Epithelial cell cultures from Botryllus schlosseri palleal buds: Accomplishments and challenges

Claudette Rabinowitz, Baruch Rinkevich

Research output: Contribution to journalArticlepeer-review


This study focuses on recent improvement in epithelial monolayer cultures originating from whole extirpated Botryllus schlosseri (Urochordata) buds. Buds (n = 2,000) were taken at different ('A' to 'D' blastogenic stages. We tested the suitability of 35 combinations of various substrates and media on attachment, cell spread, epithelial growth frequencies and on monolayer lifespans. Under favorable conditions, cultured buds at blastogenic stages 'B' to 'D' (but not stage 'A') started to attach to the substrates following a 3-day transient period that leads to formation of spheres and attached monolayers. Substrate type is important for the attachment and the development of monolayers. Under various culture conditions, some of stages 'B' and 'C' buds develop (3-20 days) one or more large (1 mm diameter) spheres. Stage 'D' buds develop monolayers (up to 20% of buds) without going through a sphere phase. Neither spheres nor attached monolayers of epithelium were observed in stage 'A' bud cultures. Spheres grew at a rate of 60 μm in diameter per day using specific medium types and did not attach unless the appropriate substrate was present. When attached, epithelial monolayers expanded at a rate of 200 μm in diameter per day, for 3-15 days, and subsequently detached and died. Sixteen types of media were tested. Medium and substrate combinations were found to determine epithelial lifespan. These results revealed significant improvements in the culture of epithelial monolayers from Botryllus palleal buds. However, an early senescence of the developed epithelial sheets (up to two weeks from onset of appearance) may indicate an internal ageing clock that should be taken into consideration in future approaches.

Original languageEnglish
Pages (from-to)137-148
Number of pages12
JournalMethods in Cell Science
Issue number3-4
StatePublished - Jan 2004
Externally publishedYes

Bibliographical note

Funding Information:
This study was supported by the NIH (ROI-DK54762) by the US-Israel Binational Science Foundation and by the Israel Science Foundation (456/01).


  • Botryllus
  • Cellular senescence
  • Epithelial cells
  • Invertebrates

ASJC Scopus subject areas

  • Cell Biology


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