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Neoblasts in Platyhelminthes are the only cells to proliferate and differentiate into all cell types. In Macrostomum lignano, the incorporation of 5'-bromo-2'-deoxyuridine (BrdU) in neoblasts confirmed the distribution of S-phase cells in two lateral bands. BrdU labeling for light and for transmission electron microscopy (TEM) identified three populations of proliferating cells: somatic neoblasts located between the epidermis and gastrodermis (mesodermal neoblasts), neoblasts located within the gastrodermis (gastrodermal neoblasts), and gonadal S-phase cells. In adults, three stages of mesodermal neoblasts (2, 2-3, and 3) defined by their ultrastructure were found. Stage 1 neoblasts where only seen in hatchlings. These stages either were phases within the S-phase of one neoblast pool or were subsequent stages of differentiating neoblasts, each with its own cell cycle. Regular TEM and immunogold labeling provided the basis for calculating the total number of neoblasts and the ratio of labeled to non-labeled neoblasts. Somatic neoblasts represented 6.5% of the total number of cells. Of these, 27% were labeled in S-phase. Of this fraction, 33% were in stage 2, 46% in stage 2-3, and 21% in stage 3. Immunogold labeling substantiated results concerning the differentiation of neoblasts into somatic cells. Non-labeled stage 2 neoblasts were present, even after a 2-week BrdU exposure. Double labeling of mitoses and FMRF-amide revealed a close spatial relationship of mesodermal neoblasts with the nervous system. Immunogold-labeled sections showed that nearly 70% of S-phase cells were in direct contact or within 5 microm from nerve cords.

Neoblasts are potentially totipotent stem cells and the only proliferating cells in adult Platyhelminthes. We have examined the cellular dynamics of neoblasts during the posterior regeneration of Macrostomum lignano. Double-labeling of neoblasts with bromodeoxyuridine and the anti-phospho histone H3 mitosis marker has revealed a complex cellular response in the first 48 h after amputation; this response is different from that known to occur during regeneration in triclad platyhelminths and in starvation/feeding experiments in M. lignano. Mitotic activity is reduced during the first 8 h of regeneration but, at 48 h after amputation, reaches almost twice the value of control animals. The total number of S-phase cells significantly increases after 1 day of regeneration. A subpopulation of fast-cycling neoblasts surprisingly shows the same dynamics during regeneration as those in control animals. Wound healing and regeneration are accompanied by the formation of a distinct blastema. These results present new insights, at the cellular level, into the early regeneration of rhabditophoran Platyhelminthes.

A survey of the current knowledge on stem cells and cell proliferation in turbellarians is presented, with special focus on recent results obtained by the authors when studying cell kinetics and cultivating neoblasts from various species. Differentiated somatic cells do not divide in flatworms. In these animals neoblasts constitute a proliferative compartment. These cells are rather uniform when viewed through a light microscope, but constitute a heterogeneous population comprising actual stem cells, progenitors and early differentiation stages. Cell kinetics were monitored by the incorporation of 5-bromo-2´-deoxyuridine and by immunocytochemical staining for mitoses in the marine microturbellarians Macrostomum, Microstomum and Convolutriloba. Distribution patterns of proliferating cells within the body were documented, and a tentative interpretation of the differences observed is given. Correlations to the nervous system and the mode of reproduction seem to exist. To analyse neoblasts quantitatively and establish primary cultures, neoblasts from the freshwater planarians (triclads) Dugesia and Schmidtea were isolated, purified and subjected to various culture conditions, with the ultimate but not yet achieved goal of establishing a permanent stem cell line. The role of stem cells in flatworms is compared with the situation in mammals, with special focus on embryogenesis, germline and strategies of cell renewal.