|Year : 2015 | Volume
| Issue : 4 | Page : 484-489
Asymmetric dimethylarginine (ADMA), nitric oxide metabolite, and estradiol levels in serum and peritoneal fluid in women with endometriosis
Maryam Kianpour1, Mehdi Nematbakhsh2, Sayad Mehdi Ahmadi3
1 Nursing and Midwifery Care Research and Department of Midwifery, Faculty of Nursing and Midwifery, Isfahan University of Medical Sciences, Isfahan, Iran
2 Water and Electrolytes Research Center and Department of Physiology, Faculty of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran
3 Isfahan Fertility and Infertility Center, Isfahan, Iran
|Date of Submission||08-May-2014|
|Date of Acceptance||07-Jan-2015|
|Date of Web Publication||17-Jul-2015|
Prof. Mehdi Nematbakhsh
Water and Electrolytes Research Center, Isfahan University of Medical Sciences, Isfahan
Source of Support: No, Conflict of Interest: This research was supported by Isfahan University of Medical Sciences.
Background: Increase in nitric oxide (NO) concentration accompanied by alteration in peritoneal immune defense reactions is involved in the pathogenesis of endometriosis. Asymmetric dimethylarginine is an endogenous competitive inhibitor of NO synthase. This study was designed to compare NO metabolite (nitrite), asymmetric dimethylarginine, and estradiol concentrations in serum and peritoneal fluid (PF) of patients with and without endometriosis.
Materials and Methods: Subjects were assigned to two groups based on their laparoscopic results. The groups consisted of women with and without endometriosis (90 and 89 participants, respectively). The serum and peritoneal levels of nitrite (stable NO metabolite), asymmetric dimethylarginine, and estradiol were measured using enzyme-linked immunosorbent assay (ELISA) kits. These parameters were analyzed and compared between the groups statistically using SPSS software version 16.
Results: Both nitrite and asymmetric dimethylarginine levels were significantly higher in the serum of the participants from both groups than those in the PF group (P < 0.05). However, no significant difference in the asymmetric dimethylarginine level was detected between the two groups. In addition, the PF level of nitrite increased significantly in patients with endometriosis when compared with non-endometriosis subjects (P < 0.05). The PF levels of estradiol in both groups were significantly higher than the serum levels of estradiol (P < 0.05).
Conclusions: The NO metabolite level of PF implies the possible role of NO in the pathogenesis of endometriosis.
Keywords: Asymmetric dimethylarginine, endometriosis, infertility, nitric oxide, peritoneal fluid
|How to cite this article:|
Kianpour M, Nematbakhsh M, Ahmadi SM. Asymmetric dimethylarginine (ADMA), nitric oxide metabolite, and estradiol levels in serum and peritoneal fluid in women with endometriosis. Iranian J Nursing Midwifery Res 2015;20:484-9
|How to cite this URL:|
Kianpour M, Nematbakhsh M, Ahmadi SM. Asymmetric dimethylarginine (ADMA), nitric oxide metabolite, and estradiol levels in serum and peritoneal fluid in women with endometriosis. Iranian J Nursing Midwifery Res [serial online] 2015 [cited 2019 Jul 20];20:484-9. Available from: http://www.ijnmrjournal.net/text.asp?2015/20/4/484/160997
| Introduction|| |
Endometriosis is a complex disease. It is defined as the presence of functional endometrial glands and stroma outside the uterine cavity. The disease is a common gynecological problem that is often associated with pelvic pain and infertility. Many women suffering from pelvic pain and dysmenorrhea have endometriosis. Furthermore, according to laparoscopic studies, endometriosis is present in more than 50% of women with unexplained fertility. A prevalence of 2-22% of endometriosis is observed among asymptomatic women, while this incidence is 40-60% in women with dysmenorrhea. In spite of extensive studies conducted and the wide acceptance of John Sampson’s theory of retrograde menstruation, the causes of endometriosis have remained ambiguous., Moreover, endometriosis is broadly known as a pelvic inflammatory problem, which is related to distorted function of immune-related cells in the peritoneal environment. Many studies comply with this notion, proposing that the peritoneal fluid (PF) of women with endometriosis has a larger number of activated macrophages that secrete different local products, such as growth factors and cytokines, as well as additional immune mediators such as nitric oxide (NO).
NO is a known free radical that is involved in various physiological and pathophysiological processes in different organs, including the human female reproductive tract. It can directly stimulate the production of vascular endothelial growth factor (VEGF); consequently, it is involved in angiogenesis of the endometrium.
Asymmetric dimethylarginine (ADMA) is an endogenous competitive inhibitor of NO synthase (NOS). Elevated level of ADMA is related to reduced systemic NO production., Furthermore, ADMA inhibits NO production in cultured human macrophages in a concentration-dependent way. Laparoscopy is a gold standard for diagnosis of endometriosis. Yet, it is an invasive procedure, requiring surgical skill and general anesthesia, also having its own risks. In addition, significant limitations exist in the visual inspection of the pelvis, particularly in the diagnosis of retroperitoneal and deep infiltrative lesions. Thus, diagnosis of endometriosis by a simple blood test would help in overcoming these problems, while significantly protecting women’s health. In this study, we attempted to compare NO and ADMA concentrations in the PF and serum of patients with and without endometriosis.
| Materials And Methods|| |
This study was approved by the ethics committee of Isfahan University of Medical Sciences (grant code 187086). Accordingly, all patients were informed about the research protocol and all necessary information was given, and they were included in the protocol after their consent was obtained. The samples were drawn from women who were subjected to laparoscopy for evaluation of infertility or pelvic pain at the Isfahan Fertility and Infertility Center. The patients with hypertension, coronary arterial diseases, diabetes, renal diseases, active pelvic inflammatory diseases, or polycystic ovarian syndrome were excluded. As the laparoscopy was done, the patients were divided into two groups, namely, women with and without endometriosis (group I, n = 90 and group II, n = 89, respectively). Written informed consent was obtained from all the participants.
Before induction of anesthesia, venous blood samples were obtained from all the participants. The samples were centrifuged, and then the serum samples were stored at a temperature of −20ºC until measurements were carried out. Before any manipulations, the PF samples were collected from pelvis. Then, the bloody fluids were discarded and the PF samples were also centrifuged. The supernatants were stored at a temperature of −20ºC for analysis.
The serum level of estradiol was measured using an enzyme-linked immunosorbent assay (ELISA) kit (Diagnostics Biochem Canada Inc., Ontario, Canada). The serum and PF levels of nitrite (stable NO metabolite) were measured using an assay (Promega Corporation, Madison, WI, USA) that involves the Griess reaction. Briefly, after adding sulfanilamide solution and incubating the mixture, N-(1- naphtyl) ethylenediamine dihydrochloride solution was added. Then, the absorbance was measured by a microreader at a wavelength of 540 nm. The nitrite (NO metabolite) concentration of the samples was determined by comparing to the nitrite standard reference curve. The serum and PF levels of ADMA were measured using ELISA kit (DLD Diagnostika GmbH, Hamburg, Germany). Briefly, ADMA in the samples competes with solid phase-bound ADMA for a fixed number of rabbit anti-ADMA. The anti-rabbit/peroxidase was used to detect the antibody bound to the solid phase ADMA, which is inversely proportional to the ADMA concentration of serum or PF.
Data are expressed as mean ± SEM. Data were analyzed using SPSS version 16. Unpaired t-test was applied to compare the parameters between the groups. Paired t-test was used to compare the serum and PF levels of ADMA and nitrite in each group. P < 0.05 were considered statistically significant.
| Results|| |
The data for estradiol, ADMA, and NO levels in serum and PF from patients with endometriosis (case) and without endometriosis (control) are given in [Figure 1]. No significant difference was observed in estradiol levels between the groups. However, the estradiol concentration in PF was significantly higher than that in the serum (P < 0.05). Both nitrite and ADMA levels in the serum of the patients from both groups were significantly higher than those in PF (P < 0.05). However, the PF level of nitrite increased significantly in endometriosis patients when compared with non-endometriosis subjects (P < 0.05). In addition, the serum levels of ADMA and nitrite and the level of ADMA in PF were not significantly different between the two groups.
|Figure 1: ADMA, NO metabolite (nitrite), and estradiol levels in serum and PF from the patients with endometriosis (case) and —without endometriosis (control). †indicates significant difference from PF (P < 0.05) and *indicates significant difference from the other group (P < 0.05)|
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| Discussion|| |
In this study, the levels of estradiol, ADMA, and nitrite in serum and PF were compared between women with and without endometriosis. We found that both ADMA and nitrite levels in PF were lower than in serum in endometriosis and non-endometriosis subjects, and endometriosis caused an increase in the nitrite level in PF but not in serum. No significant difference was observed in estradiol levels between the groups, but the estradiol concentration was higher in PF than in serum (P < 0.05).
There are several points to discuss here. The first one is NO metabolite. It was reported that PF of endometriosis patients contains higher concentrations of NO, and that the endothelial NOS located in the endometrium produces NO., The endometrium of women with endometriosis contains a larger amount of NO, and NOS., In addition, the number of peritoneal macrophages (PMs) in the peritoneal cavity is higher than other cells. In fact, 82-99% of the PF cell population belongs to the macrophage cells. Several other studies have reported an increase in the number of total PF cells, macrophages, and also cell concentration in patients with endometriosis in comparison with the control group., Furthermore, NOS activity and expression of inducible NOS (iNOS) of the PMs have been reported to be higher in these patients, and their iNOS isoform has been reported to increase in the tissues. Endothelial NOS and iNOS, are also stimulated by the cytokines that are secreted from endometrial cells, immune cells, or macrophages to release NO at higher concentrations.,, Genetic and environmental factors, as well as immunologic alterations play a role in the development of endometriosis. The irregular immune response may cause the macrophages and/or endometrial cells to inhibit implantation and also produce elevated concentration of NO. The elevated number and activity of macrophages in endometriosis was initially considered in the concept of low-grade inflammation. This has been accompanied by liberation of some immune mediators like NO,, and extra cytokines., Since NO can directly induce VEGF production, it is involved in endometrial angiogenesis., In the same way, VEGF can stimulates the release of NO from endothelial cells. Therefore, the nitrite level in PF in endometriosis patients could be increased by several pathways, and possibly this biomarker could be considered for diagnostic purposes.
The second point to discuss is estradiol levels in serum and PF. Endometriosis is known as an estrogen-dependent inflammatory disease. Estrogen stimulates the growth of endometriotic issue in primates, including humans. Estrogens can locally originate from ovary, or can be formed from the circulating estrone sulfate, or from the inactive precursor of adrenal dehydroepiandrosterone sulfate (DHEA-S), dehydroepiandrosterone (DHEA), and androstenedione.
In women with endometriosis, estrogen is produced from three main sources in the body. First, the conversion of circulating androstenedione to estrone is catalyzed by aromatase in adipose tissue and skin, and is subsequently converted to estradiol. This estradiol and estrone enter the circulation and approach the endometriosis sites. Second, the estrogen secreted from the ovary reaches the endometriotic tissue via circulation. Also, during each ovulation, large amounts of estradiol are added to pelvic implants because of follicular rupture. The last source of estradiol is cholesterol, which is changed to estradiol in endometriosis. This conversion is due to the complete set of steroidogenic genes expressed by endometriosis tissue. Therefore, the above mechanism may involve increase in the estradiol level in PF.
Angiogenesis of the endometrium is promoted by estrogen, controlling the expression of some factors like VEGF. Inflammatory and immune responses, angiogenesis, and apoptosis in women with endometriosis are altered in favor of survival and replacement of endometriotic issue. It is also reported that the increased activity or production of numerous compounds like NO causes estrogen to have direct effects on endothelial function and vascular reactivity. Thus, it seems that in patients with endometriosis, the level of NO metabolite will be increased by different immune and inflammatory responses and estradiol increase in PF is a key factor in endometriosis that elevates the NO metabolite, as estrogen promotes the persistence and survival of endometriotic tissue. Thus, because of the enhanced levels of estrogen in endometriosis, increased level of NO metabolite is observed.
The last point to be considered with regard to ADMA is endometriosis. It is reported that by stimulation of dimethylarginine dimethyl aminohydrolase (DDAH) activity, estrogen decreases plasma ADMA, which is an endogenous competitive inhibitor of NOS. Methylation of arginine residues in intracellular proteins produces ADMA via methyltransferases. Furthermore, the ADMA metabolism is catalyzed by DDAH to dimethylamine and citrulline. The study of Holden et al. illustrated three main issues. The first issue is that circulating ADMA is decreased in women after estrogen replacement therapy. Secondly, the endothelial cells reduce the release of ADMA in vitro, a nd the final issue is that the endothelial cell DDAH enzyme activity is stimulated by estrogen. At mid cycle of premenopausal women, estrogen reaches its maximum circulating concentration and plasma NO peaks. The population in our study was in reproductive age, and aging seems to be a factor affecting the ADMA levels of plasma. Moreover, the participants were in the proliferation phase. Thus, it is probable that reduction in peritoneal levels of ADMA and increase in PF levels of NO are caused by the effects of estrogen.
| Conclusion|| |
According to the data available, a hypothesis can be proposed regarding the estrogen function and the crosstalk between inflammation and proliferation in endometriosis. The high PF levels of NO (not serum levels of NO) in endometriosis patients may signify the importance of narrow increase in macrophage activating factors. As increased levels of NO and ADMA were not observed in both groups, the disease seems not to be associated with a significant modulation in the levels of circulating NO in this age range.
| Acknowledgment|| |
This research was supported by Isfahan University of Medical Sciences (grant # 187086). The authors thanks Mrs. Ayati and Mrs. Jafarzadeh for their valuable assistance.
| References|| |
Nap AW, Groothuis PG, Demir AY, Evers JL, Dunselman GA. Pathogenesis of endometriosis. Best Pract Res Clin Obstet Gynaecol 2004;18:233-44.
Mahmood TA, Templeton A. Pathophysiology of mild endometriosis: Review of literature. Hum Reprod 1990;5:765-84.
Burns WN, Schenken RS. Pathophysiology of endometriosis-associated infertility. Clin Obstet Gynecol 1999;42:586-610.
Child TJ, Tan SL. Endometriosis: Aetiology, pathogenesis and treatment. Drugs 2001;61:1735-50.
Hill JA. Immunology and endometriosis. Fertil Steril 1992;58:262-4.
Farquhar CM. Extracts from the “clinical evidence”. Endometriosis. BMJ 2000;320:1449-52.
Ishimura T, Masuzaki H. Peritoneal endometriosis: Endometrial tissue implantation as its primary etiologic mechanism. Am J Obstet Gynecol 1991;165:210-4.
Badawy SZ, Cuenca V, Marshall L, Munchback R, Rinas AC, Coble DA. Cellular components in peritoneal fluid in infertile patients with and without endometriosis. Fertil Steril 1984;42:704-8.
Bedaiwy MA, Falcone T. Peritoneal fluid environment in endometriosis. Clinicopathological implications. Minerva Ginecol 2003;55:333-45.
Ho HN, Wu MY, Yang YS. Peritoneal cellular immunity and endometriosis. Am J Reprod Immunol 1997;38:400-12.
Oral E, Olive DL, Arici A. The peritoneal environment in endometriosis. Hum Reprod Update 1996;2:385-98.
Akoum A, Kong J, Metz C, Beaumont MC. Spontaneous and stimulated secretion of monocyte chemotactic protein-1 and macrophage migration inhibitory factor by peritoneal macrophages in women with and without endometriosis. Fertil Steril 2002;77:989-94.
Arici A, Oral E, Attar E, Tazuke SI, Olive DL. Monocyte chemotactic protein-1 concentration in peritoneal fluid of women with endometriosis and its modulation of expression in mesothelial cells. Fertil Steril 1997;67:1065-72.
Cao X, Yang D, Song M, Murphy A, Parthasarathy S. The presence of endometrial cells in the peritoneal cavity enhances monocyte recruitment and induces inflammatory cytokines in mice: implications for endometriosis. Fertil Steril 2004;82(Suppl 3):999-1007.
Rong R, Ramachandran S, Santanam N, Murphy AA, Parthasarathy S. Induction of monocyte chemotactic protein-1 in peritoneal mesothelial and endometrial cells by oxidized low-density lipoprotein and peritoneal fluid from women with endometriosis. Fertil Steril 2002;78:843-8.
Anggard E. Nitric oxide: Mediator, murderer, and medicine. Lancet 1994;343:1199-206.
Dong M, Shi Y, Cheng Q, Hao M. Increased nitric oxide in peritoneal fluid from women with idiopathic infertility and endometriosis. J Reprod Med 2001;46:887-91.
Luo Q, Dong LJ, Huang HF. Increased nitric oxide levels in peritoneal fluids of minor-endometriosis patients and its relation to IVF-ET outcomes](abstract). Zhejiang Da Xue Xue Bao Yi Xue Ban 2007;36:424-8.
Rosselli M, Keller PJ, Dubey RK. Role of nitric oxide in the biology, physiology and pathophysiology of reproduction. Hum Reprod Update 1998;4:3-24.
Fukumura D, Gohongi T, Kadambi A, Izumi Y, Ang J, Yun CO, et al
. Predominant role of endothelial nitric oxide synthase in vascular endothelial growth factor-induced angiogenesis and vascular permeability. Proc Natl Acad Sci U S A 2001;98:2604-9.
Cevik D, Unay O, Durmusoglu F, Yurdun T, Bilsel AS. Plasma markers of NO synthase activity in women after ovarian hyperstimulation: Influence of estradiol on ADMA. Vasc Med 2006;11:7-12.
Boger RH, Bode-Boger SM, Thiele W, Junker W, Alexander K, Frolich JC. Biochemical evidence for impaired nitric oxide synthesis in patients with peripheral arterial occlusive disease. Circulation 1997;95:2068-74.
Boger RH, Bode-Boger SM, Szuba A, Tsao PS, Chan JR, Tangphao O, et al
. Asymmetric dimethylarginine (ADMA): A novel risk factor for endothelial dysfunction: Its role in hypercholesterolemia. Circulation 1998;98:1842-7.
Fickling SA, Williams D, Vallance P, Nussey SS, Whitley GS. Plasma concentrations of endogenous inhibitor of nitric oxide synthesis in normal pregnancy and pre-eclampsia. Lancet 1993;342:242-3.
Brosens IA, Brosens JJ. Is laparoscopy the gold standard for the diagnosis of endometriosis? Eur J Obstet Gynecol Reprod Biol 2000;88:117-9.
Brosens I, Puttemans P, Campo R, Gordts S, Brosens J. Non-invasive methods of diagnosis of endometriosis. Curr Opin Obstet Gynecol 2003;15:519-22.
Othman Eel D, Hornung D, Salem HT, Khalifa EA, El-Metwally TH, Al-Hendy A. Serum cytokines as biomarkers for nonsurgical prediction of endometriosis. Eur J Obstet Gynecol Reprod Biol 2008;137:240-6.
Osborn BH, Haney AF, Misukonis MA, Weinberg JB. Inducible nitric oxide synthase expression by peritoneal macrophages in endometriosis-associated infertility. Fertil Steril 2002;77:46-51.
Cameron IT, Campbell S. Nitric oxide in the endometrium. Hum Reprod Update 1998;4:565-9.
Omland AK, Tanbo T, Dale PO, Abyholm T. Artificial insemination by husband in unexplained infertility compared with infertility associated with peritoneal endometriosis. Hum Reprod 1998;13:2602-5.
Khorram O, Lessey BA. Alterations in expression of endometrial endothelial nitric oxide synthase and αvβ3 integrin in women with endometriosis. Fertil Steril 2002;78:860-4.
Wu MY, Ho HN. The role of cytokines in endometriosis. Am J Reprod Immunol 2003;49:285-96.
Dunselman GA, Hendrix MG, Bouckaert PX, Evers JL. Functional aspects of peritoneal macrophages in endometriosis of women. J Reprod Fertil 1988;82:707-10.
Eischen A, Duclos B, Schmitt Goguel M, Rouyer N, Bergerat JP, Hummel M, et al
. Human resident peritoneal macrophages: Phenotype and biology. Br J Haematol 1994;88:712-22.
Halme J, Becker S, Hammond MG, Raj S. Pelvic macrophages in normal and infertile women: The role of patent tubes. Am J Obstet Gynecol 1982;142:890-5.
Haney AF, Muscato JJ, Weinberg JB. Peritoneal fluid cell populations in infertility patients. Fertil Steril 1981;35:696-8.
Olive DL, Weinberg JB, Haney AF. Peritoneal macrophages andinfertility: The association between cell number and pelvicpathology. Fertil Steril 1985;44:772-7.
Gupta S, Agarwal A, krajcir N, Alvarez JG. Role of oxidative stress in endometriosis. Reprod Biomed 2006;17:126-34.
Tschugguel W, Schneeberger C, Unfried G, Brautigam G, Stonek F, Wieser F, et al
. Elevation of inducible nitric oxide synthase activity in human dring menstruation. Biol Reprod 1999;60:297-304.
Shaamash AH, Zakhari MM. Increased serum levels of nitric oxide metabolites among users: A possible role in progestin -induced bleeding. Hum Reprod 2005;20:302-6.
Nuojua-Huttunen S, Tomas C, Bloigu R, Tuomivaara L, Martikainen H. Intrauterine insemination treatment in subfertility: An analysis of factors affecting outcome. Hum Reprod 1999;14:698-703.
Van Langendonckt A, Casanas-Roux F, Donnez J. Oxidative stress and peritoneal endometriosis. Fertil Steril 2002;77:861-70.
Ota H, Igarashi S, Hatazawa J, Tanaka T. Endothelial nitric oxide synthase in the endomertium during the menstrual cycle in patients with endometriosis and adenomyosis. Fertil Steril 1998;69:303-8.
Yang WC, Chen HW, Au HK, Chang CW, Huang CT, Yen YH, et al
. Serum and endometrial markers. Best Pract Res Clin Obstet Gynaecol 2004;18:305-18.
Kim KH, Oh DS, Jeong JH, Shin BS, Joo BS, Lee KS. Follicular blood flow is a better predictor of the outcome of in vitro
fertilization-embryo transfer than follicular fl uid vascular endothelial growth factor and nitric oxide concentrations. Fertil Steril 2004;82:586-92.
Gupta S, Goldberg JM, Aziz N, Goldberg E, Krajcir N, Agarwal A. Pathogenic mechanisms in endometriosis-associated infertility. Fertil Steril 2008;90:247-57.
Grasselli F, Basini G, Bussolati S, Tamanini C. Effects of VEGF and bFGF on proliferation and production of steroids and nitric oxide in porcine granulosa cells. Reprod Domest Anim 2002;37:362-8.
Giudice LC, Kao LC. Endometriosis. Lancet 2004;364:1789-99.
Bulun SE. Endometriosis. N Engl J Med 2009;360:268-79.
Rizner TL. Estrogen metabolism and action in endometriosis: Mol Cell Endocrinol 2009;307:8-18.
Bulun SE, Lin Z, Imir G, Amin S, Demura M, Yilmaz B, et al
. Regulation of aromatase expression in estrogenresponsive breast and uterine disease: From bench to treatment. Pharmacol Rev 2005;57:359-83.
Bulun SE, Cheng YH, Yin P, Imir G, Utsnuomiya H, Attar E, et al
. Progesterone resistance in endometriosis: Link to failure to metabolize estradiol. Mol Cell Endocrinol 2006;248:94-103.
Albrecht ED, Babishkin JS, Lidor Y, Anderson LD, Udoff LC, Pepe GJ. Effect of estrogen on angiogenesis in co-culture of human endometrial cells and microvascular endothelial cells. Hum Reprod 2003;18:2039-47.
Dmowski WP, Gebel HM, Rawlins RG. Immunologic aspects of endometriosis. Obstet Gynecol Clin North Am 1989;16:93-103.
Osteen KG, Sierra-Rivera E. Does disruption of immune and endocrine systems by environmental toxins contribute to development of endometriosis? Semin Reprod Endocrinol 1997;15:301-8.
Taylor RN, Lebovic DI, Mueller MD. Angiogenic factors in endometriosis. Ann N Y Acad Sci 2002;955:89-100.
Dmowski WP, Ding J, Shen J, Rana N, Fernandez BB, Braun DP. Apoptosis in endometrial glandular and stromal cells in women with and without endometriosis. Hum Reprod 2001;16:1802-8.
Beliard A, Noel A, Foidart JM. Reduction of apoptosis and proliferation in endometriosis.Fertil Steril 2004;82:80-5.
Holden DP, Cartwright JE, Nussey SS. Whitley GS. Estrogen stimulates dimethylarginineaminohydolase activity and the metabolism of asymmetric Dimethylarginine Circulation 2003;108:1575-80.
Moncada S, Higgs A. The l-arginine-nitric oxide pathway. N Engl J Med 1993;329:2002-12.
Matsuoka H, Itoh S, Kimoto M, Kohno K, Tamai O, Wada Y, et al
. Asymmetrical dimethylarginine, an endogenous nitric oxide synthase inhibitor, in experimental hypertension. Hypertension 1997;29:242-7.
Miyazaki H, Matsuoka H, Cooke JP, Usui M, Ueda S, Okuda S, et al
. Endogenous nitric oxidesynthase inhibitor: A novel marker of atherosclerosis. Circulation 1999;99:1141-6.
Leiper J, Nandi M, Torondel B, Murray-Rust J, Malaki M, O’Hara B, et al
. Disruption of Methylarginine impairs vascular homeostasis. Nat Med 2007;13:198-203.
Palomo I, Contreras A, Alarcón LM, Leiva E, Guzmán L, Mujica V, et al
. Elevated concentration of asymmetric dimethylarginine (ADMA) in individuals with metabolic syndrome. Nitric Oxide 2011;24:224-8.
Rosselli M, Imthurm B, Macas E, Keller PJ, Dubu RK, Circulating nitrite/nitrate levels increase with follicular development: Indirect evidence for estradiol mediated NO release. Biochem Biophys Res Commun 1994;202:1543-52.