Digitum Comunidad: E0A1
http://hdl.handle.net/10201/1183
E0A1
2024-03-28T23:12:58Z
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Metabolite Profiling of Pig Seminal Plasma Identifies Potential Biomarkers for Sperm Resilience to Liquid Preservation
http://hdl.handle.net/10201/140144
Título: Metabolite Profiling of Pig Seminal Plasma Identifies Potential Biomarkers for Sperm Resilience to Liquid Preservation
Autor/es principal/es: Mateo-Otero, Yentel; Fernández-López, Pol; Ribas-Maynou, Jordi; Roca, Jordi; Miró, Jordi; Yeste, Marc; Barranco, Isabel
Resumen: Metabolomic approaches allow the study of downstream gene expression events since metabolites are considered as the products of cell signaling pathways. For this reason, many studies in humans have already been conducted to determine the influence of the metabolites present in seminal plasma (SP) on sperm physiology, and to identify putative biomarkers. However, in livestock species, these relationships are yet to be uncovered. Thus, the present study aimed to explore: (i) if concentrations of metabolites in pig SP are related to sperm quality and functionality, and (ii) if they could predict the sperm resilience to liquid storage at 17°C. To this end, 28 ejaculates were individually collected and split into three aliquots: one was used for SP analysis through nuclear magnetic resonance (NMR) spectroscopy; another served for the evaluation of sperm concentration and morphology; and the last one was utilized to determine sperm functionality parameters using computer-assisted sperm analysis (CASA) and flow cytometry after 0 h and 72 h of liquid-storage at 17°C. NMR analysis allowed the identification and quantification of 23 metabolites present in pig SP which, except for fumarate, were not observed to follow a breed-dependent behavior. Moreover, specific relationships between metabolites and sperm variables were identified: (i) glutamate, methanol, trimethylamine N-oxide, carnitine, and isoleucine were seen to be related to some sperm quality and functionality parameters evaluated immediately after semen collection; (ii) leucine, hypotaurine, carnitine and isoleucine were found to be associated to the sperm ability to withstand liquid storage; and (iii) Bayesian multiple regression models allowed the identification of metabolite patterns for specific sperm parameters at both 0 h and 72 h. The identification of these relationships opens up the possibility of further investigating these metabolites as potential sperm functional biomarkers.
Descripción: © 2021. The authors. This document is made available under the CC-BY 4.0 license http://creativecommons.org/licenses/by /4.0/
This document is the published version of a published work that appeared in final form in
Frontiers in Cell and Developmental Biology
To access the final work, see DOI: https://doi.org/10.3389/fcell.2021.669974
2021-05-28T00:00:00Z
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The Proteome of Large or Small Extracellular Vesicles in Pig Seminal Plasma Differs, Defining Sources and Biological Functions
http://hdl.handle.net/10201/140113
Título: The Proteome of Large or Small Extracellular Vesicles in Pig Seminal Plasma Differs, Defining Sources and Biological Functions
Autor/es principal/es: Barranco, Isabel; Sánchez-López, Christian M; Bucci, Diego; Alvarez-Barrientos, Alberto; Rodriguez-Martinez, Heriberto; Marcilla, Antonio; Roca, Jordi
Resumen: Seminal plasma contains many morphologically heterogeneous extracellular vesicles (sEVs). These are sequentially released by cells of the testis, epididymis, and accessory sex glands and involved in male and female reproductive processes. This study aimed to define in depth sEV subsets isolated by ultrafiltration and size exclusion chromatography, decode their proteomic profiles using liquid chromatography-tandem mass spectrometry, and quantify identified proteins using sequential window acquisition of all theoretical mass spectra. The sEV subsets were defined as large (L-EVs) or small (S-EVs) by their protein concentration, morphology, size distribution, and EV-specific protein markers and purity. Liquid chromatography-tandem mass spectrometry identified a total of 1034 proteins, 737 of them quantified by SWATH in S-EVs, L-EVs, and non-EVs-enriched samples (18-20 size exclusion chromatography-eluted fractions). The differential expression analysis revealed 197 differentially abundant proteins between both EV subsets, S-EVs and L-EVs, and 37 and 199 between S-EVs and L-EVs versus non-EVs-enriched samples, respectively. The gene ontology enrichment analysis of differentially abundant proteins suggested, based on the type of protein detected, that S-EVs could be mainly released through an apocrine blebbing pathway and be involved in modulating the immune environment of the female reproductive tract as well as during sperm-oocyte interaction. In contrast, L-EVs could be released by fusion of multivesicular bodies with the plasma membrane becoming involved in sperm physiological processes, such as capacitation and avoidance of oxidative stress. In conclusion, this study provides a procedure capable of isolating subsets of EVs from pig seminal plasma with a high degree of purity and shows differences in the proteomic profile between EV subsets, indicating different sources and biological functions for the sEVs.
Descripción: © 2023. The authors. This document is made available under the CC-BY 4.0 license http://creativecommons.org/licenses/by /4.0/
This document is the published version of a published work that appeared in final form in Molecular & Cellular Proteomics (MCP)
To access the final work, see DOI: https://doi.org/10.1016/j.mcpro.2023.100514
2023-04-01T00:00:00Z
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Immunophenotype profile by flow cytometry reveals different subtypes of extracellular vesicles in porcine seminal plasma
http://hdl.handle.net/10201/140143
Título: Immunophenotype profile by flow cytometry reveals different subtypes of extracellular vesicles in porcine seminal plasma
Autor/es principal/es: Barranco, Isabel; Alvarez-Barrientos, Alberto; Parra, Ana; Martinez-Diaz, Pablo; Lucas, Xiomara; Roca, Jordi
Resumen: Background: Porcine seminal plasma (SP) is endowed with a heterogeneous population of extracellular vesicles (sEVs). This study evaluated the immunophenotypic profile by high-sensitivity flow cytometry of eight sEV subpopulations isolated according to their size (small [S-sEVs] and large [L-sEVs]) from four different SP sources, namely three ejaculate fractions (the first 10 mL of the sperm rich fraction [SRF-P1], the remaining SRF [SRF-P2], and the post-SRF [PSRF]) and entire ejaculate (EE).
Methods: Seminal EVs were isolated using a size exclusion chromatography-based protocol from six SP pools (five ejaculates/pool) of each SP source and characterized using complementary approaches including total protein (BCA™assay), particle size distribution (dynamic light scattering), morphology (transmission electron microscopy), and purity (albumin by Western blot). Expression of CD9, CD63, CD81, CD44 and HSP90β was analyzed in all sEV subpopulations by high-sensitivity flow cytometry according to MIFlowCyt-EV guidelines, including an accurate calibration, controls, and discrimination by CFSE-labelling.
Results: Each sEV subpopulation exhibited a specific immunophenotypic profile. The percentage of sEVs positive for CD9, CD63, CD81 and HSP90β differed between S- and L-sEVs (P < 0.0001). Specifically, the percentage of sEVs positive for CD9 and CD63 was higher and that for CD81 was lower in S- than L-sEVs in the four SP sources. However, the percentage of HSP90β-positive sEVs was lower in S-sEVs than L-sEVs in the SRF-P1 and EE samples. The percentage of sEVs positive for CD9, CD63, and CD44 also differed among the four SP sources (P < 0.0001), being highest in PSRF samples. Notably, virtually all sEV subpopulations expressed CD44 (range: 88.04-98.50%).
Conclusions: This study demonstrated the utility of high-sensitivity flow cytometry for sEV immunophenotyping, allowing the identification of distinct sEV subpopulations that may have different cellular origin, cargo, functions, and target cells.
Descripción: © 2024. The authors. This document is made available under the CC-BY 4.0 license http://creativecommons.org/licenses/by /4.0/
This document is the published version of a published work that appeared in final form in Cell Communication and Signaling (CCS).
To access the final work, see DOI: https://doi.org/10.1186/s12964-024-01485-1
2024-01-23T00:00:00Z
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Untargeted metabolomic profiling of serum in dogs with hypothyroidism
http://hdl.handle.net/10201/139225
Título: Untargeted metabolomic profiling of serum in dogs with hypothyroidism
Autor/es principal/es: Muñoz-Prieto, Alberto; González-Aróstegui, Luis; Rubic, Ivana; Cerón, José Joaquín; Tvarijonavicite, Asta; Horvatic, Anita; Mrljak, Vladimir
Resumen: Hypothyroidism is one of the most commonly diagnosed endocrine disease in dogs. The clinical signs are caused
by a deficiency of the active thyroid hormones triiodothyronine (T3) and thyroxine (T4) and have a negative
impact on dog’s quality of life. We hypothesized that serum metabolic profile varies between healthy dogs and
dogs with hypothyroidism. Twenty serum samples from dogs with hypothyroidism and 20 from healthy dogs
were used for untargeted metabolomics analysis performed by LC/MS analysis. Fifteen metabolites showed
significant changes between hypothyroid and healthy dogs, being the pentose phosphate pathway (PPP),
aminoacyl-tRNA biosynthesis and pyrimidine metabolism the principal pathways altered in hypothyroidism.
Specifically, metabolites such as D-gluconic acid and L-Isoleucine may potentially act as biomarkers of disease.
2021-01-30T00:00:00Z