Meeting Abstract

29.1  Monday, Jan. 4  Bloody Whole Body Regeneration! BROWN, FD*; KEELING, EL; LE, AD; SWALLA, BJ; University of Washington, Seattle; Universidad de los Andes, Bogotá; University of Washington; California Polytechnic State University, San Luis Obispo; University of Washington, Seattle federico.brown@tuebingen.mpg.de

Colonial ascidians exhibit one of the most extreme cases of regeneration. Upon removal of all individuals in a colony of Botrylloides violaceus, the vascular network and blood left within the tunic of the colony reorganizes, and aggregates of blood differentiate into buds. One of these buds continues to complete regeneration of a whole individual. Using phase contrast microscopy, time-lapse video recording and detailed histological studies of regenerating colonies, we describe the earliest events of regeneration. We examine the pattern of cell proliferation by immunohistology using proliferating cell nuclear antigen (PCNA) antibodies. Next, we report the expression of piwi, a stem cell maintenance marker involved in microRNA processing and stem cell maintenance, in hemocytes surrounding the early regenerates. We rarely found Piwi or PCNA in differentiating tissues during vascular budding, suggesting that cells that form the epithelial tissues during budding and regeneration originate mostly from circulatory hemocyte precursors, and likely include stem cell progenitors. First attempts to establish lineage tracing are now in progress. Preliminary results using the nuclear stain DAPI (<360 nm excitation range to avoid endogenous autofluorescence) show that it can be reliably followed in the colony up to a month after labeled hemocyte injection. We find DAPI labeled cells integrate into tissue epithelia of different germ layers. Taken together, we propose that multiple stem cell types occur within the hemocytes, and that they undergo proliferation in the vasculature before differentiating into epithelial tissues.