Transplantation of 3D scaffolds seeded with human embryonic stem cells: biological features of surrogate tissue and teratoma-forming potential.

AIM: To generate complex surrogate tissue by transplanting 3D scaffolds seeded with human embryonic stem cells (hESCs) between the liver lobules of severe combined immunodeficient (SCID) mice and to assess the teratoma-forming potential. MATERIALS & METHODS: 3D poly-(lactic-co-glycolic acid) (PLGA) scaffolds coated with laminin were seeded with hESCs and then transplanted between the liver lobules of SCID mice. After a period of in vivo differentiation, the scaffolds were retrieved and analyzed using reverse transcription polymerase chain reaction, immunofluorescent staining and scanning electron microscopy. RESULTS: A proportion of the hESCs within the scaffolds differentiated into cells that produced proteins characteristic of specific tissues, including endoderm and pancreatic markers glucogon-like peptide-1 receptor, islet amyloid polypeptide and Insulin. Markers of hepatic and neuronal lineages were also investigated. Major matrix proteins abundant in multiple tissue types, including collagen I, laminin and collagen IV, were found to be profuse within the scaffold pores. Transplantation of the seeded scaffolds between liver lobules also resulted in extensive vascularization both from host blood vessel incursion and the differentiation of hESCs into endothelial progenitor cells. An investigation of teratoma-forming potential demonstrated that transplantation of 3D scaffolds seeded with hESCs will, under certain conditions, lead to the growth of teratomas. DISCUSSION: Transplantation of 3D scaffolds seeded with hESCs between liver lobules resulted in the development of surrogate tissue containing cells that produced proteins representing the pancreatic, hepatic and neuronal lineages, the assembly of an extracellular matrix structure and the formation of a vasculature. hESCs seeded within 3D scaffolds and transplanted into SCID mice were capable of forming teratomas. However, the formation and progression of teratoma growth is shown to be dependant on both the site of transplantation and the treatment of cells prior to transplantation.

Prediction of embryo developmental potential and pregnancy based on early stage morphological characteristics.

OBJECTIVE: To analyze the association between morphological details at different stages of culture with blastocyst development, with an aim to improve selection for transfer. DESIGN: Retrospective audit of data. SETTING: Tertiary referral center and university hospital. PATIENT(S): Two hundred sixty-eight couples underwent 357 treatment cycles. INTERVENTION(S): Oocyte pickups for IVF or intracytoplasmic sperm injection (ICSI) after ovarian stimulation. Embryos were individually cultured and examined on days 0-2 for morphological details and developmental characteristics, and selected for transfer, freezing, or further culture. MAIN OUTCOME MEASURES: The association of blastocyst development and pregnancy with morphological characteristics. RESULT(S): Five morphological characteristics (appearance of the cytoplasm, pronuclei and nucleoli, cytoplasmic deficit, and developmental rate) showed the strongest association with blastocyst development. By combining information from all days of culture into a cumulative score, prediction was greatly improved, compared to only using day 2 morphology. Cytoplasmic dysmorphisms of the oocyte, including accumulation of smooth endoplasmic reticulum, were associated with poor developmental performance. Differential weighting of these characteristics was included in a new embryo scoring system, which showed a strong correlation with implantation. CONCLUSION(S): Weighting individual morphological characteristics of zygotes and embryos and combining them into a cumulative embryo score can improve selection of embryos for transfer.