Reproduction

Miller, David; Brinkworth, Martin; Iles, David

doi: 10.1530/rep-09-0281pmid: 19759174

Haploid male germ cells package their DNA into a volume that is typically 10% or less that of a somatic cell nucleus. To achieve this remarkable level of compaction, spermatozoa replace most of their histones with smaller, highly basic arginine and (in eutherians) cysteine rich protamines. One reason for such a high level of compaction is that it may help optimise nuclear shape and hence support the gametes' swimming ability for the long journey across the female reproductive tract to the oocyte. Super-compaction of the genome may confer additional protection from the effects of genotoxic factors. However, many species including the human retain a fraction of their chromatin in the more relaxed nucleosomal configuration that appears to run counter to the ergonomic, toroidal and repackaging of sperm DNA. Recent research suggests that the composition of this ‘residual’ nucleosomal compartment, a generally overlooked feature of the male gamete, is far more significant and important than previously thought. In this respect, the transport and incorporation of modified paternal histones by the spermatozoon to the zygote has been demonstrated and indicates another potential paternal effect in the epigenetic reprogramming of the zygote following fertilisation that is independent of imprinting status. In this review, the most recent research into mammalian spermatozoal chromatin composition is discussed alongside evidence for conserved, non-randomly located nucleosomal domains in spermatozoal nuclei, all supporting the hypothesis that the spermatozoon delivers a novel epigenetic signature to the egg that may be crucial for normal development. We also provide some thoughts on why this signature may be required in early embryogenesis.

Visser, Liesbeth; Repping, Sjoerd

doi: 10.1530/REP-09-0229pmid: 19776097

Subfertility, defined as the inability to conceive within 1 year of unprotected intercourse, affects 10–15% of couples. Inup to 55% of couples, the male partner is diagnosed with spermatogenic failure, i.e. one or more semen parameters fall belowthe WHO criteria for normozoospermia. In these cases, assisted reproductive technology is usually used to achieve pregnancy.Both genetic and environmental factors are thought to underlie spermatogenic failure. Despite years of research, only fewgenetic factors have clearly been shown to cause spermatogenic failure, and the identification of additional genetic causesor risk factors has proven to be extremely difficult. In this review, we will present an overview of established genetic causesof spermatogenic failure, describe pitfalls in searching for novel genetic factors and discuss research opportunities forthe future.

Visser, Liesbeth; Repping, Sjoerd

doi: 10.1530/rep-09-0229pmid: 19776097

Subfertility, defined as the inability to conceive within 1 year of unprotected intercourse, affects 10–15% of couples. In up to 55% of couples, the male partner is diagnosed with spermatogenic failure, i.e. one or more semen parameters fall below the WHO criteria for normozoospermia. In these cases, assisted reproductive technology is usually used to achieve pregnancy. Both genetic and environmental factors are thought to underlie spermatogenic failure. Despite years of research, only few genetic factors have clearly been shown to cause spermatogenic failure, and the identification of additional genetic causes or risk factors has proven to be extremely difficult. In this review, we will present an overview of established genetic causes of spermatogenic failure, describe pitfalls in searching for novel genetic factors and discuss research opportunities for the future.

Rodgers, R J; Irving-Rodgers, H F

doi: 10.1530/REP-09-0177pmid: 19786400

Follicle classification is an important aid to the understanding of follicular development and atresia. Some bovine primordialfollicles have the classical primordial shape, but ellipsoidal shaped follicles with some cuboidal granulosa cells at thepoles are far more common. Preantral follicles have one of two basal lamina phenotypes, either a single aligned layer or onewith additional layers. In antral follicles <5 mm diameter, half of the healthy follicles have columnar shaped basal granulosacells and additional layers of basal lamina, which appear as loops in cross section (‘loopy’). The remainder have alignedsingle-layered follicular basal laminas with rounded basal cells, and contain better quality oocytes than the loopy/columnarfollicles. In sizes >5 mm, only aligned/rounded phenotypes are present. Dominant and subordinate follicles can be identifiedby ultrasound and/or histological examination of pairs of ovaries. Atretic follicles <5 mm are either basal atretic or antralatretic, named on the basis of the location in the membrana granulosa where cells die first. Basal atretic follicles haveconsiderable biological differences to antral atretic follicles. In follicles >5 mm, only antral atresia is observed. Theconcentrations of follicular fluid steroid hormones can be used to classify atresia and distinguish some of the differenttypes of atresia; however, this method is unlikely to identify follicles early in atresia, and hence misclassify them as healthy.Other biochemical and histological methods can be used, but since cell death is a part of normal homoeostatis, deciding whena follicle has entered atresia remains somewhat subjective.

Rodgers, R J; Irving-Rodgers, H F

doi: 10.1530/rep-09-0177pmid: 19786400

Follicle classification is an important aid to the understanding of follicular development and atresia. Some bovine primordial follicles have the classical primordial shape, but ellipsoidal shaped follicles with some cuboidal granulosa cells at the poles are far more common. Preantral follicles have one of two basal lamina phenotypes, either a single aligned layer or one with additional layers. In antral follicles <5 mm diameter, half of the healthy follicles have columnar shaped basal granulosa cells and additional layers of basal lamina, which appear as loops in cross section (‘loopy’). The remainder have aligned single-layered follicular basal laminas with rounded basal cells, and contain better quality oocytes than the loopy/columnar follicles. In sizes >5 mm, only aligned/rounded phenotypes are present. Dominant and subordinate follicles can be identified by ultrasound and/or histological examination of pairs of ovaries. Atretic follicles <5 mm are either basal atretic or antral atretic, named on the basis of the location in the membrana granulosa where cells die first. Basal atretic follicles have considerable biological differences to antral atretic follicles. In follicles >5 mm, only antral atresia is observed. The concentrations of follicular fluid steroid hormones can be used to classify atresia and distinguish some of the different types of atresia; however, this method is unlikely to identify follicles early in atresia, and hence misclassify them as healthy. Other biochemical and histological methods can be used, but since cell death is a part of normal homoeostatis, deciding when a follicle has entered atresia remains somewhat subjective.

Cleverly, Kirsty; Wu, T John

doi: 10.1530/rep-09-0117pmid: 19755481

LHRH (GNRH) was first isolated in the mammalian hypothalamus and shown to be the primary regulator of the reproductive neuroendocrine axis comprising of the hypothalamus, pituitary and gonads. LHRH acts centrally through its initiation of pituitary gonadotrophin release. Since its discovery, this form of LHRH (LHRH-I) has been shown to be one of over 20 structural variants with a variety of roles in both the brain and peripheral tissues. LHRH-I is processed by a zinc metalloendopeptidase EC 3.4.24.15 (EP24.15) that cleaves the hormone at the fifth and sixth bond of the decapeptide (Tyr5-Gly6) to form LHRH-(1–5). We have previously reported that the auto-regulation of LHRH-I (GNRH1) gene expression and secretion can also be mediated by itself and its processed peptide, LHRH-(1–5), centrally and in peripheral tissues. In this review, we present the evidence that EP24.15 is the main enzyme of LHRH metabolism. Following this, we look at the metabolism of other neuropeptides where an active peptide fragments is formed during degradation and use this as a platform to postulate that EP24.15 may also produce an active peptide fragment in the process of breaking down LHRH. We close this review by the role EP24.15 may have in regulation of the complex LHRH system.

Cleverly, Kirsty; Wu, T John

doi: 10.1530/REP-09-0117pmid: 19755481

LHRH (GNRH) was first isolated in the mammalian hypothalamus and shown to be the primary regulator of the reproductive neuroendocrineaxis comprising of the hypothalamus, pituitary and gonads. LHRH acts centrally through its initiation of pituitary gonadotrophinrelease. Since its discovery, this form of LHRH (LHRH-I) has been shown to be one of over 20 structural variants with a varietyof roles in both the brain and peripheral tissues. LHRH-I is processed by a zinc metalloendopeptidase EC 3.4.24.15 (EP24.15)that cleaves the hormone at the fifth and sixth bond of the decapeptide (Tyr5-Gly6) to form LHRH-(1–5). We have previously reported that the auto-regulation of LHRH-I (GNRH1) gene expression and secretion can also be mediated by itself and its processed peptide, LHRH-(1–5), centrally and in peripheraltissues. In this review, we present the evidence that EP24.15 is the main enzyme of LHRH metabolism. Following this, we lookat the metabolism of other neuropeptides where an active peptide fragments is formed during degradation and use this as aplatform to postulate that EP24.15 may also produce an active peptide fragment in the process of breaking down LHRH. We closethis review by the role EP24.15 may have in regulation of the complex LHRH system.

Nakai, Michiko; Kaneko, Hiroyuki; Somfai, Tamas; Maedomari, Naoki; Ozawa, Manabu; Noguchi, Junko; Ito, Junya; Kashiwazaki, Naomi; Kikuchi, Kazuhiro

doi: 10.1530/rep-09-0509pmid: 20015869

Xenografting of testicular tissue into immunodeficient mice is known to be a valuable tool for facilitating the development of immature germ cells present in mammalian gonads. Spermatogenesis in xenografts and/or in vitro embryonic development to the blastocyst stage after ICSI of xenogeneic sperm has already been reported in large animals, including pigs; however, development of the embryos to term has not yet been confirmed. Therefore, in pigs, we evaluated the in vivo developmental ability of oocytes injected after ICSI of xenogeneic sperm. Testicular tissues prepared from neonatal piglets, which contain seminiferous cords consisting of only gonocytes/spermatogonia, were transplanted under the back skin of castrated nude mice. Between 133 and 280 days after xenografting, morphologically normal sperm were recovered, and a single spermatozoon was then injected into an in vitro matured porcine oocyte. After ICSI, the oocytes were electrostimulated and transferred into estrus-synchronized recipients. Two out of 23 recipient gilts gave birth to six piglets. Here, we describe for the first time that oocytes fertilized with a sperm from ectopic xenografts have the ability to develop to viable offspring in large mammals.

Nakai, Michiko; Kaneko, Hiroyuki; Somfai, Tamas; Maedomari, Naoki; Ozawa, Manabu; Noguchi, Junko; Ito, Junya; Kashiwazaki, Naomi; Kikuchi, Kazuhiro

doi: 10.1530/REP-09-0509pmid: 20015869

Xenografting of testicular tissue into immunodeficient mice is known to be a valuable tool for facilitating the developmentof immature germ cells present in mammalian gonads. Spermatogenesis in xenografts and/or in vitro embryonic development to the blastocyst stage after ICSI of xenogeneic sperm has already been reported in large animals,including pigs; however, development of the embryos to term has not yet been confirmed. Therefore, in pigs, we evaluated thein vivo developmental ability of oocytes injected after ICSI of xenogeneic sperm. Testicular tissues prepared from neonatal piglets,which contain seminiferous cords consisting of only gonocytes/spermatogonia, were transplanted under the back skin of castratednude mice. Between 133 and 280 days after xenografting, morphologically normal sperm were recovered, and a single spermatozoonwas then injected into an in vitro matured porcine oocyte. After ICSI, the oocytes were electrostimulated and transferred into estrus-synchronized recipients.Two out of 23 recipient gilts gave birth to six piglets. Here, we describe for the first time that oocytes fertilized witha sperm from ectopic xenografts have the ability to develop to viable offspring in large mammals.