97, 3298–3303 (2000). Now is an exciting time in the field of DNA computing, as there is so much that has not been tried. In June, over 120 molecular biolo- gists, computer scientists, mathematicians TIMELINE and chemists from around the world gath- ered in Leiden8 to discuss the latest in DNA computing technology. Hayflick, his limit, and cellular ageing Clearly a next step is automation. McCaskill and colleagues in Germany have constructed a ‘microflow reactor’ on which Jerry W. Shay and Woodring E. Wright they propose to solve a 20-bit satisfiability problem in an hour and a half 8. One could also construct a microfluidic device consist- Almost 40 years ago, Leonard Hayflick in culture are immortal, and that the lack of ing of gated channels so small that only one discovered that cultured normal human cells continuous cell replication was due to igno- molecule can pass through at a time9, vastly have limited capacity to divide, after which rance on how best to cultivate the cells. improving readout8. And a team led by they become senescent — a phenomenon Carrel’s view was based on his and Albert Adleman recently solved a 6-variable, 11- now known as the ‘Hayflick limit’. Hayflick’s Ebeling’s work, done at the Rockefeller clause satisfiability problem using a ‘dry’ com- findings were strongly challenged at the Institute in New York City, in which they puter consisting of thin, gel-filled glass tubes8. time, and continue to be questioned in a few claimed that chick heart fibroblasts grew con- As for DNA chips, their future in DNA circles, but his achievements have enabled computing looks bright as well, because ‘uni- others to make considerable progress versal’ DNA chips could contain every possi- towards understanding and manipulating the ble DNA sequence of a given length (probably molecular mechanisms of ageing. about 8–12 base pairs). Hagiya and colleagues in Tokyo are finding creative uses for single- To set Hayflick’s discoveries in context, we stranded DNA molecules that fold into intra- need to go back to 1881 (TIMELINE, overleaf), strand ‘hairpins’8,10. Winfree, Seeman and col- when the German biologist August leagues — responsible for construction of Weismann1 speculated that “death takes place beautiful assemblies with DNA, such as a because a worn-out tissue cannot forever DNA nano-cube11 — have proposed the renew itself, and because a capacity for assembly of even more ordered structures that increase by means of cell division is not ever- show patterned algorithmic supramolecular lasting but finite”.This concept, which was self-assembly8,11–13. Even a handful of mathe- almost entirely forgotten by the time Hayflick maticians have lent a hand, proposing faster began his work, was later challenged by the and more efficient algorithms tailored to the French Nobel-prize-winning surgeon Alexis Figure 1 | Leonard Hayflick in 1988. needs of DNA computing8. Carrel, who suggested that all cells explanted (Photograph: Peter Argentine.) 72 | OCTOBER 2000 | VOLUME 1 www.nature.com/reviews/molcellbio PERSPECTIVES tinuously for 34 years2. This led to the general normal cells3,4. The experiment with mixed idea that all vertebrate cells could divide indef- “The largest fact to have cells further assured Hayflick and Moorhead initely in cell culture. However, Carrel’s origi- that culture artefacts could not explain their nal observations could not be reproduced by come from tissue culture in observations. They submitted a paper other scientists3,4, and may have been due to the last fifty years is that describing their findings to the Journal of an experimental error4. The cells were fed with Experimental Medicine but Peyton Rous, one a daily extract of chick embryo tissue extract- cells inherently capable of of the journal’s editors, was not easily per- ed under conditions that permitted the addi- multiplying will do so suaded. After the paper had been peer- tion of fresh living cells to the culture at each indefinitely if supplied with reviewed, Rous included the following state- feeding3. It has been suggested that Carrel ment in his covering letter:“The largest fact to knew about this error but never admitted it5,6, the right milieu in vitro.” have come from tissue culture in the last fifty but even if this explanation is untrue, no one years is that cells inherently capable of multi- has ever confirmed Carrel’s work. all, it had been 60 years since Ross Harrison plying will do so indefinitely if supplied with The Carrel experiments were of great had started the field of cell culture, and nor- the right milieu in vitro.”The article was not importance because, if valid, they meant that mal cultured cells were thought to be immor- accepted. Fortunately, the editors of normal cells freed from in vivo control mecha- tal. For Hayflick to propose that a cell-division Experimental Cell Research, where the paper nisms could function normally and, apparent- counting mechanism could be involved in was published3 in 1961, were less swayed by ly, forever. However, reports were beginning to ageing was a completely new idea. But the dogma of the day. This work and subse- emerge of difficulties in long-term cell culture Hayflick was young and ambitious, and a quent studies (TIMELINE) changed the tenor of when Leonard Hayflick (FIG. 1) and Paul series of carefully conducted experiments over research, eventually leading Sir Macfarlane Moorhead entered the field. They brilliantly about three years convinced him that the fail- Burnett, Nobel laureate from Australia, to got to the heart of the matter, demonstrating ure of his normal cells to replicate indefinitely coin the phrase “the Hayflick limit” for the finite replicative capacity of normal human was not due to technical errors. first time in his book Intrinsic Mutagenesis, fibroblasts and interpreting the phenomenon In 1961, working with the talented cytoge- published8 in 1974. as ageing at the cellular level3,4. These initial neticist Paul Moorhead, Hayflick did a series observations sparked Leonard Hayflick’s pas- of experiments that challenged Carrel’s views. Hayflick’s enduring impact sion — which has lasted his entire career — to Hayflick and Moorhead showed that popula- The durability and importance of Hayflick’s overturn the central dogma that all vertebrate tions of cultured normal human fibroblasts work are reflected in its citation history.

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