Biomedical subjects

Cheryl A Conover

Publications and source records attributed to Cheryl A Conover.

At least 19 recordsLinked to original sources

PubMed2007

Pregnancy-associated plasma protein-A (PAPP-A): a local regulator of IGF bioavailability through cleavage of IGFBPs.

Pregnancy-associated plasma protein-A (PAPP-A) was originally isolated in 1974, as one of four proteins of placental origin found in high concentrations in the blood of pregnant women. In the early 1990s several laboratories reported novel protease activity against insulin-like growth factor binding protein-4 (IGFBP-4) in media conditioned by several cell types. This activity was unique, as it appeared to require the presence of IGF to cleave IGFBP-4. In 1999, this IGF-dependent IGFBP-4 protease activity was isolated from human fibroblast conditioned media and identified as PAPP-A. Subsequently, PAPP-A was shown to be expressed by a variety of cell types, and thus no longer could be considered to be just "pregnancy-associated". This review will describe what is currently known about the structure of PAPP-A and about its function as an IGFBP protease, with a focus on new insights obtained through study of a PAPP-A knock-out mouse model and on potential clinical applications.

Amino Acid Sequence↗

PubMed2006

Cell surface detachment of pregnancy-associated plasma protein-A requires the formation of intermolecular proteinase-inhibitor disulfide bonds and glycosaminoglycan covalently bound to the inhibitor.

The metzincin metalloproteinase pregnancy-associated plasma protein-A (PAPP-A, pappalysin-1) promotes cell growth by proteolytic cleavage of insulin-like growth factor-binding proteins 4 and 5, causing the release of bound insulin-like growth factors. PAPP-A binds an unknown cell-surface heparan sulfate proteoglycan, suggesting that it controls insulin-like growth factor signaling spatially. In human pregnancy, the majority of PAPP-A circulates as a disulfide-bonded complex with its inhibitor, the proform of eosinophil major basic protein (proMBP). Interestingly, Ser-62 of proMBP is substituted with a glycosaminoglycan (GAG) chain, possibly a heparan sulfate type, and the PAPP-A.proMBP complex is unable to bind to the cell surface. We show here that proMBP detaches surface-bound PAPP-A in a process that depends on the proMBP GAG and also on the formation of intermolecular disulfide bonds between PAPP-A and proMBP. Unlike what was expected, we demonstrate that the GAG of proMBP is not required for PAPP-A.proMBP complex formation and that proMBP residues His-137, Ser-178, Arg-179, and Asn-181 are important for the recognition of PAPP-A. Using a mouse model, we find that the half-life of circulating PAPP-A and proMBP in complex is severalfold higher than both of the uncomplexed proteins, further suggesting that the PAPP-A.proMBP complex is formed at the cell surface in vivo rather than in the circulation. Further supporting this, we show that formation of the PAPP-A.proMBP complex at the cell surface proceeds rapidly compared with the slow rate of complex formation in solution. Because both PAPP-A and proMBP are expressed ubiquitously, this model may be applicable to many tissues in which insulin-like growth factor bioavailability is locally regulated.

Amino Acid Sequence↗

PubMed2006

Surface association of pregnancy-associated plasma protein-A accounts for its colocalization with activated macrophages.

Intense immunostaining for pregnancy-associated plasma protein-A (PAPP-A), a newly characterized metalloproteinase in the insulin-like growth factor system, colocalizes with activated macrophages in human atherosclerotic plaque. To determine macrophage regulation of PAPP-A expression, we developed two models of human macrophages with basal and activated phenotypes. THP-1 cells and peripheral blood monocytes could be differentiated into macrophages and activated upon specific treatment regimens with phorbol myristate acetate, macrophage colony-stimulating factor, and interleukin-1beta. Activation was assessed by cell secretion of tumor necrosis factor-alpha, which increased 30- to 100-fold with activation. Activated macrophages also secreted matrix metalloproteinase-9. However, no PAPP-A mRNA or PAPP-A antigen could be detected in these cells under any condition. Upon incubation with recombinant PAPP-A, we found that activated macrophages bound and internalized more PAPP-A than unactivated macrophages or monocytes. Internalization accounted for at least 50% of macrophage-associated PAPP-A, as assessed in studies with cytochalasin B. Membrane-bound PAPP-A retained protease activity, whereas internalized PAPP-A had little or no activity. Similar experiments carried out with a mutated variant of PAPP-A, which retains functionality as a protease but is unable to bind surface-associated glycosaminoglycan, showed no macrophage association or internalization. Absence of PAPP-A expression was confirmed in activated macrophages isolated from a hypercholesterolemic rabbit model of atherosclerosis. We therefore conclude that PAPP-A is not synthesized in, but rather is bound and internalized by, macrophages. Our findings likely account for the observed intense immunostaining for PAPP-A colocalizing with activated macrophages and may have physiological significance in the development of vulnerable plaque.

Animals↗

PubMed2006

Targeted disruption of the pregnancy-associated plasma protein-A gene is associated with diminished smooth muscle cell response to insulin-like growth factor-I and resistance to neointimal hyperplasia after vascular injury.

IGF-I is an important determinant of the vascular response to injury in large part through its ability to stimulate migration and proliferation of smooth muscle cells (SMCs). In this study, we used mice with targeted disruption of the pregnancy-associated plasma protein-A gene (PAPP-A-/-) and wild-type (WT) littermates to test the hypotheses that PAPP-A, a metalloproteinase that cleaves inhibitory IGF binding protein (IGFBP)-4, regulates vascular SMC responses to IGF-I in vitro and is critical for the development of vascular neointima after injury in vivo. Vascular SMCs from PAPP-A-/- mice lacked IGFBP-4 protease activity and failed to respond to treatment with IGF-I in the presence of IGFBP-4, whereas SMCs from WT mice with robust IGFBP-4 protease activity showed significant migratory and proliferative responses to IGF-I/IGFBP-4. For in vivo testing, PAPP-A-/- and WT mice underwent unilateral carotid ligation, a model of injury-induced neointimal hyperplasia. In WT mice, PAPP-A mRNA expression was markedly elevated 7 and 14 d after carotid ligation, associated with a progressive increase in neointimal hyperplasia and, in many cases, with complete occlusion of the vessel at 28 d. In contrast, PAPP-A-/- mice showed little evidence of progression resulting in a 75% reduction in neointimal area when compared with WT at 28 d. Cells staining for proliferating cell nuclear antigen were plentiful in the SMC-rich medial and neointimal areas of the injured WT vessel in stark contrast to the relatively few proliferating cells in the same areas of the PAPP-A-/- vessel. Expression of IGF-I and IGFBP-4 was similarly elevated in injured carotids from WT and PAPP-A-/- mice with no change in IGF-I receptor expression. IGFBP-5, an IGF-responsive gene, was increased 2-fold in WT but not in PAPP-A-/- carotids, suggesting reduced IGF activity in the absence of PAPP-A. Thus, PAPP-A-deficient mice are resistant to neointimal formation after injury, which may be explained in part by the ability of PAPP-A to enhance local IGF-I stimulation of vascular SMCs through proteolysis of IGFBP-4.

Animals↗

PubMed2006

Pregnancy associated plasma protein-A and risk stratification of patients presenting with chest pain in the emergency department.

BACKGROUND: The aim of this study was to evaluate the clinical utility of serum pregnancy associated plasma protein-A (PAPP-A) levels in assisting triage of an intermediate to high-risk patient presenting with chest pain in the Emergency Department and no definite evidence of an acute coronary syndrome. METHODS: Serum levels of PAPP-A were measured in 59 patients presenting with chest pain to the Emergency Department. The patients were independently grouped according to the presence of acute coronary syndromes or the absence thereof. RESULTS: In a multivariate model that corrected for age, sex, type of chest pain, number of risk factors, history of coronary artery disease, troponin levels, and non-specific ECG changes, PAPP-A levels were still predictive of a final diagnosis of acute coronary syndrome in patients presenting with chest pain to the Emergency Department (Odds Ratio, 2.093; 95th confidence intervals, 1.037-4.224; p=0.039). CONCLUSIONS: Elevated serum PAPP-A levels were predictive of a diagnosis of acute coronary syndrome in intermediate- to high-risk patients presenting to the Emergency Department with chest pain and no definite evidence of an acute coronary syndrome. Thus, serum PAPP-A may be valuable as an adjunct, minimally invasive marker to improve risk stratification in chest pain patients.

Acute Disease↗

PubMed2006

Prognostic value of circulating pregnancy-associated plasma protein levels in patients with chronic stable angina.

AIMS: Unstable coronary atherosclerotic plaque can be present in patients with chronic stable coronary artery disease (CAD). Our objective was to assess whether measurement of plasma pregnancy-associated plasma protein (PAPP-A) level, a reflection of plaque instability, in patients with chronic stable CAD had an independent prognostic value on the subsequent incidence of death, acute coronary syndrome (ACS), and revascularization. METHODS AND RESULTS: Patients referred for coronary angiography were recruited. A cohort of 103 patients with stable symptoms for at least 6 weeks and with a coronary angiogram showing at least a 50% luminal diameter narrowing formed our study population. Median follow-up was 4.9 years. Mean age was 65+/-10 years. In a multivariable model that included CAD traditional risk factors, ejection fraction, extent of coronary atherosclerosis, prior history of myocardial infarction, prior revascularization, discharge medications, and C-reactive protein, the plasma PAPP-A was found to be significantly associated with the endpoint of future death [adjusted hazard ratio (HR) 5.29; 95% CI 1.27-22.0; P=0.023] and with the endpoint of future death and ACS (adjusted HR 3.56; 95% CI 1.27-10.0; P=0.015), but not with the endpoint of future death and revascularization. CONCLUSION: Measurement of plasma PAPP-A level in patients with chronic stable CAD has an independent prognostic value on the occurrence of death and ACS.

Aged↗

PubMed2006

Pregnancy-associated plasma protein-a is markedly expressed by monocyte-macrophage cells in vulnerable and ruptured carotid atherosclerotic plaques: a link between inflammation and cerebrovascular events.

OBJECTIVES: The study aim was to evaluate serologic expression of pregnancy-associated protein-A (PAPP-A) in patients affected by cerebrovascular accidents and to correlate it with histopathologic carotid plaque complexity. BACKGROUND: Little is known about PAPP-A expression in carotid atherosclerotic disease and whether this protein represents a marker of plaque vulnerability also in carotid district. METHODS: Seventy-two carotid plaques from patients submitted to surgical endarterectomy (19 who suffered a major stroke, 24 transient ischemic attack, and 29 asymptomatic) were evaluated. Serologic PAPP-A levels were determined by enzyme-linked immunoadsorbent assay. Plaques were divided in three groups based on histology: 1) stable (n = 38); 2) vulnerable (n = 13); 3) ruptured with thrombus (n = 14). Immunohistochemical staining for PAPP-A, smooth muscle cells, macrophages, and T-lymphocytes was performed in all cases. Real-time polymerase chain reaction assessed local PAPP-A production, and double immunofluorescence confocal microscopy (ICM) characterized cell type expressing PAPP-A. RESULTS: Pregnancy-associated protein-A (serologic values were 4.02 +/- 0.18 mIU/l in Group 1, 7.43 +/- 0.97 mIU/l in Group 2, and 6.97 +/- 0.75 mIU/l in Group 3 [1 vs. 3, p = 0.01; 1 vs. 2, p = 0.004; 2 vs. 3, p = 0.71, respectively]). Pregnancy-associated protein-A (expression showed a mean score value of 0.62 +/- 0.06 for stable plaques, 2.54 +/- 0.14 for vulnerable plaques, and 2.71 +/- 0.12 for ruptured plaques [1 vs. 2, p = 0.001; 1 vs. 3, p = 0.001; 2 vs. 3, p = 0.37, respectively]). Real-time polymerase chain reaction demonstrated local messenger ribonucleic acid PAPP-A production, and double ICM confirmed monocyte/macrophage expression of PAPP-A in Groups 2 and 3 but not Group 1. CONCLUSIONS: This study suggests that PAPP-A is a marker of carotid plaque destabilization and rupture. Further studies are necessary to determine if PAPP-A can represents a new target for stratifying the risk of cerebrovascular events.

Aged↗

PubMed2006

In vitro and in vivo antitumor effects of the dual insulin-like growth factor-I/insulin receptor inhibitor, BMS-554417.

The insulin-like growth factor receptor (IGF-IR) and insulin receptor are either overactivated and/or overexpressed in a wide range of tumor types and contribute to tumorigenicity, proliferation, metastasis, and drug resistance. Here, we show that BMS-554417, a novel small molecule developed as an inhibitor of IGF-IR, inhibits IGF-IR and insulin receptor kinase activity and proliferation in vitro, and reduces tumor xenograft size in vivo. In a series of carcinoma cell lines, the IC50 for proliferation ranged from 120 nmol/L (Colo205) to >8.5 micromol/L (OV202). The addition of stimulatory ligands was unnecessary for the antiproliferative effect in MCF-7 and OV202 cells. BMS-554417 treatment inhibited IGF-IR and insulin receptor signaling through extracellular signal-related kinase as well as the phosphoinositide 3-kinase/Akt pathway, as evidenced by decreased Akt phosphorylation at Ser473. At doses that inhibited proliferation, the compound also caused a G0-G1 arrest and prevented nuclear accumulation of cyclin D1 in response to LR3 IGF-I. In Jurkat T-cell leukemia cells, this agent triggered apoptotic cell death via the mitochondrial pathway. BMS-554417 was orally bioavailable and significantly inhibited the growth of IGF1R-Sal tumor xenografts in vivo. BMS-554417 is a member of a novel class of IGF-IR/insulin receptor inhibitors that have potential clinical applications because of their antiproliferative and proapoptotic activity in vitro and in vivo.

Animals↗

PubMed2006

Cytokine stimulation of pregnancy-associated plasma protein A expression in human coronary artery smooth muscle cells: inhibition by resveratrol.

Through specific cleavage of proteins that bind and inhibit insulin-like growth factor-I (IGF-I), pregnancy-associated plasma protein-A (PAPP-A) enhances local IGF-I availability, and, consequently, receptor activation. PAPP-A expression is increased in experimental models of vascular injury and in human atherosclerotic plaque; however, little is known about the regulation of PAPP-A gene expression in vascular cells. In this study, we tested the hypothesis that proinflammatory cytokines involved in the vascular injury response stimulate PAPP-A gene expression in human coronary artery smooth muscle cells (hCASMC) in culture. Tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta stimulated PAPP-A gene expression in a time- and dose-dependent manner. The effect of these cytokines appears to be at the level of transcription because actinomycin D completely prevented the induction of PAPP-A gene expression. Accumulation of PAPP-A in cell-conditioned medium paralleled mRNA synthesis, as did proteolytic activity against IGF binding protein-4 (IGFBP-4). Interestingly, pretreatment of hCASMC with resveratrol, a polyphenol found in the skin of grapes and in red wine purported to underlie the "French paradox," inhibited TNF-alpha- and IL-1beta-induced PAPP-A expression and, hence, its IGFBP-4 proteolytic activity. Resveratrol had no effect on basal PAPP-A expression and protease activity. Our finding that PAPP-A gene expression in hCASMC is stimulated by TNF-alpha and IL-1beta suggests a mechanism for the regulation of PAPP-A in response to vascular injury that may contribute to the enhanced IGF-I bioactivity in intimal hyperplasia and atherosclerotic plaque development. Our results also suggest that PAPP-A may be a target of the cardiovascular system-protective effects of resveratrol.

Anti-Inflammatory Agents, Non-Steroidal↗

PubMed2005

Stress-activated signaling pathways mediate the stimulation of pregnancy-associated plasma protein-A expression in cultured human fibroblasts.

Pregnancy-associated plasma protein-A (PAPP-A) is an IGF binding protein protease that appears to function as a posttranslational modulator of IGF bioavailability in response to injury. A previous study indicated that the proinflammatory cytokines, TNFalpha and IL-1beta, were potent stimulators of PAPP-A expression in cultured human fibroblasts. In this study, we investigated the intracellular signaling pathways mediating cytokine-stimulated PAPP-A expression. Treatment of human fibroblasts with TNFalpha and IL-1beta (1 nm) had little or no effect on phosphatidylinositol 3-kinase and Erk1/2 activation, pathways commonly associated with proliferation. On the other hand, TNFalpha and IL-1beta induced p38, c-Jun N-terminal kinase (JNK), and nuclear factor (NF)kappaB activation, pathways more closely related to stress response. An inhibitor of p38 activation (SB203580) had no effect on TNFalpha- or IL-1beta-stimulated PAPP-A expression. The JNK inhibitor, SP600125, had no effect on IL-1beta- or TNFalpha-stimulated PAPP-A mRNA expression. However, SP600125 effectively inhibited IL-1beta-induced PAPP-A protein expression. MG-132, a proteasome inhibitor that blocked degradation of the intrinsic NFkappaB inhibitor, IkappaB, and thereby prevented NFkappaB activation, was a potent inhibitor of both TNFalpha- and IL-1beta-stimulated PAPP-A mRNA and protein expression and IGF binding protein-4 protease activity. MG-132 had no effect on JNK phosphorylation or p38 activation, and SB203580 and SP600125 had no effect on IkappaB degradation, documenting inhibitor specificity. BAY11-7082, another inhibitor of NFkappaB activation, also inhibited TNFalpha- and IL-1beta-stimulated PAPP-A expression and IGF binding protein-4 protease activity. These data indicate that NFkappaB activation is the primary mediator of cytokine-stimulated PAPP-A expression in human fibroblasts.

Adult↗

PubMed2005

Disruption of insulin-like growth factor-II imprinting during embryonic development rescues the dwarf phenotype of mice null for pregnancy-associated plasma protein-A.

Pregnancy-associated plasma protein-A (PAPP-A), an insulin-like growth factor-binding protein (IGFBP) protease, increases insulin-like growth factor (IGF) activity through cleavage of inhibitory IGFBP-4 and the consequent release of IGF peptide for receptor activation. Mice homozygous for targeted disruption of the PAPP-A gene are born as proportional dwarfs and exhibit retarded bone ossification during fetal development. Phenotype and in vitro data support a model in which decreased IGF-II bioavailability during embryogenesis results in growth retardation and reduction in overall body size. To test the hypothesis that an increase in IGF-II during embryogenesis would overcome the growth deficiencies, PAPP-A-null mice were crossed with DeltaH19 mutant mice, which have increased IGF-II expression and fetal overgrowth due to disruption of IgfII imprinting. DeltaH19 mutant mice were 126% and PAPP-A-null mice were 74% the size of controls at birth. These size differences were evident at embryonic day 16.5. Importantly, double mutants were indistinguishable from controls both in terms of size and skeletal development. Body size programmed during embryo development persisted post-natally. Thus, disruption of IgfII imprinting and consequent elevation in IGF-II during fetal development was associated with rescue of the dwarf phenotype and ossification defects of PAPP-A-null mice. These data provide strong genetic evidence that PAPP-A plays an essential role in determining IGF-II bioavailability for optimal fetal growth and development.

Animals↗

PubMed2004

The Lin12-notch repeats of pregnancy-associated plasma protein-A bind calcium and determine its proteolytic specificity.

The Lin12-Notch repeat (LNR) module of about 35 residues is a hallmark of the Notch receptor family. Three copies, arranged in tandem, are invariably present in the extracellular portion of the Notch receptors. Although their function is unknown, genetic and biochemical data indicate that the LNR modules participate in the regulation of ligand-induced proteolytic cleavage of the Notch receptor, a prerequisite to intramembrane cleavage and Notch signaling. Outside the Notch receptor family, the LNR module is present only in the metalloproteinase pregnancy-associated plasma protein-A (PAPP-A) and its homologue PAPP-A2, which also contain three copies. Curiously, LNR modules 1 and 2 are present within the proteolytic domain of PAPP-A/A2, but LNR3 is separated from LNR2 by more than 1000 amino acids. The growth factor antagonists insulin-like growth factor-binding protein (IGFBP)-4 and -5 are both substrates of PAPP-A. We provide here evidence that the PAPP-A LNR modules function together to determine the proteolytic specificity of PAPP-A. Analysis of C-terminally truncated PAPP-A mutants followed by the analysis of LNR deletion mutants demonstrated that each of the three PAPP-A LNR modules is strictly required for proteolytic activity against IGFBP-4 but not for proteolytic activity against IGFBP-5. Individual substitution of conserved LNR residues predicted to participate in calcium coordination caused elimination (D341A, D356A, D389A, D1484A, D1499A, and D1502A) or a significant reduction (D359A and E392A) of IGFBP-4 proteolysis, whereas IGFBP-5 proteolysis was unaffected. The activity of the latter mutants against IGFBP-4 could be partially rescued by calcium, and the addition of the calcium-binding protein calbindin D9k to wild-type PAPP-A eliminated activity against IGFBP-4 but not against IGFBP-5, demonstrating that the PAPP-A LNR modules bind calcium ions. We propose a model in which LNR3 is spatially localized in proximity to LNR1 and -2, forming a single functional unit.

Amino Acid Sequence↗

PubMed2004

Pregnancy-associated plasma protein-A (PAPP-A) expression and insulin-like growth factor binding protein-4 protease activity in normal and malignant ovarian surface epithelial cells.

Pregnancy-Associated Plasma Protein-A (PAPP-A) proteolyses insulin-like growth factor binding protein-4 (IGFBP-4), thereby regulating local IGF availability. Reduced PAPP-A mRNA expression has been reported in ovarian cancer specimens compared to normal ovarian surface epithelial cells (OSE). To characterize PAPP-A expression and proteolytic activity in OSE, we developed a lifespan-extended human cell model using a temperature-sensitive mutant of the SV40 large T antigen (SV40LT). These OSE(tsT) cells proliferate at 34 degrees C (i.e., when SV40LT-positive), but not at 39 degrees C, a temperature at which the SV40LT is unstable (SV40LT-negative). Proteolysis of radiolabeled IGFBP-4 in conditioned media from OSE(tsT) lines was IGF-dependent and blocked by anti-PAPP-A antisera. Temperature shifts that eliminated stable SV40LT induced a 7-fold increase in PAPP-A mRNA and a 4-fold increase in protein. The converse experiment (shifting to SV40LT-positive conditions) resulted in decreased levels of PAPP-A mRNA but little change in PAPP-A protein. Nevertheless, there was a marked reduction in IGF-BP-4 proteolytic activity in medium of SV40LT-positive OSE-(tsT) cells. This decreased PAPP-A activity coincided with a nearly 20-fold increase in mRNA encoding a physiological inhibitor of PAPP-A, the precursor form of eosinophil Major Basic Protein (proMBP), and 4- to 5-fold increases in proMBP protein. Primary cultures of unmodified OSE expressed high levels of PAPP-A and undetectable proMBP, and therefore produced abundant IGFBP-4 protease activity. Short-term ovarian tumor cell cultures expressed variable levels of PAPP-A and high levels of proMBP, and consequently secreted little or no IGFBP-4 protease activity. The concurrent regulation of PAPP-A and its inhibitor, proMBP, suggests that IGFBP-4 proteolysis and local regulation of IGF availability may be altered in malignant ovarian epithelial cells.

Adult↗

PubMed2004

Regulation of pregnancy-associated plasma protein-A expression in cultured human osteoblasts.

Pregnancy-associated plasma protein-A (PAPP-A) is a metalloproteinase secreted by cultured human osteoblasts that has been implicated in the regulation of local insulin-like growth factor (IGF) bioavailability during bone growth and remodeling. However, very little is known about the regulation of PAPP-A expression in bone. In this study, we determined the effect of systemic and local osteoregulatory factors on PAPP-A mRNA and protein expression in normal human osteoblasts (hOB cells). Treatment of hOB cells with particular peptide growth factors (basic fibroblast growth factor, epidermal growth factor), steroid hormones (dexamethasone, 1,25-dihydroxyvitamin D(3)), and cytokines [interleukin-6 (IL-6), IL-13, oncostatin M] with known involvement in bone cell physiology had no significant effect on PAPP-A expression. Agents that increase intracellular cyclic AMP (forskolin, prostaglandin E(2)) increased PAPP-A mRNA and protein expression approximately 3-fold. Tumor necrosis factor alpha (TNFalpha), IL-1beta, and IL-4 also increased PAPP-A expression 3- to 4-fold. Transforming growth factor beta (TGFbeta) was previously shown to stimulate PAPP-A expression in hOB cells. The effects of TGFbeta, TNFalpha, and IL-1beta were additive, whereas the effects of TGFbeta and IL-4 were synergistic. In summary, TNFalpha, IL-1beta, and IL-4 were identified as potent stimulators of PAPP-A expression in primary cultures of human osteoblasts. These findings suggest a mechanism whereby cytokines present in bone and bone marrow could augment IGF bioavailability during skeletal growth and remodeling.

Bone Remodeling↗

PubMed2004

Estrogen receptor-positive human epithelial ovarian carcinoma cells respond to the antitumor drug suramin with increased proliferation: possible insight into ER and epidermal growth factor signaling interactions in ovarian cancer.

OBJECTIVES: Development of targeted therapeutics for ovarian cancer requires a basic understanding of ovarian epithelial carcinoma cell biology. The role of estrogen and epidermal growth factor (EGF) in control of cell growth was investigated in a panel of ovarian carcinoma lines. METHODS: EGF receptor (EGFR) was detected by flow cytometry and estrogen receptor (ER) by Northern blot. Western blotting and [(3)H]thymidine incorporation were used to determine receptor activation and the effects of ligand exposure and growth factor antagonists, including the antineoplastic agent, suramin, on cell growth. RESULTS: Only one cell line, OV266, expressed ER and responded to beta-estradiol with increases in DNA synthesis and cell proliferation that could be blocked by the pure antiestrogen ICI 182,780. All cell lines possessed functional EGFR as measured by flow cytometry and phosphorylation of the receptor and mitogen-activated protein kinase after EGF treatment, but only two cell lines (OV177 and OV266) proliferated in response to exogenous EGF. Suramin had limited effectiveness in inhibiting growth in four of five cell lines and had a striking dose-dependent stimulatory effect on OV266 cell growth. The proliferative response to suramin could be inhibited with EGFR antagonists. CONCLUSION: Cultured epithelial ovarian carcinoma cell lines express EGFR (5/5) and can express ER (1/5). Differential growth responses to EGF were observed despite uniform receptor and MAPK activation. Unexpectedly, the antineoplastic agent suramin increased growth of ER positive ovarian carcinoma cells in an EGFR-dependent manner. These studies provide insight into the complex interactions of these systems in control of ovarian cancer cell growth.

Antineoplastic Agents↗

PubMed2004

Insulin and messenger ribonucleic acid expression of insulin receptor isoforms in ovarian follicles from nonhirsute ovulatory women and polycystic ovary syndrome patients.

Insulin action is mediated by two insulin receptor (IR) isoforms, differing in mitogenic and metabolic function. IR isoform expression might occur in human granulosa cells and could be altered in polycystic ovary syndrome (PCOS) from hyperinsulinemia. To determine the relationship between granulosa cell IR isoform expression and follicular fluid insulin concentration in individual follicles, 18 normal women and seven PCOS patients receiving gonadotropins for in vitro fertilization were studied. Glucose tolerance testing was performed before pituitary desensitization, and fasting serum insulin was measured at oocyte retrieval. Granulosa cells and fluid aspirated from the first follicle were used to determine IR isoform mRNA expression and insulin concentration, respectively. IR isoform A mRNA expression was greater than that of IR isoform B expression in normal mural granulosa and cumulus cells, without a cell type effect. Intrafollicular insulin levels increased with adiposity and serum insulin levels at oocyte-retrieval but did not predict IR mRNA expression. Total IR mRNA expression, but not intrafollicular insulin levels, was elevated in PCOS patients, whereas intrafollicular insulin levels were increased in women with impaired glucose tolerance. Granulosa cell IR heterogeneity, together with adiposity-dependent intrafollicular insulin availability, introduces a novel mechanism by which insulin may affect granulosa cell function within the follicle.

Adult↗

PubMed2004

Metalloproteinase pregnancy-associated plasma protein A is a critical growth regulatory factor during fetal development.

Pregnancy-associated plasma protein A (PAPPA) is a metzincin superfamily metalloproteinase in the insulin-like growth factor (IGF) system. PAPPA increases IGF bioavailability and mitogenic effectiveness in vitro through regulated cleavage of IGF-binding protein 4 (IGFBP4). To determine its function in vivo, we generated PAPPA-null mice by gene targeting. Mice homozygous for targeted disruption of the PAPPA gene were viable but 60% the size of wild-type littermates at birth. The impact of the mutation was exerted during the early embryonic period prior to organogenesis, resulting in proportional dwarfism. PAPPA, IGF2 and IGFBP4 transcripts co-localized in wild-type embryos, and expression of IGF2 and IGFBP4 mRNA was not altered in PAPPA-deficient embryos. However, IGFBP4 proteolytic activity was completely lacking in fibroblasts derived from PAPPA-deficient embryos, and IGFBP4 effectively inhibited IGF-stimulated mitogenesis in these cells. These results provide the first direct evidence that PAPPA is an essential growth regulatory factor in vivo, and suggest a novel mechanism for regulated IGF bioavailability during early fetal development.

Animals↗

PubMed2003

Pregnancy-associated plasma protein a gene expression as a target of inflammatory cytokines.

Pregnancy-associated plasma protein A (PAPP-A) cleaves IGF-binding protein-4 (IGFBP-4) and appears to enhance local IGF bioavailability in response to injury. In this study we determined the effects of growth factors and cytokines involved in the healing process on PAPP-A expression in human dermal fibroblasts. There was no effect of platelet-derived growth factor, epidermal growth factor, or basic fibroblast growth factor on PAPP-A mRNA expression in these cells. However, treatment with the proinflammatory cytokines, TNFalpha and IL-1 beta, resulted in time- and dose-dependent increases in PAPP-A mRNA and protein expression (3- to 4-fold maximal effects), which were prevented by actinomycin D. On the other hand, interferon-gamma (IFN gamma) treatment markedly inhibited PAPP-A expression. IGFBP-4 proteolytic activity was increased 4-fold in medium from TNFalpha- and IL-1 beta-treated (1 nm) cells and decreased 40% in medium from IFN gamma-treated (1 nm) cells. IGF-I-stimulated [(3)H]thymidine incorporation was significantly enhanced by pretreatment with 1 nm TNFalpha, and this enhancement was blocked in the presence of protease-resistant IGFBP-4. In conclusion, PAPP-A expression is regulated by inflammatory cytokines in adult human fibroblasts, with functional consequences on IGFBP-4 protease activity and IGF-I bioavailability. These data provide a mechanism for the regulation of PAPP-A in response to injury and further implicate PAPP-A in the wound-healing processes.

Adult↗