Recurrent pregnancy loss (RPL) is defined as three or more consecutive spontaneous miscarriages. It occurs in about 0.5–3% of women trying to conceive¹ and around 10% of all human pregnancies end in spontaneous abortion. In the majority of such cases, the aetiology remains unknown. RPL and being childless are not only a gynaecological problem but also a social stigma. Despite thorough investigations according to various clinical protocols, the underlying aetiology remains obscure in majority of patients and obstetricians are facing a challenge to determine the cause of these unexplained abortions. The process of human conception is extremely complicated and there are several factors involved in establishment and maintenance of pregnancy, placental circulation being one of them.

Identifiable causes are usually found in only about 30- 50% of these women and the treatment depends on the underlying cause of miscarriage.² Various possible causes of RPL include parental chromosomal anomaly, congenital uterine malformations, uterine fibroids, primary endometrial defects, prothrombotic states like antiphospholipid syndrome and heritable thrombophilia, endocrinological abnormalities like hypersecretion of Luteinizing Hormone (LH), hyperprolactinaemia, polycystic ovaries, hyperhomocysteinaemia smoking and alcohol, etc.³

Homocystenine (Hcy) has been under research for its effect on pregnancy as its levels are usually raised in RPL. It was found that hyperhomocysteinaemia was associated with an approximately two- to three-fold increased risk of adverse pregnancy outcome.⁴ It was confirmed by Dodds et al. that there is an association between increased Hcy level in early pregnancy and pregnancy loss.⁵ Recently, it is found that hyperhomocysteinaemia is related to low levels of vitamin B12 in pregnancy.⁶ Another study shows deficiency of both folate and cobalamin causes megaloblastic anaemia and also raises Hcy levels in body.⁷ It has also been studied that parenteral administration of cobalamin decreased the raised Hcy levels and normalised 5-methyltetrahydrofolate/total folate ratio.⁸ One study on autistic children also reported that vitamin supplementation reduced homocysteine level when measured in urine.⁹ On the contrary, there was no significant association was found between hyperhomocysteinaemia and spontaneous abortion risk, in a study by Khong et al.¹⁰

The above-mentioned references gave us a thought to find out that if in women with RPL, apart from standard treatment if an add on therapy of multivitamin is supplemented, then does it brings down the raised Hcy levels, which may improve pregnancy outcome if the gestation period crosses 18 weeks. Since there was a paucity of information regarding the effect of multivitamin supplementation in cases of RPL with hyperhomocystenine more so in an Indian population, thus the present study was planned to evaluate the effect of multivitamin supplementation on:

1. Plasma Hcy levels of the women at 18 weeks of the gestation period.

2. The number of women in whom there was no miscarriage (reached 18 weeks of gestation period).

This was an investigator-initiated, randomised, open-label, single-centre, 6-week parallel group study having two arms: standard treatment alone and standard treatment with multivitamin. The study was conducted in the Department of OBG, Adesh Institute of Medical Sciences and Research, Bathinda from 2011 to 2012. The pregnant patients visiting the OBG OPD were screened for inclusion in the study. They were included if they were 18 years and above, had early pregnancy (6–12 weeks period of gestation; gestational age calculated based on the first day of last menstrual period and/or by transvaginal ultrasonography), history of previous one or more spontaneous abortions without any detectable cause, fasting plasma Hcy levels > 15 μmol/l and were willing to give written informed consent. Exclusion criteria's were uterine anomalies, antiphospholipid syndrome, chromosomal anomalies, polycystic ovarian syndrome, any past or present medical disorder, current participation (including prior 30 days) in any other therapeutic trial and any condition that, in the opinion of investigator, may prevent the participant from adhering to the trial protocol.

Eligible women were randomised in a 1:1 ratio to either standard treatment (group I) or standard treatment with multivitamin (group II). Group I received standard treatment for RPL (bed rest, Human Chorionic Gonadotropin (HCG), supplementation, folic acid, etc.). Group II received multivitamin tablet containing homocysteine-lowering agents in the form of combination of methylating agents and folic acid in addition to standard treatment. Women were instructed to take one multivitamin tablet with 200 ml water once daily for 6 weeks

The primary outcome was alteration in mean homocysteine levels at 6 weeks from baseline in the two study groups. The secondary outcome included number of pregnancies reaching safe gestation (18 weeks). Hcy levels were estimated using florescence immunoassay technology.¹¹

The study protocol was approved by the Institutional Ethics Committee and all the participants signed a written informed consent prior to enrolment to the study. The study was conducted in accordance with Ethical Guidelines for Biomedical Research on Human Subjects, Indian Council of Medical Research.

Figure 1 depicts the flow of study participants: 54 eligible women were enrolled in the research study, out of which 6 patients were lost to follow-up and 2 patients withdrew from the study due to personal reasons. In all, 37 participants (18 in group I and 19 in group II) completed the study.

The mean age in the study group was 24±2.5 years. Majority of women in the study (67.5%) belonged to the middle socioeconomic status.

Table 1 shows that mean plasma Hcy levels decreased significantly after treatment for 6 weeks in group receiving multivitamin. This implies that treatment has a definitive role in lowering of plasma Hcy levels and improvement in pregnancy outcome.

A total of 67% (12/18) of women in group II reached a safe period of gestation of 18 weeks as compared to 47% (9/19) in group I.

Homocystiene, an amino acid, produced by the intracellular demethylation of methionine in the methylation processes. The levels of homocysteine normally fall during pregnancy. High Hcy levels are usually associated with complications like neural tube defects, placental infarcts, foetal growth retardation as well as placental abruption.

It is well know that folic acid, vitamin B12 and vitamin B6 are involved in Hcy metabolism and are required for DNA synthesis and cell growth. Our study corroborates with the finding of study conducted by Leeda et al. (1998) who studied the effects of folic acid and vitamin B6 supplements on women with hyperhomocysteinaemia and found that that vitamin B6 and folic acid correct the methionine loading test and improve the perinatal outcome.

The role of increased levels of Hcy is now being studied and discussed in detail and the proposed mechanism is that it promotes oxidative stress. Deleterious effects of these free radicals include initiation of lipid peroxidation, oxidative damage of biomolecules and cellular dysfunction and may finally initiate maternal vascular endothelial dysfunction with defective chorionic villous vascularisation.¹³

Various other similar mechanisms that have been proposed for RPL due to homocysteinaemia are:

It mimics thrombophilia that augments the relative hypercoagulable state of pregnancy and leads to thrombosis in maternal/foetal circulation.¹⁴

The placenta of women with thrombophilias and other severe complications have an increased rate of vascular lesions leading to impaired nutrition to the fetus and RPL.¹⁵

Hyperhomocysteinaemia is associated with reduced methylation, endothelial dysfunction and increased DNA damage, which can adversely affect pregnancy outcome.¹⁶

The above literature suggests that abnormality of placental vasculature and the alteration in homeostasis lead to inadequate foetal circulation finally leading to RPL but there is paucity of information in regards of management of RPL. Since hyperhomocysteinaemia is associated with reduced methylation, supplementation of multivitamin reduces the homocysteine level and can reduce the adverse pregnancy outcome in such cases.

Multivitamin reduces raised homocysteine level and reduces adverse pregnancy outcome in cases of RPL. Supplementation of multivitamin along with the standard treatment (bed rest, HCG supplementation, folic acid, etc.) of RPL during the first trimester will be helpful in improvement of pregnancy outcome in cases of RPL.

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