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PubMed1996

Expression of multiple c-kit receptor messenger ribonucleic acid transcripts during postnatal development of the rat testis.

The c-kit protooncogene is a transmembrane tyrosine kinase receptor expressed during gametogenesis. Using the polymerase chain reaction (PCR), we have identified the c-kit receptor mRNA transcripts in the rat testis and studied their expression during postnatal development of the testis. Five different transcripts were identified using sets of primers encoding within the extracellular domain. Two transcripts were obtained from primer sets encoding regions within the cytoplasmic domain and the primer set encoding the entire length of the c-kit receptor. We have compared the levels of expression of these transcripts on different days during postnatal development. The level of expression of a particular transcript varied depending upon the developmental stage of the testis. In summary, our results suggest that multiple forms of mRNAs exist for the c-kit receptor in the rat testis, and they are regulated differentially during postnatal development.

Aging↗
PubMed1979

RNA synthesis and RNA polymerase activities in germ cells of developing rainbow trout testis.

Spermatogenesis is a complex developmental process which sequentially generates several different germ cell types. These cell types from rainbow trout (Salmo gairdnerii) testis were separated by sedimentation in serum albumin gradients and characterized on the basis of their physical properties, chronological appearance, and protein synthesis. The rate of RNA synthesis, the types of RNA made, and the RNA polymerase activities present were determined for each cell type. The rate of RNA synthesis decreased from a high level in spermatogonia and spermatocytes to a low level in early spermatids and was absent in late spermatids and mature spermatozoa. Newly synthesized RNA in spermatogonia and spermatocytes consisted of a variety of molecular weight species, including 18 S and 28 S ribosomal RNAs. The synthesis of high molecular weight RNAs, especially ribosomal RNAs, decreased drastically in early spermatids, leading to the synthesis of only small molecular weight RNAs. RNA polymerase I and II were present in all cell types but the activities of both showed large decreases between spermatocytes and middle spermatids. Both RNA polymerase activities were almost absent from spermatozoa. The activities of RNA polymerase I and II from unfractionated testis cells at different stages of hormone-induced spermatogenesis were quantitated by fractionation of the solubilized extract on DEAE-cellulose. Both polymerases showed major decreases in activity which began near the chronological mid-point of development. For polymerase I the decrease in activity was over 400 fold, for polymerase II over 200 fold. The number of RNA polymerase II molecules per testis cell, quantitated by the binding of [3H]amanitin to cell extracts, also decreased markedly during spermatogenesis. The reduction in polymerase II activity was accompanied by a parallel 200-fold decrease in[3H]amanitin binding. The reduction in polymerase activity appears, therefore, to be due to an actual reduction in the cellular content of RNA polymerase II molecules. These results suggest that transcription in maturing testes is regulated, at least in part, by the concentrations of the RNA polymerases.

Amanitins↗
PubMed2001

Rabbit sex hormone-binding globulin: expression in the liver and testis during postnatal development and structural characterization by truncated proteins.

Although sex hormone binding globulin (SHBG) is found in the blood plasma of adult humans and rabbits and the gene is expressed in their livers, it is not detected in the plasma of adult rodents nor is it expressed in adult rodent livers. Thus the rabbit represents a good model to study the metabolism and function of SHBG in the blood. We have used a cloned rabbit SHBG cDNA to detect mRNA expression in rabbits during the postnatal period, and to construct truncated SHBG proteins for structure/function analysis. The SHBG mRNA appeared in the testis as early as 3 days after birth. The level increased gradually in abundance throughout postnatal development, and attained a maximum at 12 weeks of age when the gonads were fully matured. In contrast, SHBG mRNA in the livers of male and female animals increased to a maximum by 4 weeks of age, and were maintained at this level until 12 weeks before subsiding to the initial levels. The increase and decrease in SHBG mRNA levels in the liver were accompanied by similar changes in serum SHBG. This suggests that SHBG in the blood circulation comes from the liver and this might also provide a source of SHBG for the male reproductive tract before formation of the blood-testis barrier. To elucidate the minimal sequence of rabbit SHBG responsible for steroid-binding, a panel of 13 truncated SHBG proteins was constructed, expressed in Escherichia coli, and biochemically purified for study. It was shown that the complete protein sequence of rabbit SHBG was important for maintaining a stable steroid-protein complex. Unlike human SHBG for which a truncated protein of the first 206 residues of the 373 amino acid protein can still bind steroid, removal of 43 or more residues from the C-terminus of rabbit SHBG completely abolished steroid-binding.

Age Factors↗
PubMed2000

Gonocyte-Sertoli cell interactions during development of the neonatal rodent testis.

During neonatal testicular development in the rat, events critical for subsequent germ cell development occur that set the stage for fertility later in life. Some gonocytes resume mitotic activity and/or migrate to the surrounding basal lamina, and use of a carefully defined Sertoli cell-gonocyte coculture system indicates that these crucial events occur without added factors or hormones and are hence likely to depend on interaction with adjacent Sertoli cells. Coupling of the Kit receptor protein on gonocytes to stem cell factor from Sertoli cells is vital for successful migration by gonocytes, as antagonism of the former suppresses and addition of the latter stimulates gonocyte migration. During the neonatal period, intercellular adhesion is modified in a developmental manner such that neural cell adhesion molecule (NCAM) is the main adhesive molecule expressed and functioning at birth, with a progressive decline as development proceeds. This decline in NCAM expression is supported by the addition of exogenous 3,3',5-triiodothyronine in vitro, and because this factor is recognized as supporting Sertoli cell differentiation, it seems likely that changing intercellular adhesion is a function of progressive development of Sertoli cells. Other avenues whereby maturing testicular cells influence each other doubtless exist, including secretion of growth factors and other peptides and developmentally important changes in the makeup of the extracellular matrix, which Sertoli cells and gonocytes contact. Continued investigation in these areas will be very valuable in enlarging our understanding of how neonatal testicular development provides the basis for successful spermatogenesis.

Animals↗
PubMed1994

Cadherins and cadherin-associated molecules in the developing and maturing rat testis.

The calcium-dependent class of cell adhesion molecules known as cadherins mediate homotypic cell interactions in most epithelia. We have now investigated the expression and distribution of cadherins and cadherin-associated molecules in the developing and maturing rat testis. E-Cadherin was not detected in the seminiferous tubule at any time in development or in the adult. In contrast, Leydig cells expressed E-cadherin between day 15 of gestation and postnatal day 3. alpha- and beta-catenins were expressed throughout the developing testis, but were particularly prominent in Leydig cells. In the maturing testis, alpha-catenin and plakoglobin became progressively more restricted to the basal part of the seminiferous epithelium and by 23 days exhibited a pattern characteristic of the Sertoli cell junctional complex. beta-Catenin recruitment to the Sertoli cell junctional complex was not complete until 60 days. alpha-Catenin and plakoglobin were not present at sites of Sertoli cell-germ cell contacts. Northern blot analysis of testicular RNA showed three mRNA species hybridizing with N-cadherin cDNA. A pan-cadherin antibody specific for a region of the highly conserved C-terminal of all cadherins stained sites of Sertoli-spermatocyte and Sertoli-round spermatid contact in the adult rat seminiferous epithelium, but did not stain the Sertoli cell tight junctional complex. Western blots of testicular extracts indicated that the molecule(s) recognized by these antibodies had an approximate molecular mass of 120 kilodalton, typical of members of the cadherin family. Therefore, although Sertoli cells do not express E-cadherin, another member(s) of the cadherin family is present in the testis, but may not be directly involved in tight junction dynamics as in other cells. Instead, cadherin-mediated adhesion is likely to be involved in Sertoli cell-germ cell interactions. As catenins are not present at these sites, our results suggest a catenin-independent role of cadherins in germ cell adhesion to Sertoli cells.

Aging↗
PubMed1984

Sertoli cell development of pig testis in the fetal and neonatal period.

The Sertoli cells of pig fetuses from 35 days postcoitum until 1 mo after birth have been investigated by light and electron microscopy in decapitated animals and their control littermates, as well as in untreated animals. Until 52 days postcoitum, Sertoli cells change in form during the formation of sex cords but from then onwards they are rather uniform. They primarily display an elongated nonindented nucleus with one or more prominent nucleoli, a well-developed Golgi apparatus, and in the basal compartment below or beside the nucleus, a large lipid droplet. There are large quantities of rough endoplasmic reticulum (RER) from 52 days postcoitum onwards, often with complex whirl forms and a parallel arrangement of profiles with relatively few ribosomes. After birth their numbers seem to be somewhat less, and by 1 mo after birth the RER profiles are often shorter and almost free of ribosomes. Clustered ribosomes are found in large quantities throughout the period under investigation. Especially in the early fetal period, the endoplasmic reticulum (ER) profiles show prominently filled cisternae. Mitochondria are mostly long and slender, or small and ovoid. Most have lamellar cristae, but mixed lamellar-tubular cristae can also be seen. Between decapitated, control and untreated animals no obvious ultrastructural differences could be observed. The peritubular cell sheath surrounding the sex cords did not show signs of differentiation into a layer of myoid cells.

Animals↗
PubMed2005

Respiratory chain complexes and membrane fatty acids composition in rat testis mitochondria throughout development and ageing.

Throughout the maturation of germ cells, a morphological, biochemical and functional differentiation of mitochondria has been shown to occur. Ageing is known to cause changes involved in energy metabolism. These changes have been related to molecular and functional alterations in the properties of biological membranes. Variations in membrane lipid composition and lipid-protein interactions occur with ageing in several tissues. The present paper describes the relationship between these membrane alterations and the activities of lipid-dependent enzymes of isolated testis mitochondria in rats of from 10 days of age to 24 months. The specific activities of these enzymes are lower in preparations from adult and aged rats as compared to those from young rats. Temperature breaks of Arrhenius plots show age-dependent shifts to higher temperatures for the NADH-dehydrogenase, succinate-dehydrogenase, cytochrome c oxidase, and ATPase in senescent animals. Analysis of the membrane fatty acid composition reveals a distinct age-dependent fall in the content of polyunsaturated fatty acids accompanied by an increase in the proportion of saturated fatty acids and a decrease in polyunsaturated fatty acid percentage. The results suggest that during spermatogenesis and the ageing process some changes in the composition of the fatty acids in the surrounding membrane affect the protein-lipid interactions, producing a decrease in mitochondrial enzyme activities.

Adenosine Triphosphatases↗
PubMed1997

Mouse Leydig insulin-like (Ley I-L) gene: structure and expression during testis and ovary development.

Leydig insulin-like protein (Ley I-L) is a novel member of the insulin-like hormone superfamily. We report here the isolation and expression of the mouse Ley I-L gene. The gene encodes a polypeptide of 122 amino acids that shows a relatively weak homology (54%) to human and porcine prepro-Ley I-L. However, the predicted B and A chain of the mature mouse Ley I-L exhibit similarities of 73% and 71% with human and porcine Ley I-L, respectively. Alignment of the 5' flanking region of the mouse gene with those of human and porcine did not exhibit any significant sequence homology. However, it contains the conserved sequence of the Ad4 binding site that is present in all promoter regions of steroidogenic P-450 genes and the Müllerian inhibitor substance gene and is recognized by steroidogenic factor 1. The Ley I-L gene is expressed at a high level in the testis and at a much lower level in the ovary. No transcripts could be detected in placenta prepared between days 10 and 19 of pregnancy. Ley I-L transcripts were first detected in fetal testis at 13.5 dpc. After birth, transcript levels remain constant during the following 3 weeks, increasing at the stage in which the first wave of round spermatids undergo spermiogenesis suggesting a functional role of the Ley I-L in early stages of spermatogenesis and germ-cell maturation. In the ovary, the expression of Ley I-L was first detected at day 6 after birth. The pattern of Ley I-L expression at various stages of the estrous cycle and during pregnancy showed a correlation with follicle development.

Amino Acid Sequence↗
PubMed1980

A comparative study of the development of the fetal testis and ovary in the monkey (Macaca fascicularis).

The gonadal development of the Macaca fascicularis fetus was studied between 37 and 118 days on serial semi-thin and thin sections. The testis and the ovary began to differentiate at the same age (37 days); the definitive architecture of the testis was acquired at 43 days, while a cortex and a medulla did not form in the ovary until 55 to 60 days. In spite of the time-lag and the divergent development, the testis and the ovary evidenced three comparable stages; the main event of these stages was the centrifugal role of the mesonephros. The first stage (37-43 days) included the centrifugal and antero-posterior differentiation of the sex cord anlages from the mesonephric mesenchyme in contact with the proximal loops of the anterior tubules (for a detailed study see Dang and Fouquet, 1979). From 43 days (second stage), a remainder of the mesonephric mesenchymal blastema of the gonad supplied the rete system. The mesonephric tubules fused secondarily with that system which was connected to the sex cords. Whereas in the testis, the rete blastema did not play a direct role in organizing testicular structures, but only in forming excretory pathways, in the ovary, it invaded the medulla (whose initial sex cords degenerated) and penetrated to the ovigerous cords of the cortex. The rete ovarii blastema was probably the major source of periovocyte cells. The third stage included the differentiation of a steroidogenic interstitial tissue (from 50 days in the testis; at about 60 days in the ovary) and is further involution; these processes were similar in both sexes. Observation of the fine structure showed the development of the male and female gonocytes to be the same; the prespermatogonia and the oogonia could be characterized by the formation of nuclear vacuoles. The Sertoli cells and the periovogonial cells showed the same features.

Animals↗
PubMed1994

[Heterochronic occurrence of bilateral torsion of appendix testis a case report].

An 8-year-old boy who had undergone excision of the left appendix testis for torsion of the left appendix testis about one and a half years previously was brought to our department on February 18, 1991 because of right scrotal pain of 4 days' duration. Palpation revealed induration with tenderness of the superior portion of the right testicle. A scrotal ultrasonographic tomogram revealed a shadow probably representing an enlarged appendix testis. The patient was diagnosed as having torsion of the right appendix testis. There was refractory pain, and there was possibility of reactive epididymitis. So the patient underwent excision of the right appendix testis, two right appendices testis, which had enlarged to 5 mm in diameter were found, histopathological examination revealed slight bleeding and marked edema of the stroma, which may have been caused by torsion of the appendix testis. Scrotal pain subsided postoperatively. Bilateral torsion of the appendix testis is very uncommon, and our case is the 14th case reported. Torsion of the appendix testis occasionally develops. Torsion of the appendix testis occasionally develops bilaterally. When a small, tender mass at the superior pole of testis, torsion of the appendix testis should be considered.

Child↗
PubMed2002

Expression-based strategies for discovery of genes involved in testis and ovary development.

In recent years, strategies for gene identification based on differential gene expression have become increasingly popular, due in part to the development of microarray technology. These strategies are particularly well suited to the identification of genes involved in sex determination and gonadal development, which unlike the development of other organ systems, proceeds along two very different alternative courses, depending on the sex of the embryo. We have used a high-throughput, array-based expression screen to identify several genes expressed sex-specifically in developing mouse gonads. One of these, vanin 1, appears to play a role in mediating migration of mesonephric cells into the male genital ridge. Progress in characterizing other genes arising from the screen is discussed.

Animals↗
PubMed1999

Transforming growth factor beta3 in the fetal and neonatal rat testis: immunolocalization and effect on fetal Leydig cell function.

The localization of transforming growth factor beta3 (TGFbeta3) in the fetal and neonatal testis (from fetal day 13.5 to postnatal day 6) was investigated by immunohistochemical staining with a specific polyclonal antibody raised against a synthetic peptide corresponding to residues 50-75 of TGFbeta3. This antibody recognized 0.5 ng TGFbeta3 in western blot analysis, but did not detect 25 ng TGFbeta1 or TGFbeta2. The immunolocalization of TGFbeta3 in the fetal and neonatal testis changed throughout development. Immunostaining was present in the gonocytes by fetal day 13.5, persisted until postnatal day 3, and was heterogeneous in spermatogonia on postnatal day 6. The Sertoli cells contained no immunoreactivity at any age. The fetal-type Leydig cells were first immunostained for TGFbeta3 on day 16.5 and staining became very intense from day 18.5 onward. Staining disappeared when the antibody was presaturated with the synthetic peptide, but persisted when the antibody was presaturated with a tenfold excess of the corresponding peptide from TGFbeta2. Furthermore, we researched whether TGFbeta3 could act as a local regulator of fetal Leydig cell function. In a dispersed fetal testicular cell system, TGFbeta3 inhibited the LH-stimulated testosterone production by Leydig cells from 20.5-day-old fetuses. The inhibitory effect of TGFbeta3 was equal to that observed with TGFbeta1 or TGFbeta2. When compared with our previous studies showing the immunolocalization of TGFbeta1 and TGFbeta2, the present study shows that TGFbeta3 may have a specific role in the developing rat testis, but may also overlap the action of TGFbeta1 and TGFbeta2.

Animals↗
PubMed2005

Cloning and characterization of a novel intronless lactate dehydrogenase gene in human testis.

Using cDNA microarray hybridization from a human testicular cDNA library, one gene named lactate dehydrogenase A-like gene (LDHL, also known as LDHL6B) was cloned. LDHL exhibited 3.8-fold difference at expression level between adult and fetal human testes. The full cDNA length of LDHL is 1680 bp and had a 1145 bp open reading frame, which encoded a 41.9 kDa protein of 381 amino acids. Sequence analysis showed that LDHL harbors all the domains (one lactate/malate dehydrogenase, NAD binding domain and one lactate/malate dehydrogenase, alpha/beta C-terminal domain) in lactate dehydrogenase gene family. Blasting human genome database localized LDHL to human chromosome 15q22.2 and it was an intronless gene. Results of multiple-tissue PCR and real-time PCR showed that LDHL expressed mainly in testis and its mRNA abundance was testis development-related. In summary, LDHL is believed to be involved in testis development and spermatogenesis.

Amino Acid Sequence↗
PubMed1983

The effect of transection of the main vascular and nervous supply of the testis on the development of spermatogenic epithelium in the pig.

The treatment of cryptorchidism in boys by the orchiopexy procedure is variably successful due to several technical problems. In order to study some of these problems a series of experiments in the male pig were undertaken. Experimental simulations of the orchiopexy procedure in the inguinal canal in male pigs with normally descended testes gave rise to damage of the spermatogenic epithelium. Cooling experiments of abdominal testes in adult, naturally cryptorchid pigs indicate that the arrest of spermatogenesis in abdominal testes is not due to an inborn defect but is caused by maintenance of the testes at the abdominal temperature. Evaluation of the data of the interventions with the testicular artery, vein, and nerve, either separately or in combination in the abdomen, showed that there was no effect on the spermatogenic epithelium in any one of the five experimental groups. These experiments confirm that in cryptorchid boys transection of the testicular artery and vein in the abdomen may be indicated if a tension-free fixation of the testis in the scrotum is not achievable, provided that alternate routes are able to take over the main blood supply to the testis.

Animals↗