Year 3

The specific aims of the funded proposal # RL1-00670 entitled “Derivation and comparative analysis of human pluripotent hESCs, hIPSCs and hSSCs: Convergence to an embryonic phenotype” as proposed were to: 1) Derive additional hSSC (human spermatogonial stem cell) and hIPSC (human induced pluripotent stem cell) lines. 2) Compare hSSCs, hESCs and hIPSCs in terms of critical molecular, genetic and developmental characteristics. 3) Incorporate well-characterized first-generation hSSCs and IPSCs into a Stanford human pluripotent stem cell bank for broad distribution to the scientific community. Over the entire funding period, we have made substantial progress and have continued to meet or exceed our goals in many areas. However, we have also met a couple challenges that we are now addressing especially in of final characterization of lines and in banking for distribution of lines quite broadly. Our goal is to decisively address issues regarding the definition of hSSCs and pluripotency; we note that mouse SSCs clearly are pluripotent in every sense and we likely have to adjust conditions further to accomplish full pluripotency with hSSC lines. We expect that a comparison of our hSSC lines, with the hiPSCs derived from them will allow adjustments to be made, as necessary. However, we continue to observe as we also initially reported that hSSC lines do not readily form (they are “null” for) teratomas in vivo. We note that we have had several publications that indicate our progress as follows (asterisks indicate new submissions):

1. Byrne JA, Nguyen HN, Reijo Pera RA (2009) Enhanced generation of induced pluripotent stem cells from a subpopulation of human fibroblasts. PLoS One 4, e7118. PMC2744017
2. *Panula S, Medrano JV, Kee K, Bergstrom R, Nguyen HN, Byers B, Wilson KD, Wu JC, Simon C, Hovatta O, Reijo Pera RA (2010) Human germ cell differentiation from fetal- and adult-derived induced pluripotent stem cells. Hum Mol Genet 20:752-62.
3. *HN Nguyen, Byers B, Cord B, Shcheglovitov A, Byrne J, Gujar P, Kee K, Schuele B, Dolmetsch RE, Langston W, Palmer TD, Reijo Pera RA (2011) LRRK2 mutant iPSC-Derived DA neurons demonstrate increased susceptibility to oxidative stress. Cell Stem Cell 8, 1–14.
4. *B Byers, B Cord, HN Nguyen, B Schule, J Byrne, JW Langston, RA Reijo Pera, T Palmer (2011) Emergence of an early disease phenotype from Parkinson’s disease induced pluripotent stem cells. PLoS One (in press).
5. JV Medrano, C Ramathal, HN Nguyen, C Simon, RA Reijo Pera (submitted) Divergent RNA-binding proteins, DAZL and VASA, induce meiotic progression in human germ cells derived in vitro from both hESCs and iPSCs. Stem Cells.

As also noted in the last report, we have generated intellectual property on: 1) use of human SSCs for regenerative medicine and/or fertility preservation, 2) use of SSEA3 to isolate a preferentially-reprogrammable somatic cell sub-population, and 3) derivation of unique iPSC lines from patients with genetically-defined Parkinson’s Disease (PD) for pharmacological and toxicological studies. Overall progress will continue with the final establishment of banked lines that are able to be broadly distributed.