Regulation of monocyte chemotactic protein-1 expression in human endometrial endothelial cells by sex steroids: a potential mechanism for leukocyte recruitment in endometriosis.

The main aim of this study is to describe the in vivo temporal and spatial expression of monocyte chemotactic protein 1 (MCP-1) in human endometrial endothelial cells (HEECs) and to compare the in vitro regulation of MCP-1 expression by sex steroids in HEECs from women with or without endometriosis. Eutopic endometrial tissues and endometriosis implants were grouped according to the menstrual cycle phase and were examined by immunohistochemistry for MCP-1 expression. No significant difference was observed for MCP-1 immunoreactivity in the endothelial cells of eutopic endometrium of women with endometriosis when compared to endometrium of women without endometriosis and to endometriosis implants. For in vitro studies, the purity of cultured HEECs (90%-95%) was confirmed by immunocytochemistry using endothelium-specific markers CD31 and CD146. The effects of estradiol (5 x 10(- 8) mol/L), progesterone (10(-7) mol/L), or both on MCP-1 messenger RNA (mRNA) and protein levels were analyzed by reverse transcriptase-polymerase chain reaction (RT-PCR) analysis and enzyme-linked immunosorbent serologic assay (ELISA), respectively. Sex steroids did not have significant effect on MCP-1 mRNA and protein expression in HEECs from women without endometriosis. However, we observed that the sex steroid treatment stimulated MCP-1 mRNA and protein expression in HEECs from women with endometriosis (P < .05). We postulate that the stimulation of chemokine expression by sex steroids in the endometrial endothelial cells in women with endometriosis may play a central role in recruiting mononuclear cells, therefore contributing to the inflammatory aspect of endometriosis.

Differential regulation of Akt phosphorylation in endometriosis.

Protein kinase B (PKB/Akt), a serine/threonine kinase, regulates the function of many cellular proteins involved in apoptosis and proliferation. It was postulated that there is a higher Akt activity in endometriosis compared with normal endometrium, and that oestrogen may be one of the factors responsible for the high Akt activation in endometriotic cells. Phospho-Akt (pAkt) concentrations in normal, eutopic and ectopic endometrial tissues were compared by immunohistochemistry, and a higher pAkt immunoreactivity was revealed in eutopic and ectopic endometrium compared with normal endometrium, in vivo. Higher Akt phosphorylation in stromal cells from eutopic endometrium was observed, when compared with normal, in vitro (P < 0.05). Akt phosphorylation was rapidly (2-10 min) stimulated when endometrial stromal cells from normal and endometriosis patients were treated with 17 beta-oestradiol. In endometrial stromal cells from the endometriosis group, ICI 182,780 (ICI, a specific oestrogen receptor antagonist) failed to antagonize the effect of oestradiol when combined with oestradiol, and revealed a stimulatory effect on Akt phosphorylation when given alone (P < 0.05). In conclusion, since Akt affects cell survival, it is suggested that increased Akt phosphorylation may be related to the altered apoptosis/proliferation harmony in endometriosis, and therefore Akt may play a critical role in the pathogenesis of endometriosis.

CCM2 and CCM3 proteins contribute to vasculogenesis and angiogenesis in human placenta.

Placenta as an ideal model to study angiogenic mechanisms have been established in previous studies. There are two processes, vasculogenesis and angiogenesis, involved in blood vessel formation during placental development. Therefore, blood vessel formation is a crucial issue that might cause vascular malformations. One of the vascular malformations is cerebral cavernous malformation (CCM) in the central nervous system, consisting of endothelium-lined vascular channels without intervening normal brain parenchyma. Three CCM loci have been mapped as Ccm1, Ccm2, Ccm3 genes in CCM. In order to investigate whether CCM proteins participate in blood vessel formation, we report here the expression patterns of CCM2 and CCM3 in developing and term human placenta by means of immunohistochemistry and Western blot analysis. CCM2 and CCM3 were obviously detected in the vascular endothelium during early pregnancy. Moreover, vascular endothelium of stem villi revealed a moderate immunostaining for CCM2 and, to a lesser extent, in the endothelium of mature intermediate villi in term placenta. Interestingly, CCM3 immuno-staining was weakly localized in the endothelium of mature intermediate villi and showed lesser expression going toward stem villi in term placenta. The expression patterns of the proteins were clearly identified in the vascular endothelium of human placenta, suggesting that they might play roles during angiogenesis and vasculogenesis. Furthermore, with this study, CCM2 and CCM3 have been described for the first time in the human placenta.

Tenascin is highly expressed in endometriosis and its expression is upregulated by estrogen.

OBJECTIVE: To investigate the localization of tenascin expression in the endometrium of women without endometriosis and in endometriotic implants, and to determine the in vitro regulation of tenascin by E(2) in these tissues. DESIGN: Experimental laboratory study. SETTING: University medical center. PATIENT(S): Reproductive age women with or without endometriosis. INTERVENTION(S): Proliferative (n = 14), and secretory (n = 14) endometrium from women without endometriosis and endometriosis implants (n = 14) were used for immunohistochemical analysis. Endometrial and endometriotic stromal cells were grown in culture and treated with E(2), the estrogen receptor antagonist ICI 182 780 (ICI) alone, E(2) in combination with ICI, or vehicle (control) for 24 hours, and tenascin expression was analyzed by Western blotting. MAIN OUTCOME MEASURE(S): Expression levels of tenascin in normal endometrium and endometriotic implants and its regulation by E(2). RESULT(S): Tenascin immunostaining revealed an increasing intensity in the stromal cells, starting from normal secretory endometrium, then normal proliferative endometrium, and reaching the highest expression in endometriotic implants. Estradiol induced a significant increase in tenascin protein levels in the endometriotic stromal cells in culture. CONCLUSION(S): The modulation of tenascin as an extracellular matrix protein by E(2) in endometriotic stromal cells may be one of the factors playing a role in the development of endometriosis.

Matrix metalloproteinases-2, -3 and -9 in human term placenta.

Matrix metalloproteinases (MMPs) are zinc-dependent enzymes that degrade the components of the extracellular matrix (ECM) and are known to be the main mediators of human placentation and parturition. Although there are many studies on the roles and distribution of MMPs in human term placenta, so far none of the studies has investigated the distribution of MMP-2, -3 and -9 in different cells of various placental sites. In this study, we aimed to determine the distribution and enzymatic activities of MMP-2, -3 and -9 with regard to different regions of term human placenta, such as amnion, basal plate, chorionic plate, decidua, chorion laeve, Nitabuch's stria, umbilical cord and placental villi. Eighteen normal human term placentas were obtained after vaginal deliveries. Immunohistochemistry and zymography were performed for MMP-2, -3 and -9 on placental tissue sections and protein extracts, respectively. Nearly all tissues showed immunoreactivity for MMPs. The strongest enzymatic activity for MMP-2 was seen in areas where invasive trophoblast cells invaded maternal tissues. MMP-9 had the highest enzymatic activity at the contact region of fetal and maternal parts, suggesting the importance of MMP-9 in separation of the placenta from the uterine wall during labor. MMP-3 had a similar localization to MMP-9, suggesting that besides gelatinases like MMP-2 and -9, MMP-3 (stromelysin-1) may also have important roles during labor. This study describes the site-specific distribution and activities of MMPs and therefore might help in elucidating the molecular mechanisms in pathologies such as premature rupture of membranes.

Vasculogenesis and angiogenesis in the early human placenta.

Vasculogenesis and angiogenesis are two consecutive processes during blood vessel development in the human placenta. While vasculogenesis, which is the formation of first blood vessels, is achieved by differentiation of pluripotent mesenchymal cells into haemangiogenic stem cells. The subsequent step, angiogenesis, is characterized by development of new vessels from already existing vessels. In this review, we aim to give an overview of vasculogenesis and angiogenesis during the first trimester of human placental development. Recent studies have shown that at the very early stages of placental development, cytotrophoblasts trigger vasculogenesis and angiogenesis, whereas as pregnancy progresses Hofbauer and stromal cells take over the task of triggering blood vessel development. Important growth factors in this scenario are the vascular endothelial growth factor (VEGF) family and their receptors, as well as Tie-1 and Tie-2. This review depicts the molecular and morphological steps of vasculogenesis and angiogenesis, which can give further insights into human placental development and maturation disorders.

Cellular diversity of human placental stem villi: an ultrastructural and immunohistochemical study.

The aim of the study was to investigate the distribution and differentiation of cell types in the stroma of human placental stem villi (SV). A total of 14 human term placental tissues were studied. Double immunolabeling was performed for desmin-vimentin, desmin-alpha-smooth actin and vimentin-alpha-smooth actin. Cytokeratin 7, proliferating cell nuclear antigen immunolabeling was also performed. Parallel tissue samples were examined by transmission electron microscopy. HSCORE was performed for the semi-quantitative analysis of distribution of cells in the stroma of SV. Vimentin-labeled cells were mostly distributed in the subtrophoblastic area. Desmin-vimentin double immunolabeling was mainly localized in the triangular area and to a lesser degree in the perivascular area and vessel walls (p=or<0.001). However, desmin-alpha smooth actin labeling was observed predominantly in the vessel wall and perivascular area. Vimentin-alpha smooth actin immunoreactivity was significantly stronger in the triangular and perivascular areas compared to the vessel walls (p=0.003). Ultrastructurally, cells in the stroma of SV were mesenchyme cells, reticulum cells, fibroblasts, myofibroblasts, smooth muscle cells, and Hofbauer cells, filamented and vacuolated cells. The differentiation of myofibroblasts in the triangular and perivascular areas may play a role in maturation of SV and villous contractility, modulation of the intervillous space and this may have effects on maternofetal placental circulation.

Immunolocalization of glial cell-derived neurotrophic factor (GDNF) and its receptor GFR-alpha1 in varicocele-induced rat testis.

The aim of the study was to determine the immunolocalisation of glial cell-derived neurotrophic factor (GDNF) and its receptor (GFRalpha1) in testicular dysfunction induced by experimental left varicocele. Male Wistar rats were divided randomly into two groups: a varicocele-induced group and a sham-operated group for 9, 11 and 13 weeks (each group n=6). After orchiectomy, part of the left testis from each animal was fixed, processed and embedded in paraffin wax for immunohistochemistry and the other part was fixed for ultrastructural investigations. GDNF immunoreactivity was localized in the interstitial space in Leydig cell cytoplasm and there was no significant difference (P=0.5) between the varicocele-induced groups at the various time points. GFRalpha1 localization was perinuclear in spermatids and cytoplasmic in Leydig cells. The decrease of GFRalpha1 immunoreactivity was significant (P=0.001) in varicocele-induced testis at 13 weeks when compared with the age-matched sham group. This is the first study to describe the immunolocalization patterns of GDNF and GFRalpha1 in a rat model of varicocele. Although there was no change in GDNF labelling at the different time points after varicocele, GFRalpha1 was significantly decreased in the 13-week group. Distribution of GDNF and its receptor GFRalpha1 in normal and varicocele-induced rat testes suggests both autocrine and paracrine regulation of spermatogenesis.

Statins inhibit growth of human endometrial stromal cells independently of cholesterol availability.

Endometriosis is characterized by ectopic growth of endometrial tissues. Statins, inhibitors of 3-hydroxy-3methylglutaryl-coenzyme A reductase (HMGCR), have been shown to decrease proliferation of several mesenchymal tissues. Actions of statins may be related to decreased availability of cholesterol as well as intermediate metabolites of the mevalonate pathway downstream of HMGCR. This study was designed to evaluate effects of statins on growth of endometrial stromal cells and to investigate mechanisms of these effects. Human endometrial stromal cells were cultured in the absence and in the presence of serum and with or without mevastatin and simvastatin. DNA synthesis and viable cell numbers were determined. Effects of statins were also evaluated in the presence of mevalonate and squalene. Furthermore, effects on phosphorylation of mitogen-activated protein kinase 3/1 (MAPK3/1) (also known as extracellular signal-regulated kinase [ERK1/2]) were determined. Mevastatin and simvastatin induced a concentration-dependent inhibition of DNA synthesis and viable cell count in chemically defined media and in the presence of serum. Mevalonate, but not squalene, abrogated inhibitory effects of statins on cell proliferation. Statins inhibited MAPK3/1 phosphorylation. This is the first study demonstrating that statins inhibit growth of endometrial stromal cells. This effect is also demonstrable in the presence of a supply of cholesterol and may be related to decreased activation of MAPK3/1. The present observations may be relevant to potential therapeutic use of statins in conditions such as endometriosis.

Estrogen-mediated regulation of p38 mitogen-activated protein kinase in human endometrium.

CONTEXT: Estrogen predominantly exerts its biological effects through the genomic pathway by binding to its intracellular receptors, interacting with the estrogen response element located in the promoter region of the target gene, and thus regulating gene transcription. There has been an increasing body of evidence regarding nongenomic actions of estrogen. Estrogen activates multiple signaling cascades, including the MAPK pathway. p38 MAPK plays key roles in mediating apoptosis, proliferation, and inflammation, which all take place in the endometrium during cyclical changes under the influence of estrogen. OBJECTIVE: We hypothesized that estrogen might activate the p38 MAPK pathway in endometrial cells and exert some of its actions through this pathway in the endometrium. INTERVENTIONS: p38 MAPK phosphorylation was analyzed using in vivo and in vitro techniques. RESULTS: Total and phosphorylated p38 MAPK immunostainings were more intense in epithelial cells compared with stromal cells, and the phosphorylated/total p38 MAPK ratio was significantly higher in the functional endometrial layer compared with the basal layer (P < 0.05). Estradiol significantly increased p38 MAPK phosphorylation in endometrial stromal cells in culture within 2 min (P < 0.05), and this phosphorylation was blocked by a specific p38 MAPK inhibitor. Moreover, tamoxifen and raloxifene also increased phosphorylation of p38 MAPK. The estrogen receptor antagonist ICI 182,780 reversed the estrogen-induced p38 MAPK phosphorylation in endometrial stromal and epithelial cells, suggesting involvement of the estrogen receptor. CONCLUSION: Our results indicate the involvement of estrogen in regulating p38 MAPK activity in endometrial cells, suggesting a nongenomic action of estrogen through this MAPK in the endometrium.

Expression of interleukin-8 receptors in patients with adenomyosis.

OBJECTIVE: To investigate the expression of interleukin-8 (IL-8) receptors CXCR1 and CXCR2 in adenomyosis. DESIGN: Comparative immunohistochemical study. SETTING: Academic medical center. PATIENT(S): Thirty women who had undergone hysterectomy and were proved histopathologically to have adenomyosis, and 27 women without adenomyosis who had a hysterectomy for nonendometrial pathology such as leiomyomata or benign ovarian cysts. INTERVENTION(S): Tissue sections were immunostained with murine monoclonal anti-human CXCR1 and CXCR2 antibodies. MAIN OUTCOME MEASURE(S): Microscopic evaluation to assess the presence and localization of CXCR1 and CXCR2 throughout the menstrual cycle in both eutopic endometrial and adenomyotic tissues of women with adenomyosis and compare it with normal endometrium. RESULT(S): In eutopic endometrium of women with adenomyosis, proliferative phase samples showed higher epithelial CXCR1 and CXCR2 staining intensity compared with proliferative phase samples of normal endometrium. Adenomyosis foci expressed higher epithelial CXCR1 compared with the homologous eutopic endometrium and normal endometrium. On the other hand, adenomyosis foci and the homologous eutopic endometrium showed similar epithelial CXCR2 staining intensity, and this expression was higher than the normal controls. CONCLUSION(S): Intrinsic abnormalities concerning IL-8 and its receptor system may be present in the eutopic endometrium of women affected by adenomyosis. These findings suggest that IL-8 receptors may be involved in the pathogenesis and/or pathophysiology of adenomyosis.

Localization of NGF and nNOS in varicocele-induced rat testis.

Nerve growth factor (NGF) is synthesized in male germ cells. The presence of neuronal nitric oxide synthase (nNOS) in Leydig cells is related to its role in the regulation of testosterone release. Varicocele is often characterized by abnormal sperm quality and influences the fertilizing capacity of the haploid gamete. We investigated the localization of NGF and nNOS in testes of adult Wistar rats with experimentally induced varicocele after 9, 11, and 13 weeks, as well as in sham-operated controls by immunohistochemistry and Western blot. In control testis, we detected NGF in nuclei of Sertoli cells and also as small vesicular-like structures in the cytoplasm of primary spermatocytes, and in round and elongating spermatids. Varicocele-induction revealed a slight decrease of NGF at 13 weeks, especially in Sertoli cells. In control tissue, nNOS protein was present mainly in Leydig cells and in Sertoli cell cytoplasm. Additionally, nNOS immunoreactivity was present in the heads of elongated spermatids. Western blot results revealed that the decrease of NGF was not significant in the 13-week varicocele group, moreover, the amount of nNOS was not altered in any of the varicocele groups. In conclusion, NGF and nNOS have important roles for normal gametogenesis and our data for the first time indicates that varicocele induction does not necessarily affect the expression of NGF and nNOS. Thus, these two molecules do not appear to be related to varicocele induction.

Ultrastructural and immunohistochemical analysis of rat uroepithelial cell junctions after partial bladder outlet obstruction and selective COX-2 inhibitor treatment.

The present study was undertaken to evaluate alterations in uroepithelial cell junctional complexes in partial bladder outlet obstruction (PBOO) of rat bladders using ultrastructural morphometry and immunohistochemistry, and to determine whether selective COX-2 inhibitors have any effects on these structures. A total of 18 male rats were separated into three groups of six rats each: (1) sham-operated animals served as controls; (2) a PBOO group, without further treatment (3) and a group that immediately after PBOO, received treatment for 4 weeks with oral Celecoxib, a selective COX-2 inhibitor. Uroepithelial cell junctions were evaluated using transmission electron microscopy combined with morphometry. Results were also assessed by E-cadherin and alpha-catenin immunohistochemistry. Morphometrical analysis of ultrastructural evaluations revealed that 4 weeks of PBOO caused a significant reduction in the electron density of zonula adherens and zonula occludens junctional complexes. Moreover, some desmosomes located between the deeper cells of the uroepithelium showed signs of disintegration. Selective COX-2 inhibitor treatment during 4 weeks of PBOO showed protective effects on adherens and occludens junctions, as well as on desmosomes. Immunohistochemical analysis of E-cadherin confirmed that the decreased E-cadherin immunolabelling in 4 weeks of PBOO was prevented by selective COX-2 inhibitor treatment. Based on ultrastructural morphometrical analysis, we conclude that PBOO alone and in combination with selective COX-2 inhibitors can have considerable effects on uroepithelial cellular junctions. Our findings provide a novel area of investigation regarding the selective use of COX-2 inhibitors following PBOO.

Effects of excess vitamin B6 intake on cerebral cortex neurons in rat: an ultrastructural study.

The aim of this study was to investigate whether excess of vitamin B6 leads to ultrastructural changes in cerebral cortex of forty-eight healthy albino rats which were included in the study. Saline solution was injected to to the control groups (CG-10, n = 12 for 10 days; CG-15, n = 12 for 15 days; CG-20, n=12 for 20 days). The three experimental groups (EG-10, n = 12; EG-15, n = 12; EG-20, n = 12) were treated with 5 mg/kg vitamin B6 daily for 10 days (EG-10), 15 days (EG-15) and 20 days (EG-20). Brain tissues were prepared by glutaraldehyde-osmium tetroxide double fixation for ultrastructural analysis. No significant changes were observed in the control groups. The ultrastructural analysis revealed that the numbers of damaged mitochondria, lipofuscin granules and vacuoles were significantly higher in all the experimental groups than in the control groups (p < 0.05). However, synaptic density was significantly decreased in the experimental groups as compared to the control groups (p < 0.05). The results suggest that the excess of vitamin B6 intake causes damage to the cerebral cortex due to cellular intoxication and decreased synaptic density. Thus, careful attention should be paid to the time and dose of vitamin B6 recommended for patients who are supplemented with this vitamin.

Expression of interleukin-8 and monocyte chemotactic protein-1 in adenomyosis.

BACKGROUND: To clarify the inflammatory nature of adenomyosis, we aimed to investigate the expression of interleukin-8 (IL-8) and monocyte chemotactic protein-1 (MCP-1) by immunohistochemistry to determine their putative role in pathophysiology of adenomyosis. METHODS: Adenomyosis samples, with their eutopic endometrium, were collected from 30 women undergoing hysterectomy. Endometrium from 27 women without adenomyosis were also collected as a control group. Samples were grouped according to the menstrual cycle phase and examined by immunohistochemistry for IL-8 and MCP-1. RESULTS: In normal endometrium, secretory phase samples expressed higher levels of epithelial IL-8 than in proliferative phase samples (P = 0.01), and we observed a trend for an increased epithelial MCP-1 expression in the secretory phase samples compared with the proliferative phase samples (P = 0.07). Endometrial samples of women with adenomyosis did not show the same cyclic variation. In the secretory phase, eutopic endometrium of women with adenomyosis expressed lower levels of epithelial IL-8 and MCP-1 compared with normal endometrium (P < 0.05). The expression of epithelial IL-8 and MCP-1 was higher in the adenomyosis foci than the eutopic endometrium (P < 0.05). CONCLUSIONS: These findings may indicate that an intrinsic abnormality of inflammatory response may be present in eutopic endometrium of women with adenomyosis, and IL-8 and MCP-1 may contribute to the pathophysiology of adenomyosis.

Do glucose transporters have other roles in addition to placental glucose transport during early pregnancy?

Human placenta regulates the transport of maternal molecules to the fetus. It is known that glucose transport occurs via glucose transporters (GLUTs) in the feto-placental unit. Data on the expression of GLUTs during implantation are very scarce. Moreover, the question of how the decidual leukocytes obtain the energy for their activation during implantation mechanism is still under investigation. We studied the distributions of GLUT1, GLUT3, and GLUT4 in tissue sections of first trimester pregnancies the human maternal-fetal interface. GLUT1 was present in apical microvilli of the syncytiotrophoblast, in cytotrophoblast, and in vascular patterns of the villous core, whereas GLUT3 was localized in cytotrophoblasts of placental villi and in some fetal endothelial cells. Moreover, the proliferating cells of the proximal cell columns were also immunopositive for GLUT1 and GLUT3. We did not observe any positive immunoreactivity for GLUT4 in placental and decidual tissues. Essentially, GLUT3 and also to some extent GLUT1 was present in maternal leukocytes and platelets. In conclusion, our results suggest that the glucose taken up via GLUT1 and GLUT3 from the maternal circulation might not only be needed for placental functions but also for successful implantation by trophoblast invasion, proliferation and also by having a role to support energy for maternal leukocytes.

The restorative effect of a selective cyclooxygenase-2 inhibitor on urothelial cell-cell interactions after partial bladder outlet obstruction in rats.

OBJECTIVE: To determine the changes in cyclooxygenase-2 (COX-2), E-cadherin and alpha-catenin expression after partial bladder outlet obstruction (PBOO), and whether a selective COX-2 inhibitor (celecoxib) might inhibit COX-2 expression and have beneficial effects on urothelial cell-to-cell interactions in rats subjected to PBOO. MATERIALS AND METHODS: Thirty-six male rats were divided into six equal groups; celecoxib was administered after creating PBOO for 1 and 4 weeks in groups 1 and 2, respectively. Two further obstructed groups (3 and 4, PBOO for 1 and 4 weeks, respectively) received no treatment. Sham-operated animals served as controls (group 5 and 6, assessed at 1 and 4 weeks, respectively). After 1 and 4 weeks of PBOO or a sham procedure the bladder weight was recorded before sampling the bladder for Western blotting and immunohistological analysis, to assess the expressions of COX-2 and adherens proteins, E-cadherin and alpha-catenin. Urothelial cell-to-cell interactions were evaluated using electron microscopy. RESULTS: The bladder mass increased rapidly during the first 7 days after PBOO in groups 1-4 compared with 5 and 6 (P < 0.05). While the bladder mass then continued to increase for the next 21 days in group 4, it was constant in group 2 (P < 0.001). Immunohistochemical staining and Western blotting analyses showed that E-cadherin and alpha-catenin expression were reversibly decreased in rats with PBOO, while COX-2 protein expression was up-regulated. After giving celecoxib there was a significant decrease in COX-2 expression and a restoration of intercellular adherens junctions and desmosomes, as assessed on electron microscopy and expression of adherens proteins combined. CONCLUSION: The increase in COX-2 expression attributable to hypoxia and the tensile strength of bladder wall was attenuated by celecoxib. Selective COX-2 inhibitors have important restorative effects on intercellular adherens junctions and desmosomes in PBOO.

Effect of experimental varicocele on the expressions of Notch 1, 2, and 3 in rat testes: an immunohistochemical study.

OBJECTIVE: To study expressions of Notch receptor isoforms (Notch 1, 2, and 3) in normal and varicocele-induced rat testes to examine their possible functions in cell fate. DESIGN: Comparative and controlled study. SETTING: Animal Care and Operation Unit, Akdeniz University. ANIMAL(S): Wistar male rats for experimental and control groups. INTERVENTION(S): The control group underwent a sham operation (n = 6). The experimental groups underwent partial ligation of the renal vein to induce an experimental varicocele and then were killed 9 (n = 6), 11 (n = 6), and 13 (n = 6) weeks after the induction of varicocele. MAIN OUTCOME MEASURE(S): All tissues were fixed and routinely processed for paraffin embedding. Subsequent immunohistochemical studies were performed. RESULT(S): In the sham-operation rat testes, Leydig cells and elongated spermatids were immunopositive for Notch 1. Notch-2 expression was present in Leydig cells, spermatogonia, and primary spermatocytes. Notch-3 expression was limited to Leydig cells. Varicocele formation diminished the expression of both Notch-1 and Notch-2 receptors as the varicocele formation progressed over time. CONCLUSION(S): The present study suggests that Notch 1 is related to the maturation of spermatids. Notch 2 is related to both proliferation and maturation of spermatogenic cells, whereas Notch 3 seems to be related to Leydig cell functions. The decrease of both Notch-1 and Notch-2 expression depended on the degree of varicocele development over time, indicating a potential role in varicocele-associated testicular dysfunction.

Regulation of interleukin-8 expression in human endometrial endothelial cells: a potential mechanism for the pathogenesis of endometriosis.

The elevation of the proinflammatory chemoattractant cytokine levels in ectopic and eutopic endometrium of endometriosis implies an inflammatory basis for this disease. The relationship between endothelial cells and leukocytes is likely to be important in the regulation of inflammatory mediators of endometriosis. The aim of this study was to describe the temporal and spatial expression of IL-8 in human endometrial endothelial cells (HEEC) in vivo and to compare the in vitro regulation of IL-8 expression by sex steroids in HEEC from women with or without endometriosis. Eutopic endometrial tissues and endometriosis implants were grouped according to menstrual cycle phase and examined by immunohistochemistry for IL-8 expression. Endothelial cells of endometriotic implants expressed higher IL-8 immunoreactivity compared with endothelial cells of eutopic endometrium from women with or without endometriosis (P < 0.02). For in vitro studies, HEEC were isolated from women with or without endometriosis and grown to preconfluence. The purity of cultured HEEC (90-95%) was confirmed by immunocytochemistry using endothelium-specific markers, CD31 and CD146. The effects of estradiol (5 x 10(-8) m), progesterone (10(-7) m), or both on IL-8 mRNA and protein levels were analyzed by RT-PCR and ELISA, respectively. Sex steroids reduced the expression of IL-8 mRNA and protein in HEEC from women without endometriosis. In contrast, both estradiol and progesterone stimulated IL-8 mRNA and protein expression in HEEC from women with endometriosis. We postulate that the stimulation of chemokine expression by sex steroids in HEEC of women with endometriosis may play a role in the inflammatory aspect of this disease.

Expression of interleukin-8 receptors in endometriosis.

BACKGROUND: Although the etiology of endometriosis is not well understood, chemokines and their receptors are believed to play a role in its pathogenesis. Therefore, we aimed to investigate the expression and localization of interleukin-8 (IL-8) receptors CXCR1 and CXCR2 in eutopic and ectopic endometrial tissues of women with endometriosis, and in endometrium of women without endometriosis. METHODS: Ectopic (n = 27) and homologous eutopic endometrium (n = 25) from women with endometriosis and endometrium from women without endometriosis (n = 27) were used for immunohistochemical analysis of CXCR1 and CXCR2. RESULTS: In normal endometrium, epithelial CXCR1 and CXCR2 immunostaining intensities were similar in the proliferative and secretory phase. Stromal CXCR1 expression was less then epithelial expression and did not show cyclical difference. No stromal CXCR2 expression was observed. In eutopic endometrium of women with endometriosis compared to endometrium of women without endometriosis, there was a significant increase in both proliferative and secretory phases for epithelial CXCR2 expression, and in proliferative phase for CXCR1 expression (P < 0.05). Both receptor immunoreactivities were significantly increased in the epithelial cells of ectopic endometrial tissues compared to that of normal endometrium (P < 0.05). CONCLUSIONS: These findings suggest that IL-8 and its receptors may be involved in the pathogenesis of endometriosis.