Endogenous Ovarian Angiogenesis in Polycystic Ovary Syndrome-Like Rats Induced by Low-Frequency Electro-Acupuncture: The CLARITY Three-Dimensional Approach.

We sought to determine the role of ovarian vascularity and neo-angiogenesis in the development of mature follicles in polycystic ovary syndrome (PCOS) and to identify any changes induced by low-frequency electro-acupuncture (EA). Twenty-eight 21-day-old female Wistar rats were randomly divided into four groups-Control, Obesity, PCOS-like, and PCOS-like-EA (n = 7/group). Rats in the Obesity group were fed a high-fat diet throughout the experiment. Rats in the PCOS-like and PCOS-like-EA groups were implanted with a sustained-release tube containing 5α-dihydrotestosterone (DHT) beneath the skin of the neck. Rats in the PCOS-like-EA group received low-frequency EA treatment starting at 70 days for 30 min five times a week for four weeks. At the end of the experiment, all rats were euthanized and perfused with hydrogel. The ovaries were collected for clarification and imaging, and ovarian vascularity and neo-angiogenesis were analyzed. Compared with Control and Obesity rats, the ovaries in DHT-induced PCOS-like rats were smaller in size and had fewer mature follicles and corpora lutea. EA increased angiogenesis in the antral follicles of PCOS-like rats, which in turn promoted follicle maturation, ovulation, and CL formation. Therefore, endogenous ovarian angiogenesis plays a very important role in follicular maturation and might be one of the peripheral and direct mechanisms of EA on PCOS.

The effect of drospirenone (3 mg) with ethinyl estradiol (30 mcg) containing pills on ovarian blood flows in women with polycystic ovary syndrome: a case controlled study.

OBJECTIVE: To evaluate whether oral contraceptive pill (OCP) therapy has any effects on ovarian stromal blood flow by using pulsed and color Doppler at the end of 3 months follow-up period of OCP-users and non-users with or without polycystic ovary syndrome (PCOS). STUDY DESIGN: 200 patients were included in the study. The patients were designed into four groups as follows; Group 1: PCOS patients that received OCP containing 30 mcg ethinyl estradiol (EE) plus 3mg drospirenone for 3 months (DRP n=50); Group 2: PCOS patients that received no medication (n=50); Group 3: Healthy controls that received OCP (EE plus DRP) (n=50); Group 4: healthy controls that received no medication (n=50). Resistance index (RI) and pulsatility index (PI) of both ovarian arteries, hormonal, anthropometric and biochemical parameters were assessed before and after 3 months. RESULTS: There was a significant increament in RI and PI of both ovarian arteries in healthy controls (Group 3) and in women with PCOS (Group 1) who received OCP (p<0.001). The increment rate in both Doppler parameters were significantly higher in women with PCOS (Group 1) than healthy controls (Group 3) (p<0.001). Whereas RI and PI values of both ovaries remained unchanged in all untreated women with or without PCOS (Groups 2 and 4). CONCLUSION: OCP therapy reduced ovarian vascularization in both PCOS and healthy users after 3 months of therapy and this decrease is especially noticeable in women with PCOS.

The effects of laparoscopic ovarian drilling on flow mediated dilation in women with PCOS: relationship with insulin resistance and IGF-I.

AIM: The aim of this paper was to investigate whether laparoscopic ovarian drilling (LOD) has an effect on endothelial function in women with polycystic ovary syndrome (PCOS) using well validated marker of brachial artery flow-mediated dilation (FMD) and to determine which hormonal and/or metabolic parameters are responsible for postoperative endothelial response. METHODS: In a prospective pilot trial, 34 infertile women with PCOS were studied for measurement of brachial artery FMD and insulin-like growth factor (IGF)-I. Blood samples for total testosterone (T), sex hormone binding globulin (SHBG), lipid profile as well as insulin resistance index (HOMA-IR) were measured before and 3 months after LOD. Twenty-six normal fertile subjects matched for age and body mass index (BMI) served as control group. RESULTS: Baseline %FMD in patients with PCOS was significantly lower than that in control subjects. After LOD 24 women had increased %FMD (responders), whereas 10 women had decreased or unchanged %FMD (non-responders). However, neither fasting plasma glucose, insulin levels nor HOMA-IR changed after LOD. Serum levels of T, IGF-1, total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), and sex-hormone binding globulin (SHBG) significantly decreased whereas high-density lipoprotein cholesterol (HDL-C) significantly increased (P<.05). Responders had significantly lower baseline %FMD than did non-responders and both IGF-I and T significantly decreased in responders, but not in non-responders after LOD (P<0.05). Simple regression analysis revealed that the change of %FMD showed a significant negative correlation (r=-0.731, P=0.007) with that of LDL-C, but not with IGF-1 nor T in responders. CONCLUSION: Women with PCOS have evidence for endothelial dysfunction which might be improved shortly after LOD. This results from decrease in LDL-C rather than decrease in androgens and/or IGF-1 per se.

Analysis of the vascular responses in a murine model of polycystic ovary syndrome.

Polycystic ovary syndrome (PCOS) is the most common endocrine disorder in women of there productive age, but the exact pathophysiological mechanisms involved remain unclear. Cardiovascular disease risk is increased in PCOS patients and endothelial damage has been observed. We recently developed a mouse model of PCOS with reproductive and metabolic characteristics resembling those observed in women with PCOS. In this model we studied vascular function with particular emphasis on markers of vascular endothelial function. Animals were treated for 90 days with dihydrotestosterone (DHT; 27.5 mg/day) or placebo using subcutaneous continuous-release pellets. Aortas were isolated for isometric force recordings in organ baths to investigate endothelial and vascular smooth muscle characteristics. Lungs were used to analyze endothelial nitric oxide synthase (eNOS) expression and phosphorylation. Asymmetric dimethylarginine (ADMA) levels were investigated in serum to assess endothelial damage. Expression of androgen receptor (Ar) mRNA was studied in aortas. DHT treatment (compared with placebo) induced i) a significant decrease in acetylcholine-induced aortic relaxations, with no change in calcitonin gene related peptide- or sodium nitroprusside-induced relaxations, as well as 5-hydroxytryptamine-induced contractions; ii) no change in eNOS expression/phosphorylation in lungs or in plasma ADMA levels; and iii) a twofold increase in aortic AR expression. Our results suggest that, in DHT-exposed mice, hyperandrogenemia specifically decreases endothelium dependent vasorelaxation without deterioration of smooth muscle function. This study may initiate further investigations to elucidate underlying mechanism for the phenotype that is present in these animals, as well as in PCOS patients.

The blood-follicle barrier (BFB) in disease and in ovarian function.

The blood-follicle barrier (BFB) is one of the blood-tissue barriers in mammalian body found in developing follicles in the ovary. The BFB, besides the tight junction (TJ)-permeability barrier of the endothelial cells in the microvessels that surround the developing follicle, is constituted and contributed significantly by the basement membrane of the developing follicle which alters its composition rapidly during follicle development. While the concept of the BFB and its ultrastructure were described more than six decades ago, fewer than 20 reports are found in the literature that were dedicated to investigate the biology, regulation, and function of the BFB either in health or in disease. Furthermore, detailed analysis of the adhesion protein complexes and the regulation of the junction dynamics at the BFB are still missing in the literature. The goal of this short chapter is to provide an update on this important blood-tissue barrier, it is obvious that future investigation is much needed in the field to understand this ultrastructure better in order to treat and better ovarian disorders including ovarian cancer.

Brachial artery conductance during reactive hyperemia is increased in women with polycystic ovary syndrome.

OBJECTIVE: To examine changes in brachial artery conductance (BAC) during reactive hyperemia in women with polycystic ovary syndrome (PCOS) compared to controls. STUDY DESIGN: This is a pilot case-control study performed at a single academic medical center. Changes in BAC during reactive hyperemia were evaluated in 31 women with PCOS and 11 healthy control women. Fasting glucose, insulin, lipids and androgen levels were also determined. A mixed-effects model was used to compare the PCOS curve to the control curve for change in BAC from baseline during reactive hyperemia. RESULTS: Body mass index (BMI) and testosterone levels were significantly increased in the PCOS group compared to controls (P<0.05). In addition, the PCOS group had higher total and LDL cholesterol levels (P=0.05 and 0.09, respectively). Change in BAC from baseline during reactive hyperemia was significantly increased in the PCOS group compared to controls even after adjusting for age, BMI and LDL cholesterol levels (P<0.0001). There were no significant differences between the two groups in age, blood pressure, or fasting glucose or insulin levels. CONCLUSIONS: Brachial artery conductance during reactive hyperemia is significantly increased in women with PCOS compared to controls and may be a novel early indicator of increased cardiovascular risk in women with PCOS.

Immunohistochemical localization of CD31, NOTCH1 and JAGGED1 proteins in experimentally induced polycystic ovaries of immature rats.

We analyzed histomorphometrical changes and blood vessel immunohistochemical staining of CD31, NOTCH1 and JAGGED1 in induced polycystic ovaries of immature female Wistar rats, as well as serum hormone levels. The rats were randomly divided into control (n=18) and treated (n=18) groups. Treated animals received intramuscularly testosteronenantat weekly (0.1mg/g). Controls received the same amount of ricinus oil. Rats were weighed daily. Control and treated subgroups (6 rats per subgroup) were subsequently sacrificed after 21, 28 and 35 days of treatment. In ovaries of treated rats we found large cystic follicles, thick stromal tissue, many atretic preantral follicles, no ovulation and a thinner granulosa cell layer. CD31 stained blood vessels in the theca layer were reduced, with reduced JAGGED1 and NOTCH1 immunostaining. In controls, preantral and antral follicles were larger than in the treated group. Treated animals showed statistically significant lower progesterone and higher testosterone levels. They gained more weight than controls. Reduced immunostaining for NOTCH1 and JAGGED1 of reduced blood vessels of the theca layer was found in all stages of folliculogenesis with a distinct reduction in cystic and atretic follicles. Our results provide evidence of intrinsic abnormality during all stages of folliculogenesis in polycystic ovaries and this may result from crosstalk between circulating gonadotropins and follicular angiogenesis.

Superficial ovarian cortex vascularization is inversely related to the follicle reserve in normal cycling ovaries and is increased in polycystic ovary syndrome.

BACKGROUND: The superficial ovarian cortex constitutes the micro-environment where resting and early growing follicles reside. As small follicles do not possess an independent capillary network, both their survival and early growth depend on their proximity to the cortical vessels. Little is known about the possible changes in superficial ovarian cortex vascularization in normal women throughout reproductive life or in pathological conditions such as polycystic ovary syndrome (PCOS) involving abnormal early follicle growth. We studied the vascularization of the superficial and deep cortical stroma (DCS) in normal cycling ovaries from 21 to 50 years of age and in infertile women with PCOS. METHODS: We used archival ovarian samples and specific CD34 immunostaining to determine blood vessel density and to analyse correlation with age and with the ovarian follicle reserve. RESULTS: Normal cycling ovaries showed an age-related increase in the superficial cortical stroma vascularization that was inversely correlated with the density of small (primordial and primary) follicles. In contrast, blood vessel density in the DCS significantly decreased in women aged >or=40 years. Ovaries from PCOS showed a 2-fold increase in blood vessel density in both superficial cortical stroma and DCS with respect to age-matched controls. CONCLUSIONS: The increased vascularization of the superficial cortical stroma in normal ovaries in relation to age and in ovaries from PCOS could have profound effects on cortical metabolic rate, primordial follicle survival/activation and early follicle growth, and may underline changes in follicle dynamics in mid-aged women and in PCOS.

Increased arterial stiffness in nonobese women with polycystic ovary syndrome (PCOS) without comorbidities: one more characteristic inherent to the syndrome?

BACKGROUND: Polycystic ovary syndrome (PCOS) is associated with adverse metabolic effects. Some cardiovascular disease (CVD) risk markers are increased in women with PCOS. However, early markers of atherosclerosis are also associated with obesity and insulin resistance, which are related to PCOS. These markers may result either directly from PCOS or indirectly as a consequence of the comorbidities associated with the syndrome. CONTEXT: To assess the presence of early CVD markers in young, nonobese women with PCOS. PATIENTS: Forty women with PCOS and 50 healthy women with regular menstrual cycles, matched for age and body mass index (BMI). MEASUREMENTS: The following CVD markers were assessed by ultrasonography: common carotid artery (CCA) stiffness index (beta), distensibility and intima-media thickness (IMT), and brachial artery flow-mediated dilatation (FMD). Inflammatory markers, including interleukin (IL)-6, tumour necrosis factor (TNF)-alpha, homocysteine, C-reactive protein (CRP), glycaemia, lipid profile and insulin, were also assessed. RESULTS: CCA beta was higher in PCOS than in control women (3.72 +/- 0.96 vs. 3.36 +/- 0.96, P = 0.04) and CCA distensibility was lower (0.31 +/- 0.08 vs. 0.35 +/- 0.09 mmHg(-1), P = 0.02). Waist circumference, total testosterone and the Free Androgen Index (FAI) were higher in PCOS patients than in controls (78.2 +/- 10.0 vs. 71.5 +/- 7.2 cm, P = 0.001; 88.1 +/- 32.4 vs. 57.1 +/- 21.2 ng/dl, P < 0.01; 12.7 +/- 15.7%vs. 4.7 +/- 2.3%, P < 0.01, respectively), while SHBG was reduced (37.9 +/- 19.1 vs. 47.8 +/- 18.3 nmol/l, P = 0.01). The remaining variables did not differ between the groups. CONCLUSIONS: Young women with PCOS exhibit changes in vascular elasticity even in the absence of classical risk factors for CVD, such as hypertension and obesity.

Early microvascular and macrovascular dysfunction is not accompanied by structural arterial injury in polycystic ovary syndrome.

OBJECTIVE: During the last decade cardiovascular risk factors and endothelial dysfunction have been shown to be present early in life in women with Polycystic Ovary Syndrome (PCOS). The aim of the present study was a global assessment of abnormalities in the arterial bed of young women with PCOS by non-invasive, reproducible methods. DESIGN: 27 women with PCOS and 27 control women of comparable age, body mass index and waist-to-hip ratio were studied. Macrovascular function was assessed by flow-mediated dilatation (FMD) on the brachial artery. Nitrate-induced dilatation (NID) was performed to exclude a vascular smooth muscle cells injury. Microvascular function was assessed by venous occlusion plethysmography studying forearm blood flow. Arterial structure was evaluated by ultrasonographic assessment of intima-media thickness (IMT) of the carotid artery. RESULTS: FMD values were lower in women with PCOS compared to controls (PCOS: 3.84+/-0.74% vs. controls: 9.83+/-0.97%, P<0.001), but no difference was observed in NID (PCOS: 16.59+/-1.84% vs. controls: 16.64+/-2.05%, P=0.98) values. The time required for reactive hyperemia to reach peak value, a plethysmography parameter, was longer in PCOS women (PCOS: 20.63+/-4.67 sec vs. controls: 10.38+/-5.11 sec, P=0.02). No difference was observed in the combined IMT among the studied groups (PCOS: 0.49+/-0.01 mm v.s. controls: 0.51+/-0.02 mm, P=0.19). CONCLUSIONS: Using non invasive methodologies endothelial dysfunction in the macrocirculation and early impairment in the microcirculation were demonstrated in young women with PCOS who had normal profile of glycemia, lipidemia and blood pressure, and no evidence of structural arterial impairment.

Evaluation of the impact of laparoscopic ovarian drilling on Doppler indices of ovarian stromal blood flow, serum vascular endothelial growth factor, and insulin-like growth factor-1 in women with polycystic ovary syndrome.

OBJECTIVE: To study the serum levels and correlation of vascular endothelial growth factor (VEGF), insulin-like growth factor 1 (IGF-1), hormonal profile, and Doppler blood flow changes within the ovarian stroma before and after laparoscopic ovarian drilling (LOD) in women with clomiphene-resistant polycystic ovary syndrome (PCOS). DESIGN: Prospective controlled study. SETTING: University teaching hospital. PATIENT(S): Twenty-five women with clomiphene-resistant PCOS (group 1) and 20 women with regular menstrual cycles as a comparison group (group 2). INTERVENTION(S): Laparoscopic ovarian drilling. MAIN OUTCOME MEASURE(S): Serum levels of VEGF, IGF-1, and Doppler indices of ovarian stromal blood flow. RESULT(S): The serum levels of VEGF, IGF-1, T, and LH were significantly higher in group 1 before LOD than in group 2. The Doppler indices (pulsatility index and resistance index) of ovarian stromal blood flow were also significantly lower in group 1 before LOD than in group 2. The serum levels of VEGF, T, and LH were significantly reduced in group 1 after LOD compared with in group 1 before LOD. Doppler indices (pulsatility index and resistance index) of ovarian stromal blood flow were significantly increased after LOD. The VEGF levels before LOD were positively correlated with IGF-1, LH, and T. After LOD, the VEGF levels were positively correlated with LH and T. CONCLUSION(S): Higher serum levels of VEGF and IGF-1 may explain the increased vascularity that was demonstrated by Doppler blood flow measurements in PCOS. Laparoscopic ovarian drilling reduced serum VEGF, IGF-1, T, and LH and reduced ovarian blood flow velocities, which may explain the reduction of ovarian hyperstimulation syndrome in women with PCOS after LOD.

Doppler study of the uterine arteries and ovarian stroma in patients with polycystic ovary syndrome.

Doppler analysis of the uterine arteries and ovarian stroma was performed by transvaginal ultrasound in 24 patients with polycystic ovary syndrome (PCOS) and 22 ovulatory women. Vascularization of the ovarian stroma was more abundant in patients with PCOS than in control women, but no significant difference in the mean pulsatility index (PI) was observed between groups (1.14 +/- 0.28 for the PCOS group and 1.05 +/- 0.19 for the control group). The mean PI of the uterine arteries was significantly higher in the PCOS group (PI = 3.7 +/- 0.8) than in the control group (PI = 2.9 +/- 0.4). In the patients with PCOS, no correlation was observed between PI and luteinizing hormone, testosterone or androstenedione levels. Obesity had no effect on uterine artery PI, with no significant differences in this index when the 3 groups were subdivided into obese and non-obese groups.

Ovários policísticos: critérios hemodinâmicos / Polycystic ovaries: hemodynamic criteria

O advento da ultra-sonografia endovaginal de alta resolução abriu novas áreas de pesquisa nos ovários policísticos. O conhecimento da hemodinâmica ovariana é fundamental para o entendimento do comportamento fisiopatológico dos ovários policísticos. Os autores tecem considerações sobre a possibilidade da utilização do Doppler colorido na melhor definição dos ovários policísticos. Os dados sugerem que o aumento da vascularidade e a diminuição da resistência dos vasos do estroma ovariano, assim como o aumento do índice de pulsatilidade das artérias uterinas, possivelmente, são novos parâmetros sonográficos no diagnóstico da síndrome dos ovários policísticos. Esta é uma conclusão preliminar, e se fazem necessários estudos posteriores para confirmar esta proposição.

Doppler, ultrasonographic and endocrinological environment with regard to the number of small subcapsular follicles in polycystic ovary syndrome.

The aim of this study was to evaluate how, in patients with polycystic ovary syndrome, the number of small subcapsular follicles correlates with uterine and ovarian blood flow and with specific hormonal parameters. At an ultrasonographic evaluation, 30 patients with polycystic ovary syndrome showed 5-10 (group I; n = 14) or > 10 (group II; n = 16) small follicles. These patients underwent ultrasonographic (ovarian volume and stroma echodensity; number, diameter and distribution of follicles) and color Doppler (uterine and intraovarian vessels) analyses, and hormonal assay. In group II, significantly lower pulsatility index values than in group I were observed in the ovarian stromal arteries. The Ferriman-Gallwey score, plasma androstenedione level and luteinizing hormone/follicle stimulating hormone (LH/FSH) ratio results were significantly higher in group II than in group I. Androstenedione plasma levels correlated with the number of small follicles. Furthermore, the LH/FSH ratio correlated with both the number of small follicles and the stromal artery pulsatility index. The combined assessment of ovarian morphology by transvaginal ultrasound and color Doppler may provide insight into the pathological state of polycystic ovary syndrome.

The pattern of changes in ovarian stromal and uterine artery blood flow velocities during in vitro fertilization treatment and its relationship with outcome of the cycle.

OBJECTIVES: To assess the effect of short-term (2-3 weeks) pituitary suppression and controlled ovarian stimulation on ovarian and uterine artery Doppler measurements during the in vitro fertilization (IVF) treatment cycle and to compare the pattern of these changes between conception and non-conception cycles as well as between patients with normal and those with polycystic ovaries. DESIGN: Prospective observational study of women undergoing IVF treatment. SUBJECTS: Women using the long-treatment buserelin protocol who did not have uterine fibroids, ovarian cysts or endometrioma. METHODS: Serial transvaginal color and pulsed Doppler measurements of ovarian stromal and uterine artery blood flow velocity were carried out in the early follicular phase of the menstrual cycle, on the day of pituitary suppression and on the day of administration of human chorionic gonadotropin (hCG). The main outcome measures were the ovarian stromal and uterine artery blood flow peak systolic velocity (PSV) and pulsatility index (PI). RESULTS: A total of 105 patients were recruited but six patients were excluded from the analysis because they had only one stage of the measurements performed. There was a significant decline in mean ovarian stromal artery PSV after 2-3 weeks of gonadotropin releasing hormone (GnRH) agonist therapy but no effect on ovarian stromal artery PI. The mean uterine artery PSV or PI did not change significantly after 2-3 weeks of GnRH agonist therapy. There was a significantly higher mean ovarian stromal artery PSV in conception cycles compared to non-conception cycles in the early follicular phase and on the day of pituitary suppression, but not on the day of hCG administration. There were no differences between conception and non-conception cycles in the mean uterine artery PSV or PI. Women with polycystic ovaries had a higher mean ovarian artery PSV on all the three occasions of measurement. CONCLUSION: These data suggest that assessment of ovarian blood flow before commencement of gonadotropin stimulation may play a role in assessing cycles likely to result in pregnancy.