Human Corpus Research Articles

POS-01.71: Effect of aldosterone on isolated human penile corpus cavernous tissue

POS-01.71: Effect of aldosterone on isolated human penile corpus cavernous tissue

Pharmacological characterization of the novel phosphodiesterase type 5 (PDE5) inhibitor lodenafil carbonate on human and rabbit corpus cavernosum

Methods In functional studies, cavernosal strips were mounted in organ baths for isometric force recording, coupled to a PowerLab 8/SPTM data acquisition system. Rats were surgically manipulated and telemetry transmitters were implanted in the abdominal aorta for measurements of systolic, diastolic and mean arterial blood pressure. For determination of PDE activity by LC-MS/MS analysis, lodenafilc arbonate (0.005–1 μM) was preincubated in the enzymatic mixture for 5 min at room temperature. Reaction was initiated by the addition of the substrate cGMP (5 μM) at 35°C for 30 min. Pharmacokinetics was studied by tandem mass spectrometry (LC-MS/MS) with positive ion electrospray ionization using multiple reactions monitoring (NMR) method. Blood samples were collected at 0.02 to 24 h after drug administration by intravenous (1 mg/kg) and oral (10 mg) routes. The hydrolysis of lodenafil carbonate (final concentration 10 μM) was also studied in human, dog and rat plasma as well as in acid solution (100 mM hydrochloric acid).

Effects of BAY 41–2272 on smooth muscle tone, soluble guanylyl cyclase activity and NADPH oxidase activity/expression in corpus cavernosum from wild-type, neuronal and endothelial NOS null mice

Background Nitric oxide (NO) is considered to play a critical role in the control of erectile function, by activating soluble guanylyl cyclase (sGC) in cavernosal smooth muscle to generate cGMP, which, in turn, promotes relaxation leading to penile erection. The pyrazolopyridine 5-cyclopropyl-2[1-(2-fluorobenzyl)-1H-pyrazolo[3,4-b]pyridin-3-yl]pyrimidin-4-ylamine (BAY 41–2272) sensitizes sGC to NO, lowers mean arterial pressure in spontaneously hypertensive rats and increases survival in a low-NO rat model of hypertension. In vitro, BAY 41–2272 causes potent relaxation of vascular smooth muscle, rat anococcygeus muscle as well as human and rabbit corpus cavernosum (CC).

Ghrelin Affects the Release of Luteolytic and Luteotropic Factors in Human Luteal Cells

Ghrelin, well-known modulator of food intake and energy balance, is a rather ubiquitous peptide involved in several endocrine and nonendocrine actions. A possible as-yet-unknown role for ghrelin in modulating luteal function has been suggested because both ghrelin and its receptor (GRLN-R) have been immunohistochemically detected in human corpus luteum. We first investigated GRLN-R mRNA expression in midluteal phase human luteal cells. Ghrelin effect on basal and human chorionic gonadotropin (hCG)-stimulated progesterone (P) release was then analyzed. Finally, we investigated whether ghrelin could affect luteal release of vascular endothelial growth factor (VEGF), prostaglandin (PG) E(2), both luteotropic factors, and PGF(2alpha), luteolytic modulator. Ghrelin effect on both basal and hypoxia-stimulated VEGF luteal expression was analyzed. Human luteal cells were incubated for 24 h with ghrelin (10(-13) to 10(-7) m) or hCG (100 ng/ml) or CoCl(2) (10 microm), chemical hypoxia, or with hCG or CoCl(2) in combination with ghrelin. Both GRLN-R mRNA and VEGF mRNA were evaluated by real-time RT-PCR. PGs and P release was assayed by RIA, whereas VEGF release by ELISA. GRLN-R mRNA expression was demonstrated in human luteal cells. Both basal and hCG-stimulated P release was significantly decreased by ghrelin, which was able to reduce PGE(2) and increase PGF(2alpha) luteal release. Both basal and hypoxia-stimulated VEGF release was significantly decreased by ghrelin, which did not affect VEGF mRNA luteal expression. The present in vitro study provides the first evidence of a direct inhibitory influence of ghrelin on human luteal function.

The Imperial – Emancipatory Paradox of International Human Rights: How Useful is the Right to Health in Sub-Saharan Africa?

AbstractIn thinking of and formulating strategies for tackling the problem of access to good health care services in Sub-Saharan Africa, it is very tempting to look to the right to health as part of such strategies. However, given the genealogy and practices of the international human rights corpus, the question as to the value of utilising the right to health in such endeavours necessarily comes to the fore and demands investigation. Using the trajectory of the right to health, the paper analyses the usefulness or otherwise of international human rights in seeking solutions to problems relating to health in Sub-Saharan Africa. An argument is made that the right to health has low utility value despite the promises it makes. The paper concludes with the argument that an appreciation of the imperial-emancipatory paradox that is inherent in the corpus of international human rights not only enables one to expect less from the right to health but also opens up possibilities of crafting more productive strategies in the struggle to achieve the 'highest attainable standard of health' for the people of Sub-Saharan Africa.

Presence of sensory nerve corpuscles in the human corpus and cervix uteri during pregnancy and labor as revealed by immunohistochemistry

BackgroundThe uterus is exposed to changes such as enlargement and distension during pregnancy and labor. In these processes and in the process of cervical ripening, proprioceptive information is likely to be of great importance. Therefore, we wanted to study the possible existence of sensory nerve corpuscles in uterine corpus and cervix during pregnancy and labor. Studies on this aspect have not previously been perfomed.MethodsBiopsies were taken from the upper edge of the hysterotomy during caesarean section at term (n = 8), in labor (n = 5) and from the corresponding area in the non-pregnant uterus after hysterectomy (n = 7). Cervical biopsies were obtained transvaginally from the anterior cervical lip. Serial cryostat sections were prepared for immunohistochemistry using polyclonal antibodies against nerve growth factor receptor p75, protein gene product 9.5 and S-100.ResultsStructures with the characteristics of sensory nerve corpuscles were observed in several specimens after staining for p75, PGP 9.5 and S-100. They were observed in specimens of the non-pregnant corpus and cervix and also in specimens of the pregnant cervix before onset of labor. However, they were absent in all specimens during labor.ConclusionSensory corpuscles have here for the first time been detected in the human corpus and cervix uteri. Studies on the importance of the corpuscles in relation to the protective reflex actions that occur in the uterus during pregnancy should be performed in the future.

Unaltered innervation of the human cervix uteri in contrast to the corpus during pregnancy and labor as revealed by PGP 9.5 immunohistochemistry

Objective To examine the occurrence of the general neuronal marker protein gene product 9.5 (PGP 9.5) in the human corpus (isthmus region) and the cervix uteri during pregnancy and parturition Study design Biopsies were taken from the upper edge of the hysterotomy during caesarean section (CS) at term ( n = 5), in labor ( n = 5) and from the corresponding area in the non-pregnant uterus after hysterectomy ( n = 5). Cervical biopsies were obtained transvaginally from the anterior cervical lip. Serial cryostate sections were prepared for immunohistochemistry using polyclonal antibodies to PGP 9.5. Results Nerve fibers displaying PGP 9.5 immunoreactivity were observed in all sections from the three groups examined. They were identified in muscle tissue, in the stroma, and around blood vessel walls. A 30-fold decline of immunoreactive nerve fibers was observed in the isthmus part of the corpus uteri at term compared to the non-pregnant. There were no significant differences between the immunoreactivity in the cervix uteri of the three groups Conclusions The innervation of the cervix uteri is dense and unaltered throughout pregnancy and labor. In contrast, the corpus is almost denervated. Further studies are needed to clarify the reason and the impact of these findings.

Spatiotemporal Expression of Heparanase During Human and Rodent Ovarian Folliculogenesis1

Heparanase (HPSE) is an endoglycosidase that cleaves heparan sulfate proteoglycans (HSPGs), major components of the basement membrane (BM) and extracellular matrix (ECM). Heparanase activity results in release of HSPG-bound molecules, including basic fibroblast growth factor (FGF2). Structural and functional development of the corpus luteum (CL) involves tissue remodeling, active angiogenesis, and steroid production. Heparanase-induced ECM and BM breakdown as well as FGF2-stimulated endothelial proliferation may have an important role in the regulation of luteal function. Heparanase mRNA was detected by reverse-transcription-polymerase chain reaction in granulosa cells recovered from follicular fluid of in vitro fertilization patients. Using sulfate-labeled ECM, heparanase enzymatic activity was determined in human luteinized granulosa cells. Employing immunohistochemistry, heparanase protein was localized predominantly in the theca interna cell layer of the mature antral follicle, whereas in human corpora lutea, both luteinized granulosa and theca cells were immunostained for heparanase. During luteolysis, heparanase was identified in macrophages surrounding the forming corpus albicans. In serially sectioned ovaries from unstimulated rats as well as from eCG-treated rats, expression of heparanase was noted exclusively in the ovarian steroid-producing interstitial tissue. Following an ovulatory dose of hCG, heparanase was immunostained also in lutein cells of the forming corpora lutea. Temporal expression of heparanase in granulosa cells during the luteal phase and in macrophages during luteal regression supports the hypothesis that heparanase plays a role in human ovarian ECM remodeling and may potentiate cellular migration and growth factor bioavailability.

234 Immunohistochemical presence of cyclic AMP and cyclic GMP-dependent protein kinase A and G in human cavernous arteries and corpus cavernosum

234 Immunohistochemical presence of cyclic AMP and cyclic GMP-dependent protein kinase A and G in human cavernous arteries and corpus cavernosum

Distribution of DMT 1 within the human glandular system.

DMT1 is a transmembrane protein which transports the divalent metal ions Fe2+, Zn2+, Cu2+ and Mn2+. Although DMT1 has been functionally linked to duodenal absorption and cellular utilisation of iron hardly anything is known about its distribution and potential role within the human glandular system. Two polyclonal antibodies were raised to study the expression of DMT1 in tissues obtained from human corpus by the means of immunocytochemistry and Western blotting. All antibodies specifically detected a 60 kD protein band referring to human DMT1. Significant amounts of DMT1 expression were detected on the luminal site of organs, which are involved in excretion/re-absorption processes, such as salivary glands, pancreas, biliary tract and gallbladder. Our results suggest that DMT1 may be of pivotal importance for the regulation of metal ion homeostasis within organs involved in absorption and excretion of ions.

Glutathione-related enzymes in cell cultures from different regions of human epididymis.

Protection of maturing sperm from potential endogenous or exogenous harmful substances during their transit throughout the epididymis is a critical event. The authors studied the activity of gamma-glutamyl transpeptidase (GGT) and glutathione S-transferase (GST), and glutathione (GSH) levels in epithelial cell cultures from human caput, corpus, and cauda epididymides. Tissue was obtained from patients undergoing therapeutic orchidectomy for prostatic cancer. Enzymatic activity was measured in conditioned media and cellular fractions. Androgen influence was also evaluated. Both enzymatic activities were found in cellular homogenates and conditioned media from cultures of all epididymal regions. GGT activity was highest in cultures from cauda epididymis, both in conditioned media and cell fractions, while GST activity did not show regional differences in conditioned media, but exhibited higher activity in cell homogenates from cauda cultures than those obtained from corpus and caput epididymis. GSH level showed no regional difference in cell homogenates and it could not be detected in conditioned media by the method used. Presence of different concentrations of dihydrotestosterone (DHT) had no influence neither on the enzymatic activities nor GSH concentration. The results indicate that GGT and GST are present along the human epididymis and a fraction or isoform of these enzymes might be secreted to the luminal fluid to play a detoxificative role in sperm maturation.

A human vocal utterance corpus for perceptual and acoustic analysis of speech, singing, and intermediate vocalizations

In this paper we present the collection and annotation process of a corpus of human utterance vocalizations used for speech and song research. The corpus was collected to fill a void in current research tools, since no corpus currently exists which is useful for the classification of intermediate utterances between speech and monophonic singing. Much work has been done in the domain of speech versus music discrimination, and several corpora exist which can be used for this research. A specific example is the work done by Eric Scheirer and Malcom Slaney [IEEE ICASSP, 1997, pp. 1331–1334]. The collection of the corpus is described including questionnaire design and intended and actual response characteristics, as well as the collection and annotation of pre-existing samples. The annotation of the corpus consisted of a survey tool for a subset of the corpus samples, including ratings of the clips based on a speech–song continuum, and questions on the perceptual qualities of speech and song, both generally and corresponding to particular clips in the corpus.

Sex‐related differences in the development of the human fetal corpus callosum: in utero ultrasonographic study

Sex‐related differences in the development of the human fetal corpus callosum: in utero ultrasonographic study

Activity of collagen-bound matrix metalloproteinases-2 and -9 in tumours of human uterine corpus

Activity of collagen-bound matrix metalloproteinases-2 and -9 in tumours of human uterine corpus

Regulated expression and potential roles of p53 and Wilms' tumor suppressor gene (WT1) during follicular development in the human ovary.

It has been previously demonstrated that the gonadotropin-mediated inhibition ofapoptosis in rat ovarian granulosa cells is associated with changes in the expression of several cell death-regulatory genes, including p53. In addition, it has been shown that the actions of p53 may be amplified through a cooperative interaction with the Wilms' tumor suppressor gene product (WT1). Based on these findings, the present studies were conducted to determine whether p53 and WT1 are expressed and gonadotropin regulated in the human ovary and to study the relationship between tumor suppressor gene expression and apoptosis in human granulosa/lutein cells (GCs). Analysis of total RNA prepared from human GCs using the RT-PCR demonstrated the presence of p53 messenger RNA (mRNA) and four WT1 mRNA splice variants. These observations were supported by Northern blot analysis of total RNA prepared from human GCs, which revealed the presence of a single (approximately 2.8 kb) p53 mRNA transcript and two primary (approximately 1.8 and approximately 3.5 kb) WT1 mRNA transcripts. Western blot analysis of nuclear protein extracts from human GCs yielded one immunoreactive protein of the expected size (approximately 53 kDa) recognized by a p53 antibody and one immunoreactive protein of the expected size (approximately 52-54 kDa) recognized by the WT1 antibody. Immunohistochemical staining showed that both molecules were localized to nuclei of human GCs and were coordinately regulated during follicular development. Immunofluorescence analysis showed that p53 protein was localized exclusively to nuclei of GCs undergoing apoptosis during in vitro culture and was similarly localized to nuclei and cytoplasm of apoptotic granulosa cells in atretic follicles in vivo. To further evaluate whether human GC apoptosis is linked to increased expression of tumor suppressor genes, we analyzed levels of p53 and WT1 mRNA and protein in GCs induced to undergo apoptosis in vitro. Healthy (nonapoptotic) GCs snap-frozen immediately after isolation from patients undergoing in vitro fertilization-embryo transfer possessed relatively low, but detectable, levels of p53 and WT1 mRNA and protein. However, following serum-free culture to induce apoptosis, p53 mRNA and protein levels increased significantly after 24 h, paralleling the increase in the number of apoptotic GCs. The induction of both p53 mRNA and protein in GCs was inhibited by the addition of human CG to the culture medium. In contrast, WT1 mRNA and protein levels remained constitutive in GCs incubated for 24 h compared with GCs snap-frozen immediately after isolation. We conclude that the p53 and WT1 genes are expressed at the mRNA and protein levels in human GCs and that expression of p53 is regulated during follicular maturation. Nuclear accumulation of p53 protein occurs in human GCs during apoptosis in vitro and in vivo, and p53 mRNA and protein are up-regulated in GCs starved of hormonal support but down-regulated by the presence of human CG. We propose that the products of these two principal tumor suppressor genes serve as important regulators of human follicular development and corpus luteum function.

Mutations of RegIα are associated with enterochromaffin-like cell tumor development in patients with hypergastrinemia

Background & Aims: The RegIα gene ( Reg) encodes a secretory protein proposed to regulate islet β-cell and gastric mucous cell growth. Reg is expressed in rat gastric enterochromaffin-like (ECL) cells. The aim of this study was to examine Reg expression in human corpus and to determine the identity of Reg in ECL cell carcinoid tumors in hypergastrinemic patients. Methods: Reg messenger RNA (mRNA) abundance was quantified by Northern blot in extracts of gastric corpus from patients with and without ECL cell tumors and in AR4-2J cells stimulated by gastrin; cellular origins were determined by immunocytochemistry. Mutations of Reg were determined by reverse-transcription polymerase chain reaction, cloning, and sequencing, and the mutated protein was expressed in HIT-T15 cells. Results: Reg mRNA abundance was increased approximately threefold in the corpus of hypergastrinemic patients compared with controls, and was enriched in 3 of 7 ECL cell carcinoid tumors but not in non–endocrine cell gastric polyps. In AR4-2J cells, gastrin stimulated Reg mRNA abundance; this was eliminated by the gastrin/cholecystokinin B antagonist L-740,093 (10 −9 mol/L). Immunocytochemistry indicated that Reg was located in both chief cells and ECL cells in human corpus. Mutations of Reg were identified in 3 of 5 patients with ECL cell carcinoid tumors; in 2 cases, mutation of the initiator methionine residue led to exclusion of the protein from the secretory pathway. Conclusions: Gastrin regulates Reg mRNA abundance in human corpus. Mutations of Reg that prevent secretion are associated with ECL cell carcinoids, suggesting a function as an autocrine or paracrine tumor suppressor. GASTROENTEROLOGY 1999;116:1310-1318

SILDENAFIL INHIBITS PHOSPHODIESTERASE TYPE 5 IN HUMAN CLITORAL CORPUS CAVERNOSUM SMOOTH MUSCLE

Abstract Phosphodiesterases play an important physiological role by regulating the intracellular levels of cyclic nucleotides. In this study, we investigated the kinetic parameters of inhibition of phosphodiesterase (PDE) type 5 (EC 3.1.4.35, 3′,5′-cyclic GMP phosphodiesterase) by a novel, high-affinity, selective PDE type 5 inhibitor, sildenafil, in intact cells and in soluble extracts of human clitoral corpus cavernosum smooth muscle cells. Sildenafil inhibited cGMP hydrolysis in the crude extract (Ki= 7.2 ± 2.7) and in partially purified preparations (Ki= 9 nM) in a competitive manner, as determined by Dixon plots. Sildenafil was a more effective PDE type 5 inhibitor than zaprinast (Ki= 400.0 ± 76.4 nM, crude extracts; 250 nM, partially purified). Stimulation of intracellular cGMP synthesis by the nitric oxide donor sodium nitroprusside resulted in a 3.3- and 2.9-fold increase in cGMP concentration in the presence of sildenafil or zaprinast, respectively, compared to sodium nitroprusside treatment alone in intact cells at physiological temperatures. These observations suggest that human clitoral corpus cavernosum smooth muscle tone may be regulated by the synthesis and release of nitric oxide and that this pathway is dependent on PDE type 5 activity.

Levels of sex hormone-binding globulin and corticosteroid-binding globulin mRNAs in corpus luteum of human subjects: Correlation with serum steroid hormone levels

To understand regulation of the function of human ovarian corpus luteum by sex steroid-binding proteins, the levels of luteal intracellular sex hormone-binding globulin (SHBG) and corticosteroid-binding globulin (CBG) mRNAs and serum steroid hormones were simultaneously determined. The expression of SHBG and CBG mRNAs was detected in all samples analyzed. SHBG mRNA level was positively correlated with serum estradiol-17β level (p < 0.05), but not with serum progesterone level. There was a positive correlation between SHBG mRNA level and serum estradiol-17β/progesterone ratio (p < 0.01). On the other hand, CBG mRNA level was positively correlated with serum estradiol-17β and progesterone level (p < 0.01 and p < 0.01, respectively). There was no correlation between CBG mRNA level and serum estradiol-1 17β/progesterone ratio. SHBG and CBG mRNA levels were not correlated with the levels of serum testosterone, free testosterone or Cortisol. These findings suggest that the synthesis of luteal SHBG and CBG is complexly regulated by estrogen and progesterone, and that SHBG and CBG interact with estrogen and progesterone, respectively, for luteal steroidal activity.

Cytoplasmic extrusion and the switch from creatine kinase B to M isoform are completed by the commencement of epididymal transport in human and stallion spermatozoa.

Although in several species there is a relationship between epididymal sperm transport and fertility, in human in vitro fertilization (IVF), spermatozoa recovered from the caput epididymidis or even the rete testis are fertile. We studied two objective markers of sperm maturity in the sperm of men and stallions: creatine kinase (CK) concentrations, which are a measure of cytoplasmic retention in immature spermatozoa, and the ratio of CK-M and CK-B isoforms (% CK-M/[CK-M + CK-B]), which is proportional to the incidence of mature sperm. The CK markers and the fertilizing function are closely related: Immature sperm with cytoplasmic retention do not bind to the zona, because during cytoplasmic extrusion, the sperm plasma membrane is also remodeled. We examined whether changes in sperm CK values are still ongoing during epididymal transport, or if cellular maturation is completed prior to the arrival of sperm in the caput epididymidis. The incidences of mature sperm in human caput and corpus epididymidis (studied in six men with obstructive azoospermia of various pathogeneses) were (mean+/-SEM) 55.7+/-2.2 and 49.3+/-7.6%, respectively; and the sperm CK-M ratios in the caput epididymidis of three men were 72, 75, and 70%, values that are similar to those of ejaculated sperm. In four segments of the proximal and distal epididymis of three stallions (the origin of sperm was also verified by the position of the cytoplasmic droplet) and in ejaculate of five stallions, the incidences of mature sperm were 88.2+/-6.2, 89.0+/-6.7, 90.3+/-7.8, 87.6+/-5.9, and 86.7+/-0.8%, and the respective CK-M ratios were 75.0+/-8.7, 84.2+/-2.9, 87.9+/-1.2, 92.5+/-1.5, and 69.3+/-3.5%. There were no differences in the incidences of mature and immature spermatozoa or in CK-M ratios among sperm arising from the various epididymal regions or from the ejaculate in men or stallions. Thus, the cellular maturation events in sperm, as detected by the CK markers, are completed by the time the sperm commences epididymal transport. These findings are in agreement with the IVF fertility of sperm aspirated from the male reproductive tract. The data may also suggest that the primary role of sperm epididymal transport in men is to remodel the plasma membrane to enhance sperm functional integrity in the diverse environments of the male and female reproductive tracts prior to fertilization.

P-246. Co-culture of human trophoblastic and corpus luteum cells in vitro : a pilot study

P-246. Co-culture of human trophoblastic and corpus luteum cells in vitro : a pilot study