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The role of color doppler imaging in the diagnosis of polycystic ovary syndrome
The role of color Doppler imaging in the diagnosis of polycystic ovary syndrome Cesare
Battaglia,
Alessandro Modena,
D.
MD, Genazzani,
Paolo
G. Artini, MD,
and
VD, Andrea
Gerard0
D’Ambrogio,
R. Genazzani,
MD,
MD, PhD
Italy
OBJECTIVE: Our purpose was to evaluate whether intraovarian and uterine blood flow variations are associated with clinical, ultrasonographic, and endocrine polycystic ovary syndrome findings. STUDY DESIGN: Thirty-two hirsute, oligomenorrheic patients and 18 volunteer women underwent in the early follicular phase ultrasonographic evaluation of ovarian volume, echodensity, and follicle number; transvaginal color Doppler measurement of the uterine and intraovarian vessel variations; and radioimmunologic dosage of luteinizing hormone, follicle-stimulating hormone, estradiol, progesterone, testosterone, androstenedione, and other hormonal compartments. RESULTS: In the patients with polycystic ovary syndrome (increased luteinizing hormone/follicle-stimulating hormone ratio, elevated androstenedione levels, high number of subcapsular follicles by ultrasonography-augmented ovarian volume and echodensity) (n = 22) we observed, at Doppler analysis, significantly elevated uterine artery pulsatility index values associated with a typical low resistance index of stromal ovary vascularization. The pulsatility index was positively correlated with the luteinizing hormone/follicle-stimulating hormone ratio, and the resistance index was negatively correlated. The elevated uterine artery resistance was correlated with androstenedione levels. CONCLUSION: Doppler analysis can be a valuable additional tool for the diagnosis of polycystic ovary syndrome. (AM J OBSTET GYNECOL 1995;172:108-13.) Key
words:
Doppler,
polycystic
ovary syndrome,
ovarian
Polycystic ovary syndrome is a clinical disorder characterized by infertility, oligomenorrhea, hirsutism, acne or seborrhea, and obesity. The typical polycystic ovary has been traditionally described as enlarged and pearly white, with a thick capsule and numerous subcapsular follicles. A high pituitary output of luteinizing hormone (LH) associated to the secretion of excessive amounts of androgens is a frequent finding constituting the basis for chronic anovulation in polycystic ovary syndrome. Recently it has been suggested that the condition is hereditary and that the women with polycystic ovary syndrome have additional metabolic disturbances, such as hyperinsulinemia, increased insulin resistance, and hypertriglyceridemia. Furthermore, polycystic ovary syndrome can result from disturbances of various endocrine systems. It has been diagnosed in patients with Cushing’s syndrome, adrenal hyperplasia, hypothyroidism or hyperthyroidism, adrenal or ovarian tumors, and hyperprolactinemia.’ From the Department of Obstetrics and Gynecology, University of Modena. Received for publicataon Februa y 15, 1994; revised April 21, 1994; accepted June 27, 1994. Reprint requests: Cesare Battaglia, MD, Department of Obstetrics and Gynecology, University of Modena, Via de1 Pozzo, 71.-41100 Modem, Italy. Copyright 0 1995 by Mosby-Year Book, Inc. 0002-9378/95 $3.00 + 0 6/l/58724
vascularization,
hyperandrogenism
Transvaginal color flow Doppler ultrasonography gives us an accurate tool to study the female reproductive system. The reduced distance between the probe and the pelvic structures supplies better resolution, and the empty bladder gives more patient comfort. Transvaginal color Doppler imaging facilitates the detection of small vessels in the uteroovarian circulation and the measurement of impedance to flow in this vascular tree.“*4 The aim of the current study was to determine whether ovarian and uterine blood flow variations are associated with clinical, ultrasonographic, and endocrine findings typical of polycystic ovary syndrome. Material
and
methods
The study protocol was approved by the local Ethics Review Committee. Thirty-two patients with a mean (&SD) age of 24.6 * 2.9 years (range 16 to 29 years) attending the Gynecologic Endocrinological Clinic participated in the study after giving informed consent. Twenty (62.5%) patients were oligomenorrheic (cycle length ~35 days), and 12 (37.5%) were amenorrheic (no vaginal bleeding for t 3 months in individuals who previously had experienced periodic menstruation). All the patients had acne or seborrhea and hirsutism (Ferriman and Gallway score IS). Body mass index was 24.7 kg/m2 (range 20.5 to 37.5 kg/m’).
Volume 172, Number Am J Obstet Gynecol
1, Part
1
Battaglia
Fig. 1. Typical ultrasonographic evidence of polycystic ovary syndrome ovary: subcapsular follicles (10 follicles with maximum diameter < 1 cm), increased ml), and increased echodensity of ovarian stroma (score 2).
The controls consisted of 18 volunteer women with a mean age of 25.2 f 3.4 years (range 17 to 30 years) and with ovulatory cycles documented by ultrasonography and serum progesterone levels (> 5 rig/ml). In the control group the mean body mass index was similar (22.4 kg/m’, range 19.3 to 26.8 kg/m’) to that of the hirsute patients. Neither hirsute nor control patients had received hormonal treatments for ~4 months before the study. Patients were studied in the early follicular phase (cycle days 3 to 5) in controls and oligomenorrheic women or at random in amenorrheic women. Ultrasonographic examination of the ovaries was performed with a 6.5 MHz vaginal transducer (Ansaldo AU 590 Asynchronous, Genoa, Italy). Ovarian volume and follicle distribution, number, and diameter were recorded. The volume was calculated by the formula V = 7~/6 x Dl x D2 X D3, where Dl is the longitudinal diameter, D2 is the anteroposterior diameter, and D3 is the transverse diameter of the ovary. Ovarian stroma echogenicity was scored as 0 (normal), 1 (moderately increased), or 2 (markedly increased).5, 6 Doppler flow measurements of the uterine and intraovarian vessels were performed transvaginally (Ansaldo AU 590 Asynchronous Color Doppler) with a 6.5 MHz pulsed color Doppler system. All the patients rested in a waiting room for at least 15 minutes before being scanned, and they completely emptied the bladder to minimize external effects on blood flow. A 50 Hz filter was used to eliminate low-frequency signals originating from vessel wall movements. By means of color flow imaging, color signals were searched in the ovarian stroma away from the ovary surface. Although several
et al.
109
high number of small ovarian volume (11.1
blood vessels were detected inside the ovarian stroma, only the one with the lowest downstream impedance was selected for Doppler measurements. Color flow images of the ascending branches of uterine arteries were sampled lateral to the cervix in a longitudinal plane. The angle of insonation was always changed to obtain maximum color intensity. When good color signals were obtained, blood flow velocity waveforms were recorded by placing the sample volume across the vessel and entering the pulsed Doppler mode. Different index values (pulsatility and resistance indexes) were used to better compare the data with our own reference ranges. Pulsatility index, defined as the difference between peak systolic and end-diastolic flow velocity divided by the mean flow velocity, was calculated for uterine arteries. Resistance index, defined as the difference between peak systolic and end-diastolic flow divided by the peak systolic flow velocity, was calculated for intraovarian vessels. For each examination the mean value of three consecutive waveforms was obtained. No significant differences between the pulsatility index of the left and right uterine artery was observed, and therefore the average value of both arteries was used. Similarly, the right and left intraovarian lowest resistance index value was not significantly different, and the mean value was used. Ultrasonographic and Doppler analysis were performed by a single examiner (C.B.). The hormonal status of the scanned patients was unknown. An indication of the within-patient precision of the Doppler procedures was obtained by analyzing the flow velocity waveforms recorded on three occasions either from
110
Battaglia
Table
IA.
et al.
Hormonal
January 1995 Am J Obstet Gynecol
and ultrasonographic
findings
Group LH (mu/ml) FSH (mu/ml) LH/FSH ratio Testosterone (ng/lOO ml) A4-Androstenedione (ng/lOO ml) Ovarian volume (ml) Stromal echodensity (%) 0 1 2 Subcapsular follicles/ovary (%) o-5 6-10 > 10 *Comparison tComparison :p < 0.05.
between between
groups group
in hirsute
1 (n = 22)
and control
Group
12.4 k 4.4 4.0 -+ 0.7 3.1 t 0.8 60.7 t 19.9 396.7 -+ 32.7 12.8 t 2.4
2 (n = 8)*
5.4 3.1 1.7 44.2 259.0 4.2
o-+0 18 82
patients
t 2 k i ? ZL
(mean
Controls
3.9x 1.8 0.7s 15.0 51.83 1.9s
(n = 18jt
6.3 6.7 0.9 39.6 212.0 6.4
87.5 -c 10 12.5 f 2 a
+ SD)
t it f f *
Normal
1.9: 1.0 0.29 16.7 69.55 1.85
range
1.5-10 2.9-l 1 1.2-2 30-100 70-300 4-8
94.5 -+ 10 5.5 f 19 05
0 36 64
100s 01 O§
1 and 2. 1 and controls.
§p < 0.01.
Table
IB.
Hormones
in hirsute
and control
patients
I Estradiol (pgiml) Progesterone (rig/ml) 17-Hydroxyprogesterone Dehydroepiandrosterone sulfate (pg/ml) Cortisol (pg/lOO ml) Prolactin (ngiml) Thyroid-stimulating hormone (kU/ml) Free triiodothyronine (pgiml) Free iodothyroxine (pgiml)
(mean -I- SD)
Group 1 (n = 22) 19.4 1.4 1.3 2.8 16.8 14.9 2.0 5.9 11.1
k t f f f f f IL f
Group 2 (n = 8)
I
7.9 1.5 1.0 0.7 4.6 5.4 0.8 1.1 1.5
18.0 0.8 1.3 1.5 19.7 18.6 2.6 5.0 10.5
+ +i i-c 2 f f rt
I
4.1 1.0 0.4 0.7 5.4 7.3 1.3 0.6 2.0
Controls (n = 18) 16.6 f 0.9 f
6.9 0.7 1.0 k 0.4 2.0 f 1.0 19.8 2 6.8 15.1 i- 3.7 2.1 + 1.1 4.9 k 0.8 11.0 + 1.9
I
Nownal range lo-60 0.1-1.5 0.2-1.5 0.3-4.1 5.5-25 2.7-14.6 0.4-4.5 3.5-6.5 6.1-16.7
No signficant differences were observed between groups.
uterine and intraovarian arteries at l-minute intervals. An analysis of variance of the results from 15 patients gave a mean coefficient of variation of 5% for uterine and 7.9% for intraovarian arteries and showed no significant differences between the replicate analyses. Peripheral blood was obtained from all patients between 8 and 11 AM on the same day that ultrasonographic and Doppler examination took place. Many hormonal compartments were analyzed. LH, folliclestimulating hormone (FSH), estradiol, progesterone, 17-hydroxyprogesterone, testosterone, A4-androstenedione, dehydroepiandrosterone sulfate, cortisol, prolactin, thyroid-stimulating hormone, free triiodothyronine, and free iodothyroxine concentrations were determined by a radioimmunoassay (RADIM, Pomezia, Rome). The LH/FSH ratio was hrther calculated. All samples from each subject were analyzed in duplicate in the same assay. On the basis of two quality control samples the average within- and between-assay coefficients of variation were 5.1% and 7.7% for LH; 4.8% and 7.1% for FSH; 4.9% and 7.5% for estradiol, progesterone, 17hydroxyprogesterone, and testosterone, 6.8% and 8.9%
for A4-androstenedione; 5.2% and 7.7% for dehydroepiandrosterone sulfate; 5.2% and 7.5% for cortisol; 3.9% and 6.0% for prolactin; 4.1% and 6.9% for thyroid-stimulating hormone; and 3.8% and 8.8% for free triiodothyronine and free iodothyroxine. In all patients hematocrit and high-density lipoprotein were evaluated. Statistical analysis. Statistical analysis was performed with the unpaired Student t test. Relationships between the analyzed parameters were assessed with stepwise multiple linear regression. A probability of 10.05 was taken as the limit of statistical significance. Data are presented as mean 2 SD unless otherwise indicated. Results Two cluded (increased
hirsute, hyperprolactinemic from the study. On the LH/FSH
ratio
and
elevated
patients were exbasis of hormonal A4-androstene-
dione levels) and ultrasonographic (high number of subcapsular small follicles, augmented ovarian volume, and increased echogenicity of ovarian stroma) results (Fig. 1) we divided the hirsute women in polycystic
Volume 172, Number Am J Obstet Gynecol
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et al.
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Fig. 2. Intraovarian stromal vascularization in polycystic ovary syndrome: small stromal vessels with low downstream impedance (resistance index 0.51).
ovary syndrome patients (group 1, n = 22) and clinically hirsute patients (group 2, n = 8) (Table IA). In the latter group other endocrine disturbances were excluded (Table IB). In group 1 the hematocrit (39.8% k 1.8%) and high-density lipoprotein (47.7 k 15 mg/dl) was significantly different compared with group 2 (35.6% k l.l%, p < 0.05 and 70.1 t 13 mg/dl, p < 0.05, respectively) and controls (36.2% ? 1.5%, p < 0.05, and 74.2 ? 11.2 mgidl, p < 0.05, respectively). At Doppler analysis a significantly higher mean uterine pulsatility index was found in group 1 (3.58 k 0.34) compared with group 2 (2.77 t 0.45, p < 0.01) and controls (2.76 +- 0.54, p < 0.01). At the level of the ovary a stromal vascularization (Fig. 2) was found in group 1 (resistance index = 0.55 k 0.05), except in one case (in both ovaries). Similar findings were not found in group 2 or control patients. Only in one patient in the control group were a few blood vessels with high (resistance index = 0.80) downstream impedance found at the level of ovarian stroma of the left ovary. The presence of stromal ovary vascularization with low resistance had a generally high diagnostic value for polycystic ovary syndrome (Table II). In group 1 the mean uterine artery pulsatility index values were inversely correlated with mean intraovarian artery resistance index values (F = -8.34, p = 0.006). The pulsatility index values were positively correlated with the LH/FSH ratio (F = 5.65, p = 0.02); the resistance index values were negatively correlated (F =
Table II. Diagnostic syndrome of stromal
value for polycystic ovary vascularization Per patient (W
Sensitivity Specificity Positive predictive value Negative predictive value Concordaqce
95.4 96.1 95.4 96.1 95.8
ovary Per ovary @I
95.4 98.0 97.6 96.1 95.8
-4.30, p = 0.04). Furthermore, the A4-androstenedione levels resulted correlated with mean uterine pulsatility index values (F = 5.36, fl = 0.02). No significant correlations were found in other parameters of either group.
Comment This study shows that in patients with polycystic ovary syndrome important changes in ovarian vascularization occur at the level of the intraovarian arteries. Although intraovarian vessels are usually not seen before day 8 to 10 of the 28-day cycle,7, * we detected distinct arteries with characteristic low vascular impedance as early as cycle day 3 to 5. In the studied population the results were typical of polycystic ovary syndrome. Furthermore, we observed a significant inverse correlation between this finding and the LH/FSH ratio. Tonic hypersecretion of LH during the follicular
112
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et al.
January Am J Obstet
phase of the menstrual cycle occurs in polycystic ovary syndrome and is associated in theta cells and stromal hyperplasia with consequent androgen overproduction.” lo Elevated LH levels may be responsible for increased stromal vascularization by different mechanisms that may act individually or in a cumulative way. Neoangiogenesis is a possible hypothesis.“, ” Ovarian tissue from immature rats treated with pregnant mare serum gonadotropins and placed in the chick embryo become vascularized by capillaries of the chorioallantoic membrane.13 Similar effects have been reported in ovaries of mice.14 Another possible mechanism of action of LH is catecholaminergic stimulation.‘5 In fact, noradrenergic fibers innervate the ovarian blood vessels and exhibit a network of varicosities in the cortex and medulla of the ovary, in stromal tissue, and in theta interstitial cells. Electrical stimulation of the ovarian plexus of hypophysectomized rats results in morphologic transformation of the theta interstitial cells, characterized by the assumption of ultrastructural features typical of active steroid-secreting cells. Histochemical evaluation of ovarian tissue obtained from patients with polycystic ovary syndrome revealed markedly enhanced innervation of theta interstitial and stromal cell compartment. Moreover, it is known that the stimulation of the medial basal prechiasmatic area and the ventromedial hypothalamic nucleus exerts a facilitator-y action on the ovaries and increases pituitary LH release, sugesting that both the neural and hormonal elements could act in concert in regulating ovarian secretion. It is conceivable that increased LH secretion may be associated with Pi- or Pa-adrenergic stimuli responsible for contemporary vasodilation and ovarian hyperandrogenism.16 The effect of LH may also be mediated’by leukocyte and cytokine activation.17 Leukocytes and some of their products, the cytokines, appear to play important roles in the physiologic and pathologic mechanisms of the ovary. Leukocytes may be attracted in the ovary by LH-induced synthesis of interleukin-1 and tumor necrosis factor-u. The stromal liberation of vasoactive cytokines associated with mast cell degranulation may be responsible for the ovarian vasodilation. In this study, in group 1 we found higher pulsatility index value at the level of the uterine artery. This finding was correlated with LH/FSH ratio and A4androstenedione levels. In patients with polycystic ovary syndrome higher hematocrit and lower highdensity lipoprotein levels were found. Estrogens are well known to enhance blood flow in many tissues and are inversely correlated with uterine artery pulsatility index values.“, I9 Estrogen action seems mediated by a decreased function of periarterial sympathetic vasoconstrictor nerves.” All the studied patients were in hypoestrogenic state
1995 Gynecol
(estradiol ~30 pgiml); however, the highest pulsatility index values were found in patients with polycystic ovary syndrome. Androgens have direct vasoconstrictive effects on vascular tissues, mediated by specific receptors present in arterial blood vessel walls.21 Furthermore, it seems that thickening and hardening of blood vessels in vascular diseases is a fibrotic process in part mediated by androgen-dependent collagen and elastin deposition in smooth muscle cells.” Probably the process described, associated with elevated hematocrit values, in patients with polycystic ovary syndrome may be responsible for the significant higher resistances registered in uterine arteries. In women with polycystic ovary syndrome there is a significantly reduced chance of conception and an increased risk of miscarriage.“3 LH enables the reactivation of meiosis and hence the attainment of oocyte maturity before ovulation. Inappropriate release of LH might profoundly affect this process so that the released egg either is unable to be fertilized or, if fertilized, miscarriesZ4 We postulated that reduced uterine perfusion may be a further cause that does not allow normal blastocyst implantation.“. 4 Moreover, the current data with the associated alterations in lipoprotein metabolism, as suggested in group 1 by significantly lower HDL values, and the tendency to be overweight may in part explain the well-known predisposition of patients with polycystic ovary syndrome to have atherosclerosis with myocardial infarction and stroke.21. *’ In conclusion, observations from this study indicate that transvaginal color Doppler evaluation of uterine and intraovarian arteries can be added to the traditional endocrinologic and ultrasonographic parameters clinically used for the diagnosis of polycystic ovary syndrome. REFERENCES
1. Yen SSC. Chronic anovulation caused by peripheral endocrine disorders. In: Yen SSC, Jaffe BR, eds. Reproductive endocrinology. Philadelphia: WB Saunders, 1991:576-630. 2. Goswamy RK, Williams G, Steptoe PC. Decreased uterine perfusion-a cause of infertility. Hum Reprod 1988;3: 955-9. 3. Steer CV, Campbell S, Pampiglione JS, Kingsland CR, Mason BA, Collins WP. Transvaginal colour flow imaging of the uterine arteries during the ovarian and menstrual cycles. Hum Reprod 1990;5:391-5. 4. Battaglia C, Larocca E, Lanzani A, Valentini M, Genazzani AR. Doppler ultrasound studies of the uterine arteries in spontaneous and IVF stimulated ovarian cycles. Gynecol Endocrinol 1990;4:245-50. 5. Pache TD, WIadimiroff lW, Hop WCl, Fauser BClM. How to discriminate between nor&al and polycystic ovaries: transvaginal US studv. Radiolocrv 1992;183:421-3. 6. Pache ?D, de Jong FH, Hop Wk, Fauser BCJM. Association between ovarian changes assessed by transvaginal sonography and clinical and endocrine signs of the polycystic ovary syndrome. Fertil Steril 1993;59:544-9. 7. Merce LT, Garces D, Barco MJ, de la Fuente F. Intraovar-
Volume 172, Number Am J Obstet Gynecol
8.
9.
10.
11. 12.
13.
14.
15.
16.
1, Part
1
ian Doppler velocimetry in ovulatory, dysovulatory and anovulatory cycles. Ultrasound Obstet Gynecol 1992;2: 197-202. Sladkevicius P, Valentin L, Larsal K. Blood flow velocity in the uterine and ovarian arteries during the normal menstrual cycle. Ultrasound Obstet Gynecol 1993;3:199-208. Buyalos RP, Bergman RN, Geffner ME, Gornbein JA, Watanabe JM, Judd HL. The influence of luteinizing hormone and insulin on sex steroids and sex hormonebinding globulin in the polycystic ovarian syndrome. Fertil Steril 1993;60:626-33. Yoshino K, Takahashi K, Eda Y, Okada S, Kitao M. Endocrinological environment with regard to the number of microcysts in patients with polycystic ovary syndrome. Hum Reprod 1992;7:1201-4. Findlay JK. Angiogenesis in reproductive tissues. J Endocrinol 1986;111:357-66. Kurjak A, Kupesic-Urek S, Schulman H, Zalad I. Transvaginal color flow Doppler in the assessment of ovarian and uterine blood flow in infertile women. Fertil Steril 1991;56:870-3. Koos RD, Le Maire WJ. Factors that may regulate the growth and regression of blood vessels in the ovary. Semin Reprod Endocrinol 1983;1:295-307. Sato E, Ishibashi J, Koide SS. Inducement of blood vessel formation by ovarian extracts from mice injected with gonadotrophins. Experientia 1982;38:1248-9. Kawakami M, Kubo K, Uemura T, Nagase M, Hayashi R. Involvement of ovarian innervation in steroid secretion. Endocrinology 1981;109:136-45. Hernandez ER, Jemenez JL, Payne DW, Adashi E. Adrenergic regulation of ovarian androgen biosynthesis is mediated via BB-adrenergic theta-interstitial cell recognition sites. Endocrinology 1988;122:1592-602.
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et al.
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17. Brannstrom M, Norman RJ. Involvment of leukocytes and cytokines in the ovulatory process and corpus luteum function. Hum Reprod 1993;8:1762-75. 18. Dickson WM, Bose MJ, Locatelli A. Effect of estrogen and progesterone on uterine blood flow in castrate sows. Am J Physiol 1969;217:1431-6. 19. Killam AP, Rosenfeld CR, Battaglia FC, Makowski EL, Meschia G. Effect of estrogens on the uterine blood flow of oophorectomized ewes. AM J OBSTET GYNECOL 1973;115: 1045-5 1. 20. Ford SP, Reynolds LP, Farley DB, Bhatngar RK, Van Orden DE. Interaction of ovarian steroids and periarterial or,-adrenergic receptors in altering uterine blood flow during the estrous cycle of gilts. AM J OBSTET GYNECOL 1984;150:480-4. 21. Horwitz KB, Horwitz LD. Canine vascular tissue are targets for androgens, estrogens, progestins, and glucocorticoids. J Clin Invest 1982;69:740-8. 22. Fisher GM, Swain ML. Effect of sex hormones on blood pressure and vascular connective tissue in castrated and non castrated rats. Am J Physiol 1977;232:617-21. 23. Chappel SC, Howles C. Reevaluation of the roles of luteinizing hormone and follicle-stimulating hormone in the ovulatory process. Hum Rerod 1991;6:1206-12. 24. Balen AH. Hypersecretion of luteinizing hormone and the polycystic ovary syndrome. Hum Reprod 1993;8(suppl 2): 123-8. 25. Dahlgren E, Janson PO, Johansson S, Lapidus L, Oden A. Polycystic ovary syndrome and risk for myocardial infarction-evaluated from a risk factor model based on a prospective population study of women. Acta Obstet Gynecol Stand 1992;71:599-604.
Battaglia,
Alessandro Modena,
D.
MD, Genazzani,
Paolo
G. Artini, MD,
and
VD, Andrea
Gerard0
D’Ambrogio,
R. Genazzani,
MD,
MD, PhD
Italy
OBJECTIVE: Our purpose was to evaluate whether intraovarian and uterine blood flow variations are associated with clinical, ultrasonographic, and endocrine polycystic ovary syndrome findings. STUDY DESIGN: Thirty-two hirsute, oligomenorrheic patients and 18 volunteer women underwent in the early follicular phase ultrasonographic evaluation of ovarian volume, echodensity, and follicle number; transvaginal color Doppler measurement of the uterine and intraovarian vessel variations; and radioimmunologic dosage of luteinizing hormone, follicle-stimulating hormone, estradiol, progesterone, testosterone, androstenedione, and other hormonal compartments. RESULTS: In the patients with polycystic ovary syndrome (increased luteinizing hormone/follicle-stimulating hormone ratio, elevated androstenedione levels, high number of subcapsular follicles by ultrasonography-augmented ovarian volume and echodensity) (n = 22) we observed, at Doppler analysis, significantly elevated uterine artery pulsatility index values associated with a typical low resistance index of stromal ovary vascularization. The pulsatility index was positively correlated with the luteinizing hormone/follicle-stimulating hormone ratio, and the resistance index was negatively correlated. The elevated uterine artery resistance was correlated with androstenedione levels. CONCLUSION: Doppler analysis can be a valuable additional tool for the diagnosis of polycystic ovary syndrome. (AM J OBSTET GYNECOL 1995;172:108-13.) Key
words:
Doppler,
polycystic
ovary syndrome,
ovarian
Polycystic ovary syndrome is a clinical disorder characterized by infertility, oligomenorrhea, hirsutism, acne or seborrhea, and obesity. The typical polycystic ovary has been traditionally described as enlarged and pearly white, with a thick capsule and numerous subcapsular follicles. A high pituitary output of luteinizing hormone (LH) associated to the secretion of excessive amounts of androgens is a frequent finding constituting the basis for chronic anovulation in polycystic ovary syndrome. Recently it has been suggested that the condition is hereditary and that the women with polycystic ovary syndrome have additional metabolic disturbances, such as hyperinsulinemia, increased insulin resistance, and hypertriglyceridemia. Furthermore, polycystic ovary syndrome can result from disturbances of various endocrine systems. It has been diagnosed in patients with Cushing’s syndrome, adrenal hyperplasia, hypothyroidism or hyperthyroidism, adrenal or ovarian tumors, and hyperprolactinemia.’ From the Department of Obstetrics and Gynecology, University of Modena. Received for publicataon Februa y 15, 1994; revised April 21, 1994; accepted June 27, 1994. Reprint requests: Cesare Battaglia, MD, Department of Obstetrics and Gynecology, University of Modena, Via de1 Pozzo, 71.-41100 Modem, Italy. Copyright 0 1995 by Mosby-Year Book, Inc. 0002-9378/95 $3.00 + 0 6/l/58724
vascularization,
hyperandrogenism
Transvaginal color flow Doppler ultrasonography gives us an accurate tool to study the female reproductive system. The reduced distance between the probe and the pelvic structures supplies better resolution, and the empty bladder gives more patient comfort. Transvaginal color Doppler imaging facilitates the detection of small vessels in the uteroovarian circulation and the measurement of impedance to flow in this vascular tree.“*4 The aim of the current study was to determine whether ovarian and uterine blood flow variations are associated with clinical, ultrasonographic, and endocrine findings typical of polycystic ovary syndrome. Material
and
methods
The study protocol was approved by the local Ethics Review Committee. Thirty-two patients with a mean (&SD) age of 24.6 * 2.9 years (range 16 to 29 years) attending the Gynecologic Endocrinological Clinic participated in the study after giving informed consent. Twenty (62.5%) patients were oligomenorrheic (cycle length ~35 days), and 12 (37.5%) were amenorrheic (no vaginal bleeding for t 3 months in individuals who previously had experienced periodic menstruation). All the patients had acne or seborrhea and hirsutism (Ferriman and Gallway score IS). Body mass index was 24.7 kg/m2 (range 20.5 to 37.5 kg/m’).
Volume 172, Number Am J Obstet Gynecol
1, Part
1
Battaglia
Fig. 1. Typical ultrasonographic evidence of polycystic ovary syndrome ovary: subcapsular follicles (10 follicles with maximum diameter < 1 cm), increased ml), and increased echodensity of ovarian stroma (score 2).
The controls consisted of 18 volunteer women with a mean age of 25.2 f 3.4 years (range 17 to 30 years) and with ovulatory cycles documented by ultrasonography and serum progesterone levels (> 5 rig/ml). In the control group the mean body mass index was similar (22.4 kg/m’, range 19.3 to 26.8 kg/m’) to that of the hirsute patients. Neither hirsute nor control patients had received hormonal treatments for ~4 months before the study. Patients were studied in the early follicular phase (cycle days 3 to 5) in controls and oligomenorrheic women or at random in amenorrheic women. Ultrasonographic examination of the ovaries was performed with a 6.5 MHz vaginal transducer (Ansaldo AU 590 Asynchronous, Genoa, Italy). Ovarian volume and follicle distribution, number, and diameter were recorded. The volume was calculated by the formula V = 7~/6 x Dl x D2 X D3, where Dl is the longitudinal diameter, D2 is the anteroposterior diameter, and D3 is the transverse diameter of the ovary. Ovarian stroma echogenicity was scored as 0 (normal), 1 (moderately increased), or 2 (markedly increased).5, 6 Doppler flow measurements of the uterine and intraovarian vessels were performed transvaginally (Ansaldo AU 590 Asynchronous Color Doppler) with a 6.5 MHz pulsed color Doppler system. All the patients rested in a waiting room for at least 15 minutes before being scanned, and they completely emptied the bladder to minimize external effects on blood flow. A 50 Hz filter was used to eliminate low-frequency signals originating from vessel wall movements. By means of color flow imaging, color signals were searched in the ovarian stroma away from the ovary surface. Although several
et al.
109
high number of small ovarian volume (11.1
blood vessels were detected inside the ovarian stroma, only the one with the lowest downstream impedance was selected for Doppler measurements. Color flow images of the ascending branches of uterine arteries were sampled lateral to the cervix in a longitudinal plane. The angle of insonation was always changed to obtain maximum color intensity. When good color signals were obtained, blood flow velocity waveforms were recorded by placing the sample volume across the vessel and entering the pulsed Doppler mode. Different index values (pulsatility and resistance indexes) were used to better compare the data with our own reference ranges. Pulsatility index, defined as the difference between peak systolic and end-diastolic flow velocity divided by the mean flow velocity, was calculated for uterine arteries. Resistance index, defined as the difference between peak systolic and end-diastolic flow divided by the peak systolic flow velocity, was calculated for intraovarian vessels. For each examination the mean value of three consecutive waveforms was obtained. No significant differences between the pulsatility index of the left and right uterine artery was observed, and therefore the average value of both arteries was used. Similarly, the right and left intraovarian lowest resistance index value was not significantly different, and the mean value was used. Ultrasonographic and Doppler analysis were performed by a single examiner (C.B.). The hormonal status of the scanned patients was unknown. An indication of the within-patient precision of the Doppler procedures was obtained by analyzing the flow velocity waveforms recorded on three occasions either from
110
Battaglia
Table
IA.
et al.
Hormonal
January 1995 Am J Obstet Gynecol
and ultrasonographic
findings
Group LH (mu/ml) FSH (mu/ml) LH/FSH ratio Testosterone (ng/lOO ml) A4-Androstenedione (ng/lOO ml) Ovarian volume (ml) Stromal echodensity (%) 0 1 2 Subcapsular follicles/ovary (%) o-5 6-10 > 10 *Comparison tComparison :p < 0.05.
between between
groups group
in hirsute
1 (n = 22)
and control
Group
12.4 k 4.4 4.0 -+ 0.7 3.1 t 0.8 60.7 t 19.9 396.7 -+ 32.7 12.8 t 2.4
2 (n = 8)*
5.4 3.1 1.7 44.2 259.0 4.2
o-+0 18 82
patients
t 2 k i ? ZL
(mean
Controls
3.9x 1.8 0.7s 15.0 51.83 1.9s
(n = 18jt
6.3 6.7 0.9 39.6 212.0 6.4
87.5 -c 10 12.5 f 2 a
+ SD)
t it f f *
Normal
1.9: 1.0 0.29 16.7 69.55 1.85
range
1.5-10 2.9-l 1 1.2-2 30-100 70-300 4-8
94.5 -+ 10 5.5 f 19 05
0 36 64
100s 01 O§
1 and 2. 1 and controls.
§p < 0.01.
Table
IB.
Hormones
in hirsute
and control
patients
I Estradiol (pgiml) Progesterone (rig/ml) 17-Hydroxyprogesterone Dehydroepiandrosterone sulfate (pg/ml) Cortisol (pg/lOO ml) Prolactin (ngiml) Thyroid-stimulating hormone (kU/ml) Free triiodothyronine (pgiml) Free iodothyroxine (pgiml)
(mean -I- SD)
Group 1 (n = 22) 19.4 1.4 1.3 2.8 16.8 14.9 2.0 5.9 11.1
k t f f f f f IL f
Group 2 (n = 8)
I
7.9 1.5 1.0 0.7 4.6 5.4 0.8 1.1 1.5
18.0 0.8 1.3 1.5 19.7 18.6 2.6 5.0 10.5
+ +i i-c 2 f f rt
I
4.1 1.0 0.4 0.7 5.4 7.3 1.3 0.6 2.0
Controls (n = 18) 16.6 f 0.9 f
6.9 0.7 1.0 k 0.4 2.0 f 1.0 19.8 2 6.8 15.1 i- 3.7 2.1 + 1.1 4.9 k 0.8 11.0 + 1.9
I
Nownal range lo-60 0.1-1.5 0.2-1.5 0.3-4.1 5.5-25 2.7-14.6 0.4-4.5 3.5-6.5 6.1-16.7
No signficant differences were observed between groups.
uterine and intraovarian arteries at l-minute intervals. An analysis of variance of the results from 15 patients gave a mean coefficient of variation of 5% for uterine and 7.9% for intraovarian arteries and showed no significant differences between the replicate analyses. Peripheral blood was obtained from all patients between 8 and 11 AM on the same day that ultrasonographic and Doppler examination took place. Many hormonal compartments were analyzed. LH, folliclestimulating hormone (FSH), estradiol, progesterone, 17-hydroxyprogesterone, testosterone, A4-androstenedione, dehydroepiandrosterone sulfate, cortisol, prolactin, thyroid-stimulating hormone, free triiodothyronine, and free iodothyroxine concentrations were determined by a radioimmunoassay (RADIM, Pomezia, Rome). The LH/FSH ratio was hrther calculated. All samples from each subject were analyzed in duplicate in the same assay. On the basis of two quality control samples the average within- and between-assay coefficients of variation were 5.1% and 7.7% for LH; 4.8% and 7.1% for FSH; 4.9% and 7.5% for estradiol, progesterone, 17hydroxyprogesterone, and testosterone, 6.8% and 8.9%
for A4-androstenedione; 5.2% and 7.7% for dehydroepiandrosterone sulfate; 5.2% and 7.5% for cortisol; 3.9% and 6.0% for prolactin; 4.1% and 6.9% for thyroid-stimulating hormone; and 3.8% and 8.8% for free triiodothyronine and free iodothyroxine. In all patients hematocrit and high-density lipoprotein were evaluated. Statistical analysis. Statistical analysis was performed with the unpaired Student t test. Relationships between the analyzed parameters were assessed with stepwise multiple linear regression. A probability of 10.05 was taken as the limit of statistical significance. Data are presented as mean 2 SD unless otherwise indicated. Results Two cluded (increased
hirsute, hyperprolactinemic from the study. On the LH/FSH
ratio
and
elevated
patients were exbasis of hormonal A4-androstene-
dione levels) and ultrasonographic (high number of subcapsular small follicles, augmented ovarian volume, and increased echogenicity of ovarian stroma) results (Fig. 1) we divided the hirsute women in polycystic
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Fig. 2. Intraovarian stromal vascularization in polycystic ovary syndrome: small stromal vessels with low downstream impedance (resistance index 0.51).
ovary syndrome patients (group 1, n = 22) and clinically hirsute patients (group 2, n = 8) (Table IA). In the latter group other endocrine disturbances were excluded (Table IB). In group 1 the hematocrit (39.8% k 1.8%) and high-density lipoprotein (47.7 k 15 mg/dl) was significantly different compared with group 2 (35.6% k l.l%, p < 0.05 and 70.1 t 13 mg/dl, p < 0.05, respectively) and controls (36.2% ? 1.5%, p < 0.05, and 74.2 ? 11.2 mgidl, p < 0.05, respectively). At Doppler analysis a significantly higher mean uterine pulsatility index was found in group 1 (3.58 k 0.34) compared with group 2 (2.77 t 0.45, p < 0.01) and controls (2.76 +- 0.54, p < 0.01). At the level of the ovary a stromal vascularization (Fig. 2) was found in group 1 (resistance index = 0.55 k 0.05), except in one case (in both ovaries). Similar findings were not found in group 2 or control patients. Only in one patient in the control group were a few blood vessels with high (resistance index = 0.80) downstream impedance found at the level of ovarian stroma of the left ovary. The presence of stromal ovary vascularization with low resistance had a generally high diagnostic value for polycystic ovary syndrome (Table II). In group 1 the mean uterine artery pulsatility index values were inversely correlated with mean intraovarian artery resistance index values (F = -8.34, p = 0.006). The pulsatility index values were positively correlated with the LH/FSH ratio (F = 5.65, p = 0.02); the resistance index values were negatively correlated (F =
Table II. Diagnostic syndrome of stromal
value for polycystic ovary vascularization Per patient (W
Sensitivity Specificity Positive predictive value Negative predictive value Concordaqce
95.4 96.1 95.4 96.1 95.8
ovary Per ovary @I
95.4 98.0 97.6 96.1 95.8
-4.30, p = 0.04). Furthermore, the A4-androstenedione levels resulted correlated with mean uterine pulsatility index values (F = 5.36, fl = 0.02). No significant correlations were found in other parameters of either group.
Comment This study shows that in patients with polycystic ovary syndrome important changes in ovarian vascularization occur at the level of the intraovarian arteries. Although intraovarian vessels are usually not seen before day 8 to 10 of the 28-day cycle,7, * we detected distinct arteries with characteristic low vascular impedance as early as cycle day 3 to 5. In the studied population the results were typical of polycystic ovary syndrome. Furthermore, we observed a significant inverse correlation between this finding and the LH/FSH ratio. Tonic hypersecretion of LH during the follicular
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phase of the menstrual cycle occurs in polycystic ovary syndrome and is associated in theta cells and stromal hyperplasia with consequent androgen overproduction.” lo Elevated LH levels may be responsible for increased stromal vascularization by different mechanisms that may act individually or in a cumulative way. Neoangiogenesis is a possible hypothesis.“, ” Ovarian tissue from immature rats treated with pregnant mare serum gonadotropins and placed in the chick embryo become vascularized by capillaries of the chorioallantoic membrane.13 Similar effects have been reported in ovaries of mice.14 Another possible mechanism of action of LH is catecholaminergic stimulation.‘5 In fact, noradrenergic fibers innervate the ovarian blood vessels and exhibit a network of varicosities in the cortex and medulla of the ovary, in stromal tissue, and in theta interstitial cells. Electrical stimulation of the ovarian plexus of hypophysectomized rats results in morphologic transformation of the theta interstitial cells, characterized by the assumption of ultrastructural features typical of active steroid-secreting cells. Histochemical evaluation of ovarian tissue obtained from patients with polycystic ovary syndrome revealed markedly enhanced innervation of theta interstitial and stromal cell compartment. Moreover, it is known that the stimulation of the medial basal prechiasmatic area and the ventromedial hypothalamic nucleus exerts a facilitator-y action on the ovaries and increases pituitary LH release, sugesting that both the neural and hormonal elements could act in concert in regulating ovarian secretion. It is conceivable that increased LH secretion may be associated with Pi- or Pa-adrenergic stimuli responsible for contemporary vasodilation and ovarian hyperandrogenism.16 The effect of LH may also be mediated’by leukocyte and cytokine activation.17 Leukocytes and some of their products, the cytokines, appear to play important roles in the physiologic and pathologic mechanisms of the ovary. Leukocytes may be attracted in the ovary by LH-induced synthesis of interleukin-1 and tumor necrosis factor-u. The stromal liberation of vasoactive cytokines associated with mast cell degranulation may be responsible for the ovarian vasodilation. In this study, in group 1 we found higher pulsatility index value at the level of the uterine artery. This finding was correlated with LH/FSH ratio and A4androstenedione levels. In patients with polycystic ovary syndrome higher hematocrit and lower highdensity lipoprotein levels were found. Estrogens are well known to enhance blood flow in many tissues and are inversely correlated with uterine artery pulsatility index values.“, I9 Estrogen action seems mediated by a decreased function of periarterial sympathetic vasoconstrictor nerves.” All the studied patients were in hypoestrogenic state
1995 Gynecol
(estradiol ~30 pgiml); however, the highest pulsatility index values were found in patients with polycystic ovary syndrome. Androgens have direct vasoconstrictive effects on vascular tissues, mediated by specific receptors present in arterial blood vessel walls.21 Furthermore, it seems that thickening and hardening of blood vessels in vascular diseases is a fibrotic process in part mediated by androgen-dependent collagen and elastin deposition in smooth muscle cells.” Probably the process described, associated with elevated hematocrit values, in patients with polycystic ovary syndrome may be responsible for the significant higher resistances registered in uterine arteries. In women with polycystic ovary syndrome there is a significantly reduced chance of conception and an increased risk of miscarriage.“3 LH enables the reactivation of meiosis and hence the attainment of oocyte maturity before ovulation. Inappropriate release of LH might profoundly affect this process so that the released egg either is unable to be fertilized or, if fertilized, miscarriesZ4 We postulated that reduced uterine perfusion may be a further cause that does not allow normal blastocyst implantation.“. 4 Moreover, the current data with the associated alterations in lipoprotein metabolism, as suggested in group 1 by significantly lower HDL values, and the tendency to be overweight may in part explain the well-known predisposition of patients with polycystic ovary syndrome to have atherosclerosis with myocardial infarction and stroke.21. *’ In conclusion, observations from this study indicate that transvaginal color Doppler evaluation of uterine and intraovarian arteries can be added to the traditional endocrinologic and ultrasonographic parameters clinically used for the diagnosis of polycystic ovary syndrome. REFERENCES
1. Yen SSC. Chronic anovulation caused by peripheral endocrine disorders. In: Yen SSC, Jaffe BR, eds. Reproductive endocrinology. Philadelphia: WB Saunders, 1991:576-630. 2. Goswamy RK, Williams G, Steptoe PC. Decreased uterine perfusion-a cause of infertility. Hum Reprod 1988;3: 955-9. 3. Steer CV, Campbell S, Pampiglione JS, Kingsland CR, Mason BA, Collins WP. Transvaginal colour flow imaging of the uterine arteries during the ovarian and menstrual cycles. Hum Reprod 1990;5:391-5. 4. Battaglia C, Larocca E, Lanzani A, Valentini M, Genazzani AR. Doppler ultrasound studies of the uterine arteries in spontaneous and IVF stimulated ovarian cycles. Gynecol Endocrinol 1990;4:245-50. 5. Pache TD, WIadimiroff lW, Hop WCl, Fauser BClM. How to discriminate between nor&al and polycystic ovaries: transvaginal US studv. Radiolocrv 1992;183:421-3. 6. Pache ?D, de Jong FH, Hop Wk, Fauser BCJM. Association between ovarian changes assessed by transvaginal sonography and clinical and endocrine signs of the polycystic ovary syndrome. Fertil Steril 1993;59:544-9. 7. Merce LT, Garces D, Barco MJ, de la Fuente F. Intraovar-
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