Analysis of pregnancy-associated plasma protein a production in human adult cardiac progenitor cells

IGF-binding proteins (IGFBPs) and their proteases regulate IGFs bioavailability in multiple tissues. Pregnancy-associated plasma protein A (PAPP-A) is a protease acting by cleaving IGFBP2, 4, and 5, regulating local bioavailability of IGFs. We have previously shown that IGFs and IGFBPs are produced by human adult cardiac progenitor cells (haCPCs) and that IGF-1 exerts paracrine therapeutic effects in cardiac cell therapy with CPCs. Using immunofluorescence and enzyme immunoassays, we firstly report that PAPP-A is produced and secreted in surprisingly high amounts by haCPCs. In particular, the homodimeric, enzymatically active, PAPP-A is secreted in relevant concentrations in haCPC-conditioned media, while the enzymatically inactive PAPPA/proMBP complex is not detectable in the same media. Furthermore, we show that both homodimeric PAPP-A and proMBP can be detected as cell associated, suggesting that the previously described complex formation at the cell surface does not occur easily, thus positively affecting IGF signalling. Therefore, our results strongly support the importance of PAPP-A for the IGFs/IGFBPs/PAPP-A axis in CPCs biology

Toxicity assessment of (99m)technetium-labeled human beta-defensin-3 in CD1 mice

Objective: Human beta-defensin-3 (HBD-3) is an antimicrobial peptide which is up-regulated during 99m inflammation. Based on the previously demonstrated capacity of technetium-99m ( Tc) labelled HBD-3 of distinguishing infection from inflammation in rats, we have decided to collect information on the potential toxicity of the tracer in view of its possible use for imaging in humans. Materials and Methods: 99m Recombinant HBD-3 underwent labeling with Tc. The CD1 mice were selected as standard rodent species. Ten mice, 5 male and 5 female, were subjected to physical examination and housed in a dedicated room in 5 per cage. After 9 days pre-test period, all mice were weighted for dose adjustment and received 99m intravenously 6mcg/mouse of Tc-HBD-3. Mortality was recorded daily, while body weight was registered once a week. Clinical observation of animals was performed daily for sickness symptoms due to 99m the drug treatment. At day 19 a second dose of 6mcg/mouse Tc-HBD-3, was administered. Twenty-four hours after the second dose (day 20) the animals were euthanized. A piece of liver, kidneys, heart and lungs was collected for histopathological analysis. Results:Our results showed that the labelled-HBD-3 dose did not induce significant toxicity in mice. Of course these parameters were not sufficient to authorize use in humans. This non-toxic dose of HBD-3 when translated from animals to humans resulted in an equivalent dose of approximately 25 times higher than that needed for imaging. Conclusion:Our non toxicity data of 99m using Tc-beta-defensin-3 in mice offer a further indication in favour of the clinical use of this radiopharmaceutical in all cases where discrimination between infection and inflammation is needed

Labelling of Granulocytes by Phagocytic Engulfment with (64)Cu-Labelled Chitosan-Coated Magnetic Nanoparticles.

Purpose: The aim of the present work was to perform the labelling of granulocytes by theirengulfment with chitosan-coated magnetic 64Cu nanoparticles (MNPs) in order to obtain aradiopharmaceutical suitable for dual imaging (PET–MRI) of inflammatory/infective diseases.Procedures: Specimens of 5–20 mg MNPs were washed with saline–isotonic solution andrecuperated by magnetic decantation; assessment of the release of 64Cufrom labelled granulocytes in plasma was performed by measuring the radioactivity of both the cellularpellet and the supernatant solution.Results: Our data showed the binding capacity of chitosan-coated MNPs for cationic metal. Theamount of Cu+2 chelated captured per milligram of MNPs was constant and independent of thereagent concentrations. In all cases, more than 90% of the engulfed granulocytes were positiveto the TBE test. The MNPs were localised within the cells.Conclusion: In our in vitro model, MNPs are taken up by granulocytes through phagocytosis,whereas previously described methods were based on the use of a chelating agent that permitCu to cross the cell membrane. Moreover, the 64Cu-engulfed granulocytes showed a highstability of up to 80% of retained radioactivity after 24 h of incubation.; 10 mg MNPs wasallowed to reactwith 16 μg 64Cu [64Ni(p,n) at 12–9 MeV] followed by anion exchange chromatographywith a specific activity of 56 MBq/μg. Pellets of granulocytes were obtained from peripheral blood;MNPs engulfment by granulocytes was obtained and granulocyte-engulfed viability was assessed bythe trypan blue exclusion (TBE) test performed at 5 min, 2 h and 4 h; 15–58 μg Cu2+ (CuCl2·H2O) in 50 μl of acidified (pH 5.5)saline solution was added to the MNPs re-suspended saline–isotonic solutio

Effect of External Magnetic Field on IV 99mTc-Labeled Aminosilane-Coated Iron Oxide Nanoparticles: Demonstration in a Rat Model: Special Report

ABSTRACT: Among the most interesting applications of ferromagnetic nanoparticles (NPs) in medicine is the potential for localizing pharmacologically or radioactively tagged agents directly to selected tissues selected by an adjustable external magnetic field. This concept is demonstrated by the application external magnetic field on IV Tc-labeled aminosilane-coated iron oxide NPs in a rat model. In a model comparing a rat with a 0.3-T magnet over a hind paw versus a rat without a magnet, a static acquisition at 45 minutes showed that 27% of the administered radioactivity was in the area subtended by the magnet, whereas the liver displays a percentage of binding of 14% in the presence of the magnet and of 16% in the absence of an external magnetic field. These preliminary results suggest that the application of an external magnetic field may be a viable route for the development of methods for the confinement of magnetic NPs labeled with radioactive isotopes targeted for predetermined sites of the body

Performance Metrics of the Scoring System for the Diagnosis of the Beckwith-Wiedemann Spectrum (BWSp) and Its Correlation with Cancer Development

Different scoring systems for the clinical diagnosis of the Beckwith-Wiedemann spectrum (BWSp) have been developed over time, the most recent being the international consensus score. Here we try to validate and provide data on the performance metrics of these scoring systems of the 2018 international consensus and the previous ones, relating them to BWSp features, molecular tests, and the probability of cancer development in a cohort of 831 patients. The consensus scoring system had the best performance (sensitivity 0.85 and specificity 0.43). In our cohort, the diagnostic yield of tests on blood-extracted DNA was low in patients with a low consensus score (similar to 20% with a score = 2), and the score did not correlate with cancer development. We observed hepatoblastoma (HB) in 4.3% of patients with UPD(11)pat and Wilms tumor in 1.9% of patients with isolated lateralized overgrowth (ILO). We validated the efficacy of the currently used consensus score for BWSp clinical diagnosis. Based on our observation, a first-tier analysis of tissue-extracted DNA in patients with <4 points may be considered. We discourage the use of the consensus score value as an indicator of the probability of cancer development. Moreover, we suggest considering cancer screening for negative patients with ILO (risk similar to 2%) and HB screening for patients with UPD(11)pat (risk similar to 4%)

Importance of Baseline Prognostic Factors With Increasing Time Since Initiation of Highly Active Antiretroviral Therapy: Collaborative Analysis of Cohorts of HIV-1-Infected Patients

Background: The extent to which the prognosis for AIDS and death of patients initiating highly active antiretroviral therapy (HAART) continues to be affected by their characteristics at the time of initiation (baseline) is unclear. Methods: We analyzed data on 20,379 treatment-naive HIV-1- infected adults who started HAART in 1 of 12 cohort studies in Europe and North America (61,798 person-years of follow-up, 1844 AIDS events, and 1005 deaths). Results: Although baseline CD4 cell count became less prognostic with time, individuals with a baseline CD4 count 350 cells/μL (hazard ratio for AIDS = 2.3, 95% confidence interval [CI]: 1.0 to 2.3; mortality hazard ratio = 2.5, 95% CI: 1.2 to 5.5, 4 to 6 years after starting HAART). Rates of AIDS were persistently higher in individuals who had experienced an AIDS event before starting HAART. Individuals with presumed transmission by means of injection drug use experienced substantially higher rates of AIDS and death than other individuals throughout follow-up (AIDS hazard ratio = 1.6, 95% CI: 0.8 to 3.0; mortality hazard ratio = 3.5, 95% CI: 2.2 to 5.5, 4 to 6 years after starting HAART). Conclusions: Compared with other patient groups, injection drug users and patients with advanced immunodeficiency at baseline experience substantially increased rates of AIDS and death up to 6 years after starting HAART