Polycystic Ovary Syndrome Dr. Heidi Fritz 17 June 2013 Polycystic ovary syndrome - a complex problem By: Heidi Fritz MA, ND Bolton Naturopathic Clinic 64 King St W, Bolton, ON L7E1C7 www.boltonnaturopathic.ca email@example.com Jump to: Part 1 Part 2 Part 3Part 4 Part I: What is Polycystic Ovary Syndrome? Polycystic ovary syndrome (PCOS) is a common women’s health concern, affecting between 5-20% of women of reproductive age.(1) Polycystic ovary syndrome is so named as a reflection of the multiple cysts that form in the ovaries of these women, and which are visible on ultrasound. Although PCOS primarily affects ovarian function, it is important to note that it is a diffuse syndrome affecting many aspects of a woman’s physical function, including skin, hair, cardiovascular health, and risk of diabetes. Having PCOS carries long-term health implications. Furthermore, because it masquerades as so many symptoms, PCOS can be challenging to diagnose. This article discusses the underlying biochemical problems contributing to PCOS, health implications, and how natural medicines can help. Signs and symptoms of PCOS can include: irregular menstrual cycles (oligomenorrhea), being overweight, acne, male-pattern hair growth (hirsuitism), loss of hair from the scalp (alopecia), and darkening of the skin in areas behind the neck or elbow creases (acanthosis nigricans).(2) It is important to note that irregular or missed menstrual cycles are actually an indicator that ovulation is not occurring, and this leads to problems with fertility in many women with PCOS.(3) In fact, what is happening is that as the follicle cells develop in the ovary, they fail to grow appropriately to release an egg, but grow into large unruptured cysts instead; they do regress over time but cause the cystic appearance of the ovary on ultrasound. The criteria for diagnosing PCOS are twofold: 1) the presence of hyperandrogenism, and 2) ovarian dysfunction, with the exclusion of other related disorders.(2) Hyperandrogenism refers to an increase in either blood levels of free testosterone and/ or the hormone DHEAS; or it can refer to symptoms of excess testosterone activity such as acne, excess hair growth, or hair loss, without elevated blood levels. Ovarian dysfunction refers to either lack of ovulation (anovulation), defined as greater than 35 days between cycles or under 10 menses per year;(2) or the presence of at least 12 cysts on the ovary on ultrasound. The manifestations of PCOS listed above can therefore be explained by two common underlying biochemical problems present among these women. First, women with PCOS are affected by inherent defects of androgen hormone synthesis and metabolism. This is not yet well studied, however, it appears that these women may overproduce DHEA or testosterone from the adrenal glands and ovaries.(2, 4) In addition, the ovaries of women with PCOS express higher numbers of androgen hormone receptors, meaning that these women may also be hyper-responsive to DHEA and/ or testosterone.(5) Secondly, most women affected by PCOS suffer from insulin resistance.(6) This means that the body needs to over-produce the hormone insulin in order to properly control blood glucose levels. Insulin is the hormone that allows the cells to use glucose from the blood. After a meal, it is normal for blood glucose to increase; however in response the body secretes insulin, which tells the cells to take glucose up out of the blood, and this results in normalization of blood glucose levels. In women with PCOS, the cells do not respond efficiently to insulin, causing the body to secrete increasing amounts in order to compensate. We can compare this to a mom yelling more loudly at her teenager when he or she refuses to listen! This is important because high insulin is akey contributor to both the ovulation defects and excess androgens (testosterone, DHEA) characteristic of PCOS, and strategies to increase insulin sensitivity play a crucial role in managing PCOS. In fact, medical management of PCOS is primarily directed toward regulating insulin with the medication, metformin. The two most commonly prescribed medications for PCOS include the birth control pill in order to regulate menstruation, and metformin to increase insulin sensitivity. It is important to realize that while the birth control pill induces menses, it does not correct ovulation problems, and may worsen insulin sensitivity .(7) Correcting insulin resistance and ovarian dysfunction through nutritional strategies offers a comprehensive way to address the underlying cause of PCOS and reduce long-term risk of diabetes and heart disease. References 1. Yildiz BO, Bozdag G, Yapici Z, Esinler I, Yarali H. Prevalence, phenotype and cardiometabolic risk of polycystic ovary syndrome under different diagnostic criteria. Hum Reprod. 2012;27(10):3067-73. 2. Azziz R, Carmina E, Dewailly D, Diamanti-Kandarakis E, Escobar-Morreale HF, Futterweit W, et al.; Task Force on the Phenotype of the Polycystic Ovary Syndrome of The Androgen Excess and PCOS Society. The Androgen Excess and PCOS Society criteria for the polycystic ovary syndrome: the complete task force report. FertilSteril. 2009;91(2):456-88. 3. Roy KK, Baruah J, Singla S, Sharma JB, Singh N, Jain SK, et al. A prospective randomized trial comparing the efficacy of Letrozole and Clomiphene citrate in induction of ovulation in polycystic ovarian syndrome. J Hum Reprod Sci. 2012;5(1):20-5. 4. Hogg K, Young JM, Oliver EM, Souza CJ, McNeilly AS, Duncan WC. Enhanced thecal androgen production is prenatally programmed in an ovine model of polycystic ovary syndrome. Endocrinology. 2012;153(1):450-61. 5. Catteau-Jonard S, Jamin SP, Leclerc A, Gonzalès J, Dewailly D, di Clemente N. Anti-Mullerian hormone, its receptor, FSH receptor, and androgen receptor genes are overexpressed by granulosa cells from stimulated follicles in women with polycystic ovary syndrome. J ClinEndocrinolMetab. 2008;93(11):4456-61. 6. Stepto NK, Cassar S, Joham AE, Hutchison SK, Harrison CL, Goldstein RF, et al. Women with polycystic ovary syndrome have intrinsic insulin resistance on euglycaemic-hyperinsulaemic clamp. Hum Reprod. 2013;28(3):777-84. 7. Watanabe RM, Azen CG, Roy S, Perlman JA, Bergman RN. Defects in carbohydrate metabolism in oral contraceptive users without apparent metabolic risk factors.JClinEndocrinolMetab. 1994;79(5):1277-83. Polycystic ovary syndrome - a complex problem Part II The Role of Diet in Insulin Resistance by: Heidy Fritz, MA, ND Bolton Naturopathic Clinic 64 King St W, Bolton, ON L7E1C7 www.boltonnaturopathic.ca firstname.lastname@example.org The goals of a therapeutic diet for PCOS are two-fold: 1) improve insulin sensitivity, which as we discussed in Part I, is a major factor contributing to anovulation and hyperandrogenism; and 2) assisting with moderate weight loss, if appropriate. In the vast majority of women, these two endpoints are closely related. For instance, even a small degree of healthy weight loss will in itself lead to improvements in insulin sensitivity in most people, apart from specific diet changes that we will discuss.(1,2) However, a small number of women with PCOS suffer from what is called thin-variant PCOS.(3) These women are not overweight, and therefore weight is not necessarily related to their problems with insulin sensitivity; however following the principles of a low-glycemic diet, such as we will describe below, will nonetheless improve their insulin sensitivity.(3) First, we will discuss the principles of healthy weight loss, followed by a discussion of glycemic index application. Healthy weight loss is achieved by inducing a state of caloric deficiency relative to caloric output. The word “calorie” or caloric simply refers to a measure of energy; certain types of foods have more energy than others. If more energy is consumed than is used on a day-to-day basis, then the net excess is stored as fat. A safe and sustainable rate of weight loss is between 1-2 lb per week; more than this will not likely last. To lose the equivalent of 1lb of fat tissue, a deficiency of 3500 kilocalories needs to be created. By comparison, 1 cup of salad (before dressing) contains about 40 kcal; one slice of bread contains roughly 100 kcal; and 1 cup ice cream contains 250 kcal. On average, a woman requires 2000 kcal per day to maintain her body weight. Therefore, to achieve weight loss of 1lb per week, she needs to restrict her caloric intake to 1500 kcal per day. This creates a deficiency of 500 kcal per day, which over a period of seven days equals 3500 kcal. This woman therefore needs to keep track of the types and amounts of foods that she eats on a daily basis in order to meet this number. This may sound daunting, however a healthcare provider trained in nutrition, such as a naturopathic doctor, can help you put together a diet with meal options that you can combine throughout the day in order to achieve this number; having a meal plan with options laid out can reduce some of the scare factor associated with such an undertaking, but leaves you with a clear program. The second crucial component of healthy weight loss is engaging in moderate exercise for at least 30 minutes each day. This does not mean that someone who has not been active in years should go take up marathon running. Something as simple as going for a ½ hour brisk walk (but not speed walking!) increases your caloric expenditure, burns fat, maintains muscle mass, and elevates you metabolic rate – allowing you to burn more fat even after you finish exercising! Exercise will make you feel better overall so you won’t feel the same need for habitual “comfort foods,” and it will make you feel energized, which will make your diet program much easier to stick with. If someone is going to engage in a weight loss program, there must, must, must be a commitment to daily exercise. The other important aspect of exercise for women with PCOS is that it increases insulin sensitivity, independently of weight loss.(4) Exercise induces changes at the level of the muscle cells themselves that allows them to respond to insulin more effectively, upregulating the GLUT-4 insulin receptor on the cell surface that allows that cell to take up more glucose.(5) In addition to the general principles of weight loss described above, caloric restriction and exercise, another factor to consider with respect to PCOS is the concept of glycemic load. Simply put, glycemic load refers to how quickly the food we eat is transformed to “sugar” or glucose in the blood. Glycemic load also refers to how much glucose it is turned into, with a small amount being less problematic than large amounts. Eating a meal high in simple sugars or refined carbohydrates (ie. pastries, bread, pasta, white rice) leads to large and rapid spikes in blood glucose levels.In order to control this, the body must secrete large amounts of insulin, telling the cells to take up this glucose and get it out of the blood. However, not only do these cells store the glucose as fat, but this increased insulin production complicates the underlying problem facing women with PCOS: high insulin levels and insulin resistance. A recent study has shown that any diet resulting in weight loss will result in net benefit on PCOS.(6) However, a diet limiting sources of refined carbohydrates and emphasizing carbohydrates from nuts, legumes, fruits, and vegetables instead results in greater reductions in insulin resistance in women with PCOS compared to the conventional diet.(6,7) This kind of low-glycemic diet may also be of particular benefit among thin women with PCOS. References 1. Kahleova H, Mari A, Nofrate V, Matoulek M, Kazdova L, Hill M, et al. Improvement in β-cell function after diet-induced weight loss is associated with decrease in pancreatic polypeptide in subjects with type diabetes. J Diabetes Complications. 2012;26(5):442-9. 2. Petersen KF, Dufour S, Morino K, Yoo PS, Cline GW, Shulman GI.Reversal of muscle insulin resistance by weight reduction in young, lean, insulin-resistant offspring of parents with type 2 diabetes.ProcNatlAcadSci U S A. 2012;109(21):8236-40. 3. Stepto NK, Cassar S, Joham AE, Hutchison SK, Harrison CL, Goldstein RF, TeedeHJ. Women with polycystic ovary syndrome have intrinsic insulin resistance on euglycaemic-hyperinsulaemic clamp. Hum Reprod. 2013 Mar;28(3):777-84. 4. Lee S, Bacha F, Hannon T, Kuk JL, Boesch C, Arslanian S. Effects of aerobic versus resistance exercise without caloric restriction on abdominal fat, intrahepatic lipid, and insulin sensitivity in obese adolescent boys: a randomized, controlled trial. Diabetes. 2012;61(11):2787-95. 5. Etgen GJ Jr, Jensen J, Wilson CM, Hunt DG, Cushman SW, Ivy JL. Exercise training reverses insulin resistance in muscle by enhanced recruitment of GLUT-4 to the cell surface. Am J Physiol. 1997;272(5 Pt 1):E864-9. 6. Moran LJ, Ko H, Misso M, Marsh K, Noakes M, Talbot M, et al. Dietary Composition in the Treatment of Polycystic Ovary Syndrome: A Systematic Review to Inform Evidence-Based Guidelines. J AcadNutrDiet.2013 Feb 15.doi:pii: S2212-2672(12)01925-9. 10.1016/j.jand.2012.11.018. 7. Mehrabani HH, Salehpour S, Amiri Z, Farahani SJ, Meyer BJ, Tahbaz F. Beneficial effects of a high-protein, low-glycemic-load hypocaloric diet in overweight and obese women with polycystic ovary syndrome: a randomized controlled intervention study. J Am CollNutr. 2012;31(2):117-25. Polycystic ovary syndrome - a complex problem Part III Nutritional Agents for Managing PCOS: Inositol by: Heidy Fritz, MA, ND Bolton Naturopathic Clinic 64 King St W, Bolton, ON L7E1C7 www.boltonnaturopathic.ca email@example.com Part II has addressed essential dietary strategies for controlling insulin resistance. In Part III we discuss the role of inositol in the management ofpolycystic ovary syndrome (PCOS). Inositol is a nutrient that belongs to the family of B-vitamins, however it plays a unique role in the cell, acting as a second messenger in the insulin-signaling pathway.(1,2) As such, inositol acts as a natural insulin-sensitizing agent. Currently, inositol is the most well researched natural agents for treating PCOS, and has been shown to improve many aspects of this condition including inducing ovulation, improving insulin resistance, improving hormone levels and reducing acne and hirsuitism. Ovulation Several studies have shown that inositol supplementation increases ovulation rates. A randomized controlled trial by Raffone et al found that inositol restored spontaneous ovulation in 65% of women, compared to 50% by metformin.(3) In the inositol group, ovulation occurred a mean 14.8 days from day 1 of the menstrual cycle. In another randomized, double blind, placebo controlled study, the authors found that 16 out of 23 women in the myo-inositol group ovulated, compared to just 4 out of 19 in placebo group.(1) A third study found that inositol restored menstrual cycling in all the women who had no or irregular menses at baseline.(4) Infertility Inositol has been shown to improve egg quality parameters as well as pregnancy rates in women with PCOS undergoing ovulation induction or in vitro fertilization (IVF). In a study of ovulation induction using FSH administration, women receiving inositol had a pregnancy rate to 28.9% (11 out of 38 women), compared to 26.1% with metformin (11 out of 42 women).(3) In a study of women undergoing IVF, inositol was shown to reduce the amount of FSH stimulation required for follicle maturation, decrease the number of degenerated oocytes, and increase the quality of oocytes retrieved (MII oocytes).(5) Metformin As described above, studies to date suggest that inositol may be equally effective as metformin in inducing ovulation and pregnancy. Metformin has been shown to work in part by increasing release of the inositol mediator in the insulin-signaling cascade.(6) This suggests that metformin may increase utilization of inositol, and there benefit from supplementing with inositol to maintain optimal levels. One study has shown that co-administration of metformin plus inositol plus a dietary intervention led to significantly better improvements in menstrual cycle regularity and percent of body fat weight loss in the group receiving combined treatment compared to metformin alone .(7) Hyperandrogenism Inositol has been shown to improve hyperandrogenism, both in reducing testosterone levels and in reducing symptoms associated with excess testosterone or DHEAS. For instance, when given to women on the birth control pill, inositol resulted in greater improvements in hirsuitism after one year compared to the birth control pill alone (8) Inositol plus the birth control pill was also shown to improve fasting glucose, fasting insulin, testosterone and DHEAS levels compared to the birth control pill alone. Insulin Resistance Inositol’s primary mechanism is theimprovement of insulin signaling. As such, it has been shown to improve the insulin sensitivity index as well as overall insulin levelsin women with PCOS.(1) A good “side effect” of this is that inositol also reduced blood pressure and cholesterol levels in these women.(1) It is worth noting however, that inositol may be of benefit even among women with PCOS but who do not have insulin resistance. According to a recent study, inositol supplementation improved egg quality in women with PCOS who showed no evidence of insulin resistance.(9) This opens up exciting possibilities for new applications and further research. Dosing Most studies have used the myo-inositol form, as opposed to the D-chiro-inositol form, at a dosage of 2 to 4g per day. There have been no reported side effects, but in general it is a good idea to start with a low dose and gradually increase to avoid any gastrointestinal upset. References 1. Costantino D, Minozzi G, Minozzi E, Guaraldi C. Metabolic and hormonal effects of myo-inositol in women with polycystic ovary syndrome: a double-blind trial. Eur Rev Med Pharmacol Sci. 2009;13(2):105-10. 2. Unfer V, Carlomagno G, Dante G, Facchinetti F. Effects of myo-inositol in women with PCOS: a systematic review of randomized controlled trials. GynecolEndocrinol. 2012;28(7):509-15. 3. Raffone E, Rizzo P, Benedetto V. Insulin sensitiser agents alone and in co-treatment with r-FSH for ovulation induction in PCOS women. GynecolEndocrinol. 2010;26(4):275-80. 4. Genazzani AD, Lanzoni C, Ricchieri F, Jasonni VM. Myo-inositol administration positively affects hyperinsulinemia and hormonal parameters in overweight patients with polycystic ovary syndrome. GynecolEndocrinol. 2008;24(3):139-44. 5. Papaleo E, Unfer V, Baillargeon JP, Fusi F, Occhi F, De Santis L. Myo-inositol may improve oocyte quality in intracytoplasmic sperm injection cycles. A prospective, controlled, randomized trial.FertilSteril. 2009;91(5):1750-4. 6. Baillargeon JP, Iuorno MJ, Jakubowicz DJ, Apridonidze T, He N, Nestler JE. Metformin therapy increases insulin-stimulated release of D-chiro-inositol-containing inositolphosphoglycan mediator in women with polycystic ovary syndrome. JClinEndocrinolMetab. 2004;89(1):242-9. 7. Le Donne M, Alibrandi A, Giarrusso R, Lo Monaco I, Muraca U. [Diet, metformin and inositol in overweight and obese women with polycystic ovary syndrome: effects on body composition]. Minerva Ginecol. 2012;64(1):23-9. Italian. 8. Minozzi M, Costantino D, Guaraldi C, Unfer V. The effect of a combination therapy with myo-inositol and a combined oral contraceptive pill versus a combined oral contraceptive pill alone on metabolic, endocrine, and clinical parameters in polycystic ovary syndrome.GynecolEndocrinol. 2011;27(11):920-4. 9. Unfer V, Carlomagno G, Rizzo P, Raffone E, Roseff S. Myo-inositol rather than D-chiro-inositol is able to improve oocyte quality in intracytoplasmic sperm injection cycles. A prospective, controlled, randomized trial.Eur Rev Med Pharmacol Sci. 2011;15(4):452-7. Polycystic ovary syndrome - a complex problem Part IV Nutritional Agents for Managing PCOS: NAC and Vitamin D by: Heidy Fritz, MA, ND Bolton Naturopathic Clinic 64 King St W, Bolton, ON L7E1C7 www.boltonnaturopathic.ca firstname.lastname@example.org In Part III we discussed the evidence showing benefit from supplementation with inositol on a wide variety of clinical endpoints related to PCOS. Given the strength of this evidence, inositol deserves to be a first-line natural agent for use in PCOS. Additional, secondary agents that have shown potential in improving aspects of PCOS include N-acetylcysteine (NAC), vitamin D, and fish derived omega-3 fatty acids. Here we will discuss the additional benefits that can be derived from these agents. N-acetylcysteine N-acetylcysteine or NAC is a fascinating little amino acid-like molecule. Its role in PCOS is not well defined, but there are a few different ways in which it is thought to act. First, NAC is a precursor for the universal antioxidant glutathione.(1) Increasing glutathione means that the body is better equipped to handle oxidative stress and minor toxic exposures that we encounter on a day to day basis. Theoretically, this might mean that oocytes are also more protected from cellular damage. Secondly and more specifically, NAC may be able to improve insulin sensitivity and ovulation rates in women with PCOS (2,3). In a recent randomized, controlled trial, NAC was studied in 180 infertile women with PCOS who were undergoing ovulation induction with clomiphene citrate.(3) Co-administration of NAC alongside clomiphene citrate was shown to significantly improve the number of follicles >18 mm and endometrial thickness on the day of hCG administration. In addition, ovulation and pregnancy rates were also significantly higher in the combined treatment group compared to clomiphene citrate plus placebo. Another study found that NAC after surgical intervention for PCOS resulted in similarly better pregnancy rates, as well as lower miscarriage rates and higher rate of live births.(4) Another study compared the effects of NAC to metformin in women with PCOS. NAC was found to be equivalent to metformin in all areas that were assessed, including the following: decrease in body mass index, hirsutism, fasting insulin levels, insulin sensitivity, free testosterone levels, and menstrual irregularity.(5) The dose of NAC used ranged from 1200 to 1800 mg per day. Vitamin D Low levels of vitamin D have been reported in a large proportion of women with PCOS.(6) In one study, 37% of patients had vitamin D levels below 50 nmol/L, whereas the recommended level in Canada is 75 nmol/L.(6) Vitamin D is thought to play a role in maintaining normal blood glucose control and insulin function.(7) Early studies have shown that vitamin D may improve glucose metabolism and menstrual regularity in women with PCOS.(8) In particular one study administering the equivalent of between 2000-3000 IU per day found that there was a significant decrease in fasting glucose and insulin secretion after vitamin D treatment. In addition, approximately 50% of women who had irregular cycles reported improvement of menstrual frequency after 24 weeks. Given that so many women with PCOS appear to have sub-optimal levels of vitamin D, it seems prudent to make use of this safe agent that is also so important for other areas of health. For more information on a dosing schedule that is appropriate for you, please consult your naturopathic doctor. Omega-3 fatty acids The fish-derived omega-3 fatty acids eicosapentanoic acid (EPA) and docosahexanoic acid (DHA) are well known for their anti-inflammatory and cardioprotective effects.(9) This is important for PCOS because women with this condition are at elevated risk of heart disease as a result of several factors, including impaired insulin sensitivity, higher cholesterol and blood pressure, and a tendency to be overweight.(10) In addition, an increased omega-6 to omega-3 fatty acid ratio has been identified in women with PCOS.(11) Supplementation with omega-3 fatty acids may help correct this imbalance. EPA and DHA have also been shown to improve some of the hormonal and metabolic disturbances characteristic of PCOS. For instance, supplementation of approximately 2g of combined EPA+DHA, has been shown to significantly reduce plasma free testosterone, with the greatest effect being among women who had the greatest reductions in their omega-6:omega-3 ratios in response to supplementation.(11) In other words, the more their omega-6 to omega-3 ratio improved, the more their testosterone levels went down. Interestingly, in lab models, stimulation of cells by omega-6 was actually shown to increase production of androgen hormones!(11) Another study found that fish oil supplementation was able to improve blood glucose and insulin levels, as well as improve cholesterol fractions.(12) Supplementation with a good quality fish oil appears to be beneficial for women with CPOS on the basis that it corrects an imbalance in omega 3 fatty acid levels, reduces long-term risk of cardiovascular disease, improves metabolic parameters such as insulin resistance, and reduces free testosterone. References 1. Uraz S, Tahan G, Aytekin H, Tahan V. N-acetylcysteine expresses powerful anti-inflammatory and antioxidant activities resulting in complete improvement of acetic acid-induced colitis in rats. Scand J Clin Lab Invest. 2013;73(1):61-6. 2. Fulghesu AM, Ciampelli M, Muzj G, Belosi C, Selvaggi L, Ayala GF, Lanzone A. N-acetyl-cysteine treatment improves insulin sensitivity in women with polycystic ovary syndrome. FertilSteril. 2002;77(6):1128-35. 3. Salehpour S, Sene AA, Saharkhiz N, Sohrabi MR, Moghimian F. N-Acetylcysteine as an adjuvant to clomiphene citrate for successful induction of ovulation in infertile patients with polycystic ovary syndrome. J ObstetGynaecol Res. 2012;38(9):1182-6. 4. Nasr A. Effect of N-acetyl-cysteine after ovarian drilling in clomiphene citrate-resistant PCOS women: a pilot study. Reprod Biomed Online. 2010;20(3):403-9. 5. Oner G, Muderris II. Clinical, endocrine and metabolic effects of metformin vs N-acetyl-cysteine in women with polycystic ovary syndrome.Eur J ObstetGynecolReprod Biol. 2011;159(1):127-31. 6. Muscogiuri G, Policola C, Prioletta A, Sorice G, Mezza T, Lassandro A, Della Casa S, Pontecorvi A, Giaccari A. Low levels of 25(OH)D and insulin-resistance: 2 unrelated features or a cause-effect in PCOS? ClinNutr. 2012;31(4):476-80. 7. Lerchbaum E, Obermayer-Pietsch B. Vitamin D and fertility: a systematic review. Eur J Endocrinol. 2012;166(5):765-78. 8. Wehr E, Pieber TR, Obermayer-Pietsch B. Effect of vitamin D3 treatment on glucose metabolism and menstrual frequency in polycystic ovary syndrome women: a pilot study. J Endocrinol Invest. 2011;34(10):757-63. 9. No authors. Dietary supplementation with n-3 polyunsaturated fatty acids and vitamin E after myocardial infarction: results of the GISSI-Prevenzione trial. GruppoItaliano per lo Studio dellaSopravvivenzanell'Infartomiocardico.Lancet. 1999;354(9177):447-55. 10. Mani H, Levy MJ, Davies MJ, Morris DH, Gray LJ, Bankart J, et al. Diabetes and cardiovascular events in women with polycystic ovary syndrome; a 20 years retrospective cohort study. ClinEndocrinol (Oxf). 2012. doi: 10.1111/cen.12068. [Epub ahead of print]. 11. Phelan N, O'Connor A, KyawTun T, CorreiaN, Boran G, Roche HM, et al. Hormonal and metabolic effects of polyunsaturated fatty acids in young women with polycystic ovary syndrome: results from a cross-sectional analysis and a randomized, placebo-controlled, crossover trial. Am J ClinNutr. 2011;93(3):652-62. 12. Mohammadi E, Rafraf M, Farzadi L, Asghari-Jafarabadi M, Sabour S. Effects of omega-3 fatty acids supplementation on serum adiponectin levels and some metabolic risk factors in women with polycystic ovary syndrome. Asia Pac J ClinNutr. 2012;21(4):511-8.