Mononuclear cells modulate the activity of pancreatic stellate cells which in turn promote fibrosis and inflammation in chronic pancreatitis

Background: Interactions between mononuclear cells and activated pancreatic myofibroblasts (pancreatic stellate cells; PSC) may contribute to inflammation and fibrosis in chronic pancreatitis (CP). Methods: Markers of fibrosis and inflammation were concomitantly analysed by immunohistochemistry in chronic pancreatitis tissues. In vitro, PSC were stimulated with TNFalpha and LPS. Primary human blood mononuclear cells (PBMC) and PSC were cocultured, followed by analysis of cytokines and extracellular matrix (ECM) proteins. PBMC were derived from healthy donors and CP and septic shock patients. Results: In areas of mononuclear cell infiltration in chronic pancreatitis tissues, there was decreased immunoreactivity for collagen1 and fibronectin, in contrast to areas with sparse mononuclear cells, although PSC were detectable in both areas. LPS and TNFalpha induced collagen1 and fibronectin levels as well as the matrix degradation enzyme MMP-1. Coculture experiments with PSC and PBMC revealed increased fibronectin secretion induced by PBMC. In addition, donor and CP PBMC significantly induced an increase in IL-6, MCP-1 and TGFbeta levels under coculture conditions. Determination of the source of cytokines and ECM proteins by mRNA expression analysis confirmed PSC as major contributors of ECM production. The increase in cytokine expression was PBMC- and also PSC-derived. Conclusion: Mononuclear cells modulate the activity of pancreatic stellate cells, which may in turn promote fibrosis and inflammation

Inflammation-Associated Nitrotyrosination Affects TCR Recognition through Reduced Stability and Alteration of the Molecular Surface of the MHC Complex

Nitrotyrosination of proteins, a hallmark of inflammation, may result in the production of MHC-restricted neoantigens that can be recognized by T cells and bypass the constraints of immunological self-tolerance. Here we biochemically and structurally assessed how nitrotyrosination of the lymphocytic choriomeningitis virus (LCMV)-associated immunodominant MHC class I-restricted epitopes gp33 and gp34 alters T cell recognition in the context of both H-2Db and H-2Kb. Comparative analysis of the crystal structures of H-2Kb/gp34 and H-2Kb/NY-gp34 demonstrated that nitrotyrosination of p3Y in gp34 abrogates a hydrogen bond interaction formed with the H-2Kb residue E152. As a consequence the conformation of the TCR-interacting E152 was profoundly altered in H-2Kb/NY-gp34 when compared to H-2Kb/gp34, thereby modifying the surface of the nitrotyrosinated MHC complex. Furthermore, nitrotyrosination of gp34 resulted in structural over-packing, straining the overall conformation and considerably reducing the stability of the H-2Kb/NY-gp34 MHC complex when compared to H-2Kb/gp34. Our structural analysis also indicates that nitrotyrosination of the main TCR-interacting residue p4Y in gp33 abrogates recognition of H-2Db/gp33-NY complexes by H-2Db/gp33-specific T cells through sterical hindrance. In conclusion, this study provides the first structural and biochemical evidence for how MHC class I-restricted nitrotyrosinated neoantigens may enable viral escape and break immune tolerance

The role of nitric oxide in the emotional learning of rats in the plus-maze

The present study evaluated the role of nitric oxide (NO) in the transfer latency (TL) paradigm in the elevated plus-maze. Male Wistar rats received i.p. injections of either 0.9% Saline, N-w Nitro-L-arginine-methyl-ester (L-NAME, an inhibitor of NO synthesis), D-NAME (inert isomer), scopolamine (SCO, antagonist of muscarinic receptors), or MK-801 (antagonist of NMDA receptors) and, after 30 min, were submitted to TL procedure. In an independent experiment, the ability of the same L-NAME treatments in changing the arterial pressure and blood glucose level (BGL) was evaluated in conscious rats. The treatment with SCO (1 mg kg(-1)), MK-801 (0.15 mg kg(-1)) and L-NAME (10 and 50 mg kg(-1)), but not with D-NAME, impaired the TL learning. The L-NAME-induced TL deficit was counteracted by L-ARG (100 and 200 mg kg(-1)), while the co-administration of sub-effective doses of L-NAME and MK-801 failed to impair the TL learning. The L-NAME (50 mg kg(-1)) treatment failed to alter the BGL. All treatments with L-NAME induced hypertension, but the rats treated with L-NAME (5 mg kg(-1)) were still able to learn the TL task. The data indicate that the TL deficit induced by L-NAME (10 and 50 mg kg(-1)) is not due to either hypertension or changes in the BGL. It is also possible to establish that NO production is important for emotional learning underlying the TL procedure in rats. (c) 2004 Elsevier Inc. All rights reserved.84335135