A Survey and Critical Evaluation of Isolation, Culture, and Cryopreservation Methods of Human Amniotic Epithelial Cells

Expansion of human amniotic epithelial cells using condition cell reprogramming technology

“…The use of human serum (in DMEM/F12) has been reported previously [10]; however, the cells plateaued at passage 2, and no molecular characterization was performed on the cells cultured in human serum. We have evaluated the impact of various cryopreservation media on stem cell markers and post-thaw viability in a recent comparative study [6]. As compared to commercially available and other tested cryopreservation media (e.g., amniotic fluid, CryoStor CS10, Stem Cell banker), cryopreservation media containing human serum used in this study (see Methods) also provides an easy and promising solution to preserve these cells effectively.…”

“…While the conditions that promote the indefinite proliferation of hAECs in XF conditions have not been adequately explored, there are several recent studies that address possible improvements in both cryopreservation and expansion of hAECs [ 6 ]. In this study, we demonstrated that hAECs are amenable to ex vivo expansion under CRC growth conditions in XF conditions.…”

“…As discussed in a recent study, several factors affect the isolation, yield and purity of hAECs [ 6 ]. All phases of sample collection are vulnerable to errors; therefore, precise and timely handling is critically important.…”

“…Despite the remarkable stem-like pluripotency of hAECs and ease of collection, as well as ethical considerations, the use of these cells is still restricted due to in part inefficient and limited ex vivo expansion [5]. To date, many protocols have been published to culture hAECs; however, an inability to scale and expand hAECs without significantly compromising their unique and privileged nature precludes the widespread use of hAECs for regenerative therapies [6]. This limitation is primarily due to the use of sub-optimal culture conditions that limit expansion through the induction of immunostimulatory markers, a concomitant reduction in pluripotency, and ultimately, the cessation of proliferation due to cellular senescence, mostly after passage 2-3 [7][8][9][10].…”

Amniotic stem cells as a source of regenerative medicine to treat female infertility

“…Due to the differentiation ability into germ cells and oocyte-like cells, stem cells may adopt the following mechanisms to repair ovarian functions; i) heal injured reproductive tissues by replenishing healthy cells, ii) restore or increase the number of secondary and mature follicles, iii) improve microenvironment by secreting paracrine factors and ameliorate ovarian function, iv) immune regulation by secreting anti-inflammatory factors, and v) regulate the hormonal levels that maintain estrous reproductive cycles and stimulate ovulation such as E2 (Estradiol), AMH (Anti-Müllerian hormone), and FSH (Follicle-stimulating hormone). Despite the great success of stem cell therapeutics in reproductive disease management, the ethical concerns, heterological nature, low yield, and lower ex-vivo proliferation rate of adult stem cells limit their clinical translation [3]. Conversely, easily accessible placental-derived amniotic stem cells (ASCs) represent a viable therapeutic option due to their successful uses in other diseases and their differentiation ability toward cells of germline lineage [4][5][6] Stem cells isolated from the umbilical cord have wellestablished therapeutic uses and are discussed in literature excessively [7].…”