M Barragán1, J Pons2, A Ferrer-Vaquer1, D Cornet-Bartolomé1, A Schweitzer3, J Hubbard3, H Auer4, A Rodolosse2, R Vassena1. 1. Clínica EUGIN, Travessera de les Corts 322, 08029 Barcelona, Spain. 2. Functional Genomics Core, Institute for Research in Biomedicine (IRB) Barcelona, Parc Científic de Barcelona, Baldiri Reixac 10, 08028 Barcelona, Spain. 3. Thermo Fisher Scientific, 3450 Central Expressway, Santa Clara, CA 95051, USA. 4. Functional GenOmics Consulting, Bellavista 53, 08753 Pallejà, Spain.
Abstract
STUDY QUESTION: How does the human oocyte transcriptome change with age and ovarian reserve? SUMMARY ANSWER: Specific sets of human oocyte messenger RNAs (mRNAs) and non-coding RNAs (ncRNAs) are affected independently by age and ovarian reserve. WHAT IS KNOWN ALREADY: Although it is well established that the ovarian reserve diminishes with increasing age, and that a woman's age is correlated with lower oocyte quality, the interplay of a diminished reserve and age on oocyte developmental competence is not clear. After maturation, oocytes are mostly transcriptionally quiescent, and developmental competence prior to embryonic genome activationrelies on maternal RNA and proteins. STUDY DESIGN, SIZE, DURATION: A total of 36 vitrified/warmed MII oocytes from 30 women undergoing oocyte donation were included in this study, processed and analyzed individually. PARTICIPANTS/MATERIALS, SETTING, METHODS: Total RNA from each oocyte was independently isolated, amplified, labeled, and hybridized on HTA 2.0 arrays (Affymetrix). Data were analyzed using TAC software, in four groups, each including nine oocytes, according to the woman's age and antral follicular count (AFC) (mean ± SD): Young with High AFC (YH; age 21 ± 1 years and 24 ± 3 follicles); Old with High AFC (OH; age 32 ± 2 years and 29 ± 7 follicles); Young with Low AFC (YL; age 24 ± 2 years and 8 ± 2 follicles); Old with Low AFC (OL; age 34 ± 1 years and 7 ± 1 follicles). qPCR was performed to validate arrays. MAIN RESULTS AND THE ROLE OF CHANCE: We identified a set of 30 differentially expressed mRNAs when comparing oocytes from women with different ages and AFC. In addition, 168 non-coding RNAs (ncRNAs) were differentially expressed in relation to age and/or AFC. Few mRNAs have been identified as differentially expressed transcripts, and among ncRNAs, a set of Piwi-interacting RNAs clusters (piRNAs-c) and precursor microRNAs (pre-miRNAs) were identified as increased in high AFC and old groups, respectively. Our results indicate that age and ovarian reserve are associated with specific ncRNA profiles, suggesting that oocyte quality might be mediated by ncRNA pathways. LARGE SCALE DATA: Data can be found via GEO accession number GSE87201. LIMITATIONS, REASONS FOR CAUTION: The oldest woman included in the study was 35 years old, thus our results cannot readily be extrapolated to women older than 35 or infertile women. WIDER IMPLICATIONS OF THE FINDINGS: We show, for the first time, that several non-coding RNAs, usually regulating DNA transcription, are differentially expressed in relation to age and/or ovarian reserve. Interestingly, the mRNA transcriptome of in vivo matured oocytes remains remarkably stable across ages and ovarian reserve, suggesting the possibility that changes in the non-coding transcriptome might regulate some post-transcriptional/translational mechanisms which might, in turn, affect oocyte developmental competence. STUDY FUNDING AND COMPETING INTEREST(S): This work was supported by intramural funding of Clinica EUGIN and by the Secretary for Universities and Research of the Ministry of Economy and Knowledge of the Government of Catalonia. J.H. and A.S. are employees of Affymetrix, otherwise there are no competing interests.
STUDY QUESTION: How does the human oocyte transcriptome change with age and ovarian reserve? SUMMARY ANSWER: Specific sets of human oocyte messenger RNAs (mRNAs) and non-coding RNAs (ncRNAs) are affected independently by age and ovarian reserve. WHAT IS KNOWN ALREADY: Although it is well established that the ovarian reserve diminishes with increasing age, and that a woman's age is correlated with lower oocyte quality, the interplay of a diminished reserve and age on oocyte developmental competence is not clear. After maturation, oocytes are mostly transcriptionally quiescent, and developmental competence prior to embryonic genome activationrelies on maternal RNA and proteins. STUDY DESIGN, SIZE, DURATION: A total of 36 vitrified/warmed MII oocytes from 30 women undergoing oocyte donation were included in this study, processed and analyzed individually. PARTICIPANTS/MATERIALS, SETTING, METHODS: Total RNA from each oocyte was independently isolated, amplified, labeled, and hybridized on HTA 2.0 arrays (Affymetrix). Data were analyzed using TAC software, in four groups, each including nine oocytes, according to the woman's age and antral follicular count (AFC) (mean ± SD): Young with High AFC (YH; age 21 ± 1 years and 24 ± 3 follicles); Old with High AFC (OH; age 32 ± 2 years and 29 ± 7 follicles); Young with Low AFC (YL; age 24 ± 2 years and 8 ± 2 follicles); Old with Low AFC (OL; age 34 ± 1 years and 7 ± 1 follicles). qPCR was performed to validate arrays. MAIN RESULTS AND THE ROLE OF CHANCE: We identified a set of 30 differentially expressed mRNAs when comparing oocytes from women with different ages and AFC. In addition, 168 non-coding RNAs (ncRNAs) were differentially expressed in relation to age and/or AFC. Few mRNAs have been identified as differentially expressed transcripts, and among ncRNAs, a set of Piwi-interacting RNAs clusters (piRNAs-c) and precursor microRNAs (pre-miRNAs) were identified as increased in high AFC and old groups, respectively. Our results indicate that age and ovarian reserve are associated with specific ncRNA profiles, suggesting that oocyte quality might be mediated by ncRNA pathways. LARGE SCALE DATA: Data can be found via GEO accession number GSE87201. LIMITATIONS, REASONS FOR CAUTION: The oldest woman included in the study was 35 years old, thus our results cannot readily be extrapolated to women older than 35 or infertile women. WIDER IMPLICATIONS OF THE FINDINGS: We show, for the first time, that several non-coding RNAs, usually regulating DNA transcription, are differentially expressed in relation to age and/or ovarian reserve. Interestingly, the mRNA transcriptome of in vivo matured oocytes remains remarkably stable across ages and ovarian reserve, suggesting the possibility that changes in the non-coding transcriptome might regulate some post-transcriptional/translational mechanisms which might, in turn, affect oocyte developmental competence. STUDY FUNDING AND COMPETING INTEREST(S): This work was supported by intramural funding of Clinica EUGIN and by the Secretary for Universities and Research of the Ministry of Economy and Knowledge of the Government of Catalonia. J.H. and A.S. are employees of Affymetrix, otherwise there are no competing interests.
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