Quercetin and Bornyl Acetate Regulate T-Lymphocyte Subsets and INF-γ/IL-4 Ratio In Utero in Pregnant Mice

Hindawi Publishing Corporation Evidence-Based Complementary and Alternative Medicine Volume 2011, Article ID 745262, 7 pages doi:10.1155/2011/745262 Research Article Quercetin and Bornyl Acetate Regulate T-Lymphocyte Subsets and INF-γ/IL-4 Ratio In Utero in Pregnant Mice Xiaodan Wang,1 Aituan Ma,1 Wanyu Shi,2 Meiying Geng,1 Xiuhui Zhong,1 and Yantao Zhao2 1 College of Traditional Chinese Veterinary Medicine, Agricultural University of Hebei, Baoding 071001, China 2 College of Animal Science and Veterinary Medicine, Agricultural University of Hebei, Baoding 071001, China Correspondence should be addressed to Xiuhui Zhong, Received 25 April 2010; Revised 21 June 2010; Accepted 10 September 2010 Copyright © 2011 Xiaodan Wang et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The objective of this study is to investigate the antiabortive effects of Quercetin and Bornvl Acetate and their immunological modulation at maternal-fetal interface. Lipopolysaccharide (LPS) was injected via tail vein to induce abortion in mice which received Quercetin and Bornvl Acetate at days 4–7 of gestation. Uterine CD4+/CD8+ T lymphocytes and IFN-γ/IL-4 of each group (n = 10) were detected by immunohistochemistry and enzyme-linked immunosorbent assay, respectively. The ratio of CD4+/CD8+ increased significantly (P < .01) in the uterus of LPS-induced abortion mice. In the Quercetin and Bornvl Acetate pretreated mice followed by LPS administration, the ratio of CD4+/CD8+ dropped to 0.562 ± 0.021, lower than that of LPSabortion group (P < .01). The mean value of IFN-γ/IL-4 in LPS-treated mice was 0.310 ± 0.066, higher than that of Quercetin and Bornyl Acetate group. The results indicate that Quercetin and Bornyl Acetate have an antiabortive effect through modulation of immunological balance at maternal-fetal interface. 1. Introduction Pregnancy is a complex, sophisticated physiological process. During this process, the embryo is viewed as a semiallograft due to the expression of paternal antigens that are foreign to the mother and a total allograft in fertilization in vitro and embryo transfer (IVF-ET). The semiallogenic nature of the fetus is not usually rejected by the maternal immune system, which includes complicated immune modulation. In recent years, the contribution of focalized immunity at the maternofetal interface to successful embryo implantation and growth has been studied to a great extent; maintaining the maternofetal tolerance is a main concern for reproductive immunologists. Cases of pregnancy failure, such as spontaneous abortion, have been studied for association with altered immune responses [1, 2]. In 1993, Wegmann et al. [3] first raised the hypothesis that there was a pregnancypreferred Th2 bias involving the downregulation of the cellular immune response, and the upregulation of humoral immune response. There have been a great number of studies which support Wegmann’s hypothesis [4, 5]. T lymphocytes involved in gestation immunity account for the third most abundant leukocyte population in normal early pregnancy decidua, accounting for 20%–30% of stromal leukocytes. Endometrial T lymphocytes participate in the defense mechanism of uterus, and either secreting cytokines when stimulated or acting as local suppressor cells. The human decidua contains an unusually high proportion of lymphocytes, mainly NK and T cells, which are potentially cytotoxic to the trophoblast when they are stimulated with certain cytokines, resulting in embryo loss. Olivares et al. [6] reported that activated decidual lymphocytes participate in human spontaneous abortion by inducing apoptosis but not necrosis of the trophoblast. Quercetin is an herbal flavonoid derived from various kinds of plants and has been found to possess various biological activities, such as antioxidative [7, 8], antimicrobial [9], wound-healing [10], anticancer [11, 12] and immunomodulatory activities [13]. In Chinese herbal medicine, Quercetin is the major constituent of many herbs, such as Semen Cuscutae, Herba Taxilli, Cortex Eucommiae. Bornyl Acetate constitutes the major components in the oil of Fructus Amomi. It is recorded in the Chinese Veterinary Pharmacopoeia (the 2005 edition) that Semen Cusutae, Herba Taxilli, Cortex Eucommiae, and Fructus Amomi are antiabortives. So far, there have been few reports on whether Quercetin and Bornyl Acetate are the effective components of the antiabortive herbs, and how these components affect 2 the survival of the fetus. In the present study, Lipopolysaccharide (LPS) was administered by injection to pregnant mice to induce embryo resorption. Uterine CD4+ and CD8+ T cells were detected immunohistochemically, and IFN-γ and IL-4 contents were measured by Enzyme-linked immunosorbent assay (ELISA) with an aim to elucidate the antiabortive effects and the mechanisms of Quercein and Bornyl Acetate. 2. Materials and Methods 2.1. Preparation of Reagents. LPS (Lipopolysaccharide, Sigma Chemicals) from Escherichia coli was dissolved in sterile phosphate-buffered saline (PBS) (0.01 M, pH 7.4) at a concentration of 0.5 μg/mL. Ten mg of Quercetin (Sigma products, purity = 98%) was dissolved with a small amount of dehydrated alcohol and then diluted with PBS to 4 mL, the final concentration being 2.5 mg/ml. An amount of 10 μL Bornyl Acetate (Fluka products, purity ≥99%) was dissolved with small amount of dehydrated alcohol, and then diluted with PBS to 4 mL. Quercetin (5 mg) and Bornyl Acetate (5 μL) was mixed and dissolved with a small amount of dehydrated alcohol, and then diluted with PBS to 4 mL. 2.2. Treatment of Animals. BALB/c virgin mice aged 10 weeks (18–22 g BW) were purchased from the Laboratory Animal Center of Hebei Medical University, China and were given free access to mouse chow and water, with a 12 hours light cycle from 7:00–19:00. Pregnancies were obtained by housing one virgin female with one male, and the females were examined each day in the early morning for the presence of a vaginal plug. The day that the vaginal plug was detected was designated as day 0 of pregnancy. The pregnant mice were randomly divided into 5 groups. Group A was kept as a control group, group B as a LPS model group, group C as a Que (Quercetin) group, group D as a BA (Bornyl Acetate) group, and group E as a Que + BA group. Mice in groups B, C, D, and E were given intravenous injections (i.v.) of 0.2 mL (0.1 μg) LPS/mice via the lateral tail vein on day 7. Mice in groups C, D, and E received Que, BA, and Que + BA respectively at a dose of 0.4 mL per day via oral gavage on gestation days 4–7. Animals in group A received an oral gavage of PBS on the same days of gestation as in group C, and were administrated with PBS at 0.4 mL at day 7 of gestation. Animals were treated according to the Guidelines for Keeping Experimental Animals issue (...truncated)