Nanoparticles, including lipopolyamines leading to lipoplexes, liposomes, and polyplexes are targeted drug carrier systems in the current search for a successful delivery system for polynucleic acids. This review is focused on the impact of gene and siRNA delivery for studies of efficacy, pharmacodynamics, and pharmacokinetics within the setting of the wide variety of in vivo animal models now used. This critical appraisal of the recent literature sets out the different models that are currently being investigated to bridge from studies in cell lines through towards clinical reality. Whilst many scientists will be familiar with rodent (murine, fecine, cricetine, and musteline) models, few probably think of fish as a clinically relevant animal model, but zebrafish, madake, and rainbow trout are all being used. Larger animal models include rabbit, cat, dog, and cow. Pig is used both for the prevention of foot-and-mouth disease and human diseases, sheep is a model for corneal transplantation, and the horse naturally develops arthritis. Non-human primate models (macaque, common marmoset, owl monkey) are used for preclinical gene vector safety and efficacy trials to bridge the gap prior to clinical studies. We aim for the safe development of clinically effective delivery systems for DNA and RNAi technologies.

The persistent overexpression of PTPN22 and the transient reduction of CREB activity, associated with impaired Treg differentiation, might play a role in ACS.

Tissue factor (TF), the key activator of the blood coagulation cascade and of thrombus formation, is also expressed by circulating human platelets. Despite the documented in-depth characterization of platelet TF carried out in the past 15 years, some authors still fail to identify TF in platelets, especially when assessment in platelet-rich plasma (PRP) or washed platelets is carried out. This study aims to extend the characterization of the subset of TF-positive platelets in PRP from healthy subjects and to verify how different centrifugation forces, used to prepare the PRP, could affect the analysis of TF-positive platelets. Data indicate that large-size platelets express significantly higher amount of TF compared to small-size cells, in terms of both TF protein and TF mRNA. Upon stimulation, large platelets readily expose on the cell membrane TF, which is functionally active, i.e., able to generate factor Xa (FXa) as well as thrombin. By contrast, TF activity in small platelets is almost completely quenched by tissue factor pathway inhibitor (TFPI), becoming indeed detectable only after treatment with an anti-TFPI antibody. Our data highlight that particular attention must be paid to the preparation and collection of the PRP since such preanalytical variables may influence the platelet recovery and in turn affect subsequent analysis, whether it is flow cytometry, functional activity tests, proteome, or transcriptome analysis. Indeed, the TF-positive subset of large platelets can easily be lost if centrifugation protocols are not optimized, thus erroneously leading to a false-negative result.

In acute coronary syndrome (ACS), T cell abnormalities are associated to a worse outcome. Loss of inhibitory activity of CD31, an Ig-like adhesion molecule, on peripheral leukocytes has been found to enhance atherosclerosis in experimental models. In this study, we examined the expression of CD31 on T cells, and its role on TCR signaling in 35 patients with non-ST elevation ACS, in 35 patients with stable angina (SA), and in 35 controls. Furthermore, 10 ACS and 10 SA patients were re-analyzed at 1-year follow-up. Flow-cytometry analysis showed that in ACS patients, CD31 expression was reduced on total CD4(+) and CD4(+)CD28(null) (P < 0.001, ACS vs. SA), on naïve (P < 0.001, ACS vs. SA) and on central-memory and effector-memory CD4(+) T cells (P < 0.05, ACS vs. SA and controls). The immunomodulatory effect of CD31 on TCR signaling of CD4(+) and CD4(+)CD28(null) T cells, was lower in ACS than SA patients (P < 0.05, for both comparisons). At 1-year follow-up, CD31 expression and function increased in ACS becoming similar to that found in SA. CD31 recruitment in the immunological synapse was lower in ACS than controls (P = 0.012). Moreover, CD31 modulated MAPK signaling and reduced the expression of T bet and Rorγ-t, necessary for Th1 and Th17 differentiation. Finally, we studied TCR signaling in CD31(+) naïve and primed T cell subsets observing a different pattern of protein phosphorylation. A CD31-mediated regulatory pathway is enhanced in SA and temporarily downregulated in ACS. As CD31 modulates both T cell activation, by increasing the threshold for TCR stimulation, and T cell differentiation, it might represent a novel molecular target to treat T cell abnormalities in ACS.

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