Insulin stimulation of target cells elicits a burst of H 2 O 2 that enhances tyrosine phosphorylation of the insulin receptor and its cellular substrate proteins as well as distal signaling events in the insulin action cascade. The molecular mechanism coupling the insulin receptor with the cellular oxidant-generating apparatus has not been elucidated. Using reverse transcription-PCR and Northern blot analyses, we found that Nox4, a homolog of gp91phox, the phagocytic NAD(P)H oxidase catalytic subunit, is prominently expressed in insulin-sensitive adipose cells. Adenovirus-mediated expression of Nox4 deletion constructs lacking NAD(P)H or FAD/NAD(P)H cofactor binding domains acted in a dominant-negative fashion in differentiated 3T3-L1 adipocytes and attenuated insulin-stimulated H 2 O 2 generation, insulin receptor (IR) and IRS-1 tyrosine phosphorylation, activation of downstream serine kinases, and glucose uptake. Transfection of specific small interfering RNA oligonucleotides reduced Nox4 protein abundance and also inhibited the insulin signaling cascade. Overexpression of Nox4 also significantly reversed the inhibition of insulin-stimulated IR tyrosine phosphorylation induced by coexpression of PTP1B by inhibiting PTP1B catalytic activity. These data suggest that Nox4 provides a novel link between the IR and the generation of cellular reactive oxygen species that enhance insulin signal transduction, at least in part via the oxidative inhibition of cellular protein-tyrosine phosphatases (PTPases), including PTP1B, a PTPase that has been previously implicated in the regulation of insulin action.

Vascular remodeling within the aorta results in a loss of structural integrity with consequent aneurysm formation. This degradation is more common in the abdominal aorta, but also occurs above the diaphragm in the thoracic aorta. Conventionally, the aorta has been considered a large vascular conduit with uniform cellular and extracellular structure and function. Evidence is accumulating, however, to suggest that variations exist between the thoracic and abdominal aorta, thereby demonstrating regional heterogeneity. Further pathophysiologic studies of aortic dilation in each of these regions have identified disparities in atherosclerotic plaque deposition, vessel mechanics, protease profiles, and cell signaling pathways. Improved understanding of this spatial heterogeneity may promote evolution in the management of aneurysm disease through computational models of aortic wall stress, imaging of proteolytic activity, targeted pharmacologic treatment, and the application of region-specific gene therapy.

Thoracic aortic aneurysm (TAA) is a clinically silent and potentially fatal disease whose pathophysiology is poorly understood. Application of data derived from animal models and human tissue analysis of abdominal aortic aneurysms may prove misleading given current evidence of structural and biochemical aortic heterogeneity above and below the diaphragm. Genetic predisposition is more common in TAA and includes multi-faceted syndromes such as Marfan, Loeys-Dietz, and type IV Ehlers-Danlos as well as autosomal-dominant familial patterns of inheritance. Investigation into the consequences of these known mutations has provided insight into the cell signaling cascades leading to degenerative remodeling of the aortic medial extracellular matrix (ECM) with TGF-β playing a major role. Targeted research into modifying the upstream regulation or downstream effects of the TGF-β1 pathway may provide opportunities for intervention to attenuate TAA progression.

Background-The effect of multiple integrated stimuli on vascular wall expression of matrix metalloproteinases (MMPs) remains unknown. Accordingly, this study examined the influence of the vasoactive peptide angiotensin II (Ang II) on wall tension-induced promoter activation of MMP-2, MMP-9, and membrane type-1 MMP (MT1-MMP). Methods and Results-Thoracic aortic rings harvested from transgenic reporter mice containing the MMP-2, MMP-9, or MT1-MMP promoter sequence fused to a reporter gene were subjected to 3 hours of wall tension at 70, 85, or 100 mm Hg, with or without 100 nM Ang II. Total RNA was harvested from the aortic rings, and reporter gene transcripts were quantified by quantitative real-time polymerase chain reaction to measure MMP promoter activity. MT1-MMP promoter activity was increased at both 85 and 100 mm Hg, compared with baseline tension of 70 mm Hg, whereas treatment with Ang II stimulated MT1-MMP promoter activity to the same degree at all tension levels (PϽ0.05). Elevated tension and Ang II displayed a potential synergistic enhancement of MMP-2 promoter activation at 85 and 100 mm Hg, whereas the same stimuli caused a decrease in MMP-9 promoter activity (PϽ0.05) at 100 mm Hg. Conclusions-This study demonstrated that exposure to a relevant biological stimulus (Ang II) in the presence of elevated tension modulated MMP promoter activation. Furthermore, these data suggest that a mechanical-molecular set point exists for the induction of MMP promoter activation and that this set point can be adjusted up or down by a secondary biological stimulus.

Background Congenital bicuspid aortic valves (BAVs) result from fusion of two valve cusps, resulting in left-noncoronary (L-N), right-left (R-L), and right-noncoronary (R-N) morphologies. BAVs predispose to ascending thoracic aortic aneurysms (ATAAs). This study hypothesized that ATAAs with each BAV morphology group possess unique signatures of matrix metalloproteinases (MMPs) and endogenous tissue inhibitors (TIMPs). Methods ATAA tissue from 46 BAV patients was examined for MMP/TIMP abundance and global MMP activity compared to normal aortic specimens (n=15). Proteolytic balance was calculated as the ratio of MMP abundance to a composite TIMP score (TS). Results were stratified by valve morphology group (L-N (n=6), R-L (n=31), and R-N(n=9)). Results The BAV specimens (p<0.05 vs. normal aorta, 100%) displayed elevated global MMP activity (273±63%), MMP-9 (263±47%), and decreased MMP -7 (56±10%), -8 (58±11%), TIMP -1 (63±7%) and -4 (38±3%). The R-L group showed increased global MMP activity (286±89%) and MMP-9 (267±55%) with reduced MMP -7 (45±7%) -8 (68±15%), TIMP -1 (58±7%) and -4 (35±3%). The L-N group showed elevated global MMP activity (284±71%), and decreased MMP-8 (37±17%) and TIMP-4 (48±14). In the R-N group, MMP -7 (46±13%) and -8 (36±17%), and TIMP -1 (59±10) and -4 (42±5%) were decreased. The R-L group demonstrated an increased proteolytic balance for MMP-1, MMP-9, and MMP-12 relative to L-N and R-N. Conclusion Each BAV morphology group possesses a unique signature of MMPs and TIMPs. MMP/TIMP score ratios suggest that the R-L group may be more aggressive, justifying earlier surgical intervention.

The prevalence of atherosclerotic disease continues to increase, and despite significant reductions in major cardiovascular events with current medical interventions, an additional therapeutic window exists. Atherosclerotic plaque growth is a complex integration of cholesterol penetration, inflammatory cell infiltration, vascular smooth muscle cell (VSMC) migration, and neovascular invasion. A family of matrix-degrading proteases, the matrix metalloproteinases (MMPs), contributes to all phases of vascular remodeling. The contribution of specific MMPs to endothelial cell integrity and VSMC migration in atherosclerotic lesion initiation and progression has been confirmed by the increased expression of these proteases in plasma and plaque specimens. Endogenous blockade of MMPs by the tissue inhibitors of metalloproteinases (TIMPs) may attenuate proteolysis in some regions, but the progression of matrix degeneration suggests that MMPs predominate in atherosclerotic plaque, precipitating vulnerability. Plaque neovascularization also contributes to instability and, coupling the known role of MMPs in angiogenesis to that of atherosclerotic plaque growth, interest in targeting MMPs to facilitate plaque stabilization continues to accumulate. This article aims to review the contributions of MMPs and TIMPs to atherosclerotic plaque expansion, neovascularization, and rupture vulnerability with an interest in promoting targeted therapies to improve plaque stabilization and decrease the risk of major cardiovascular events.

Thoracic aortic aneurysms (TAAs) develop as a result of dysregulated extracellular matrix remodeling mediated by several matrix metalloproteinases (MMPs). Membrane type-1 MMP (MT1-MMP) is the prototypical member of a unique family of membrane-bound MMPs, possessing multiple substrates and functions. The present study tested the hypothesis that MT1-MMP expression, abundance, and activity would be elevated during TAA development and that this protease is produced primarily by mesenchymal cells within the thoracic aorta. Descending thoracic aortas were harvested from C57BL/6J mice at multiple time points (2, 4, 8, and 16 wk, n = 15 each) post-TAA induction (0.5M CaCl(2), 15 min) and compared with reference controls (n = 15). The expression and abundance of MT1-MMP, MMP-2, and tissue inhibitor of metalloproteinase (TIMP)-2 were assessed by quantitative PCR and immunoblot analysis. MT1-MMP activity was determined by fluorescent peptide assay. MT1-MMP was localized within the aortic wall by immunohistochemistry. MT1-MMP abundance and localization in live animals (8 wk post-TAA induction vs. control) was determined by micro-ultrasound imaging with an MT1-MMP-targeted microbubble contrast agent. Aortic diameter was increased 172 +/- 7% at 16 wk post-TAA induction (P < 0.05). MT1-MMP and MMP-2 mRNA levels were elevated at 2 wk post-TAA induction (P < 0.05). MT1-MMP protein abundance increased progressively to a maximum of 178 +/- 26% at 16 wk post-TAA induction, whereas MMP-2 and TIMP-2 peaked at 2 wk post-TAA induction (526 +/- 93% and 376 +/- 48%, respectively, P < 0.05). MT1-MMP colocalized with fibroblasts, and MT1-MMP-targeted contrast binding was elevated in 8-wk TAA-induced mice versus control mice (217 +/- 53% vs. 81 +/- 8%, P < 0.05). In conclusion, these novel results suggest that MT1-MMP plays a dynamic multifunctional role in TAA development and, therefore, may provide a significant target for therapeutic strategies.

Percutaneous endoscopic gastrostomy (PEG) is the procedure of choice for establishing enteral access in patients unable to take oral feedings. Serious complications are rare; however, misplaced PEGs and PEG/ Jejunums can lead to hollow viscus injuries with intra-abdominal contamination and subsequent peritonitis, septicemia, and death. The presence of free intra-abdominal air is a reliable indicator of a perforated viscus and often points to a surgical emergency; however, in the case of PEGs, pneumoperitoneum without a perforated viscus, or “benign pneumoperitoneum” creates a diagnostic dilemma. To determine the incidence and clinical significance of pneumoperitoneum after PEG or PEG/Jejunum (J) we reviewed the records of 722 patients who underwent these procedures at our institution. Of 39 patients found to have free air after PEG/PEG/J placement, 33 (85%) had “benign pneumoperitoneum” and were discharged without complication or surgical intervention. Of the six patients with serious complications related to their procedure, five (83%) had clinical signs of intra-abdominal complications (peritonitis) that helped guide their management. Of these six patients, the two receiving abdominal radiographs instead of abdominal CT scanning had a 50 per cent negative laparotomy rate. We present an algorithm for the management of patients found to have pneumoperitoneum after PEG or PEG/J placement.