The human inducible nitric oxide synthase (iNOS) gene is overexpressed in a number of human inflammatory diseases. Previously, we observed that the human iNOS gene is transcriptionally regulated by cytokines and demonstrated that the cytokine-responsive regions are upstream of ؊3.8 kilobase pairs (kb). Therefore, the purpose of this study was to further localize the functional enhancer elements and to assess the role of the transcription factor NF-B in both human liver (AKN-1) and human lung (A549) epithelial cell lines. The addition of NF-B inhibitors significantly suppressed cytokinestimulated iNOS mRNA expression and NO synthesis, indicating that NF-B is involved in the induction of the human iNOS gene. Analysis of the first 4.7 kb of the 5-flanking region demonstrated basal promoter activity and failed to show any cytokine-inducible activity. However, promoter constructs extending to ؊5.8 and ؊7.2 kb revealed 2-3-fold and 4 -5-fold induction, respectively, in the presence of cytokines. DNA sequence analysis from ؊3.8 to ؊7.2 kb identified five putative NF-B cis-regulatory transcription factor binding sites upstream of ؊4.7 kb. Site-directed mutagenesis of these sites revealed that the NF-B motif at ؊5.8 kb is required for cytokine-induced promoter activity, while the sites at ؊5.2, ؊5.5, and ؊6.1 kb elicit a cooperative effect. Electromobility shift assays using a site-specific oligonucleotide and nuclear extracts from cells stimulated with cytokine-mixture, tumor necrosis factor-␣ or interleukin-1␤, but not interferon-␥, exhibited inducible DNA binding activity for NF-B. These data indicate that NF-B activation is required for cytokine induction of the human iNOS gene and identifies four NF-B enhancer elements upstream in the human iNOS promoter that confer inducibility to tumor necrosis factor-␣ and interleukin-1␤.

The human inducible nitric oxide synthase (hiNOS) gene is expressed in several disease states and is also important in the normal immune response. Previously, we described a cytokine-responsive enhancer between ؊5.2 and ؊6.1 kb in the 5-flanking hiNOS promoter DNA, which contains multiple nuclear factor ␤ (NF-B) elements. Here, we describe the role of the IFN-Jak kinase-Stat (

Rationale Cardiac progenitor cells are an attractive cell type for tissue regeneration but their mechanism for myocardial remodeling is still unclear. Objective This investigation determines how chronological age influences the phenotypic characteristics and the secretome of human cardiac progenitor cells (CPCs), as well as their potential to recover injured myocardium. Methods and Results Adult (aCPCs) and neonatal (nCPCs) cells were derived from patients more than 40 years or less than one month of age, respectively, and their functional potential was determined in a rodent myocardial infarction (MI) model. A more robust in vitro proliferative capacity of nCPCs, compared to aCPCs, correlated with significantly greater myocardial recovery mediated by nCPCs in vivo. Strikingly, a single injection of nCPC-derived total conditioned media (nTCM) was significantly more effective than nCPCs, aCPC-derived TCM (aTCM), or nCPC-derived exosomes in recovering cardiac function, stimulating neovascularization, and promoting myocardial remodeling. High resolution accurate mass spectrometry (HRAMS) with reverse phase liquid chromatography fractionation and mass spectrometry (LC-MS/MS) was employed to identify proteins in the secretome of aCPCs and nCPCs, and literature-based networking software identified specific pathways affected by the secretome of CPCs in the setting of MI. Examining the TCM, we quantified changes in the expression pattern of 804 proteins in nTCM and 513 proteins in aTCM. Literature-based proteomic network analysis identified that 46 and 6 canonical signaling pathways were significantly targeted by nTCM and aTCM, respectively. One leading candidate pathway is heat shock factor-1 (HSF-1), potentially affecting 8 identified pathways for nTCM but none for aTCM. To validate this prediction, we demonstrated that modulation of HSF-1 by knockdown in nCPCs or overexpression in aCPCs significantly altered the quality of their secretome. Conclusions In conclusion, a deep proteomic analysis revealed both detailed and global mechanisms underlying the chronological age-based differences in the ability of CPCs to promote myocardial recovery via the components of their secretome.

The Bard Recovery and Bard G2 filters had high prevalences of fracture and embolization, with potentially life-threatening sequelae.

These data indicate that NO down-regulates iNOS gene transcription, and that the effect is mediated in part by inhibiting NF-kappa B activity. These results identify a novel negative feedback mechanism whereby NO down-regulates iNOS gene expression, possibly to limit overproduction during the septic response.