High-throughput sperm differential proteomics suggests that epigenetic alterations contribute to failed assisted reproduction  

Oleh:

Azpiazu, Ruben ; Amaral, Alexandra ; Castillo, Judit ; Estanyol, Josep Maria ; Guimera, Marta

Jenis: Article from Journal - ilmiah internasional
Dalam koleksi: Human Reproduction vol. 29 no. 06 (Jun. 2014), page 1225-1237.
Topik: human sperm; differential proteomics; isotopic; tandem mass tags; epigenetics; assisted reproduction

Ketersediaan

Perpustakaan FK Nomor Panggil: H07.K.2014.02 Non-tandon: 1 (dapat dipinjam: 0) Tandon: tidak ada

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STUDY QUESTION Are there quantitative alterations in the proteome of normozoospermic sperm samples that are able to complete IVF but whose female partner does not achieve pregnancy? SUMMARY ANSWER Normozoospermic sperm samples with different IVF outcomes (pregnancy versus no pregnancy) differed in the levels of at least 66 proteins. WHAT IS KNOWN ALREADY The analysis of the proteome of sperm samples with distinct fertilization capacity using low-throughput proteomic techniques resulted in the detection of a few differential proteins. Current high-throughput mass spectrometry approaches allow the identification and quantification of a substantially higher number of proteins. STUDY DESIGN, SIZE, DURATION This was a case–control study including 31 men with normozoospermic sperm and their partners who underwent IVF with successful fertilization recruited between 2007 and 2008. PARTICIPANTS/MATERIALS, SETTING, METHODS Normozoospermic sperm samples from 15 men whose female partners did not achieve pregnancy after IVF (no pregnancy) and 16 men from couples that did achieve pregnancy after IVF (pregnancy) were included in this study. To perform the differential proteomic experiments, 10 no pregnancy samples and 10 pregnancy samples were separately pooled and subsequently used for tandem mass tags (TMT) protein labelling, sodium dodecyl sulphate–polyacrylamide gel electrophoresis, liquid chromatography tandem mass spectrometry (LC-MS/MS) identification and peak intensity relative protein quantification. Bioinformatic analyses were performed using UniProt Knowledgebase, DAVID and Reactome. Individual samples (n = 5 no pregnancy samples; n = 6 pregnancy samples) and aliquots from the above TMT pools were used for western blotting. MAIN RESULTS AND THE ROLE OF CHANCE By using TMT labelling and LC-MS/MS, we have detected 31 proteins present at lower abundance (ratio no pregnancy/pregnancy < 0.67) and 35 at higher abundance (ratio no pregnancy/pregnancy > 1.5) in the no pregnancy group. Bioinformatic analyses showed that the proteins with differing abundance are involved in chromatin assembly and lipoprotein metabolism (P values < 0.05). In addition, the differential abundance of one of the proteins (SRSF protein kinase 1) was further validated by western blotting using independent samples (P value < 0.01). LIMITATIONS, REASONS FOR CAUTION For individual samples the amount of recovered sperm not used for IVF was low and in most of the cases insufficient for MS analysis, therefore pools of samples had to be used to this end. WIDER IMPLICATIONS OF THE FINDINGS Alterations in the proteins involved in chromatin assembly and metabolism may result in epigenetic errors during spermatogenesis, leading to inaccurate sperm epigenetic signatures, which could ultimately prevent embryonic development. These sperm proteins may thus possibly have clinical relevance.

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