Basement Membrane Promotes Ciliary Body Derived Cell Spheres Formation

Controversy still exists whether ciliary body contains stem cells (CBSCs) or epithelial cells (CBECs). CBSCs were reported for differentiation capacity and CBECs for high reprogramming efficiency to produce induced Pluripotent Stem (iPS) cells. However, ciliary body derived cells (CBCs) application as a cell therapy needs a guaranteed isolation and culture protocol from ciliary body tissue strip. Four protocols were compared; one (I) belonged to a published article (Gu et al., 2007) and the rest three, protocols II, III and IV, were developed. Protocol II, III and IV used trypsin-EDTA and mechanical scraping to isolate single CBCs, basement membrane (BM) fragments and other debris. Protocols used 40 and/or 100 μm cell strainers at different steps to remove different scraped tissue fragments and isolated single CBCs. Protocol III and IV contained CBCs attached on BM. Success/failure numbers in out of 5 repeated experiments per protocol (n=5) were 2/3 (I), 3/2 (II), 4/1 (III) and 5/0 (IV). Cultures were very clean (I), partially clean (II and III) and dirty (IV). Protocol III and IV based cultures showed clumps of cell spheres. This study results confirmed that BM promotes cell proliferation and primary cell spheres formation. CBCs are in controversy for recognition as epithelial or stem cells but their successful isolation and culture, and iPS cells reprogramming protocols can open a new era for reinforcing their applicability.