Tachykinin and Kisspeptin Expression in Human Granulosa and Cumulus Cells

Study Overview
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Brief Summary
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The purpose of this project is to study the presence of expression differences - at RNA and protein level - of different members of the tachykinin family and kisspeptin and their receptors, between fertile women and infertile patients with different etiologies.

Official Title
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Development of a New Diagnostic Tool to Assess Oocyte Quality Based on Tachykinin and Kisspeptin Expression Analysis in Human Granulosa and Cumulus Cells

Conditions
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Infertility

Intervention / Treatment
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* Procedure: Ovarian puncture


Participation Criteria
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Eligibility Criteria
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Two groups of subjects (each one with different criteria): Oocyte donors Inclusion Criteria: Age between 18 and 34 years Normal caryotype Normal psychological test Exclusion Criteria: Presence of hereditary diseases Beta-thalassemia Cystic fibrosis Human immunodeficiency virus (HIV), Hepatitis B virus (HBV), Hepatitis C virus (HCV) Infertility patients Inclusion Criteria: Age between 18 and 45 years One of the following etiologies: Patient with PCOS according to Rotterdam Criteria or Patients with endometriosis stage I-IV or Patients with low ovarian response according to Bologna criteria or Patients with advances maternal age (between 38 and 45 years)

Ages Eligible for Study
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Minimum Age: 18 Years
Maximum Age: 45 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

Primary Purpose: Basic Science
Allocation: Non-Randomized
Intervention Model: Single Group Assignment
Masking: None (Open Label)


Arms and Interventions

| Participant Group/Arm | Intervention/Treatment |
| --- | --- |
| Active Comparator: Donors<br>Oocyte donors without infertility problems | Procedure: Ovarian puncture<br>* Donors and patients are subjected to ovarian puncture as part of their donation procedure or IVF treatment. Oocyte are retrieved in this procedure along with granulosa and cumulus cells. Donors and patients sign an informed consent in order to donate their granulosa and cumulus cells for the study.<br>|
| Experimental: PCOS<br>Patients with Polycystic Ovary Syndrome (PCOS) according to Rotterdam criteria | Procedure: Ovarian puncture<br>* Donors and patients are subjected to ovarian puncture as part of their donation procedure or IVF treatment. Oocyte are retrieved in this procedure along with granulosa and cumulus cells. Donors and patients sign an informed consent in order to donate their granulosa and cumulus cells for the study.<br>|
| Experimental: Endometriosis<br>Patients with endometriosis (I-IV) | Procedure: Ovarian puncture<br>* Donors and patients are subjected to ovarian puncture as part of their donation procedure or IVF treatment. Oocyte are retrieved in this procedure along with granulosa and cumulus cells. Donors and patients sign an informed consent in order to donate their granulosa and cumulus cells for the study.<br>|
| Experimental: Age<br>Patients with advanced maternal age (38 years or more) | Procedure: Ovarian puncture<br>* Donors and patients are subjected to ovarian puncture as part of their donation procedure or IVF treatment. Oocyte are retrieved in this procedure along with granulosa and cumulus cells. Donors and patients sign an informed consent in order to donate their granulosa and cumulus cells for the study.<br>|
| Experimental: Low ovarian response<br>Patients with low ovarian response according to Bologna criteria | Procedure: Ovarian puncture<br>* Donors and patients are subjected to ovarian puncture as part of their donation procedure or IVF treatment. Oocyte are retrieved in this procedure along with granulosa and cumulus cells. Donors and patients sign an informed consent in order to donate their granulosa and cumulus cells for the study.<br>|

What is the study measuring?
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Primary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Expression level of Substance P (SP) | Expression analysis at mRNA (mRNA = messenger ribonucleic acid) and protein level using techniques of real time quantitative PCR (PCR = polymerase chain reaction), immunocytochemistry and western blot | 4 years | 
| Expression level of neurokinin A (NKA) | Expression analysis at mRNA and protein level using techniques of real time quantitative PCR, immunocytochemistry and western blot | 4 years | 
| Expression level of neurokinin B (NKB) | Expression analysis at mRNA and protein level using techniques of real time quantitative PCR, immunocytochemistry and western blot | 4 years | 
| Expression level of hemokinin 1 (HK-1) | Expression analysis at mRNA and protein level using techniques of real time quantitative PCR, immunocytochemistry and western blot | 4 years | 
| Expression level of neurokinin 1 receptor (NK1R) | Expression analysis at mRNA and protein level using techniques of real time quantitative PCR, immunocytochemistry and western blot | 4 years | 
| Expression of neurokinin 2 receptor (NK2R) | Expression analysis at mRNA and protein level using techniques of real time quantitative PCR, immunocytochemistry and western blot | 4 years | 
| Expression level of neurokinin 3 receptor (NK3R) | Expression analysis at mRNA and protein level using techniques of real time quantitative PCR, immunocytochemistry and western blot | 4 years | 
| Expression level of kisspeptin 1 (KISS1) | Expression analysis at mRNA and protein level using techniques of real time quantitative PCR, immunocytochemistry and western blot | 4 years | 
| Expression level of kisspeptin 1 receptor (KISS1R) | Expression analysis at mRNA and protein level using techniques of real time quantitative PCR, immunocytochemistry and western blot | 4 years | 

Secondary Outcome Measures

| Outcome Measure | Measure Description | Time Frame |
| --- | --- | --- |
| Patient or donor age | Years | 4 years | 
| Patient or donor body mass index | kg/m2 | 4 years | 
| Years of infertility | Number of years that the patient has been trying to have a child without success | 4 years | 
| Estradiol level (day of ovulation induction) | Estradiol level in blood (pg/mL), measured the day in which ovulation is induced | 4 years | 
| Progesterone level (day of ovulation induction) | Progesterone level in blood (ng/mL), measured the day in which ovulation is induced | 4 years | 
| Number of oocytes retrieved in the ovarian puncture |  | 4 years | 
| Number of mature oocytes (metaphase II) obtained in the ovarian puncture |  | 4 years | 
| Fertilization rate | Proportion of oocytes correctly fertilized (2 pronuclei and 2 polar bodies) after the ICSI (ICSI = intracytoplasmic sperm injection) procedure | 4 years | 
| Embryo quality | Embryo quality at day 3 of embryo development (of each embryo of the cohort). Quality assessed according to ASEBIR (ASEBIR = Asociación Española para el estudio de la Biología de la Reproducción = Spanish society for reproductive biology) criteria. Embryo quality grade A to D. | 4 years | 
| Daily and total dose of FSH (FSH = follicle stimulating hormone) | Daily dose of FSH for controlled ovarian stimulation. International Units (IU) | 4 years | 
| Daily and total dose of HMG (HMG = human menopausal gonadotropin) | Daily dose of HMG for controlled ovarian stimulation. International Units (IU) | 4 years | 
| Type of ovulation induction | Ovulation induction using 6500 IU of hCG (hCG = human chorionic gonadotropin) or 0.2 mg of triptorelin (Decapeptyl) | 4 years | 

 Terms related to the study
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Keywords Provided by Centre Hospitalier Valida
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Infertility, Tachykinin, Kisspeptin