As the SAZ functions in PR in a similar way in Hediste diversicolor, Phylo foetida and Syllis amica, we will focus essentially on Syllis amica and other Syllidae species, which offer the opportunity to compare PR with AR. PR and AR include 2 phases: the first phase is the regeneration of the terminal part of the worm (the pygidium for PR, the prostomium for AR); the second phase is represented by the differentiation of segments from the SAZ, which corresponds to the metamerization of the regenerate. While the first phase needs around the same time to be achieved in PR and AR (9–10 dpa for Syllis amica), the timing of the second phase is different between PR and AR. In PR, it is characterized by its rapidity and relatively high number of regenerated segments, compared to AR (e.g. 23 segments are regenerated by 60/70 dpa in PR, while only 3–4 segments by 40/45 dpa in AR). Moreover, regenerative segment formation grades into normal growth segment formation at the end of PR, while segment formation stops in AR. The first phase (regeneration of the terminal part of the worm) differs from the second phase (metamerization of the regenerate), because it does not need the presence of mesodermal cells. In PR, the presence of epidermal and intestinal cells is sufficient to build a normal pygidium, a situation similar to what we observed in Hediste diversicolor, whose pygidium also regenerates after X-irradiation while segmentation is inhibited (Boilly 1969d), as well in Phylo foetida (Boilly 1968b). However, in AR of S. amica the epidermis alone is enough to regenerate the prostomium. The pygidium and the prostomium might represent the two parts of the trochophore larva before the formation of the mesodermal primordia. Thus, this first regeneration phase might reproduce what happens during the trochophore development (at least in those species with this kind of larva). These situations clearly show that the regeneration of the pygidium, as that of the prostomium, are independent from that of segments, and consequently is not linked to the activity of a SAZ during regeneration.

The second phase (metamerization of the regenerate) needs the presence of mesodermal regeneration cells. When these cells are selectively destroyed, either with an appropriate dose of X-rays or intracoelomic injection of Thorotrast, no segment is regenerated, while epidermal and intestinal epithelia are not affected. Mesodermal cells are vital for the segmentation of the regenerate. This condition is also found during the development of the trochophore, whose metamerization starts only after the appearance of mesodermal primordia. For lower X-ray dose, or diluted Thorotrast intracoelomic injection, segment regeneration was possible thanks to viable blastema mesodermal cells not killed by the treatment. This suggests that the number of regenerating segments depends on the prevalence of cell death after X-irradiation or Thorotrast injection (Boilly 1969b, 1969c). One of the first effects of mesodermal cell loss concerns the beginning of segmentation, which is delayed in proportion with the intensity of treatment both in AR and PR. However, the following segmentation course distinguishes PR from AR.

In PR, segmentation speed decreased with increasing X-ray dose. However, this loss of regenerated segments was not counterbalanced by a significative lengthening of the segmentation phase since regeneration stopped around 60–70 dpa in Syllis amica whatever the dose used (Boilly 1969b). This result shows that the extent of regeneration (number of regenerated segments) is not linked to the number of segments missing after amputation, as it was sometimes proposed (review in Abeloos 1932). The meaning of this regeneration stop is still not understood; the SAZ is always active as the normal growth takes over from regeneration but at a lower speed. The hypothetic “regeneration signal” (Abeloos 1932) resulting from the discontinuity between the regenerated pygidium and the last segment of the stump, with a lifespan limited to 60–70 days in our model, could initiate the regeneration process which should be maintained only during this time. Nevertheless, discontinuity is not enough to allow regeneration, as growth factors from nerves are necessary to sustain cell proliferation, as demonstrated in amphibians (Stocum 2017) and cancer models.

Although the beginning of segmentation was delayed in the same manner in AR as in PR, contrary to PR, segmentation showed to be less or not responsive to X-ray dose increase. Segmentation speed did not change significantly, and for all X-ray doses used (except for doses which stop segmentation both in PR and AR), the number of regenerated segments was the same (around 4). It is possible that the low capacity of AR is linked to a limited availability of mesodermal regeneration cells. Two reasons could explain this situation: (1) the absence of digestive tube participation in AR, (2) the limited stimulatory action of nerves on these cells. Indeed, we showed in S. amica that the digestive tube does not participate in AR, contrary to PR where the presence of the digestive tube is necessary for segmentation. We consider that the absence of a digestive tube in AR could lower the mesodermal regenerative cell population, as the coelomic epithelium surrounding the digestive tube is an important source of mesodermal regenerative cells. Some observations support this hypothesis. While S. amica is unable to regenerate the digestive tube anteriorly, two other syllids (Syllis gracilis and Syllis malaquini), which regenerate easily 14 anterior segments at 59 dpa (Boilly and Thibault 1974), and 8 segments 14 dpa, respectively (Ribeiro et al. 2021) can do it. They are even able to regenerate the whole anterior part of the digestive tube derived from the stomodeum (pharynx, proventricle, ventricle and its caeca). This situation was described also in 2 other Syllidae: Procerastea halleziana (Langhammer 1928) and Epigamia alexandri, which regenerate 10 segments anteriorly together with the complete anterior digestive tube (Malaquin 1893). These species show therefore stronger regenerative abilities, contrary to many other syllids like S. amica, which do not regenerate the digestive tube anteriorly and instead regenerate only a few segments (3–4 for S. amica). In PR, the situation is quite different because, contrary to AR, no segment is regenerated in the absence of the digestive tube (Boilly 1969d), a result which suggests that the morphogenetic function of this organ takes priority over its function as a source of mesodermal regeneration cells (Boilly 1973). This could also explain why PR decreased in relation to Thorotrast poisoning of intestinal cells (Boilly 1969d), a treatment which certainly reduced the regeneration of the digestive tube.

Nevertheless, if the presence of regeneration cells close to the wound is a prerequisite for regeneration, their multiplication is a necessity to the rebuilt of lost tissues after amputation. This might involve growth factors, and especially those released from nerves as described in other regeneration models (Sinigaglia and Averof 2019). Knowing the essential role of regenerating nerve fibres on proliferation during regeneration (Boilly and Bauduin 1988; Taban et al. 1996; Zenjari et al. 1997), as well as in cancer (Guo and Gil 2022), we consider that a difference of nerve activity in PR vs AR could explain the difference of function of the corresponding SAZ. In PR, regenerated nerve fibres from the amputated nerve cord are always in contact with mesodermal cells; consequently, mesodermal cells, always maintained in activity, constantly bring new cells for regeneration and normal growth. On the other hand, in AR, nerve fibres regenerate from the severed nerve cord into the blastema, as during PR, but rapidly connect with the newformed brain of the regenerated prostomium as it was already observed in another syllid (Weidhase et al. 2017). As a consequence, the number of regenerating nerve fibres decreases in AR-SAZ, which will be relatively denervated compared to the PR-SAZ. Therefore, without regenerating nerve fibres, the proliferation of mesodermal cells decreases correlatively with the activity of AR-SAZ, and regeneration ultimately stops. This would explain why AR ends largely before PR, the SAZ of which remains always functional, first for posterior regeneration, then for normal growth. Behind these observations, the nature of the nerve factor controlling the proliferation of regeneration cells (Sinigaglia and Averof 2019), as well as cancer cells (Guo and Gil 2022), needs to be known within the scope of regeneration and cancer control research.

These results mean that PR-SAZ functions differently from AR-SAZ. However, one could object that sexual maturation/stolonization, which is normally induced in posterior segments,—those used for AR study—could disturb AR (Durchon 1959, 1952). We checked this possibility on S. amica by using posterior segments re-amputated 50- and 75-days post-amputation, X-irradiated or not, and even after stolon release. All were able to regenerate anteriorly 3 to 4 segments at the same time, suggesting that sexual maturation did not disturb regeneration. Two factors (digestive tube, nerves) which play a role in the availability of mesodermal regeneration cells might explain the difference of SAZ function in PR vs AR. This problem warrants further investigations, especially at the molecular level. This was explored recently at the transcriptomics level on two syllid species: Sphaerosyllis hystrix, which exhibits limited AR and Syllis gracilis able to well regenerate anterior segments, and the entire part of the anterior digestive tube (Ribeiro et al. 2019). This study has shown that gene expression during PR and normal growth are similar but not identical to AR. However, this preliminary study does indicate the genes more particularly concerned with PR vs AR, and more investigations are needed to know which gene(s) are involved in the different regeneration phases, and how they can be modulated in experimental situations to clarify how they participate in regeneration.