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Biomedical subjects

Diana Valbuena

Publications and source records attributed to Diana Valbuena.

7 recordsLinked to original sources
PubMed2006

Heteroparental blastocyst production from microsurgically corrected tripronucleated human embryos.

OBJECTIVE: To prove the efficiency of identification and removal of one of the surplus paternal pronuclei in dispermic IVF zygotes to obtain heteroparental blastocysts. DESIGN: Experimental. SETTING: One hundred fourteen tripronucleated (3PN) embryos from conventional IVF. PATIENT(S): After informed and signed consent, the patients from Instituto Valenciano Infertilidad (IVI), Valencia, donated their abnormally fertilized embryos. INTERVENTION(S): Seventy-two embryos were diploidized by microsurgical removal of the pronucleus located at the farthest position to the second polar body. Forty-two 3PN embryos served as controls. MAIN OUTCOME MEASURE(S): Survival and correction rate; in vitro development up to the blastocyst stage; X, Y, and 18 chromosome determination by triple fluorescent in situ hybridization and, inheritance analysis for 10 polymorphic repeat regions using polymerase chain reaction (PCR) amplification and sequencing. RESULT(S): Seventy-eight percent of 3PN zygotes (56/72) survived manipulation and eventually 51 zygotes had two pronuclei (71%). Forty-one percent of manipulated embryos progressed in vitro to the blastocyst stage (21/51). Fluorescent in situ hybridization analysis performed on eight manipulated embryos confirmed their diploid state; all four controls were triploid. Heteroparental inheritances were also confirmed in four of six manipulated embryos. CONCLUSION(S): Heteroparental blastocysts can be derived from corrected dispermic zygotes.

Adult↗
PubMed2006

Human embryo culture.

Human embryonic stem cells (hESCs) are derived from preimplantation embryos. Approximately 60% of human embryos are blocked during in vitro development. Although statistics are inconclusive, experience demonstrates that hESCs are more effectively derived from high-quality embryos. In this way, optimal human embryo culture conditions are a crucial aspect in any derivation laboratory. Embryos can be cultured solely with sequential media or cocultured on a monolayer of a given cell type. This chapter explores general aspects of human embryonic development, the concept of sequential culture and coculture, and specific protocols and procedures in which the authors are experienced, including the results obtained.

Coculture Techniques↗
PubMed2006

Derivation of clinical-grade human embryonic stem cells.

Embryonic stem cells proliferate in vitro while maintaining an undifferentiated state, and are capable of differentiating into most cell types under appropriate conditions. These properties imply great potential in the treatment of various diseases and disabilities. In fact, the first clinical trials with hESC for treating spinal cord injuries will begin next year. However, therapeutic application of human embryonic stem cell derivatives is compromised by the exposure of existing lines to animal and human components, with the subsequent risk of contamination with retroviruses and other pathogens, which can be transmitted to patients. The scientific community is striving to avoid the use of xenogeneic or allogeneic components in the process of derivation new hESC lines. This review summarizes attempts that have been made to avoid these contaminants and the breakthroughs achieved in the derivation of clinical-grade hESC that could be used for therapeutic purposes.

Animals↗
PubMed2006

Derivation and characterization of three new Spanish human embryonic stem cell lines (VAL -3 -4 -5) on human feeder and in serum-free conditions.

A total of 184 human embryos, frozen for >5 years, were donated; informed consent was obtained according to Spanish law 45/2003. Survival rate was 40% and three out of 24 blastocysts (12.5%) developed into putative hESC lines, named VAL-3, VAL-4, and VAL-5. The derivation process was performed on microbiologically tested and irradiated human foreskin fibroblasts and designed to minimize contact with xeno-components in knockout DMEM supplemented with knockout serum replacement, and basic fibroblast growth factor. Fingerprinting and HLA typing of the cell lines allowed their identification and traceability. Karyotype was normal for VAL-3 (46XY), VAL-4 (46XX) and VAL-5 (46XX). All three hESC lines expressed specific markers for non-differentiation (Nanog, stage-specific embryonic antigen-4 [SSEA-4], tumour-related antigen [TRA]-1-60, and TRA-1-81) and were negative for SSEA-1. RT-PCR further demonstrated the expression of Oct-4, Sox2, Rex-1, Nanog, Cripto, Thy-1, and Lefty-A. Furthermore, they were found to be negative for classical differentiation markers such as neurofilament heavy chain (ectoderm), renin (mesoderm), and amylase (endoderm). All three cell lines displayed high levels of telomerase activity, and were shown to successfully overcome cryopreservation and thawing. Finally, these three new hESC lines have demonstrated the potential to differentiate in vitro and in vivo (teratoma formation) into cell types originating from all three germ layers.

Animals↗
PubMed2005

First derivation in Spain of human embryonic stem cell lines: use of long-term cryopreserved embryos and animal-free conditions.

The first two human embryonic stem cell lines (VAL-1 and VAL-2) have been derived in Spain with long-term cryopreserved embryos under animal-free conditions. In the first series, 40 human embryos that had been cryopreserved at day 2 of development were thawed after >5 years. A derivation efficiency of 5% per frozen embryo or 12.5% per blastocyst was obtained.

Cryopreservation↗