Biological foundation for epigenetic investigations of ART derived human oocytes and embryos

Lead Research Organisation: University of Leeds
Department Name: Lds Inst Genetics Health & Therapeutics

Abstract

A growing body of evidence suggests that some of the technologies used to treat infertility in humans and/or the infertility itself may compromise the function of key genes which are associated with imprinting. The imprint is a unique mark found on a small number of genes. The imprint is established during the growth of eggs and sperm and it is vital for the development of healthy embryos before they implant to produce a pregnancy. Importantly, the imprinted mark can, in some cases, be changed erroneously during the laboratory procedures used to treat infertility. Errors in how the gene imprints are established and read may predispose an individual to diseases such as cancer and/or specific disorders associated with imprinted gene disruption including rare mental retardation syndromes such as Angelman syndrome, and Beckwith-Wiedemann syndrome. This is a major cause of concern as over a million babies have been born worldwide following assisted conception. Furthermore, it is possible that the more invasive assisted reproductive technologies (ART) which are increasingly being developed and used to treat infertile patients may predispose human embryos and the children born subsequently to imprinting errors. It is therefore of paramount importance to understand how imprinted genes work in healthy human eggs, sperm and embryos and to investigate how their programming can go wrong. This project will predominantly use highly sensitive and sophisticated molecular biology tools called microarrays and follow-up analyses to simultaneously study multiple imprinted genes and their regulators throughout the development of human eggs and embryos. The information generated by these unique studies will be used to confirm the safety of a new treatment for the in vitro growth of eggs- a technique which involves an extended period of culture in the laboratory during which time the eggs are ripened before they are fertilised. It is anticipated that status of the imprinted genes and their regulators can be used to test the safety of new ARTs, such as the in vitro maturation of oocytes, before they are introduced into routine clinical practise. The proposed work will therefore help us to understand and hopefully prevent imprinting errors and diseases being passed onto the next generation of children born following assisted conception.

Technical Summary

Imprinted genes are expressed predominantly or exclusively from one parental allele. This mode of regulation is uniquely sensitive to disruption, and therefore imprinted genes themselves are cellular biomarkers of epigenetic disruption, cellular stress and cancer. In assisted reproductive technology (ART) in humans, gametes and embryos are manipulated in vitro at the exact time when the complex and delicate processes of epigenetic reprogramming occur. Recent evidence suggests a range of epigenetic mutations leading to diseases such as Angelman syndrome and Beckwith-Wiedemann syndrome have arisen in children born following assisted conception. Although some of these of imprinting defects may be caused by infertility itself, experimental evidence also indicates that ARTs can disrupt epigenetic mechanisms and alter the regulation of imprinted genes. The need to directly quantify the epigenetic consequences of ART and infertility on the health of the human preimplantation embryos is therefore becoming increasingly important. Due to the limited availability of human gametes and embryos for research and the requirement for analytical methods which are sensitive at the single oocyte/embryo level, the epigenetic processes that regulate early human development remain largely unknown. Despite these limitations we established a programme to investigate early epigenetic processes in humans and have developed appropriate tools and techniques to permit studies of oogenesis and embryogenesis. This project therefore aims to conduct a robust series of investigations of the imprinted genes in human oocytes and preimplantation embryos and to relate these observations to the epigenetic health of embryos derived following ART. This application will test the hypothesis that disruption of imprinting attributable to in vitro manipulations of human gametes and embryos will result in aberrant expression of imprinted genes in the preimplantation embryos so produced. Specifically this application will: 1) establish whether the imprinted genes and epigenetic regulators serve as biomarkers of epigenetic disruption in human blastocysts derived by conventional ART including IVF, ICSI and the in vitro maturation (IVM) of oocytes; 2) monitor allelic expression of imprinted gene transcripts in blastocysts to determine whether expression is monoallelic or otherwise; 3) use bisulphite genomic sequencing to analyse methylation imprints in human ART-derived oocytes and embryos; and 4) conduct a detailed analysis of imprinted gene expression and methylation in a cohort of IVM blastocysts. The data generated will significantly advance our knowledge of the mechanism(s) behind ART-induced imprinting defects in human oocytes and embryos and will help us understand how to alleviate these problems in the future.

Publications

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Huntriss J (2018) Epigenetics and Reproductive Medicine: Scientific Impact Paper No. 57. in BJOG : an international journal of obstetrics and gynaecology

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Huntriss J (2015) Analysis of DNA Methylation Patterns in Single Blastocysts by Pyrosequencing®. in Methods in molecular biology (Clifton, N.J.)

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Huntriss JD (2013) Variable imprinting of the MEST gene in human preimplantation embryos. in European journal of human genetics : EJHG

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Ntostis P (2017) Potential sperm contributions to the murine zygote predicted by analysis. in Reproduction (Cambridge, England)

 
Description The findings of this project have been utilized in several ways including: (i) to advance scientific knowledge and understanding as evidenced by multiple publications in the form of original, peer reviewed research papers, research reviews and book chapters; (ii) to prime future research grant applications and studentships; (iii) for the pubic engagement of science as evidenced by lay publications/magazine articles and talks to patient groups, politicians/policy makers and schools; (iv) to inform research led teaching of undergraduate and postgraduate students; (v) to improve patient treatments; and (vi) to support the career progression of both early career researchers and established scientists.
First Year Of Impact 2010
Sector Healthcare,Pharmaceuticals and Medical Biotechnology
Impact Types Societal

 
Description MRC Ph.D studentship
Amount £50,000 (GBP)
Organisation Medical Research Council (MRC) 
Sector Public
Country United Kingdom
Start 01/2010 
End 12/2013
 
Description Testing the health and therapeutic potential of in vitro derived oocytes for the restoration of female fertility and the treatment of infertility- MRC Biomedical Catalyst Developmental Pathway Funding Scheme
Amount £1,474,143 (GBP)
Funding ID MR/T025654/1 
Organisation Medical Research Council (MRC) 
Sector Public
Country United Kingdom
Start 04/2020 
End 09/2023
 
Description The Safety and Efficacy Of Microfluidic Embryo Culture As A Means To Improve The Efficiency Of Human Infertility Treatment, MRC Confidence In Concept Grant
Amount £79,404 (GBP)
Organisation University of Leeds 
Sector Academic/University
Country United Kingdom
Start 04/2020 
End 03/2021
 
Description Pyrosequencing collaboration with Dr A Murrell 
Organisation Cancer Research UK Cambridge Institute
Country United Kingdom 
Sector Academic/University 
PI Contribution Preparation and provision of samples for analysis and the conduct of the pyrosequencing assays in the Cambridge laboratory
Collaborator Contribution This collaboration has facilitated the application and validation of pyrosequencing as a means to analyse gene methylation status in individual embryos
Impact Research publication - Huntriss et al 2011, Quantitative analysis of DNA methylation of imprinted genes in single human blastocysts by pyrosequencing. Fertil Steril. 30;95(8):2564-7 PMID:21575943
Start Year 2010
 
Description Conference presentation 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Invited symposium presentation at the 1st world congress of artificial gametogenesis. Presentation entitled "Advances in the in vitro growth and maturation of oocytes" presentation generated significant debate and discussion and follow-up meetings about research strategy
Year(s) Of Engagement Activity 2017
 
Description Fertility conference poster presentation 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Poster presentation entitled "A consistent set of imprinted gene transcripts are expressed in human blastocysts: Preliminary evidence for an imprinted gene network operating in human preimplantation development". Poster sparked questions and discussion.
Year(s) Of Engagement Activity 2017
URL http://fertilityconference.org
 
Description Higher eduction college visit (York), student interest and careers 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact Lecture to Human Biology students at York College. Assisted Reproductive Technologies: Present and Future Uses- talk sparked questions and discussion.

College requested similar talk for students in future years
Year(s) Of Engagement Activity 2014
 
Description Invited Seminar at Hull York Medical School 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Participants in your research and patient groups
Results and Impact Hull York Medical School invited talk on -Assisted Reproductive Technologies and their Effects on Epigenetic Regulation in Early Mammalian Development, generated questions and discussion

Research collaboration
Year(s) Of Engagement Activity 2014
 
Description Invited lecture 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Postgraduate students
Results and Impact 25 MSc student attended invited lecture which sparked questions and discussion afterwards
Year(s) Of Engagement Activity 2016
 
Description Invited lecture 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Postgraduate students
Results and Impact Invited lecture on "Epigenetic Programming in Gametes" 1 for Module MED6111 on Gametes for the MSc in Reproductive and Developmental Medicine, University of Sheffield
Year(s) Of Engagement Activity 2015
 
Description Invited lecture 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact Invited lecture for annual scientific meeting of the sub-specialist trainees in reproductive medicine and surgery which sparked questions and discussions afterwards.
Year(s) Of Engagement Activity 2016
 
Description Invited plenary paper 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Professional Practitioners
Results and Impact Symposium paper entitled "Understanding the Epigenetic Effects of ART on the Human Preimplantation Embryo" delivered at the annual conference of the Association of Clinical Embryologists . This presentation sparked questions and discussion afterwards.
Year(s) Of Engagement Activity 2016
 
Description Lecture to Biology students at Ampleforth College, N.Yorks 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact Lecture to Biology students at Ampleforth College, N.Yorks. Talk on Assisted Reproductive Technologies: Present and Future Uses

Talk generated interest amongst biology students
Year(s) Of Engagement Activity 2014
 
Description Lecture to MSc students at the University of Sheffield 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact Lecture to students on MSc in Reproductive and Developmental Medicine. University of Sheffield . Lecture entitled- Ovarian tissue cryopreservation and fertility preservation

n/a
Year(s) Of Engagement Activity 2013
 
Description Lecture to students at the University of Sheffield 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact Lecture to students on MSc in Reproductive and Developmental Medicine. University of Sheffield . Lecture entitled: Epigenetic Reprogramming in the Mammalian Germline.

n/a
Year(s) Of Engagement Activity 2014
 
Description Leeds University Horizons Programme 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? Yes
Type Of Presentation Workshop Facilitator
Geographic Reach Local
Primary Audience Public/other audiences
Results and Impact 12 members of the public from different backgrounds attended a series of 6 short talks and a practical workshop.

One member of the attending group who was a bank manager subsequently registered with the Open University to register for a Bsc in Biological Science
Year(s) Of Engagement Activity 2007
 
Description Plenary conference presentation 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Professional Practitioners
Results and Impact Invited plenary presentation at the "The Summit on Organ Banking through Converging Technologies" Presentation entitled "XXX "sparked numerous questions and follow-up discussions with a number of interested parties including professionals working in the field and the media. The summit was attended by 225 participants with interests in transplantation, biopreservation, tissue engineering, tissue banking, and philanthropy.
Year(s) Of Engagement Activity 2017
URL https://obs2017.dryfta.com/en/
 
Description Progress Education Trust 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? Yes
Type Of Presentation Keynote/Invited Speaker
Geographic Reach National
Primary Audience Public/other audiences
Results and Impact Predominantly lay audience of ~ 100-150 people including media, undergraduate students and policy makers with some academics attended a days conference of multiple short talks and discussions
PET 2011 annual conference 23rd Nov 2011, London : www.progress.org.uk/conference2011
Talk by Prof H Picton, Title "Measuring metabolism: a new tool for our trade?"

Received excellent audience feedback from PET participant survey which stated that the talk was very accessible and was well received and the take home message was understood . This claim is substantiated by several media reports on media websites
Year(s) Of Engagement Activity 2011
 
Description Research profile in UK Science and Technology Publication 
Form Of Engagement Activity A magazine, newsletter or online publication
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Media (as a channel to the public)
Results and Impact Provision of a profile research article published in:
Public Service Review: UK Science and Technology 04. pp156-7
This was published both in hard copy and online
websites: www.publicservice.co.uk/
www.publicservice.co.uk/pub_selectissue.asp?publication=UK%edition.pagesuite-professional.co.uk/Launch.aspx?EID=55ff6b04-1e0d-4d3a-91ad-11f2bd7ea09420Science%20and%20Technologyhttp://edition.pagesuite-professional.co.uk/Launch.aspx?EID=55ff6b04-1e0d-4d3a-91ad-11f2bd7ea094
http://edition.pagesuite-professional.co.uk/Launch.aspx?EID=55ff6b04-1e0d-4d3a-91ad-11f2bd7ea094


This magazine (published in hard copy and online) is disseminated to the public, and parliament and national and international policy makers, international funding organisations and industry. This research profile was selected by Public Sector UK Science and Technology as amongst the top 10 research profiles from 2011 and invited for follow-up editorial in January 2013 and for inclusion as key research profile in the delegate packs for the SciTech Europe Conference, Brussels 23rd Nov 2012
Year(s) Of Engagement Activity 2011
 
Description School pupil work placements 
Form Of Engagement Activity Participation in an activity, workshop or similar
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Schools
Results and Impact An A level pupil spent a 3 week work placement in our research laboratories and observed research undertaken for MRC funded research and also received some laboratory-based research training.

This experience assisted the pupil with her career choices and helped with her interviews for a University place to study Medicine.
Year(s) Of Engagement Activity 2011
 
Description Work experience placements for undergraduate students 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach Regional
Primary Audience Undergraduate students
Results and Impact This activity has provided 48 week laboratory work placements in the form of honorary, trainee research technician posts for 10 undergraduate science students from Huddersfield University

The direct experience of laboratory work, research, and laboratory management not only improved the skill base f the students, but it has also supported their attainment of improved degree marks and has assisted them in their quest for jobs in a laboratory environment following graduation from their degree. Some of the placement students have now taken up Ph.D studentships in other Universities
Year(s) Of Engagement Activity 2009,2010,2011,2012