Pub Date : 2023-01-01DOI: 10.1590/1984-3143-AR2023-0017
Luanna Lorenna Vieira Rodrigues, Yasmin Beatriz França Moura, João Vitor da Silva Viana, Lhara Ricarliany Medeiros de Oliveira, Érika Almeida Praxedes, José de Brito Vieira, Sarah Leyenne Alves Sales, Herlon Victor Rodrigues Silva, Maria Claudia Dos Santos Luciano, Claudia Pessoa, Alexsandra Fernandes Pereira
The puma population is constantly decreasing, and cloning by somatic cell nuclear transfer can be used to conserve the species. One of the factors determining the success of the development of cloned embryos is the cell cycle stage of the donor cells. We evaluated the effects of full confluency (~100%), serum starvation (0.5% serum), and roscovitine (15 µM) treatments on the cell cycle synchronization in G0/G1 of puma skin-derived fibroblasts by flow cytometric analysis. Also, we assessed the effects of these synchronization methods on morphology, viability, and apoptosis levels using microscopy tools. The results showed that culturing the cells to confluence for 24 h (84.0%), 48 h (84.6%), and 72 h (84.2%) and serum starvation for 96 h (85.4%) yielded a significantly higher percentage of cells arrested in the G0/G1 (P 0.05) phase than cells not subjected to any cell cycle synchronization method (73.9%). Nevertheless, while serum starvation reduced the percentage of viable cells, no difference was observed for the full confluence and roscovitine treatments (P 0.05). Moreover, roscovitine for 12 h (78.6%) and 24 h (82.1%) was unable to synchronize cells in G0/G1 (P 0.05). In summary, full confluency induces puma fibroblast cell cycle synchronization at the G0/G1 stage without affecting cell viability. These outcomes may be valuable for planning donor cells for somatic cell nuclear transfer in pumas.
{"title":"Full confluency, serum starvation, and roscovitine for inducing arrest in the G<sub>0</sub>/G<sub>1</sub> phase of the cell cycle in puma skin-derived fibroblast lines.","authors":"Luanna Lorenna Vieira Rodrigues, Yasmin Beatriz França Moura, João Vitor da Silva Viana, Lhara Ricarliany Medeiros de Oliveira, Érika Almeida Praxedes, José de Brito Vieira, Sarah Leyenne Alves Sales, Herlon Victor Rodrigues Silva, Maria Claudia Dos Santos Luciano, Claudia Pessoa, Alexsandra Fernandes Pereira","doi":"10.1590/1984-3143-AR2023-0017","DOIUrl":"https://doi.org/10.1590/1984-3143-AR2023-0017","url":null,"abstract":"<p><p>The puma population is constantly decreasing, and cloning by somatic cell nuclear transfer can be used to conserve the species. One of the factors determining the success of the development of cloned embryos is the cell cycle stage of the donor cells. We evaluated the effects of full confluency (~100%), serum starvation (0.5% serum), and roscovitine (15 µM) treatments on the cell cycle synchronization in G<sub>0</sub>/G<sub>1</sub> of puma skin-derived fibroblasts by flow cytometric analysis. Also, we assessed the effects of these synchronization methods on morphology, viability, and apoptosis levels using microscopy tools. The results showed that culturing the cells to confluence for 24 h (84.0%), 48 h (84.6%), and 72 h (84.2%) and serum starvation for 96 h (85.4%) yielded a significantly higher percentage of cells arrested in the G<sub>0</sub>/G<sub>1</sub> (P 0.05) phase than cells not subjected to any cell cycle synchronization method (73.9%). Nevertheless, while serum starvation reduced the percentage of viable cells, no difference was observed for the full confluence and roscovitine treatments (P 0.05). Moreover, roscovitine for 12 h (78.6%) and 24 h (82.1%) was unable to synchronize cells in G<sub>0</sub>/G<sub>1</sub> (P 0.05). In summary, full confluency induces puma fibroblast cell cycle synchronization at the G<sub>0</sub>/G<sub>1</sub> stage without affecting cell viability. These outcomes may be valuable for planning donor cells for somatic cell nuclear transfer in pumas.</p>","PeriodicalId":7889,"journal":{"name":"Animal Reproduction","volume":null,"pages":null},"PeriodicalIF":1.7,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10124155/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9413760","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-01-01DOI: 10.1590/1984-3143-AR2022-0103
Sven Wuertz, Axel Orban, Fabian Johannes Schaefer, Julia Lynne Overton, Angela Krüger
Carotenoids are determinants of reproductive fitness and egg quality. Here we studied the accumulation of astaxanthin (AX), canthaxanthin (CA) zeaxanthin (ZX), lutein (LU), retinol (RX) and dehydroretinol (DR) during vitellogenesis comparing previtellogenic and vitellogenic pikeperch (Sander lucioperca) eggs (n = 5 each), as well as selected tissues (liver, fat and muscles) in first süawning females (1176-1450 g). Futhermore, we compared egg batches with high (88-99% hatching rate, n = 5) or low (40-67% hatching rate, n= 5) egg quality. Vitellogenic follicles revealed higher concentrations of DR, RX, ZX and LU compared to previtellogenic follicles. Neither CA nor AX was detectable. In parallel, DR and RX were mobilized in the liver. In adipose and muscle tissue, comparing previtellogenic and vitellogenic females, no significant differences in carotenoid/retinoid content were observed. In high quality egg batches, both DR and RX were increased. LU was lower in high quality than in low quality eggs. In a conclusion, the amount of retinoids seems suboptimal in low quality egg batches and increased DR and RX are desirable in pikeperch. Since hypervitaminosis of retinoids can be problematic though, supplementation of the food with carotenoids, which can serve as precursors for retinoids, has to be carried out carefully.
{"title":"Carotenoids and retinoids in the gonad of brood-stock pikeperch: accumulation during vitellogenesis and influence on egg quality in farmed pikeperch <i>Sander lucioperca</i>.","authors":"Sven Wuertz, Axel Orban, Fabian Johannes Schaefer, Julia Lynne Overton, Angela Krüger","doi":"10.1590/1984-3143-AR2022-0103","DOIUrl":"https://doi.org/10.1590/1984-3143-AR2022-0103","url":null,"abstract":"<p><p>Carotenoids are determinants of reproductive fitness and egg quality. Here we studied the accumulation of astaxanthin (AX), canthaxanthin (CA) zeaxanthin (ZX), lutein (LU), retinol (RX) and dehydroretinol (DR) during vitellogenesis comparing previtellogenic and vitellogenic pikeperch (<i>Sander lucioperca</i>) eggs (n = 5 each), as well as selected tissues (liver, fat and muscles) in first süawning females (1176-1450 g). Futhermore, we compared egg batches with high (88-99% hatching rate, n = 5) or low (40-67% hatching rate, n= 5) egg quality. Vitellogenic follicles revealed higher concentrations of DR, RX, ZX and LU compared to previtellogenic follicles. Neither CA nor AX was detectable. In parallel, DR and RX were mobilized in the liver. In adipose and muscle tissue, comparing previtellogenic and vitellogenic females, no significant differences in carotenoid/retinoid content were observed. In high quality egg batches, both DR and RX were increased. LU was lower in high quality than in low quality eggs. In a conclusion, the amount of retinoids seems suboptimal in low quality egg batches and increased DR and RX are desirable in pikeperch. Since hypervitaminosis of retinoids can be problematic though, supplementation of the food with carotenoids, which can serve as precursors for retinoids, has to be carried out carefully.</p>","PeriodicalId":7889,"journal":{"name":"Animal Reproduction","volume":null,"pages":null},"PeriodicalIF":1.7,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10205056/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9530276","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-01-01DOI: 10.1590/1984-3143-AR2023-0058
Marcia de Almeida Monteiro Melo Ferraz, Giuliana de Avila Ferronato
Traditional methods of gamete handling, fertilization, and embryo culture often face limitations in efficiency, consistency, and the ability to closely mimic in vivo conditions. This review explores the opportunities presented by microfluidic and 3D culture systems in overcoming these challenges and enhancing in vitro embryo production. We discuss the basic principles of microfluidics, emphasizing their inherent advantages such as precise control of fluid flow, reduced reagent consumption, and high-throughput capabilities. Furthermore, we delve into microfluidic devices designed for gamete manipulation, in vitro fertilization, and embryo culture, highlighting innovations such as droplet-based microfluidics and on-chip monitoring. Next, we explore the integration of 3D culture systems, including the use of biomimetic scaffolds and organ-on-a-chip platforms, with a particular focus on the oviduct-on-a-chip. Finally, we discuss the potential of these advanced systems to improve embryo production outcomes and advance our understanding of early embryo development. By leveraging the unique capabilities of microfluidics and 3D culture systems, we foresee significant advancements in the efficiency, effectiveness, and clinical success of in vitro embryo production.
{"title":"Opportunities involving microfluidics and 3D culture systems to the <i>in vitro</i> embryo production.","authors":"Marcia de Almeida Monteiro Melo Ferraz, Giuliana de Avila Ferronato","doi":"10.1590/1984-3143-AR2023-0058","DOIUrl":"https://doi.org/10.1590/1984-3143-AR2023-0058","url":null,"abstract":"<p><p>Traditional methods of gamete handling, fertilization, and embryo culture often face limitations in efficiency, consistency, and the ability to closely mimic <i>in vivo</i> conditions. This review explores the opportunities presented by microfluidic and 3D culture systems in overcoming these challenges and enhancing <i>in vitro</i> embryo production. We discuss the basic principles of microfluidics, emphasizing their inherent advantages such as precise control of fluid flow, reduced reagent consumption, and high-throughput capabilities. Furthermore, we delve into microfluidic devices designed for gamete manipulation, <i>in vitro</i> fertilization, and embryo culture, highlighting innovations such as droplet-based microfluidics and on-chip monitoring. Next, we explore the integration of 3D culture systems, including the use of biomimetic scaffolds and organ-on-a-chip platforms, with a particular focus on the oviduct-on-a-chip. Finally, we discuss the potential of these advanced systems to improve embryo production outcomes and advance our understanding of early embryo development. By leveraging the unique capabilities of microfluidics and 3D culture systems, we foresee significant advancements in the efficiency, effectiveness, and clinical success of <i>in vitro</i> embryo production.</p>","PeriodicalId":7889,"journal":{"name":"Animal Reproduction","volume":null,"pages":null},"PeriodicalIF":1.7,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10449241/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10107882","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-01-01DOI: 10.1590/1984-3143-AR2022-0086
Breno Queiroz Pinheiro, Francisco Felipe de Magalhães, Francisco Wesley da Silva Alves, Isaac Neto Goes Silva, Augusto Manuel Rodrigues Faustino, Lúcia Daniel Machado da Silva
The identification of putative prognostic factors in canine mammary neoplasms (CMNs) has been focused on tissue-specific biomarkers, but the serum biomarkers, including cancer antigen 15-3 (CA 15-3), c-reactive protein (CRP), and lactate dehydrogenase (LDH) have been demonstrated to display clinical application in cases of CMNs. The aim of the study was to evaluate the levels of these serum biomarkers and their association with well-established prognostic factors in CMNs. Samples from 15 female canines with CMNs and 15 clinically healthy ones were collected. The results were evaluated using the Tukey's, Pearson, or Spearman tests. The cut-off point, sensitivity, specificity, and area under curve (AUC) were evaluated using the receiver operating characteristic (ROC) curve analysis in a logistic regression model (P<0.05). The levels of CA 15-3, CRP and LDH were significantly higher in the serum of female dogs with CMNs compared to the healthy ones. Moreover, these factors were positively correlated with ulceration, tumor size, histopathological grade, metastatic lymph node, and clinical staging. Female dogs with CMNs were found to exhibit highest serum levels of CA 15-3, CRP, and LDH. Therefore, they can be applied to improve the efficacy of the diagnosis and prognostic evaluation in casas of CMNs.
犬乳腺肿瘤(CMNs)推定预后因素的鉴定主要集中在组织特异性生物标志物上,但血清生物标志物,包括癌抗原15-3 (CA 15-3)、c反应蛋白(CRP)和乳酸脱氢酶(LDH)已被证明在CMNs病例中具有临床应用价值。该研究的目的是评估这些血清生物标志物的水平及其与CMNs预后因素的关系。采集了15只CMNs母犬和15只临床健康犬的标本。使用Tukey’s、Pearson或Spearman检验对结果进行评估。采用logistic回归模型(P . 0)的受试者工作特征(ROC)曲线分析,评估分界点、敏感性、特异性和曲线下面积(AUC)
{"title":"CA 15-3, CRP, and LDH correlates with prognostic parameters in canine mammary neoplasms.","authors":"Breno Queiroz Pinheiro, Francisco Felipe de Magalhães, Francisco Wesley da Silva Alves, Isaac Neto Goes Silva, Augusto Manuel Rodrigues Faustino, Lúcia Daniel Machado da Silva","doi":"10.1590/1984-3143-AR2022-0086","DOIUrl":"https://doi.org/10.1590/1984-3143-AR2022-0086","url":null,"abstract":"<p><p>The identification of putative prognostic factors in canine mammary neoplasms (CMNs) has been focused on tissue-specific biomarkers, but the serum biomarkers, including cancer antigen 15-3 (CA 15-3), c-reactive protein (CRP), and lactate dehydrogenase (LDH) have been demonstrated to display clinical application in cases of CMNs. The aim of the study was to evaluate the levels of these serum biomarkers and their association with well-established prognostic factors in CMNs. Samples from 15 female canines with CMNs and 15 clinically healthy ones were collected. The results were evaluated using the Tukey's, Pearson, or Spearman tests. The cut-off point, sensitivity, specificity, and area under curve (AUC) were evaluated using the receiver operating characteristic (ROC) curve analysis in a logistic regression model (P<0.05). The levels of CA 15-3, CRP and LDH were significantly higher in the serum of female dogs with CMNs compared to the healthy ones. Moreover, these factors were positively correlated with ulceration, tumor size, histopathological grade, metastatic lymph node, and clinical staging. Female dogs with CMNs were found to exhibit highest serum levels of CA 15-3, CRP, and LDH. Therefore, they can be applied to improve the efficacy of the diagnosis and prognostic evaluation in casas of CMNs.</p>","PeriodicalId":7889,"journal":{"name":"Animal Reproduction","volume":null,"pages":null},"PeriodicalIF":1.7,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10023068/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9201077","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-01-01DOI: 10.1590/1984-3143-AR2023-0064
Clara Slade Oliveira, Luiz Sergio Almeida Camargo, Marcos Vinicius Gualberto Barbosa da Silva, Naiara Zoccal Saraiva, Carolina Capobiango Quintão, Marco Antonio Machado
Genomic selection has transformed the livestock industry, enabling early-life selection of animals. Biopsy sampling of pre-implantation embryos has been described since 1968. However, it was only after 2010, with the advancement of molecular biology techniques such as whole genomic amplification and SNP Chips, that next-generation sequencing became commercially available for bovine embryos. It is now possible to make decisions about which embryos to transfer not only based on recipients' availability or embryo morphology but also on genomic estimates. This technology can be implemented for a wide spectrum of applications in livestock. In this review, we discuss the use of embryo biopsy for genomic selection and share our experience with Gir and Girolando Brazilian breeding programs, as well as future goals for implementing it in Brazilian bovine in vitro embryo production practices.
{"title":"Embryo biopsies for genomic selection in tropical dairy cattle.","authors":"Clara Slade Oliveira, Luiz Sergio Almeida Camargo, Marcos Vinicius Gualberto Barbosa da Silva, Naiara Zoccal Saraiva, Carolina Capobiango Quintão, Marco Antonio Machado","doi":"10.1590/1984-3143-AR2023-0064","DOIUrl":"https://doi.org/10.1590/1984-3143-AR2023-0064","url":null,"abstract":"<p><p>Genomic selection has transformed the livestock industry, enabling early-life selection of animals. Biopsy sampling of pre-implantation embryos has been described since 1968. However, it was only after 2010, with the advancement of molecular biology techniques such as whole genomic amplification and SNP Chips, that next-generation sequencing became commercially available for bovine embryos. It is now possible to make decisions about which embryos to transfer not only based on recipients' availability or embryo morphology but also on genomic estimates. This technology can be implemented for a wide spectrum of applications in livestock. In this review, we discuss the use of embryo biopsy for genomic selection and share our experience with Gir and Girolando Brazilian breeding programs, as well as future goals for implementing it in Brazilian bovine <i>in vitro</i> embryo production practices.</p>","PeriodicalId":7889,"journal":{"name":"Animal Reproduction","volume":null,"pages":null},"PeriodicalIF":1.7,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10399131/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9944786","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-01-01DOI: 10.1590/1984-3143-AR2023-0021
Bruno Tomazele Rovani, Vitor Braga Rissi, Monique Tomazele Rovani, Bernardo Garziera Gasperin, Tadeu Baumhardt, Vilceu Bordignon, Liliane de Freitas Bauermann, Daniele Missio, Paulo Bayard Dias Gonçalves
Abstract Radiotherapy causes destruction of tumor cells, but also threatens the integrity and survival of surrounding normal cells. Then, woman submitted to irradiation for cancer treatment may present permanent ovary damage, resulting in impaired fertility. The objective of this study was to investigate the effects of therapeutic doses of ionizing radiation (IR), used for ovarian cancer treatment in humans, on bovine cumulus-oocyte complexes (COCs) as experimental model. Bovine ovaries were exposed to 0.9 Gy, 1.8 Gy, 3.6 Gy or 18.6 Gy IR, and then COCs were collected and used to evaluate: (a) oocyte nuclear maturation; (b) presence of phosphorylated H2A.X (γH2AX), as an indicator of DNA double-strand breaks (DSBs); and (c) expression of genes involved in DNA repair (TP53BP1, RAD52, ATM, XRCC6 and XRCC5) and apoptosis (BAX). The radiation doses tested in this study had no detrimental effects on nuclear maturation and did not increase γH2AX in the oocytes. However, IR treatment altered the mRNA abundance of RAD52 (RAD52 homolog, DNA repair protein) and BAX (BCL2-associated X protein). We conclude that although IR doses had no apparent effect on oocyte nuclear maturation and DNA damage, molecular pathways involved in DNA repair and apoptosis were affected by IR exposure in cumulus cells.
{"title":"Analysis of nuclear maturation, DNA damage and repair gene expression of bovine oocyte and cumulus cells submitted to ionizing radiation.","authors":"Bruno Tomazele Rovani, Vitor Braga Rissi, Monique Tomazele Rovani, Bernardo Garziera Gasperin, Tadeu Baumhardt, Vilceu Bordignon, Liliane de Freitas Bauermann, Daniele Missio, Paulo Bayard Dias Gonçalves","doi":"10.1590/1984-3143-AR2023-0021","DOIUrl":"https://doi.org/10.1590/1984-3143-AR2023-0021","url":null,"abstract":"Abstract Radiotherapy causes destruction of tumor cells, but also threatens the integrity and survival of surrounding normal cells. Then, woman submitted to irradiation for cancer treatment may present permanent ovary damage, resulting in impaired fertility. The objective of this study was to investigate the effects of therapeutic doses of ionizing radiation (IR), used for ovarian cancer treatment in humans, on bovine cumulus-oocyte complexes (COCs) as experimental model. Bovine ovaries were exposed to 0.9 Gy, 1.8 Gy, 3.6 Gy or 18.6 Gy IR, and then COCs were collected and used to evaluate: (a) oocyte nuclear maturation; (b) presence of phosphorylated H2A.X (γH2AX), as an indicator of DNA double-strand breaks (DSBs); and (c) expression of genes involved in DNA repair (TP53BP1, RAD52, ATM, XRCC6 and XRCC5) and apoptosis (BAX). The radiation doses tested in this study had no detrimental effects on nuclear maturation and did not increase γH2AX in the oocytes. However, IR treatment altered the mRNA abundance of RAD52 (RAD52 homolog, DNA repair protein) and BAX (BCL2-associated X protein). We conclude that although IR doses had no apparent effect on oocyte nuclear maturation and DNA damage, molecular pathways involved in DNA repair and apoptosis were affected by IR exposure in cumulus cells.","PeriodicalId":7889,"journal":{"name":"Animal Reproduction","volume":null,"pages":null},"PeriodicalIF":1.7,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10247184/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9982459","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Nucleotide-binding oligomerization domain receptors (NOD-like receptors, NLRs) have critical effects on interfaces of the immune and reproductive systems, and the spleen plays a key role in both innate and adaptive immune functions. It is hypothesized that NLR family participates in maternal splenic immune regulation during early pregnancy in sheep. In this study, maternal spleens were collected on day 16 of the estrous cycle, and days 13, 16 and 25 of gestation (n = 6 for each group) in ewes. Expression of NLR family, including NOD1, NOD2, class II transactivator (CIITA), NLR family apoptosis inhibitory protein (NAIP), nucleotide-binding oligomerization domain, Leucine rich repeat and Pyrin domain containing 1 (NLRP1), NLRP3 and NLRP7, was analyzed using quantitative real-time PCR, Western blot and immunohistochemistry analysis. The results revealed that expression levels of NOD1, NOD2, CIITA and NLRP3 were downregulated at days 13 and 16 of pregnancy, but expression of NLRP3 was increased at day 25 of pregnancy. In addition, expression values of NAIP and NLRP7 mRNA and proteins were improved at days 16 and 25 of pregnancy, and NLRP1 was peaked at days 13 and 16 of pregnancy in the maternal spleen. Furthermore, NOD2 and NLRP7 proteins were limited to the capsule, trabeculae and splenic cords. In summary, early pregnancy changes expression of NLR family in the maternal spleen, which may be related with the maternal splenic immunomodulation during early pregnancy in sheep.
{"title":"Changes in expression levels of Nod-like receptors in the spleen of ewes.","authors":"Jiaxuan Wu, Shengya Fang, Pengfei Feng, Chunjiang Cai, Leying Zhang, Ling Yang","doi":"10.1590/1984-3143-AR2022-0093","DOIUrl":"https://doi.org/10.1590/1984-3143-AR2022-0093","url":null,"abstract":"<p><p>Nucleotide-binding oligomerization domain receptors (NOD-like receptors, NLRs) have critical effects on interfaces of the immune and reproductive systems, and the spleen plays a key role in both innate and adaptive immune functions. It is hypothesized that NLR family participates in maternal splenic immune regulation during early pregnancy in sheep. In this study, maternal spleens were collected on day 16 of the estrous cycle, and days 13, 16 and 25 of gestation (n = 6 for each group) in ewes. Expression of NLR family, including NOD1, NOD2, class II transactivator (CIITA), NLR family apoptosis inhibitory protein (NAIP), nucleotide-binding oligomerization domain, Leucine rich repeat and Pyrin domain containing 1 (NLRP1), NLRP3 and NLRP7, was analyzed using quantitative real-time PCR, Western blot and immunohistochemistry analysis. The results revealed that expression levels of NOD1, NOD2, CIITA and NLRP3 were downregulated at days 13 and 16 of pregnancy, but expression of NLRP3 was increased at day 25 of pregnancy. In addition, expression values of NAIP and NLRP7 mRNA and proteins were improved at days 16 and 25 of pregnancy, and NLRP1 was peaked at days 13 and 16 of pregnancy in the maternal spleen. Furthermore, NOD2 and NLRP7 proteins were limited to the capsule, trabeculae and splenic cords. In summary, early pregnancy changes expression of NLR family in the maternal spleen, which may be related with the maternal splenic immunomodulation during early pregnancy in sheep.</p>","PeriodicalId":7889,"journal":{"name":"Animal Reproduction","volume":null,"pages":null},"PeriodicalIF":1.7,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10205055/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9525073","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
The domestic pig breeds are in danger of extinction whereas the erosion of their gene pool is a serious concern because they significantly contribute to the rich biodiversity. Overall aim of this study was to determine the protocol for preserving the semen of the Windsnyer boars for conservation. A total of 18 ejaculates (6 replications/boar) were collected from three Windsnyer boars of proven fertility with the use of hand-gloved approach method, twice per week. Boars semen were pooled and extended with Beltsville Thawing Solution [(BTS) IMV Technologies, France], held at 18°C for 3 hours and centrifuged. The sperm pellet was re-suspended with Fraction A (20% egg yolk + BTS) and cooled at 5°C for 1 hour. Following cooling, semen was divided and diluted into different cryoprotectants (ethylene glycol, glycerol, propanediol, ethylene glycol + glycerol + propanediol) at equal contribution to make the total concentrations [4, 8, 12 and 16% and the 0% (control; without cryoprotectant)] and loaded into 0.25 mL straws. Two cryopreservation methods (liquid nitrogen vapour and controlled rated) were used to cryopreserve the semen straws. Semen straws were thawed at different temperatures (5, 18, 37 and 40°C) and evaluated for sperm motility, viability, and morphology traits. Post-thawed sperm total motility (36.0±5.3) and live normal sperm (49.5±8.3) percentages were recorded to be higher in the treatment supplemented with 16% glycerol (P<0.05). The highest sperm total motility percentage was recorded at 40°C (26.8±3.2) thawing temperature for liquid nitrogen vapour treatment (P<0.05). In conclusion, 16% glycerol was found to be the suitable cryoprotectant concentration for semen cryopreserved with liquid nitrogen vapour method and thawed at 40°C.
{"title":"Sperm cryopreservation in Windsnyer boars; principles, technique, and updated outcomes","authors":"Mamonene Angelinah Thema, Masindi Lottus Mphaphathi, Mahlatsana Ramaesela Ledwaba, Tshimangadzo Lucky Nedambale","doi":"10.1590/1984-3143-ar2022-0100","DOIUrl":"https://doi.org/10.1590/1984-3143-ar2022-0100","url":null,"abstract":"The domestic pig breeds are in danger of extinction whereas the erosion of their gene pool is a serious concern because they significantly contribute to the rich biodiversity. Overall aim of this study was to determine the protocol for preserving the semen of the Windsnyer boars for conservation. A total of 18 ejaculates (6 replications/boar) were collected from three Windsnyer boars of proven fertility with the use of hand-gloved approach method, twice per week. Boars semen were pooled and extended with Beltsville Thawing Solution [(BTS) IMV Technologies, France], held at 18°C for 3 hours and centrifuged. The sperm pellet was re-suspended with Fraction A (20% egg yolk + BTS) and cooled at 5°C for 1 hour. Following cooling, semen was divided and diluted into different cryoprotectants (ethylene glycol, glycerol, propanediol, ethylene glycol + glycerol + propanediol) at equal contribution to make the total concentrations [4, 8, 12 and 16% and the 0% (control; without cryoprotectant)] and loaded into 0.25 mL straws. Two cryopreservation methods (liquid nitrogen vapour and controlled rated) were used to cryopreserve the semen straws. Semen straws were thawed at different temperatures (5, 18, 37 and 40°C) and evaluated for sperm motility, viability, and morphology traits. Post-thawed sperm total motility (36.0±5.3) and live normal sperm (49.5±8.3) percentages were recorded to be higher in the treatment supplemented with 16% glycerol (P<0.05). The highest sperm total motility percentage was recorded at 40°C (26.8±3.2) thawing temperature for liquid nitrogen vapour treatment (P<0.05). In conclusion, 16% glycerol was found to be the suitable cryoprotectant concentration for semen cryopreserved with liquid nitrogen vapour method and thawed at 40°C.","PeriodicalId":7889,"journal":{"name":"Animal Reproduction","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"136202625","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-01-01DOI: 10.1590/1984-3143-AR2023-0009
Pedro Nacib Jorge-Neto, Thiago Cavalheri Luczinski, Gediendson Ribeiro de Araújo, Letícia Alecho Requena, Rogério Silva de Jesus, Larissa Schneider Brandão Souza, Ricardo Zanella, Eliane Vianna da Costa E Silva, Thyara de Deco-Souza, Cristiane Schilbach Pizzutto
The cryopreservation of jaguar semen must be improved to produce high-quality biobanking doses. Until now, the rare studies of semen freezing in the species have only evaluated glycerol, always with a significant reduction in sperm quality in thawed semen. The purpose of this study was to assess the efficacy of three cryoprotectants, dimethylsulfoxide (DMSO), glycerol (GLY), and methanol (MET), in the cryopreservation of jaguar semen in an LDL-based extender, as well as the effect of thawing temperature on dosage quality. Five mature males with a history of reproduction were used. On the males, an infrared thermal image (IRT) was captured, the spicules and testes were analyzed, and the CASA system was used to evaluate the quality of fresh and thawed sperm. The superficial IRT was 4.6 ± 1.2 °C cooler than the anal sphincter, and the semen measured between 27.3 and 28.7 °C shortly after exiting the urethra. The total motility of fresh sperm was 55.3 ± 22.6%, and progressive motility was 36.3 ± 18%. The total motility of thawed sperm was 5.28 ± 2.51%, 4.49 ± %2.49, and 0.51 ± 0.62% for DMSO, GLY, and MET, respectively. DMSO and GLY performed better than MET, and there was no difference in thawing temperature (37°C 30 s vs. 50°C 12 s). All animals exhibit a considerable level of morphological changes in sperm. Low amounts of total and progressive motility were found in the thawed sperm. Males with a high level of sperm morphological changes were found to be fertile, but the lone male with normospermia was infertile. Thus, we contest the applicability of the commonly used morphological classification for bovines to felid species.
美洲豹精液的冷冻保存必须得到改进,以生产高质量的生物库剂量。到目前为止,对该物种精液冷冻的罕见研究只评估了甘油,解冻精液中的精子质量总是显著降低。本研究的目的是评估三种冷冻保护剂二甲基亚砜(DMSO)、甘油(GLY)和甲醇(MET)在低密度脂蛋白基填充剂中冷冻保存美洲豹精液的效果,以及解冻温度对剂量质量的影响。研究对象为5只有繁殖史的成年雄性。对雄性进行红外热成像(IRT),分析精子针状体和睾丸,并利用CASA系统对新鲜和解冻精子的质量进行评价。浅表IRT比肛门括约肌低4.6±1.2℃,精液在离开尿道后不久测得27.3 ~ 28.7℃。新鲜精子总活动力为55.3±22.6%,进行性活动力为36.3±18%。DMSO、GLY和MET解冻后精子总活力分别为5.28±2.51%、4.49±% 2.49%和0.51±0.62%。DMSO和GLY的表现优于MET,解冻温度(37°C 30 s vs 50°C 12 s)没有差异。所有动物的精子都表现出相当程度的形态变化。在解冻的精子中发现了少量的总运动性和进行性运动性。精子形态改变程度高的雄虫是可育的,而无精子症的雄虫是不育的。因此,我们对牛的常用形态分类在猫科动物中的适用性提出了质疑。
{"title":"Cryopreservation of jaguar (<i>Panthera onca</i>) sperm cells using different cryoprotectants and different thawing temperatures.","authors":"Pedro Nacib Jorge-Neto, Thiago Cavalheri Luczinski, Gediendson Ribeiro de Araújo, Letícia Alecho Requena, Rogério Silva de Jesus, Larissa Schneider Brandão Souza, Ricardo Zanella, Eliane Vianna da Costa E Silva, Thyara de Deco-Souza, Cristiane Schilbach Pizzutto","doi":"10.1590/1984-3143-AR2023-0009","DOIUrl":"https://doi.org/10.1590/1984-3143-AR2023-0009","url":null,"abstract":"<p><p>The cryopreservation of jaguar semen must be improved to produce high-quality biobanking doses. Until now, the rare studies of semen freezing in the species have only evaluated glycerol, always with a significant reduction in sperm quality in thawed semen. The purpose of this study was to assess the efficacy of three cryoprotectants, dimethylsulfoxide (DMSO), glycerol (GLY), and methanol (MET), in the cryopreservation of jaguar semen in an LDL-based extender, as well as the effect of thawing temperature on dosage quality. Five mature males with a history of reproduction were used. On the males, an infrared thermal image (IRT) was captured, the spicules and testes were analyzed, and the CASA system was used to evaluate the quality of fresh and thawed sperm. The superficial IRT was 4.6 ± 1.2 °C cooler than the anal sphincter, and the semen measured between 27.3 and 28.7 °C shortly after exiting the urethra. The total motility of fresh sperm was 55.3 ± 22.6%, and progressive motility was 36.3 ± 18%. The total motility of thawed sperm was 5.28 ± 2.51%, 4.49 ± %2.49, and 0.51 ± 0.62% for DMSO, GLY, and MET, respectively. DMSO and GLY performed better than MET, and there was no difference in thawing temperature (37°C 30 s vs. 50°C 12 s). All animals exhibit a considerable level of morphological changes in sperm. Low amounts of total and progressive motility were found in the thawed sperm. Males with a high level of sperm morphological changes were found to be fertile, but the lone male with normospermia was infertile. Thus, we contest the applicability of the commonly used morphological classification for bovines to felid species.</p>","PeriodicalId":7889,"journal":{"name":"Animal Reproduction","volume":null,"pages":null},"PeriodicalIF":1.7,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10075188/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9273557","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2023-01-01DOI: 10.1590/1984-3143-AR2022-0048
Willian Vaniel Alves Dos Reis, Raiza Rocha Pereira, Mozarth Vieira, Cibele Cristina Tavares da Cunha, Bianca Rodrigues Acácio, Gustavo Guerino Macedo, Eliane Vianna da Costa-E-Silva, Breno Fernandes Barreto Sampaio
The objective of this study was to reduce the effects of cryoinjury caused in bovine semen by cryopreservation. Ejaculates were collected from Nellore bulls and subjected to freezing in C (control), ozone (15, 30, and 60 µg mL-1 of ozone), quercetin (25, 50, and 100 µg mL-1 of quercetin), and carnosine groups (100, 200, and 300 ng mL-1 of carnosine). Samples were evaluated post-thaw (M0) and post-rapid thermoresistance test (M30) for sperm kinetics (total motility, progressive motility, curvilinear speed, linearity and amplitude of lateral head displacement) and cell structure viability (plasma membrane integrity, acrosomal integrity, mitochondrial potential, membrane fluidity, and lipid peroxidation). There were no differences (P > 0.05) between the control, quercetin, and carnosine-treated groups for the parameters evaluated at M0 and M30. In turn, supplementation with ozone resulted in lower values for sperm kinetics (P < 0.05) and lower mitochondrial potential at M30 (P < 0.05). Quercetin and carnosine at the concentrations used did not promote significant gains in frozen semen, nor did they demonstrate cytotoxicity. We expected to obtain positive results with the use of ozone. Nonetheless, the addition was harmful to the parameters of sperm kinetics, and its effect was not observed as a possible pro-antioxidant. We believe that the fact that the gas did not harm the sperm structure opens avenues for tests with lower dosages, since, by reducing its concentration, we could minimize the damage to sperm kinetics.
{"title":"Impact of quercetin, carnosine, and ozone in the cryopreservation on Nellore (<i>Bos indicus</i>) semen.","authors":"Willian Vaniel Alves Dos Reis, Raiza Rocha Pereira, Mozarth Vieira, Cibele Cristina Tavares da Cunha, Bianca Rodrigues Acácio, Gustavo Guerino Macedo, Eliane Vianna da Costa-E-Silva, Breno Fernandes Barreto Sampaio","doi":"10.1590/1984-3143-AR2022-0048","DOIUrl":"https://doi.org/10.1590/1984-3143-AR2022-0048","url":null,"abstract":"<p><p>The objective of this study was to reduce the effects of cryoinjury caused in bovine semen by cryopreservation. Ejaculates were collected from Nellore bulls and subjected to freezing in C (control), ozone (15, 30, and 60 µg mL<sup>-1</sup> of ozone), quercetin (25, 50, and 100 µg mL<sup>-1</sup> of quercetin), and carnosine groups (100, 200, and 300 ng mL<sup>-1</sup> of carnosine). Samples were evaluated post-thaw (M0) and post-rapid thermoresistance test (M30) for sperm kinetics (total motility, progressive motility, curvilinear speed, linearity and amplitude of lateral head displacement) and cell structure viability (plasma membrane integrity, acrosomal integrity, mitochondrial potential, membrane fluidity, and lipid peroxidation). There were no differences (<i>P > 0.05</i>) between the control, quercetin, and carnosine-treated groups for the parameters evaluated at M0 and M30. In turn, supplementation with ozone resulted in lower values for sperm kinetics (<i>P < 0.05</i>) and lower mitochondrial potential at M30 (<i>P < 0.05</i>). Quercetin and carnosine at the concentrations used did not promote significant gains in frozen semen, nor did they demonstrate cytotoxicity. We expected to obtain positive results with the use of ozone. Nonetheless, the addition was harmful to the parameters of sperm kinetics, and its effect was not observed as a possible pro-antioxidant. We believe that the fact that the gas did not harm the sperm structure opens avenues for tests with lower dosages, since, by reducing its concentration, we could minimize the damage to sperm kinetics.</p>","PeriodicalId":7889,"journal":{"name":"Animal Reproduction","volume":null,"pages":null},"PeriodicalIF":1.7,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10075187/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9273558","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}