Glutathione levels and miscarriage For a pregnancy to succeed, the maternal immune system must adapt to the presence of foreign cells. Within the immune system, T cells polarize to differential functions characterized by their pattern of cytokine expression. T helper 1 (TH1) cells produce IL-12, IFN-g, and TNF-a, whereas T helper 2 (TH 2) cells produce IL-4, 5, and 10. Rodent studies have shown that if TH2-type cytokines are produced at the maternal–fetal interface, then the pregnancy goes to term, but if TH1-type cytokines are produced, then the pregnancy ends in miscarriage. Human studies have confirmed that the TH1-type cytokines are harmful to a pregnancy (1). What is unclear, however, is what triggers either a TH1- or TH2-type response. Recent murine studies have implicated glutathione (2). Glutathione plays an important role in many basic processes, detoxifies free radicals and endogenous toxins, and helps preserve the intracellular redox balance. Glutathione depletion was found to inhibit TH1 cytokine production from murine antigen–presenting cells (2). More recent murine studies have found that a deficiency of glucose 6 phosphate dehydrogenase (G6PD), the rate-limiting enzyme in the hexose monophosphate shunt pathway, led to an increase in perinatal deaths (3). G6PD is involved in the regeneration of NADPH, which at times of oxidative stress maintains glutathione levels for the essential detoxification of free radicals and lipid peroxides. The aim of this study was to measure glutathione levels in pregnant women with a history of recurrent miscarriage, some of whose pregnancies went to term and some of whose did not. Our institution’s ethics committee approved this study, and all women gave informed consent before blood samples were taken. Eleven first-trimester pregnant women with a history of recurrent miscarriage (at least three miscarriages) were enrolled into the study. None of the women were on any medication. Six of the pregnancies went successfully to term (mean gestation at time of sampling 6.2 6 0.9 wks), whereas five ended in miscarriage again later in the first trimester (mean gestation at time of sampling 6.8 6 1.2 wks). Serum samples were stored at 220°C until required for analysis. Glutathione levels were measured according to the method of Punchard et al. (4). Briefly, 100-ml aliquots of serum or glutathione standard (0 –1 nmol/100 mL) were analyzed in flat-bottomed 96-well plates. 100 mL of solution containing 1.5 mMl 21 DTNB and 0.5mMl 21 NDPH were added. The reaction was initiated by addition of 50 mL IU/ml glutathione reductase (173 U/mg protein) to all wells excluding the blank, and the reaction was followed over 10 min at 410 nm using an MR5000 Dynatech plate reader. Linear rates of reaction were obtained over the range of the standard curve. The results obtained are given in Figure 1. These show that in pregnant women who later miscarried, glutathione levels were significantly higher than in those women whose pregnancies went successfully to term. At the time of sampling, all were pregnant. As far as we are aware, this is the first time that data showing a link between glutathione levels and the TH 1/TH2 response have been shown in humans. These results support those found in the earlier murine studies (2). If the results presented here are combined with our earlier findings (1) made in the same patients, then elevated glutathione levels were associated with increased IFN-g production. The lower glutathione levels were associated with increased IL-10 levels. At the time of sampling, all patients were pregnant. Glutathione is required to detoxify free radicals and lipid peroxides and to maintain the intracellular balance. All are affected by oxidative stress. Recent studies have shown that female mice with two defective copies of the gene for the rate-limiting enzyme G6PD had significantly reduced litters compared with mice in whom the gene was not faulty. Further work showed that if the fetus was able to produce G6PD, then the pregnancies continued, even if the womb was unable to produce G6PD (3). G6PD is intracellular, and deficiency of this enzyme will result in reduced levels of GSH within the cell. The GSH that was measured and found to be elevated in this study was extracellular, implying inverted oxidative stress. In mice where Received March 6, 2000; revised and accepted June 8, 2000. Reprint requests: Dr. R. Wilson, Department of Medicine, Glasgow Royal Infirmary, 10 Alexandra Parade, Glasgow G31 2ER, United Kingdom (FAX: 0141-211-4010; E-mail: gcl025@clinmed.gla.ac.uk). CORRESPONDENCE FERTILITY AND STERILITYt VOL. 74, NO. 6, DECEMBER 2000 Copyright ©2000 American Society for Reproductive Medicine Published by Elsevier Science Inc. Printed on acid-free paper in U.S.A. 0015-0282/00/$20.00 PII S0015-0282(00)01585-5 1257