G Pellegrini is a member of the Board of Directors of Holostem Terapie Avanzate S.r.l. among entrepreneurs AF-DX 384 investing in the sector and thus slowing down their diffusion. In the meantime, scientific papers C in addition to huge research investments in this area of medicine C increase expectations with regards to clinical trials and routine clinical treatments. In Europe and the USA, few ATMPs have been approved; though they are still in their infancy, it is useful to understand their procedures and share experiences, in order to facilitate the development of this area of personalized medicine. The first example of a therapeutic use of cells extracted from human tissue was the use of selected hematopoietic cells in hematologic diseases and in oncology . However, this tissue is considered a typical transplant and not an ATMP, since transplanted cells are not extensively manipulated. Legally speaking, selecting and transplanting cells is considered a minimal manipulation to the same extent as an organ transplant, since the same risks do not apply to cell pathway stimulation and extensive proliferation manipulation of cells destined for transplantation in tissue reconstruction was successfully obtained with epidermal cells for the treatment of severely burnt patients [2,3]. Corneal cell therapy After these first trials, in 1997 the human limbo-corneal epithelium was reconstructed and transplanted into two patients . It was obtained by culturing cells extracted from a small biopsy of the ocular surface; the two patients were suffering AF-DX 384 from a limbal stem cell deficiency with a resultant defect in corneal repair and AF-DX 384 opacification of the anterior surface of the eye. The clinical outcome was good and results were proven to be stable up to the 2 2 years of documented follow-up period, suggesting that the selected biopsy area contained the cells needed for a long-term regeneration. This was one of the first examples of application of regenerative medicine in the field of ophthalmology, and it was successfully reproduced, with some modifications, by Ivan Schwab in the USA , Ray Tsai in Taiwan , and later by Geeta Vemuganti in India . The treatment became a routine patient treatment in Italy in 2004 (reimbursed by the National Health System), was accepted in India in 2008, but never spread out of these specific countries. In the USA, the treatment was stopped due to regulatory requirements. After the first proof CDK4 of principle, lengthy studies were needed AF-DX 384 to investigate the mechanisms controlling stable regeneration and enabling high reproducibility in the manufacturing process. Location/identification of stem cells Since adult stem cells maintain a population of highly differentiated but short-lived cells such as epithelia, data suggest that human corneal stem cells can be found in the limbus. The first step of investigation was mapping the stem cells of the two epithelia covering the ocular surface: the cornea and the conjunctiva (Figure 1). Since the two epithelia behave differently, it had to be assessed whether a single stem cell could produce both epithelia, under the control of a local microenvironment, or whether two stem cells committed to different cell fates could be located in the areas to be defined . Open in a separate window Figure 1.? Corneal and conjunctival areas selected for biopsy retrieval. Superior and inferior fornix (yellow line), superior, inferior, nasal and temporal bulbar conjunctiva (black asterisk), superior, inferior, temporal and nasal limbus (light blue line), paracentral cornea (blue line), central cornea (white asterisk). All defined areas were cultured and analyzed for stem cell content. Studies in humans have shown the presence of two distinct adult stem cells, capable of corneal and conjunctival tissue regeneration, respectively; in particular, stem cells from the conjunctiva were shown to be bipotent (able to generate epithelium and goblet cells) and ubiquitous in the tissue, whereas corneal stem cells were proven to be segregated in the limbus , giving rise to progenitors covering the AF-DX 384 corneal surface. Different results on other mammals were reported by Majo (2016) Manuscript in Preparation]. Carrier selection In recent years, the environment of keratinocyte stem cells.