The expression of selenium-binding protein 1 is decreased in uterine leiomyoma

Diagnostic Pathology The expression of selenium-binding protein 1 is decreased in uterine leiomyoma Peng Zhang 0 Cunxian Zhang 0 Xudong Wang 0 Fang Liu 0 C James Sung 0 M Ruhul Quddus 0 W Dwayne Lawrence 0 0 Department of Pathology and Laboratory Medicine, Women & Infants Hospital of Rhode Island, Warren Alpert Medical School of Brown University , 101 Dudley Street, Providence, Rhode Island 02905 , USA Background: Selenium has been shown to inhibit cancer development and growth through the mediation of selenium-binding proteins. Decreased expression of selenium-binding protein 1 has been reported in cancers of the prostate, stomach, colon, and lungs. No information, however, is available concerning the roles of seleniumbinding protein 1 in uterine leiomyoma. Methods: Using Western Blot analysis and immunohistochemistry, we examined the expression of seleniumbinding protein 1 in uterine leiomyoma and normal myometrium in 20 patients who had undergone hysterectomy for uterine leiomyoma. Results and Discussion: The patient age ranged from 34 to 58 years with a mean of 44.3 years. Proliferative endometrium was seen in 8 patients, secretory endometrium in 7 patients, and atrophic endometrium in 5 patients. Two patients showed solitary leiomyoma, and eighteen patients revealed 2 to 5 tumors. Tumor size ranged from 1 to 15.5 cm with a mean of 4.3 cm. Both Western Blot analysis and immunohistochemistry showed a significant lower level of selenium-binding protein 1 in leiomyoma than in normal myometrium. Larger tumors had a tendency to show a lower level of selenium-binding protein 1 than smaller ones, but the difference did not reach a statistical significance. The expression of selenium-binding protein 1 was the same among patients with proliferative, secretory, and atrophic endometrium in either leiomyoma or normal myometrium. Also, we did not find a difference of selenium-binding protein 1 level between patients younger than 45 years and older patients in either leiomyoma or normal myometrium. Conclusions: Decreased expression of selenium-binding protein 1 in uterine leiomyoma may indicate a role of the protein in tumorigenesis. Our findings may provide a basis for future studies concerning the molecular mechanisms of selenium-binding protein 1 in tumorigenesis as well as the possible use of selenium in prevention and treatment of uterine leiomyoma. - Introduction Uterine leiomyoma, the most common neoplasm of the female genital tract, probably occurs in the majority of women by age 50 and is responsible for significant morbidity in patients [1-3]. Symptoms include pelvic pressure, pelvic pain, abnormal uterine bleeding, infertility, and miscarriage [4,5]. Uterine leiomyoma represents a major indication for hysterectomy among women in the United States, accounting for one-third of about 600,000 hysterectomy procedures performed annually [6,7]. Not only is hysterectomy associated with morbidity and mortality, but it also has a huge economic impact on healthcare systems [1]. The scientific literature contains a large body of information concerning the epidemiology, hormonal influence, genetics, and molecular alterations in uterine leiomyoma. Risk factors include early menarche, nulliparity, obesity, African-American ethnicity, and temoxifen use [8-13]. Many of these factors are associated with increased levels of estrogen and progesterone. Estrogen and progesterone act through the mediation of estrogen receptor and progesterone receptor, respectively. The majority of literature revealed higher concentrations of estrogen and progesterone receptors in leiomyoma than in normal myometrium [3]. Leiomyoma of the uterus also overexpresses various growth factors including transforming growth factor, fibroblastic growth factor, epidermal growth factor receptor, and platelet-derived growth factor [3]. Inherent abnormality of myometrium in patients has also been implicated in the development of leiomyoma since the myometrium in the uterus harboring leiomyoma shows a significantly higher level of estrogen receptor than that without tumor [14]. Leiomyoma of the uterus has been shown to be monoclonal by studies using X-linked glucose 6-phosphate dehydrogenase isozymes [15], X-linked androgen receptor [16,17], and X-linked phosphoglycerokinase [18]. Cytogenetic studies have identified several chromosomal alterations, including t(12;14), del(7q), 6p21, and trisomy 12 (3). However, it is unclear whether the genetic alterations occur before the genesis of leiomyoma or they are secondary events. Despite numerous studies concerning the molecular and genetic changes in uterine leiomyoma, the mechanisms of development remain unknown. Further work is needed to elucidate the pathogenesis that would lead to the discovery of effective prevention and treatment of the tumor. Selenium, an essential trace element, has been shown to have an anti-cancer effect. Many reports have described a relationship between insufficient selenium intake and increased risk of cancer [19-21]. The anticancer action of selenium is thought to be mediated by selenium-binding protein 1 (SELENBP1), a 56 kDa intracellular protein, that binds covalently to selenium. The gene of SELENBP1 is located at chromosome 1q21-22 [22]. The expression of SELENBP1 has been shown to be decreased in several tumors including cancers of the prostate, lungs, colon, and ovary [23-26]. However, little information exists concerning the role of SELENBP1 in tumorigenesis of uterine leiomyoma. In this study, we examined the expression of SELENBP1 in uterine leiomyoma and normal myometrium. Materials and methods The study consisted of 20 consecutive specimens of hysterectomy performed for leiomyoma at our institution in July 2004. We recorded the number and size of leiomyoma as well as the endometrial pattern in each patient. Using a monoclonal antibody against human SELENBP1 (Medical Biological Laboratory International Corporation, Watertown, MA), we evaluated the expression of SELENBP1 by Western Blot and immunohistochemistry. For Western Blot, 100 mg sample was taken from each leiomyoma of an unfixed uterine specimen. We selected areas of leiomyoma without degenerative changes. Also sampled was 100 mg of tissue from normal myometrium in the same uterus. The sample was immediately placed in 1 ml radioimmunoprecipitation assay buffer containing 50 mM Tris-HCl (pH7.4), 150 mM NaCl, 1% Triton X-100, 1% sodium deoxycholate, 0.1% SDS, 1 mM PMSF, 1 mM EDTA, 5 ug/ml aprotinin, 5 ug/ml leupeptin, 1 mM Na3VO4, and 5 mM NaF. After being cut into smaller pieces with scissors, the sample was homogenized on ice with a motor-driven tissue Tearor for 5 times, each for 10 seconds. The homogenate was placed on an orbital shaker at 4C for 30 minutes and then centrifuged at 4C with 14,000 g for 15 minutes. The supernatant was collected in a fresh tube and stored at -80C for later use. Protein concentration was determined by a Bradford protein assay (Bio-Rad, (...truncated)