Abstract
BackgroundAlpha 2 macroglobulin (A2M; also known as ovostatin), a homotetrameric protein with four disulfide-linked subunits, has the unique feature of inactivating/inhibiting most known proteases including serine-, threonine-, cysteine-, aspartic- and metalloproteases. In chickens, A2M has been identified and characterized biochemically, but little is known of its functional role(s) in the oviduct, hormonal regulation of expression or its expression in ovarian carcinomas in chickens. Therefore, we investigated estrogen regulation of A2M gene expression during development of the chicken oviduct, and its expression in normal and cancerous ovaries from chickens.MethodsTo determine tissue-specific expression of A2M in chickens, we collected various organs from male and female chickens and performed RT-PCR analyses. To examine A2M gene expression in the oviduct of 1-week-old female chicks that received a subcutaneous implant of 15 mg DES in the abdominal region for 20 days, we performed RT-PCR, qPCR and in situ hybridization analyses using cDNAs from control- (n = 5) and DES-treated oviducts (n = 5), and then each segment of the oviduct from DES-treated chicks. To determine if A2M is a biomarker of ovarian cancer in hens, we collected cancerous (n = 10) ovaries from a total of 136 chickens which had completely stopped egg-laying and performed RT-PCR and in situ hybridization analyses.ResultsWe found that A2M is most abundant in the chicken oviduct, specifically luminal (LE) and glandular epithelia (GE), but it was not detected in any other tissues of either sex. We then determined that DES (dietylstilbestrol, a synthetic nonsteroidal estrogen) increased A2M mRNA only in LE and GE of the oviduct of chicks. Further, expression of A2M was most abundant in GE of endometrioid adenocarcinoma of cancerous, but not normal ovaries of hens.ConclusionsCollectively, results of the present study indicate that A2M is novel estrogen-stimulated gene expressed in LE and GE of the chicken oviduct and may be used for monitoring effects of therapies for ovarian cancer in laying hens.
Highlights
Alpha 2 macroglobulin (A2M; known as ovostatin), a homotetrameric protein with four disulfidelinked subunits, has the unique feature of inactivating/inhibiting most known proteases including serine, threonine, cysteine, aspartic- and metalloproteases
The gene consists of 35 exons and the mRNA has 4,715 bp encoding a protein with 1,454 amino acid residues
Chicken A2M protein contains the highly conserved macroglobulin 1 (MG1) and MG2 domains, A2M family N-terminal region, and A2M receptor binding domain found in mammalian A2M proteins (Additional file 1, Supplemental Figure S1)
Summary
Alpha 2 macroglobulin (A2M; known as ovostatin), a homotetrameric protein with four disulfidelinked subunits, has the unique feature of inactivating/inhibiting most known proteases including serine-, threonine-, cysteine-, aspartic- and metalloproteases. A2M has been identified and characterized biochemically, but little is known of its functional role(s) in the oviduct, hormonal regulation of expression or its expression in ovarian carcinomas in chickens. We investigated estrogen regulation of A2M gene expression during development of the chicken oviduct, and its expression in normal and cancerous ovaries from chickens. A2M is known as an ovostatin or ovomacroglobulin. It is found in the oviduct and egg white, but not in other tissues or serum [13] and it has a strong anti-collagenase activity [14,15]. Little is known about regulation of its expression by steroid hormones in the oviduct or its expression in normal and cancerous ovaries
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