Abstract
Menstruation is the expulsion of the endometrial lining of the uterus following a nearly month long preparation for embryo implantation and pregnancy. Increasingly, the health of the endometrium is being recognized as a critical factor in female fertility, and proteomes and transcriptomes from endometrial biopsies at different stages of the menstrual cycle have been studied for both diagnostic and therapeutic purposes (1 Kao, L. C., et al. 2003 Endocrinology 144, 2870-2881; Strowitzki, Tet al. 2006 Hum. Reprod. Update 12, 617-630; DeSouza, L., et al. 2005 Proteomics 5, 270-281). Disorders of the uterus ranging from benign to malignant tumors, as well as endometriosis, can cause abnormal menstrual bleeding and are frequently diagnosed through endometrial biopsy (Strowitzki, Tet al. 2006 Hum. Reprod. Update 12, 617-630; Ferenczy, A. 2003 Maturitas 45, 1-14). Yet the proteome of menstrual blood, an easily available noninvasive source of endometrial tissue, has yet to be examined for possible causes or diagnoses of infertility or endometrial pathology. This study employed five different methods to define the menstrual blood proteome. A total of 1061 proteins were identified, 361 were found by at least two methods and 678 were identified by at least two peptides. When the menstrual blood proteome was compared with those of circulating blood (1774 proteins) and vaginal fluid (823 proteins), 385 proteins were found unique to menstrual blood. Gene ontology analysis and evaluation of these specific menstrual blood proteins identified pathways consistent with the processes of the normal endometrial cycle. Several of the proteins unique to menstrual blood suggest that extramedullary uterine hematopoiesis or parenchymal hemoglobin synthesis may be occurring in late endometrial tissue. The establishment of a normal menstrual blood proteome is necessary for the evaluation of its usefulness as a diagnostic tool for infertility and uterine pathologies. Identification of unique menstrual blood proteins should aid the forensic community in distinguishing menstrual blood from circulating blood.
Highlights
Menstrual blood is a complex biological fluid composed of blood, vaginal secretions, and the endometrial cells of the uterine wall as they exist immediately prior to menses
The nucleolar canal system (NCS) is believed necessary for the significant amount of protein synthesis required by the maturing endometrium in preparation for blastocyst implantation including the production of glycoproteins and extensive amounts of glycogen [6, 8]
Comparison of Menstrual Blood, Vaginal Fluid, and Venous Blood Proteomes—Menstrual blood is composed of three distinct body fluids: blood, vaginal fluid, and the cells and fluid of the late secretory phase of the uterine endometrial lining which is shed during menstruation
Summary
Materials—Chemical reagents including salts, chaotropic salts, buffers, detergents, organics, alpha-cyano-4-hydroxycinnamic acid (CHCA), Bradford protein assay kit and albumin and IgG immunodepletion ProteoPrep Immunoaffinity kit were purchased from Sigma (St. Louis, MO). RIPA buffer was from Fisher (Pittsburgh, PA). Ni-nitrilotriacetic acid (NTA) agarose beads were purchased from Qiagen (Valencia, CA). Sequencing grade modified trypsin was from Promega (Madison, WI). Precast 15% PAGE gels and immobilized pH
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