Effects of Isoflavones on Gene-expression

Study Overview
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Brief Summary
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Alleged benefits experienced by the consumption of soy in Asian countries have been attributed to the isoflavone content of soy products. Amongst other benefits, isoflavones are believed to relieve menopausal symptoms and are therefore often consumed in supplement form in Western countries. These supplements contain relatively high amounts of isoflavones, and the question is if these concentrations still exert beneficial effects or whether negative effects become dominant. Therefore, the investigators will study the effect of intake of one dose of isoflavones, as compared to placebo, for eight weeks on gene-expression in Peripheral Blood Mononuclear Cells (PBMCs) in post-menopausal, equol-producing women.

Detailed Description
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Primary Objective: to determine the effect of intake of one dose of isoflavones, as compared to placebo, for eight weeks on gene-expression in Peripheral Blood Mononuclear Cells (PBMCs) in post-menopausal, equol-producing women. Secondary Objectives: to determine the association between isoflavone plasma levels and gene-expression in PBMCs; to determine the variation of isoflavone plasma levels between subjects after intake of isoflavones for four and eight weeks; to explore whether PBMC gene-expression markers identified after 8 weeks isoflavone intervention are already present after 4 weeks intervention; to explore whether the severity of previous menopausal complaints is related to the effect of isoflavones on PBMC gene-expression; and to explore the association between isoflavone levels in plasma and spot urine. Study design: Double-blind placebo controlled crossover intervention study Study population: Thirty-six healthy females, 45-70 years, post-menopausal and equol-producing Intervention: Two intervention periods of eight weeks with a isoflavone supplement or a placebo and a washout period of 8 weeks in between. Main study parameters/endpoints: The main study parameter is gene-expression in PBMCs measured by micro-arrays.

Official Title
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The Effect of Isoflavone Supplement Intake on Gene-expression in Postmenopausal Women

Conditions
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Postmenopause

Intervention / Treatment
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* Dietary Supplement: Isoflavone supplement


Participation Criteria
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Eligibility Criteria
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Inclusion Criteria: 45-70 years Equol producer Post-menopausal (Follicle Stimulating Hormone (FSH) >40 UI/L) or menstrual cycle absent for more than 1 year. Exclusion Criteria: current use of contraceptives containing hormones current use of hormone replacement therapy regular soy product use (more than once a week) regular isoflavone supplement use (more than once a week) current use of medication containing sex hormones or sex hormone-triggering compounds current use of anti-inflammatory medicines use of antibiotics in the past 6 months severe heart disease thyroid disorders removed thyroid gland complete ovariectomy prior diagnosis of cancer in medical history alcohol and drug abuse current smoker Body Mass Index (BMI) >35 kg/m2 allergy to soy (products)

Ages Eligible for Study
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Minimum Age: 45 Years
Maximum Age: 70 Years

Sexes Eligible for Study
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Female

Accepts Healthy Volunteers
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Accepts Healthy Volunteers

Study Plan
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How is the study designed?
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Design Details

Allocation: Randomized
Intervention Model: Crossover Assignment
Masking: Triple


Arms and Interventions

| Intervention/Treatment |
| --- |
|Dietary Supplement: Isoflavone supplement|The participants will consume 2 times 2 supplements per day, which will lead to a daily dose of 114 mg. (HPLC analysis confirmed an aglycone isoflavone content of 28.41 mg per supplement)|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Gene-expression measured by micro-arrays | Gene expression changes will be assessed in PBMCs using whole genome Affymetrix microarrays. The gene expression changes after exposure to the isoflavone supplement will be compared to the changes after exposure to the placebo. | gene-expression after 8 weeks of exposure to the isoflavone supplement | 
| Gene-expression measured by micro-arrays | Gene expression changes will be assessed in PBMCs using whole genome Affymetrix microarrays. The gene expression changes after exposure to the isoflavone supplement will be compared to the changes after exposure to the placebo. | after 8 weeks of exposure to the placebo | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Isoflavone levels in plasma and spoturine | levels of the isoflavones genistein, dihydrogenistein, daidzein, dihydrodaidzein, equol and glycitein will be measured with High performance liquid chromatography (HPLC). Following the intention to treat analysis, these concentration markers will be determined in order to correlate the gene-expression measured with quantitative real-time polymerase chain reaction (QPCR) with the isoflavone plasma levels, and to stratify for the plasma levels in per protocol analysis. | after 0 weeks of exposure to the isoflavone supplement | 
| Isoflavone levels in plasma and spoturine | levels of the isoflavones genistein, dihydrogenistein, daidzein, dihydrodaidzein, equol and glycitein will be measured with HPLC. Following the intention to treat analysis, these concentration markers will be determined in order to correlate the gene-expression measured with QPCR with the isoflavone plasma levels, and to stratify for the plasma levels in per protocol analysis. | after 4 weeks of exposure to the isoflavone supplement | 
| Isoflavone levels in plasma and spoturine | levels of the isoflavones genistein, dihydrogenistein, daidzein, dihydrodaidzein, equol and glycitein will be measured with HPLC. Following the intention to treat analysis, these concentration markers will be determined in order to correlate the gene-expression measured with QPCR with the isoflavone plasma levels, and to stratify for the plasma levels in per protocol analysis. | after 8 weeks of exposure to the isoflavone supplement | 
| Isoflavone levels in plasma and spoturine | levels of the isoflavones genistein, dihydrogenistein, daidzein, dihydrodaidzein, equol and glycitein will be measured with HPLC. Following the intention to treat analysis, these concentration markers will be determined in order to correlate the gene-expression measured with QPCR with the isoflavone plasma levels, and to stratify for the plasma levels in per protocol analysis. | after 0 weeks of exposure to the placebo | 
| Isoflavone levels in plasma and spoturine | levels of the isoflavones genistein, dihydrogenistein, daidzein, dihydrodaidzein, equol and glycitein will be measured with HPLC. Following the intention to treat analysis, these concentration markers will be determined in order to correlate the gene-expression measured with QPCR with the isoflavone plasma levels, and to stratify for the plasma levels in per protocol analysis. | after 4 weeks of exposure to the placebo | 
| Isoflavone levels in plasma and spoturine | levels of the isoflavones genistein, dihydrogenistein, daidzein, dihydrodaidzein, equol and glycitein will be measured with HPLC. Following the intention to treat analysis, these concentration markers will be determined in order to correlate the gene-expression measured with QPCR with the isoflavone plasma levels, and to stratify for the plasma levels in per protocol analysis. | after 8 weeks of exposure to the placebo | 
| Confirmation of gene-expression with quantitative real time polymerase chain reaction (QPCR) | Ten genes per sample will be verified with QPCR. PBMCs from the subjects are available at time points before, halfway and after the two intervention periods. On the basis of the micro-array results it will be decided which genes from which samples will be quantified. | after 0 weeks of exposure to the isoflavone supplement | 
| Confirmation of gene-expression with quantitative real time polymerase chain reaction (QPCR) | Ten genes per sample will be verified with QPCR. PBMCs from the subjects are available at time points before, halfway and after the two intervention periods. On the basis of the micro-array results it will be decided which genes from which samples will be quantified. | after 4 weeks of exposure to the isoflavone supplement | 
| Confirmation of gene-expression with quantitative real time polymerase chain reaction (QPCR) | Ten genes per sample will be verified with QPCR. PBMCs from the subjects are available at time points before, halfway and after the two intervention periods. On the basis of the micro-array results it will be decided which genes from which samples will be quantified. | after 0 weeks of exposure to the placebo | 
| Confirmation of gene-expression with quantitative real time polymerase chain reaction (QPCR) | Ten genes per sample will be verified with QPCR. PBMCs from the subjects are available at time points before, halfway and after the two intervention periods. On the basis of the micro-array results it will be decided which genes from which samples will be quantified. | after 8 weeks of exposure to the isoflavone supplement | 
| Confirmation of gene-expression with quantitative real time polymerase chain reaction (QPCR) | Ten genes per sample will be verified with QPCR. PBMCs from the subjects are available at time points before, halfway and after the two intervention periods. On the basis of the micro-array results it will be decided which genes from which samples will be quantified. | after 4 weeks of exposure to the placebo | 
| Confirmation of gene-expression with quantitative real time polymerase chain reaction (QPCR) | Ten genes per sample will be verified with QPCR. PBMCs from the subjects are available at time points before, halfway and after the two intervention periods. On the basis of the micro-array results it will be decided which genes from which samples will be quantified. | after 8 weeks of exposure to the placebo | 

 Terms related to the study
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Keywords Provided by Centre Hospitalier Valida
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health, women