bluuebunny/crossref_metadata_2025_split huggingface dataset:
Effect of age on seminiferous tubule protein secretion and the adverse effects of testicular toxicants in the rat
Authors: MCLAREN T. T. , FOSTER P. M. D. , SHARPE R. M. 
Published: May 2008

SummaryThis study has assessed the effect of age on protein secretion by seminiferous tubules (ST) isolated from rats and their response to Sertoli cell toxicants. ST were isolated from immature (aged 28 days), late pubertal (aged 45 days) and young adult (aged 70 days) rats and cultured in vitro for 24 h with 35S‐methionine in the presence or absence of FSH (1 mg/ml), m‐dinitrobenzene (m‐DNB) or nitrobenzene (NB) (both at 10‐4M). Incorporation of 35S‐methionine into newly synthesized proteins in the culture medium (secreted proteins) was assessed and the pattern of protein secretion evaluated using two‐dimensional sodium dodecylsulphate polyacrylamide gel electrophoresis (2‐D SDS‐PAGE). These data were compared with those obtained using cultures of immature rat Sertoli cells + germ cells (SC+GC). Addition of FSH, m‐DNB or NB in vitro either had no effect (NB) or had a small stimulatory effect (FSH and m‐DNB) on the incorporation of 35S‐methionine into overall secreted proteins by ST isolated from immature rats. At the same doses, addition of either FSH, m‐DNB or NB to SC+GC co‐cultures resulted in increased incorporation of radiolabel into secreted proteins in all instances. In contrast, the same additions to ST isolated from adult rats resulted in a 20–34% decrease in the overall incorporation of 35S‐methionine into secreted proteins. ST isolated from late pubertal rats showed a similar response to adult rats except that the decreases in incorporation induced by FSH, m‐DNB and NB were smaller. Analysis by 2‐D SDS‐PAGE revealed considerable age‐dependent differences in the proteins secreted by ST from immature and adult rats, of which 13 were identified as being of potential importance. Most of these proteins were prominent secretory products of ST from adult rats, but were minor or non‐detectable products of cultures of ST or SC+GC from immature rats. Most of these proteins disappeared or decreased in abundance after culture of ST with m‐DNB or NB. Two proteins showed the reverse pattern, being more prominent secretory products in immature than mature rats, and their secretion was unaffected or was increased by toxicant exposure. These results demonstrate that there are major age‐dependent differences in the secretion of total and specific proteins by isolated ST and that these are probably related to changes in the germ cell complement with age. The susceptibility of many of these proteins to perturbation by m‐DNB and NB may explain the adverse effects of these compounds on spermatogenesis. Finally, the present results suggest strongly that studies of the regulation of spermatogenesis should not use cells/tissue isolated from immature rats.
The influence of pentoxifylline on motility and viability of spermatozoa from normozoospermic semen samples
Authors: TOURNAYE HERMAN , JANSSENS RONNY , DEVROEY PAUL , VAN STEIRTEGHEM ANDRK 
Published: May 2008

SummaryIn order to evaluate the effects of pentoxifylline on sperm motility and longevity, a controlled in‐vitro study was conducted on normozoospermic donor semen samples using the Cellsoft automated system for sperm motility analysis. After incubation and selection, pentoxifylline was found to improve the recovery of spermatozoa and to increase their velocity. In the subgroup of progressively motile spermatozoa, curvilinear velocity was also enhanced. It is concluded that pentoxifylline has an effect on the vigour, but not on the pattern, of sperm motion. Pentoxifylline did not improve the motility characteristics of senescent spermatozoa in normozoospermic sperm samples. Sperm survival, as shown by supra‐vital staining, and motility longevity both decreased with time after pentoxifylline treatment.
Adenosine triphosphate and motility characteristics of fresh and cryopreserved human spermatozoa
Authors: McLAUGHLIN E. A. , FORD W. C. L. , HULL M. G. R. 
Published: May 2008

SummaryThe concentration of ATP and the motility of human spermatozoa was measured in fresh and cryopreserved cells from the same 15 ejaculates. No coherent picture of the relationship between motility and ATP concentration emerged in whole semen or in spermatozoa washed by repeated centrifugation and resuspension in Biggers Whitten and Whttingham medium. This may have been due to the presence of dead spermatozoa and contaminating cells. After preparation on a Percoll gradient, the ATP concentration in fresh and cryopreserved spermatozoa was the same (6±0.7 nmol/108 spermatozoa) but 85±2.5% of the fresh spermatozoa were progressively motile with an average path velocity of 55±3.5 pm/s compared to corresponding values of 33±5.3% and 44±3.4 μm/s in frozen/thawed spermatozoa. This suggests that the poor motility of cryopreserved spermatozoa does not result from deficient ATP production. No relationship was found between ATP concentration and the ability of motile spermatozoa in the ejaculate to survive freezing.
The value of quantitative DNA flow cytometry of testicular fine‐needle aspirates in assessment of spermatogenesis: a study of 137 previously maldescended human testes
Authors: GIWERCMAN A. , CLAUSEN O. P. F. , BRUUN E. , FRIMODT‐MOLLER C. , SKAKKEBAEK N. E. 
Published: May 2008

SummaryIn order to assess the suitability of DNA flow cytometry of fine‐needle aspirates for quantiftring spermatogenesis, the results from DNA flow cytometry were compared to histological evaluation of testicular biopsies taken concomitantly from 171 previously maldescended testes. In 137 of 171 cases, sufficient material for flow cytometric as well as histological evaluation was obtained.Histological analysis of surgical biopsy specimens revealed spermatogenesis including the spermatid stage in 117 of the 137 gonads. In six of the 117 gonads no haploid cells were found using flow cytometry. On the other hand, surgical biopsies failed to reveal spermatogenesis in five cases in which the corresponding aspirates contained haploid cells. Both methods therefore seem equally sensitive in detection of spermatogenesis.Other types of histological patterns also corresponded to distinct DNA histograms.Thus, in 11 of 12 cases with Sertoli‐cell‐only pattern in all tubules, at least 95% of the cells had a diploid DNA content. Furthermore, predominance of tubules with maturation arrest at the primary spermatocyte level corresponded to an increased proportion of tetraploid cells.When compared to surgical biopsy, DNA flow cytometry of testiclar fine‐needle aspirates is a more objective, easy and rapid method, which is more convenient for the patient. This study has indicatedthat DNA flow cytometry is a suitable method of quantitative assessment of spermatogenesis. One of the first target groups might be men with azoospermia. In such men, DNA flow cytometric analysis of fine‐needle aspirates and surgical biopsy are apparently of equal sensitivity in detecting gonads with spermatogenesis. We conclude that DNA flow cytometry may become an alternative method for the quantification of spermatogenesis.
The hamster Sertoli cell in early testicular regression and early recrudescence: a stereological and endocrine study
Authors: RUSSELL LONNIE D. , CHANDRASHEKAR V. , BARTKE ANDRZEJ , HIKIM AMIYA F. SINHA 
Published: May 2008

SummaryUtilizing stereological techniques, the hamster Sertoli cell at Stage VII of the spermatogenic cycle was examined for early signs of regressive changes after short‐term photoperiod‐induced testicular regression and for early signs of stimulation after short‐term photoperiod‐induced recrudescence. After 4–6 weeks of exposure to short photoperiod, germ cell degeneration was prominent but there were only minimal changes in Sertoli cell structure. These included a decline in volume of multivesicular bodies and in the surface area of mitochondrial membranes and adluminal plasma membrane, while no changes were noted in the other parameters measured. Of the endocrine changes, only plasma FSH levels had declined at this time and there were virtually no significant correlations between plasma FSH levels and the various structural parameters. The general lack of Sertoli cell changes in response to a short exposure to inhibitory photoperiod in the seasonally breeding hamster is similar to the early response to hypophysectomy in the none‐seasonal rat. During early photoperiod‐related recrudescence, testis weight, tubular lumen volume and interstitial parameters were increased. Significantly more preleptotene spermatocytes (about a 50% increase) were present in stage VII tubules, though adluminal germ cells did not increase in number. At this time some structural features of the Sertoli cells were enhanced in volume (cytoplasm, smooth and rough endoplasmic reticulum) and surface area (outer and inner mitochondrial membranes, smooth and rough endoplasmic reticulum and the basal compartment plasma membrane). Blood levels of FSH and testosterone were increased significantly but correlated only with rough endoplasmic reticulum, suggesting that initiation of the synthetic processes was imporant in reestablishment of spermatogenesis. Thus, similar to our findings in the Leydig cell, the structural responses of the Sertoli cell during regression and after recrudescence do not involve the same organelles. During re‐stimulation, we suggest that hormonal stimulation affects primarily synthetic processes in the nucleus and rough endoplasmic reticulum of the Sertoli cell, and that its functions can be re‐established only after these processes have been stimulated.
Improvement in quality of cryopreserved human spermatozoa by swim‐up before freezing
Authors: PÉREZ‐SÉNCHEZ F. , COOPER T. G. , YEUNG C. H. , NIESCHLAG E. 
Published: May 2008

SummarySelecting a population of spermatozoa by the swim‐up technique yields, after freezing and thawing, a population of cells that contains proportionally more spermatozoa which are morphologically normal, fewer spermatozoa with damaged tail membranes, and a greater percentage of progressively motile spermatozoa with greater velocities and amplitudes of head displacement than those obtained after freezing and thawing the same semen samples in the normal way. This pattern was found for the semen from 10 patients and five volunteers. However, the cells selected by swim‐up were as susceptible to the stresses caused by freezing and thawing as unselected spermatozoa in the original semen sample, and the improvement came from the better quality of the initial sample.
Peroxidase‐positive round cells and microorganisms in human semen together with antibiotic treatment adversely influence the outcome of in‐vitro fertilization and embryo transfer
Authors: DE GEYTER CH. , DE GEYTER M. , BEHRE H. M. , SCHNEIDER H. P. G. , NIESCHLAG E. 
Published: May 2008

SummaryHuman semen contains not only spermatozoa but also other cells routinely differentiated as being peroxidase‐positive (e.g. leucocytes) and peroxidase‐negative (eg. immature germ cells and lymphocytes) cells. Considerable uncertainty exists about their role in male fertility. To assess the clinical value of both parameters, and of microorganisms in semen, 391 treatments with in‐vitro fertilization and embryo transfer were analysed retrospectively, and the concentrations of both peroxidase‐positive and ‐negative cells, together with the presence of microorganisms in semen, were compared with both the fertilization and pregnancy rates. The data indicate that the results of treatment were affected only by excessively elevated concentrations of peroxidase‐positive cells (>6×106/ml) and only marginally by the presence of microorganisms in the semen. The pregnancy rate after in‐vitro fertilization and embryo transfer was not improved by antibiotic treatments preceding gamete recovery by several weeks. The increased presence of peroxidase‐negative cells (e.g. germ cells) in semen was not associated with a significant change in the pregnancy rate. However, the concentration of peroxidase‐negative cells in semen correlated significantly with sperm numbers (p<0.01), sperm concentration (p<0.01), and normal morphology rates (p<0.01). It is concluded that short‐term antibiotic treatment of asymptomatic patients before assisted reproduction should be handled with caution. The widespread view that peroxidase‐negative cells in semen are harmful is rejected.
Localization of mRNAs by in‐situ hybridization to the residual body at stages IX‐X of the cycle of the rat seminiferous epithelium: fact or artefact?
Authors: MILLAR M. R. , SHARPE R. M. , MAGUIKE S. M. , GAUGHAN J. , WEST A. P. , SAUNDERS P. T. K. 
Published: May 2008

SummarySeveral recent articles have reported localization of specific mRNAs in the rat testis to stage IX and X seminiferous tubules using in‐situ hybridization. In all cases the expression was located basally in the tubules and appeared as discrete round clusters of grains close to the lamina propria. The localization was interpreted as being in Sertoli cells or leptotene spermatocytes. In this study we demonstrate that this pattern is most probably due to artefactual binding of probes to the residual body (RB).In the present study testicular tissue, perfusion‐fixed with Bouin's and embedded in paraffin, was used, as this resulted in excellent morphological preservation such that RBs within tubules at stages VIII—X were clearly distinguishable. RNA content of the FU3s was demonstrated at stages VIII—X using methyl green pyronin staining, and could be eliminated by pretreatment with RNAse or trichloroacetic acid. Localization of mRNAs for 11 seminiferous tubule proteins was assessed using 35S‐labelled and digoxigenin‐labelled riboprobes (activin receptor‐11, α‐inhibin, transferrin, androgen‐binding protein (ABP), cyclic protein‐2 (CP‐2), CREM, sulphated glycoproteins 1 and 2 (SGP‐1 and SGP‐2), transition protein 2 (TP‐2) and cystatin‐C), and digoxigenin‐labelled oligonucleotide probes (transition protein‐1 (TP‐l), TP‐2 and protamine‐1). All of these probes showed localization to the correct cell type(s) within the seminiferous epithelium. In addition, six antisense riboprobes (activin receptor‐II, CREM, SGP‐2, CP‐2, cystatin C and α‐inhibin) showed hybridization to basally located residual bodies in tubules at stages IX—X on one or more occasions, whereas residual bodies around the edge of the lumen (stage VIII) or in transit through the seminiferous epithelium showed no hybridization; sense probes showed no localization to residual bodies.A common feature of the probes which localized to the basal RBs was that they had been prepared using cDNA cloned into Bluescript SK— vector such that the antisense strand was generated from the T7 polymerase promotor. A cRNA prepared using T7 polymerase and Bluescript vector alone and a GC‐rich 27mer oligonucleotide corresponding to the region of the multiple cloning site of Bluescript adjacent to the T7 site both localized uniquely to basal RB.It is concluded that the hybridization seen within RBs is probably a subtle artefact unique to RBs undergoing dissolution following fusion with Sertoli cell lysosomes, and may reflect nonspecific hybridization to GC‐rich fragments of RNA.
Expression and localization of N‐ and E‐cadherin in the human testis and epididymis
Authors: ANDERSSON ANNA‐MARIA , EDVARDSEN KLAUS , SKAKKEBÆK NIELS E. 
Published: May 2008

SummaryCellular interactions in the testis and epididymis are an important prerequisite for spermatogenesis and sperm maturation, and involve a well‐developed complex of intercellular junctions. Cadherins are cell surface proteins which mediate intercellular Ca2+ ‐dependent adhesion and are believed to be fundamentally important for maintaining multicellular structures. In the present study we report the expression of a 135 kDa N‐cadherin polypeptide in the human seminiferous epithelium by immunoblotting. The presence of N‐cadherin was demonstrated by immunohistochemistry on the surface of spermatogonia and primary spermatocytes, and possibly also around some early spermatids, whereas late spermatids were always negative. Endothelial cells also stained for N‐cadherin, whereas peritubular cells and Leydig cells did not. No expression of E‐cadherin could be demonstrated in the human testis. In the human epididymis E‐cadherin, but not N‐cadherin, was expressed and localized to the surface of the principal epithelial cells as shown by immunohistochemistry. These observations indicate that cadherins play an important role in the organization of the seminiferous and epididymal epithelium.
Increased 17‐OH‐Progesterone levels following hCG stimulation in men with idiopathic oligozoospermia and raised FSH levels
Authors: ANAPLIOTOU M. L. G. , LIPARAKI M. , AMERICANOS N. , GOULANDRIS N. , PAPAIOANNOU D. 
Published: May 2008

SummaryLeydig cell function was investigated in 71 men with idiopathic oligospermia and compared to 14 fertile controls by assessing the steroidogenic response to GnRH and the repetitive administration of hCG (1500 IU X3). The oligospermic men were divided into two groups according to their basal serum FSH values (FSH<8, n=35; FSH>8, n=36), this level being defined by the mean + 3 SD of the levels in normal men (3.71+4.08 mIU/ml). Oversecretion of LH was supported by the findings of: (a) higher basal LH levels (p<0.0001) in both oligospermic groups, although still within the normal range; (b) higher Dmax LH and area LH (p<0.0001) levels in the FSH>8 group; (c) a strong position correlation (p<0.001) of the above parameters with the respective levels of FSH. No difference in basal testosterone levels was observed between the three groups, whereas basal levels of 17‐OHP were sigdcantly higher (p<0.05) in the group with FSH>8. The testosterone/LH ratio was significantly (p<0.0001) lower in the FSH>8 group, and was correlated inversely to the basal blood levels of FSH <0.0001) and to the area LH (p<0.04). After the hCG test, there was no difference in the testosterone and oestradiol response between the groups, whereas the secretion of 17‐OHP and the ratio of 17‐OHP/testosterone was sigdcantly higher (p<0.0001) in the group with FSH>8 compared with the other two groups. Using multiple regression, the total production of 17‐OHP and the 17‐OHP/testosterone ratio were found to be correlated positively with FSH levels. These results support the view that in men with idiopathic oligozoospermia associated with severe Sertoli cell dysfunction there is parallel oversecretion of LH and compensated dysfunction of the Leydig cells, as indicated by oversecretion or accumulation of 17‐OHP after hCG administration, and also by the low testosterone/LH ratio. This is possibly due to alterations in intratesticular paracrine factors deriving from the Sertoli cells, as suggested by the positive correlation between the altered steroidogenic indices and blood FSH levels.
Immunohistochemical localization of proliferating cell nuclear antigen as a tool to study cell proliferation in rodent and primate testes
Authors: SCHLATT STEFAN , WEINBAUER GERHARD E. 
Published: May 2008

SummaryThe immunohistochemical localization of proliferating cell nuclear antigen (PCNA) has become a widely used method for the detection of proliferating cells in cancerous tissues. PCNA expression is maximal around the S phase of the cell cycle. This study has evaluated the applicability of PCNA localization for the analysis of germ cell proliferation in rats, Djungarian hamsters, rhesus and cynomolgus monkeys and men, using Bouin's‐fixed, paraplast‐embedded tissue. In addition, involuted testes from photoinhibited hamsters, testes from immature rhesus monkeys and from GnRH antagonist‐treated rats and cynomolgus monkeys were included. Monoclonal mouse anti‐PCNA antibody (clone: PC10) was used for detection of the antigen. Visualization was performed by immunogold‐silver staining or avidin‐biotin staining. PCNA labelling was confined to the nuclei of spermatogonia and early spermatocytes within the seminiferous epithelium of all species. The distribution of PCNA among the different types of A‐spermatogonia in primates is in good agreement with the previously described proliferation pattern of these cells. No staining was observed in resting A‐dark spermatogonia, while differentiating A‐pale spermatogonia were positive at distinct stages of the seminiferous epithelium cycle. In the rodent species the pattern of labelled A‐spermatogonia was stage‐specific, but agreed only partly with the previously described pattern of mitotic figures of A‐spermatogonia. Hormonal withdrawal induced a decrease in the number of PCNA‐positive cells in adult rats, hamsters and monkeys. In immature testes from rhesus monkeys positive staining was present in spermatogonia but also in some Sertoli cells, indicating proliferative activity of Sertoli cells in the prepubertal stage. It is concluded that PCNA localization can be used to assess the proliferative status of the renewing spermatogonia in the primate testis and to analyse the proliferative activity of the seminiferous epithelium under various endocrine conditions.
Effects of antioxidants on human sperm preparationtechniques
Authors: GRIVEAU J. E. , LANNOU D. LE 
Published: May 2008

SummaryThe effect of two different sperm preparation techniques, Percoll gradient centrifugation and swim‐up from a washed pellet were tested on the functional competence of the selected spermatozoa. Percoll gradient centrifugation brought about an improvement in sperm motility parameters such as curvilinear velocity and straight‐line velocity, an increase in the rates of hyperactivation and the acrosome reaction and an increase in the percentage of motile spermatozoa after 24 h of incubation compared to the centrifugation, swim‐up procedure. The effects of antioxidants such as dithiothreitol (DTT) or reduced glutathione (GSH), and reactive oxygen species‐scavenging enzymes such as catalase or superoxide dismutase (SOD) during the stage of centrifugation before the swim‐up procedure were also studied. Though all of these agents prevented the fall in sperm motility after 24 h incubation, only DTT and SOD improved the rates of both hyperactivation and the acrosome reaction. GSH also improved the acrosome reaction, whereas catalase was without significant effect on the rates of hyperactivation or the acrosome reaction. These results indicate that Percoll gradient centrifugation selects spermatozoa with better functional competence than does centrifugation swim‐up. The damage caused by the centrifugation can be prevented by the addition of antioxidants, suggesting that the differences noted with the Percoll gradient method was due to better protection against peroxidative damage due to the centrifugation of unselected spermatozoa. However, the use of DTT is limited by virtue of the fact that this sulphydryl reducing agent leads to destabilization of the sperm chromatin. In contrast, GSH and SOD could have therapeutic potential.