Young women living with obesity (OB) have a greater risk of developing iron deficiency. Plus, the risk of anemia and/or ID in young women living with overweight (OW) and obesity (OB) is further increased by inadequate dietary intake and/or poor bioavailability of iron, as well as gastrointestinal and menstrual iron losses. It is not certain whether women living with OW/OB can meet their iron requirements from their day-to-day diet.

The aim of this study is to compare iron absorption and losses over a long period between women living with and without OW and OB. Secondary outcomes include iron and inflammation status, as well as dietary iron intake.

Adiposity-related inflammation stimulates an increase in secretion of hepcidin hormone thus limiting the absorption of iron and increasing the risk for iron deficiency (ID). In addition, the risk of anemia and/or ID in young women living with overweight (OW) or obesity (OB) is further increased by inadequate dietary intake and/or poor bioavailability of iron, as well as gastrointestinal and menstrual iron losses. Although one study quantified iron losses and absorption in young African women using the stable isotope dilution method the focus was on normal-weight women. Thus, whether women living with OW/OB can meet their iron requirements from their habitual diet remains uncertain.

This is a longitudinal study including an iron absorption study (Phase 1) and a follow-up period (Phase 2) in women living with and without OW and OB.

In phase 1, 70 healthy, mildly- and non-anemic young women living with and without OW or OB will receive a test drink labelled with 15 mg of labelled ferrous sulfate (57Fe) (Visit 1). Fractional iron absorption from the test drink will be determined by measuring the incorporation of the isotopic label into red blood cells 14 days after administration (Visit 2), and will be compared between the two groups. Participants will then undergo a one-year equilibration period. During this time, they will be contacted monthly for an electronically completed, telephonic or in-person health check, chronic medication and supplement use,, blood donation or transfusion, and whether they have fallen pregnant. In addition, they will be asked if they are on a weight-loss diet, a urine pregnancy test done and weight and haemoglobin measured once every 3-4 months. Halfway through the equilibration period we will ask participants not taking hormonal contraceptives (and not intending to do so in phase 2 of the study), to track their menstrual cycle by providing them with a monthly paper- or electronic calendar.

About 1 year after isotope administration, participants will be contacted again and screened (Visit 3) for phase 2 of the study. During this visit, a health and medical history check including the use of proton pump inhibitors,antacids, H2 blockers or iron supplements, contraceptive use and menstruation intensity will be conducted. Then, anthropometric measurements will be performed to determine BMI, and a urine sample collected for pregnancy test. If the participant qualifies to proceed with phase 2, they will be required to consume an antihelminthic dose. Phase 2 of the study will involve 4 visits (Visits 4 to 7), where they will be followed up for 6 months. During this time, blood samples will be drawn every 8 weeks for the determination of isotopic composition. In addition, iron and inflammation status will be assessed. Dietary intake will be assessed using three 24-hour recalls conducted within visits 4, 6 and 7.

Additionally, for willing participants not using hormonal contraceptives and having a regular cycle (26 to 30 days), Visit 4 (or subsequent convenient visit) will be planned to coincide with day 2 of their menstrual phase where 6 ml of blood will be drawn to quantify sex hormones, hepcidin, iron and inflammation status and IL-6. These participants will be invited again during the follicular phase (day 7), early luteal phase (day 18) and late luteal phase (day 23) for additional blood samples. For each of these sampling points, a window period of +/- 1 day will be allowed.

Inclusion Criteria:

• Of African descent
• BMI of 18.5 to 24.9 kg/m2 for participants living without OW/OB and BMI ≥ 28 kg/m2 for participants living with OW/OB
• Having low to moderate inflammation-adjusted iron stores (ferritin ≤50 µg/L)
• Absence of low-grade inflammation (CRP<2 mg/l) for participants living without OW/OB and presence of low-grade inflammation CRP 2-20 mg/l) for participants living with OW/OB
• Planning to reside in the study area for at least 2 years

Exclusion Criteria:

• Hemoglobin < 11 g/dl
• Treated or self-reported chronic or malabsorptive disorder
• Current use of chronic anti-inflammatory medication, like corticosteroids or non-steroidal anti-inflammatory medication
• Pregnancy or planning to become pregnant in the next 2 years
• Lactation
• Fear of needles or experiencing vaso-vagal episodes when exposed to blood
• Difficulty drawing blood due to poor quality veins
• Blood donation in the past 4 months or plans to donate blood during the study
• On a weight-loss diet or program or planning to start the same during the study
• Smoking
• Unwillingness to stop taking iron and /or vitamin C-containing supplements during phase 1 of the study
• Regular use of antacids, proton pump inhibitors or H2 blockers

Additional inclusion criteria applicable to phase 2 of the study

• Labelled with stable iron isotope (tracer) for a minimum of one year
• Willingness not to start or stop contraceptive use during the 6 months
• BMI ≥ 18.5 to 24.9 kg/m2 for participants living without OW/OB and BMI ≥ 28 kg/m2 for participants living with OW/OB

Additional exclusion criteria applicable to phase 2 of the study

- Blood transfusion, intravenous iron infusion, blood donation or significant blood loss during the equilibration period