Comparative study of nerve fiber density between adenomyosis patients with moderate to severe pain and mild pain.

BACKGROUND: Since some retrospective studies have given inconsistent findings about innervation in adenomyosis, its role in the pain mechanism is still inconclusive. OBJECTIVE: Define the nerve fiber density in adenomyotic tissue as it correlated to pain symptoms. MATERIAL AND METHOD: A cross-sectional study was performed in twenty-five uterine samples from reproductive age women with adenomyosis who underwent either laparotomy or laparoscopic surgery. The nerve fiber density from hysterectomized specimens as measured by immunohistochemistry staining for Protein gene product (PGP) 9.5 and Neurofilament (NF) were compared with the level of pain in the patients as defined by a visual analogue scale and a verbal rating scale. RESULTS: Nerve fibers as detected by PGP9.5 and NF staining in the myometrium were significantly increased in the group of women with adenomyosis experiencing moderate and severe pain as compared to the group experiencing less pain (4 (0, 7) vs. 1.55 (0, 7)/mm2, p-value <0.001, and 6 (3, 10) vs. 0 (0, 4)/mm2, p-value <0.001 respectively). At both phases of the menstrual cycle, the densities of nerve fibers stained with PGP9.5 and NF showed no significant difference. CONCLUSION: These results suggested that the increased of nerve fibers shown in the more severe pain group might play a role in the pathogenesis or symptoms of adenomyosis.

[Expression of nerve growth factor produced by ectopic endometrium from patients with adenomyosis and its relationship with pain scales and innervation].

OBJECTIVE: To investigate the expression of nerve growth factor (NGF) in the ectopic endometrium in adenomyosis patients, and explore the relationship between NGF expression and innervation or pain scales. METHODS: From Mar. 2009 to Oct. 2009, 45 adenomyosis patients undergoing hysterectomy in Obstetrics and Gynecology Hospital of Fudan University were enrolled in this study, which were classified into 33 cases in pain group and 12 cases in non-pain group based on symptom. The degree of dysmenoreal, chronic pelvic pain and dyspareunia was evaluated by visual analogue scale, including no pain, mild to moderate pain and severe pain group. In the mean time, 26 patients with leiomyoma or cervical intraepithelial neoplasia III (CIN III) undergoing hysterectomy were defined as control group. Ectopic endometrium from experimental group and eutopic endometrium from control group were collected in the surgery. The expression of NGF was examined by immunohistochemistry. The density of protein gene product (PGP) 9.5 positive nerve fibers was detected by immuno-fluorescence. RESULTS: The NGF level and the density of PGP 9.5 positive nerve fibers in adenomyosis pain group (0.25 ± 0.08, 16 ± 8 )were higher than adenomyosis painless (0.19 ± 0.05, P = 0.007;11 ± 5, P = 0.018) and control group (0.18 ± 0.05, P = 0.000; 9 ± 4, P = 0.000) . The NGF level and the density of PGP9.5 positive nerve fibers in severe dysmenorrheal group (0.29 ± 0.07, 19 ± 10) were higher than mild to moderate dysmenorrheal (0.22 ± 0.07, P = 0.018;13 ± 4, P = 0.035) and painless group (0.18 ± 0.05, P = 0.000;11 ± 5, P = 0.006) of adenomyosis patients. There was no difference of NGF level and the density of PGP 9.5 positive nerve fibers in chronic pelvic pain group and no chronic pelvic pain group of adenomyosis patients, so was dyspareunia group and no dyspareunia group. CONCLUSION: The increased NGF level of adenomyosis nodules and improving innervation might be involved in the mechanism of adenomyosis related pain.

NT, NPY and PGP 9.5 presence in myomeytrium and in fibroid pseudocapsule and their possible impact on muscular physiology.

The uterine myoma pseudocapsule is a neurovascular bundle surrounding fibroid, containing neuropeptides, probably involved in uterine scar healing. We studied neurotensin (NT), neuropeptide tyrosine (NPY), and protein gene product 9.5 (PGP 9.5) nerve fibres in the pseudocapsule neurovascular bundle of intramural uterine fibroids on 67 no pregnant women by intracapsular myomectomy sparing the neurovascular bundle, sampling full thickness specimens of the pseudocapsule of uterine fibroids (PUF) and normal myometrium (NM) obtained from the fundus uteri (FU) and the uterine body (UB). The samples were sent for histological and immunofluorescent analyses and compared by morphometrical quantification. The Conventional Unit (C.U.) difference of NT, NPY, and PGP 9.5 nerve fibres was statistically analyzed. Our results showed that NT, NPY, and PGP 9.5 neurofibers are almost equally present in PUF as in NM of a no pregnant uterus. As all of these neuropeptides are present in the uterine muscle and can affect muscle contractility, uterine peristalsis and muscular healing. A myomectomy respecting the pseudocapsule neurofibers should facilitate smooth muscle scarring and promote restoration of normal uterine peristalsis with a possible positive influence on fertility.

Uterine distension differentially affects remodelling and distensibility of the uterine vasculature in non-pregnant rats.

During pregnancy the mammalian uterine circulation undergoes significant expansive remodelling necessary for normal pregnancy outcome. The underlying mechanisms are poorly defined. The goal of this study was to test the hypothesis that myometrial stretch actively stimulates uterine vascular remodelling by developing a new surgical approach to induce unilateral uterine distension in non-pregnant rats. Three weeks after surgery, which consisted of an infusion of medical-grade silicone into the uterine lumen, main and mesometrial uterine artery and vein length, diameter and distensibility were recorded. Radial artery diameter, distensibility and vascular smooth muscle mitotic rate (Ki67 staining) were also measured. Unilateral uterine distension resulted in significant increases in the length of main uterine artery and vein and mesometrial segments but had no effect on vessel diameter or distensibility. In contrast, there were significant increases in the diameter of the radial arteries associated with the distended uterus. These changes were accompanied by reduced arterial distensibility and increased vascular muscle hyperplasia. In summary, this is the first report to show that myometrial stretch is a sufficient stimulus to induce significant remodelling of uterine vessels in non-pregnant rats. Moreover, the results indicate differential regulation of these growth processes as a function of vessel size and type.

Surgical management of neurovascular bundle in uterine fibroid pseudocapsule.

The uterine fibroid pseudocapsule is a fibro-neurovascular structure surrounding a leiomyoma, separating it from normal peripheral myometrium. The fibroid pseudocapsule is composed of a neurovascular network rich in neurofibers similar to the neurovascular bundle surrounding a prostate. The nerve-sparing radical prostatectomy has several intriguing parallels to myomectomy. It may serve either as a useful model in modern fibroid surgical removal, or it may accelerate our understanding of the role of the fibrovascular bundle and neurotransmitters in the healing and restoration of reproductive potential after intracapsular myomectomy. Surgical innovations, such as laparoscopic or robotic myomectomy applied to the intracapsular technique with magnification of the fibroid pseudocapsule surrounding a leiomyoma, originated from the radical prostatectomy method that highlighted a careful dissection of the neurovascular bundle to preserve sexual functioning after prostatectomy. Gentle uterine leiomyoma detachment from the pseudocapsule neurovascular bundle has allowed a reduction in uterine bleeding and uterine musculature trauma with sparing of the pseudocapsule neuropeptide fibers. This technique has had a favorable impact on functionality in reproduction and has improved fertility outcomes. Further research should determine the role of the myoma pseudocapsule neurovascular bundle in the formation, growth, and pathophysiological consequences of fibroids, including pain, infertility, and reproductive outcomes.

Mysteries of endometriosis pain: Chien-Tien Hsu Memorial Lecture 2009.

The more that one looks at the condition endometriosis, the more one realises that it is a unique and complex condition exhibiting a bizarre range of deviations from normal endometrial and myometrial physiology, and presenting with a challenging range of pain-related symptoms. The changing nature of the pain is not well defined, and the molecular mechanisms leading to pain generation are far from clear. Recent research has begun to reveal some of these links between expression of unusual molecules in the eutopic endometrium and ectopic lesions, microanatomical changes in the pelvic nervous system, neuronal dysfunction and the later development of neuropathic pain. A better understanding of these mechanisms will undoubtedly lead to improved use of current medical and surgical treatments, and to the development of novel treatments in the future.

Endometriosis-associated nerve fibers and pain.

The assessment and diagnosis of endometriosis remain elusive targets. Patient and medical-related factors add to delays in the detection and treatment. Recently, investigators have revealed specific nerve fibers present in endometriotic tissue, with existing parallels between density and pain severity. The aim of this review is to compile a comprehensive review of existing literature on endometriosis-related nerve fiber detection, and the effects of medical therapy on these neural fibers. We performed a systematic literature-based review using Medline and PubMed of nerve fibers detected in eutopic endometrium, endometriotic lesions, and the peritoneum. Various arrangements of significant medical terms and phrases consisting of endometriosis, pelvic pain, nerve fiber detection/density in endometriosis, and diagnoses methodology, including treatment and detection were applied in the search. Subsequent references used were cross-matched with existing sources to compile all additional similar reports. Similar nerve fibers were detected within lesions, endometrium, and myometrium, though at varying degrees of density. Hormonal therapy is widely used to treat endometriosis and was shown to be related to a reduction in fiber density. A direct result of specific nerve fiber detection within eutopic endometrial layers points to the use of a minimally invasive endometrial biopsy technique in reducing delay in diagnosis and subsequent possible preservation of fertility.

Effects of experimentally induced diabetes mellitus on pharmacologically and electrically elicited myometrial contractility.

1. Diabetes is one of the most frequent complications of gestation, affecting approximately 7% of pregnancies. However, little is known about its effects on electrically and pharmacologically stimulated myometrial contractility. The aim of the present study was to investigate the consequences of streptozotocin (STZ)-induced diabetes on: (i) electrical field stimulation (EFS)-evoked contraction of isolated uterine rings as a function of gestational age; and (ii) the uterotonic and tocolytic actions of α- and ß-adrenoceptor stimulation, respectively. The effects of oxytocin in late pregnancy were also investigated. 2. During pregnancy, EFS-evoked contractions of isolated uterine rings from intact rats declined, whereas isolated uterine rings from diabetic rats exhibited continuously low sensitivity to EFS. 3. In non-pregnant rats, diabetes resulted in increased noradrenaline-mediated contractility and a decreased relaxation response to terbutaline. At the mRNA level, diabetes enhanced the expression of α1B-adrenoceptors in non-pregnant rats from 14.65 to 18.39 µg/mL (P < 0.05), whereas the expression of α1D-adrenoceptors decreased (from 42.87 to 35.67 µg/mL; P < 0.05). During pregnancy, the responses to these sympathomimetics did not differ between diabetic and intact rats. 4. In late pregnancy (on Days 15 and 21), oxytocin caused greater maximum contractility of uterine rings from diabetic rats without affecting the EC(50). In addition, on Day 15 of pregnancy, the expression of oxytocin receptors in the myometrium of diabetic rats was higher than that in intact rats. 5. The results of the present study indicate that experimental diabetes facilitates gestation-induced denervation and increases myometrial sensitivity to oxytocin in late pregnancy. If similar mechanisms operate in humans, this could contribute to a tendency to premature uterine contractions in diabetes-complicated pregnancies.

[Distribution of nerve fibers in endometrium and its clinical significance in adenomyosis].

OBJECTIVE: To investigate nerve fibers distribution in endometrium of adenomyosis and their relationship with dysmenorrhea. METHODS: Endometrial tissue was sampled from 74 hysterectomy specimens including 32 cases with adenomyosis and 42 cases with uterine fibroids. Two-step Envision immunohistochemical staining was used to detect distribution of nerve fibers in endometrium. Highly specific polyclonal rabbit anti-protein gene product 9.5 (PGP9.5) and monoclonal mouse anti-neurofilament protein (NF) were used to demonstrate both myelinated and unmyelinated nerve fibers in endometrium in women with adenomyosis and uterine fibroids. RESULTS: The positive rate of PGP9.5 immunoreactive nerve fibers in the functional layer of endometrium of pain patients were with 64% (14/22) in adenomyosis and 67% (10/15) in uterine fibroids. And their density were 0.6 (0 - 9.4)/mm(2) and 0.6 (0 - 6.0)/mm(2) without reaching statistical difference (P > 0.05). No expression of NF could be detected in the functional layer of endometrium of adenomyosis and uterine fibroids. There were no PGP9.5 immunoreactive nerve fibers in the functional layer of endometrium in non-pain women with adenomyosis and uterine fibroids. Moreover, No NF immunoreactive nerve fibers in the functional layer of endometrium were shown in non-pain patients with adenomyosis and uterine fibroids. PGP9.5 immunoreactive nerve fibers and the nerve density in the basal layer of endometrium were 64% (14/22), 1.1 (0 - 12.0)/mm(2) in pain adenomyosis and 50% (5/10), 0.6 (0 - 3.0)/mm(2) in non-pain adenomyosis. NF immunoreactive nerve fibers and the density in the basal layer of endometrium were 23% (5/22), (0 - 0.6)/mm(2) in pain adenomyosis and 20% (2/10), (0 - 1.0)/mm(2) in non-pain adenomyosis. PGP9.5 immunoreactive nerve fibers and the nerve density in the basal layer of endometrium were 80% (12/15) and 1.6 (0 - 10.0)/mm(2) in pain fibroids and 44% (12/27), 0 (0 - 5.0)/mm(2) in non-pain fibroids. NF immunoreactive nerve fibers and the nerve density in the basal layer of endometrium were 40% (6/15), 0 (0 - 0.4)/mm(2) in pain fibroids and 15% (4/27), 0 (0 - 1.0)/mm(2) in non-pain fibroids. There was no statistical different PGP9.5 and NF immunoreactive nerve fibers distribution in basal layer of endometrium between pain adenomyosis and pain fibroids or between non-pain adenomyosis and non-pain fibroids (all P > 0.05). However, PGP9.5 immunoreactive nerve fibers density in basal layer of endometrium was higher in pain adenomyosis and fibroids when compared with non-pain adenomyosis and fibroids (P < 0.05). CONCLUSIONS: PGP9.5 immunoreactive nerve fibers might confer the occurrence of pelvic pain, however, NF immunoreactive nerve fibers may not involved in the pathogenesis of pain.

The importance of pseudocapsule preservation during hysteroscopic myomectomy.

Minimally invasive techniques for myomectomy are based on the rationale of preserving the myometrial integrity, in order to spare muscular and fibro-neurovascular myometrial fibers and ensure complete and bloodless myoma removal. Post-operative myometrial vascularization is crucial in injured muscle regeneration. The post-surgical myometrial healing is needful for uterine reproductive function. Neurotransmitters and neurofibers were analyzed in the myoma pseudocapsule surrounding fibroid. They activate signaling molecule synthesis and release which, in turn, promote cell activation and induce muscle regeneration and growth. Pseudocapsule damage during myomectomy may lead to a reduction of neuropeptides and neurofibers at the hysterotomic site, to a poor physiological myometrial healing, with more fibrosis due to hypoxia, ischemia and necrosis. These pathophysiological events cause deficit in myometrial neurotransmission, muscular impulse and contractility, with ultimately impaired uterine muscle function during pregnancy, labor and delivery. Hence, during myomectomy, all manipulations should be performed as precisely and bloodlessly as possible, avoiding extensive, high wattage diathermocoagulation or excessive tissue manipulation or muscular trauma. Any iatrogenic pseudocapsule damage may alter neurotransmitter function during successive myometrial healing, impacting negatively on uterine repair and on eventual pregnancies. Hence the reasoned myomectomy on a biological basis, the "intracapsular myomectomy", satisfied these surgical and physiological requirements. It was described precisely and firstly by the hysteroscopy, with the image magnification of the preservation of the myoma pseudocapsule. The "intracapsular hysteroscopic myomectomy" demonstrated the safe and effective removal of submucous myomas with intramural development. It allowed to completely remove the myoma in one or two surgical steps, saving the pseudocapsule and the surrounding healthy myometrium. The respect of the myometrium and the reduced thermal injury, a part the excellent outcomes in terms of surgical complications prevention, post-surgical fibrosis and intrauterine synechiae reduction, highlighted the physiological development of a successive pregnancy, without any myometrial complications during pregnancy, labor and delivery.

Neurotrimin is an estrogen-regulated determinant of peripheral sympathetic innervation.

Mechanisms underlying axon degeneration in peripheral neuropathies and during normal remodeling are poorly understood. Because estrogen induces widespread sympathetic axon degeneration in female reproductive tract smooth muscle, we surveyed estrogen-regulated genes in rat myometrium. Microarray analysis revealed that the neural cell adhesion protein neurotrimin (Ntm) was markedly up-regulated 6 hr and down-regulated 24 hr after injection of 17beta-estradiol, and real time RT-PCR confirmed this pattern of expression. Protein analysis by Western blotting showed that uterine Ntm protein is also up-regulated in vivo 6-24 hr following estrogen injection and that Ntm protein is increased selectively in the myometrium during the high-estrogen phase of the estrous cycle. Cultured myometrial smooth muscle cells display perinuclear accumulations of Ntm protein, and 17beta-estradiol also increases intracellular levels of Ntm and its secretion into the culture medium. To determine if neurotrimin is required for estrogen-induced sympathetic pruning, sympathetic neurons were cocultured with uterine smooth muscle cells transfected with siRNA directed against Ntm. Although estrogen inhibited neurite outgrowth in nontransfected cocultures, estrogen's ability to reduce sympathetic outgrowth was impaired substantially following Ntm down-regulation. This supports a role for neurotrimin in mediating estrogen-induced sympathetic pruning in some peripheral targets. Together with earlier studies, these findings support the idea that physiological sympathetic axon degeneration is a multifactorial process requiring dynamic regulation of multiple repellant proteins.

Characterization of prostanoid receptors present on adrenergic neurons innervating the porcine uterine longitudinal muscle.

The cyclooxygenase-prostanoid pathway regulates myometrial contractility through activation of prostanoid receptors on uterine smooth muscles. However, the possible expression of prostanoid receptors on autonomic nerves cannot be excluded completely. The aim of the present study was to clarify the presence of neural prostanoid receptors on adrenergic nerves in the porcine uterine longitudinal muscle. In [(3)H]-noradrenaline-loaded longitudinal muscle strips of porcine uterus, electrical field stimulation (EFS) evoked [(3)H]-noradrenaline release in a stimulation frequency-dependent manner. The EFS-evoked release was completely abolished in Ca(2+)-free (EGTA, 1mM) incubation medium and by tetrodotoxin or omega-conotoxin GVIA, suggesting that [(3)H]-noradrenaline was released from neural components. The EFS-evoked [(3)H]-noradrenaline release was significantly enhanced by treatment with indomethacin. In the presence of indomethacin, PGE(2) and PGF(2alpha), but not PGD(2), inhibited the EFS-evoked [(3)H]-noradrenaline release. Of synthetic prostanoid receptor agonists examined, both U46619 (TP) and sulprostone (EP(1)/EP(3)) decreased the EFS-evoked [(3)H]-noradrenaline release in a concentration-dependent manner, while fluprostenol (FP), BW245C (DP) and butaprost (EP(2)) were almost ineffective. SQ29548 (TP receptor antagonist) blocked the effect of U46619, but SC19220 (EP(1) receptor antagonist) did not change the inhibition by sulprostone or PGE(2). Double immunofluorescence staining using protein gene product 9.5, tyrosine hydroxylase, EP(3) receptor and TP receptor antibodies suggested the localization of EP(3) or TP receptors on adrenergic nerves in the porcine uterus. These results indicated that neural EP(3) and TP receptors are present on adrenergic nerves of the porcine uterine longitudinal muscle. Endogenous prostanoid produced by cyclooxygenase can regulate noradrenaline release in an inhibitory manner through activation of these neural prostanoid receptors.

Monosodium glutamate impairs the contraction of uterine visceral smooth muscle ex vivo of rat through augmentation of acetylcholine and nitric oxide signaling pathways.

The aim of the study was to examine the toxic effects of Monosodium glutamate (MSG), an extensively used food additive, on the contraction of uterine visceral smooth muscle (UVSM) in rat and to elucidate the probable neurocrine mechanism involved in it. MSG produced significant potentiation of the force and inhibition of frequency of uterus recorded ex vivo in chronic MSG exposure and in single dose acute experiments. MSG also produced significant potentiation of force of acetylcholine induced contraction and no alterations in atropine induced contraction of uterus. Further, MSG produced significant increase in force and frequency of contraction of neostigmine incubated uterus. We have found significant potentiation of the post pause force of contraction of uterus when MSG was applied in adrenaline incubated uterus. MSG also produced significant decrease in frequency of contraction of sodium nitroprusside incubated uterus; increase in frequency of N-ω-Nitro-l-Arginine Methyl Ester incubated uterus and no significant changes in frequency of contraction of methylene blue incubated uterus. These results indicate that MSG potentiates the force of contraction of UVSM predominantly by augmenting the activity of cholinergic intrinsic efferents and inhibits the frequency of contraction probably by augmenting the activity of nitrergic efferents. In conclusion, MSG potentiates the force and inhibits the frequency of contraction of UVSM, and the MSG induced effect is probably mediated through the augmentation of acetylcholine and nitric oxide signaling pathways.

Postablation neuroma of the myometrium-a report of 5 cases.

When hysterectomy is performed for chronic pelvic pain, routine pathology examination often provides no explanation. However, analysis of small uterine nerves using immunostains may help to address this deficiency. Small uterine nerves tend to be sparse or absent in wide areas of normal myometrium. Some studies of uterine nerves have suggested that endometriosis, adenomyosis, and fibroids are not inherently painful, with increased small nerves in the inner uterine wall associated with the history of pelvic pain. Although such areas may appear normal on hematoxylin and eosin (H&E), we have found a subtle inner wall lesion termed inner myometrial elastosis, best detected with trichrome or elastic stains, which may be a reaction to microscopic tears of inner myometrium. Such tears may induce increased inner wall innervation via the generation of nerve growth factor in granulation tissue. In the course of studying uterine nerves with immunostains, we found 5 cases with florid nerve proliferation, after deep endometrial ablation for abnormal uterine bleeding led to increased pelvic pain. We suggest that immunostains for postablation neuromas should be done in hysterectomies when pelvic pain increases after endometrial ablation. This may offer gynecologists and their patients an objective finding with a rational, scientific explanation for the pelvic pain.

A quantitative study of rat uterine sympathetic innervation during pregnancy and post partum.

In mammals, pregnancy induces a transient and extensive degeneration of uterine sympathetic innervation. We used the models of unilateral oviduct ligation and in oculo myometrium transplant in pregnant rats to address the role of stretching forces and/or hormone milieu in the loss of sympathetic innervation. The sympathetic fibres of the uterine horn and in oculo myometrial transplants were quantified on tissue sections processed by the glyoxylic acid technique. In normal pregnant rats, the density of uterine horn innervation was significantly reduced at late pregnancy and recovery took place during post partum. The empty horn of pregnant rats showed no significant changes in density of myometrial innervation during pregnancy or post partum. In oculo myometrial transplants were organotypically reinnervated in virgin animals. When the transplants were exposed to gestational hormonal milieu, few or no fibres were observed to the end of pregnancy; however, a significant increase at post partum was observed. Results showed that both the effects of stretching and the hormone milieu derived from the fetus-placenta complex play a role as inductors of changes on sympathetic myometrial innervation during pregnancy and support the idea that immature muscular uterine fibres are more susceptible to the effects of pregnancy than those originating from adult animals.

Estrogen-induced collagen reorientation correlates with sympathetic denervation of the rat myometrium.

Estrogen inhibits the growth and causes the degeneration (pruning) of sympathetic nerves supplying the rat myometrium. Previous cryoculture studies evidenced that substrate-bound signals contribute to diminish the ability of the estrogenized myometrium to support sympathetic nerve growth. Using electron microscopy, here we examined neurite-substrate interactions in myometrial cryocultures, observing that neurites grew associated to collagen fibrils present in the surface of the underlying cryosection. In addition, we assessed quantitatively the effects of estrogen on myometrial collagen organization in situ, using ovariectomized rats treated with estrogen and immature females undergoing puberty. Under low estrogen levels, most collagen fibrils were oriented in parallel to the muscle long axis (83% and 85%, respectively). Following estrogen treatment, 89% of fibrils was oriented perpendicularly to the muscle main axis; while after puberty, 57% of fibrils acquired this orientation. Immunohistochemistry combined with histology revealed that the vast majority of fine sympathetic nerve fibers supplying the myometrium courses within the areas where collagen realignment was observed. Finally, to assess whether depending on their orientation collagen fibrils can promote or inhibit neurite outgrowth, we employed cryocultures, now using as substrate tissue sections of rat-tail tendon. We observed that neurites grew extensively in the direction of the parallel-aligned collagen fibrils in the tendon main axis but were inhibited to grow perpendicularly to this axis. Collectively, these findings support the hypothesis that collagen reorientation may be one of the factors contributing to diminish the neuritogenic capacity of the estrogen-primed myometrial substrate.

Dienogest reduces proliferation, NGF expression and nerve fiber density in human adenomyosis.

OBJECTIVES: To evaluate the in vivo effect of dienogest on proliferation, apoptosis, aromatase expression, vascular density, nerve growth factor (NGF) expression and nerve fiber density in human adenomyosis tissue. STUDY DESIGN: Twelve women who underwent hysterectomy for adenomyosis were enrolled. Six patients received dienogest treatment prior to hysterectomy (dienogest group), and age-matched six patients who had not received any hormonal treatment for ≥3 months before surgery (control group). Cell proliferation, vascular and nerve fiber density in adenomyosis tissue were evaluated by staining for Ki67, von Willebrand factor and PGP9.5, respectively. Apoptosis was detected using the TUNEL assay. The expression aromatase and NGF were evaluated by staining for corresponding antibodies. RESULTS: The proportion of Ki67 positive epithelial cells was significantly lower in samples from dienogest-treated patients in comparison with controls (p<0.05). The density of blood vessels in adenomyosis was marginally lower in the dienogest group in comparison with controls but statistical significance was not reached (p=0.07). The intensity of NGF expression and the density of nerve fibers were significantly lower in the dienogest group compared with controls (p<0.05 for both). CONCLUSION: This study demonstrates that adenomyosis, taken from patients treated with dienogest, shows remarkable histological features, such as reductions in proliferation, NGF expression and nerve fiber density. These findings indicate the impact of dienogest on local histological events, and explains its therapeutic effect on adenomyosis.

Pre-eclampsia and partial uterine denervation.

Pre-eclampsia is characterised by a maternal syndrome of hypertension and proteinuria, that is frequently associated with reduced fetal growth. The characteristic histopathological observation in the placental bed is narrowing and atherosis of the distal branches of the uterine arteries at, and around, the deciduo-myometrial interface. In the maternal kidneys there is swelling of the glomerular capillaries and mesangium with some inclusions in the basement membrane ("glomeruloendotheliosis") and evidence of vasoconstriction in many other organs. Untreated maternal hypertension leads to convulsions (eclampsia) which may result in maternal and fetal death. The nerve plexus at the endometrial(decidual)--myometrial interface was first reported in 1959 though has received little attention in the intervening years. It appears to play an important role in maintaining the separation of two tissues with intrinsic proliferative potential (endometrium and myometrium). The present hypothesis proposes that damage to the nerve plexus at the endometrial-myometrial interface causes impaired control of a third proliferative tissue (invading trophoblast) resulting in the characteristic histological changes. Growth factors produced by nerves and blood vessels may contribute to the process of normal placentation e.g. nerve growth factor, vascular endothelial growth factor, etc. and these processes may be compromised in areas of denervation. Neural connections between the uterine and renal innervations (L1, 2) result in renal vasoconstriction and widespread systemic maternal vasoconstriction in an attempt to provide increased blood flow for the uteroplacental circulation (maternal hypertension, small-for-gestational age fetus). Loss of these neural connections through the same process of partial uterine denervation may cause reduced fetal growth without the maternal circulatory changes of pre-eclampsia (maternal normotension, small-for-gestational age, fetus). Variations in maternal circulatory compliance alter the "phenotype" of the condition such that prior maternal hypertension may cause pre-eclampsia through intrarenal mechanisms without significant fetal growth restriction. Increases in circulatory compliance in multiparity prevent the typical features of the condition, or, if they are expressed then they present in a different sequence with haematological and hepatic consequences presenting before the renal manifestations (HELLP syndrome, haemolysis, elevated liver enzymes, low platelets).

The importance of pelvic nerve fibers in endometriosis.

Several lines of recent evidence suggest that pelvic innervation is altered in endometriosis-affected women, and there is a strong presumption that nerve fibers demonstrated in eutopic endometrium (of women with endometriosis) and in endometriotic lesions play roles in the generation of chronic pelvic pain. The recent observation of sensory C, sensory A-delta, sympathetic and parasympathetic nerve fibers in the functional layer of endometrium of most women affected by endometriosis, but not demonstrated in most women who do not have endometriosis, was a surprise. Nerve fiber densities were also greatly increased in myometrium of women with endometriosis and in endometriotic lesions compared with normal peritoneum. Chronic pelvic pain is complex, and endometriosis is only one condition which contributes to this pain. The relationship between the presence of certain nerve fibers and the potential for local pain generation requires much future research. This paper reviews current knowledge concerning nerve fibers in endometrium, myometrium and endometriotic lesions, and discusses avenues of research that may improve our knowledge and lead to enriched understanding and management of endometriotic pain symptoms.

Estrogen modulates myometrium-induced sympathetic neurite formation through actions on target and ganglion.

Estrogen induces rapid and extensive degeneration of rodent uterine myometrial sympathetic innervation. To clarify the underlying mechanisms, we used explant cultures to assess whether estrogen affects the myometrium's ability to induce sympathetic neuritogenesis and the sympathetic neuron's ability to respond. Superior cervical ganglion explants from ovariectomized adult donors extended neurites when cultured alone in serum-free medium, and their numbers increased 2.3-fold in the presence of myometrial explants from ovariectomized adult rats. The myometrium's ability to induce neuritogenesis was abolished by injection of myometrium donors with 17beta-estradiol 24 h prior to tissue harvest. Myometrial neurite-promoting effects were also abolished by adding 2x10(-8)M estradiol to the culture medium. Because outgrowth from ganglia of ovariectomized rats cultured alone was not affected by estrogen in the culture medium, this indicates that estrogen acts directly on myometrium to abrogate its neurite-promoting effects. However, estrogen injection of ganglion donor rats also inhibited neurite extension toward ovariectomized myometrium, suggesting that some factor in ovariectomized rats normally acts on the ganglion to prevent estrogen from inhibiting neurite outgrowth. When ganglia from hypophysectomized ovariectomized donors were cultured alone, neuritogenesis was normal but estrogen added to the culture medium now attenuated outgrowth. Prolactin but not other pituitary-derived hormones reversed the suppression of neuritogenesis induced by estrogen. We conclude that estrogen acts directly on myometrium to inhibit its neuritogenic effects on sympathetic neurons. Estrogen can also attenuate neurite formation by acting directly on the ganglion; this effect normally is not apparent at low estrogen levels because a pituitary hormone (possibly prolactin) prevents the ganglion from responding fully to estrogen. With high in vivo estrogen, this pituitary hormone's effects are abated, possibly through diminished release, and estrogen directly reduces ganglion neuritogenesis. Thus, estrogen regulates uterine sympathetic nerve remodeling through actions on myometrium, ganglion, and intermediary pituitary factors.