Generation and proteome profiling of PBMC-originated, iPSC-derived lentoid bodies.

Abstract

Here, we report proteome profiling of peripheral blood mononuclear cell (PBMC)-originated, induced pluripotent stem cell (iPSC)-derived, lens-like organoids termed lentoid bodies at two differentiation time points. A small aliquot of the blood sample was ascertained to collect PBMCs that were reprogrammed to iPSCs. The PBMC-originated, iPSCs were differentiated to lentoid bodies employing the "fried egg" method. Quantitative real-time PCR (qRT-PCR) analysis revealed increased expression levels of lens-associated markers in lentoid bodies while transmission electron microscopy identified closely packed lens epithelial- and differentiating fiber-like cells in lentoid bodies. Total cellular protein was extracted from lentoid bodies at differentiation day 25 and mass spectrometry identified a total of 9,473 proteins. The low counts of crystallin proteins at differentiation day 25 prompted us to re-examine the proteome at differentiation day 35 as we reasoned that 10 additional days of differentiation will increase the crystallin count. However, we did not detect any substantial increase in crystallin protein counts at differentiation day 35. In conclusion, we report generation and proteome profiles of PBMC-originated, iPSC-derived lentoid bodies at multiple differentiation time points.