Relaxin and Nitric Oxide Signalling - IngentaConnect

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Current Protein and Peptide Science, 2008, 9, 638-645

Relaxin and Nitric Oxide Signalling M.C. Baccari1,* and D. Bani2 1

Departments of Physiological Sciences and 2Anatomy, Histology and Forensic Medicine, University of Florence, Florence, Italy Abstract: The peptide hormone relaxin (RLX) has been shown to exert a variety of functions in both reproductive and non-reproductive tissues. The molecular mechanism of RLX on its target cells appears to involve multiple intracellular signalling systems, including the nitric oxide (NO) pathway. NO is an ubiquitous molecule synthesised from L-arginine under the catalytic action of different nitric oxide synthase (NOS) isoforms and its altered production has been reported to be involved in several diseases. RLX has been demonstrated to promote NO biosynthesis by up-regulating NOS expression; its influence on the different NOS appears to depend on the cell type studied. In addition to its physiological roles, RLX has been postulated as a potential therapeutic agent in several diseases. In particular, based on its property to promote NO biosynthesis, RLX may be regarded as a therapeutic tool in diseases characterized pathogenically by an impaired NO production. The aim of the present mini-review is to summarize and discuss the pathophysiological actions of RLX, strictly related to its ability to activate the endogenous NO pathway in reproductive and non-reproductive target organs.

Keywords: Relaxin, nitric oxide. INTRODUCTION Relaxin (RLX), identified in 1926 as a peptide hormone [1], has been long considered pertinent to reproduction, based on early studies demonstrating its ability to promote elongation of the interpubic ligament in mammals [2], cervical softening [3], growth of the mammary gland [4] and to inhibit spontaneous contractions of the myometrium [5]. The finding that RLX was primarily produced in the pregnant ovary and the placenta further confirmed its role as a reproductive hormone. Major steps in the knowledge of RLX physiology were the isolation and purification of RLXs from different species [6,7], that allowed identification of their amino-acid sequences [8-10], and the set-up of specific radioimmunoassay methods for RLX dosage in biological samples [11-13]. Identification of RLX binding sites [14,15] and, more recently, of RLX genes in reproductive as well as non-reproductive organs and tissues [reviewed in 16] has led to a revision of the former concept of RLX as a reproductive hormone, opening the possibility that this hormone could exert a broad range of biological effects on many organs and apparatuses, such as the central nervous system, the gastrointestinal tract, the cardiovascular system, the kidney, the lungs, etc. [16-22]. It is a tempting hypothesis that these effects of RLX may also contribute to the tuning of extrareproductive organs to the physiological needs of pregnancy. Formerly included in the insulin superfamily based on structure similarities, RLX has been recently re-assigned to a distinct hormone family, the RLX peptide family that, in humans, encompasses 3 RLX isoforms and 4 insulin like peptides (INSLs) [16,23,24]. In fact, the molecular interaction of RLX with its target cells is different from that of in*Address correspondence to this author at the Department of Physiological Sciences, University of Florence, Viale G.B. Morgagni 63, I-50134 Florence, Italy; Tel: (+39) 055-4237339; Fax: (+39) 055-4379506; E-mail: [email protected] 1389-2037/08 $55.00+.00

sulin, whose receptors are membrane-associated tyrosine kinases, and involves multiple intracellular signalling systems, including cAMP [17, 25-34]. In 2002, upon an accurate screening of a human and mouse DNA library of genes encoding for orphan receptors, Hsu and co-workers eventually identified two surface G protein-coupled receptors that fulfil the requirements for RLX receptors, i.e. the ability to mediate the action of RLX through a cAMP-dependent pathway distinct from that of insulin and IGFs [35]. These molecules were then termed relaxin family peptide receptors (RXFP) 1 & 2, RXFP1 being the most specific receptor for circulating RLX [36]. Therefore RLX may induce elevation of cAMP in cells and tissues by activation of adenylate cyclase through Gs proteins [35]. Nevertheless, elevation of cAMP levels may also involve tyrosine kinase activation: in human endometrial stromal cells and in monocytic cell line THP-1, Ivell and co-workers demonstrated that RLXinduced activation of G protein-coupled receptor leads to tyrosine phosphorylation which, in turn, inhibits phosphodiesterase activity and further up-regulates cAMP levels [27, 37-39]. In more recent years, other receptors of the RLX family peptides have been identified and localized in reproductive and non-reproductive organs [36]. An additional intracellular signalling system shown to be targeted by RLX is guanylate cyclase. In fact, increased cGMP levels have been found in different cell types [40-43] in response to RLX. This effect results from the activation by RLX of the endogenous nitric oxide (NO) pathway, first described in isolated rat serosal mast cells [44]. NO is synthesized from the amino acid L-arginine under the catalytic action of NO synthases (NOS), with L-citrulline as a co-product, by almost all mammalian cells [45, 46]. Three major NOS isoforms are usually expressed by different cell types: the constitutive isoforms, endothelial NOS (eNOS or NOS III) and neuronal NOS (nNOS or NOS I) and © 2008 Bentham Science Publishers Ltd.

Relaxin and Nitric Oxide Signalling

the inducible NOS (iNOS or NOS II). The latter one can be either expressed constitutively or induced by a variety of stimuli, including pro-inflammatory mediators and hormones. NO is a ubiquitous autacoid molecule involved in the regulation of many physiological processes; therefore, its altered production, in excess or defect, has been related to the pathophysiology of several diseases [47]. There is convincing evidence that NOS expression is up-regulated by RLX in several target cells and organs with either reproductive or non-reproductive functions. Thus, the widespread actions of RLX are not so surprising taking into account the ubiquitous distribution of NO in mammalian cells. One of the most interesting features is that the influence of RLX on the different NO synthases appears to depend on the cell type studied [42, 48-50]. In this view, two possible pathways by which RLX-activated G protein-coupled receptors can stimulate NO biosynthesis have been suggested [51]: activation of eNOS by a mechanism involving phosphoinositide 3kinase/protein kinase B and induction of iNOS expression mediated by cAMP elevation. It has been also suggested that RLX may indirectly activate the NO pathway through the stimulation of extracellular matrix remodelling. In fact, RLX-induced collagenase activation could convert big endothelin (ET)-1 into bioactive ET1-32, which in turn binds to ETB receptors thereby inducing eNOS activation and NO release [18]. In addition to its physiological roles, RLX has been postulated as a potential therapeutic agent in several diseases affecting both reproductive and non-reproductive systems, mainly emerging from its antifibrotic and anti-inflammatory actions, as well as its ability to promote vasodilatation and angiogenesis [17, 19-21, 24, 52-57]. This article aims at reviewing and discussing the physiopathological effects of RLX strictly related to the activation by this hormone of the endogenous NO pathway in the reproductive and non-reproductive systems, as summarized in Table 1. RLX AND NO SIGNALLING IN THE REPRODUCTIVE SYSTEM RLX received its name owing to its firstly described effect, i.e. the marked lengthening of the guinea pig interpubic ligament and softening of the tissues of the birth canal aimed at facilitating delivery of the fetus [1, 3]. Later studies in animals evidenced that, during pregnancy, RLX causes uterine muscle quiescience and promotes the remodelling of the connective tissue extracellular matrix of the organs of the reproductive tract; these effects are probably aimed at favouring accommodation of the growing fetus [58, 59]. The mechanisms that regulate softening of the uterine cervix during pregnancy have been widely investigated: in particular, clinical studies have suggested an involvement of NO in cervical ripening [60] especially during late pregnancy [61]. In the rat, NO synthesis inhibition by means of L-nitroarginine methyl ester (L-NAME) throughout the last 6 days of pregnancy prolonged the duration of delivery [62]. When cervices were removed on day 20 of gestation and incubated overnight with the NO synthesis inhibitor LNAME, they were less extensible than the untreated controls [62]. The involvement of NO in cervical ripening is further

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supported from the observation that increased expression of all three NOS isoforms has been found in the rat cervix during pregnancy as compared with the non-pregnant state [62, 63]. However, the possibility that the effects of RLX on cervical softening may be mediated - at least in part - through NO, is matter of controversy. This view is indeed supported by the studies of Shi and co-workers on pregnant rats, in which administration of NOS inhibitors such as L-NAME significantly blunted RLX-induced cervical softening in respect to the controls [64]. On the other hand, NOindependent softening of the rat cervix upon acute RLX administration has been also reported [65]. To explain these conflicting results in the same animal model, these authors have postulated a two-stage process of cervical softening, with NO being only involved in late pregnancy. Accordingly, in the rat cervix, the levels of iNOS protein and mRNA increase markedly at term pregnancy and during labor, i.e., during the ultimate stage of cervical softening [62, 63]. NOS activation also appears to be required for RLX in order to increase wet weight in the rat cervix [65]: in this study, administration of L-NAME has been reported to reduce this typical effect of RLX, which could be mediated through eNOS. Increased expression of eNOS has been also shown in surface epithelium, glands, endometrial stromal cells, and myometrium of the uterus of mice upon systemic RLX treatment (Fig. 1). The related increase in endogenous NO appears to contribute to the property of RLX to inhibit myometrial contractility: the direct muscular relaxant responses to RLX, recorded in strips from mouse uteri were blunted by the NO synthesis inhibitor L-N G-nitro arginine (L-NNA) [49]. A decrease of frequency and amplitude of spontaneous or evoked (chemically or electrically induced) myometrial contractions by RLX has been reported in isolated preparations from various animal species [66-69]. In some cases, the RLX effect was reduced by NO synthesis inhibitors, thus indicating the involvement of the L-arginine-NO pathway in the relaxant action of this hormone on the myometrium. Thus, the presence of a RLX-driven NO pathway in the uterus appears to be involved in cervical softening and growth as well as uterine muscle quiescience, conceivably required to facilitate accommodation of the growing fetus. In humans, however, RLX appears to be of little importance to this regard, as it shows negligible effects on myometrial quiescence [20]. RLX has been reported to have mammotrophic properties in rodents [4, 70]. In particular, systemic administration of RLX in combination with estrogens promoted growth and differentiation of the mammary parenchyma and stroma [71]. RLX was also shown to influence the behaviour of human breast cancer cells in vitro [41]: in hormone-dependent MCF-7 breast carcinoma cells, RLX, at proper concentrations and exposure times, has growth-inhibiting and differentiation-promoting effects, which are mediated through the activation of the NO pathway. In fact, RLX increased iNOS activity in the whole range of concentrations employed (nmol-μmol/L), whereas it showed a biphasic effect on constitutive eNOS, which was stimulated at the lower concentrations and inhibited at the higher ones. Thus, endogenous NO generation seems to be involved in the regulation of mam-

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Table 1.

Baccari and Bani

Modulation of NO Generation by RLX in Different Targets

Organs, tissues, cells

Species

Model

Evidence for NO production

NO synthase up-regulation nNOS eNOS iNOS

Biological effects

Ref.

Uterus (endometrium, myometrium)

mouse

in vivo

NOS inhibition (L-NNA)

protein

myometrial quiescence

[49]

Uterine cervix

mouse rat

in vivo

NOS inhibition (L-NAME)

protein mRNA

ripening, increased weight

[64] [65]

Mammary cancer MCF-7 cells

human

in vitro

NOS activity (nitrites, 3H-citrulline)

protein

growth inhibition, differentiation

[41]

Kidney arteries

rat

ex vivo in vivo

NOS inhibition (L-NAME)

vasodilatation, increased hyperfiltration

[80] [103]

Ileum

mouse

ex vivo in vivo


decreased motility, normalization of hypermotility

[115] [124]

Stomach

mouse

ex vivo in vivo


decreased motility

[50] [118]

Aortic smooth muscle cells

ox

in vitro

NOS activity (nitrites, cGMP)

protein

relaxation

[42]

Coronary endothelial cells

rat guinea pig

ex vivo in vitro

NOS inhibition (L-NMMA) NOS activity (nitrites, cGMP)

protein mRNA

vasorelaxation, reduced cell adhesion

[48] [85] [93]

Umbilical vein endothelial cells

human

in vitro

NOS activity (nitrites)

protein mRNA

vasorelaxation

[88]

Neutrophils

human

in vitro

NOS inhibition (L-NMMA) NOS activity (nitrites)

protein

functional inhibition, decreased chemotaxis

[94]

Basophils

human

in vitro

NOS inhibition (L-NMMA) NOS activity (3H-citrulline)

functional inhibition, decreased histamine release

[89]

Platelets

human rabbit

in vitro

NOS inhibition (L-NMMA) NOS activity (nitrites, cGMP)

functional inhibition

[40] [95]

Mast cells

rat guinea pig

in vitro

NOS inhibition (L-NMMA, L-NAME)

decreased histamine release

[44]

mary cancer cell growth and differentiation. On the other hand, whether the NO pathway plays a similar functional role in the normal mammary gland remains largely to be defined. For instance, the three isoforms of NOS are constitutively present in the lactating mammary gland; however, while iNOS expression and, consequently, NO production are increased during weaning, eNOS is diminished [72]. Clearly, further studies are needed to clarify this issue.

protein

protein

protein

protein

RLX AND NO SIGNALLING IN NON-REPRODUCTIVE SYSTEMS Cardiovascular System The notion that RLX is a cardiovascular hormone dates back to the late 1950s, when an impure porcine RLX preparation, injected to patients suffering for peripheral vascular diseases and Raynaud’s syndrome, was shown to cause a dramatic, albeit transient, amelioration of symptoms and



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trations which are in the nanomolar range, similar to the RLX blood levels of normal human pregnancy [87]. RLX is extremely potent as a vasorelaxant: in the isolated, perfused rat and guinea pig heart, the dose-dependent increase in coronary flow induced by RLX is significantly higher than that obtained with similar doses of typical vasodilatatory agents such as acetylcholine or sodium nitroprusside [85]. This latter concept indirectly suggested that these effects of RLX would be mediated by mechanisms involving a potent vascular effector and, once again, NO appeared as the perfect candidate. Sound experimental evidence exists that the vasoactive action of RLX can be chiefly ascribed to the stimulation of NO biosynthesis by blood vessel cells. In vitro studies have shown that addition of RLX to the culture medium of endothelial cells from rat coronaries and human umbilical vein and of bovine artery smooth muscle cells causes a dose-dependent increase in the production of NO and in the intracellular levels of cGMP [42, 48, 88], which mediates the cell response to NO. This results in a decrease in cytosolic Ca2+ that leads to de-activation of contractile filaments and cell relaxation [42]. On the other hand, the RLX-induced vasodilatation does not depend on increased histamine release, as mast cells and basophils, which are the main tissue and circulating repository of histamine, respectively, are rather inhibited by RLX, once again through a NO-mediated mechanism [44, 89].

Fig. (1). Uterotropic effects of RLX. Representative images of the uteri (insets) and relevant histological sections of sexually mature, ovariectomized mice treated with subcutaneous injections of estrogens (E2: 5 μg 17-estradiol for 7 days) or estrogens plus RLX (E2+RLX: 5 μg 17-estradiol for 7 days plus 2 μg pure porcine RLX for the last 24 hrs.). RLX, besides increasing the overall size of the uterus due to both growth and interstitial fluid accumulation of the uterine tissues (see insets), also enhances eNOS immunoreactivity, particularly in surface epithelium, glands and the outer, longitudinal myometrium. Bars = 200 μm.

signs of ischemia [73, 74]. In some patients who also suffered for ischemic heart disease, RLX treatment led to the reduction of the daily glyceryl trinitrate requirements. Retrospectively, this important observation clearly suggests that RLX has a direct, dilatory effect on peripheral and coronary vasculature. Recognition of the blood vessels as specific RLX targets came from the identification of RLX binding sites/receptors on cells of the vascular wall [15, 35, 75 ]. Vasodilatation is a specific effect of RLX and has been observed in many target organs and tissues, in both females and males. Briefly, RLX promotes vasodilatation in reproductive organs, such as the uterus [76, 77] and the mammary gland [78], as well as in non-reproductive targets, including mesocaecum [79], kidney [80-82], liver [83], lung [84] and heart [85,86]. Vasodilatation appears as a physiological effect of RLX since it is fully manifested at hormone concen-

It is well known that, in endothelial cells, eNOS can be stimulated by vasoactive agonists and hormones, such as acetylcholine, bradykinin and estrogens, acting through G protein-coupled receptors [90-92], similarly to the RLXRXFP1 signaling complex. On the other hand, studies on vascular endothelial and smooth muscle cells suggest that RLX can promote vascular NO generation by an alternate pathway to the constitutive eNOS, i.e. by up-regulating the expression of iNOS [42, 48, 88]. From a pathophysiological perspective, sustained vascular NO generation by RLX can counteract vasoconstriction as well as leukocyte and platelet adhesion and vascular inflammation, which are the major causes of atherogenesis. Recently, it has been demonstrated that RLX can reduce endothelial adhesiveness to neutrophils in pro-inflammatory conditions in vitro by down-regulating endothelial cell adhesion molecules [93]. Besides vascular cells, RLX can directly influence other cells involved in vascular dysfunction and atherosclerosis, i.e. leukocytes and platelets. In fact, once again by a NO-dependent mechanism, RLX has been shown to strongly inhibit the activation of neutrophils induced by pro-inflammatory mediators, thereby reducing reactive oxygen species (ROS) generation and chemotaxis [94], and to dose-dependently reduce the aggregation of platelets [40, 95]. As a whole, these findings afford RLX to be considered a true vascular hormone and physiological blood vessel effector. Its functional role in reproductive physiology could be to participate in the cardiovascular adjustments of pregnancy needed to sustain utero-placental blood flow demand. On a broader perspective, based on its NO-dependent protective effects against hypertension and atherogenesis, RLX could be viewed as a natural defence against cardiovascular disease, especially in women during fertile life. As a matter of fact, fertile women have better endothelial function, vascular reactivity and higher NO biosynthesis than age-matched men

642 Current Protein and Peptide Science, 2008, Vol. 9, No. 6

[96]. Furthermore, epidemiologic studies indicate that the incidence of cardiovascular disease in women is very low until menopause and increases sharply thereafter [97, 98], concurrently with cessation of ovarian activity and hence of RLX secretion in blood. Kidney The identification of RLX and RLX-3 gene transcripts by RT-PCR in the rodent medulla and cortex of the kidney [99], later confirmed by immunohistochemistry [56] strongly suggests that the kidney is a potential source and/or target organ for RLX. In conscious rats, peak renal vasodilatation and hyperfiltration occur at mid-gestation, thus suggesting a major role of the pregnancy hormones in modulating renal circulation dynamics [100]. In this context, Danielson and co-workers showed that prolonged administration of porcine or recombinant human RLX to conscious rats increased effective renal plasma flow and glomerular filtration rate (GFR) to the levels observed during mid-term pregnancy [80]. The response to recombinant human RLX was not genderdependent, as it was later confirmed in male animals [81]. Chronic administration of porcine RLX diminished the renal vasoconstrictor response to angiotensin II infusion [80], another typical phenomenon occurring in rat gestation [101, 102]. More recently, myogenic reactivity of small renal arteries isolated from rats treated with recombinant human RLX was found reduced [103] to a comparable extent as that observed in small renal arteries isolated from mid-term pregnant rats [104]. The essential role played by RLX in renal circulation also comes from the observation that all the above-mentioned effects (renal vasodilatation, hyperfiltration and reduced myogenic reactivity of small renal arteries) were abolished by neutralizing anti-RLX antibodies or by removing the ovaries, the main source of circulating RLX, in mid-term pregnant rats [82]. In RLX-pretreated non-pregnant rats, RLX has been shown to stimulate renal vasodilatation and hyperfiltration [57, 105, 106] likely by increasing NO production. In fact, blockade of the renal effects of RLX by NOS inhibitors strongly accounts for a NO-dependent mechanism [80, 103]. Furthermore, acute administration of NOS inhibitors to chronically instrumented conscious mid-term pregnant rats decreased GFR and effective renal plasma flow, and caused a rise in effective renal vascular resistance [102]. In keeping with these in vivo observations, myogenic reactivity of small renal arteries isolated from mid-term pregnant rats was reduced in comparison with control virgin animals, and NOS inhibitors were able to bring the myogenic reactivity back to the control levels [104]. These studies support the view of an essential role for NO in pregnancy- and RLX-induced adaptations of renal small arteries. In the kidney, the vasodilator action of RLX may also occur through activation of endothelin (ET)B receptor by ET1-32, this latter generated in the extracellular mileu upon RLX-induced up-regulation of collagenases [18]. In turn, ETB can lead to increased NO production [57]. NO could also contribute to the pro-angiogenic effects of RLX: of note, substantial loss of blood capillaries parallels the evolution of progressive fibrosis in renal disease [107]. In this view, re-

Baccari and Bani

cent in vivo studies have shown that exogenous RLX is able to slow down the progression of renal fibrosis in experimental rodent models [99, 108-111]. Gastrointestinal Tract It is widely reported in the basic and clinical literature that sex female hormones, such as estrogen and progesterone, are involved in gastrointestinal motor disturbances [112]. In addition to ovarian steroids, RLX also appears to influence gut motility. Early reports have indicated that a purified ovarian RLX preparation reduces the strength and frequency of contractions in the isolated rat ileum [113] and that RLX had disruptive effects on the migrating myoelectric complex of the rat small intestine in vivo [114]. These findings have been confirmed and extended by recent functional studies on mouse ileal preparations using highly purified porcine RLX. In these studies, RLX was shown to depress ileal spontaneous contractions by a direct action on smooth muscle [115]. The physiological relevance of the above described effects of RLX on small bowel motility remains matter of speculation: it is conceivable that RLX could contribute to increase transit time of ingesta, thereby facilitating digestion and absorption of food nutrients to fulfil the increased needs of the mother and the growing fetus. A major step in the knowledge on the mechanism of action of RLX on gastrointestinal motility came from the observation that, in ex vivo preparations from mouse ileum, NOS inhibitors reversed the effects of the hormone [115], indicating the involvement of endogenous NO biosynthesis in the RLX effect. Evaluation of NOS expression by immunohistochemistry and Western Blot analysis confirmed that an 18-h pre-treatment of intact mice of both genders with systemically administered RLX resulted in an overexpression of iNOS and eNOS by the ileal smooth muscle coat. NO has been demonstrated to play an important role in the inhibitory regulation of gut motility. NO, besides being generated from smooth muscle cells of the gastrointestinal tract, is also considered the main inhibitory neurotransmitter released from non-adrenergic, non-cholinergic (NANC) nervous fibres [116]. Gastrointestinal motor responses are indeed controlled by myogenic, nervous (excitatory and inhibitory) and hormonal mechanisms [117]. On these grounds, the influence of RLX was also tested on neurallyinduced excitatory and inhibitory responses elicited in strips from the mouse gastric fundus [50, 118]. Functional studies confirmed the results obtained in the ileum: RLX, through a NO-mediated mechanism, was able to depress the amplitude of the neurally-induced contractile responses and to increase the amplitude of the inhibitory ones. At variance with the ileum, evaluation of NOS expression showed an increase in both nNOS and eNOS. These findings allowed us to conclude that RLX can act on the inhibitory neural control of the stomach by up-regulating NO biosynthesis at the nervous level. Thus, in the gastrointestinal tract, RLX can depress either myogenic contractions, as occurs in the ileum, or neurally-induced responses, as it does in the stomach. Accordingly, in strips dissected from pregnant rats, a reduction of gastric motility strictly related to an increased NO production release from NANC nerves was observed, whereas such


mechanism is not operating in the ileum [119]. From a physiological perspective, an increased adaptive fundic relaxation and a decrease of the contractile responses of the stomach fit well with the delayed gastric emptying often reported to occur during pregnancy. Thus, we can tentatively speculate that RLX, by up-regulating NO production, could be involved in pregnancy-related reduction of gastric motility. Although excess NO has been reported to be involved in gastrointestinal motor disorders, the same also occurs for defective NO generation. In this view, an altered production/release of endogenous NO has been reported to occur in some inherited or acquired gastrointestinal motor dysfunctions [120]. In particular, impaired NO production and release has been related to gastrointestinal dysmotilities observed in ex vivo preparations from mice with muscular distrophy due to mutated dystrophin gene (mdx mice) [121123]. Based on the above ability of RLX to up-regulate gastrointestinal NO biosynthesis, we were intrigued by the possibility to counteract the motor dysfunction of the ileum in the mdx mice by therapeutic administration of RLX. For this purpose, the effects of RLX were tested on the abnormally elevated spontaneous contractions observed in ileal preparations from male mdx mice. Direct exposure of the ileal strips to RLX, as well as a 18-h systemic pre-treatment of the mdx mice with the hormone, greatly reduced the amplitude of the spontaneous contractions, shifting the abnormal motility pattern of the mdx mice close to that of the normal control mice. The reversal of the effects of RLX by the NOS inhibitor NG-nitro-L-arginine or the guanylate cyclase inhibitor ODQ, indicated the involvement of NO. Evaluation of the expression of NOS isoforms by immunohistochemistry and Western blot strongly supported the functional data: in fact, increased levels of iNOS in the ileal muscle coat of the RLX-pretreated mdx mice was observed as compared with the vehicle-pretreated, control mdx animals [124]. These findings provide support to the concept that RLX, owing to its ability to up-regulate the intrinsic NO biosynthetic machinery in the gastrointestinal tract, could represent an useful tool in the treatment of motor abnormalities related to defective NO production.


NO

= Nitric oxide

NOS

= Nitric oxide synthase

RLX

= Relaxin

ROS

= Reactive oxygen species

RXFP = Relaxin family peptide receptors VEGF = Vascular endothelial growth factor REFERENCES [1] [2] [3] [4] [5] [6] [7] [8] [9] [10] [11] [12] [13] [14] [15] [16] [17] [18] [19] [20] [21] [22] [23] [24]

CONCLUSIONS In this mini-review we have emphasized the role played by RLX, acting through activation of the NO pathway, in the pathophysiology of reproductive and non-reproductive systems. Taken together, the data available in the literature converge to suggest that some of the reported effects of RLX on its target organs could be ascribable to its ability to upregulate NOS expression in several target cells and organs. This observation provides background to the challenging hypothesis that RLX could be a potential therapeutic tool in NO deficiency-related diseases, an issue which deserves to be further investigated.

[25] [26] [27] [28] [29] [30] [31] [32]

ABBREVIATIONS ET

= Endothelin

IGF

= Insulin-like growth factor

INSL = Insulin-like peptide

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[33] [34]

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Received: March 27, 2008

Revised: May 21, 2008

Accepted: June 27, 2008

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