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Kaposi's sarcoma following chemotherapy for testicular cancer in a homosexual man: demonstration of cytomegalovirus RNA in sarcoma cells.

A 35-year-old white Jewish homosexual man who had undergone surgery and chemotherapy for an embryonal carcinoma of the testis subsequently developed Kaposi's sarcoma. The neoplasm involved the skin as well as visceral tissues. Tissue derived from a biopsy specimen of one of the skin lesions was used in the in situ hybridization technique for the detection of genetic material. Cytomegalovirus messenger RNA was identified in the neoplastic Kaposi cells in the skin. The significance of this finding is discussed.

Cytomegalovirus↗

Isolated testicular polyarteritis nodosa mimicking testicular neoplasm-long-term follow-up.

Polyarteritis nodosa is a systemic vasculitis characterized by segmental necrotizing lesions of medium and small-size arteries. Rarely, the inflammatory process is isolated and involves a single organ without systemic manifestations. We describe a patient with isolated polyarteritis nodosa of the testis who presented with a testicular mass mimicking primary testicular tumor. The postoperative laboratory evaluation was negative. Long-term follow-up, without systemic treatment, showed no evidence of recurrence in the remainder of the testis or development of systemic vasculitis.

Adult↗

Promoter characterization of the human and mouse epilysin (MMP-28) genes.

Epilysin (MMP-28) is a recently cloned member of the matrix metalloproteinase family (Lohi et al., J. Biol. Chem. 276 (2001) 10134). It is expressed at highest levels in the skin by basal and suprabasal keratinocytes, and in testis by developing germ cells. To characterize the epilysin promoter, we isolated a 3.0 kb fragment of human genomic DNA containing 5'-flanking sequence of the epilysin gene, and a corresponding 660 bp fragment from the mouse. The 5'-flanking sequences contain no typical TATA-boxes or CCAAT sequences close to the translation initiation sites. RNase protection assay revealed that two transcription start sites are utilized in the human epilysin gene, situated 210 and 230 bp upstream from the translation start site. The promoter contains a GT-box, situated 300 bp upstream from the translation start site, with homology to the consensus binding site for transcription factors of the Sp family. This site is perfectly conserved between the human and mouse promoters. For reporter gene assays a series of constructs with fragments of increasing length of the epilysin promoter were coupled to the firefly luciferase gene. Reporter gene assays indicated that deletion or mutation of the GT-box dramatically reduces the transcriptional activity both in keratinocytes and in spermatogonia. Gel mobility shift assays showed that several nuclear proteins bind specifically to this sequence. Supershift assays with antibodies specific for members of the Sp family identified Sp1 and Sp3 as components of these protein/DNA complexes and hence as possible regulators of the epilysin gene. Our results indicate that the epilysin promoter has distinctive structural and functional features, which may control the unique expression and regulation patterns of the epilysin gene.

Animals↗

Expression of an antigen associated with basal bodies of human ciliated epithelial cells.

The process and regulation of ciliogenesis in human epithelia is little understood and many components of the cilium and associated structures have not been characterised. We have identified a monoclonal antibody, LhS28, which recognises a 44,000-45,000 M(r) protein specifically associated with human ciliated epithelial cells. Immunoperoxidase labelling of formalin-fixed paraffin wax-embedded human tissues showed that LhS28 was expressed in the sub-apical zone of ciliated epithelial cells of the Fallopian tube and upper respiratory tract, but not ciliated ependyma, non-ciliated epithelia or testis containing developing spermatozoa. Immunoelectron microscopy demonstrated that the antigen recognised by LhS28 was associated with the basal body structure of the cilium and specifically with the 9 + 0 microtubule arrays. LhS28 should be a useful tool in the identification of ciliated cells in pathological specimens and for investigating mechanisms of ciliogenesis.

Antibodies, Monoclonal↗

Expression of a linear Sry transcript in the mouse genital ridge.

Sry is the Y-chromosomal gene that is pivotal in the determination of sex in mammals, however the structure of the Sry transcript produced in the embryo has not been determined. We show here that the transcript expressed in the developing mouse gonad at the sex determining stage of development is linear, polyadenylated and encoded by a single exon, in contrast to the circular, apparently untranslated transcript produced in adult testes. The linear transcript was not detected in any other fetal tissue nor in any adult tissue tested, and was expressed only in the genital ridge portion of the urogenital ridge. The spatial and temporal profile of Sry expression suggests that its role in the mouse fetus is limited to initiating Sertoli cell development during testis determination.

Animals↗

Gamma delta T cells in infection-induced and autoimmune-induced testicular inflammation.

In a previous report, we investigated inflammatory responses induced by injecting Listeria monocytogenes into one testis of a mouse. We demonstrated that the contralateral testis also developed an orchitis despite the absence of bacteria, indicating that the inflammation on the uninfected, contralateral side was of autoimmune character. In both infected and autoimmune testes, gammadelta and alphabeta T cells infiltrated during the inflammation. In this paper, we present the data of a comparison of the character of gammadelta T cells of the infected and autoimmune testes. In both testes, gammadelta T cells appeared to be activated, as assessed by high CD44 and low l-selectin expression. Analysis of T-cell receptor (TCR) usage in both inflammation types revealed the same gammadelta TCR repertoire. Finally, the semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) demonstrated that gammadelta T cells in both types of inflammation were capable of producing interleukin-2 (IL-2), IL-4, interferon-gamma (IFN-gamma), IL-10 and transforming growth factor-beta (TGF-beta). These results imply that gammadelta T cells present in infected-induced and autoimmunity-induced inflammation have the same characteristics and could work as immunoregulatory cells.

Animals↗

Development of the blood-testis barrier in the mouse is delayed by neonatally administered diethylstilbestrol but not by beta-estradiol 3-benzoate.

A group of newborn mice were treated with 1 micro g dose-1 individual-1 of diethylstilbestrol (DES) on alternate days, from days 1 to 11 postnatally. Another group of mice were treated similarly with 125 ng dose-1 individual-1 of beta-estradiol 3-benzoate (E2B). The testes were sequentially examined up to 84 days of age using light and electron microscopy. Spermatogenic cells in the DES-treated mice differentiated normally from birth until 17 days of age, when they differentiated into pachytene spermatocytes and remained at this meiotic prophase for the next 10 days approximately. The cells then began to differentiate further, ultimately forming spermatozoa by 49 days of age. Confocal and electron microscopy showed that the blood-testis barrier did not form until 28 days of age in the DES-treated mice, and a delay in the functional maturation of this structure, as the blood-testis barrier, was confirmed by intercellular tracer experiments. The arrest of spermatogenesis at the meiotic prophase may have been attributable to the DES-induced defective formation of the blood-testis barrier. No delay of the blood-testis barrier formation was detected in the E2B-treated mice. Thus, DES and E2B, both of which are known as potent oestrogenic compounds, had different effects on the Sertoli cells.

Animals↗

Isoforms of angiotensin I-converting enzyme in the development and differentiation of human testis and epididymis.

Angiotensin I-converting enzyme (ACE; CD143, Kininase II, EC 3.4.15.1) is known to be crucial for male fertility in animal models. We therefore studied its testicular (tACE) and somatic (sACE) isoforms in foetal and adult human testis and epididymis using monoclonal antibodies and cRNA probes. During spermatogenesis, tACE was found only in differentiating germ cells and was the only isoform within the seminiferous tubules of adult men. Although tACE mRNA was present in spermatocytes, tACE protein was initially found in post-meiotic step 3 spermatids and increased markedly during further differentiation. The enzyme was strictly confined to the adluminal membrane site of elongating spermatids and was localized at the neck and midpiece region of released and ejaculated spermatozoa. In contrast, sACE was expressed heterogeneously in Leydig cells and endothelial cells of the testicular interstitium, and homogeneously along the luminal surface of epithelial cells lining the ductuli efferents, corpus and cauda of epididymis, and vas deferens. The cell- and site-restricted pattern of sACE corresponded to that found in foetal tissues except an additional and transient expression of sACE in foetal germ cells and foetal Sertoli cells. Our study documents for the first time in humans the regulation and unique cellular distribution of ACE isoforms during the ontogenesis of the lower male genital tract.

Aging↗

The effect of changing the distance between the TATA-box and cap site by up to three base pairs on the selection of the transcriptional start site of a cloned eukaryotic gene in vitro and in vivo.

We have studied how small changes in the distance between the TATA-box and cap site affect transcription of a eukaryotic gene in vitro and in vivo. The trout protamine gene TPG-3 [Gregory et al. (1982) Nucl. Acids Res. 10, 7581-7592] is a good model for such a study as it has (i) a consensus TATA-box 32 base pairs (bp) upstream from an A-residue which is the natural cap site (designated +1) (ii) two further A-residues at -5 and +5, providing alternative transcriptional start sites which are in significantly different sequence environments and (iii) a unique AvaII restriction site immediately downstream from the TATA-box which is ideal for the insertion or deletion of up to 3bp. Transcripts of the wild type and mutant genes were generated in vitro using a HeLa whole cell extract or 'in vivo' by transient expression following their transfection into HeLa cells. These 'spacer' mutations did not affect the efficiency of transcription of the gene in vitro but they did affect the selection of transcriptional start site both in vitro and 'in vivo'. Analysis of 5'-ends by S1-mapping and primer extension showed that the A-residue(s) selected are those which, by insertion or deletion, come to lie on the same face of the DNA double helix as the TATA-box, although the DNA sequence in the immediate vicinity of the potential start sites influences their utilisation. Comparison of the TPG-3 wild type transcripts in these experimental systems with natural mRNA suggests that cap site selection is more stringent in the developing trout testis.

Animals↗

Determining transcript number using the polymerase chain reaction: Pgk-2, mP2, and PGK-2 transgene mRNA levels during spermatogenesis.

We describe a technique that uses reverse transcription and the polymerase chain reaction (pcr) to rapidly quantitate numbers of specific mRNA transcripts from nanogram quantities of total cellular RNA. Linearity of input molecules to output signal was maintained by limiting the cycle number and the amount of input RNA and by minimizing the number of manipulations. Absolute levels of specific transcripts were determined by the inclusion of a separate standard curve composed of serially diluted in vitro transcribed RNA run alongside the experimental samples. This allowed rapid quantitation of many samples simultaneously. We applied this technique to measuring the expression of phosphoglycerate kinase 2 (Pgk-2) transgenes in the mouse testis during development. A human PGK-2 transgene, a PGK-2/CAT transgene, and the endogenous mPgk-2 gene all displayed similar patterns and levels of expression, consistent with the conclusion that peak RNA accumulation occurs in pachytene spermatocytes. Mouse protamine 2 (mP2) is expressed at a level approximately tenfold higher than Pgk-2 and displays a different pattern of expression consistent with initiation of transcription occurring in haploid round spermatids.

Animals↗

Development of the blood-testis barrier in the domestic fowl (Gallus domesticus).

The formation of the blood-testis barrier (BTB) in the domestic fowl was studied at the electronmicroscopic level employing lanthanum as a tracer. No effective barrier could be demonstrated in testes before puberty, although several components of the Sertoli junctional complex such as focal tight junctions and desmosomes were already existent. The time of onset of meiosis after hatching showed great individual variation and meiosis did not occur synchronously in the tubules of a given testis. An effective barrier could first be detected in tubules containing early spermatids, and in which spermatogonia and primary spermatocytes at the leptotene stage were still within the open compartment. Thus, barrier formation was correlated with the occurrence of haploid germ cells. Complete compartmentation of seminiferous tubules, leaving only spermatogonia within the open compartment, was attained in tubules containing elongated spermatids of the maturation phase. In these tubules, primary spermatocytes at the leptotene stage were situated in an intermediate compartment.

Animals↗

Effect of reserpine on growth and sexual development of chickens.

One-month-old male chickens were given injections of 1 mg, 2 mg and 4 mg/kg of reserpine. The injections were repeated at weekly intervals for three months. The chickens receiving reserpine grew at half the rate of untreated control chickens and failed to grow combs. At 4 months of age the testes of reserpine-treated chickens were only about one-tenth the weight of the testes of the control chickens and histologically they showed extreme hypoplasia of the seminiferous tubules. Two large doses of reserpine one month apart could cause atrophy of the testes of adult roosters.

Animals↗

Serum spermagglutinins and semen quality in the bull.

Three bulls injected with homologous semen or testis material developed spermagglutinins in serum while one control bull inoculated with egg white did not. In one bull a high spermagglutinin titre was associated with allergic epididymitis, a decreased proportion of motile spermatozoa, and an increased proportion of tall abnormalities. The palpable lesion persisted for 35 days, the spermagglutinin titre and semen abnormalities for a further 2 months. No change in semen quality was associated with the lower serum spermagglutinin titres in the other two treatment bulls or over comparable periods of time in the control bull. No naturally occurring spermagglutinins were found in a population of 50 bulls with varying semen qualities at an artificial breeding centre.

Agglutinins↗

Proliferative activity of gonocytes, Sertoli cells and interstitial cells during testicular development in mice.

Developing mouse testis was studied from Day 14 post coitum (p.c.) until Day 35 post partum (p.p.) by [3H]thymidine autoradiography. The gonocytes proliferated actively at Day 14 p.c., the [3H]thymidine labelling index (L.I.) being 7.5%, and were quiescent from Day 16 p.c. up to the first day of life, when spermatogenesis started. The L.I. increased to 20% at Day 2 p.p. The L.I. for the Sertoli cells was approximately 20% before birth. After birth the proliferative activity decreased. After Day 11 p.p., the Sertoli cells showed their typical adult appearance. After Day 17 p.p. no labelled Sertoli cells were observed. The Leydig cells featured a very low proliferative activity up to Day 21 p.p. (L.I. of maximal 1.9%). At Day 29 p.p. there was a peak of 7.4% in L.I., followed by a sharp decrease to 0.35% at Day 35 p.p. The L.I. of mesenchymal cells decreased from 11.4% at Day 14 p.c. to 1.1% at Day 14 p.p. and remained more or less constant thereafter. The proliferative activity of myoid, endothelial and perivascular cells followed a similar course to that of mesenchymal cells, their L.I.s being high before birth (16, 12.5 and 19%, respectively, decreasing until Day 14 p.p. (0.6, 2.0 and 1.2%, respectively) and thereafter being more or less constant. There was an increase in the relative number of Leydig cells from approximately 4% of the total interstitial cell number at Day 14 p.p. to 29.5% at Day 35 p.p. At the same time, the relative number of mesenchymal cells decreased from 55 to 13%. The diameter of the seminiferous tubules showed a peak of 92 microns at Day 16 p.c., decreased to 44 microns at Day 1 p.p. and increased again to 204 microns at Day 33 p.p. These results show that, except for the Leydig cells, the proliferative activity of testicular cell types is highest during the pre- and early postnatal period. The major outgrowth of the Leydig cell population occurs around the fourth week after birth. The results are in accordance with the hypothesis that the mesenchymal cells are the progenitors of Leydig cells.

Animals↗