The Regenerative Potential of Crinoids

Crinoids are well known for their spectacular regenerative capabilities extensively and successfully employed to reconstruct both external parts, namely arms, pinnules and cirri, and internal organs, such as digestive apparatus, gonads and even complete visceral mass, which can be frequently lost following traumatic injury, predation or spontaneous autotomy. Specimens collected in nature always show regenerating arms at different stages of growth (Fig. 1a). These regenerative phenomena can be easily reproduced in the laboratory by mimicking the autotomy conditions and amputating the arms at the level of the autotomy plane (sutures) (Fig. 1b-d).Arm regeneration in crinoids represents the most thoroughly explored model in echinoderm regeneration studies (Candia Carnevali and Bonasoro 2001b). Recently, we have carried out a comprehensive study of the overall process of arm regeneration in the comatulid Antedon mediterranea, a valuable and flexible experimental model previously successfully employed by old classical studies of developmental biology (Per-rier 1873;Minckert 1905;Reichensperger 1912),which was re-explored in all its aspects from the macroscopic to the molecular level.

This phenomenon can be described on the whole as a typical blastemal regeneration in which new structures develop from migratory pluripotential, actively proliferating cells in the presence of presumptive regulatory factors. The overall process can be subdivided into three main phases (Fig. 2a): a repair phase, an early regenerative phase and an advanced regenerative phase,

Experiments Crinoids

Fig. 1. Schematic presentation of the experimental model. a Specimen of Antedon mediterranea with three regenerating arms at different stages (arrows). b Main anatomical features of the arm in sagittal section. ag Ambulacral groove; cc coelomic canal; li ligament; mu muscle; os ossicle; su suture. c Normal arm, experimental amputations and regenerating arm. d Stereomicroscopic view of regenerating arm (2 weeks post-amputation). Bar 1 mm

Fig. 1. Schematic presentation of the experimental model. a Specimen of Antedon mediterranea with three regenerating arms at different stages (arrows). b Main anatomical features of the arm in sagittal section. ag Ambulacral groove; cc coelomic canal; li ligament; mu muscle; os ossicle; su suture. c Normal arm, experimental amputations and regenerating arm. d Stereomicroscopic view of regenerating arm (2 weeks post-amputation). Bar 1 mm whose crucial aspects are related to common fundamental mechanisms such as (1) intervention of stem cells and/or employment of dedifferentiated cells (Fig. 2 b-e), (2) cell migration and proliferation, (3) contribution of putative growth factors, particularly in terms of specific neurally derived factors and (4) mechanisms of pattern formation. The data obtained so far are derived from an integrated approach which utilizes different methods (first of all classical methods of microscopy - LM, confocal, TEM, SEM - and specific methods of immunocytochemistry, but also basic methods of biochemistry and molecular biology) on experimentally induced arm regenerations (standard or abnormal) obtained under significantly different experimental conditions, including extreme mutilations (explants) or exposure to specific types of environmental contaminants. In particular, the normal mechanisms and pattern of the regenerative processes under standard conditions have been established in serial experiments of regeneration at different stages following pseudo-autotomic amputations (Candia Carnevali et al. 1993, 1995, 1997). A parallel analysis has been carried out on the regenerative processes of both the normal regenerating arms and the respective amputated arm segments

Experiments Crinoids

Fig. 2. a Schematic reconstruction of the main phases of arm regeneration. Top downwards Repair phase [0-24 h post-amputation (pa)], early regenerative phase (24-72 h pa), advanced regenerative phase (72 h to 3 weeks pa). b Regenerative blastema; cl cicatricial layer; ra: regrowing arm. b-e TEM: migratory cells involved in regenerative processes. Presumptive stem cells: amoebocyte (b),coelomocyte (c),phagocyte (d), granulocyte (e). Bars 4 |m

Fig. 2. a Schematic reconstruction of the main phases of arm regeneration. Top downwards Repair phase [0-24 h post-amputation (pa)], early regenerative phase (24-72 h pa), advanced regenerative phase (72 h to 3 weeks pa). b Regenerative blastema; cl cicatricial layer; ra: regrowing arm. b-e TEM: migratory cells involved in regenerative processes. Presumptive stem cells: amoebocyte (b),coelomocyte (c),phagocyte (d), granulocyte (e). Bars 4 |m

(explants, Fig. 1 c; Candia Carnevali et al. 1998), which can be maintained under good living conditions for about 3 weeks and represent excellent models for testing the arm regenerative potential in terms of autonomy of resources and control and for comparing regenerative mechanisms in the same individual. Different types of isolated explants have been successfully employed: during the culture period they are able to undergo extensive repair and regenerative processes in parallel with their donor arms. Comparison between the regenerative processes of arm explants and normal regenerating arms of corresponding stages highlights that beside general similarities in the basic regenerative processes there are some meaningful differences in terms of mechanisms employed and cellular/tissue elements involved. The regenerative potential, mechanisms and pattern have also been explored and compared under other experimental conditions, particularly with regard to aberrant regenerations resulting from arms deliberately subjected to traumatic mutilations which do not reproduce autotomy (Candia Carnevali and Bona-soro 2001b). The bulk of the results obtained so far in crinoids not only throw light on the most relevant aspects related to wound healing, morphogenesis, differentiation and growth in echinoderm regeneration, but also strongly suggest employing this fascinating and promising experimental model for a successful applied approach.

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