The ascending testis and the testis undescended since birth share the same histopathology

PURPOSE: The etiology of the ascending testis is controversial. We propose that ascending testis, defined as a testis previously thought to be descended and later noted to be out of the scrotum, is due to mild hypogonadotropic hypogonadism affecting both testes. The diagnosis of these low types of true undescended testes is difficult to make clinically in children since they are frequently confused with retractile testes. In this study we compared testicular biopsies in a group of boys with ascending testes with those in boys who had an undescended testis since birth (primary undescended testis). MATERIALS AND METHODS: Between 1985 and 1995, 91 patients with ascending testes underwent orchiopexy and bilateral testis biopsy. The total germ cell count, processus vaginalis status, age at surgery and whether followup was done by a pediatrician or pediatric urologist were compared in patients with ascending and unilateral primary undescended testes. RESULTS: The total germ cell count was similar in the undescended and the contralateral descended testis in patients with ascending and primary undescended testes. The processus vaginalis was more likely to be closed in ascending testes (57% versus 36%, p = 0.0001). Age at surgery and the total germ cell count were similar in patients followed by pediatricians and pediatric urologists. CONCLUSIONS: The ascending testis has the same germ cell count as the primary undescended testis. Yearly followup by the primary care physician is recommended for patients with retractile testes.     

Assessment of the role of Leydig cell products other than testosterone in spermatogenesis and fertility in adult rats

Adult male rats were treated with ethane dimethanesulphonate (EDS) to destroy the Leydig cells and were then supplemented for 3-10 weeks with testosterone esters (TE) by injection every 3 days. The latter treatment prevented Leydig cell regeneration but maintained quantitatively the androgen-dependent aspects of spermatogenesis, as judged by germ cell counts at stage VII of the spermatogenic cycle. Other than the absence of Leydig cells, the testes of EDS-treated, TE-supplemented rats showed only two morphological changes, (1) the appearance of mast cells throughout the interstitium, and (2) a 3- to 4-fold increase in the number of degenerating germ cells (secondary spermatocytes) at stages XIV-I; this was reflected in a significant decrease in the ratio of spermatids to pachytene spermatocytes at stage VII. These changes were not observed in either oil-treated or TE-treated control rats although similar, but less marked, changes in cell degeneration at stages XIV-I were observed in rats actively immunized against oxytocin. Epididymal sperm number was reduced marginally (approximately 15%) in EDS-treated, TE-supplemented rats while sperm motility was affected even less. In a serial mating trial, some of these treated rats showed evidence of subfertility/infertility, but this was mostly transient and may have been the result of epididymal effects of EDS. These results suggest that Leydig cell products other than testosterone are not essential for maintenance of spermatogenesis and fertility in rats, although because of increased germ cell degeneration during the final stages of meiosis (perhaps as the result of oxytocin withdrawal), a small reduction in sperm count may occur.     

Expression of cubilin in mouse testes and Leydig cells

Cubilin (cubn) is a receptor for vitamins and various protein ligands. Cubn lacks a transmembrane domain but anchors to apical membranes by forming complexes with Amnionless or Megalin. In an effort to better understand the uptake of nutrients in testis, we analysed cubn expression in the developing mice testes. In testes, cubn mRNA increased from birth to adulthood. In the inter‐stitium and isolated seminiferous tubules, neonatal increase in cubn mRNA until 14 days post‐partum (pp) was followed by a marked increase at puberty (28 days pp). Cubn was found in the gonocytes, spermatogonia, spermatocytes and spermatids in the developing testes. In adult testes, strong Cubn immunoreactivity was found in the elongating spermatids, suggesting the role of Cubn in endocytosis during early spermiogenesis. In Sertoli cells and peritubular cells, Cubn immunoreactivity was weak throughout the testis development. In the inter‐stitium, Cubn immunoreactivity was found in foetal Leydig cells, was weak to negligible in the stem cells and progenitor Leydig cells and was strong in immature and adult Leydig cells, demonstrating a positive association between Cubn and steroidogenic activity of Leydig cells. Collectively, these results suggest that Cubn may participate in the endocytotic uptake of nutrients in germ cells and somatic cells, supporting the spermatogenesis and steroidogenesis in mouse testes.     

Mn2+-dependent Adenylyl Cyclase (AC) in Rat Testis: Kinetic Properties and Optimalization of Assay Conditions

The aim of the present study was to investigate some kinetic properties of the soluble Mn²⁺-dependent adenylyl cyclase (AC) in rat testis and to examine the viability of this enzyme under various assay conditions.
1. Constant AC activity was found during the first 35 min of incubation, and AC activity was proportional to protein concentrations in the range tested (10–45 μg/tube).
2. The enzyme activity was constant for at least 24 h at 4°C and for at least 2 h at 35°C. At higher temperatures (above 45°C) the AC activity rapidly declines.
3. The enzyme displays a pH optimum in the psysiological range (7.0–7.4) and increasing ionic strenght (0-0.4 M NaCl/KCl) caused only a marginal decrease in enzyme activity.
5. The soluble Mn²⁺-dependent AC in rat testis could not be stimulated by hormones (hCG, FSH, prostaglandins), guanyl-nucleotides (GMP-P(NH)P, 2–4 times 10^-5 M) or F^-. Ca²⁺ (0.1–5 mM) caused a dose-dependent stimulation of AC activity; the most pronounced effect was seen at low Mn²⁺ concentrations.
6. The K_m for MnATP^2- was estimated to 3.0 mM and the K_m for Mn²⁺ to 7.2 mM. Increasing concentrations of Mn²⁺ did not alter the affinity for MnATP^2- and neither did varying levels of ATP affect the affinity for Mn²⁺.     

Preliminary physico-chemical characterization of the soluble Mn2+-dependent adenylate cyclase in the rat testis

This paper reports some physico-chemical properties of the Mn⁺-dependent adenylate cyclase (AC) of the rat testis. The AC activity in the crude cytosol (106 000 × g ) migrates as a single peak in a linear sucrose gradient (5–20%) with a sedimentation coefficient (S_{20, w}) of 4,1. The enzyme exhibits an Einstein-Stokes radius (r_s) of 30.6 Å as assessed by gel filtration (K_av= 0.50) on a calibrated Sephadex G-200 column. The molecular shape is close to spherical (f/f_o= 1.25). Isoelectric focussing of the AC peak (K_av= 0.50) from the Sephadex G-200 column revealed a pI of 5.7. Ion exchange chromatography of the crude cytosol on a Whatman DE-52 column gave a single peak eluting between 0.13-0.17 M NaCl. Electrophoresis on polyacrylamide gels resulted in a migrating AC peak with an apparent Rf of 0.40 relative to bromophenol blue. The Mn²⁺-dependent AC is not retained on a Con-A Sepharose 4B column indicating that the enzyme molecule does not contain sugars possessing α-D-manno- or α-D-glucopyranosyl terminal groups. (NH₄)₂SO₄ precipitates the AC activity (4°C) between 30–60% saturation. The yield approximates 85% of total AC activity with the specific activity reaching a plateau (110 pinoles cAMP/mg protein/min.) at 50% saturated (NH₄)₂SO₄. The soluble Mn⁺-dependent AC of the rat testis thus appears to be relatively symmetrical with a size of approximately 52 000 D and a negative charge at physiological pH. Apparently it does not contain sugar moieties and it exhibits little molecular heterogeneity.     

Kinetic Properties of Hexokinase of Germ Cells in Rat Testis

The Michaelis-Menten constants (Km) of glucose, fructose, galactose, 2-deoxy-D-glucose and ATP as substrate for hexokinase of spermatocyte, spermatid and cauda epididymal spermatozoa extracts were measured. The Km value of glucose was very similar for all three germ cells. It was also true for all other substrates. The affinity of glucose for this enzyme was the highest while that of fructose was the lowest. The Km values were in general agreement with characteristics of hexokinase extracted from other tissues of rat. The Vmax values were also determined. The Vmax ratio of fructose to glucose showed the highest values of 1.1. The Vmax ratios of other substrates to glucose were below 1.0. The results suggest that hexokinase in the germ cells is similar to that in other tissues in its kinetic properties.     

A matter of death and life: the significance of germ cell death during spermatogenesis

The significance of cell death occurring during spermatogenesis is a subject of interest because of its potential medical importance. Unfortunately, the field has been difficult for andrologists to penetrate, in part because of the difficulties of studying germ cells in vitro and the complexity of designing suitable models in which to dissect the molecular signalling pathways involved in control of germ cell apoptosis. As a result, the reasons for these deaths remain unclear despite considerable investigative effort. As developments which have occurred over the last few years in understanding of apoptosis can shed light on this important topic, this review focuses on what is currently known about germ cell apoptosis and outlines the emerging picture of what might be the causes and biological role of germ cell deaths in spermatogenesis.     

Morphology of normal and malignant germ cells*

By means of semithin section histology and electron microscopy the following male germ cells are described: gonocytes and prospermatogonia in the embryonic and fetal testes; spermatogonia and primary spermatocytes in the testis of the adult; abnormal spermatogonia and spermatocytes, which are not tumour cells. In contrast to these cells, four main tumour cell types which on the basis of morphological criteria can be said to be derived from germ cells, are classified and designated as Tc 1 to Tc 4. The various tissues accompanying the tumour cells are briefly described.     

Foetal meiosis in the testis of the rodent Octodon degus

Different stages of meiotic prophase have been studied in foetal testes of the rodent, Octodon degus, using the light and electron microscope. Special attention was focused on the ultrastructural morphology of these meiotic cells in comparison to pre-spermatogonia of foetal testes and meiotic spermatocytes of the adult male testis. Meiosis occurs in only a few cells located among fibroblasts of the tunica albuginea or in the region of the gonadal blastema. The foetal meiotic process resembles adult meiosis in its ultrastructural characteristics; typical pachytene synaptonemal complexes and leptotene or diplotene axial elements appear associated to the chromatin. This process occurs at the same foetal age that meiosis commences in the ovary, thus reinforcing the idea that both meiosis-inducing and meiosis-preventing substances are secreted in both sexes. The intra-or extracordonal localization of the germ cells would be an important factor in determining the cells' response to these substances.     

Prenatal development of murine gonads with special reference to germ cell differentiation: a morphological and immunohistochemical study

The prenatal differentiation of male and female gonads of the mouse was investigated both morphologically and immunohistochemically. Sexual dimorphism could be detected as early as 12 days post-coitum (dpc) by the appearance of the primary elements of the tunica albuginea and positive immunoreactivity for anti-Muellerian hormone in the Sertoli cells of the male gonad. Male germ cells passed two waves of mitotic activity, a first wave between 12 and 14 dpc, which is followed by apoptosis of the old germ cell generation, and a second wave between 17 and 20 dpc. Oct-4 was expressed as a juxtanuclear ring in the cytoplasm of germ cells up to 17 dpc. Subsequently, it was down-regulated and completely disappeared in 20 dpc full-term fetuses. By contrast, M2A antigen revealed only a weak immunoreaction in some germ cells of 14 dpc gonads, but exhibited strong signals in all germ cells of 20 dpc full-term fetuses. Therefore, we postulate that, in the mouse, prenatal germ cells represent two populations: the first is immunopositive for Oct-4 and disappeared in full-term fetuses, whereas the second appeared in 14 dpc and is immunopositive for M2A antigen.     

Germline-competent stem cell in avian species and its application

Germ cells are the only cell type in the body that can transfer genetic information to the next generation. Germline-competent stem cells can self-renew and contribute to the germ cell lineage giving rise to pluripotent stem cells under specific conditions. Hence far, studies on germline-competent stem cells have contributed to the generation of avian model systems and the conservation of avian genetic resources. In this review, we focus on previous studies on germline-competent stem cells from avian species, mainly chicken germline-competent stem cells, which have been well established and characterized. We discuss different sources of germline-competent stem cells and recent advances for the future applications in birds.     

全反式视黄酸影响小鼠胚胎干细胞向原始生殖细胞分化的相关基因表达

目的 探讨全反式视黄酸(all-trans retinoic acid,atRA)诱导小鼠胚胎干细胞(mouse embryonic stem cells,mESCs)向原始生殖细胞(Primordial germ cellS,PGCs)分化的相关基因表达改变及其机制.方法 mESCs分化形成拟胚体(embryoid bodies,Ebs),不同浓度atRA(1 μM,2 μM,5 μM)持续诱导Ebs 16h、2d和5d,观察Ebs形态变化,实时PCR和Western blot检测PGCs分化相关基因和蛋白表达变化.分析基因肩动子中RA反应元件,以Stra8(Stimulated by Retinoic Acid Gene 8,Stra8)基因为阳性对照,确定各基因对atRA刺激的原发性反应和继发性反应.结果 不同浓度atRA诱导Ebs 16h和2d后形态无明显变化.诱导后Ebs的PGCs分化相天基因mRNA表达分4种情况:16h内(Stra8)表达量达最大,然后下降;2d内(Scp3)表达量达最大,然后下降;5d达到最大量(Mvh);表达量无明显变化(Fragilis,Blimp-1和Prm1).Mvh表达符合继发反应特点;Stra8和Scp3表达符合原发反应特点;Blimp1,Prm1和Fragilis表达可能与atRA调节无关.Stra8蛋白变化与mRNA变化相一致,而Mvh则不一致.结论 atRA诱导mESCs向PGCs分化,各阶段标志基因表达变化均在相对短的时间内完成(2～5d),atRA诱导浓度可选用1 μM.根据基因表达模式初步得出,atRA调控的靶基因为Mvh,具备继发反应特点,与特异性分化有关,而Blimp-1、Fragilis和Prm1调控PGCs分化可能与atRA无关.     

咖啡因对胎鼠及乳鼠睾丸生殖细胞数量及合成DNA的影响

为了深入了解咖啡因对胎儿、新生儿生殖细胞的数量及其合成ＤＮＡ的影响，本实验采用低（３×１０－４ｍｏｌ／Ｌ）、中（６×１０－４ｍｏｌ／Ｌ）、高（１．２×１０－３ｍｏｌ／Ｌ）浓度咖啡因体外培养ＳＤ孕１８天胎鼠、０天及４天乳鼠睾丸组织块，培养时间分别为１周、２周、３周，观察咖啡因作用后睾丸内生殖细胞数量的变化，并且利用计算机图象分析系统测定生殖细胞ＤＮＡ的含量，以探讨两者的关系。结果如下：（１）１８天胎鼠睾丸培养组织的生殖细胞受咖啡因影响最少，０天乳鼠次之，４天乳鼠受影响较大。（２）低浓度的咖啡因对生殖细胞数量影响较少；中等浓度的咖啡因在培养３周后才有生殖细胞数量的降低；而高浓度的咖啡因在培养２周后，已有下降，３周更明显。（３）生殖细胞数量的减少往往伴随ＤＮＡ含量的降低，由此推测高浓度咖啡因长时间培养后使生殖细胞数量减少，可能通过抑制ＤＮＡ复制发挥作用。     

Sex Determination and Differentiation of the Mammalian Gonad

There is growing evidence to suggest that the presence of a single gene on the Y chromosome, which also codes for the expression of the H-Y antigen, causes the primordial gonad to develop structurally as a testis: in the absence of this gene, an ovary is formed. Whether the gonad subsequently becomes capable of producing viable gametes depends on the sex chromosome makeup of the germ cells that migrate into it; XY or XO germ cells seem to be capable of developing into either oocytes or spermatocytes, depending on the environment, whereas germ cells with two X chromosomes can only develop as oocytes, and will not survive in a testicular environment. The H-Y antigen probably plays no role in the sexual differentiation of the germ cell.
There is another gene on the Y chromosome that determines testis size, and testes: body weight ratios vary widely in different mammals. Amongst the higher primates the gorilla has the lowest ratio (0.018%), man is intermediate (0.079%) and the chimpanzes have the highest (0.269%). Testis size is predominantly determined by the amount of tubular tissue, and appears to be closely related to copulatory frequency, and hence to the mating behaviour of the species. It is important to take these marked species differences in testis size into account when using primate models for human male contraceptive research and development.     

糖尿病对小鼠睾丸组织中PCNA表达的影响

为研究糖尿病对睾丸生殖细胞DNA合成的影响，应用免疫组化ABC法检测增殖细胞核抗原（proliferation cell nuleus antigen PCNA)在睾丸组织的表达。结果显示免疫阳性细胞包括精原细胞和初级精母细胞，少数支持细胞和肌样细胞胞核也有着色。PCNA阳性率随糖尿病病程进展呈明显下降趋势，而且每年龄段糖尿病组PCNA阳性率均明显低于相应正常对照组。由此得出结论，糖尿病对PCNA在睾丸组织中表达起抑制作用，且与糖尿病病程呈正相关，即糖尿病时生精过程受到明显抑制，这种抑制作用可能与糖尿病所导致的雄性生殖能力下降有关。