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Hawkins Bressler L, Fritz MA, Wu SP, Yuan L, Kafer S, Wang T, DeMayo FJ, Young SL. Poor Endometrial Proliferation After Clomiphene is Associated With Altered Estrogen Action. J Clin Endocrinol Metab 2021; 106:2547-2565. [PMID: 34058008 PMCID: PMC8372647 DOI: 10.1210/clinem/dgab381] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/13/2021] [Indexed: 12/25/2022]
Abstract
CONTEXT Suboptimal endometrial thickening is associated with lower pregnancy rates and occurs in some infertile women treated with clomiphene. OBJECTIVE To examine cellular and molecular differences in the endometrium of women with suboptimal vs optimal endometrial thickening following clomiphene. METHODS Translational prospective cohort study from 2018 to 2020 at a university-affiliated clinic. Reproductive age women with unexplained infertility treated with 100 mg of clomiphene on cycle days 3 to 7 who developed optimal (≥8mm; n = 6, controls) or suboptimal (<6mm; n = 7, subjects) endometrial thickness underwent preovulatory blood and endometrial sampling. The main outcome measures were endometrial tissue architecture, abundance and location of specific proteins, RNA expression, and estrogen receptor (ER) α binding. RESULTS The endometrium of suboptimal subjects compared with optimal controls was characterized by a reduced volume of glandular epithelium (16% vs 24%, P = .01), decreased immunostaining of markers of proliferation (PCNA, ki67) and angiogenesis (PECAM-1), increased immunostaining of pan-leukocyte marker CD45 and ERβ, but decreased ERα immunostaining (all P < .05). RNA-seq identified 398 differentially expressed genes between groups. Pathway analysis of differentially expressed genes indicated reduced proliferation (Z-score = -2.2, P < .01), decreased angiogenesis (Z-score = -2.87, P < .001), increased inflammation (Z-score = +2.2, P < .01), and ERβ activation (Z-score = +1.6, P < .001) in suboptimal subjects. ChIP-seq identified 6 genes bound by ERα that were differentially expressed between groups (P < .01), some of which may play a role in implantation. CONCLUSION Women with suboptimal endometrial thickness after clomiphene exhibit aberrant ER expression patterns, architectural changes, and altered gene and protein expression suggesting reduced proliferation and angiogenesis in the setting of increased inflammation.
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Affiliation(s)
- Leah Hawkins Bressler
- Department of Obstetrics & Gynecology, Division of Reproductive Endocrinology & Infertility, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
| | - Marc A Fritz
- Department of Obstetrics & Gynecology, Division of Reproductive Endocrinology & Infertility, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
| | - San-Pin Wu
- Reproductive and Developmental Biology Laboratory, National Institute of Environmental Health Sciences, Research Triangle Park, NC, USA
| | - Lingwen Yuan
- Reproductive and Developmental Biology Laboratory, National Institute of Environmental Health Sciences, Research Triangle Park, NC, USA
| | - Suzanna Kafer
- Reproductive and Developmental Biology Laboratory, National Institute of Environmental Health Sciences, Research Triangle Park, NC, USA
| | - Tianyuan Wang
- Integrative Bioinformatics Support Group, National Institute of Environmental Health Sciences, Research Triangle Park, NC, USA
| | - Francesco J DeMayo
- Reproductive and Developmental Biology Laboratory, National Institute of Environmental Health Sciences, Research Triangle Park, NC, USA
| | - Steven L Young
- Department of Obstetrics & Gynecology, Division of Reproductive Endocrinology & Infertility, University of North Carolina at Chapel Hill, Chapel Hill, NC, USA
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Ali FF, Abdelzaher WY, Ibrahim RA, Elroby Ali DM. Amelioration of estrogen-induced endometrial hyperplasia in female rats by hemin via heme-oxygenase-1 expression, suppression of iNOS, p38 MAPK, and Ki67. Can J Physiol Pharmacol 2019; 97:1159-1168. [PMID: 31505119 DOI: 10.1139/cjpp-2019-0287] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/26/2022]
Abstract
Although heme oxygenase-1 (HO-1) is part of an endogenous defense system implicated in the homeostatic response, its role in cell proliferation and tumor progression is still controversial. Endometrial hyperplasia (EH) is associated with high risk of endometrial cancer (EC). Therefore, we aimed to evaluate the effect of hemin, a HO-1 inducer, against EH. Thirty-two female rats (60-70 days old) were divided into 4 groups treated for 1 week: vehicle control group, hemin group (25 mg/kg; i.p. 3 times/week), estradiol valerate (EV) group (2 mg/kg per day, p.o.), and hemin plus EV group. Sera were obtained for reduced glutathione level. Uterine malondialdehyde, superoxide dismutase, total nitrite/nitrate, and interleukin-1β levels were estimated. HO-1 and p38 mitogen-activated protein kinase expressions were obtained in uterine tissue. Uterine histological and immunohistochemical assessment of iNOS and Ki67 were also done. Results demonstrated that upregulation of HO-1 expression in hemin plus EV rats led to amelioration of EH which was confirmed with histological examination. This was associated with significant decrease in oxidative stress parameters, p38 mitogen-activated protein kinase expression, and interleukin-1β level. Also, uterine iNOS and Ki67 expressions were markedly suppressed. In conclusion, upregulation of HO-1 expression via hemin has ameliorative effect against EH through its antioxidant, anti-inflammatory, and antiproliferative actions.
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Affiliation(s)
- Fatma F Ali
- Department of Medical Physiology, Faculty of Medicine, Minia University, Egypt
| | | | - Randa Ahmed Ibrahim
- Department of Histology and Cell Biology, Faculty of Medicine, Minia University, Egypt
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Autophagy is upregulated in ovarian endometriosis: a possible interplay with p53 and heme oxygenase-1. Fertil Steril 2015; 103:1244-51.e1. [DOI: 10.1016/j.fertnstert.2015.02.007] [Citation(s) in RCA: 51] [Impact Index Per Article: 5.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/04/2014] [Revised: 01/27/2015] [Accepted: 02/05/2015] [Indexed: 12/31/2022]
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Higashimura Y, Naito Y, Takagi T, Mizushima K, Hirai Y, Harusato A, Ohnogi H, Yamaji R, Inui H, Nakano Y, Yoshikawa T. Oligosaccharides from agar inhibit murine intestinal inflammation through the induction of heme oxygenase-1 expression. J Gastroenterol 2013. [PMID: 23188093 DOI: 10.1007/s00535-012-0719-4] [Citation(s) in RCA: 60] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/04/2023]
Abstract
BACKGROUND Agarose is hydrolyzed easily to yield oligosaccharides, designated as agaro-oligosaccharides (AGOs). Recently, it has been demonstrated that AGOs induce heme oxygenase-1 (HO-1) expression in macrophages and that they might lead to anti-inflammatory property. Nevertheless, the molecular mechanism of AGO-mediated HO-1 induction remains unknown, as does AGOs' ability to elicit anti-inflammatory activity in vivo. This study was undertaken to uncover the mechanism of AGO-mediated HO-1 induction and to investigate the therapeutic effect of AGOs on intestinal inflammation. METHODS Mice were treated with 2,4,6-trinitrobenzene sulfonic acid (TNBS) to induce colitis. The respective degrees of mucosal injury of mice that had received AGO and control mice were compared. We investigated HO-1 expression using Western blotting, quantitative real-time PCR (qRT-PCR), and immunohistochemistry. The expression of tumor necrosis factor-α (TNF-α) was measured using qRT-PCR and enzyme-linked immunosorbent assay. RESULTS AGO administration induced HO-1 expression in colonic mucosa. The induction was observed mainly in F4/80 positive macrophages. Increased colonic damage and myeloperoxidase activity after TNBS treatment were inhibited by AGO administration. TNBS treatment induced TNF-α expression, and AGO administration suppressed induction. However, HO inhibitor canceled AGO-mediated amelioration of colitis. In RAW264 cells, AGOs enhanced HO-1 expression time-dependently and concentration-dependently and suppressed lipopolysaccharide-induced TNF-α expression. Furthermore, agarotetraose-mediated HO-1 induction required NF-E2-related factor 2 function and phosphorylation of c-jun N-terminal kinase. CONCLUSIONS We infer that AGO administration inhibits TNBS-induced colitis in mice through HO-1 induction in macrophages. Consequently, oral administration of AGOs might be an important therapeutic strategy for inflammatory bowel disease.
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Affiliation(s)
- Yasuki Higashimura
- Molecular Gastroenterology and Hepatology, Graduate School of Medical Science, Kyoto Prefectural University of Medicine, 465 Kajii-cho, Kawaramachi-Hirokoji, Kamigyo-ku, Kyoto, Kyoto, 602-8566, Japan
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Abstract
The ovarian steroid hormones progesterone and estradiol are well established regulators of human endometrial function. However, more recent evidence suggests that androgens and locally generated steroids, such as the glucocorticoids, also have a significant impact on endometrial breakdown and repair. The temporal and spatial pattern of steroid receptor presence in endometrial cells has a significant impact on the endometrial response to steroids. Furthermore, regulation of steroid receptor function by modulatory proteins further refines local responses. This review focuses on steroid regulation of endometrial function during the luteo-follicular transition with a focus on menstruation and endometrial repair.
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Affiliation(s)
- Jacqueline A Maybin
- MRC Centre for Reproductive Health, The University of Edinburgh, The Queen's Medical Research Institute, Edinburgh, UK
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Iwahara Y, Nagai A, Yoshiki N, Igarashi K, Yamashita K, Kubota T. Expression of heme oxygenase in the eutopic and ectopic endometrium in patients with adenomyosis. Gynecol Endocrinol 2012; 28:892-6. [PMID: 22559824 DOI: 10.3109/09513590.2012.683064] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/28/2023] Open
Abstract
Heme oxygenase (HO) is the rate-limiting enzyme that catalyzes the degradation of heme into iron, carbon monoxide, and biliverdin. This enzyme has important functions in cellular homeostasis, including the regulation of oxidative load, apoptosis, and inflammation. Two isoforms of HO, the inducible HO-1 and the constitutive HO-2, are expressed and are known to play a role in the normal human endometrium throughout the menstrual cycle, but there is little evidence for HO expression and behavior in adenomyosis, which is the occurrence of intramural ectopic endometrial tissue. The aim of this study was to investigate the presence and localization of the two HO isoforms in both eutopic and ectopic endometrium of women with adenomyosis during the menstrual cycle. The oxidative stress and apoptosis related to HO-1 expression were also assessed. The expression of HO-1 and HO-2 in both eutopic and ectopic endometrium was confirmed, and their levels in the ectopic endometrium were lower than those in the eutopic endometrium. The cyclic variability of HO expression was lost in the ectopic endometrium during the menstrual cycle, whereas this variability was apparent in the eutopic endometrium. Moreover, HO-1 expression corresponded to apoptotic events in the eutopic endometrium. Constitutive HO-2 expression corresponded to endometrial proliferation and degradation. These results reveal that both HO-1 and HO-2 contribute little in the pathophysiology of adenomyosis.
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Affiliation(s)
- Yuki Iwahara
- Comprehensive Reproductive Medicine, Graduate School, Tokyo Medical and Dental University, Yushima, Bunkyo-ku, Tokyo, Japan
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Rudolph M, Döcke WD, Müller A, Menning A, Röse L, Zollner TM, Gashaw I. Induction of overt menstruation in intact mice. PLoS One 2012; 7:e32922. [PMID: 22412950 PMCID: PMC3296749 DOI: 10.1371/journal.pone.0032922] [Citation(s) in RCA: 35] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/20/2011] [Accepted: 02/06/2012] [Indexed: 11/19/2022] Open
Abstract
The complex tissue remodeling process of menstruation is experienced by humans and some primates, whereas most placental mammals, including mice, go through an estrous cycle. How menstruation and the underlying mechanisms evolved is still unknown. Here we demonstrate that the process of menstruation is not just species-specific but also depends on factors which can be induced experimentally. In intact female mice endogenous progesterone levels were raised by the induction of pseudopregnancy. Following an intrauterine oil injection, the decidualization of the endometrium was reliably induced as a prerequisite for menstruation. The natural drop of endogenous progesterone led to spontaneous breakdown of endometrial tissue within an average of 3 days post induction of decidualization. Interestingly, morphological changes such as breakdown and repair of the endometrial layer occurred in parallel in the same uterine horn. Most importantly, endometrial breakdown was accompanied by vaginally visible (overt) bleeding and flushing out of shed tissue comparable to human menstruation. Real-time PCR data clearly showed temporal changes in the expression of multiple factors participating in inflammation, angiogenesis, tissue modulation, proliferation, and apoptosis, as has been described for human menstruating endometrium. In conclusion, human menstruation can be mimicked in terms of extravaginally visible bleeding, tissue remodeling, and gene regulation in naturally non-menstruating species such as intact female mice without the need for an exogenous hormone supply.
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Affiliation(s)
- Marion Rudolph
- Target Discovery, Global Drug Discovery, Bayer HealthCare, Berlin, Germany.
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Naito Y, Takagi T, Uchiyama K, Yoshikawa T. Heme oxygenase-1: a novel therapeutic target for gastrointestinal diseases. J Clin Biochem Nutr 2011; 48:126-33. [PMID: 21373265 PMCID: PMC3045685 DOI: 10.3164/jcbn.10-61] [Citation(s) in RCA: 58] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/01/2010] [Accepted: 07/01/2010] [Indexed: 12/18/2022] Open
Abstract
Heme oxygenase-1 (HO-1) is the rate-limiting enzyme in the catabolism of heme, followed by production of biliverdin, free iron and carbon monoxide (CO). HO-1 is a stress-responsive protein induced by various oxidative agents. Recent studies demonstrate that the expression of HO-1 in response to different inflammatory mediators may contribute to the resolution of inflammation and has protective effects in several organs against oxidative injury. Although the mechanism underlying the anti-inflammatory actions of HO-1 remains poorly defined, both CO and biliverdin/bilirubin have been implicated in this response. In the gastrointestinal tract, HO-1 is shown to be transcriptionally induced in response to oxidative stress, preconditioning and acute inflammation. Recent studies suggest that the induction of HO-1 expression plays a critical protective role in intestinal damage models induced by ischemia-reperfusion, indomethacin, lipopolysaccharide-associated sepsis, trinitrobenzene sulfonic acid, and dextran sulfate sodium, indicating that activation of HO-1 may act as an endogenous defensive mechanism to reduce inflammation and tissue injury in the gastrointestinal tract. In addition, CO derived from HO-1 is shown to be involved in the regulation in gastro-intestinal motility. These in vitro and in vivo data suggest that HO-1 may be a novel therapeutic target in patients with gastrointestinal diseases.
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Affiliation(s)
- Yuji Naito
- Department of Molecular Gastroenterology and Hepatology, Kyoto Prefectural University of Medicine, Kamigyo-ku, Kyoto 602-8566, Japan
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Takagi T, Naito Y, Uchiyama K, Yoshikawa T. The role of heme oxygenase and carbon monoxide in inflammatory bowel disease. Redox Rep 2011; 15:193-201. [PMID: 21062534 DOI: 10.1179/174329210x12650506623889] [Citation(s) in RCA: 29] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
Inflammatory bowel disease (IBD), including ulcerative colitis (UC) and Crohn's disease, is a chronic and recurrent inflammatory disorder of the intestinal tract. Since the precise pathogenesis of IBD remains unclear, it is important to investigate the pathogenesis of IBD and to evaluate new anti-inflammatory strategies. Recent evidence suggests that heme oxygenase-1 (HO-1) plays a critical protective role during the development of intestinal inflammation. In fact, it has been demonstrated that the activation of HO-1 may act as an endogenous defensive mechanism to reduce inflammation and tissue injury in various animal intestinal injury models induced by ischemia-reperfusion, indomethacin, lipopolysaccharide-associated sepsis, trinitrobenzene sulfonic acid or dextran sulfate sodium. In addition, carbon monoxide (CO) derived from HO-1 has been shown to be involved in the regulation of intestinal inflammation. Furthermore, administration of a low concentration of exogenous CO has a protective effect against intestinal inflammation. These data suggest that HO-1 and CO may be novel therapeutic molecules for patients with gastrointestinal inflammatory diseases. In this review, we present what is currently known regarding the role of HO-1 and CO in intestinal inflammation.
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Affiliation(s)
- Tomohisa Takagi
- Molecular Gastroenterology and Hepatology, Graduate School of Medical Science, Kyoto Prefectural University of Medicine, Kyoto, Japan
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Khorram O, Han G, Magee T. Cigarette smoke inhibits endometrial epithelial cell proliferation through a nitric oxide-mediated pathway. Fertil Steril 2010; 93:257-63. [PMID: 19022425 DOI: 10.1016/j.fertnstert.2008.09.074] [Citation(s) in RCA: 27] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/09/2008] [Revised: 09/10/2008] [Accepted: 09/25/2008] [Indexed: 11/24/2022]
Abstract
OBJECTIVE To determine the direct effects of cigarette smoke (CS) on human endometrial surface epithelial cell line proliferation. DESIGN In vitro study using HES cells and primary human endometrial cells. SETTING University-based academic center. PATIENT(S) Premenopausal women in the proliferative phase of the cycle. INTERVENTION(S) The HES cells and primary human endometrial cells were exposed to cigarette smoke-saturated solution. MAIN OUTCOME MEASURE(S) Cell proliferation and expression of different isoforms of nitric oxide synthase. RESULT(S) Cigarette smoke inhibited HES cell proliferation in a dose- and time-dependant manner. The inhibitory effect of CS was blocked by hemoglobin and enhanced by L-arginine (L-Arg). Cigarette smoking and nicotine stimulated the expression of endothelial NO synthase (eNOS) and inducible NO synthase (iNOS) whereas benzo[a]pyrene (BP) only stimulated the expression of eNOS in HES cells. Cigarette smoke stimulated the expression of eNOS/iNOS in primary human endometrial cells, comprised of epithelial and stromal cells. The effect of CS on eNOS/iNOS expression in HES cells was blocked by ascorbic acid but not by glutathione. CONCLUSION(S) Cigarette smoke inhibits endometrial cell proliferation through a nitric oxide-mediated pathway.
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Affiliation(s)
- Omid Khorram
- Department of Obstetrics and Gynecology, Harbor-UCLA Medical Center, Torrance, California, USA.
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Ovári L, Aranyosi J, Balla G. Acute effect of cigarette smoking on placental circulation - a study by carbon-monoxide measurement and Doppler assessment. ACTA ACUST UNITED AC 2009; 96:243-50. [PMID: 19457768 DOI: 10.1556/aphysiol.96.2009.2.9] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/19/2022]
Abstract
OBJECTIVE Carbon-monoxide (CO) decreases placental vascular impedance. We assessed the consequences of smoking-induced temporary maternal CO-increase on fetal and placental circulation. STUDY DESIGN In a prospective study twenty-nine smoking pregnant women and their fetuses were evaluated. We determined the changes in maternal blood CO levels after smoking, and the concomitant changes in maternal and fetal circulation. Changes in fetal heart rate, uterine artery (UTA), middle cerebral artery (MCA), and descending aorta (DA) flow were measured by Doppler velocimetry. Changes in maternal CO level and umbilical flow value were assessed by paired t-test. The correlation between CO level and placental flow was assessed by partial correlation test. RESULTS CO level increased (mean +/-SD 1.7 +/- 0.065% vs. 2.36 +/- 0.89, p<0.0001). Nicotine-related maternal circulatory parameters changed significantly, but uterine flow values remained unchanged. Fetal heart rate increased, while flow in MCA and DA showed no change. CO-dependent umbilical artery impedance remained unchanged (Pulsatility Index: 0.956 +/- 0.18 vs. 0.948 +/- 0.21). Partial correlation between CO level and umbilical arterial impedance showed no significance (r:-0.324). CONCLUSION Despite significant CO elevation, the mainly CO-regulated placental flow remained unchanged.
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Affiliation(s)
- László Ovári
- University of Debrecen, Department of Obstetrics and Gynecology, Medical and Health Sciences Center H-4012 Debrecen Hungary.
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Kobayashi H, Yamada Y, Kanayama S, Furukawa N, Noguchi T, Haruta S, Yoshida S, Sakata M, Sado T, Oi H. The role of iron in the pathogenesis of endometriosis. Gynecol Endocrinol 2009; 25:39-52. [PMID: 19165662 DOI: 10.1080/09513590802366204] [Citation(s) in RCA: 56] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
BACKGROUND Endometriosis may cause symptoms including chronic pelvic pain and infertility, and increases susceptibility to the development of ovarian cancer. Genomic studies have started to delineate the wide array of mediators involved in the development of endometriosis. Understanding the mechanisms of endometriosis development and elucidating its pathogenesis and pathophysiology are intrinsic to prevention and the search for effective therapies. METHOD OF STUDY The present article reviews the English language literature for biological, pathogenetic and pathophysiological studies on endometriosis. Several recent genomic studies are discussed in the context of endometriosis biology. RESULTS Severe hemolysis occurring during the development of endometriosis results in high levels of free heme and iron. These compounds oxidatively modify lipids and proteins, leading to cell and DNA damage, and subsequently fibrosis development. Recent studies based on genome-wide expression analysis technology have noted specific expression of heme/iron-dependent mediators in endometriosis. The heme/iron-dependent signaling pathway of endometriosis, which is providing new insights into the regulation of inflammation, detoxification and survival, is discussed. CONCLUSION Several important endometriosis-specific genes overlap with those known to be regulated by iron. Other genes are involved in oxidative stress. Iron has a significant impact on endometriotic-cell gene expression. This review summarizes recent advances in the heme/iron-mediated signaling and its target genes, outlines the potential challenges to understanding of the pathogenesis and pathophysiology of endometriosis, and proposes a possible novel model.
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Affiliation(s)
- Hiroshi Kobayashi
- Department of Obstetrics and Gynecology, Nara Medical University, 840 Shijo-cho, Kashihara, Nara, Japan.
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Takagi T, Naito Y, Mizushima K, Nukigi Y, Okada H, Suzuki T, Hirata I, Omatsu T, Okayama T, Handa O, Kokura S, Ichikawa H, Yoshikawa T. Increased intestinal expression of heme oxygenase-1 and its localization in patients with ulcerative colitis. J Gastroenterol Hepatol 2008; 23 Suppl 2:S229-33. [PMID: 19120903 DOI: 10.1111/j.1440-1746.2008.05443.x] [Citation(s) in RCA: 43] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/09/2022]
Abstract
BACKGROUND Heme oxygenase-1 (HO-1) is regarded as a sensitive and reliable indicator of cellular oxidative stress. Two end products of heme degradation, carbon monoxide (CO) and bilirubin, are involved in the protective role of HO-1 against oxidative injury. We have demonstrated enhanced expression of this enzyme and increased concentration of CO in experimental models of colitis, but the role of HO-1 in patients with ulcerative colitis (UC) has not been extensively investigated. The aim of the present study was to determine the intestinal levels and localization of ho-1 mRNA and HO-1 protein in patients with UC. METHODS Eighteen patients with UC and 13 patients with colon cancer were prospectively selected from subjects who underwent colonoscopy. Biopsy specimens were obtained from the inflamed mucosa of UC patients and from the normal mucosa at least 5 cm from the margin of carcinoma. The expression of ho-1 mRNA was assayed by real-time polymerase chain reaction (PCR). The colonic expression of HO-1 was determined by immunohistochemistry and western blotting using a monoclonal antibody against HO-1. RESULTS The expression of ho-1 mRNA and HO-1 protein was significantly increased in the colonic mucosa of patients with active UC compared with normal mucosa. In the patients with active UC, mononuclear cells in the submucosa of the colon were positive for HO-1, and there was negligible staining in the epithelial cells. CONCLUSION The present findings are evidence of the induction of HO-1 in the colon of UC patients.
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Affiliation(s)
- Tomohisa Takagi
- Biomedical Safety Science, Graduate School of Medical Science, Kyoto Prefectual University of Medicine, Kawaramachi-Hirokoji, Kamigyo-ku, Kyoto, Japan
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Kubota T. Role of vasoactive substances on endometrial and ovarian function. Reprod Med Biol 2007; 6:157-164. [PMID: 29662409 DOI: 10.1111/j.1447-0578.2007.00179.x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022] Open
Abstract
In this review, it is proposed that the vasoactive agents endothelin (ET), nitric oxide (NO)/NO synthase (NOS) and carbon monoxide(CO)/heme oxygenase(HO) act directly on human endometrial functions and on ovarian functions in the normal menstrual cycle and in implantation periods. These vasoactive substances are likely to be important autocrine/paracrine factors that regulate a variety of physiological and pathological processes. The main actions of these agents are differentiation and implantation in the endometrial functions, and follicular growth, luteinization and atresia in the ovarian functions, in the tight connection between endometrial and ovarian systems during normal menstrual periods and during implantation (Reprod Med Biol 2007; 6: 157-164).
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Affiliation(s)
- Toshiro Kubota
- Comprehensive Reproductive Medicine, Graduate School, Tokyo Medical and Dental University, Tokyo, Japan
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Abstract
AIM: To purify and identify heme oxygenase (HO) isomers which exist in rat liver, spleen and brain treated with hematin and phenylhydrazine and in untreated rat liver and to investigate the characteristics of HO isomers, to isolate and confirm the rat HO-1 cDNA that actually encodes HO-1 by expressing cDNA in monkey kidney cells (COS-1 cells), to prepare the rat heme oxygenase-1 (HO-1) mutant and to detect inhibition of HO-1 mutated enzyme.
METHODS: First, rat liver, spleen and brain microsomal fractions were purified by DEAE-Sephacel and hydroxylapatite. The characteristics including activity, immunity and inducibility of two isomers (HO-1 and HO-2), and their apparent molecular weight were measured by detecting enzymatic activities, SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting analysis, respectively. Second, plasmid pcDNA3HO1 containing native rat HO-1 cDNA and pcDNA3HO1D25 carrying mutated rat HO-1 cDNA (His25Ala) were constructed by site-directed mutagenesis. COS-1 cells transfected with pcDNA3HO1 and pcDNA3HO1D25 were collected and disrupted by sonication, the microsomes were prepared by ultracentrifugation. Third, the inhibition of rat HO-1 mutant was analyzed.
RESULTS: Two isomers were purified and identified in treated rat liver, spleen, brain and untreated rat liver. HO-1 was the predominant form with a ratio of 2.0:1 and 3.2:1 of HO-1 and HO-2 in liver and spleen, respectively, but only the activity of HO-2 in the brain and untreated liver could be detected. The apparent molecular weights of HO-1 and HO-2 were about Mr 30000 and Mr 36000 under reducing conditions, respectively. The antiserum against liver HO-2 was employed in Western blotting analysis, the reactivity of HO-1 in the liver was not observed. The plasmid pcDNA3HO1 was highly expressed in endoplasmic reticulum of transfected COS-1 cells. The specific activity was ≈5-fold higher than that of the control. However, the enzyme activity of mutated HO-1 declined. While an equal amount of mutant was added to the enzyme reaction system, the levels of bilirubin decreased 42%.
CONCLUSION: The studies suggest that HO-1 and HO-2 exist in the hematin and phenylhydrazine treated rat liver and spleen, but only HO-2 in the brain and untreated liver. Two constitutive forms are different in molecular weight, inducibility and immunochemical properties. The activity of expressed HO-1 in COS-1 cells is higher than that of purified enzyme from rat spleen tissue. It suggests that this clone has an insert of 1030 base-pairs encodes HO-1. His25Ala mutant reduced the formation of bilirubin and it suggests that the mutant could competely bind the heme with native enzyme.
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Affiliation(s)
- Zhen-Wei Xia
- Department of Pediatrics, Rui Jin Hospital, Shanghai Second Medical University, China.
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Casanas-Roux F, Van Langendonckt A, Dolmans MM, Donnez J. Expression of inducible heme oxygenase in human endometrium. Fertil Steril 2002; 78:1327-8. [PMID: 12477535 DOI: 10.1016/s0015-0282(02)04395-9] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
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