Local delivery of IL-2 reduces atherosclerosis via expansion of regulatory T cells.

OBJECTIVE: Recent studies indicate that regulatory T cells (Tregs) attenuate murine atherosclerosis. Since interleukin (IL)-2 induces Tregs proliferation, we tested the impact of L19-IL2, a fusion antibody specific to extra-domain B of fibronectin (ED-B) containing an active human IL-2 molecule, in experimental atherosclerosis. METHODS AND RESULTS: L19-IL2 or appropriate controls were given intravenously to 6 month old Western diet-fed apoE(-/-) mice on day 1, 3, and 5. Human IL-2 was detected on day 7 within atherosclerotic plaques of L19-IL2-treated mice, and magnetic resonance imaging of the plaques showed a significant adventitial gadolinium enhancement on day 7 and 13, suggesting microvascular leakage as a result of the pharmacodynamic activity of L19-IL2. Treatment with L19-IL2 significantly reduced the size of pre-established atherosclerotic plaques at the thoracic aorta (Sudan III stained area) and in the aortic root area (microscopic, morphometric analysis) on day 7 as compared to controls (L19, D1.3-IL2, NaCl) as well as compared to baseline (day 0). Tregs markers Foxp3 and CTLA4 were highly increased in plaques after L19-IL2 treatment compared to controls (p<0.01), whereas the macrophage marker Mac3 was significantly reduced (p<0.03). Co-treatment with IL-2-receptor blocking antibody PC61 abrogated L19-IL2-induced plaque reduction compared with IgG control (p<0.03). CONCLUSION: L19-IL2 delivers functional IL-2 to pre-established atherosclerotic plaques of WD-fed apoE(-/-) mice resulting in significant plaque size reduction mediated by local Tregs.

Transient receptor potential channel TRPM8 agonists stimulate calcium influx and neurotensin secretion in neuroendocrine tumor cells.

TRPM8 is a member of the melastatin-type transient receptor potential ion channel family. Activation by cold or by agonists (menthol, icilin) induces a transient rise in intracellular free calcium concentration ([Ca(2+)](i)). Our previous study demonstrated that Ca(2+)-permeable cation channels play a role in IGF-1-induced secretion of chromogranin A in human neuroendocrine tumor (NET) cell line BON [Mergler et al.: Neuroendocrinology 2006;82:87-102]. Here, we extend our earlier study by investigating the expression of TRPM8 and characterizing its impact on [Ca(2+)](i) and the secretion of neurotensin (NT). We identified TRPM8 expression in NET BON cells by RT-PCR, Western blotting and immunofluorescence staining. Icilin increased [Ca(2+)](i) in TRPM8-transfected human embryonic kidney cells (HEK293) but not in mock-transfected cells. Icilin and menthol induced Ca(2+) transients in BON cells as well as in primary NET cell cultures of two different pancreatic NETs as detected by single cell fluorescence imaging. Icilin increased non-selective cation channel currents in BON cells as detected by patch-clamp recordings. This activation was associated with increased NT secretion. Taken together, this study demonstrates for the first time the expression TRPM8 in NET cells and its role in regulating [Ca(2+)](i) and NT secretion. The regulation of NT secretion in NETs by TRPM8 may have a potential clinical implication in diagnosis or therapy.