Introduction

Introduction

Preeclampsia is a major cause of maternal, fetal and neonatal morbidity and mortality, particularly in developing countries. Considering the burden of preeclampsia and its associated complications, it is important to understand the underlying risk factors and mechanisms involved in its etiology. There is considerable interest in the potential for dietary long chain polyunsaturated fatty acids (LCPUFA) as a therapeutic intervention to prevent preeclampsia, as they are involved in angiogenesis, oxidative stress, and inflammatory pathways. The study hypothesizes that LCPUFA and one carbon metabolite status among Indian women play a causal role in the etiology of preeclampsia, mediated by changes in oxidative stress and growth factors which alter placental development. This study proposes to examine the associations of maternal LCPUFA and one carbon micronutrient status in early gestation with clinical outcome in preeclampsia and to understand the underlying key biochemical and molecular mechanisms.

The ICMR Centre for Advanced Research (CAR) study follows a cohort of pregnant women from early pregnancy until delivery to examine longitudinally the associations of maternal LCPUFA with clinical outcome in preeclampsia. A multisite centre for advanced research was established and pregnant women coming to Bharati hospital and Gupte hospital, Pune, India for their first antenatal visit are recruited and followed up at 11-14 weeks, 18-22 weeks, 26-28 weeks, and at delivery. Their personal, obstetric, clinical, and family history are recorded. Anthropometric measures (height, weight), food frequency questionnaire (FFQ), physical activity, socioeconomic status, fetal ultrasonography, and color Doppler measures are recorded at different time points across gestation. Maternal blood at all time points, cord blood, and placenta at delivery are collected, processed and stored at -80°C. The children’s anthropometry is assessed serially up to the age of 2 years, when their neurodevelopmental scores will be assessed.

The Outcome of Proposed Study

The Outcome of Proposed Study

The present study will be useful in:

• Development/validation of biomarkers for early prediction of preeclampsia.
• Providing information on mechanistic aspects of preeclampsia.
• Understanding the neurodevelopmental outcome of children born to women with preeclampsia.
• Correlation of USG and color Doppler findings and histopathological examination of placenta.
• Establishing a biorepository: The establishment of a biorepository will provide an invaluable resource to be shared with other investigators for future research on other factors influencing adverse pregnancy outcomes.

Objectives

Objectives

The REVAMP study proposes to examine the associations of maternal LCPUFA and one carbon micronutrient status in early gestation with clinical outcome in preeclampsia and to understand the underlying key biochemical and molecular mechanisms. It consists of 4 sub-projects:

Project 1: Longitudinal changes in metabolites of the one carbon cycle in preeclampsia

The study will examine baseline levels (in early pregnancy) at 11–14 weeks of gestation (visit 1 –V1) and changes across gestation [18–22 weeks (visit 2 – V2), 26–28 weeks (visit 3 – V3) and at delivery] in components of the one carbon cycle [(folate, vitamin B12, methionine, glutathione, homocysteine, SAM, S-adenosylhomocysteine (SAH)] from early pregnancy until delivery, and placental global DNA methylation levels. It will also examine the relationship of altered one carbon metabolism to methylation potential (SAM: SAH ratio) in the placenta. Furthermore, these changes in the one carbon cycle metabolites will be associated with dietary and lifestyle factors, USG and color Doppler measures.

Project 2: Role of oxidative stress and metabolites of LCPUFA metabolic pathway in preeclampsia

This study will examine levels of oxidative stress markers (protein carbonyl, 8- hydroxyl (OH) guanidine, malondialdehyde (MDA); LCPUFA status and their bioactive metabolites (thromboxane B2; prostaglandin E2) at baseline and across gestation. In addition, measurements of placental phospholipids and gene expression and DNA methylation patterns of PPAR gamma, PEMT, and DNA methyl transferase (DNMT) genes from the placenta will be undertaken.

Project 3: Growth factors and regulation of their gene expression in preeclampsia

This study will analyze levels of angiogenic growth factors – [VEGF, PlGF, sFlt-1, sEng], HIF-1, neurotrophins – [brain derived neurotrophic factor (BDNF), nerve growth factor (NGF) and their receptors] across gestation. It will also examine the associations between altered expression of growth factors involved in placental development, their regulation by methylation as a response to altered one carbon metabolism, and placental histopathological changes. These molecular as well as structural changes in the placenta will further be correlated with clinical parameters for better understanding the etiopathology of preeclampsia.

Project 4: Association of maternal nutrition, biochemical and epigenetic changes with anthropometry and developmental changes in children born to mothers with preeclampsia

This study aims to follow-up the babies for their postnatal anthropometric measurements at birth, 6, 10, and 14 weeks, 6, 9, 12, 15, 18, and 24 months and developmental scales at 2 years of age. The associations of maternal nutrition, biochemical and epigenetic changes, USG and Doppler measures with anthropometry and developmental scores in children will also be undertaken to better understand the mechanisms involved in the fetal programming of developmental disorders in children born to mothers with preeclampsia.

Figure 1 depicts the connectivity between the various objectives.

Key questions

The proposed study will answer the following questions:

• Do levels of LCPUFA and their metabolites, and one carbon nutrients, differ in early pregnancy between women who do and do not go on to develop preeclampsia?
• Is there evidence that these differences are dietary in origin?
• Do the molecular and biochemical analyzes provide a plausible mechanistic pathway to explain the pathophysiology of preeclampsia?
• Do the early pregnancy biomarkers identified provide a clinically useful prediction of later preeclampsia?
• Does the postnatal growth and cognitive development of children of women with preeclampsia differ from that of controls?
• Are these differences in the children explained by changes in growth factors?

Mission

Mission

Reducing maternal and child morbidity and mortality by exploring mechanisms involved in preeclampsia.

Major Activities

Major Activities

Meetings with Collaborators and Advisers

Prof. Caroline Fall (University of Southampton)

Dr. K. Kumaran (MRC Lifecourse Epidemiology Unit & Holdsworth Memorial Hospital, Mysore)

Dr. Giriraj Chandak (CSIR- Centre for Cellular and Molecular Biology (CSIR-CCMB), Hyderabad)

Discussion with Clinicians from Bharati Hospital and Gupte Hospital

Training in Administering Questionnaires

Staff Trained in Administering Nutritional Questionnaires at National Institute of Nutrition (NIN), Hyderabad

Staff Trained in Administering Nutritional Questionnaires at National Institute of Nutrition (NIN), Hyderabad

Staff Trained in Administering Standard of Living Index (SLI) and Physical Activity Questionnaire at IRSHA

Training in Placental Histopathology

Training at St. John’s Research Institute, Bangalore

Staff Counseling the Patients and Administering Various Questionnaires at Bharati Hospital and Gupte Hospital

Follow up Meetings with all the Collaborators

Yearly Advisory Meeting

Establishment of Biorepository

Laboratory Analysis

1. Gas Chromatography (GC)

Fatty acid analysis using GC

2. ELISA

Estimation of protein levels of growth factors using ELISA

3. PCR and Real Time-PCR

mRNA expression of various genes using RT-PCR

4. High Performance Liquid Chromatography (HPLC)

S-Adenosyl Methionine (SAM) and S-Adenosyl-L-Homocysteine (SAH) estimations using HPLC

5. Ultracentrifuge

Isolation of plasma membranes from placental homogenates

Management team and Staff

Management team

• Dr. Sadhana Joshi, Professor and Head
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• Dr. Nisha S. Wadhwani, Scientist “B”,
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ICMR CAR Staff

• Ms. Madhura Sarda, Psychologist,
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• Ms. Shweta D Madiwale, Research Assistant,
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• Ms. Rutuja Tope, Nutritionist,
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• Ms. Sakshi Selukar, Nutritionist,
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• Mr. Sagar Bhosale, Social Worker,
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• Ms. Prachi Joshi, Social Worker,
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• Ms. Anupam Poddar, Lab Assistant,
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• Ms. Aditi Mane, Lab Assistant,
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• Ms. Aditi Godhamgaonkar, Lab Assistant,
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• Ms. Nikita Joshi, Lab Assistant,
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• Ms. Juilee Deshpande, Lab Assistant,
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• Ms. Aboli Ballal, Lab Assistant,
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• Mr. Mahesh Funde, Data Entry Operator,
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• Mr. Nganthoi Elangbam, Data Entry Operator,
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• Mr. Aniket Shelar, Field Attendant,
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Contact us

ICMR- Centre for Advanced Research (CAR)

Interactive Research School for Health Affairs (IRSHA),
Bharati Vidyapeeth (Deemed to be University),
Pune-Satara Road, Dhankawadi,
Pune 4110043,
Maharashtra, India
Tel No.:- 020-24366920
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