The purpose of this article is to determine whether implantation of a ‘cleavage stage’(2-3 days) embryo is more likely to result in a live birth or if implantation should occur later when the embryo has reached the ‘blastocysts’ (5-6 days) stage of development. While there have been papers published on the success of ‘blastocysts’ implantation, it is noted that these trials were not randomised nor a consensus about the best practice for blastocyst culture. This paper delves into the results of a clinical randomised trial on blastocysts against cleavage stage. The merits found towards blastocysts are based on 2 arguments. 1. That most successful pregnancies in vivo, the embryo does not reach the uterus at day 3, which is the typical implantation time for cleavage transfer. 2. That blastocysts have higher implantation potential as the embryo have reach the blastocysts stage, have the most potential to survive. It is this second point, the one of self-selection which is seen a negative in the use of blastocysts implantation. This study found that although the live birth rate for the randomised controlled trail for blastocysts was a small but significant difference, the miscarriage rate between cleavage stage and blastocysts remained the same. On a whole this paper found that the benefits between cleavage stage and blastocysts are still unclear.
citing finding from original paper, correct permission for images, no textbook info, research articles not review articles!!
http://www.nei.nih.gov/lca/timeline.asp, "Courtesy: National Eye Institute, National Institutes of Health (NEI/NIH)."
1869 Dr. Theodor Leber (1840-1917), German ophthalmologist, first describes what is now known as Leber congenital amaurosis, an inherited retinal disease that causes severe visual impairment early in childhood
1932-34 George Wald, Ph.D. first identified vitamin A in the retina during a National Research Council fellowship in biology
1965 Human adeno-associated virus (AAV) was discovered.
1967 Dr. George Wald, Harvard University, received The Nobel Prize in Physiology or Medicine for his discovery of the role of vitamin A in the visual process
1984 Drs. Nicolas Muzyczka and Paul Hermonat, published an article demonstrating that adeno-associated virus (AAV) can be used to introduce foreign DNA into human and murine tissue culture cells.
1990 Dr. T. Michael Redmond of the NEI's Laboratory of Retinal Cell and Molecular Biology, Section on Gene Regulation began work on RPE-specific monoclonal antibody.
1993 Dr. T. Michael Redmond of the NEI's Laboratory of Retinal Cell and Molecular Biology, Section on Gene Regulation cloned RPE65, a protein necessary for processing vitamin A in the visual cycle
1997 RPE65 gene mutations identified as the cause of congenital blindness in some children with Leber congenital amaurosis
1997 The knockout mouse model was created by Dr. T. Michael Redmond's team in the NEI's Laboratory of Retinal Cell and Molecular Biology, Section on Gene Regulation to study RPE65.
Dr. T. Michael Redmond talks about the knock-out mouse model:
When the knock-out mouse was first developed, we were able to understand in very great extent, to a very sophisticated extent, the biochemistry of the disorder, and very shortly in the heels of the discovery of the development of the mouse model, a dog model was discovered, and this was a previously existing blind dog which was established to be an RPE65 mutation, based on the finding of human mutations and also based on the mouse, the development of the mouse knock-out, and within very short order, we had two animal models for this disease. The mouse gave us a terrific understanding of the biochemistry and physiology.
1998 Dr. T. Michael Redmond's team in the NEI's Laboratory of Retinal Cell and Molecular Biology, Section on Gene Regulation use the knockout mouse model to establish RPE65’s role in vitamin A metabolism
1998 RPE65 gene mutation was discovered as causing congenital blindness in Briard dogs
2001 NEI-supported researchers at the University of Pennsylvania and University of Florida restore vision in a Briard dog using gene transfer RPE65 therapy
2001 NEI/NIH expands support for pre-clinical studies of RPE65 gene therapy development
2005 Restored vision in Briard dog persists longer than four years following RPE65 gene transfer
2007 NEI-supported RPE65 human clinical trial began. This open label, single dose, dose escalation study is designed to assess the safety of using a modified adeno-associated viral vector (rAAV2-hRPE65) to deliver the normal RPE65 gene to the retina.
Principal Investigator: Dr. Samuel Jacobson, University of Pennsylvania Principal Investigator: Dr. Barry Byrne, University of Florida Gene Vector Specialist: Dr. William Hauswirth, University of Florida Surgeon: Dr. Shalesh Kaushal, University of Florida Dr. Shalesh Kaushal explains the cause of Leber congenital amaurosis (LCA):
LCA as I was mentioning is an inherited retinal degeneration but it's unusual it's severe it affects children early on. Most retinal degenerations manifest themselves a little bit later in life, many times in the teens, 20s and 30s. And I think that's one of the defining characteristics of LCA although there are other inherited severe forms of retinal degeneration that are similar to LCA. There are many different or at least a set of genes that have been identified to cause LCA. The most common is the RPE65 gene. This is a gene that produces a protein that helps process vitamin A in the cells that nourish the retina. Those cells are called the retinal pigment epithelial cells or RPE cells for short. And we know in these patients because of these mutations it doesn't produce enough of vitamin A, which is important for allowing the visual proteins that detect light to sense light.
2008 Initial results from two clinical trials conducted by the Children’s Hospital of Philadelphia (CHOP) and the University College of London published in the April 27 online version of New England Journal of Medicine.
2008 First cohort of the NEI-supported RPE65 human clinical trial completed. (Three patients treated.) Promising 3-month results published in Proceeding of the National Academy of Sciences (September 22) and Human Gene Therapy (September 7).
Dr. Samuel Jacobson shares what patients report seeing after gene therapy:
So what do they notice, well, generally speaking within a week, they will tell you that they see things brighter. They will tell you that they see things. Some are more specific observers and others are less specific. Some will tell you where exactly where they see it brighter. One person calls it a headlight. The headlight got turned on on day five and it just hasn't turned off. That's a really interesting headlight. I mean it's not a headlight that's there without vision, stimulation but there's a view of the world through that headlight that wasn't there before.
2009 One-year results from the NEI-supported RPE65 human clinical trial published August online in Human Gene Therapy and in an August 13 letter to the editor in the New England Journal of Medicine. All three patients remained healthy and maintained previous visual gains. One patient also noticed a visual improvement that helped her perform daily tasks.
How RPE65 gene transfer therapy works:
In people with Leber congenital amaurosis, the RPE65 protein, critical for visual function, is not made due to mutations in the corresponding RPE65 gene. Fixing the problem requires isolating the segment of DNA corresponding to the normal RPE65 gene and packaging it into a vector made from a virus. The gene–containing vector is injected into the eyeball under the retina, between the retinal pigment epithelium and choroid where it enters the cells of the retina. The vector releases the normal RPE65 gene segment. The normal RPE65 protein is now manufactured by the cell. The RPE65 protein is released from the cell to vision receptors, restoring normal visual function.
2009 Results from early LCA clinical studies represent one of the first steps toward the use of gene transfer therapy for an inherited for of blindness. Researchers are hopeful that similar techniques will be applied to other genetic disease affecting vision.
Dr. Redmond discusses the significance of the trial:
The LCA gene therapy trial is highly significant to the field of vision in that it was proof of principle that genetic blindness could be reversed. It provides a template, for future work in gene therapy, not only of the eye, but also of other systems in the body. In particular, it allows one to think of more complex and more difficult avenues to go down.