Tamara Garrido-Gómez1, Alicia Quiñonero2, Oreto Antúnez3, Patricia Díaz-Gimeno2, Jose Bellver2, Carlos Simón4, Francisco Domínguez5. 1. Fundación Instituto Valenciano de Infertilidad (FIVI), Instituto Universitario IVI (IUIVI), Valencia, Spain INCLIVA Biomedical Research Institute, Valencia, Spain. 2. Fundación Instituto Valenciano de Infertilidad (FIVI), Instituto Universitario IVI (IUIVI), Valencia, Spain. 3. Proteomics Unit, Valencia University, Valencia, Spain. 4. Fundación Instituto Valenciano de Infertilidad (FIVI), Instituto Universitario IVI (IUIVI), Valencia, Spain Department of Pediatrics, Obstetrics and Gynecology, Valencia University, INCLIVA, Valencia, Spain Department of Obstetrics and Gynecology, Stanford University, Stanford, CA, USA. 5. Fundación Instituto Valenciano de Infertilidad (FIVI), Instituto Universitario IVI (IUIVI), Valencia, Spain INCLIVA Biomedical Research Institute, Valencia, Spain francisco.dominguez@ivi.es.
Abstract
STUDY QUESTION: Are there any proteomic differences between receptive (R) and non-receptive (NR) endometrial receptivity array (ERA)-diagnosed endometria obtained on the same day of a hormonal replacement therapy (HRT) treatment cycle? SUMMARY ANSWER: There is a different proteomic signature between R and NR ERA-diagnosed endometrium obtained on the same day of HRT cycles. WHAT IS KNOWN ALREADY: The human endometrial transcriptome has been extensively investigated in the last decade resulting in the development of a new diagnostic test based on the transcriptomic signature of the window of implantation (WOI). Much less is known about the proteomics derived from the transcripts present during the WOI. STUDY DESIGN, SIZE, AND DURATION: This study was a basic proteomic analysis of human endometrial biopsies taken from twelve IVF patients. PARTICIPANTS/MATERIALS, SETTING, AND METHODS: Human endometrial biopsies were collected during HRT cycles after 5 days of progesterone (P) administration, and diagnosed as receptive (R; n = 6) or non-receptive (NR; n = 6) by the ERA test. Endometrial proteins were extracted, labelled and separated using differential in-gel electrophoresis (DIGE). Proteins were identified using mass spectrometry, followed up by in silico analysis. Validation studies using western blots and immunolocalization were performed for the progesterone receptor membrane component 1 (PGRMC1) and annexin A6 (ANXA6) proteins. MAIN RESULTS AND THE ROLE OF CHANCE: DIGE analysis followed by protein identification by MALDI-MS and database searches revealed 24 differentially expressed proteins in R versus NR samples. In silico analysis showed two pathways which were significantly different between R and NR samples. Expression of PGRMC1 and ANXA6 was validated and localized by western blots and immunohistochemistry. These results highlight these proteins as key targets likely to be important in the comprehension of human endometrial receptivity. LIMITATIONS, REASONS FOR CAUTION: This was mainly a descriptive study with no functional studies on the proteins found. We also used a low number of human endometrial samples for the DIGE analysis. WIDER IMPLICATIONS OF THE FINDINGS: This study identified the proteomic profile associated with receptive or non-receptive human endometria. Our findings suggest that although histological dating indicates a putative 'receptive' status within the WOI, a different transcriptomic and proteomic profile is observed in these samples. We should move towards using more personalized WOIs, where identification of the correct endometrial receptivity status, and consequently the success of IVF, relies on individual molecular signatures rather than traditional endometrial dating. STUDY FUNDING/COMPETING INTERESTS: F.D.'s participation in this work was supported by the Spanish Ministry of Economy and Competitiveness, through the Miguel Servet Programme (CP13/00075) co-founded by FEDER. The project was also supported by a grant from the Spanish Ministry of Economy and Competitiveness, through the FIS Programme (PI12/00450). The authors have no financial/commercial conflicts of interest to declare.
STUDY QUESTION: Are there any proteomic differences between receptive (R) and non-receptive (NR) endometrial receptivity array (ERA)-diagnosed endometria obtained on the same day of a hormonal replacement therapy (HRT) treatment cycle? SUMMARY ANSWER: There is a different proteomic signature between R and NR ERA-diagnosed endometrium obtained on the same day of HRT cycles. WHAT IS KNOWN ALREADY: The human endometrial transcriptome has been extensively investigated in the last decade resulting in the development of a new diagnostic test based on the transcriptomic signature of the window of implantation (WOI). Much less is known about the proteomics derived from the transcripts present during the WOI. STUDY DESIGN, SIZE, AND DURATION: This study was a basic proteomic analysis of human endometrial biopsies taken from twelve IVFpatients. PARTICIPANTS/MATERIALS, SETTING, AND METHODS: Human endometrial biopsies were collected during HRT cycles after 5 days of progesterone (P) administration, and diagnosed as receptive (R; n = 6) or non-receptive (NR; n = 6) by the ERA test. Endometrial proteins were extracted, labelled and separated using differential in-gel electrophoresis (DIGE). Proteins were identified using mass spectrometry, followed up by in silico analysis. Validation studies using western blots and immunolocalization were performed for the progesterone receptor membrane component 1 (PGRMC1) and annexin A6 (ANXA6) proteins. MAIN RESULTS AND THE ROLE OF CHANCE: DIGE analysis followed by protein identification by MALDI-MS and database searches revealed 24 differentially expressed proteins in R versus NR samples. In silico analysis showed two pathways which were significantly different between R and NR samples. Expression of PGRMC1 and ANXA6 was validated and localized by western blots and immunohistochemistry. These results highlight these proteins as key targets likely to be important in the comprehension of human endometrial receptivity. LIMITATIONS, REASONS FOR CAUTION: This was mainly a descriptive study with no functional studies on the proteins found. We also used a low number of human endometrial samples for the DIGE analysis. WIDER IMPLICATIONS OF THE FINDINGS: This study identified the proteomic profile associated with receptive or non-receptive human endometria. Our findings suggest that although histological dating indicates a putative 'receptive' status within the WOI, a different transcriptomic and proteomic profile is observed in these samples. We should move towards using more personalized WOIs, where identification of the correct endometrial receptivity status, and consequently the success of IVF, relies on individual molecular signatures rather than traditional endometrial dating. STUDY FUNDING/COMPETING INTERESTS: F.D.'s participation in this work was supported by the Spanish Ministry of Economy and Competitiveness, through the Miguel Servet Programme (CP13/00075) co-founded by FEDER. The project was also supported by a grant from the Spanish Ministry of Economy and Competitiveness, through the FIS Programme (PI12/00450). The authors have no financial/commercial conflicts of interest to declare.
Authors: Melissa L McCallum; Cindy A Pru; Yuichi Niikura; Siu-Pok Yee; John P Lydon; John J Peluso; James K Pru Journal: Endocrinology Date: 2016-06-16 Impact factor: 4.736
Authors: Julian A Gingold; Joseph A Lee; Jorge Rodriguez-Purata; Michael C Whitehouse; Benjamin Sandler; Lawrence Grunfeld; Tanmoy Mukherjee; Alan B Copperman Journal: Fertil Steril Date: 2015-06-13 Impact factor: 7.329
Authors: Ioanna Kosteria; Athanasios K Anagnostopoulos; Christina Kanaka-Gantenbein; George P Chrousos; George T Tsangaris Journal: In Vivo Date: 2017 May-Jun Impact factor: 2.155
Authors: Dunja Maria Baston-Buest; Olga Altergot-Ahmad; Sarah Jean Pour; Jan-Steffen Krüssel; Udo Rudolf Markert; Tanja Natascha Fehm; Alexandra Petra Bielfeld Journal: Mediators Inflamm Date: 2017-02-15 Impact factor: 4.711