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19‐Norandrostenedione (4‐estrene‐3,17‐dione) Inhibits Porcine Oocyte Maturation In Vitro 19‐去甲雄烯二酮(4‐estene‐3,17‐dione)抑制猪卵母细胞体外成熟
Pub Date : 1986-02-01 DOI: 10.1002/MRD.1120130210
S. Daniel, M. Khalil, D. Armstrong
Recent work has shown that 19-norandrostenedione is a major steroidal component of porcine follicular fluid; however, little is known about its role(s) in the regulation of follicular function. This study was designed to examine the effect of 19-norandrostenedione on porcine oocyte maturation in vitro. Oocyte-cumulus complexes were isolated from medium (3–6-mm diameter)-sized prepubertal pig follicles and incubated for 12 h in medium with or without dibutyryl cyclic AMP ((Bu)2cAMP, 1 mM) with or without testosterone (5 x 10−7 M) or 19-norandrostenedione (5 x 10−7 M). In medium alone, 70.8% of oocytes spontaneously resumed meiosis as evidenced by the occurrence of germinal vesicle breakdown. Oocyte maturation was inhibited by (Bu)2cAMP (44.6% of oocytes matured). Although neither steroid alone affected maturation, both testosterone and 19-norandrostenedione enhanced the effect of (Bu)2cAMP (22.5 and 19.6%, respectively, resumed meiosis). The effects of testosterone and 19-norandrostenedione on (Bu)2cAMP-inhibited oocyte maturation were dose dependent and there was no significant difference between the actions of the steroids. The effect of 19-norandrostenedione was reversible and dependent on the presence of an intact cumulus. Hydroxyflutamide (SCH-16423), a nonsteroidal compound known to block androgen receptors, abolished the effects of both testosterone and 19-norandrostenedione on germinal vesicle breakdown, indicating that the actions of these steroids are truly androgenic. The results of this study suggest that 19-norandrostenedione may be of physiological importance in the regulation of porcine oocyte maturation.
最近的研究表明,19-去甲雄烯二酮是猪卵泡液的主要甾体成分;然而,对其在卵泡功能调节中的作用知之甚少。本研究旨在探讨19-去甲雄烯二酮对猪卵母细胞体外成熟的影响。从中等(直径3 - 6毫米)大小的青春期前猪卵泡中分离卵母细胞-卵丘复合物,在有或没有二丁基环AMP ((Bu)2cAMP, 1毫米)和有或没有睾酮(5 × 10−7米)或19-去甲雄烯二酮(5 × 10−7米)的培养基中孵育12小时。仅在培养基中,70.8%的卵母细胞自发恢复减数分裂,这表明发生了生发囊泡破裂。(Bu)2cAMP抑制卵母细胞成熟(44.6%成熟)。虽然类固醇单独影响成熟,但睾酮和19-去甲雄烯二酮都增强了(Bu)2cAMP的作用(分别为22.5%和19.6%)。睾酮和19-去甲雄烯二酮对(Bu) 2camp抑制的卵母细胞成熟的影响呈剂量依赖性,两种激素的作用无显著差异。19-去甲雄烯二酮的作用是可逆的,并且依赖于完整积云的存在。羟氟他胺(SCH-16423)是一种已知能阻断雄激素受体的非甾体化合物,它能消除睾丸激素和19-去甲雄烯二酮对生发囊泡破裂的影响,表明这些类固醇的作用确实是雄激素性的。本研究结果提示,19-去甲雄烯二酮可能在调节猪卵母细胞成熟过程中具有重要的生理意义。
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引用次数: 11
Adenosine and gpp(NH)p modulate mouse sperm adenylate cyclase 腺苷和gpp(NH)p调节小鼠精子腺苷酸环化酶
Pub Date : 1986-02-01 DOI: 10.1002/MRD.1120130208
D. M. Stein, L. Fraser, N. Monks
The possible roles of adenosine and the GTP analogue Gpp(NH)p in regulating mouse sperm adenylate cyclase activity were investigated during incubation in vitro under conditions in which after 30 min the spermatozoa are essentially uncapacitated and poorly fertile, whereas after 120 min they are capacitated and highly fertile. Adenylate cyclase activity, assayed in the presence of 1 mM ATP and 2 mM Mn2+, was determined by monitoring cAMP production. When adenosine deaminase (1 U/ml) was included in the assay to deplete endogenous adenosine, enzyme activity was decreased in the 30-min suspensions but increased in the 120-min samples (P 10−5 M) of adenosine. Similar inhibition was also observed in the absence of Gpp(NH)p, suggesting the presence of an inhibitory P site on the enzyme. In further experiments, the effects of Gpp(NH)p in the presence and absence of adenosine deaminase were examined. Activity in 30-min suspensions was stimulated by the guanine nucleotide and in the presence of adenosine deaminase this stimulation was marked, reversing the inhibition seen with adenosine deaminase alone. In capacitated suspensions the opposite profile was observed, with Gpp(NH)p plus adenosine deaminase being inhibitory; again, this was a reversal of the effects obtained in the presence of adenosine deaminase alone, which had stimulated enzyme activity. These results suggest the existence of a stimulatory adenosine receptor site (Ra) on mouse sperm adenylate cyclase that is expressed in uncapacitated spermatozoa and an inhibitory receptor site (Ri) that is expressed in capacitated cells, with guanine nucleotides modifying the final response to adenosine. It is concluded that adenosine and guanine nucleotides may regulate mouse sperm adenylate cyclase activity during capacitation.
在体外培养条件下,研究了腺苷和GTP类似物Gpp(NH)p在调节小鼠精子腺苷酸环化酶活性中的可能作用。在体外培养条件下,精子在30分钟后基本丧失能力和低生育能力,而在120分钟后则具有能力和高生育能力。腺苷酸环化酶活性,在1mm ATP和2mm Mn2+存在下测定,通过监测cAMP的产生来确定。当加入腺苷脱氨酶(1 U/ml)以消耗内源性腺苷时,酶活性在30分钟的悬液中下降,但在120分钟的腺苷样品(P 10−5 M)中增加。在Gpp(NH)p缺失的情况下也观察到类似的抑制作用,这表明在酶上存在抑制p位点。在进一步的实验中,我们考察了Gpp(NH)p在腺苷脱氨酶存在和不存在时的作用。鸟嘌呤核苷酸刺激了30分钟悬液的活性,在腺苷脱氨酶存在的情况下,这种刺激是明显的,逆转了单独使用腺苷脱氨酶所见的抑制作用。在能化悬液中观察到相反的情况,Gpp(NH)p +腺苷脱氨酶具有抑制作用;再一次,这与单独存在腺苷脱氨酶时获得的效果相反,后者刺激了酶的活性。这些结果表明,小鼠精子腺苷酸环化酶上存在一个在失能精子中表达的刺激性腺苷受体位点(Ra)和一个在失能细胞中表达的抑制性受体位点(Ri),其中鸟嘌呤核苷酸修饰了对腺苷的最终反应。由此可见,腺苷和鸟嘌呤核苷酸可能在获能过程中调控小鼠精子腺苷酸环化酶活性。
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引用次数: 43
A comparison of fertility in vitro and in vivo after exposure of male mice to high environmental pressure 雄性小鼠暴露于高环境压力后体外和体内生育能力的比较
Pub Date : 1986-02-01 DOI: 10.1002/MRD.1120130207
L. Fraser, S. Monk, B. Wardley‐Smith, S. B. Cohen, M. Halsey
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引用次数: 2
Oviducal pars recta‐induced degradation of vitelline coat proteins in relation to acquisition of fertilizability of toad eggs 卵磷脂诱导卵黄膜蛋白降解与蟾蜍卵获得受精性的关系
Pub Date : 1986-01-01 DOI: 10.1002/MRD.1120140305
K. Takamune, N. Yoshizaki, C. Katagiri
In the toad Bufo bufo japonicus the vitelline coat (VC) of the uterine egg (UEVC) is more readily lysed by the sperm lysin than the VC of coelomic egg (CEVC). Fluorometric determinations of released proteins after incubation of the VC with the sperm lysin in vitro revealed that the CEVC is not completely refractory to the lysin but increases in susceptibility after treatment with a pars recta extract (PRE). Experiments employing isolated pars recta granules showed that both this increase of VC susceptibility and the acquisition of egg fertilizability are ascribable to the contents of the granules. SDS-PAGE analyses of VC proteins revealed that in comparison with CEVC, UEVC lacks 40–52K proteins concomitant with the increased stainability of 39K protein and the appearance of 36K protein. These changes in SDS-PAGE profiles were observed either in oviducal eggs after passage through the pars recta or in coelomic eggs treated with PRE but were inhibited when coelomic eggs were treated with PRE containing soybean trypsin inhibitor (SBTI) and leupeptin. Likewise, the acquisition of fertilizability by treatment of coelomic egg with PRE was inhibited by SBTI. Dejellied uterine eggs were successfully fertilized when pretreated with trypsin inhibitors before insemination but were not fertilized when pre-treated with concanavalin A. We propose that the hydrolytic degradation of certain VC proteins due to the tryptic activity of pars recta granules renders the VC susceptible to the sperm lysin, so that the eggs are made receptive to a fertilizing sperm.
蟾(Bufo Bufo japonicus)子宫卵(UEVC)卵黄膜(vitelline coat, VC)比体腔卵(coelomic egg, CEVC)卵黄膜(vitelline coat, VC)更容易被精子溶酶溶解。体外对VC与精子溶素孵育后释放的蛋白质进行荧光测定,发现CEVC对溶素并非完全不耐受,但在用直肠精提取物(PRE)处理后,其敏感性增加。实验采用孤立pars直肠颗粒表明VC易感性的增加和收购蛋可受精是由于颗粒的内容。VC蛋白的SDS-PAGE分析显示,与CEVC相比,UEVC缺乏40-52K蛋白,同时39K蛋白的染色性增加,36K蛋白出现。这些SDS-PAGE谱的变化既发生在通过直肠部的卵中,也发生在经PRE处理的体腔卵中,但在经含有大豆胰蛋白酶抑制剂(SBTI)和胰肽的PRE处理的体腔卵中,SDS-PAGE谱的变化受到抑制。同样,用PRE处理体腔卵获得受精率也被SBTI抑制。在授精前用胰蛋白酶抑制剂预处理后,脱胶的子宫卵可以成功受精,但用豆豆蛋白a预处理后则不能受精。我们认为,由于直肠蛋白颗粒的胰蛋白酶活性,某些VC蛋白的水解降解使VC对精子溶酶敏感,因此卵子可以接受受精的精子。
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引用次数: 36
From the cellular to the molecular dimension: the actual challenge for human fertilization research 从细胞到分子维度:人类受精研究的实际挑战
Pub Date : 1986-01-01 DOI: 10.1002/MRD.1120130106
J. Tesarik
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引用次数: 21
The mechanism of cumulus cell-oocyte uncoupling: evidence for the participation of both cumulus cells and oocytes. 积云细胞-卵母细胞解偶联的机制:积云细胞和卵母细胞共同参与的证据。
Pub Date : 1982-01-01 DOI: 10.1002/MRD.1120060208
J. Eppig, P. Ward-Bailey
Cumulus cells are metabolically coupled to oocytes via heterologous gap junctions. This coupling terminates near the time of ovulation, and the termination appears to be correlated with the mucification of the cumulus cells lying immediately adjacent to the oocytes. The first objective of this project was to determine whether follicle stimulating hormone (FSH) induction of cumulus cell-oocyte uncoupling could occur independently of FSH-stimulated cumulus mucification (expansion). Intercellular coupling was measured as a percentage of radiolabeled choline (or its metabolites) that was incorporated into the oocyte relative to the total amount of radiolabel incorporated into the entire cumulus cell-oocyte complex. It was found that the complete suppression of FSH-stimulated cumulus expansion with chondroitin sulfate B had no suppressive effect on FSH-stimulated cumulus cell-oocyte uncoupling. This finding showed that FSH-stimulated cumulus expansion was not required for cumulus cell-oocyte uncoupling. Since 17β-estradiol, testosterone, or progesterone could not induce maximal cumulus cell uncoupling, it was concluded that the uncoupling-promoting action of FSH was probably not mediated by steroid hormones. A partial uncoupling of cumulus cells and oocytes was found when spontaneous oocyte maturation had occurred in the absence of FSH. This partial uncoupling was prevented by incubation of cumulus cell-oocyte complexes in concentrations of dibutyryl cyclic adenosine monophosphate (dbcAMP) or 3-isobutyl-1-methyl xanthine (IBMX) (0.25 and 0.10 mM respectively) that suppressed spontaneous oocyte maturation without inducing cumulus expansion. These inhibitors also prevented the maximal induction of uncoupling that would have been provoked by biological grade preparations of either FSH or luteinizing hormone (LH). It was concluded that two factors were required to bring about maximal cumulus cell-oocyte uncoupling: one factor was dependent upon the action of gonadotropins on cumulus cell function, the other factor appeared to be a function of the oocytes, since maximal uncoupling could occur only after the germinal vesicles had broken down.
卵丘细胞通过异源间隙连接与卵母细胞代谢偶联。这种偶联在排卵时终止,终止似乎与紧邻卵母细胞的积云细胞的粘液化有关。该项目的第一个目的是确定卵泡刺激素(FSH)诱导的积云细胞-卵母细胞解偶联是否可以独立于FSH刺激的积云粘液化(扩张)发生。细胞间偶联测量为放射性标记胆碱(或其代谢物)掺入卵母细胞的百分比,相对于整个卵丘细胞-卵母细胞复合物中放射性标记物的总量。结果发现,硫酸软骨素B完全抑制fsh刺激的积云扩张,对fsh刺激的积云细胞-卵母细胞解耦没有抑制作用。这一发现表明fsh刺激的积云扩张不是积云细胞-卵母细胞解耦所必需的。由于17β-雌二醇、睾酮、孕酮均不能诱导最大程度的卵泡刺激素解偶联,提示促卵泡刺激素的解偶联作用可能不是由类固醇激素介导的。在没有促卵泡刺激素的情况下,卵母细胞自发成熟时,发现卵丘细胞和卵母细胞部分解偶联。在二丁基环腺苷单磷酸(dbcAMP)或3-异丁基-1-甲基黄嘌呤(IBMX)浓度(分别为0.25和0.10 mM)中孵育积云细胞-卵母细胞复合物,抑制卵母细胞的自发成熟,而不诱导积云膨胀,从而阻止了这种部分解偶联。这些抑制剂还能最大限度地防止生物级FSH或黄体生成素(LH)制剂引起的解偶联。结果表明,最大程度的卵-积云解偶联需要两个因素:一个因素依赖于促性腺激素对积云细胞功能的作用,另一个因素似乎是卵母细胞的功能,因为最大程度的解偶联只有在生发囊泡被破坏后才能发生。
{"title":"The mechanism of cumulus cell-oocyte uncoupling: evidence for the participation of both cumulus cells and oocytes.","authors":"J. Eppig, P. Ward-Bailey","doi":"10.1002/MRD.1120060208","DOIUrl":"https://doi.org/10.1002/MRD.1120060208","url":null,"abstract":"Cumulus cells are metabolically coupled to oocytes via heterologous gap junctions. This coupling terminates near the time of ovulation, and the termination appears to be correlated with the mucification of the cumulus cells lying immediately adjacent to the oocytes. The first objective of this project was to determine whether follicle stimulating hormone (FSH) induction of cumulus cell-oocyte uncoupling could occur independently of FSH-stimulated cumulus mucification (expansion). Intercellular coupling was measured as a percentage of radiolabeled choline (or its metabolites) that was incorporated into the oocyte relative to the total amount of radiolabel incorporated into the entire cumulus cell-oocyte complex. It was found that the complete suppression of FSH-stimulated cumulus expansion with chondroitin sulfate B had no suppressive effect on FSH-stimulated cumulus cell-oocyte uncoupling. This finding showed that FSH-stimulated cumulus expansion was not required for cumulus cell-oocyte uncoupling. Since 17β-estradiol, testosterone, or progesterone could not induce maximal cumulus cell uncoupling, it was concluded that the uncoupling-promoting action of FSH was probably not mediated by steroid hormones. A partial uncoupling of cumulus cells and oocytes was found when spontaneous oocyte maturation had occurred in the absence of FSH. This partial uncoupling was prevented by incubation of cumulus cell-oocyte complexes in concentrations of dibutyryl cyclic adenosine monophosphate (dbcAMP) or 3-isobutyl-1-methyl xanthine (IBMX) (0.25 and 0.10 mM respectively) that suppressed spontaneous oocyte maturation without inducing cumulus expansion. These inhibitors also prevented the maximal induction of uncoupling that would have been provoked by biological grade preparations of either FSH or luteinizing hormone (LH). It was concluded that two factors were required to bring about maximal cumulus cell-oocyte uncoupling: one factor was dependent upon the action of gonadotropins on cumulus cell function, the other factor appeared to be a function of the oocytes, since maximal uncoupling could occur only after the germinal vesicles had broken down.","PeriodicalId":12668,"journal":{"name":"Gamete Research","volume":"31 1","pages":"145-154"},"PeriodicalIF":0.0,"publicationDate":"1982-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"88267388","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 40
Circular DNA in human and boar spermatozoa 人类和野猪精子中的环状DNA
Pub Date : 1982-01-01 DOI: 10.1002/MRD.1120050408
J. McGrath, D. Evenson
Circular DNA molecules were isolated from human and boar whole spermatozoa or spermatozoal nuclei and measured for size by electron microscopy. The DNA molecules derived from both mammals were heterogeneous in size ranging from 0.07 to 17 μm; nearly 75% of the molecules were ⩽0.5 μm in length. The mean lengths were 1.0 μm and 1.5 μm for circular DNAs isolated from human and boar spermatozoa, respectively. The origin and function of these molecules remains unknown.
从人和猪的整个精子或精子核中分离出环状DNA分子,并用电子显微镜测量其大小。来自两种哺乳动物的DNA分子大小差异较大,在0.07 ~ 17 μm之间;近75%的分子长度≤0.5 μm。从人和猪精子中分离的环状dna的平均长度分别为1.0 μm和1.5 μm。这些分子的起源和功能尚不清楚。
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引用次数: 3
The effect of nicotine on sperm-Egg interaction in the sea urchin: Polyspermy and electrical events 尼古丁对海胆精子-卵子相互作用的影响:多精和电事件
Pub Date : 1982-01-01 DOI: 10.1002/MRD.1120050203
B. Dale, A. Santis, B. Hagström
We have studied some of the effects of nicotine on sea urchin eggs, spermatozoa, and their interaction using electrical recording techniques and fertilization-rate experiments. Pretreating eggs with nicotine enhances the fertilization rate, whereas this drug has an inhibitory effect on spermatozoa. Pulse-treated eggs or eggs fertilized in the presence of nicotine give rise to attenuated step depolarizations, which may be attributed to a decrease in membrane resistance (Rm) of the egg or, in the latter case, to an alteration to the spermatozoon. Concurrently, with the change in the step depolarization there is a reduction in amplitude of the fertilization potential (FP) suggesting that the cortical reaction is in some way altered. Nicotine has no effect on the Rm of fertilized eggs or oocytes, where there are no cortical granules. We suggest that nicotine alters the cortex of sea urchin eggs–possibly by causing a partial dissolution of cortical granules–which renders the eggs more receptive to spermatozoa. The reductions in amplitude of the step depolarization and the FP are consequences of this alteration.
我们利用电记录技术和受精率实验研究了尼古丁对海胆卵、精子的影响及其相互作用。用尼古丁预处理卵子可以提高受精率,而该药物对精子有抑制作用。脉冲处理的卵子或在尼古丁存在下受精的卵子会产生减弱的阶跃去极化,这可能是由于卵子膜抗性(Rm)的降低,或者在后一种情况下,是由于精子的改变。同时,随着阶跃去极化的变化,受精势(FP)的振幅也在减小,这表明皮质反应在某种程度上发生了改变。尼古丁对没有皮质颗粒的受精卵或卵母细胞的Rm没有影响。我们认为,尼古丁改变了海胆卵的皮质——可能是通过引起皮质颗粒的部分溶解——从而使海胆卵更容易接受精子。阶跃去极化和FP振幅的减小是这种改变的结果。
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引用次数: 11
Microdensitometric evaluation of the DNA content, as ploidy parameter, of spermatozoa in the polymorphic chromosomal system of Akodon molinae cabrera (Rodentia, Cricetidae) 用微密度法评价鼠目蟋蟀多态染色体系统中精子DNA含量的倍性参数
Pub Date : 1982-01-01 DOI: 10.1002/MRD.1120050405
C. Redi, S. Garagna, M. S. Merani, E. Capanna, N. Bianchi, M. Romanini
The influence of chromosome variability on the production of euploid spermatozoa was investigated in a suitable biological model, the Akodon molinae system. This consists of individuals whose chromosome constitution is 2n = 42, 2n = 43, or 2n = 44. The only difference between these three karyotypes occurs through a Robertsonian rearrangement combined with two pericentric inversions. Thus, the animals with 2n = 42 (simple homozygotes or SH) have two large metacentric chromosomes number 1; animals with 2n = 43 (heterozygotes or Ht) have a chromosome 1 and two subterminal chromosomes la and lb homologues of the long and short arms of the chromosome 1, respectively; animals with 2n = 44 (double homozygotes or DH) have a pair of la and a pair of lb chromosomes. The gametic euploidy frequency correlated with each chromosome constitution was evaluated on the basis of the DNA content of spermatozoa, which was determined microdensitometrically after the Feulgen reaction, taking into account the site of the spermatozoa along the male genital tract. A comparative assessment of gametic aneuploidy frequency in caput epididymis versus vas deferens demonstrated (1) a falloff in euploid production in passing from the 2n = 42 to the 2n = 44 chromosome forms, alongside a high degree of intragroup variability, and (2) a lower aneuploidy frequency in the vas deferens than in caput epididymis in all the forms considered. These two features, taken together with similar results in the mouse chromosome variability system, suggest that a selection mechanism is operative against aneuploid spermatozoa in the epididymis. This finding is of interest in a wider perspective, since it might turn out to be valid for many mammals.
研究了染色体变异对整倍体精子产生的影响。这包括染色体构成为2n = 42、2n = 43或2n = 44的个体。这三种核型之间的唯一区别是通过罗伯逊重排结合两个中心倒位。因此,具有2n = 42(简单纯合子或SH)的动物具有两条大的1号异心染色体;2n = 43(杂合子或Ht)的动物在1号染色体的长臂和短臂上分别有一条1号染色体和两条亚末端染色体la和lb同源物;2n = 44(双纯合子或DH)的动物有一对la和一对lb染色体。考虑到精子沿男性生殖道的位置,根据Feulgen反应后用微密度法测定的精子DNA含量,评估与各染色体构成相关的配子整倍体频率。对附睾头和输精管配子非整倍体频率的比较评估表明:(1)从2n = 42染色体形式到2n = 44染色体形式的整倍体产生下降,同时具有高度的群体内变异性;(2)在所有考虑的形式中,输精管的非整倍体频率低于附睾头。这两个特征,再加上小鼠染色体变异系统的类似结果,表明附睾中存在一种针对非整倍体精子的选择机制。从更广泛的角度来看,这一发现很有趣,因为它可能对许多哺乳动物都有效。
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引用次数: 9
Evaluation of premeiotic activity in the developing rabbit testis using a modified squash technique 改良挤压法评价兔睾丸早衰活性
Pub Date : 1982-01-01 DOI: 10.1002/MRD.1120060206
M. Randolph, B. Gondos
Germ cells in the developing rabbit testis were studied using a modified squash technique. Preleptotene figures present in the postnatal testis were examined and compared with corresponding stages in ovarian germ cells. The findings indicated differences in the pattern of preleptotene changes in the developing ovary and testis.
采用改良挤压法对兔睾丸生殖细胞进行了研究。产前瘦素数字存在于出生后的睾丸进行了检查,并与相应阶段的卵巢生殖细胞进行了比较。研究结果表明,在发育中的卵巢和睾丸中,preleptene的变化模式有所不同。
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引用次数: 4
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Gamete Research
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