Rikke Hjortebjerg

Objective: Anabolic androgenic steroids (AAS) may exert effects through the insulin-like growth factor (IGF) system. We compared the IGF system in male and female recreational athletes who were current or former AAS users and non-users. Methods: The cross-sectional study included 80 active AAS users (19 women), 26 former users (8 women), and 58 age- and sex-matched non-users (16 women). Participants underwent clinical examination, echocardiography, and measurement of circulating IGF-1, IGF-2, IGF bioactivity, IGF-binding proteins (IGFBPs), pregnancy-associated plasma protein-A (PAPP-A) and -A2 (PAPP-A2), and stanniocalcin-1 (STC1) and −2 (STC2). Results: In active AAS users, IGF-2 levels were reduced by 36.1% vs. former users and 38.4% vs. non-users (P < .001), and IGFBP-3 levels were 13.6% and 35.8% lower (P < .001). STC2 was increased in AAS-using men only, being 24.5% higher than former users (P = .029) and 45.7% higher than non-users (P < .001). PAPP-A was higher in non-using men than women, whereas levels were similar in active users. Differences remained after adjustment for age, sex, body fat, exercise and strength training, family history of coronary disease, smoking, alcohol, and drug use. No group differences were observed for IGF-1, bioactive IGF, IGFBP-2, or PAPP-A2. Cumulative lifetime AAS exposure was associated with IGF-2, IGFBP-3, and STC2 in multivariable models. Higher IGF-2 and IGFBP-3 associated with lower left ventricular mass, higher left ventricular ejection fraction, and better diastolic function, whereas higher STC2 associated with adverse outcomes (all P < .05). Conclusions: Illicit AAS use is associated with persistent, sex-specific alterations in the IGF–PAPP-A–STC2 axis, potentially contributing to cardiovascular risk in recreational athletes.

Pregnancy-associated plasma protein-A (PAPP-A) and PAPP-A2 modulate insulin-like growth factor (IGF) action and are inhibited by the stanniocalcins (STC1 and STC2). We previously demonstrated increased PAPP-A and IGF activity in ascites from women with ovarian carcinomas. In this prospective, longitudinal study of 107 women with ovarian cancer and ascites accumulation, we determined corresponding serum and ascites levels of IGF-1, IGF-2, PAPP-A, PAPP-A2, STC1, and STC2 and assessed their relationship with mortality. As compared to serum, we found highly increased ascites levels of PAPP-A (51-fold) and PAPP-A2 (4-fold). Elevated levels were also observed for IGF-1 (12%), STC1 (90%) and STC2 (68%). In contrast, IGF-2 was reduced by 29% in ascites. Patients were followed for a median of 38.4 months (range: 45 days to 8.9 years), during which 73 patients (68.2%) died. Overall survival was longer for patients with high serum IGF-1 (hazard ratio (HR) per doubling in protein concentration: 0.60, 95% CI: 0.40–0.90). However, patients with high ascites levels of IGF-1 showed a poorer prognosis (HR: 2.00 (1.26–3.27)). High serum and ascites IGF-2 levels were associated with increased risk of mortality (HR: 2.01 (1.22–3.30) and HR: 1.78 (1.24–2.54), respectively). Similarly, serum PAPP-A2 was associated with mortality (HR: 1.26 (1.08–1.48)). Our findings demonstrate the presence and activity of the IGF system in the local tumor ecosystem, which is likely a characteristic feature of malignant disease and plays a role in its peritoneal dissemination. The potential clinical implications are supported by our finding that serum levels of the proteins are associated with patient prognosis.

Pregnancy-associated plasma protein-A (PAPP-A) and its homolog PAPP-A2 are enzymes that modulate the availability and mitogenic activity of insulin-like growth factor-I (IGF-I). PAPP-A has been implicated in numerous cancers but reports on PAPP-A2 in malignancy are non-existent. In a prospective observational study of 689 patients under suspicion of lung cancer, we examined levels of PAPP-A and PAPP-A2 and their relationship with mortality. Serum PAPP-A and PAPP-A2 concentrations were determined in pre-diagnostic blood samples using ELISA, and immunohistochemical staining of PAPP-A and PAPP-A2 was performed in malignant tissue from five operable patients. A total of 144 patients were diagnosed with lung cancer, whereas the diagnosis was rejected in 545 subjects, who served as a control group. PAPP-A2 concentrations were higher in patients with lung cancer [median (IQR): 0.33 (0.21–0.56) ng/mL] than in controls [0.27 (0.17–0.39) ng/mL], p < 0.001, whereas PAPP-A levels did not differ. Presence of PAPP-A and PAPP-A2 were confirmed in tumor specimens, and staining occurred in a heterogeneous pattern. Patients were observed for a median (range) of 7 (6; 8) years, during which 114 patients (79.2%) died. Patient mortality differed according to PAPP-A2 tertile (p < 0.001). PAPP-A2 was associated with mortality with an unadjusted hazard ratio (95% CI) per doubling in protein concentration of 1.30 (1.12; 1.53), p = 0.001. In a multivariable model adjusted for age, sex, and BMI, PAPP-A2 remained predictive of the endpoint with a hazard ratio per doubling in protein concentration of 1.25 (1.05; 1.48), p = 0.013. Collectively, PAPP-A2, but not PAPP-A, is elevated in patients with lung cancer and associated with mortality. This novel role of PAPP-A2 in cancer warrants further functional studies as well as validation in external cohorts.

Objective: Pregnancy-associated plasma protein-A (PAPP-A) has been suggested as a proatherogenic enzyme by its ability to locally increase insulin-like growth factor (IGF) activity through proteolytic cleavage of IGF binding protein-4 (IGFBP-4). Recently, stanniocalcin-2 (STC2) was discovered as an inhibitor of PAPP-A. This study aimed to investigate IGFBP-4, PAPP-A, and STC2 as local regulators of IGF bioactivity in the cardiac microenvironment by comparing levels in the pericardial fluid with those in the circulation of patients with cardiovascular disease. Methods: Plasma and pericardial fluid were obtained from 39 patients undergoing elective cardiothoracic surgery, hereof 15 patients with type 2 diabetes. Concentrations of IGF-I, intact and fragmented IGFBP-4, PAPP-A, and STC2 were determined by immunoassays and IGF bioactivity by a cell-based assay. Results: In pericardial fluid, the concentrations of total IGF-I, intact IGFBP-4, and STC2 were 72 ± 10%, 91 ± 5%, and 40 ± 24% lower than in plasma, while PAPP-A was 15 times more concentrated. The levels of the 2 IGFBP-4 fragments generated by PAPP-A and reflecting PAPP-A activity were elevated by more than 25%. IGF bioactivity was 62 ± 81% higher in the pericardial fluid than plasma. Moreover, pericardial fluid levels of both IGFBP-4 fragments correlated with the concentration of PAPP-A and with the bioactivity of IGF. All protein levels were similar in pericardial fluid from nondiabetic and diabetic subjects. Conclusions: PAPP-A increases IGF bioactivity by cleavage of IGFBP-4 in the pericardial cavity of cardiovascular disease patients. This study provides evidence for a distinct local activity of the IGF system, which may promote cardiac dysfunction and coronary atherosclerosis.

Patients with inflammatory bowel disease (IBD) present with reduced serum insulin-like growth factor I (IGF-I). Anti-inflammatory treatment with prednisolone or infliximab ameliorates symptoms and increases circulating IGF-I, but prednisolone induces catabolism, whereas infliximab may promote protein synthesis. Recently, stanniocalcin-2 (STC2) was discovered as a novel inhibitor of the enzyme pregnancy-associated plasma protein-A (PAPP-A), which modulates IGF-I activity. PAPP-A can cleave IGF binding protein-4 (IGFBP-4), upon which IGF-I is liberated. We hypothesized that prednisolone and infliximab exert different effects on levels of STC2, PAPP-A, and IGFBP-4, thereby explaining the distinct metabolic effects of prednisolone and infliximab. Thirty-eight patients with active IBD treated with either prednisolone (n = 17) or infliximab (n = 21) were examined before and after 7 days of treatment. Circulating levels of IGF-I, IGF-II, IGFBP-3, PAPP-A, and STC2 were measured by immunoassays. Intact IGFBP-4 and two IGFBP-4 fragments were determined by a novel immunoassay. Bioactive IGF was assessed by cell-based IGF receptor activation assay. Concentrations of IGFBP-4, PAPP-A, and STC2 on day 0 and 7 were compared to healthy control subjects. Following seven days of prednisolone treatment, total and bioactive IGF-I were increased (p < 0.001 and p < 0.05, respectively). Upon infliximab treatment, total IGF-I levels were augmented (p < 0.05), yet IGF bioactivity remained unaltered. Intact IGFBP-4 and the two IGFBP-4 fragments generated upon cleavage by PAPP-A were all decreased following treatment with either prednisolone or infliximab (all p < 0.05). PAPP-A levels were only increased by infliximab (p = 0.005), whereas the inhibitor STC2 did not respond to any of the treatments. IGF-I and IGFBP-4 concentrations were markedly altered in patients with IBD and near-normalized with disease remission following treatment with prednisolone or infliximab. Thus, IGFBP-4 may modulate IGF bioavailability in IBD. The effect of immunosuppression did not appear to extend beyond the regulation of IGF and IGFBP-4, as neither PAPP-A nor STC2 were discernibly affected. ClinicalTrials.gov: NCT00955123 . Date of registration: August 7, 2009 (retrospectively registered).