Direct Differentiation of Human Pluripotent Stem Cells into Haploid Spermatogenic Cells

TLDR

Human embryonic and induced pluripotent stem cells can generate primordial germ cells but have not previously been shown to produce spermatogonia, haploid spermatocytes, or spermatids. We demonstrate that human embryonic and induced pluripotent stem cells can differentiate directly, without genetic manipulation, into advanced male germ cell lineages—including spermatogonia‑like, spermatocyte‑like, and haploid spermatid‑like cells expressing characteristic markers—and that these spermatids exhibit uniparental genomic imprints at H19 and IGF2 comparable to human sperm, indicating a novel in‑vitro model for spermatogenesis.

Abstract

Human embryonic stem cells (hESCs) and induced pluripotent stem cells (hiPSCs) have been shown to differentiate into primordial germ cells (PGCs) but not into spermatogonia, haploid spermatocytes, or spermatids. Here, we show that hESCs and hiPSCs differentiate directly into advanced male germ cell lineages, including postmeiotic, spermatid-like cells, in vitro without genetic manipulation. Furthermore, our procedure mirrors spermatogenesis in vivo by differentiating PSCs into UTF1-, PLZF-, and CDH1-positive spermatogonia-like cells; HIWI- and HILI-positive spermatocyte-like cells; and haploid cells expressing acrosin, transition protein 1, and protamine 1 (proteins that are uniquely found in spermatids and/or sperm). These spermatids show uniparental genomic imprints similar to those of human sperm on two loci: H19 and IGF2. These results demonstrate that male PSCs have the ability to differentiate directly into advanced germ cell lineages and may represent a novel strategy for studying spermatogenesis in vitro.