{"title":"使用猪卵泡液衍生的外泌体进行体外成熟,以替代传统方法。","authors":"","doi":"10.1016/j.theriogenology.2024.08.030","DOIUrl":null,"url":null,"abstract":"<div><p>Extracellular vesicles, also known as exosomes, influence numerous cellular functions by regulating different signaling pathways. However, their role in animal reproduction remains understudied. This study aimed to evaluate the effects of porcine follicular fluid-derived exosomes (pff-Exos) on porcine oocyte <em>in vitro</em> maturation and parthenogenetic embryo development. We obtained pff-Exos through mixed-method ultracentrifugation and size-exclusion chromatography. Transmission electron microscopy revealed an increase in the expression of exosome markers in the first four of thirteen fractions. The number of pff-Exo was 2.2 × 10<sup>6</sup> particles per microliter. The highest maturation rate of porcine oocytes treated with pff-Exo was observed with 1.1 × 10<sup>7</sup> particles of pff-Exo in the absence of porcine follicular fluid (pFF) culture conditions. Moreover, increased expression of <em>Gdf9</em> and <em>Bmp15</em> was observed. The developmental rate was the highest upon treatment with 1.1 × 10<sup>7</sup> particles of pff-Exo, which increased the total cell number in blastocysts. Embryonic development to the 2-cell stage was similar between the control and pff-Exo groups; however, development to the 4-cell stage and blastocyst was significantly increased in the pff-Exo group (61.6 ± 6.08 % and 29.72 ± 1.41 %, respectively; <em>P</em> < 0.05) compared with that in the control group (42.0 ± 5.19 % and 18.14 ± 1.78 %, respectively). The expression levels of <em>Oct4</em>, <em>Sox2</em>, <em>Bcl2</em>, <em>Elf4</em>, and Gcn5 significantly increased at the pff-Exo 2-cell stage, whereas those of <em>Bax, Hdac1, Hdac6</em>, and <em>Sirt6</em> decreased. Specifically, the Oct4, Sox2, Elf4, Gcn5, and Hdac6 levels remained stable in pff-Exo 4-cell embryos, whereas those of p53 and <em>Hat1</em> were reduced and increased, respectively. Treatment with pffExos significantly increased H3K9 and H3K14 acetylation levels. These results demonstrate that pff-Exo affects the <em>in vitro</em> maturation of porcine oocytes and early embryonic development by regulating gene expression.</p></div>","PeriodicalId":23131,"journal":{"name":"Theriogenology","volume":null,"pages":null},"PeriodicalIF":2.4000,"publicationDate":"2024-08-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S0093691X24003583/pdfft?md5=3ec3871d35eae886aef5453074fef629&pid=1-s2.0-S0093691X24003583-main.pdf","citationCount":"0","resultStr":"{\"title\":\"In vitro maturation using porcine follicular fluid-derived exosomes as an alternative to the conventional method\",\"authors\":\"\",\"doi\":\"10.1016/j.theriogenology.2024.08.030\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><p>Extracellular vesicles, also known as exosomes, influence numerous cellular functions by regulating different signaling pathways. However, their role in animal reproduction remains understudied. This study aimed to evaluate the effects of porcine follicular fluid-derived exosomes (pff-Exos) on porcine oocyte <em>in vitro</em> maturation and parthenogenetic embryo development. We obtained pff-Exos through mixed-method ultracentrifugation and size-exclusion chromatography. Transmission electron microscopy revealed an increase in the expression of exosome markers in the first four of thirteen fractions. The number of pff-Exo was 2.2 × 10<sup>6</sup> particles per microliter. The highest maturation rate of porcine oocytes treated with pff-Exo was observed with 1.1 × 10<sup>7</sup> particles of pff-Exo in the absence of porcine follicular fluid (pFF) culture conditions. Moreover, increased expression of <em>Gdf9</em> and <em>Bmp15</em> was observed. The developmental rate was the highest upon treatment with 1.1 × 10<sup>7</sup> particles of pff-Exo, which increased the total cell number in blastocysts. Embryonic development to the 2-cell stage was similar between the control and pff-Exo groups; however, development to the 4-cell stage and blastocyst was significantly increased in the pff-Exo group (61.6 ± 6.08 % and 29.72 ± 1.41 %, respectively; <em>P</em> < 0.05) compared with that in the control group (42.0 ± 5.19 % and 18.14 ± 1.78 %, respectively). The expression levels of <em>Oct4</em>, <em>Sox2</em>, <em>Bcl2</em>, <em>Elf4</em>, and Gcn5 significantly increased at the pff-Exo 2-cell stage, whereas those of <em>Bax, Hdac1, Hdac6</em>, and <em>Sirt6</em> decreased. Specifically, the Oct4, Sox2, Elf4, Gcn5, and Hdac6 levels remained stable in pff-Exo 4-cell embryos, whereas those of p53 and <em>Hat1</em> were reduced and increased, respectively. Treatment with pffExos significantly increased H3K9 and H3K14 acetylation levels. These results demonstrate that pff-Exo affects the <em>in vitro</em> maturation of porcine oocytes and early embryonic development by regulating gene expression.</p></div>\",\"PeriodicalId\":23131,\"journal\":{\"name\":\"Theriogenology\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":2.4000,\"publicationDate\":\"2024-08-29\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.sciencedirect.com/science/article/pii/S0093691X24003583/pdfft?md5=3ec3871d35eae886aef5453074fef629&pid=1-s2.0-S0093691X24003583-main.pdf\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Theriogenology\",\"FirstCategoryId\":\"97\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S0093691X24003583\",\"RegionNum\":2,\"RegionCategory\":\"农林科学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q3\",\"JCRName\":\"REPRODUCTIVE BIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Theriogenology","FirstCategoryId":"97","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0093691X24003583","RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q3","JCRName":"REPRODUCTIVE BIOLOGY","Score":null,"Total":0}
In vitro maturation using porcine follicular fluid-derived exosomes as an alternative to the conventional method
Extracellular vesicles, also known as exosomes, influence numerous cellular functions by regulating different signaling pathways. However, their role in animal reproduction remains understudied. This study aimed to evaluate the effects of porcine follicular fluid-derived exosomes (pff-Exos) on porcine oocyte in vitro maturation and parthenogenetic embryo development. We obtained pff-Exos through mixed-method ultracentrifugation and size-exclusion chromatography. Transmission electron microscopy revealed an increase in the expression of exosome markers in the first four of thirteen fractions. The number of pff-Exo was 2.2 × 106 particles per microliter. The highest maturation rate of porcine oocytes treated with pff-Exo was observed with 1.1 × 107 particles of pff-Exo in the absence of porcine follicular fluid (pFF) culture conditions. Moreover, increased expression of Gdf9 and Bmp15 was observed. The developmental rate was the highest upon treatment with 1.1 × 107 particles of pff-Exo, which increased the total cell number in blastocysts. Embryonic development to the 2-cell stage was similar between the control and pff-Exo groups; however, development to the 4-cell stage and blastocyst was significantly increased in the pff-Exo group (61.6 ± 6.08 % and 29.72 ± 1.41 %, respectively; P < 0.05) compared with that in the control group (42.0 ± 5.19 % and 18.14 ± 1.78 %, respectively). The expression levels of Oct4, Sox2, Bcl2, Elf4, and Gcn5 significantly increased at the pff-Exo 2-cell stage, whereas those of Bax, Hdac1, Hdac6, and Sirt6 decreased. Specifically, the Oct4, Sox2, Elf4, Gcn5, and Hdac6 levels remained stable in pff-Exo 4-cell embryos, whereas those of p53 and Hat1 were reduced and increased, respectively. Treatment with pffExos significantly increased H3K9 and H3K14 acetylation levels. These results demonstrate that pff-Exo affects the in vitro maturation of porcine oocytes and early embryonic development by regulating gene expression.
期刊介绍:
Theriogenology provides an international forum for researchers, clinicians, and industry professionals in animal reproductive biology. This acclaimed journal publishes articles on a wide range of topics in reproductive and developmental biology, of domestic mammal, avian, and aquatic species as well as wild species which are the object of veterinary care in research or conservation programs.