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The effects of mitoquinol mesylate on post-thaw characteristics and fertilizing ability of rooster semen cryopreserved in the Beltsville medium 甲磺酸米托喹啉对Beltsville培养基中冷藏公鸡精液解冻后特性及受精能力的影响。
IF 2.4 2区 农林科学 Q3 REPRODUCTIVE BIOLOGY Pub Date : 2025-01-11 DOI: 10.1016/j.theriogenology.2025.01.008
N. Asadzadeh , R. Masoudi , R. Nateghi , N.D. Davachi , H.J. Barfourooshi , P.M. Bartlewski
Cryopreservation of rooster semen is a reproductive technology carried out to boost genetic gain and productivity in commercial flocks of chicken. However, semen freezing significantly reduces the quality and fertilizing potential of spermatozoa. This study examined cryoprotective effects of the mitochondria-targeted antioxidant mitoquinol mesylate added to the freezing extender by assessing post-thaw characteristics of rooster sperm. Semen samples were diluted in the Beltsville extender supplemented with 0, 1, 10, 100 or 1000 nM of mitoquinol mesylate. Following the thawing of cryopreserved semen doses, we evaluated the following sperm parameters: motility and morphology, membrane integrity and mitochondrial function, acrosome integrity, apoptosis status, lipid peroxidation, DNA fragmentation, reactive oxygen species (ROS) accumulation, epigenetic patterns (DNA methylation and histone modifications), and fertilizing ability. Semen diluted in extenders containing 10 or 100 nM of mitoquinol mesylate significantly exceeded all other groups of frozen-thawed semen samples in sperm motility, average path velocity as well as membrane/acrosome integrity, mitochondrial function indices and DNA/histone modification. Moreover, the addition of 10 and 100 nM of mitoquinol mesylate significantly reduced lipid peroxidation, apoptosis rate, DNA fragmentation and ROS concentrations compared with all other dilutions and was associated with a higher (P ≤ 0.05) fertilization rate compared with a non-supplemented control group, albeit it was still significantly lower compared with that obtained using fresh semen. It can be, however, concluded that mitoquinol mesylate significantly improved an array of quality parameters and fertilizing potential of rooster semen, and hence can be recommended for use as a diluent additive in semen cryopreservation procedures employed in commercial poultry operations.
公鸡精液的低温保存是一种生殖技术,用于提高商业鸡群的遗传增益和生产力。然而,精子冷冻会显著降低精子的质量和受精潜力。本研究通过评估公鸡精子的解冻后特性,研究了在冷冻扩展剂中添加线粒体靶向抗氧化剂甲磺酸米托喹啉的冷冻保护作用。精液样品在添加0、1、10、100或1000 nM甲磺酸米托喹啉的Beltsville扩展器中稀释。在冷冻保存的精液解冻后,我们评估了以下精子参数:活力和形态、膜完整性和线粒体功能、顶体完整性、细胞凋亡状态、脂质过氧化、DNA片段化、活性氧(ROS)积累、表观遗传模式(DNA甲基化和组蛋白修饰)和受精能力。在含有10或100 nM甲磺酸米托喹啉的延长剂中稀释的精液在精子活力、平均路径速度、膜/顶体完整性、线粒体功能指数和DNA/组蛋白修饰方面显著优于其他所有冻融精液样品。此外,与其他稀释剂相比,添加10和100 nM甲甲酸米托喹啉显著降低了脂质过氧化、细胞凋亡率、DNA片段化和ROS浓度,与未添加对照组相比,受精率更高(P≤0.05),但与使用新鲜精液相比仍显著降低。然而,可以得出结论,甲磺酸米托喹啉显著改善了公鸡精液的一系列质量参数和受精潜力,因此可以推荐在商业家禽经营中使用的精液冷冻保存过程中作为稀释剂添加剂。
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引用次数: 0
Antagonistic interaction between miR-143 and KRAS gene regulating male mouse germ cell apoptosis miR-143与KRAS基因调控雄性小鼠生殖细胞凋亡的拮抗相互作用
IF 2.4 2区 农林科学 Q3 REPRODUCTIVE BIOLOGY Pub Date : 2025-01-11 DOI: 10.1016/j.theriogenology.2024.12.024
Yu Lu , Shudong Niu , Guisheng Zhang, Yanfeng Guo, Baotong Fu, Miaomiao Wang, Jianan Liu, Haili Zhang, Wen Lu, Ming Zhang
Precisely regulated spermatocyte growth, differentiation, and apoptosis are crucial for sustainable male fertility. miR-143 has been demonstrated to regulate gene expression and cell apoptosis in various human cancers. However, the function of mmu-mir-143 (miR-143) in mammalian testes and its underlying mechanism remains unexplored. In this study, the expression of miR-143 was detected in C57BL/6 mice spermatocytes by in situ hybridization (ISH) and immunofluorescence (IF) co-staining and transfecting miR-143 inhibitor into GC-2 cells (mouse spermatogenic cells) shows that miR-143 inhibits cleaved Caspase 3 (CC3)-induced male germ cell death. The current study used IF co-staining of KI67 and γ-H2A.X in the testes of C57BL/6 mice at different developmental stages, revealing that active proliferation and apoptosis of spermatocytes occurred simultaneously in the testes at 14 day post-partum (dpp). Kras was predicted as a potential target of miR-143 in mice using of the online database TargetScan, verified by quantitative real-time PCR (qPCR), western blotting (WB), and Dual-luciferase reporter gene assay. Co-transfection of miR-143 inhibitor and Kras siRNA into GC-2 cells revealed an antagonistic correlation between miR-143 and Kras in regulating male germ cell death. Finally, miR-143 inhibitor and mimics were administered into the seminiferous tubule of 3-week-old C57BL/6 mice. The histomorphology, IF co-staining, and WB data indicated that the testes treated with the miR-143 inhibitor showed significantly aberrant phenotypes, including damaged seminiferous tubules, reduced spermatocyte quantity, and elevated levels of apoptosis. This study uncovered the mechanism by which miR-143 inhibits male germ cell apoptosis through the repression of Kras/KRAS levels and the inhibition of Caspase 3 activation, providing insight into the role of miRNA in spermatogenesis and the maintenance of male fertility.
精确调节精母细胞的生长、分化和凋亡是维持男性生育能力的关键。miR-143已被证明在多种人类癌症中调节基因表达和细胞凋亡。然而,mmu-mir-143 (miR-143)在哺乳动物睾丸中的功能及其潜在机制仍未被探索。本研究通过原位杂交(ISH)和免疫荧光(IF)共染色检测了miR-143在C57BL/6小鼠精细胞中的表达,并将miR-143抑制剂转染到GC-2细胞(小鼠生精细胞)中,结果表明miR-143抑制cleaved Caspase 3 (CC3)诱导的雄性生殖细胞死亡。本研究采用IF共染色KI67和γ-H2A。X在C57BL/6小鼠不同发育阶段睾丸中的表达,揭示了产后14天睾丸中精子细胞的活跃增殖和凋亡同时发生。利用在线数据库TargetScan预测Kras是miR-143在小鼠中的潜在靶标,并通过定量实时PCR (qPCR)、western blotting (WB)和双荧光素酶报告基因测定进行验证。将miR-143抑制剂和Kras siRNA共转染GC-2细胞,发现miR-143和Kras在调节男性生殖细胞死亡方面存在拮抗相关性。最后,将miR-143抑制剂和模拟物注入3周龄C57BL/6小鼠的精管中。组织形态学、IF共染色和WB数据表明,miR-143抑制剂处理的睾丸显示出明显的异常表型,包括精管损伤、精细胞数量减少和细胞凋亡水平升高。本研究揭示了miR-143通过抑制Kras/ Kras水平和抑制Caspase 3激活抑制男性生殖细胞凋亡的机制,为miRNA在精子发生和维持男性生育能力中的作用提供了新的视角。
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引用次数: 0
R-spondin1 plays an indispensable role in ovarian development of Qi River crucian carp (Carassius auratus) by regulating estrogen synthesis R-spondin1通过调节雌激素的合成,在祁河鲫鱼卵巢发育中起着不可缺少的作用。
IF 2.4 2区 农林科学 Q3 REPRODUCTIVE BIOLOGY Pub Date : 2025-01-11 DOI: 10.1016/j.theriogenology.2025.01.006
Limin Wu , Mengfan Wu , Yongjing Li , Qingqing Xin , Yuchi Wang , Xi Shi , Xuejun Li
R-spondin1 (Rspo1) is a member of the secreted furin-like domain-containing protein family, and it is recognized for its significance in mammalian ovarian development. However, its role in teleost ovarian development remains largely uninvestigated. The Qi River crucian carp (Carassius auratus) is a species capable of gynogenesis, and it encounters challenges of premature ovarian maturation in aquaculture settings. Previous research established the essential involvement of Rspo1 in oocyte growth in Qi River crucian carp, but the precise molecular mechanisms underlying its role remain poorly understood. In this study, we categorized the pre-spawning ovarian development process of premature Qi River crucian carp into five stages through meticulous examination of morphology and histology. Immunofluorescence analysis revealed colocalization of Rspo1 with Vasa protein in oogonia, primary growth stage, and cortical vacuolar stage oocytes, and it was also detected in somatic cells. After a 60-day period of RNA interference via injection of Rspo1 double-stranded RNA into late-previtellogenesis stage ovaries, a substantial proportion of oocytes were arrested in the primary growth stage and exhibited a marked reduction in the expression of germ cell marker genes and an increase in apoptosis signaling. RNA-sequencing and real-time PCR analyses indicated a potential association between genes involved in hormone synthesis, lipid storage, and cell proliferation with ovary development in Qi River crucian carp. Furthermore, a significant decrease in levels of serum estrogens and vitellogenin was observed after Rspo1 knockdown. Dual-fluorescence in situ hybridization analysis demonstrated co-expression of Rspo1 with cyp19a1a in ovarian germ and surrounding somatic cells. Furthermore, results of a promoter assay indicated that Rspo1 can dose-dependently activate cyp19a1a expression. Collectively, these findings suggest that Rspo1 plays a role in ovarian development and oocyte growth by modulating cyp19a1a expression and influencing estrogen synthesis. These results provide valuable insights into the molecular mechanisms underlying the involvement of Rspo1 in ovarian development in Qi River crucian carp.
R-spondin1 (Rspo1)是分泌的含furin样结构域蛋白家族的成员,在哺乳动物卵巢发育中具有重要意义。然而,它在硬骨鱼卵巢发育中的作用在很大程度上仍未被研究。齐河鲫鱼(Carassius auratus)是一种能够雌核发生的物种,它在水产养殖环境中遇到卵巢早熟的挑战。以往的研究确定了Rspo1在祁河鲫鱼卵母细胞生长中的重要作用,但其作用的确切分子机制尚不清楚。在本研究中,我们通过细致的形态学和组织学检查,将早熟齐河鲫鱼产卵前卵巢发育过程分为五个阶段。免疫荧光分析显示,在卵原细胞、初生长期和皮质空泡期卵母细胞中,Rspo1与Vasa蛋白共定位,在体细胞中也检测到。在胚胎发育晚期卵巢注射Rspo1双链RNA干扰60天后,大量卵母细胞停留在初级生长阶段,生殖细胞标记基因表达明显减少,凋亡信号明显增加。rna测序和实时PCR分析表明,与激素合成、脂质储存和细胞增殖有关的基因与齐河鲫鱼卵巢发育存在潜在关联。此外,Rspo1敲除后,血清雌激素和卵黄蛋白原水平显著下降。双荧光原位杂交分析证实Rspo1与cyp19a1a在卵巢胚和周围体细胞中共表达。此外,启动子实验结果表明,Rspo1可以剂量依赖性地激活cyp19a1a的表达。综上所述,这些发现表明Rspo1通过调节cyp19a1a表达和影响雌激素合成在卵巢发育和卵母细胞生长中起作用。这些结果为揭示Rspo1参与鲫鱼卵巢发育的分子机制提供了有价值的见解。
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引用次数: 0
Influence of antifreeze protein III on canine sperm cryopreservation 抗冻蛋白III对犬精子冷冻保存的影响。
IF 2.4 2区 农林科学 Q3 REPRODUCTIVE BIOLOGY Pub Date : 2025-01-07 DOI: 10.1016/j.theriogenology.2025.01.004
Abbas Farshad, Emilia Diel, Axel Wehrend
Sperm cryopreservation is crucial in reproductive biotechnology; however, the longevity of frozen and thawed semen is limited by the deterioration of sperm cell integrity. This study aimed to examine the effects of adding antifreeze protein III (AFP III) to the diluent, using samples from eight healthy mature dogs. The ejaculates were divided into aliquots and diluted with a standard Tris-fructose-egg yolk extender containing AFP III at concentrations of 0, 0.75, 1.0, and 2.0 μg/ml. After thawing, the samples were analyzed for kinematic parameters, membrane Integrity, lipid peroxidation, viability, acrosome integrity, intracellular hydrogen peroxide, mitochondrial membrane potential and apoptotic metrics. The results show that while motility and velocity were not significantly different between the treated and control groups (p > 0.05), the treated groups generally performed better. Specifically, the 0.75 and 1.0 μg/ml groups exhibited better movement compared to the 2.0 μg/ml group. Additionally, there was a significant difference (p < 0.05) in membrane integrity between the control and treated groups, though no differences were observed among the treated groups. Significant differences (p < 0.05) were also observed in viability and acrosome integrity, with the 0.75 and 1.0 μg/ml groups outperforming the control and 2.0 μg/ml groups. There were no significant variations (p > 0.05) in phosphatidylserine translocation, lipid peroxidation, mitochondrial membrane potential, or hydrogen peroxide levels. However, the 0.75 and 1.0 μg/ml groups demonstrated superior effects compared to both the control and the 2.0 μg/ml groups. These results suggest that the addition of antifreeze proteins, specifically AFP III, markedly improves the protection of canine sperm during cryopreservation. This enhancement is evident in various parameters, underscoring the beneficial effects of AFP III in maintaining sperm quality.
精子冷冻保存在生殖生物技术中至关重要;然而,冷冻和解冻精液的寿命受到精子细胞完整性恶化的限制。本研究旨在检测在稀释液中添加抗冻蛋白III (AFP III)的效果,使用来自8只健康成年犬的样本。将射精液分成等分,用含有AFP III的标准三果糖蛋黄扩展剂稀释,浓度分别为0、0.75、1.0和2.0 μg/ml。解冻后,分析样品的运动学参数、膜完整性、脂质过氧化、活力、顶体完整性、细胞内过氧化氢、线粒体膜电位和凋亡指标。结果表明,虽然治疗组和对照组之间的运动和速度差异不显著(p < 0.05),但治疗组总体表现较好。其中,与2.0 μg/ml组相比,0.75和1.0 μg/ml组表现出更好的运动。此外,磷脂酰丝氨酸易位、脂质过氧化、线粒体膜电位和过氧化氢水平也有显著差异(p 0.05)。然而,与对照组和2.0 μg/ml组相比,0.75和1.0 μg/ml组表现出更好的效果。这些结果表明,添加抗冻蛋白,特别是AFP III,可以显著提高犬精子在冷冻保存过程中的保护作用。这种增强在各种参数中都很明显,强调了AFP III在维持精子质量方面的有益作用。
{"title":"Influence of antifreeze protein III on canine sperm cryopreservation","authors":"Abbas Farshad,&nbsp;Emilia Diel,&nbsp;Axel Wehrend","doi":"10.1016/j.theriogenology.2025.01.004","DOIUrl":"10.1016/j.theriogenology.2025.01.004","url":null,"abstract":"<div><div>Sperm cryopreservation is crucial in reproductive biotechnology; however, the longevity of frozen and thawed semen is limited by the deterioration of sperm cell integrity. This study aimed to examine the effects of adding antifreeze protein III (AFP III) to the diluent, using samples from eight healthy mature dogs. The ejaculates were divided into aliquots and diluted with a standard Tris-fructose-egg yolk extender containing AFP III at concentrations of 0, 0.75, 1.0, and 2.0 μg/ml. After thawing, the samples were analyzed for kinematic parameters, membrane Integrity, lipid peroxidation, viability, acrosome integrity, intracellular hydrogen peroxide, mitochondrial membrane potential and apoptotic metrics. The results show that while motility and velocity were not significantly different between the treated and control groups (p &gt; 0.05), the treated groups generally performed better. Specifically, the 0.75 and 1.0 μg/ml groups exhibited better movement compared to the 2.0 μg/ml group. Additionally, there was a significant difference (p &lt; 0.05) in membrane integrity between the control and treated groups, though no differences were observed among the treated groups. Significant differences (p &lt; 0.05) were also observed in viability and acrosome integrity, with the 0.75 and 1.0 μg/ml groups outperforming the control and 2.0 μg/ml groups. There were no significant variations (p &gt; 0.05) in phosphatidylserine translocation, lipid peroxidation, mitochondrial membrane potential, or hydrogen peroxide levels. However, the 0.75 and 1.0 μg/ml groups demonstrated superior effects compared to both the control and the 2.0 μg/ml groups. These results suggest that the addition of antifreeze proteins, specifically AFP III, markedly improves the protection of canine sperm during cryopreservation. This enhancement is evident in various parameters, underscoring the beneficial effects of AFP III in maintaining sperm quality.</div></div>","PeriodicalId":23131,"journal":{"name":"Theriogenology","volume":"235 ","pages":"Pages 86-93"},"PeriodicalIF":2.4,"publicationDate":"2025-01-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142972185","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
PARL regulates porcine oocyte meiotic maturation by mediating mitochondrial activity PARL通过介导线粒体活性调控猪卵母细胞减数分裂成熟。
IF 2.4 2区 农林科学 Q3 REPRODUCTIVE BIOLOGY Pub Date : 2025-01-07 DOI: 10.1016/j.theriogenology.2025.01.005
Naru Zhou , Zongliang Liu , Zhenhu Shi , Lei Luo , Mengqing Xuan , Ruiqing Zhu , Kunlong Hu , Xinyue Zhu , Wenhuan Xu , Yunsheng Li , Zubing Cao , Yunhai Zhang
PARL is a rhomboid membrane protein that plays a crucial role in regulating the metabolism and maintaining the homeostasis of mitochondria which provide important energy and material reserves for oocyte maturation. However, the impact of PARL on oocyte maturation remains poorly understood. Here, we elucidated the pivotal role of PARL in oocyte maturation through its regulatory effects on mitochondrial activity. Specifically, our findings revealed that inhibiting PARL expression by interfering with RNA transcription in oocytes led to a substantial decrease in the rate of first polar body extrusion and early development of parthenogenetically activated embryos. Moreover, PARL deficiency disrupted mitochondrial distribution and activity, leading to the accumulation of ROS, abnormal distribution of CGs and actin, increased tubulin acetylation modification, disturbed spindle assembly and chromosome alignment, ultimately caused DNA damage in porcine oocytes at the metaphase II stage. Intriguingly, PARL deficiency did not cause occurrence of apoptosis in oocytes. Furthermore, our study highlighted that PARL deficiency caused the aberrant expression of genes associated with oocyte maturation, particularly those genes associated with mitochondrial function and DNA integrity. Collectively, these results demonstrate that the indispensable role of PARL in orchestrating porcine oocyte meiotic maturation though its modulation of mitochondrial activity.
PARL是一种菱形膜蛋白,在调节线粒体代谢和维持线粒体稳态中起着至关重要的作用,为卵母细胞成熟提供重要的能量和物质储备。然而,PARL对卵母细胞成熟的影响仍然知之甚少。在这里,我们通过对线粒体活性的调节作用阐明了PARL在卵母细胞成熟中的关键作用。具体来说,我们的研究结果表明,通过干扰卵母细胞中的RNA转录来抑制PARL的表达,可以显著降低第一极体挤出率和孤雌激活胚胎的早期发育。此外,PARL缺乏破坏了线粒体的分布和活性,导致ROS的积累,CGs和肌动蛋白的异常分布,微管蛋白乙酰化修饰增加,纺锤体组装和染色体排列受到干扰,最终导致猪卵母细胞在II中期出现DNA损伤。有趣的是,PARL缺乏并未导致卵母细胞发生凋亡。此外,我们的研究强调,PARL缺陷导致与卵母细胞成熟相关的基因表达异常,特别是与线粒体功能和DNA完整性相关的基因。总的来说,这些结果表明PARL通过调节线粒体活性在协调猪卵母细胞减数分裂成熟中发挥了不可或缺的作用。
{"title":"PARL regulates porcine oocyte meiotic maturation by mediating mitochondrial activity","authors":"Naru Zhou ,&nbsp;Zongliang Liu ,&nbsp;Zhenhu Shi ,&nbsp;Lei Luo ,&nbsp;Mengqing Xuan ,&nbsp;Ruiqing Zhu ,&nbsp;Kunlong Hu ,&nbsp;Xinyue Zhu ,&nbsp;Wenhuan Xu ,&nbsp;Yunsheng Li ,&nbsp;Zubing Cao ,&nbsp;Yunhai Zhang","doi":"10.1016/j.theriogenology.2025.01.005","DOIUrl":"10.1016/j.theriogenology.2025.01.005","url":null,"abstract":"<div><div>PARL is a rhomboid membrane protein that plays a crucial role in regulating the metabolism and maintaining the homeostasis of mitochondria which provide important energy and material reserves for oocyte maturation. However, the impact of PARL on oocyte maturation remains poorly understood. Here, we elucidated the pivotal role of PARL in oocyte maturation through its regulatory effects on mitochondrial activity. Specifically, our findings revealed that inhibiting PARL expression by interfering with RNA transcription in oocytes led to a substantial decrease in the rate of first polar body extrusion and early development of parthenogenetically activated embryos. Moreover, PARL deficiency disrupted mitochondrial distribution and activity, leading to the accumulation of ROS, abnormal distribution of CGs and actin, increased tubulin acetylation modification, disturbed spindle assembly and chromosome alignment, ultimately caused DNA damage in porcine oocytes at the metaphase II stage. Intriguingly, PARL deficiency did not cause occurrence of apoptosis in oocytes. Furthermore, our study highlighted that PARL deficiency caused the aberrant expression of genes associated with oocyte maturation, particularly those genes associated with mitochondrial function and DNA integrity. Collectively, these results demonstrate that the indispensable role of PARL in orchestrating porcine oocyte meiotic maturation though its modulation of mitochondrial activity.</div></div>","PeriodicalId":23131,"journal":{"name":"Theriogenology","volume":"235 ","pages":"Pages 75-85"},"PeriodicalIF":2.4,"publicationDate":"2025-01-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142972186","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
The mechanism of Se in regulating the proliferation and apoptosis of sheep Leydig cells through the miR-200a/NRF2 pathway Se通过miR-200a/NRF2途径调节绵羊亮德细胞增殖和凋亡的机制
IF 2.4 2区 农林科学 Q3 REPRODUCTIVE BIOLOGY Pub Date : 2025-01-06 DOI: 10.1016/j.theriogenology.2025.01.003
Kexin Li , Xiaolei Wang , Liang Ma , Youshe Ren , Lei Shi
This study aimed to investigate the mechanism by which Se in regulates the proliferation and apoptosis of sheep Leydig cells via the miR-200a/NRF pathway. The cells were isolated and purified from the testes of 8-month-old sheep via a Percoll density gradient. After the cells were treated with different concentrations of Se (0, 2.0, 4.0, 6.0, and 8.0 μmol/L of Se) for 18 h, the miR-200a levels was detected. MiR-200a mimics and inhibitors were transfected into the cells, resulting in five groups (control, NC mimics, miR-200a mimics, NC inhibitor and miR-200a inhibitor). Cell viability and antioxidant status were measured via CCK8 and antioxidant assays, respectively. The abundances of pro-apoptotic (BAX, CASPASE 3 and CASPASE 8), cell cycle (P21, P27 and CDK1), and NRF2-related (NRF2, HO-1, NQO1 and KEAP1) genes were detected by real-time PCR and Western blot analysis.
The results revealed that miR-200a mimics group presented greater (P < 0.05) abundances of NRF2, HO-1 and NQO1 mRNA transcripts and proteins. Compared with those both in the NC mimics and the miR-200a inhibitor groups, the activities of GSH-Px and SOD, as well as cell viability in the miR-200a mimics group were significantly greater (P < 0.05). In contrast, the ROS levels, MDA content and abundances of KEAP1, P21, P27 and apoptosis-related genes mRNA transcripts and proteins were decreased (P < 0.05). The highest (P < 0.05) miR-200a expression level was detected in the Se6.0 group. Compared with that in the Se (6.0 μmol/L) group, cell viability in the Se + miR-200a inhibitor group was lower (P < 0.05). The abundances of NRF2, HO-1 and NQO1 in the Se + miR-200a inhibitor group were lower (P < 0.05) than those in the Se (6.0 μmol/L) group but greater (P < 0.05) than those in the inhibitor group, while KEAP1 displayed the opposite trend (P < 0.05).
These results indicate that Se can activate the NRF2 antioxidant signaling pathway to regulate the proliferation and apoptosis of sheep Leydig cells and that miR-200a plays a vital role in this process. The regulatory effect of Se on male reproduction and spermatogenesis may be related to the number of Leydig cells. This study aimed to provide experimental data for Se regulation of spermatogenesis.
本研究旨在探讨硒通过miR-200a/NRF通路调控绵羊间质细胞增殖和凋亡的机制。通过Percoll密度梯度从8月龄羊的睾丸中分离纯化细胞。不同浓度Se(0、2.0、4.0、6.0、8.0 μmol/L Se)处理细胞18 h后,检测miR-200a水平。将MiR-200a模拟物和抑制物转染到细胞中,分为五组(对照组、NC模拟物、MiR-200a模拟物、NC抑制剂和MiR-200a抑制剂)。通过CCK8和抗氧化试验分别测定细胞活力和抗氧化状态。采用real-time PCR和Western blot检测促凋亡基因(BAX、CASPASE 3和CASPASE 8)、细胞周期基因(P21、P27和CDK1)和NRF2相关基因(NRF2、HO-1、NQO1和KEAP1)的丰度。结果显示,miR-200a模拟组与对照组相比,差异有统计学意义(P < 6.0)。与Se (6.0 μmol/L)组相比,Se + miR-200a抑制剂组细胞活力降低(P < 0.05)
{"title":"The mechanism of Se in regulating the proliferation and apoptosis of sheep Leydig cells through the miR-200a/NRF2 pathway","authors":"Kexin Li ,&nbsp;Xiaolei Wang ,&nbsp;Liang Ma ,&nbsp;Youshe Ren ,&nbsp;Lei Shi","doi":"10.1016/j.theriogenology.2025.01.003","DOIUrl":"10.1016/j.theriogenology.2025.01.003","url":null,"abstract":"<div><div>This study aimed to investigate the mechanism by which Se in regulates the proliferation and apoptosis of sheep Leydig cells via the miR-200a/NRF pathway. The cells were isolated and purified from the testes of 8-month-old sheep via a Percoll density gradient. After the cells were treated with different concentrations of Se (0, 2.0, 4.0, 6.0, and 8.0 μmol/L of Se) for 18 h, the miR-200a levels was detected. MiR-200a mimics and inhibitors were transfected into the cells, resulting in five groups (control, NC mimics, miR-200a mimics, NC inhibitor and miR-200a inhibitor). Cell viability and antioxidant status were measured via CCK8 and antioxidant assays, respectively. The abundances of pro-apoptotic (<em>BAX</em>, <em>CASPASE 3</em> and <em>CASPASE 8</em>), cell cycle (<em>P21</em>, <em>P27</em> and <em>CDK1</em>), and NRF2-related (<em>NRF2</em>, <em>HO-1</em>, <em>NQO1</em> and <em>KEAP1</em>) genes were detected by real-time PCR and Western blot analysis.</div><div>The results revealed that miR-200a mimics group presented greater (<em>P</em> &lt; 0.05) abundances of <em>NRF2</em>, <em>HO-1</em> and <em>NQO1</em> mRNA transcripts and proteins. Compared with those both in the NC mimics and the miR-200a inhibitor groups, the activities of GSH-Px and SOD, as well as cell viability in the miR-200a mimics group were significantly greater (<em>P</em> &lt; 0.05). In contrast, the ROS levels, MDA content and abundances of <em>KEAP1</em>, <em>P21</em>, <em>P27</em> and apoptosis-related genes mRNA transcripts and proteins were decreased (<em>P</em> &lt; 0.05). The highest (<em>P</em> &lt; 0.05) miR-200a expression level was detected in the Se<sub>6.0</sub> group. Compared with that in the Se (6.0 μmol/L) group, cell viability in the Se + miR-200a inhibitor group was lower (<em>P</em> &lt; 0.05). The abundances of <em>NRF2</em>, <em>HO-1</em> and <em>NQO1</em> in the Se + miR-200a inhibitor group were lower (<em>P</em> &lt; 0.05) than those in the Se (6.0 μmol/L) group but greater (<em>P</em> &lt; 0.05) than those in the inhibitor group, while <em>KEAP1</em> displayed the opposite trend (<em>P</em> &lt; 0.05).</div><div>These results indicate that Se can activate the NRF2 antioxidant signaling pathway to regulate the proliferation and apoptosis of sheep Leydig cells and that miR-200a plays a vital role in this process. The regulatory effect of Se on male reproduction and spermatogenesis may be related to the number of Leydig cells. This study aimed to provide experimental data for Se regulation of spermatogenesis.</div></div>","PeriodicalId":23131,"journal":{"name":"Theriogenology","volume":"235 ","pages":"Pages 103-113"},"PeriodicalIF":2.4,"publicationDate":"2025-01-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142984804","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Relationships between activity monitoring device data and ovarian, uterine, hormonal, and pregnancy variables in beef cows 肉牛活动监测装置数据与卵巢、子宫、激素和妊娠变量之间的关系。
IF 2.4 2区 农林科学 Q3 REPRODUCTIVE BIOLOGY Pub Date : 2025-01-06 DOI: 10.1016/j.theriogenology.2025.01.001
Cecilia Constantino Rocha , Ana Beatriz Montevecchio , Alexandra Bennett , Abdul Waheed , Mackenzie Mazziotta , Tatiane S. Maia , McKenzie Lane-Jackson Haimon , Quinn A. Hoorn , Masroor Sagheer , Camila J. Cuellar , Oscar Alejandro Ojeda-Rojas , Rebecca Lynn Krisher , Marcello Rubessa , Ky G. Pohler , Peter J. Hansen , Philipe Moriel , Ricardo C. Chebel , Mario Binelli
Implementing accelerometer technologies in beef operations is an alternative to increase precision in estrous detection. We hypothesized that (1) the accelerometer algorithm has similar accuracy in detecting behavioral estrus as does visual observation of pressure-sensitive sensors (estrus patches) in grazing beef cows; (2) variables measured by the accelerometer, such as estrus intensity, are associated with hormonal, ovarian, and uterine variables monitored before, during, and after estrus; and (3) the accelerometer variables are associated with the probability of pregnancy in grazing beef cows submitted to embryo transfer (ET). Fifty cows were fitted with accelerometer and patches to detect estrus after a synchronization protocol in eight subsequent rounds. For each round, only cows that showed estrus (day 0; D0) received ET. Follicular diameter, endometrial thickness, corpus luteum (CL) area, and estradiol (E2) and progesterone (P4) concentrations were measured during proestrus, estrus, and early diestrus. On D7, ET was performed. Pregnancies were diagnosed on D46 and cows recovered for 35D before a new replicate. Patches had a greater accuracy (98 % vs. 91 %) of detection of behavioral estrus than accelerometer algorithm. Cows with lower estrus intensity in the accelerometer had greater follicular diameter on D0 (P = 0.022), CL area on D4 and D7 (P = 0.05), endometrial thickness on D-1 (P = 0.10), and reduced E2 concentrations on D-1 (P = 0.0032). The accelerometer variables did not predict accurately the probability of pregnancy/ET. In conclusion, visual observation of patches was more accurate in detecting estrus than the accelerometer algorithm and most of the associations between accelerometers and physiological variables were for characteristics measured at proestrus.
在牛肉操作中实施加速度计技术是提高发情检测精度的另一种选择。我们假设:(1)加速度计算法在检测放牧肉牛的行为发情方面与视觉观察压力敏感传感器(发情斑块)具有相似的准确性;(2)加速度计测量的变量,如发情强度,与发情之前、期间和之后监测的激素、卵巢和子宫变量相关;(3)加速度计变量与胚胎移植(ET)放牧肉牛的怀孕概率相关。在随后的8轮同步协议后,50头奶牛安装了加速计和补丁来检测发情。每轮,只有发情的奶牛(第0天;分别于发情前期、发情期和早衰期测定卵泡直径、子宫内膜厚度、黄体面积、雌二醇(E2)和黄体酮(P4)浓度。D7行ET。在第46天诊断出怀孕,奶牛在35天恢复,然后进行新的复制。与加速度计算法相比,补丁在检测行为发情方面具有更高的准确性(98%对91%)。加速度计显示,发情强度较低的奶牛在D0时卵泡直径增大(P = 0.022), D4和D7时CL面积增大(P = 0.05), D-1时子宫内膜厚度增大(P = 0.10), D-1时E2浓度降低(P = 0.0032)。加速度计变量不能准确预测怀孕/ET的概率。综上所述,视觉观察斑块比加速度计算法更准确地检测发情,加速度计与生理变量之间的关联大部分是在发情前测量的特征。
{"title":"Relationships between activity monitoring device data and ovarian, uterine, hormonal, and pregnancy variables in beef cows","authors":"Cecilia Constantino Rocha ,&nbsp;Ana Beatriz Montevecchio ,&nbsp;Alexandra Bennett ,&nbsp;Abdul Waheed ,&nbsp;Mackenzie Mazziotta ,&nbsp;Tatiane S. Maia ,&nbsp;McKenzie Lane-Jackson Haimon ,&nbsp;Quinn A. Hoorn ,&nbsp;Masroor Sagheer ,&nbsp;Camila J. Cuellar ,&nbsp;Oscar Alejandro Ojeda-Rojas ,&nbsp;Rebecca Lynn Krisher ,&nbsp;Marcello Rubessa ,&nbsp;Ky G. Pohler ,&nbsp;Peter J. Hansen ,&nbsp;Philipe Moriel ,&nbsp;Ricardo C. Chebel ,&nbsp;Mario Binelli","doi":"10.1016/j.theriogenology.2025.01.001","DOIUrl":"10.1016/j.theriogenology.2025.01.001","url":null,"abstract":"<div><div>Implementing accelerometer technologies in beef operations is an alternative to increase precision in estrous detection. We hypothesized that (1) the accelerometer algorithm has similar accuracy in detecting behavioral estrus as does visual observation of pressure-sensitive sensors (estrus patches) in grazing beef cows; (2) variables measured by the accelerometer, such as estrus intensity, are associated with hormonal, ovarian, and uterine variables monitored before, during, and after estrus; and (3) the accelerometer variables are associated with the probability of pregnancy in grazing beef cows submitted to embryo transfer (ET). Fifty cows were fitted with accelerometer and patches to detect estrus after a synchronization protocol in eight subsequent rounds. For each round, only cows that showed estrus (day 0; D0) received ET. Follicular diameter, endometrial thickness, corpus luteum (CL) area, and estradiol (E2) and progesterone (P4) concentrations were measured during proestrus, estrus, and early diestrus. On D7, ET was performed. Pregnancies were diagnosed on D46 and cows recovered for 35D before a new replicate. Patches had a greater accuracy (98 % vs. 91 %) of detection of behavioral estrus than accelerometer algorithm. Cows with lower estrus intensity in the accelerometer had greater follicular diameter on D0 (P = 0.022), CL area on D4 and D7 (P = 0.05), endometrial thickness on D-1 (P = 0.10), and reduced E2 concentrations on D-1 (P = 0.0032). The accelerometer variables did not predict accurately the probability of pregnancy/ET. In conclusion, visual observation of patches was more accurate in detecting estrus than the accelerometer algorithm and most of the associations between accelerometers and physiological variables were for characteristics measured at proestrus.</div></div>","PeriodicalId":23131,"journal":{"name":"Theriogenology","volume":"235 ","pages":"Pages 64-74"},"PeriodicalIF":2.4,"publicationDate":"2025-01-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142967048","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
In vitro study of carbetocin, an oxytocin receptor agonist, and 4-phenylfuroxan-3-carbonitrile, a NO-releasing agent, as cervical dilatators in sheep
IF 2.4 2区 农林科学 Q3 REPRODUCTIVE BIOLOGY Pub Date : 2025-01-06 DOI: 10.1016/j.theriogenology.2025.01.002
García-Barcelo Pedro , Romero Angel , Rodríguez-Piñón Marcelo , Cerecetto Hugo
The aim was to study the effect of 4-phenylfuroxan-3-carbonitrile (Fx), a NO-releasing agent, and carbetocin, an oxytocin receptor agonist, on matrix metalloproteinases-2 (MMP-2) activity and PGE2 production in cervix from cycling sheep. Cervical explants were incubated during 12 h with MEM supplemented with increasing concentrations of Fx in DMSO (2 %) (0 to 300 μg/mL) with Cb (100 ng/mL) (Experiment 1, n = 15) and DMSO (2 %), DMSO + Cb (100 ng/mL) or DMSO + Fx (30 μg/mL) (Experiment 2, n = 10), and their respective controls. In the supernatants, activated (A) and latent (L) MMP-2 activities were determined by a SDS-PAGE zymography, PGE2 concentration by immunoassay and NO production indirectly as nitrites by spectrophotometry. Data were analyzed by ANOVA. The Cb treatment increase the A MMP-2 activity in DMSO (Experiment 1 at follicular phase and Experiment 2) or alone (Experiment 2) and increase the L MMP-2 activity (Experiments 1 and 2) (P < 0.02). The DMSO treatment also increase the L MMP2 activity (Experiment 2) (P < 0.0001). Treatment with Fx + DMSO increased the concentration of accumulated nitrites in the supernatant (P < 0.0001) (Experiment 1), but did not affect or decrease the activity of A and L MMP-2 (P < 0.04) (Experiments 1 and 2). The PGE2 concentration trend to increase with Cb treatment (P = 0.0614) and decrease with Fx+DMSO treatment (P < 0.0001) (Experiment 2). In conclusion, Cb and/or DMSO treatment of cervical explants increase the MMP-2 activity through PGE2-independent mechanisms, but Fx in DMSO fail in this, suggesting that the pre-treatment with Cb and/or DMSO could be used to increase cervical dilation in ewes.
{"title":"In vitro study of carbetocin, an oxytocin receptor agonist, and 4-phenylfuroxan-3-carbonitrile, a NO-releasing agent, as cervical dilatators in sheep","authors":"García-Barcelo Pedro ,&nbsp;Romero Angel ,&nbsp;Rodríguez-Piñón Marcelo ,&nbsp;Cerecetto Hugo","doi":"10.1016/j.theriogenology.2025.01.002","DOIUrl":"10.1016/j.theriogenology.2025.01.002","url":null,"abstract":"<div><div>The aim was to study the effect of 4-phenylfuroxan-3-carbonitrile (Fx), a NO-releasing agent, and carbetocin, an oxytocin receptor agonist, on matrix metalloproteinases-2 (MMP-2) activity and PGE2 production in cervix from cycling sheep. Cervical explants were incubated during 12 h with MEM supplemented with increasing concentrations of Fx in DMSO (2 %) (0 to 300 μg/mL) with Cb (100 ng/mL) (Experiment 1, n = 15) and DMSO (2 %), DMSO + Cb (100 ng/mL) or DMSO + Fx (30 μg/mL) (Experiment 2, n = 10), and their respective controls. In the supernatants, activated (A) and latent (L) MMP-2 activities were determined by a SDS-PAGE zymography, PGE2 concentration by immunoassay and NO production indirectly as nitrites by spectrophotometry. Data were analyzed by ANOVA. The Cb treatment increase the A MMP-2 activity in DMSO (Experiment 1 at follicular phase and Experiment 2) or alone (Experiment 2) and increase the L MMP-2 activity (Experiments 1 and 2) (P &lt; 0.02). The DMSO treatment also increase the L MMP2 activity (Experiment 2) (P &lt; 0.0001). Treatment with Fx + DMSO increased the concentration of accumulated nitrites in the supernatant (P &lt; 0.0001) (Experiment 1), but did not affect or decrease the activity of A and L MMP-2 (P &lt; 0.04) (Experiments 1 and 2). The PGE2 concentration trend to increase with Cb treatment (P = 0.0614) and decrease with Fx+DMSO treatment (P &lt; 0.0001) (Experiment 2). In conclusion, Cb and/or DMSO treatment of cervical explants increase the MMP-2 activity through PGE2-independent mechanisms, but Fx in DMSO fail in this, suggesting that the pre-treatment with Cb and/or DMSO could be used to increase cervical dilation in ewes.</div></div>","PeriodicalId":23131,"journal":{"name":"Theriogenology","volume":"235 ","pages":"Pages 168-174"},"PeriodicalIF":2.4,"publicationDate":"2025-01-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143024767","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
ReBreed21-ET: Evaluation of a rapid resynchronization program that allows timed embryo transfer every 21 days ReBreed21-ET:快速再同步程序的评估,允许每21天定时胚胎移植。
IF 2.4 2区 农林科学 Q3 REPRODUCTIVE BIOLOGY Pub Date : 2025-01-02 DOI: 10.1016/j.theriogenology.2024.12.031
João Paulo N. Andrade , Victor E. Gomez-León , Guilherme Madureira , Luma C. Sartori , Gustavo F. Grillo , Rafael R. Domingues , Meliton Fosado , Rodrigo V. Sala , Milo C. Wiltbank
This study evaluated the efficiency of a rapid resynchronization of ovulation program to allow timed embryo transfer (TET) every 21d in heifer embryo recipients. Holstein heifers (n = 510) had synchronized ovulation using a modified 5d CoSynch program for a TET (D7) after induced ovulation (D0). After TET, heifers were blocked by number of previous TETs and randomized into one of two resynchronization of ovulation programs: Resynch28 (n = 279), a traditional resynch program for TET 35d after previous TET; or ReBreed21-ET, a program designed to allow TET 21d after previous TET. Once assigned into one of the two programs, heifers were kept in the same program during a 105-d TET period. In Resynch28, heifers received an intravaginal progesterone (P4) insert on D28, on D33 the P4 was removed, and pregnancy diagnosis was performed using rectal ultrasound to determine embryonic heartbeat. Nonpregnant heifers received a PGF2α treatment with a later GnRH treatment on D35 for a potential TET on D42 (35d after previous TET). Heifers in ReBreed21-TET received an intravaginal P4 insert on D14, on D19 the P4 was removed, and on D21 a GnRH treatment was given to synchronize a new ovulation. On D28, pregnancy diagnosis was performed using transrectal ultrasound to detect an embryonic heartbeat and nonpregnant heifers that had a corpus luteum (CL) ≥18 mm in diameter received a TET (21d after the previous TET). Pregnancy per ET (P/ET) from the first TET was greater for heifers in ReBreed21-ET (52 %) than Resynch28 (39.4 %). In contrast, the subsequent TET (second and later) had similar P/ET for ReBreed21-ET (40.4 %) and Resynch28 (40.8 %). The overall pregnancy loss from D28 to D63 did not differ between programs (ReBreed21-ET [18.5 %] and Resynch28 (16.3 %]). Nevertheless, there were fewer pregnancy losses from D28 to D33 for Resynch28 (3.5 %) than ReBreed21-ET (10.1 %), while from D33 to D47, there was greater pregnancy loss for Resynch28 (10.1 %) than ReBreed21-ET (4.9 %). Time to pregnancy was approximately 8d earlier for ReBreed21-ET (45 ± 3d) than Resynch28 (53 ± 3d). Overall cumulative pregnancies at the end of a 105-d TET season were greater for ReBreed21-ET (75.2 %) than Resynch28 (64 %). Thus, the ReBreed21-ET program can improve the efficiency of TET programs.
本研究评估了在母牛胚胎受体中每21天进行定时胚胎移植(TET)的快速再同步排卵程序的效率。在诱导排卵(D0)后,使用改良的5d cosync程序进行TET (D7)同步排卵(n = 510)。TET后,根据先前TET的数量,将母牛随机分配到两个重新同步排卵计划中的一个:Resynch28 (n = 279),这是一个传统的TET再同步计划,在上次TET后35d进行;或ReBreed21-ET,这是一个旨在允许在前一次TET之后进行TET 21天的程序。一旦分配到两个项目中的一个,小母牛在105天的TET期间被保持在同一个项目中。在Resynch28中,母牛在D28时接受阴道内孕酮(P4)插入,在D33时取出P4,并通过直肠超声检测胚胎心跳进行妊娠诊断。未怀孕的母牛在第35天接受PGF2α治疗和随后的GnRH治疗,以期在第42天(前一次TET后35天)进行潜在的TET治疗。ReBreed21-TET组的小母牛在第14天阴道内插入P4,在第19天移除P4,在第21天给予GnRH治疗以同步新的排卵。D28妊娠诊断采用经直肠超声检测胚胎心跳,对黄体(CL)直径≥18 mm的未妊娠母牛进行TET(在前一次TET后21d)。ReBreed21-ET第一次TET的妊娠率(P/ET)高于Resynch28(39.4%)(52%)。相比之下,ReBreed21-ET(40.4%)和Resynch28(40.8%)的后续TET(第二次及以后)的P/ET相似。从D28到D63的总体妊娠损失在两个方案之间没有差异(ReBreed21-ET[18.5%]和Resynch28(16.3%])。然而,从D28到D33, Resynch28的妊娠损失(3.5%)比ReBreed21-ET(10.1%)少,而从D33到D47, Resynch28的妊娠损失(10.1%)比ReBreed21-ET(4.9%)大。ReBreed21-ET的妊娠时间(45±3d)比Resynch28(53±3d)早约8d。在105 d TET季节结束时,ReBreed21-ET的总累积妊娠率(75.2%)高于Resynch28(64%)。因此,ReBreed21-ET计划可以提高TET计划的效率。
{"title":"ReBreed21-ET: Evaluation of a rapid resynchronization program that allows timed embryo transfer every 21 days","authors":"João Paulo N. Andrade ,&nbsp;Victor E. Gomez-León ,&nbsp;Guilherme Madureira ,&nbsp;Luma C. Sartori ,&nbsp;Gustavo F. Grillo ,&nbsp;Rafael R. Domingues ,&nbsp;Meliton Fosado ,&nbsp;Rodrigo V. Sala ,&nbsp;Milo C. Wiltbank","doi":"10.1016/j.theriogenology.2024.12.031","DOIUrl":"10.1016/j.theriogenology.2024.12.031","url":null,"abstract":"<div><div>This study evaluated the efficiency of a rapid resynchronization of ovulation program to allow timed embryo transfer (TET) every 21d in heifer embryo recipients. Holstein heifers (n = 510) had synchronized ovulation using a modified 5d CoSynch program for a TET (D7) after induced ovulation (D0). After TET, heifers were blocked by number of previous TETs and randomized into one of two resynchronization of ovulation programs: Resynch28 (n = 279), a traditional resynch program for TET 35d after previous TET; or ReBreed21-ET, a program designed to allow TET 21d after previous TET. Once assigned into one of the two programs, heifers were kept in the same program during a 105-d TET period. In Resynch28, heifers received an intravaginal progesterone (P4) insert on D28, on D33 the P4 was removed, and pregnancy diagnosis was performed using rectal ultrasound to determine embryonic heartbeat. Nonpregnant heifers received a PGF2α treatment with a later GnRH treatment on D35 for a potential TET on D42 (35d after previous TET). Heifers in ReBreed21-TET received an intravaginal P4 insert on D14, on D19 the P4 was removed, and on D21 a GnRH treatment was given to synchronize a new ovulation. On D28, pregnancy diagnosis was performed using transrectal ultrasound to detect an embryonic heartbeat and nonpregnant heifers that had a corpus luteum (CL) ≥18 mm in diameter received a TET (21d after the previous TET). Pregnancy per ET (P/ET) from the first TET was greater for heifers in ReBreed21-ET (52 %) than Resynch28 (39.4 %). In contrast, the subsequent TET (second and later) had similar P/ET for ReBreed21-ET (40.4 %) and Resynch28 (40.8 %). The overall pregnancy loss from D28 to D63 did not differ between programs (ReBreed21-ET [18.5 %] and Resynch28 (16.3 %]). Nevertheless, there were fewer pregnancy losses from D28 to D33 for Resynch28 (3.5 %) than ReBreed21-ET (10.1 %), while from D33 to D47, there was greater pregnancy loss for Resynch28 (10.1 %) than ReBreed21-ET (4.9 %). Time to pregnancy was approximately 8d earlier for ReBreed21-ET (45 ± 3d) than Resynch28 (53 ± 3d). Overall cumulative pregnancies at the end of a 105-d TET season were greater for ReBreed21-ET (75.2 %) than Resynch28 (64 %). Thus, the ReBreed21-ET program can improve the efficiency of TET programs.</div></div>","PeriodicalId":23131,"journal":{"name":"Theriogenology","volume":"235 ","pages":"Pages 145-151"},"PeriodicalIF":2.4,"publicationDate":"2025-01-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143012172","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
GM-CSF treatment of frozen bovine sperm improves function, fertilization, and subsequent embryo development
IF 2.4 2区 农林科学 Q3 REPRODUCTIVE BIOLOGY Pub Date : 2025-01-02 DOI: 10.1016/j.theriogenology.2024.12.022
Annie L. Whitty , Karen L. Kind , Kylie R. Dunning , Nicole O. McPherson , Mark B. Nottle
In vitro embryo production (IVP) is used in the cattle industry to increase the rate of genetic gain. IVP uses semen that has been frozen and thawed, a process that renders sperm less viable than sperm from fresh semen. Granulocyte macrophage colony stimulating factor (GM-CSF) is present in bovine seminal plasma, while its receptor is present on bovine sperm. The present study aimed to determine if GM-CSF could improve the function and quality of frozen-thawed bovine sperm and embryo development following in vitro fertilization. Thawed bovine sperm (n = 3 bulls with 3 replicates per bull) was incubated with 0, 0.1, 1, 2 or 10 ng/ml of recombinant bovine GM-CSF in buffered wash media for 45 min and assessed for motility, glucose uptake, mitochondrial activity, intracellular calcium, capacitation, DNA integrity, and in vitro embryo development. The addition of 1, 2, and 10 ng/ml GM-CSF increased total motility (P = 0.02, P = 0.007, P = 0.01), progressive (P = 0.02, P = 0.03, P = 0.01), and rapid motility (P = 0.01, P = 0.01, P = 0.01), while 10 ng/ml increased glucose uptake (P = 0.003), and 1, 2, and 10 ng/ml increased capacitation (P = 0.003, P = 0.001, P = 0.0003). There was no difference between groups for mitochondrial activity, intracellular calcium, or DNA integrity. GM-CSF treatment of sperm prior to in vitro insemination increased fertilization rate (P = 0.01), hatching blastocyst rate (P = 0.05), and blastocyst inner cell mass cell number (P = 0.03) compared with control. In conclusion, GM-CSF treatment of frozen-thawed bovine sperm improves sperm function and quality resulting in increased fertilization capacity and subsequent embryo development, suggesting it may improve cattle IVP efficiencies.
{"title":"GM-CSF treatment of frozen bovine sperm improves function, fertilization, and subsequent embryo development","authors":"Annie L. Whitty ,&nbsp;Karen L. Kind ,&nbsp;Kylie R. Dunning ,&nbsp;Nicole O. McPherson ,&nbsp;Mark B. Nottle","doi":"10.1016/j.theriogenology.2024.12.022","DOIUrl":"10.1016/j.theriogenology.2024.12.022","url":null,"abstract":"<div><div>In vitro embryo production (IVP) is used in the cattle industry to increase the rate of genetic gain. IVP uses semen that has been frozen and thawed, a process that renders sperm less viable than sperm from fresh semen. Granulocyte macrophage colony stimulating factor (GM-CSF) is present in bovine seminal plasma, while its receptor is present on bovine sperm. The present study aimed to determine if GM-CSF could improve the function and quality of frozen-thawed bovine sperm and embryo development following in vitro fertilization. Thawed bovine sperm (n = 3 bulls with 3 replicates per bull) was incubated with 0, 0.1, 1, 2 or 10 ng/ml of recombinant bovine GM-CSF in buffered wash media for 45 min and assessed for motility, glucose uptake, mitochondrial activity, intracellular calcium, capacitation, DNA integrity, and in vitro embryo development. The addition of 1, 2, and 10 ng/ml GM-CSF increased total motility (P = 0.02, P = 0.007, P = 0.01), progressive (P = 0.02, P = 0.03, P = 0.01), and rapid motility (P = 0.01, P = 0.01, P = 0.01), while 10 ng/ml increased glucose uptake (P = 0.003), and 1, 2, and 10 ng/ml increased capacitation (P = 0.003, P = 0.001, P = 0.0003). There was no difference between groups for mitochondrial activity, intracellular calcium, or DNA integrity. GM-CSF treatment of sperm prior to in vitro insemination increased fertilization rate (P = 0.01), hatching blastocyst rate (P = 0.05), and blastocyst inner cell mass cell number (P = 0.03) compared with control. In conclusion, GM-CSF treatment of frozen-thawed bovine sperm improves sperm function and quality resulting in increased fertilization capacity and subsequent embryo development, suggesting it may improve cattle IVP efficiencies.</div></div>","PeriodicalId":23131,"journal":{"name":"Theriogenology","volume":"235 ","pages":"Pages 46-55"},"PeriodicalIF":2.4,"publicationDate":"2025-01-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143130055","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
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Theriogenology
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