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Home , Abraham Trembley, Lazzaro Spallanzani, Regeneration (biology)

DEVELOPMENTAL DYNAMICS 238:2357–2363, 2009

REVIEWS–A PEER REVIEWED FORUM

Regeneration According to Spallanzani

Panagiotis A. Tsonis* and Timothy P. Fox

In this report, we elaborate on a letter that Spallanzani wrote to Bonnet reporting his findings on in worms, snails, tadpoles, and . The letter (original in French and translated in English; see Supplementary Material, which is available online) was written to discuss whether or not regeneration in these supports Bonnet’s theory on germs. The letter includes several drawings by Spallanzani, which were not published in the Prodromo, his book on Reproduction. Spallanzani made important observations, which he described with considerable detail, but overall he was unable to confidently support Bonnet’s theory. This letter reflects the way of thinking in the 18th century that shaped the important scientific fields of regeneration and reproduction. Developmental Dynamics 238:2357–2363, 2009. © 2009 Wiley-Liss, Inc.

Key words: Spallanzani; regeneration; animal reproduction

Accepted 4 June 2009

INTRODUCTION of Creation. On the contrary, epigene- 1740 and on worm regeneration in sis allowed space for questioning the 1744 (Savioz, 1948; Dinsmore, 1991) Rooted in Aristotelian philosophy, the role of God. As expected when the first shocked the scientific world. Re´aumur belief that lower animals were gener- experiments in the 18th century re- and Bonnet were preformationists ated spontaneously from decay pre- vealed the regenerative power of ani- and, in fact, Re´aumur believed that vailed until the 17th century when mals, these two competing theories germs were contained within parts re- Redi in 1668 carried out well-con- trolled experiments that provided the were called upon to explain this new sponsible for regeneration. In his writ- property of animals. ings, Bonnet argued that Trembley’s first proof against it (Redi, 1668, th 1671). A similar view, epigenesis, de- The 18 century could very well be experiments with and his with veloped by Harvey, stated that new considered as the golden era in regen- worms supported the pre-existence of appear from undifferenti- eration research. Many studies during germs and became a leading propo- ated matter. These ideas were sup- the previous century had led the way nent of the preformation theory. ported later in the 18th century by to understand how reproduction oc- Among these intellectual giants, Needhan and Buffon who believed curs and many theories and thoughts is also credited that organic molecules organized by were developed to explain how ani- as being one of the pioneers in regen- internal molds were responsible for mals (and humans) reproduce (Cobb, eration research. In 1768, Spallanzani animal generation (Benson, 1991). 2006). Naturally, experi- published the Prodromo, his historical The rival position to epigenesis, which mented with regeneration in animals book on Animal Reproduction. At that prevailed at the end of the 17th cen- (which then was called actually repro- time, the word “reproduction” was tury, was preformation. According to duction). Pivotal discoveries by Re´au- used to denote “regeneration” as well. this, the organisms were preformed in mur on appendage regeneration in in- In his book, Spallanzani described the . Preformation was also in sects in 1712 (Wheeler, 1926), by several types of regeneration with favor when it came to religious beliefs Tremblay on Hydra regeneration in mention of regeneration of the frog because it fit the notion that all gen- 1744 (Baker, 1952; Dawson, 1987), tail and limbs. The ac- erations were established at the time and by Bonnet on parthenogenesis in counts were surprisingly quite short

Additional Supporting Information may be found in the online version of this article. Department of and Center for Tissue Regeneration and Engineering at Dayton, University of Dayton, Dayton, Ohio *Correspondence to: Panagiotis A. Tsonis, Department of Biology and Center for Tissue Regeneration and Engineering at Dayton, University of Dayton, Dayton, OH 45469-2320. E-mail: [email protected] DOI 10.1002/dvdy.22057 Published online 3 August 2009 in Wiley InterScience (www.interscience.wiley.com).

© 2009 Wiley-Liss, Inc. 2358 TSONIS AND FOX

Figs. 1–6. Anatomy and regeneration in worms. Fig. 1: The gross artery (red) attached to the intestines. Fig. 2: The artery branches into five vessels (red) as it reaches the head. Spallanzani describes the vessels as small bags composed of one or many constrictions. Fig. 3: The vein that runs from head to tail is winding. It is intersected by another vein, which is straightened. Fig. 4: The “boat” worm, a fresh water worm, whose tail is on the surface of the water (light blue), while the head is hidden in the mud (dark blue). Fig. 5: Regeneration in fresh water worm. A cone is produced. The reddish color at the base of the cone is the anus. Fig. 6: At the posterior part of the anus, a red line appears in the same direction as the artery of the animal (o). Spallanzani describes that at this stage he could not see circulation in the red line but later blood flows into the artery. with not much detail or any drawings sponded on September 14 and was nistic view of animal biology or how the that were common in publications of quite happy that Spallanzani shared issue of soul fits in a regenerating ani- that time. However, in a long letter to his views and that he was also work- mal). In addition, this letter also reflects Bonnet written on September 21, ing on earthworm regeneration. Bon- the clarity of thought that Spallanzani 1766, Spallanzani outlined in great net in a sense invited Spallanzani to possessed in order to explain his find- detail regeneration in many organ- pursue the work on earthworm regen- ings. We, therefore, strongly believe isms, including earthworm, snail, eration. An exchange of letters ensued that the contents of this letter should be frog, and salamander (Biagi, 1958; in which Spallanzani outlined a series of great historical value to scholars in Dinsmore, 1991). The letter, which of experiments that were inspired by and regenera- reads as a research paper, is also dec- Bonnet’s influence. Bonnet mentioned tion research. Here we will highlight his orated with many drawings by Spal- that regeneration research should be major experiments and points. Also, we lanzani’s hand (original in French and able to resolve the germ concept, thus have reproduced the drawings and we translated in English can be found in inviting Spallanzani to elaborate on present them with added color as well. Supplementary Material). One of the this (Biagi, 1958; Dinsmore, 1991). The color is in accordance with the de- major reasons Spallanzani wrote this In his letter of September 21, Spal- scription that Spallanzani provides in letter was to discuss Bonnet’s theory lanzani casts doubts on the existence of on germs, according to which repro- germs for some types of regeneration his letter. Figures 1–17 have been set in duction and regeneration were medi- and obviously he does this with great the order in which they appear in the ated by germs that existed in the or- care not to offend Bonnet. Thus, this letter, but have been grouped according ganism. Spallanzani initiated letter bears enormous significance to the . We will end this review correspondence with Bonnet on July about how scientists were thinking at by reflecting on how these ideas shaped th 18, 1765 by sending him copies of two that time and how they dealt with these scientific thought in the 18 century of his publications. On August 24, obstacles imposed by their social/reli- and how they relate to our thoughts in 1765, Spallanzani wrote again to Bon- gious status as well as by the prevailing the present day. Text in italics contains net critically commenting on Need- way of thinking of the time (for exam- comments by us related to Spallanza- ham and Buffon’s ideas. Bonnet re- ple, it questioned the Cartesian mecha- ni’s observations. REGENERATION ACCORDING TO SPALLANZANI 2359

REGENERATION IN ends of the head are dead but all the more or less equal parts. Almost all EARTHWORMS AND posterior parts containing the ovary the anterior parts regenerated (obvi- FRESHWATER WORMS and the small bags generated a head, ously these parts contained the repro- which in the end was equal to the orig- ductive system) while most of the pos- After a short salutation and com- inal one. terior parts (36/50) died. In the second ments on how some scientists view experiment, he cut the worm into three parts in a way that the anterior Bonnet’s research, Spallanzani sets Experiment 4 the stage by describing certain exper- part contained 3–6 rings and no re- iments he performed to learn about Divide longitudinally the body into productive system, and the middle the anatomy and physiology of the two parts from the head to two thirds part contained the reproductive sys- earthworm. Spallanzani provides of its length and then remove the pos- tem and the posterior part. At the end three drawings of the anatomy of ar- terior third (tail). Results: The two of the experiment, all the anterior teries and veins (Figs. 1–3). He then longitudinally divided parts and the parts were dead, the middle parts re- proceeds by outlining six different ex- tail part died. generated and many of the posterior periments to study regeneration in parts were alive but without signs of regeneration. Spallanzani also men- earthworm. Drawings outlining these Experiment 5 experiments and the results are tions that at the level of the tail, the shown in Supplementary Material. Divide the worm longitudinally into “boat” structure never failed to form, two parts from the tail to two thirds of and he also concludes that these Experiment 1 its length and then remove the ante- worms regenerate naturally because Transversal section of the earthworm rior part containing the head. Results: artificial regeneration (when he cut into three parts in a way that the The two parts, which were separated, the worms) was identical with natural ovary, and the small bags (he means died, and the third part, which re- regeneration (when he found them in vessels; Fig. 2) remain in the anterior mained and contained the head, gener- nature). part, or in the part where the head is ated the tail (obviously the part con- Having established that worms can located. Results: After a few months, tained part of or the whole reproductive regenerate, Spallanzani then at- the posterior parts, or tails, and the system). tempts to explain the mechanism. middle parts all died, with the excep- Most of his observations are through the circulatory system, because this tion of 5 or 6 of them. The anterior Experiment 6 part, which contained the ovary and can be easily observed. First, he iden- the small bags, after 16 to 20 days Divide longitudinally the whole worm tifies a red dot by the edges of the started to generate a very small bud; from the head to tail. Results: All the anus. This shape of the anus stretches this small bud stopped growing during worms died. out. A cone starts to form, but the winter, but continued its development Spallanzani then proceeds to dis- anus he claims is not at the tip of the during spring. cuss how earthworms grow. The ques- cone but more at the rear of the cone tion is: Is growth mediated by the ad- (Fig. 5). At the posterior end of the dition of new rings or by the expansion animal (stump), a red line appears in Experiment 2 of old ones? This question is related to the same direction as the artery of the Divide the worm transversally into 3 preformation notions. In order to an- animal (denoted as O artery in Fig. 6). parts in a way that the ovary and the swer this question, Spallanzani de- In the beginning, the line does not small bags belong to the middle part. cided to count rings in 11 worms that show any circulation but later it does, Results: All anterior parts or the ends hatched from eggs and in 11 fully and the blood flows into the O artery. of the head died. The tails (posterior grown ones. He observed 84, 100, 96, Thus, the red line is the regenerated parts) also died with the exception of 90, 102, 104, 88, 93, 96, 104, 90 in the artery. The cone grows and the edges three of them. Many of the middle former and 88, 94, 73, 101, 87, 83, 96, of the anus acquire more redness. As parts generated a head at their ante- 80, 89, 95, 101 in the latter. Based on the cone grows, rings become obvious. rior extremity, and a tail at their pos- this, he concluded that growth is the Spallanzani also attempted repeated terior extremity. result of expansion of old rings. amputations, which were successful. Spallanzani attempts to provide Spallanzani then turns to describe He finally concludes that the shape some explanation of the few survivors regeneration in another species, the (boat) is always formed and that the that did not contain the reproductive fresh water worm. As for its appear- repeated amputations lead to shorten- system in experiments 1 and 2. Obvi- ance, he states that the part that con- ing of the animal. Spallanzani does ously, the pieces survived for several tains the head is hidden in the mud, not elaborate here how germs could months and regeneration of the tail while the tail is pushed at the surface account for regeneration in earth- was more obvious. of the water forming a structure that worms, but he compares this later with resembles a groove or a boat (Fig. 4). head regeneration in snails. At the end He also describes the anatomy of the of this section, Spallanzani develops Experiment 3 circulatory system for this worm. As some ideas about the respiratory sys- Remove from the worm the part of the with earthworms, Spallanzani experi- tem, but his observations are rather head that remains above the ovary mented with regeneration. In one ex- inconclusive. Despite the interesting and the small bags. Results: All the periment, he cut 50 worms into two way that earthworms regenerate and 2360 TSONIS AND FOX

Figs. 7–9. Regeneration in snails. Fig. 7: Head regeneration. A bud with two lips and two small horns. Spallanzani claims that at this stage the long horns are not visible. Figs. 8-9: Tail regeneration. After amputation (8) the stump retracts and in the middle (o) a whitish beak appears in the center of the musculature. the intriguing necessity of the repro- horn, the rest of the horn gradually emerged in parts, which appeared one ductive system, which implies depen- becomes rounded into a small bud, after another and reconstituted the dence on some specific factor, not much which grows and its color darkens. Fi- head with time. Spallanzani describes is known today at the molecular level. nally, a dark spot appears at the tip of one case where regeneration consisted Nevertheless, after Spallanzani, many the bud. The bud grows, becomes of a bud made of two lips of the mouth scientists experimented with earth- longer, and after some time the muti- and two small horns. The bud was worm regeneration. The necessity of lated horn acquires its normal length. “implanted” in the piece (the stump) certain segments, which include the re- Spallanzani notes that at the bound- (Fig. 7). The long horns were the next productive system, has been verified ary between the old and the regener- part that was regenerated. Spallan- (Morgan, 1901). In the earthworm, ated piece, there is a whitish color. zani makes the case that not all parts Eisenia foetida, which has 100 seg- When he cut both horns, both regen- are regenerating during the same ments, amputation between segments erated as well. Next, Spallanzani dis- time frame. He then proceeded to dis- 20–35 results in the failure of regener- cusses the nature of the dark spots at sect the regenerated head and fol- ation (Supp. Fig. 1). In this worm, re- the tip of the long horns. He disagrees lowed the direction of the four nerves generation is mediated by the forma- with other authors that they are eyes. that are inserted in the horns. He tion of , and and Rather, he believes that they are like claims that he could not find where endoderm maintain their identities. It antennas, which direct the snail dur- the old part united with the new one, also seems that in general ing motion. Obviously Spallanzani and this was the case for the skin rely on dedifferentiation of older tis- was wrong about this because it is now (outer surface of the horns) and esoph- sues, rather than pluripotent reserve well known that the spots are eyes and agus. With these observations and the cells (Goss, 1969). Nevertheless, stud- their regeneration has been well docu- comparison he mentions with the ies on in anne- mented (Eakin and Ferlatte, 1973). earthworm, Spallanzani sets the stage lids have indicated the regeneration of But most amazingly, Spallanzani ex- for discussing Bonnet’s theory on primordial germ cells from parts of the perimented with regeneration of the germs. body devoid of gonads, arising from snail’s head! The severed pieces con- Next, Spallanzani describes regen- piwi-expressing germile stem cells tained two lips, mouth, part of the eration of the tail. He noticed that af- (Weisblat, 2006). esophagus, a tooth, muscles, and four ter amputation, the piece left behind horns (two long and two small ones). retracts (Figs. 8 and 9). In the middle REGENERATION IN He observed regeneration in many of of the tissue (denoted as O in Fig. 9), them (he used 200) after 26 days. He there is formation of a “beak,” its color GARDEN SNAILS (SLUGS) then attempted to compare these re- being white. That was in the center of The next part of the letter deals with sults with regeneration in earth- the tail’s musculature and it contin- regeneration of head structures in worms. He attests that regeneration ued to grow in all directions until it snails. He first describes regeneration of the head from a sectioned posterior became equal to the removed part. of the long horns (stalks), which “ bear part of the earthworm is mediated by The regenerated part maintained its at their extremity a nice bud adorned a cone, which is a whole organ, a min- white color and it was also recognized by a blackhead.” iature of a head waiting to fully de- for having a non-stressed gross skin Following partial sectioning of a velop. However, in the snail the head that is seen in the remainder of the REGENERATION ACCORDING TO SPALLANZANI 2361

Figs. 10–14. Tail regeneration in salamanders. Fig. 10: Beginning of regeneration with the thin crest of tissue denoted as rst. Fig. 11: Growth of the blastema (reproduction). Fig. 12: Appearance of the spinal cord and notochord (Spallanzani refers to them as bands). Fig. 13: Regeneration after the whole tail was cut. Two small mounts of flesh (ab and bc) were elevated on each side of the spine (b). Fig. 14: Progression of regeneration from the case shown in Figure 13. Growth only occurred at the b side to form an elevation. tail (maybe he is observing here a kind are so perfectly united in the repro- surface). He then goes on to describe the of wound , which has not duction (regenerated part), so that the beginning of a formation from the thin become full-thickness skin). Even parts are in continuity with the ones surface, which could be round or elliptic when the amputation was on an in the old animal. He discusses that he (what is known now as blastema). He oblique surface, it was completed. Ex- has difficulty seeing how germs can observed that along the longer axis of periments with other snails, which he account for this. Likewise, he con- the elliptic surface appears a thin crest calls Limaconi Ignudi, were not as cludes the same for snail head regen- of flesh, which is the beginning of the good in that they could regenerate the eration because, as mentioned above, reproduction (regenerating tail). For horns but not their heads. the parts regenerated separately and anyone who has studied tail or limb re- Spallanzani then embarks on ex- the regenerated nerves looked like ex- generation, this is the early cone forma- plaining regeneration based on Bon- tensions of the old ones. In order to tion, which is most obvious at the center net’s germ theory. As briefly men- gain more insight and information on of the stump. Spallanzani in fact notices tioned above for earthworm and fresh this, Spallanzani turns next to his ex- the different colors (a thin yellow line) water worms, Spallanzani is unable to periments on salamander and tadpole between the old tail and the regenerat- explain it as “a simple extension”; tail regeneration. ing one (Fig. 10). He then describes how therefore, he strongly believes that that central crest of flesh grows in all pre-existing germs must be involved. TAIL REGENERATION IN directions (clearly the formation of He based this on his observation that blastema) (Fig. 11). From the center of SALAMANDERS the cone at the site of amputation re- the regenerate, two small bands appear sembles the part that was removed. First, Spallanzani describes tail regen- and they go up to the tip of the crest. Even though Spallanzani tries to eration in adult salamanders. After am- These bands are brownish and the re- please Bonnet by supporting his the- putation, he observed a lot of blood flow mainder of the regenerating tail is ory, he cannot be quite sure that and retraction of the skin at the edges of white (Fig. 12). In fact, Spallanzani germs are responsible for the whole the stump. Then he observed that a notes that these two bands belong to regeneration: “let me have a small kind of thin surface of reddish white two different planes in the regenerate. doubt to your sound judgment.”. color appear and the edges of the skin The bands that Spallanzani refers to are The doubt is cast from his observa- seem implanted on this surface. Spal- the regenerating spinal cord and noto- tions that the gross artery, the two lanzani, in fact, describes here the for- chord. After 3 months, the color has veins, intestines, and the musculature mation of a wound epithelium (the thin changed to become darker. Spallanzani 2362 TSONIS AND FOX

Figs. 15–17. Tail regeneration in tadpoles. Fig. 15: Anatomy of the tail. Spallanzani proposes that the tail is made of two parts. An elevated “tongue” (light brown; omu) and two lateral membranes (vom and umn). Spallanzani denotes the spine as a kind of marrow (dark brown; cm) and ae “small parallel ribs,” obviously referring to the musculature. Fig. 16: The circulatory system in intact and regenerated tail. In intact tail, it forms a big red thread. In the regenerated part, the red thread is not present. Fig. 17: The regenerated part, consisting of skin muscles and spinal cord (lighter colors), is described as a continuation of the old (intact) part. notes here that when he dissected the parts, a major pointed one, which he each side toward the extremity of the regenerating tail, it was very soft and calls “tongue,” and two lateral mem- tail, and then they turn upward, still he could not tell that the spine was branes (transparent tissue) (Fig. 15). being divided from each other. The di- made up of small bones. For him, this is In Figure 15, the tongue (light brown vision remains until they reach back an important observation because since color) is demarcated by o, m, u, while at the amputation plane where they he cannot see the small bones and the the two membranes (yellowish-white all unite into a vein. Older tadpoles spine develops with time and is quite color) are denoted by v, o, m and u, m, also regenerated the tail but Spallan- similar to the spine of the old tail, he n. Inside the tongue is a kind of mar- zani was unable to observe circula- concludes that simple extension cannot row (c, m; dark brown color) that runs tion. The same conclusions were made explain regeneration. Thus, this exper- across. This must be the cartilaginous for the continuity between the old and iment would rather support develop- spine because he even points out that it new filaments. Also the skin that en- ment of germs. He then describes exper- is not bony but is made up of a sub- velops the muscles of the regenerating iments where the whole tail was cut. He stance more consistent than the mus- tail was continuous with the old part, explains that for 26 days he saw no sign cle. Because the tongue is still opaque, but had a different color (Fig. 17). He of regeneration. Eventually, the thin he cannot see circulation in it, but he briefly mentions that regeneration of surface appeared. Then at each side of can observe many small vessels run- limbs in old tadpoles was never seen the spine two mounts of flesh appeared ning from the tongue to the lateral (obviously, he amputated them after a (ab and bc), which covered the spine edges of the membranes and back. He particular stage in (Fig. 13). There was some growth at the then goes on to describe the circula- which regeneration is not permitted). junction of these two mounts (b in Fig. tory organization in the tail and he The differences in regeneration 13) and not much more (Fig. 14). Spal- points out that there are two main (morphology) of the vessels allow lanzani correctly concluded that if too vessels. Here he disagrees with Leeu- much of the tail is removed, regenera- wenhoek who claimed to have discov- Spallanzani to declare that tail regen- tion is greatly retarded or inhibited. He ered four vessels. After tail amputa- eration in tadpoles originates from a then reports that in young salamanders tion, he describes two main events. lengthening (extension) of the old (one month old), regeneration follows One is the regeneration of the vessels parts and not from development of the same steps but is much faster. He and the other accumulation of “fila- germs. He also thinks that the way also suggests that some bony small ments” at the edges of the tongue. He that the membranes regenerated grains seen along the small band make points out that there are very remark- gives more ground to his conclusion. the spine. able differences between circulation in Spallanzani then tries very tactfully the natural part and the regenerated to elicit Bonnet’s opinion based on his results: “On which side is your incli- REGENERATION OF one (Fig. 16). In the former, the artery and vein make one vessel and run par- nation Dear Sir? In your very kind TADPOLES’ TAIL allel to the length of the tail, but in the letter (the one Bonnet had sent to Spallanzani begins by describing the latter when the artery enters the re- Spallanzani on August 8) you are gross macroscopic anatomy of the tad- generating part, it divides into many nicely asking me if my observations pole’s tail. He divides the tail in two branches, which go on winding on confirm your principles. Now you can REGENERATION ACCORDING TO SPALLANZANI 2363 see with your own eyes and judge a tion in the regenerating limb is differ- wise, if he knew all these marvelous thousand times better than I can.” ent than the original and that some- examples of reserved stem cells or Spallanzani then states that Bonnet times there would be cases with more even the powerful potential of embry- assigns two sources of repair. One or less bones. Even though he does not onic stem cells that can in principle that proceeds through the develop- speculate, that observation also goes build an , he might have ment of small filaments and explains against germ theory. written to Bonnet with joy that the regeneration of animal skin or plant germ theory is valid! peel and a second that involves the CONCLUSIONS development of germs to explain re- generation of a whole organ or body It is obvious from studying this letter REFERENCES parts. In a sense, Spallanzani at- that Spallanzani based his conclu- sions on macroscopic observations Anderson L. 1982. and the tempts to tell Bonnet that germs can- order of the known. Dordrecht, The not account for regeneration in snails made with different criteria. For ex- Netherlands: D. Reidel Publishing Com- and the tadpole’s tail. Spallanzani ample, several conclusions are based pany. ends his letter confessing that to be on the circulatory system, which could Baker JR. 1952. . Sci- certain he needs to do more experi- be hindered by the transparency of the entist and philosopher. London: Edward tissue. To scientists in our time, it is Arnold and Co. ments, especially to identify if he Benson KR. 1991. Observation versus phil- could observe the union between the easy to see where Spallanzani failed osophical commitment in the eighteenth- germ (which he now calls an animal or succeeded in supporting Bonnet’s century ideas of regeneration and gener- graft) and the stump. Spallanzani germ theory. Going against the prefor- ation. In: Dinsmore CE, editor. History speculates that for circulation to be mation theory was a real moral prob- of regeneration research. New York: lem for these scientists, because it had Cambridge University Press. explained by germs (the regenerated Biagi B. 1958. Lazzaro Spallanzani: Epis- part uniting with the old part, which dominated their scientific beliefs and tolario. Vol 1. Florence: Sansoni Anti- results in common circulation), then a was rooted in their social status. The quariato. fossil of the graft should be found as in reader has only to consider that even Cobb M. 2006. Generation. New York: the case of the plant shunt. It is our today, when it comes to regenerative Bloomsbury. impression that Spallanzani tries to medicine and stem cells, opinions vary Dawson VP. 1987. Nature’s enigma. The based on social status and religion. problem of the in the letters of leave room to support Bonnet’s theory. Bonnet, Trembley and Re´aumur. Phila- Bonnet was a who had Notwithstanding this great problem, delphia: American Philosophical Society. learned not to succumb to dogmatism Spallanzani and the other regenera- Del Rio-Tsonis K, Tsonis PA. 2003. Eye and thus he knew how to accept criti- tion pioneers made outstanding con- regeneration at the molecular age. Dev cism. On January 17, 1771 he wrote to tributions to science and many of their Dyn 226:211–224. observations were correct. Theories Dinsmore CH, editor. 1991. A history of Spallanzani: “I have always sincerely regeneration research. New York: Cam- sought the truth and I have warned a always are there to explain results. bridge University Press. hundred times that I never flatter my- Even today, the mechanisms of regen- Eakin RM, Ferlatte MM. 1973. Studies on self with the thought that I have al- eration are still debated. Is there true eye regeneration in a snail, Helix as- ways found it. So don’t spare me my transdifferentiation or is regeneration persa. J Exp Zool 184:81–96. mediated by undifferentiated stem Goss RJ. 1969. Principles of regeneration. errors, and criticize me whenever you New York: Academic Press. cells? The has again been proven judge it necessary. When nature pro- Morgan TH. 1901. Regeneration. London: nounces against me you must not in- an indispensable tool. Studies con- Macmillan Company. th dulge in the language of friendship; ducted from the 19 century on have Redi F. 1668. Esperienze intorno alla ge- and I will be the first to submit to its shown that dedifferentiation of so- nerazione degl’insetti (Florence, Stella) decisions.” (quoted in Savioz, 1948, matic cells accounts for tissue repair and the version in Latin: De generatonis during limb regeneration (Morgan, insectorium, Amsterdam (1671). and translated by Anderson, 1982). Savioz R. 1948. The philosophy of Charles 1901). Thus, muscle, nervous tissue, Bonnet of . Paris: Libraire Philos- and bone dedifferentiate to give rise to THE PRODROMO ophique J. Vrin (trans. from French). the regenerated muscle, nerves, and Spallanzani L. 1768. Prodromo di un opera Two years later, in 1768, Spallanzani bone (Tsonis, 1996). During lens re- da imprimersi sopra la riproduzioni ana- published the Prodromo, his book on generation, the iris pigment epithelial mali. Giovanni Montanari, Modena. Translated in English by Maty M. 1769. Animal reproduction (Regeneration). cells from the dorsal iris form a con- An essay on animal reproduction. Lon- Surprisingly, the book was quite short tinuous vesicle that then transdiffer- don: T. Becket & DeHondt. with not much detail or illustrations entiates to lens (Del Rio-Tsonis and Tsonis PA. 1996. Limb regeneration. Cam- as he had suggested he would do. Nev- Tsonis, 2003). If Spallanzani had seen bridge: Cambridge University Press. ertheless, it is considered to be a sem- histological preparations from these Weisblat DA. 2006. regenera- inal publication for regeneration re- events, he would have concluded that tion: the worms’ turn. Curr Biol 16: R453–R455. search. In the book, Spallanzani also regeneration is most likely explained Wheeler WM. 1926. Translation of Re´au- describes regeneration of the limbs in through simple extension and he mur’s The of Ants. New salamanders. He noted that circula- would have discounted germs. Like- York: Alfred A. Knopf.