Alpha-2-macroglobulin is, like alpha-1-antitrypsin, alpha-2-antiplasmin, and antithrombin III, a protease inhibitor. It inhibits many proteases, including trypsin, thrombin and collagenase. Webb et al. (1996) demonstrated that murine alpha-2-macroglobulin binds TGF-beta and inhibits TGF-beta-receptor interactions.
NCBI Summary:
The protein encoded by this gene is a protease inhibitor and cytokine transporter. It uses a bait-and-trap mechanism to inhibit a broad spectrum of proteases, including trypsin, thrombin and collagenase. It can also inhibit inflammatory cytokines, and it thus disrupts inflammatory cascades. Mutations in this gene are a cause of alpha-2-macroglobulin deficiency. This gene is implicated in Alzheimer's disease (AD) due to its ability to mediate the clearance and degradation of A-beta, the major component of beta-amyloid deposits. A related pseudogene, which is also located on the p arm of chromosome 12, has been identified. [provided by RefSeq, Nov 2016]
General function
Metabolism
Comment
alpha 2-Macroglobulin (alpha 2M) is a broad spectrum protease inhibitor associated with inflammatory responses and proposed to be important in tissue remodeling. alpha 2 M also functions as a carrier of specific growth factors and cytokines, including platelet-derived growth factor, transforming growth factor-beta, basic fibroblast growth factor, interleukin-1, interleukin-6. Vaughan et al. reported that alpha 2-macroglobulin is a binding protein of inhibin and activin. Mather (1996) reviewed the literature on alpha 2-macroglobulin as a soluble binding protein for inhibin and activin.
Cellular localization
Secreted
Comment
Using the follicular fluid of porcine ovaries as a source, Kohyama et al. (1998) purified a high-molecular-weight protein having enzyme activity toward t-butyloxycarbonyl-Gln-Arg-Arg-4-methylcoumaryl-7-amide. Its molecular weight was estimated to be approximately 720,000. The protein was immunoprecipitated with antihuman alpha 2-macroglobulin antibody and was visualized in Western blot
analysis using the same antibody. The complexed enzyme of alpha 2-macroglobulin and follipsin activated a single-chain precursor tissue-type plasminogen activator at a drastically reduced rate compared with free follipsin.
Hampered cumulus expansion of porcine cumulus-oocyte complexes by excessive presence of alpha2 -macroglobulin is likely mediated via inhibition of zinc-dependent metalloproteases. Appeltant R et al. (2017) In vitro maturation (IVM) in serum causes hampered expansion of porcine cumulus-oocyte complexes (COCs) due to excessive alpha2 -macroglobulin (A2M). This study investigated two hypotheses that could explain the effect of A2M: (i) binding of epidermal growth factor (EGF) to A2M, followed by its decreased availability; and (ii) inhibition of zinc-dependent metalloproteases. Cumulus expansion was evaluated based on the diameter of the COCs, the proportion of COCs participating in a floating cloud and the proportion of COCs with loss of cumulus cells. The first hypothesis of decreased EGF availability was tested by increasing the EGF concentration (20 and 50 ng/mL vs. 10 ng/mL), but was not confirmed because cumulus expansion did not improve. To verify the second hypothesis of inhibited zinc-dependent metalloproteases, the effect of tissue inhibitor of metalloproteases-3 (TIMP-3) on cumulus expansion during IVM with and without A2M was investigated. To immuno-neutralize A2M, serum was pre-incubated with A2M antibodies. Impaired cumulus expansion because of TIMP-3 could only be observed during IVM in 10% of serum with A2M antibodies. No effect of TIMP-3 was observed in medium without A2M antibodies. These results indicate that A2M and TIMP-3 share a common target, a zinc-dependent metalloprotease. Future research is directed toward the identification of the protease involved.//////////////////
Curry et al reported the presence of alpha 2-macroglobulin in follicular fluid. It was proposed that TIMP produced in the granulosa cell compartment in conjunction with alpha 2-macroglobulin from the serum may act to control the site and extent of ovarian connective tissue remodeling.Ireland JL, et al reported evidence for for autocrine or paracrine roles of {alpha}2-macroglobulin in regulation of estradiol production by granulosa cells and development of dominant follicles.
alpha2-Macroglobulin (alpha2-M) inhibits proteinases and modulates the actions of growth factors and cytokines. Despite the key roles proteinases, growth factors, and cytokines have in folliculogenesis, the role of alpha2-M in follicular development is unknown. Our objectives were to: a) determine if granulosa cells produce alpha2-M and have alpha2-M receptors, b) examine the effect of alpha2-M on estradiol production by granulosa cells, c) establish whether amounts of alpha2-M and alpha2-M receptors were altered during dominant non-ovulatory follicle development, and d) examine alpha2-M's mechanism of action. The results demonstrated that bovine granulosa cells contain 5.2 Kb and 15 Kb mRNAs, and 720 kDa and 500 kDa proteins that correspond respectively to sizes of mRNAs and proteins for alpha2-M and the alpha2-M receptor. Treatment of granulosa cells with alpha2-M resulted in a specific dose responsive increase in estradiol production. Cell viability, cell number, and the amount of aromatase in granulosa cells were not altered by alpha2-M. Treatment of granulosa cells with factors that bind alpha2-M or its receptor did not mimic alpha2-M action. Although intrafollicular amounts of alpha2-M remained unchanged, amounts of alpha2-M receptor in granulosa cells were strongly inversely associated with concentrations of estradiol in dominant and subordinate follicles. Based on these results, we concluded that alpha2-M may have autocrine or paracrine roles in granulosa cells potentially important for regulation of estradiol production and development of dominant follicles.
Comparative proteome analysis of porcine follicular fluid and serum reveals that excessive alpha(2)-macroglobulin in serum hampers successful expansion of cumulus-oocyte complexes. Bijttebier J et al. Follicular fluid (pFF) constitutes the micro-environment of the maturing oocyte. Although pFF is a transudate of serum, in pigs, it is superior to serum in promoting in vitro expansion of the cumulus cells, a specialized cell population surrounding the oocyte. A comparative proteome analysis of autologous serum and pFF was performed to investigate proteins involved in successful cumulus expansion of porcine oocytes. iTRAQ labeling followed by 2-D LC ESI-Q-TOF MSMS revealed 63 proteins common to both fluids of which the abundance of 13 proteins was significantly different (P<0.05). Seven proteins were more concentrated in serum whereas 6 proteins were more abundant in pFF. To investigate the importance of these proteins, the cumulus matrices of cumulus-oocyte complexes (COCs) were collected after in vitro maturation (IVM) in media supplemented with either of both biologically fluids and then subjected to 2-D PAGE analysis. Alpha(2)-macroglobulin (A2M) and CH4 and secrete domains of swine IgM, which were both less abundant in pFF, were absent from cumulus matrix extracts after IVM in pFF. Although both proteins were incorporated in the matrices of COCs matured in serum, depletion of A2M from serum could significantly compensate for the impaired cumulus expansion of oocytes matured in serum.
Expression regulated by
FSH, LH, Steroids, prolactin
Comment
Gaddy-Kurten et al. (1991) demonstrate that alpha 2M mRNA and protein are synthesized in thecal cells of developing follicles in response to low levels of LH. Conversely, alpha 2M mRNA and protein are only synthesized by granulosa cells of follicles that have been stimulated to luteinize either in vivo by the LH surge or in vitro by FSH and testosterone and are also exposed to PRL. The obligatory requirement for PRL is specific; associated with increased numbers of PRL-binding sites; mediated by time-dependent appearance of alpha 2M in the endoplasmic reticulum (12 h), Golgi apparatus (24 h), and secretion vesicles (48 h); and involves in part increased transcription of the alpha 2M gene. Dajee et al (1996) reported the prolactin induction of the alpha 2-Macroglobulin gene in rat ovarian granulosa cells and the association with stat 5 activation and binding to the interleukin-6 response element. They documented that Stat 5b, SF-1, and COUP-TF each exert specific effects on the function of the alpha2M promoter: basal activity is controlled by the balance of SF-1 (positive) and COUP-TF (negative) activities and PRL inducibility is mediated by activation of Stat 5b.
Ovarian localization
Granulosa, Theca, Luteal cells
Comment
Gaddy-Kurten et al. (1989) studied the expression of alpha 2M mRNA and protein in specific ovarian cell types during ovulation, luteinization, and luteolysis. Before ovulation, alpha 2M mRNA is not detectable in granulosa cells. Twelve hours after injection of an ovulatory dose of hCG alpha 2M mRNA is detectable in luteinizing follicles, which is increased further by 48 h and maintained in corpora lutea (CL) for up to 96 h. Administration of PRL from 24-96 h results in both inhibition of luteolysis and marked increases in alpha 2M mRNA in CL, but not in residual tissues.
Follicle stages
Comment
Gaddy-Kurten et al. (1989) reported that alpha 2M protein (188K monomer) is present (immunoblot and immunofluorescence data) in small antral and preovulatory follicles even though mRNA is not detectable; after an ovulatory dose of hCG the protein level transiently increases by 12 h and declines thereafter through 96 h.
Phenotypes
PCO (polycystic ovarian syndrome)
Mutations
3 mutations
Species: mouse
Mutation name: None
type: null mutation fertility: fertile Comment:Umans et al. (1995) reported that alpha 2-Macroglobulin-deficient mice were viable and produced normally sized litters with normal sex ratio over 3 generations. Knock-out mice were more resistant to endotoxin but more sensitive to a choline-free diet supplemented with ethionine.
Species: human
Mutation name: None
type: naturally occurring fertility: fertile Comment: On screening of a normal materia, Bergqvist et al.(1979) reported that 1 man was found to have a low alpha 2M, namely 25 % with electroimmuno assay. Investigation of the family revealed that the mother (age 69) and one daughter (age 5) had low values too. All other coagulation and fibrinolytic components were normal. They had no signs of increased fibrinolysis and normal levels of alpha 2-antiplasmin, alpha 1-antitrypsin and antithrombin III. Liver function tests were normal. Poller et al. (1989) reported the detection of an alteration of the alpha-2-macroglobulin gene in a patient with chronic lung disease and serum alpha-2-macroglobulin deficiency.
Species: human
Mutation name: type: None fertility: None Comment: Proteomic Analysis of Plasma in the Polycystic Ovary Syndrome Identifies Novel Markers Involved in Iron Metabolism, Acute-Phase Response, and Inflammation. Insenser M et al. Context: Hypothesis-free approaches such as proteomic analysis may identify novel biomarkers for disease. Objective: The objective of the study was to compare the plasma proteome of patients presenting with the polycystic ovary syndrome (PCOS) with that of women without hyperandrogenism. Design: This was a case-control study. Settings: The study was conducted at an academic hospital. Patients: Patients included 12 PCOS patients and 12 women without hyperandrogenism. Interventions: Interventions included basal blood sampling. Main Outcome Measures: Two-dimensional difference in-gel electrophoresis, matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry, Western blot, and ELISA analyses were measured. Results: Two-dimensional difference in-gel electrophoresis and matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry analyses identified haptoglobin beta-chain and alpha2-macroglobulin as proteins underexpressed in PCOS samples, whereas transferrin and kappa-free light chain were overexpressed. We were able to confirm only the underexpression of haptoglobin beta-chain in subsequent Western blot and ELISA analyses. Conclusions: Proteomic analysis of plasma from PCOS patients revealed changes in protein expression in several acute-phase response proteins including isoforms of plasma haptoglobin, alpha2-macroglobulin, and transferrin and in kappa-free light chain. In addition to their role as inflammatory markers, some of these molecules play major roles in iron metabolism, further suggesting that iron metabolism and low-grade chronic inflammation may be involved in the pathogenesis of insulin-resistant disorders such as PCOS.//////////////Validation of proteomic biomarkers previously found to be differentially expressed in women with Polycystic Ovary Syndrome: a cross-sectional study. Haoula Z 2014 et al.
Abstract Objectives: The aim of this study was to independently validate proteomic biomarkers previously reported to be differentially expressed in women with Polycystic Ovary Syndrome (PCOS) compared with controls. This study focused on plasma proteomic biomarkers. Methods: This was a cross-sectional study at the University of Nottingham, in which samples from 30 PCOS and 30 control women were analysed by Western blotting. Results: Mean abundance ratios from Western blots of plasma total haptoglobin and haptoglobin beta proteins were 1.25 (CI 1.11-1.4) and 1.24 (CI 1.04-1.44). The mean abundance ratio from the blots of alpha-2 macroglobulin was however 1.05 (CI, 1-1.1). The mean PCOS/control BMI ratio was 1.18 (CI 1.17-1.20). There was no correlation between PCOS/control BMI ratio and alpha-2 macroglobulin, total haptoglobin and haptoglobin beta abundance ratios. There was also no correlation between PCOS/control insulin ratio and alpha-2 macroglobulin, total haptoglobin and haptoglobin beta abundance ratios. Conclusions: Total haptoglobin and haptoglobin beta chain protein abundance was found to be elevated in women with PCOS compared with controls. We were unable to confirm decreased alpha-2 macroglobulin levels as reported in a previous study. Independent validation studies are required to validate early promising proteomic biomarkers in PCOS.
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