bluuebunny/crossref_metadata_2025 huggingface dataset:
The Human Bulbo‐Urethral Glands: A Transmission Electron Microscopy and Scanning Electron Microscopy Study
Authors: RIVA ALESSANDRO , USAI ENZO , COSSU MARGHERITA , SCARPA ROBERTO , TESTA‐RIVA FRANCESCA
Published: December 2015

This paper investigates the ultrastructure of human bulbo‐urethral glands using specimens obtained at surgery. The tubulo‐alveolar endpieces of these glands are lined by typical mucous cells in different stages of the secretory cycle. The most interesting features of their cytoplasm are membrane‐bounded bodies with a filamentous texture that usually fuse with the mucous droplets before they discharge into the lumen. Cells with apical dark granules are sometimes encountered in the ductal portions of the gland, which are probably ductal cells endowed with a scanty synthetic and secretory activity. Myoepithelial cells are not very numerous and are observed around mucous cells only.
Changes in Testicular Morphology in Boars Actively Immunized Against Gonadotropin Hormone‐Releasing Hormone
Authors: AWONIYI CALEB A. , CHANDRASHEKAR VARADARAJ , ARTHURS ROBERT D. , SCHANBACHER BRUCE D. , FALVO RICHARD E.
Published: December 2015

Alterations in testicular morphology were studied in boars actively immunized against gonadotropin hormone releasing hormone (GnRH). Ten boars were divided equally into two experimental groups (five GnRH‐immunized, and five controls). Antibody production was achieved by conjugating GnRH to human serum globulin (hSG). The GnRH‐hSG conjugate was emulsified in complete Freund's adjuvant, and administered to boars at 12 weeks of age. Boars were given a booster in incomplete Freund's adjuvant on week 18 and 20. The presence of high antibody titers to GnRH caused luteinizing hormone and testosterone to decline to nondetectable levels. Morphometric examination showed a reduction in percentage volume in Leydig cells/unit testis, seminiferous tubule diameter and seminiferous epithelial height, and an increase in non‐Leydig cell interstitial tissue in GnRH‐immunized boars compared with controls. Histologic evaluation displayed severe damage of the seminiferous epithelium, absence of spermatids, incomplete cell associations, disruption of Sertoli cells, formation of multinucleated giant cells, and a striking reduction in size and cytoplasmic structures of Leydig cells in GnRH‐immunized animals. These results demonstrate the potent inhibitory effects of GnRH immunoneutralization on the boar reproductive system.
Isolation and Characterization of the Plasma Membrane of Rat Cauda Epididymal Spermatozoa
Authors: AGRAWAL PRABHA , MAGARGEE SUSAN F. , HAMMERSTEDT ROY H.
Published: December 2015

Cauda epididymal rat spermatozoa were isolated by flushing the excised epididymis and the plasma membrane was detached by a nitrogen cavitation treatment (500 psi, 10 minutes equilibration at 4 C). Membrane vesicles were recovered after sucrose gradient centrifugation. Portions of the sperm surface releasing the plasma membrane were assessed by light microscopy of fluoroscein isothiocyanate‐succinylated concanavalin A‐treated spermatozoa and by transmission electron microscopy. Plasma membrane was detached from the region overlying the acrosome from most spermatozoa and from the middle‐piece overlying the mitochondria from some cells. Thus, the fraction analyzed was derived from at least two portions of the sperm surface. The fractions from the sucrose density gradient were analyzed for gross chemical composition (phospholipid, protein and sterol) and the protein components were detected after electrophoresis under denaturing conditions; the peak fractions (at density ∼1.13 g/ml) were judged homogeneous. Replicate analyses of such preparations established mass ratios of protein to phospholipid of 0.63, total sterol to phospholipid of 0.18, and demosterol to cholesterol of 0.32. The molecular composition of the phospholipid fraction was determined to be 10% phosphatidylserine (mole percent), 3% phosphatidylinosital, 3% sphingomyelin, 31% phosphatidylethanolamine, 27% phosphatidylcholine, 10% diphosphatidylglycerol and 5% of an unknown component. Fatty acyl analyses of the phospholipid fraction revealed that ∼70% of the residues consisted of palmitoyl (16:0) and stearoyl (18:0) acyl groups, with the balance distributed among various unsaturated acyl groups (18:1, 22:3, 22:4 and 22:5); about 40% of the recovered phospholipids represented ether acyl phosphatides. Differences in the lipid composition of rat vesicles described here and similar vesicles isolated from ram and boar spermatozoa (described previously) are discussed. The partitioning of the nitroxyl spin label 3‐doxylheptane into vesicles isolated from rat and ram spermatozoa was assessed by electron paramagnetic resonance spectroscopy at temperatures between 4 C and 26 C; no difference in the response of the spin label in the two vesicle preparations was detected.
Electron Microscopic Study of the Secretion Process in Bovine Reproductive Organs
Authors: AGRAWAL YASHPAL , VANHA‐PERTTULA TAPANI
Published: December 2015

The origin and mechanism of the secretion of membrane‐bound particles in bovine seminal plasma were studied with transmission (TEM) and scanning (SEM) electron microscopy of the epididymis, vas deferens, ampulla, and seminal vesicle of adult bulls. In the SEM study, all these organs were found to contain apical protrusions in the lining of the epithelial cells. Eventually the protrusions became detached and formed secretory bodies within the lumina of these organs. In the epididymis, the TEM study disclosed a granular and rather homogeneous content in the protrusions and bodies, whereas in the vas deferens they contained dilated cisternae of smooth endoplasmic reticulum. In the ampulla and seminal vesicle, the formation of the apical protrusions was associated with an accumulation of membrane‐bound vesicles. These vesicles were found to be released from the storage bodies into the secretory fluid of the lumen. Both could be harvested from isolated seminal vesicle secretions by Percoll gradient centrifugation. It was concluded that various parts of the bovine reproductive organs discharge their secretory products at least partly by an apocrine mechanism. The membrane‐bound particles in the seminal plasma, however, appear to be mainly derived from the ampulla and seminal vesicle.
Study of the Accuracy of Physical and Biochemical Markers in Semen to Detect Infectious Dysfunction of the Accessory Sex Glands
Authors: COMHAIRE FRANK H. , VERMEULEN LUTGARD , PIETERS ONNY
Published: December 2015

Infection of the male accessory sex glands may result in impaired secretory function and alteration of the composition of seminal plasma. Using receiver operating characteristic curves and accuracy tests, the power of several biochemical and physical markers was evaluated for their ability to discriminate between semen of infected and noninfected infertile men. The total output of citric acid had the strongest discriminating power, followed by acid phosphatase and gamma‐glutamyltranspeptidase. Measurement of the concentration of fructose was found to be nondiscriminatory.
Effect of Immobilization Stress on Testosterone and Inhibin in Male Rats
Authors: DEMURA R. , SUZUKI T. , NAKAMURA S. , KOMATU H. , ODAGIRI E. , DEMURA H.
Published: December 2015

Adult male Wistar rats were subjected to acute and chronic immobilization stress. Changes in plasma levels of LH, FSH and testosterone and in the content of testicular inhibin and testosterone were studied. Plasma LH levels decreased significantly in response to both acute and chronic stress. Significant decreases in plasma testosterone content were also observed after chronic stress. In contrast, plasma FSH and testicular bioassayable inhibin content did not change after acute or chronic stress. These findings indicate that in male rats immobilization stress induced a dissociation in LH and FSH responses, and decreased testosterone while inhibin remained unaffected.
Normal Gonadal Functions and Fertility After 23 Months of Treatment of Prepubertal Male and Female Dogs with the GnRh Agonist [D‐Trp6, des‐Gly‐NH210]GnRH Ethylamide
Authors: LACOSTE DANIEL , DUBÉ DONALD , TRUDEL CLAUDE , BÉLANGER ALAIN , LABRIE FERNAND
Published: December 2015

The GnRH agonist [D‐Trp6, des‐Gly‐NH210]GnRH ethylamide was administered to prepubertal male and female dogs daily for 23 months by subcutaneous injection. During GnRH agonist treatment, plasma steroid levels, namely dehydroepiandrosterone, androst‐5‐ene‐3β‐17β‐diol, androstenedione, testosterone, dihydrotestosterone, 5α‐androstane‐3α,17β‐diol, 5α‐androstane, 3β,17β‐diol were markedly inhibited in male animals, whereas in female animals, the plasma concentrations of DHEA and Δ5‐diol were decreased. Within 2 months following cessation of therapy, all steroids increased to normal adult levels. Morphological studies reveal that treatment of male animals with the GnRH agonist is accompanied by a small volume of seminiferous tubules, Leydig cells, and prostate gland, whereas in the ovaries of female animals, there is a large number of primordial follicles, a few primary follicles, but no secondary follicles. In the pituitary gland of animals of both sexes, LH‐secreting cells have high levels of glycogen particles in their cytoplasm and tend to be either of normal appearance with dilated rough endoplasmic reticulum (RER) or strongly atrophied with a dark‐stained cytoplasm, a contraction of RER, and a decrease in the number of secretory granules. Reticular cells of the connective tissue also show high levels of glycogen particles. After the 14 month recovery period, spermatogenesis has a normal adult appearance, the prostate gland shows a normal secretory epithelium, and secondary follicles are easily observed in the ovary. Gonadotrophs are free of glycogen accumulation, but reticular cells continue to show an accumulation of glycogen particles in their cytoplasm. Two male and two female animals were mated after the recovery period and produced normal offspring with normal fertility. The present results indicate that GnRH agonist treatment achieves a blockade of sexual maturation and that following cessation of treatment, normal pituitary‐gonadal functions resume, with appararently normal fertility and normal offspring.
Monoclonal Antibodies to Epididymis‐specific Proteins Using Mice Rendered Immune Tolerant to Testicular Proteins
Authors: Joshi Saurabh A. , Ranpura Sandeep A. , Khan Shagufta A. , Khole Vrinda V.
Published: December 2015

ABSTRACT: Monoclonal antibodies (mabs) have been used as a powerful tool for identification of newer sperm proteins. However, conventional hybridoma technology rarely provides chance to obtain mabs to epididymal proteins. To increase this chance, we have used an alternate method of neonatal tolerization. In this protocol, animals were tolerized at birth using testicular proteins followed by immunization with cauda epididymal sperm protein (which is a cocktail of proteins both from testicular and epididymal origin). This protocol induced a specific immune response to epididymal sperm proteins. Spleen from one of these animals was then used for preparation of mabs. This fusion resulted in a number of mabs reacting specifically to epididymal proteins. Although mabs identified a protein of approximately similar molecular weight on 1‐dimensional Western blot analysis, there were differences in regional localization on rat sperm as seen by indirect immunofluorescence. Immunohistochemical localization of these proteins in rat epididymis showed region specific synthesis. The synthesis of proteins was seen in the distal caput epididymis, and maximum expression was seen in supranuclear region of corpus epithelium. The proteins were localized on sperm from corpus and cauda region. Epididymis specific synthesis of the proteins and agglutinating nature of the mabs to these underlines the functional importance of these proteins in sperm maturation in epididymis. These antibodies could therefore, be used as tools for understanding the physiology of maturation of sperm in epididymis and role of the epididymal protein in fertilization.
Absolute Polymorphic Teratozoospermia in Patients With Oligo‐Asthenozoospermia Is Associated With an Elevated Sperm Aneuploidy Rate
Authors: Vicari Enzo , de Palma Adele , Burrello Nunziatina , Longo Giusi , Grazioso Caterina , Barone Nunziata , Zahi Mohamed , D'agata Rosario , Calogero Aldo E.
Published: December 2015

ABSTRACT: Infertile patients with abnormal sperm parameters have an increased sperm aneuploidy rate, despite a normal blood karyotype. The evaluation of sperm chromosome aberrations in patients with teratozoospermia only has shown a rate similar to that found in patients exhibiting oligo‐astheno‐teratozoospermia, which suggests that teratozoospermia is the critical parameter associated with aneuploidy. However, it is not known which alteration of the sperm morphology is associated with chromosome aberrations. The few cases reported so far have shown an association with the presence of abnormal head morphology and particularly with enlarged heads. We report the sperm aneuploidy rate of 3 patients with oligo‐asthenozoospermia who have absolute teratozoospermia (100% abnormal forms) and a different percentage of sperm head abnormalities. Fourteen healthy men with normozoospermia served as control subjects. Sperm aneuploidy and diploidy rates were calculated by using triple‐color fluorescence in situ hybridization (FISH) for chromosomes 12, X, and Y, and double‐color FISH was used for chromosomes 8 and 18. Patient K53, who had the highest number of spermatozoa with enlarged heads (54.3%), also had the highest aneuploidy and diploidy rates. The other 2 patients, K56 and K61, had sperm aneuploidy and diploidy rates lower than those of patient K53 but still well above the range found in normal men. Sperm chromosome abnormalities were intermediate in patient K61 and lower in patient K56, who had the lowest rate of spermatozoa with enlarged heads (18.9%). These data add further evidence that patients with teratozoospermia have an increased sperm aneuploidy rate and that this is particularly high in presence of an elevated percentage of spermatozoa with enlarged heads. For this reason, germ cells exhibiting this abnormality should not be used in in vitro fertilization programs.
Development of an Inexpensive Artificial Vagina for Semen Collection from Rabbits
Authors: Naughton Cathy K. , Nelson David R. , Thomas Anthony J.
Published: December 2015

ABSTRACT: The rabbit is well suited for infertility research because of the animal's size, relatively low cost, and accessibility of the genitalia. There are several studies reporting sperm count and motility in rabbits; however, no easily reproducible or inexpensive device for semen collection has been reported. Herein, we report the construction of an inexpensive, effective artificial vagina assembled from easily obtainable products that may be used to collect rabbit ejaculates. This device was used to perform 243 ejaculation trials on 17 rabbits. Eighty‐six percent (209/243) of the trials resulted in rabbit mounts. Overall, 91% (191/209) of mounts resulted in successful semen collection. After 60 mounts, a 98% successful ejaculate collection rate was achieved. The cost of the reusable device is $2.64, plus silicone condoms at $7.50 each. This practical artificial vagina for semen collection is an inexpensive and effective means of obtaining semen from rabbits for andrologic study.
Pharmacokinetics and Degree of Aromatization Rather Than Total Dose of Different Preparations Determine the Effects of Testosterone: A Nonhuman Primate Study in Macaca fascicularis
Authors: Weinbauer Gerhard F. , Partsch Carl‐Joachim , Zitzmann Michael , Schlatt Stefan , Nieschlag Eberhard
Published: December 2015

ABSTRACT: Currently available testosterone (T) preparations differ substantially in their pharmacokinetic profile that might influence their androgenic properties in terms of suppression of the gonadal axis, effects on anabolic parameters, lipid metabolism, and erythropoiesis. The present work was undertaken to determine the physiological effects of three T preparations with different serum kinetics. Twenty adult male cynomolgus monkeys (Macaca fascicularis) were randomly assigned to receive treatment for 28 weeks with either T enanthate (TE) every 4 weeks, T buciclate (TB) every 7 weeks, or T undecanoate (TU) every 10 weeks or remaining untreated (controls). Each injection delivered 20 mg pure T per kilogram body weight. Pharmacokinetic profiles demonstrated higher peak levels of T for TE‐treated animals; serum half‐lives were longer for TU or TB. Estradiol levels (area under the curve) were significantly higher in TB vs TU or TE. All T regimens suppressed serum luteinizing hormone bioactivity and testicular volumes declined (all P < .001 vs controls). Sperm counts were markedly lowered in all animals but least in TE (P < .01 vs TB or TU). During recovery phase, return to normal for all three parameters occurred significantly earlier in TE‐treated animals, followed by those given TU, compared with TB (all P < .001 between groups). Body weight increased significantly during T exposure. This effect was stronger and more sustained in TB vs TU or TE (both P < .001). Serum creatinine and hemoglobin increased with high significance in all T‐treated animals (all P < .001 vs controls). The lowering impact of T on serum lipids was markedly stronger in the longer‐acting T preparations in comparison with TE, as were effects on purine metabolism (all P < .001). The pattern of exposure and degree of aromatization rather than overall exposure to T determine its effects in the preclinical primate model. Both fluctuations of androgen concentrations and the conversion rate to estradiol influence gonadal suppression as well as metabolism. These results have to be considered in men receiving treatment for hypogonadism or regimens for hormonal contraception.
Evaluation of Chromosome Breakage and DNA Integrity in Sperm: An Investigation of Remote Semen Collection Conditions
Authors: Young Karen E. , Robbins Wendie A. , Xun Lin , Elashoff David , Rothmann Susan A. , Perreault Sally D.
Published: December 2015

ABSTRACT: Collection of ejaculated semen at a remote site (outside of the laboratory) would facilitate participation rates and geographic diversity in reproductive epidemiology studies. Our study addressed concerns that remote collection and overnight mail return might induce chromosome/DNA damage. We collected semen from 10 healthy men. Part of each sample was snap frozen in liquid nitrogen and the rest held at 22 ± 1°C for 24 hours in a transport container (simulating ambient temperature during overnight return) then snap frozen. DNA breakage and fragmentation were measured using tandem‐label sperm‐fluorescence in situ hybridization (FISH), terminal deoxynucleotidyl transferase‐mediated dUTP nick end‐labeling (TUNEL), and neutral comet assay. Tandem‐label sperm‐FISH and TUNEL detected no statistically significant difference between sperm fresh frozen (FF) and those frozen after 24 hours (F24). The mean frequency of chromosome breakage per 10 000 cells scored in sperm‐FISH for FF and F24 was 10.5 ± 1.3 breaks and 11.2 ± 1.1 breaks, respectively (P = .69, Student's t test). The mean frequency of TUNEL‐positive cells per 2000 cells scored in FF and F24 was 136 ± 29 and 213 ± 28 cells, respectively, which approached but did not reach statistical significance (P = 0.07, Student's t test). The neutral comet assay detected a statistically significant difference in DNA strand breakage between the 2 groups (percentage of DNA in the tail P = 0.037; tail moment P = 0.006; and tail length P = 0.033, all Student's t test). The mean frequency of damage denoted by tail length in μm per 100 cells scored in FF and F24 was 175.0 ± 15.5 and 152.2 ± 17.6 μm, respectively. Tandem‐label sperm‐FISH, TUNEL, and neutral comet assay are useful analytical techniques for laboratory‐based studies of human sperm genomic integrity; however, for field studies incorporating the nonrefrigerated return of semen after 24 hours, only chromosome breakage at a level that can be detected using tandem‐label sperm‐FISH was unaffected. TUNEL and neutral comet assay need further study before they are used in specimens collected at remote sites and transported to a central laboratory.
Semen Quality of Men With Asymptomatic Chlamydial Infection
Authors: Hosseinzadeh Saeid , Eley Adrian , Pacey Allan A.
Published: December 2015

ABSTRACT: We have shown previously that the in vitro exposure of spermatozoa to elementary bodies (EBs) of Chlamydia trachomatis can lead to sperm death over a number of hours of incubation. As such, we have hypothesized that the ejaculates of men with a chlamydial infection could contain increased numbers of nonmotile (dead) spermatozoa if they are exposed to EBs prior to ejaculation. To test this hypothesis, the ejaculates of 642 men undergoing diagnostic semen analysis as part of ongoing infertility investigations with their partner were examined. All men were without symptoms of genitourinary infections and semen analysis was performed according to World Health Organisation (WHO) 1999 methods after a 3–5 day abstinence period. In addition to semen analysis, nested plasmid polymerase chain reaction (PCR) was undertaken on the ejaculate to detect the presence of C trachomatis DNA. A total of 31 semen specimens (4.9%) were found to be positive, and in 28 of these, the diagnosis was confirmed using the ligase chain reaction (LCR). Men whose ejaculates were PCR positive for chlamydial DNA had a significantly (P < .05) higher mean concentration of leukocytes (1.71 ± 2.20 × 106 per mL) and a higher mean ejaculate volume (3.45 ± 1.52 mL) than in those whose ejaculates were PCR negative (leukocyte concentration: 0.67 ± 2.59 × 106 per mL; volume 2.93 ± 1.38 mL). Leukocytospermia was twice as common in men that were PCR positive for chlamydial DNA (P < .05) but it was not always associated with the presence of chlamydial DNA in semen. However, there was no difference in the mean percent motility between the 2 groups and the proportion of asthenozoospermia also did not differ. Because these results do not confirm the hypothesis proposed from our in vitro experiments, further work needs to be undertaken to understand whether human spermatozoa are actually exposed to elementary bodies of C trachomatis in an infected individual prior to ejaculation.
Gonadotropin‐Mediated Regulation of the Murine VEGF Expression in MA‐10 Leydig Cells
Authors: Schwarzenbach Heidi , Chakrabarti Gopa , Paust Hans Joachim , Mukhopadhyay Amal K.
Published: December 2015

ABSTRACT: Presence of vascular endothelial growth factor (VEGF) is not only limited to cells directly involved in angiogenesis but has also been demonstrated in steroidogenic cells like testicular Leydig cells. Because Leydig cells are subjected to regulation by gonadotropic hormones and produce steroid hormones, we have investigated here the effects of human chorionic gonadotropin (hCG) or steroid hormones on VEGF expression in cultured mouse tumor Leydig cells (MA‐10 cells) and have then analyzed the underlying molecular mechanisms. Northern blot analysis and enzyme‐linked immunosorbent assays revealed increases in VEGF mRNA and protein levels, respectively, over 3–20 hours in MA‐10 cells after stimulation with hCG or 8‐Br‐cAMP. Although MA‐10 cells lack the classical progesterone receptor, progesterone was able to stimulate VEGF expression. Promoter analyses and antibody supershift experiments suggested that the proximal region is able to constitutively bind the transcription factors Sp1 and Sp3. Mutations of 2 potential Sp1 binding sites in the proximal region showed the requirement of these motifs for stimulation of VEGF by hCG and 8‐Br‐cAMP. The distal cytosine‐rich sequence interacts with so far‐unidentified faster migrating factors. Following stimulation with hCG or 8‐Br‐cAMP, the binding of these proteins was increased in the complexes formed in the proximal and distal regions. VEGF expression in Leydig cells is regulated by gonadotropin via a cAMP‐dependent mechanism, and the transcription factors Sp1 and Sp3 appear to be involved in the activation of the promoter. Progesterone also appears to play a role in the regulation of VEGF, acting presumably via a nonconventional receptor that remains to be characterized yet.
Limited Processing of Pro‐Matrix Metalloprotease‐2 (Gelatinase A) Overexpressed by Transfection in PC‐3 Human Prostate Tumor Cells: Association With Restricted Cell Surface Localization of Membrane‐Type Matrix Metalloproteinase‐1
Authors: Wilson Michael J. , Jiang Aixiang , Wiehr Carol , Wang Xing , Sinha Akhouri A. , Pei Duanqing
Published: December 2015

ABSTRACT: The expression and activation of matrix metalloproteinases (MMPs) by tumor cells is correlated with progression to invasive and metastatic status. The purpose of this study was to examine the role of increased MMP‐2 (gelatinase A) expression in prostate cancer progression utilizing human prostate PC‐3 cancer cells that overexpress MMP‐2 using gene transfection. PC‐3 cells were transfected with pCR‐3 vector only and pCR‐3 MMP‐2 plasmids employing the LipofectAMINE method, and stable transfectants were selected with G418. The expression of MMP‐2, tissue inhibitor of metalloproteinase‐2 (TIMP‐2), and membrane‐type MMP 1 (MT1‐MMP) in PC‐3 parental and transfected cells under serum‐free conditions was determined by zymography, immunoblotting, immunofluorescent microscopy, Northern blotting, and/or reverse transcriptase‐polymerase chain reaction (RT‐PCR). MMP‐2 transfected cells produced primarily the proenzyme form of MMP‐2; the parental and vector control transfected PC‐3 cells did not express any MMP‐2 that was detectable by the methods we employed. Treatment of PC‐3 MMP‐2 transfected cells with Concanavalin A (Con A), in contrast to HT‐1080 cells, processed only a small amount of the secreted 72‐kd proenzyme to a 62‐kd intermediate and a cell‐associated 59‐kd active form. The low level of secreted pro‐MMP‐2 processing induced by Con A was inhibited by serine protease inhibitors and was unaffected by cyclic adenosine monophosphate (cAMP). Immunoblotting showed that these cells produced abundant TIMP‐2 and lower amounts of MT1‐MMP in comparison with Con A—responding HT‐1080 cells. HT‐1080 cells respond to Con A by translocating MT1‐MMP from intracellular localization sites to the plasma membrane, an effect not observed in PC‐3 cells. The molecular basis for the low level of processing of pro‐MMP‐2 by PC‐3 cells may be due to an overabundance of TIMP‐2 and/or a low level of cell surface active MT1‐MMP.
Cell Specificity of Aquaporins 0, 3, and 10 Expressed in the Testis, Efferent Ducts, and Epididymis of Adult Rats
Authors: Hermo Louis , Krzeczunowicz Dominic , Ruz Ricardo
Published: December 2015

ABSTRACT: Aquaporins (AQPs) are transmembrane protein channels that allow the rapid passage of water across an epithelium at a low energy requirement, though some also transport glycerol, urea, and solutes of various sizes. At present, 11 members of the AQP family of proteins have been described in mammals, with several being localized to the testis (AQP‐7 and AQP‐8), efferent ducts (AQP‐1 and AQP‐9), and epididymis (AQP‐1 and AQP‐9) of adult rats. With the discovery of expression of multiple AQPs in different tissues, we undertook a systematic analysis of several other members of the AQP family on Bouin‐fixed tissues of the male reproductive tract employing light microscope immunocytochemistry. In the testis, AQP‐0 expression in the seminiferous epithelium was restricted to Sertoli cells and to Leydig cells of the interstitial space; no reaction was observed in the efferent ducts or epididymis. In Sertoli cells, a semicircular pattern of staining was noted, with only one fourth or one half of the Sertoli cells of a given tubule showing a reaction product. Furthermore, while Sertoli cells at stages VI—VIII of the cycle showed intense staining, those at stages IX—XIV were least reactive, with Sertoli cells at stages I—V showing intermediate levels of reaction product. The epithelial expression of AQP‐10 was restricted to the microvilli of the nonciliated cells and the cilia of the ciliated cells of the efferent ducts; however, the endothelial cells of vascular channels of the efferent ducts and epididymis were also intensely reactive. AQP‐3 expression was localized exclusively to the epididymis, where intense staining was noted exclusively over basal cells. Examination of orchidectomized rats revealed that AQP‐3 expression was abolished over basal cells and that it was greatly diminished after efferent duct ligation. As the reaction was not fully restored in orchidectomized animals supplemented with high levels of testosterone, we suggest that AQP‐3 expression in basal cells is regulated in part by testosterone, in addition to a luminal factor emanating from the testis. Together, the data indicate a cell‐ and tissue‐specific expression for AQP‐0, AQP‐3, and AQP‐10 in the testis, efferent ducts, and epididymis, as well as differential regulating factors for the expression of AQP‐3 in basal cells.
Inhibitors of Phosphoinositide 3‐Kinase, LY294002 and Wortmannin, Affect Sperm Capacitation and Associated Phosphorylation of Proteins Differently: Ca2+‐Dependent Divergences
Authors: Nauc Veronica , de Lamirande Eve , Leclerc Pierre , Gagnon Claude
Published: December 2015

ABSTRACT: Sperm capacitation is regulated by multiple pathways that also control sperm motility and tyrosine (Tyr) phosphorylation of several sperm proteins. Among the reported pathways, phosphoinositide 3‐kinase (PI3K) signaling and its role in modulating sperm postejaculatory changes and motility remain elusive. It was shown that wortmannin, a selective inhibitor of PI3K, prevents human sperm acrosome reaction. Using LY294002 (2‐(4‐morphlinyl)‐8‐phenyl‐4H‐1‐benzopyran‐4‐one), another chemically different inhibitor of PI3K, it was suggested that this enzyme inhibits human sperm motility. In this study, we used the 2 known inhibitors of PI3K to investigate their effect on sperm capacitation and associated protein phosphorylation events. Our data show that sperm incubated with LY294002 undergo capacitation and increased Tyr phosphorylation of specific sperm proteins in a manner similar to that promoted by the capacitation inducer fetal cord serum ultrafiltrate (FCSu), as well as double phosphorylation of the threonine (Thr)‐glutamine (Glu)‐Tyr motif. Under similar conditions, wortmannin did not affect these sperm functions on its own, although it did prevent the effect induced by FCSu. Consistently, wortmannin decreased the phospho (P)‐Tyr content of sperm proteins and prevented the phosphorylation of their Thr‐Glu‐Tyr motif. We also show by means of immunoblotting and cell fractionation experiments the presence of PI3K and its downstream effector Akt (protein kinase B) at the membrane level, as well as sperm heads and flagella. Our data show that human spermatozoa contain a consensus motif usually phosphorylated by Akt and that its P‐serine (Ser)/Thr content is increased by both LY294002 and FCSu, while it is decreased by wortmannin. In addition, the 2 inhibitors differently affected the intracellular calcium concentration, [Ca2+]i. While LY294002 increased [Ca2+]i, wortmannin did not affect its content and did not prevent the LY294002 effect. Thus, we propose that the LY294002‐promoted increase in [Ca2+]i operates independently of PI3K. In conclusion, we suggest that special care be taken when using LY294002 to investigate the role that PI3K plays in a cellular phenomenon.
Semen Characteristics After Overnight Shipping: Preservation of Sperm Concentrations, HspA2 Ratios, CK Activity, Cytoplasmic Retention, Chromatin Maturity, DNA Integrity, and Sperm Shape
Authors: Huszar Gabor , Celik‐Ozenci Ciler , Cayli Sevil , Kovacs Tamas , Vigue Lynne , Kovanci Ertug
Published: December 2015

ABSTRACT: We tested several approaches that can be used to preserve sperm attributes and the objective biochemical markers of sperm maturity and function for assessment in a remote centralized laboratory after overnight shipping of semen samples. Addition of phenyl‐methyl‐sulfonyl‐fluoride (PMSF) to a final concentration of 20 μg/mL semen at 4°C has preserved sperm concentrations and HspA2 isoform ratios, even at room temperature, simulating a shipping delay in moderate ambient temperatures. Regarding the attributes of individual spermatozoa, the patterns of CK‐immunocytochemistry (demonstrates cytoplasmic retention in diminished‐maturity spermatozoa); aniline blue staining pattern (tests chromatin maturity); sperm shape assessed by both Kruger strict morphology and computer assisted morphometry; and sperm DNA integrity, as tested by DNA nick translation, all remained unchanged. Thus, the PMSF‐4°C conditions preserved sperm concentrations and the cytoplasmic and nuclear biomarkers of sperm cellular maturity and function for next‐day analysis. This shipping method will facilitate the early detection of subtle changes in semen quality that can affect sperm function, even when there has been no decline in sperm concentrations to signal possible toxic effects. Furthermore, sample preservation will enable investigators to evaluate semen for toxicology studies and for diagnosis of male infertility from remote locations. Home collection of semen should enhance study participation, and semen assessment in centralized laboratories will address concerns regarding interlaboratory variations and quality control.
Changes in Sperm Glycogen Synthase Kinase‐3 Serine Phosphorylation and Activity Accompany Motility Initiation and Stimulation
Authors: Somanath Payaningal R. , Jack Shannan L. , Vijayaraghavan Srinivasan
Published: December 2015

ABSTRACT: Sperm motility is regulated by protein phosphorylation. We have shown that the signaling kinase, glycogen synthase kinase‐3α (GSK‐3α), is present in spermatozoa. In somatic cells, GSK‐3 is regulated by serine and tyrosine phosphorylation. In this report, we document that both GSK‐3α and GSK‐β isoforms are present in spermatozoa, with GSK‐3α being the predominant isoform. The relationship between GSK‐3 serine phosphorylation and motility was investigated. Serine phosphorylation of GSK‐3 increases significantly in spermatozoa during their passage through the epididymis. Initiation and stimulation of motility in vitro by isobutyl‐methyl‐xanthine, 2‐chloro‐2′‐deoxy‐adenosine, and calyculin A lead to a dramatic increase in GSK‐3 serine phosphorylation. The concentration‐dependent induction of motility by calyculin A is closely associated with GSK‐3 serine phosphorylation. Immunoprecipitation of GSK‐3α and GSK‐3β shows that both of the GSK‐3 isoforms are more active in caput than in caudal spermatozoa. Calyculin A treatment decreased the activity of both isoforms. Column chromatography was used to purify inactive GSK‐3α from the caudal sperm extracts. This GSK‐3α species was phosphorylated at amino acid residues serine 21 and tyrosine 214. Inactive GSK‐3α is present in caudal but not in caput epididymal spermatozoa. The enzymes protein kinase B (PKB; also known as cAkt) and phosphoinositide 3‐kinase (PI3‐kinase), the upstream signaling proteins involved in GSK‐3 phosphorylation, are both present in spermatozoa. Fluorescence immunocytochemistry showed that GSK‐3 is present in the head and tail regions of sperm. Our work suggests a novel role for the signaling system involving GSK‐3 in the regulation of sperm motility.
Dominant Prostasome Immunogens for Sperm‐Agglutinating Autoantibodies of Infertile Men
Authors: Carlsson Lena , Ronquist Gunnar , Nilsson B. Ove , Larsson Anders
Published: December 2015

ABSTRACT: The presence of naturally occurring anti‐sperm antibodies (ASA) is a well‐known cause of infertility in men and women, but the antigens for these antibodies are poorly characterized. We have previously shown that prostasomes adhere to sperm cells and that prostasomes are major targets for ASA associated with infertility. These autoantigens have not been characterized. We used 2‐dimensional electrophoresis, immunoblotting, and mass‐spectrometry to identify the prostasome antigens for these autoantibodies. By these techniques, we revealed that prolactin‐inducible protein (PIP) and clusterin were dominant prostasome immunogens for spermagglutinating autoantibodies of 20 patients with immunological infertility. PIP was identified by 19 of 20 (95%) patient sera and clusterin by 17 of 20 (85%). In addition, 10 sporadically occurring prostasomal antigens were identified in this context, viz alcohol dehydrogenase [NADP+], annexin I, annexin III, BRCA1‐associated ring domain protein 1, heat shock 27‐kd protein, isocitrate dehydrogenase, lactoylglutathione lyase, NG,NG‐dimethylarginine dimethylaminohydrolase 1, peroxiredoxin 2, and syntenin 1.
Effects of Maternal Exposure to 3,3′,4,4′,5‐Pentachlorobiphenyl (PCB126) or 3,3′,4,4′,5,5′‐Hexachlorobiphenyl (PCB169) on Testicular Steroidogenesis and Spermatogenesis in Male Offspring Rats
Authors: Yamamoto Masako , Narita Akiko , Kagohata Mari , Shirai Mitsuyuki , Akahori Fumiaki , Arishima Kazuyoshi
Published: December 2015

ABSTRACT: On days 7–21 of gestation, Sprague‐Dawley rats were orally administered 3 or 30 μg/kg/d of 3,3′,4,4′,5‐pentachlorobiphenyl (PCB126) or 3,3′,4,4′,5,5′‐hexachlorobiphenyl (PCB169) daily. Their male offspring were autopsied at 3, 6, and 15 weeks after birth to investigate the effects of the 2 polychlorinated biphenyls (PCBs) on spermatogenesis and steroidogenesis in their testes. PCB treatment caused a decrease in the area ratio of 3β‐hydroxysteroid dehydrogenase (HSD)‐expressing cells (Leydig cells)/testis at 3 weeks after birth. When PCB126 was administered to pregnant rats, the plasma testosterone levels in their offspring were decreased at 3 weeks. The expression levels of P450scc, 3β‐HSD, and P45017α mitochondrial RNAs (mRNAs) were unchanged, although the StAR (steroidogenic acute regulatory protein) mRNA expression level was increased at 6 weeks. On the other hand, when PCB169 was administered, plasma testosterone levels were decreased at 3 and 6 weeks and were increased at 15 weeks. Plasma luteinizing hormone (LH) levels were decreased at 6 weeks, and plasma follicle‐stimulating hormone (FSH) levels were increased at 15 weeks. The expression levels of 3β‐HSD and P45017α were increased, and the mRNA level of 5α‐reductase 1 was decreased at 15 weeks. PCB169 treatment suppressed the conversion of round spermatids between stages VII and VIII. These results indicate that in utero and lactational exposure to PCB126 or PCB169 decreases plasma testosterone levels in 3‐week‐old rats, with no change in the expression levels of the mRNAs of enzymes, and that PCB169 inhibits testicular steroid synthesis more strongly than PCB126. PCB169 greatly altered the concentration of testosterone, indicating a stronger inhibitory effect on spermatogenesis. Low testosterone and LH levels in prenatally PCB169‐exposed rats until 6 weeks after birth presumably retard the functional differentiation of testicular Leydig cells; however, the increased testosterone levels at 15 weeks suggest that Leydig cells in PCB‐exposed rats are virtually mature by the 15th week.
Could Sperm Aneuploidy Rate Determination Be Used as a Predictive Test Before Intracytoplasmic Sperm Injection?
Authors: Petit François M. , Frydman Nelly , Benkhalifa Moncef , Le Du Anne , Aboura Azzedine , Fanchin Renato , Frydman Rene , Tachdjian Gerard
Published: December 2015

ABSTRACT: Chromosome abnormalities in embryos are a major cause of implantation and development failures. Some couples with normal karyotypes have repeated implantation failures after intracytoplasmic sperm injection (ICSI). In order to value patients at risk for genetic ICSI failures and the validity of sperm aneuploidy analysis, we have studied cytogenetic abnormalities in sperm from ICSI patients. Twentynine patients with normal karyotypes were included. Ten patients had at least 4 ICSI treatments without pregnancy (group A). Nine patients had a pregnancy after 1 to 3 ICSI treatments (group B). Ten fertile men with normal semen parameters were studied as controls (group C). Fluorescent in situ hybridization (FISH) was used for sperm nucleus cytogenetic analysis using chromosomes 8, 9, 13, 18, 21, X, and Y specific probes. Aneuploidy for each chromosome and diploidy rates were significantly higher in group A than in group B and in group B than in group C (P < .05). Considering each patient in groups A and B, aneuploidy rate for each chromosome was too variable to be considered as a significant test. We proposed analysis of the total sperm aneuploidy. Chromosomal sperm nuclei profile could be used as a predictive biological test before ICSI in order to improve genetic counseling for oligoasthenoteratozoospermia patients.
2D or not 2D‐Layered Functional (C, N) Materials “Beyond Silicon and Graphene”
Authors: Bojdys Michael J.
Published: December 2015

Thin layers of the semiconductor silicon are omnipresent in all electronic devices. Ongoing, exponential improvement in size, performance, and cost of a single transistor is predicted to face serious challenges in the near feature. With the era of “postsilicon electronics” on the horizon, researchers are racing to address issues of material security, less energy‐intensive manufacturing, and design of device properties. The chemistry of carbon and nitrogen (C, N) provides an attractive avenue toward the modular design of new 2D materials “beyond silicon and graphene.”

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Anatomical variants of lower limb vasculature and implications for free fibula flap: Systematic review and critical analysis
Authors: Abou‐Foul Ahmad K. , Borumandi Farzad
Published: December 2015

Sound knowledge of the anatomical patterns of lower limb vasculature is crucial for raising the free fibula flap (FFF). The tibial arteries are particularly susceptible to anatomical variations with a dominant role of peroneal artery (PR) for lower leg perfusion, and hence precluding fibular flap harvest. In this systematic review we study the prevalence of lower limbs with dominant peroneal artery (dPR). A systematic review of the MEDLINE and EMBASE databases was conducted using a comprehensive combination of keywords and search algorithm according to PRISMA guidelines. Articles describing the branching patterns of lower limb arteries, in cadaveric or angiographic studies, were included. A total of 5,790 limbs were included from 26 studies. dPR was found in 5.2% of all limbs. The combination of dPR with hypoplastic or aplastic posterior tibial artery was the most common pattern (3.3%) followed by hypoplastic anterior tibial artery (1.5%). Peronea arteria magna (PAM) was seen in 0.4% of limbs. Bilateral variants were seen in 20% of all cases with a dPR. A greater awareness of dPR as anatomical variation and its prevalence among healthy subjects can prevent ischemic donor‐site complications after FFF harvest. Clinical examination alone may not suffice to detect anatomical variations, hence preoperative imaging of lower limb vascular system is recommended. © 2015 Wiley Periodicals, Inc. Microsurgery 36:165–172, 2016.
Enhanced thermal conductivity of PE/BN composites through controlling crystallization behavior of PE matrix
Authors: Zhao Tianbao , Zhang Xianlong
Published: December 2015

In this article, in order to enhance the thermal conductivity of the polyethylene (PE)/boron nitride (BN) composites through controlling the crystallization behavior of the PE matrix, the crystallization and melting behavior of the PE in the PE/BN composites was investigated. When the BN content was more than 10 wt%, an extra weak exothermic peak (Th) at 130°C was observed. Moreover, after the annealing of the PE/BN composites at 130°C, the extra weak melting peaks (Tmh) of the PE in the PE/BN composites were also observed and shifted to the high temperature with increasing annealing time, which proved that the Th was induced by PE crystallization. Meanwhile, the results of temperature‐dependent absorbance IR spectra of the PE/BN composites showed that the crystallization peak (729 cm−1) remarkably appeared at 130.2°C, indicating that the crystallization of the PE in the PE/BN composites can occur at 130.2°C. When the annealing time and temperature were 20 min and 130°C, the thermal conductivity of the PE/BN composite was 16% higher than that of the unannealed PE/BN composites. In addition, the results of the wide angle X‐ray diffraction (WAXD) showed that the BN particles had no influence on the PE crystalline form in the PE/BN composites. POLYM. COMPOS., 38:2806–2813, 2017. © 2015 Society of Plastics Engineers
Error dynamics in extended‐range forecasts
Authors: Nicolis C.
Published: December 2015

General expressions for the dynamical behaviour of prediction errors of time‐averaged observables involved in extended‐range forecasts are derived and applied to systems giving rise to chaotic dynamics, systems close to criticalities and stochastically forced systems. The role of the initial conditions, of the parameters present and of the averaging period, in the properties of the error, and in particular in the predictability gain with respect to the error associated with the instantaneous observables, are revealed, providing insights on optimal choices of this period depending on the intrinsic properties of the system of interest.
Note on the biochemical preparation of inosose
Authors: Kluyver A. J. , Boezaardt A. G. J.
Published: December 2015

It is shown that inosose, a cyclose prepared fairly recently by Posternak by oxidation of meso‐inositol with concentrated nitric acid, can be obtained in a much higher yield by the biochemical oxidation of the said cyclic polyalcohol with the aid of the acetic acid bacterium Acetobacter suboxydans.
Das Ramanspektrum des flüssigen Dimethyl‐disulfids
Authors: Gerding H. , Westrik R.
Published: December 2015

Das Ramanspektrum des flüssigen Dimethyldisulfids wurde erneut aufgenommen und der Polarisationszustand von den wichtigsten Ramanlinien bestimmt. Das Molekül besitzt die Struktur CH3 ‐ S ‐ S ‐ CH3. Es befindet sich bei Zimmertemperatur hauptsächlich in der cis‐Form

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Die Rotation um die S‐S Bindung ist bedeutend gehemmt.
The chemistry of small‐ring compounds. Part 50. Anomalous ozonolysis of methylenecyclobutanes
Authors: van Velzen J. C. , van Tunen W. C. J. , de Boer Th. J.
Published: December 2015

Methylenecyclobutane and higher alkylidenecyclobutanes containing no substituents in the ring undergo Criegee ozonolysis with formation of cyclobutanone as a major product. When the ring contains α‐substituents, a primary ozonide is formed in which the (substituted) cyclobutyl ring is ruptured simultaneously with O‐O fission. The overall result is a rearrangement to a seven‐membered cyclic 3,4‐dioxa ketone (cf. Scheme 3). This rearrangement is mechanistically related to previously studied cyclopropyl systems in which the small ring is ruptured without the assistance of a ring substituent.

At room temperature, the 3,4‐dioxacycloheptanone derivative slowly loses formaldehyde, leaving a γ;‐butyrolactone. Structurally related 3,4‐dioxacyclohexanones are relatively stable, i.e. they do not decompose into corresponding β‐lactones.
Crystal structure determination of 1‐[N‐benzyloxycarbonyl‐1‐amino‐(S‐4‐methoxybenzyl)‐2‐thio‐ethyl] ‐2‐hydroxy‐3‐ethoxycarbonyl‐1,2,3,4 ‐tetrahydro‐β‐carboline, C32H35N3O6S
Authors: Behm Helmut , Beurskens Paul T. , Plate Ralf , Ottenheijm Harry C. J.
Published: December 2015

The structure of the title compound, C32H35N3O6S, was determined by X‐ray analysis. Mr=589.71; triclinic; Space group: P‐1; Z=2; a=11.1173(7), b=12.0068(13), c=13.1270(8)Å, α=104.612(8), β=103.833(5), γ=109.706(6)°, γ;=1491.80Å3, DC =1.31 Mg/m3. CuKα radiation, graphite crystal monochromator, λ=1.5418, μ(CuKα) = 13.27 cm−1, F(000)=620.00, T=290°K. Final conventional R‐factor=0.0428, RW=0.0463 for 3099 unique reflections and 519 variables. The structure was solved using MULTAN and DIRDIF.