An orchestrated intron retention program in meiosis controls timely usage of transcripts during germ cell differentiation

Global transcriptome reprogramming during sper-matogenesis ensures timely expression of factors in each phase of male germ cell differentiation. Sper-matocytes and spermatids require particularly exten-sive reprogramming of gene expression to switch from mitosis to meiosis and to support gamete morphogenesis. Here, we uncovered an extensive alternative splicing program during this transmeiotic differentiation. Notably, intron retention was largely the most enriched pattern, with spermatocytes showing generally higher levels of retention compared with spermatids. Retained introns are characterized by weak splice sites and are enriched in genes with strong relevance for gamete func-tion. Meiotic intron-retaining transcripts (IRTs) were exclusively localized in the nucleus. However, differ-ently from other developmentally regulated IRTs, they are stable RNAs, showing longer half-life than properly spliced transcripts. Strikingly, fate-mapping experiments revealed that IRTs are recruited onto polyribosomes days after synthesis. These studies reveal an unexpected function for regulated intron retention in modulation of the timely expression of select transcripts during spermatogenesis

TNF-α inhibits GDNF levels in Sertoli cells, through a NF-κB-dependent, HES1-dependent mechanism

Glial cell line-derived neurotrophic factor (GDNF) is a soluble molecule crucial for the regulation of the spermatogonial stem cells (SSC) of the testis. The effects of GDNF on target cells have been extensively described, but mechanisms underlying GDNF regulation are currently under investigation. In the nervous system, GDNF expression is regulated by pro-inflammatory cytokines including lipopolysaccharide (LPS), interleukin 1 beta (IL-1β), and tumor necrosis factor alpha (TNF-α) but the effect of these cytokines on GDNF expression in the testis is unclear

Spermatogonial kinetics in humans

The human spermatogonial compartment is essential for daily production of millions of sperm. Despite this crucial role, the molecular signature, kinetic behavior and regulation of human spermatogonia are poorly understood. Using human testis biopsies with normal spermatogenesis and by studying marker protein expression, we have identified for the first time different subpopulations of spermatogonia. MAGE-A4marks all spermatogonia, KITmarks all Bspermatogonia and UCLH1 all Apale-dark (Ap-d) spermatogonia. We suggest that at the start of the spermatogenic lineage there are Ap-d spermatogonia that are GFRA1High, likely including the spermatogonial stem cells. Next, UTF1 becomes expressed, cells become quiescent and GFRA1 expression decreases. Finally, GFRA1 expression is lost and subsequently cells differentiate into B spermatogonia, losing UTF1 and acquiring KIT expression. Strikingly, most human Ap-d spermatogonia are out of the cell cycle and even differentiating type B spermatogonial proliferation is restricted. A novel scheme for human spermatogonial development is proposed that will facilitate further research in this field, the understanding of cases of infertility and the development of methods to increase sperm output

The AGILE Mission

AGILE is an Italian Space Agency mission dedicated to observing the gamma-ray Universe. The AGILE's very innovative instrumentation for the first time combines a gamma-ray imager (sensitive in the energy range 30 MeV-50 GeV), a hard X-ray imager (sensitive in the range 18-60 keV), a calorimeter (sensitive in the range 350 keV-100 MeV), and an anticoincidence system. AGILE was successfully launched on 2007 April 23 from the Indian base of Sriharikota and was inserted in an equatorial orbit with very low particle background. Aims. AGILE provides crucial data for the study of active galactic nuclei, gamma-ray bursts, pulsars, unidentified gamma-ray sources, galactic compact objects, supernova remnants, TeV sources, and fundamental physics by microsecond timing. Methods. An optimal sky angular positioning (reaching 0.1 degrees in gamma- rays and 1-2 arcmin in hard X-rays) and very large fields of view (2.5 sr and 1 sr, respectively) are obtained by the use of Silicon detectors integrated in a very compact instrument. Results. AGILE surveyed the gamma- ray sky and detected many Galactic and extragalactic sources during the first months of observations. Particular emphasis is given to multifrequency observation programs of extragalactic and galactic objects. Conclusions. AGILE is a successful high-energy gamma-ray mission that reached its nominal scientific performance. The AGILE Cycle-1 pointing program started on 2007 December 1, and is open to the international community through a Guest Observer Program

Cross- talk between testicular steroidogenesis and spermatogenesis

Spermatogenesis is regulated by a complex interplay between endocrine signals and local interactions of tubules with the interstitial tissue, mainly androgen producing Leydig cells (LC). We hypothesized that the functional phenotype of LC reflects the developmental environment in which these cells grow and interact with other testicular cells, including both somatic and germ cells. As experimental model, we used the organotypic culture of 2.5dpp mouse testicular fragments in the presence of 10% KSR (Knockout Serum Replecement), which is essential to maintain the entire spermatogenetic process from spermatogonia to spermatids in vitro up to 5-6 weeks. At different times we analyzed: 1) testicular morphology, to assess germ cell differentiation; 2) LC proliferation by KI67 immunostaining; and 3) LC maturation, by measuring androgens and INSL3 secreted into the medium. LC were identified by immunostaining of specific markers, including 3βHSD, c-Kit and INSL3. In the presence of KSR, testosterone significantly increased from the first to the second week and was maintained up to the fifth week, in association with germ cell differentiation up to haploid elongated spermatids. On the contrary, in the absence of KSR, testosterone markedly declined during the fourth week, together with an altered spermatogenesis. These results indicate that LC undergo normal development and function in vitro, provided that culture conditions maintained spermatogenesis. We also addressed the regulation of LC function in humans, using fresh and cryopreserved testicular biopsies obtained from beating heart organ donors. Having assessed several culture conditions, we set up a new system biology model allowing to successfully in vitro culture testicular fragments for three hours and to analyze the differential LC response to LH and hCG in terms of testosterone secretion. The study of the age-dependent LC response is in progress, comparing young (15-40 years) and elderly (60-85 years) donors

Retinoic acid down-regulates GDNF expression in immature Sertoli cells

The balance between retinoic acid (RA) and GDNF (Glial cell-derived neurotrophic factor) determines the behavior of the spermatogonial stem cell compartment. Importantly, RA inhibits GDNF expression during the cycle of the seminiferous epithelium [1]. However, the molecular mechanism by which RA impinge on GDNF regulation within the testis is not known. The aim of the present project is to clarify the molecular mechanisms underlying RA-dependent gdnf regulation in mouse testis. Deepening these dynamics could ameliorate the comprehension of SSCs niche and the mechanism behind SSCs self-renewal

Retinoic acid regulation of human spermatogonial genes

The retinoic acid (RA) is a vitamin A metabolite that, in the mouse, has been demonstrated to regulate the spermatogonial differentiation, the meiotic entry, the initiation of the spermatid elongation and the release of spermatozoa. Over the years several RA responsive genes have been identified. They have been involved in the mediation of one or more events. Among others Stra8 is crucial to induce the spermatogonial differentiation and the meiotic entry [1]. In our study we tested the hypothesis that the RA is able to regulate the same genes in the human spermatogonia

Spermatogonial cells: mouse, monkey and man comparison

In all mammals, spermatogonia are defined as constituting the mitotic compartment of spermatogene-sis including stem, undifferentiated and differentiating cell types, possessing distinct morphological and molecular characteristics. Even though the real nature of the spermatogonial stem cell and its regulation is still debated the general consensus holds that in steady-state spermatogenesis the stem cell compart-ment needs to balance differentiation versus self-renewal. This review highlights current understanding of spermatogonial biology, the kinetics of amplification and the signals directing spermatogonial differ-entiation in mammals. The focus will be on relevant similarities and differences between rodents and non human and human primates