92 MICHELA SUGNI1,2, CINZIA FERRARIO1,2, FRANCESCO BONASORO1,
MARIA DANIELA CANDIA CARNEVALI1
1 Dipartimento di Scienze e Politiche Ambientali, Università degli Studi di Milano 2 Center for Complexity & Biosystems, Università degli Studi di Milano, Milano, Italia
PLURIPOTENT VERSUS REPROGRAMMED CELLS:
CELL PLASTICITY IN ECHINODERM REGENERATION
Echinoderms display remarkable regenerative capabilities and offer a variety of models to study this phenomenon widely distributed throughout the Phylum. Although their regenerative phenomena have been traditionally attributed to two different mechanisms (i.e. epimorphosis and morphallaxis), the true origin and fate of the involved cells are still unclear. An up-to-date overview of cell recruitment processes in the different echinoderm classes is here provided in order to define the state of the art, including the main unsolved issues, as well as the necessary future steps to cover the knowledge gap. Among stellate echinoderms, crinoids are the only group clearly displaying the recruitment of morphologically undifferentiated cells stocked in the stump tissues (i.e. coelomic canals and brachial nerve): these undifferentiated cells actively migrate to form a true blastema where they undergo proliferation and differentiation up to to regenerate the lost tissues. Reprogramming of differentiated cells occurs only in stress conditions. In contrast, a true regenerative blastema is missing in brittle stars and starfish, which apparently mainly rely on recruitment of dedifferentiated cells from mature tissues. In starfish, dedifferentiation is massively employed at the level of muscle tissues, also in location far from the wound site. In both these classes progenitor-like cells are provided and recruited via epithelial-mesenchymal transition (EMT) from coelomic epithelium. In sea cucumbers neural and intestinal regeneration are the main process under investigation. In the former, the absence of “stemness” marker in the transcriptome suggests that radial nerve cord regeneration depends on dedifferentiation of the supporting glial cells that re-differentiate in both the same cytotype and new neurons. Myocyte dedifferentiation markedly occurs during gut regeneration. In sea urchins, damaged test and broken spines are reformed through dedifferentiation of stump cells with only minor local cell proliferation, whereas totally removed spines are regenerated via undifferentiated (pluripotent) cells. Overall, echinoderm regeneration appears mainly to rely on dedifferentiation phenomena rather than recruitment of pluripotent cells already stocked in the stump tissues, the exact origin, identity and fate of the involved cells being still unknown in most cases. Echinoderm tissues, especially coelomic epithelium and muscles, show a high potential of plasticity in terms of cell differentiation/dedifferentiation and activity (proliferation, migration), and EMT plays key roles in this plasticity. Cell tracking coupled to molecular and microscopy approaches will be strongly needed to unravel in detail the strikingly effective cellular mechanisms and pathways (from cell origin to fate) employed by echinoderms in their regeneration processes.
93
Simposio 4
Cellule staminali, differenziamento e riprogrammazione cellulare:
modelli tradizionali e modelli innovativi
Coordinatori
Loriano Ballarin, Luciana Dini
94 FRANCESCA BALLIN, NICOLA FRANCHI, ANNA PERONATO, LORIANO BALLARIN
Dipartimento di Biologia, Università di Padova
EXPRESSION STUDY OF MOLECULAR MARKERS INVOLVED IN STEMNESS MAINTEINANCE AND DIFFERENTIATION IN THE COLONIAL ASCIDIAN
BOTRYLLUS SCHLOSSERI
Ascidians are invertebrate chordates, members of the subphylum Tunicata, and represent the sister group of vertebrates. They offer the opportunity to investigate and compare the behaviour of both embryonic and adult stem cells. Morphological data suggest the presence of undifferentiated haemocytes (haemoblasts) able to proliferate and give rise to terminally differentiated cells. Relevant studies were also carried out in the neural lineage, in which neural progenitor cells regenerate the brain after extirpation. In B. schlosseri, during the cyclical generation change, bud primordial cells, probably deriving from a pool of long-living stem cells, are able to give rise to the neural complex. We screened the B. schlosseri genome and transcriptome, looking for transcripts/genes showing similarity to vertebrate molecular markers of haematopoietic and neural stem cells. Four sequences, orthologous to mammalian transcripts considered markers of haematopoietic progenitor cells, were identified in B. schlosseri. They are: bsabcg2, bscd133, bsgata1/2/3 and bsgata4/5/6. ISH on haemocyte monolayers and colony sections, resulted in labelling of cells in the sub-endostylar haemolymph lacunae. This results matches previously morphological data that identified the endostyle as a stem cell niche. Quantitative real time PCR (qRT-PCR) highlighted the overexpression of the considered genes in the mid-cycle phase of the blastogenetic cycle. During this phase, there is the formation of new secondary buds emerging from the primary buds. The high expression levels of bsabcg2, bscd133, bsgata1/2/3 and bsgata4/5/6 genes in the mid-cycle phase reflect the presence of undifferentiated cells involved in proliferative and differentiation events required for the new blastogenetic generation.
For the neural lineage, we identified two transcripts orthologues of vertebrate neural stem cell markers (BsSox2 and BsMsi2). ISH with riboprobes for BsSox2 and BsMsi2 revealed a common labelling in the endostyle niche. The presence of bssox2 and bsmsi2 transcripts in the cells of the region known to be a stem cell niche, led us to conclude that in B. schlosseri a single population of pluripotent stem cells is probably present that could differentiate into haematopoietic or neural cells.
95
ALESSANDRA SALVETTI1, LEONARDO ROSSI1, ANDREA DEGL’INNOCENTI2, FRANCO VERNI3, SANDRA GHELARDONI4, PAOLA IACOPETTI1, JACK VAN LOON5,
GIANNI CIOFANI1,6
1 Unità di Biologia e Genetica, Dipartimento di Medicina Clinica e Sperimentale, Università di Pisa 2 Smart Bio-Interfaces, Istituto Italiano di Tecnologia
3 Dipartimento di Biologia, Università di Pisa
4 Dipartimento di Patologia Chirurgica, Medica, Molecolare e dell'Area Critica, Università di Pisa
5a DESC (Dutch Experiment Support Center), Department of Oral and Maxillofacial Surgery / Oral Pathology, VU University Medical Center & Academic Centre for Dentistry Amsterdam (ACTA), Amsterdam Movement Sciences, Amsterdam, The Netherlands and 5b
European Space Agency Technology Center (ESA-ESTEC), TEC-MMG LIS Lab, Noordwijk, The Netherlands 6 Department of Mechanical and Aerospace Engineering, Politecnico di Torino