Vasodilation of Isolated Vessels and the Isolation of the Extracellular Matrix of Tight-skin Mice

The interferon regulatory factor 5 (IRF5) is crucial for cells to determine if they respond in a pro-inflammatory or anti-inflammatory fashion. IRF5's ...

Non-thermal Infrared Light Treatment of Ischemia/Reperfusion Injury and Subsequent Analysis of Macrophage Differentiation

Tissue damage and necrosis from inflammatory processes are a consequence of ischemia reperfusion injury (IRI). In skeletal muscle, ischemia reduces the ...

Medical College of Wisconsin

Systemic sclerosis (SSc) is an autoimmune connective tissue disorder characterized by oxidative stress, impaired vascular function, and attenuated angiogenesis. The tight-skin (Tsk(-/+)) mouse is a model of SSc that displays many of the cellular features of the clinical disease. We tested the hypotheses that abnormal fibrillin-1 expression and chronic phospholipid oxidation occur in Tsk(-/+) mice and, furthermore, that these factors precipitate a prooxidant state, collagen-related protein expression, apoptosis, and mesenchymal transition in endothelial cells cultured on Tsk(-/+) extracellular matrix. Human umbilical vein endothelial cells were seeded on microfibrils isolated from skin of C57BL/6J (control) and Tsk(-/+) mice in the presence or absence of chronic pretreatment with the apolipoprotein Apo A-I mimetic D-4F (1 mg·kg(-1)·day(-1) ip for 6 to 8 wk). Nitric oxide-to-superoxide anion ratio was assessed 12 h after culture, and cell proliferation, apoptosis, and phenotype were studied 72 h after culture. Tsk(-/+) mice demonstrated abnormal "big fibrillin" expression (405 kDa) by Western blot analysis compared with control. Endothelial cells cultured on microfibrils prepared from Tsk(-/+) mice demonstrated reduced proliferation, a prooxidant state (reduced nitric oxide-to-superoxide anion ratio), increased apoptosis, and collagen-related protein expression associated with mesenchymal transition. Chronic D-4F pretreatment of Tsk(-/+) mice attenuated many of these adverse effects. The findings demonstrate that abnormal fibrillin-1 expression and chronic oxidative stress mediate endothelial mesenchymal transition in Tsk(-/+) mice. This mesenchymal transition may contribute to the reduction in angiogenesis that is known to occur in this model of SSc.

Abnormal fibrillin-1 expression and chronic oxidative stress mediate endothelial mesenchymal transition in a murine model of systemic sclerosis.

The tight skin mouse (Tsk(-/+)) is a model of scleroderma characterized by impaired vasoreactivity, increased oxidative stress, attenuated angiogenic response to VEGF and production of the angiogenesis inhibitor angiostatin. Low-level light therapy (LLLT) stimulates angiogenesis in myocardial infarction and chemotherapy-induced mucositis. We hypothesize that repetitive LLLT restores vessel growth in the ischemic hindlimb of Tsk(-/+) mice by attenuating angiostatin and enhancing angiomotin effects in vivo. C57Bl/6J and Tsk(-/+) mice underwent ligation of the femoral artery. Relative blood flow to the foot was measured using a laser Doppler imager. Tsk(-/+) mice received LLLT (670 nm, 50 mW cm(-2), 30 J cm(-2)) for 10 min per day for 14 days. Vascular density was determined using lycopersicom lectin staining. Immunofluorescent labeling, Western blot analysis and immunoprecipitation were used to determine angiostatin and angiomotin expression. Recovery of blood flow to the ischemic limb was reduced in Tsk(-/+) compared with C57Bl/6 mice 2 weeks after surgery. LLLT treatment of Tsk(-/+) mice restored blood flow to levels observed in C57Bl/6 mice. Vascular density was decreased, angiostatin expression was enhanced and angiomotin depressed in the ischemic hindlimb of Tsk(-/+) mice. LLLT treatment reversed these abnormalities. LLLT stimulates angiogenesis by increasing angiomotin and decreasing angiostatin expression in the ischemic hindlimb of Tsk(-/+) mice.

Transient repetitive exposure to low level light therapy enhances collateral blood vessel growth in the ischemic hindlimb of the tight skin mouse.

Metal on metal articulations in hip arthroplasty offer advantages, including lower volumetric wear compared to conventional metalonpolyethylene bearings, and increased resistance to dislocation. Reports described early failures, with histologic features similar to a Type IV immune response. Mechanisms by which metal wear products cause this reaction are not completely understood. We hypothesized a mechanism through direct activation of endothelial cells (ECs) by metal ions, resulting in both vasculitis and accumulation of lymphocytes without prior immune sensitization. Effects of metal ions were evaluated using human ECs in culture. Alterations in chemotactic proteins IL8 and MCP1 were assessed, as was upregulation of the adhesion molecule ICAM-1 and lymphocyte binding to ECs. Cobalt increased secretion of IL8 and MCP1 significantly, and upregulated the expression of ICAM-1 in ECs compared to stimulation by chromium and controls. Binding of lymphocytes to ECs and transEC migration were both significantly increased by cobalt but not chromium. These findings suggest that cobalt contributes more to the activation of ECs and lymphocyte binding than chromium without an allergic response. Some of the adverse tissue reactions to implants with components made of cobalt-chromium-molybdenium alloys may be due in part to activation of the endothelium by metal ions.

Metal ions activate vascular endothelial cells and increase lymphocyte chemotaxis and binding.

Implant fixation through osseointegration is essential for the success of uncemented total joint arthroplasty, and nature and composition of implant surface play a critical role in this process. Despite widespread use of uncemented implants, the extent of bone ingrowth into implants is generally only a small percentage of the total implant surface. An understanding of the processes whereby bone cells grow into and multiply on porous surfaces is critical for the design and manufacture of implants that maximize ingrowth and implant fixation. A wide variety of implant materials are currently utilized for uncemented total joint arthroplasty, including titanium mesh, cobalt chromium beads, and tantalum deposited on a carbon network. Despite differences in physical and chemical properties of these materials, all have functioned well clinically. Therefore, the goals of this study were to compare and contrast the effects of these materials on the proliferation, phenotypic maturation, and mineralization of osteoblasts. Disks of porous tantalum, titanium mesh, and cobalt chromium beaded surfaces were fabricated and processed employing the same methods used to produce implants, including packaging and sterilization. Preosteoblasts were plated on disks, cellular morphology was evaluated by scanning electron microscopy. Osteoblast proliferation was significantly higher on the porous tantalum compared to other implant surfaces. Alkaline phosphatase activity, osteocalcin secretion, and upregulation of RUNX2 were inversely proportional to the rate of proliferation. Mineralization of osteoblasts paralleled the rate of proliferation. These findings suggest that proliferation of osteoblasts into the interstices of implant materials along with delayed maturation were favorable for increased bone ongrowth and ultimately implant stabilization.

Porous ongrowth surfaces alter osteoblast maturation and mineralization.

Interferon regulatory factor 5 (IRF5) has been called a "master switch" for its ability to determine whether cells mount proinflammatory or anti-inflammatory responses. Accordingly, IRF5 should be an attractive target for therapeutic drug development. Here we report on the development of a novel decoy peptide inhibitor of IRF5 that decreases myocardial inflammation and improves vascular endothelial cell (EC) function in tight-skin (Tsk/+) mice. Biolayer interferometry studies showed the Kd of IRF5D for recombinant IRF5 to be 3.72 ± 0.74x10-6M. Increasing concentrations of IRF5D (0-100 μg/mL, 24h) had no significant effect on EC proliferation or apoptosis. Treatment of Tsk/+ mice with IRF5D (1mg/kg/d subcutaneously, 21d) reduced IRF5 and ICAM-1 expression and monocyte/macrophage and neutrophil counts in Tsk/+ hearts compared to expression in hearts from PBS-treated Tsk/+ mice (p<0.05). EC-dependent vasodilatation of facialis arteries isolated from PBS-treated Tsk/+ mice was reduced (~15%). IRF5D treatments (1mg/kg/d, 21d) improved vasodilatation in arteries isolated from Tsk/+ mice nearly 3-fold (~45%, p<0.05), representing nearly 83% of the vasodilatation in arteries isolated from C57Bl/6J mice (~55%). IRF5D (50μg/mL, 24h) reduced nuclear translocation of IRF5 in myocytes cultured on both Tsk/+ cardiac matrix and C57Bl/6J cardiac matrix (p<0.05). These data suggest that IRF5 plays a causal role in inflammation, fibrosis and impaired vascular EC function in Tsk/+ mice and that treatment with IRF5D effectively counters IRF5-dependent mechanisms of inflammation and fibrosis in the myocardium in these mice.

An IRF5 Decoy Peptide Reduces Myocardial Inflammation and Fibrosis and Improves Endothelial Cell Function in Tight-Skin Mice.

Diabetes mellitus is a significant risk factor for peripheral artery disease. Diabetes mellitus induces chronic states of oxidative stress and vascular inflammation that increase neutrophil activation and release of myeloperoxidase. The goal of this study is to determine whether inhibiting myeloperoxidase reduces oxidative stress and neutrophil infiltration, increases vascularization, and improves blood flow in a diabetic murine model of hindlimb ischaemia.

Janine Struve

Department of Orthopedic Surgery

Medical College of Wisconsin

Vasodilation of Isolated Vessels and the Isolation of the Extracellular Matrix of Tight-skin Mice

Non-thermal Infrared Light Treatment of Ischemia/Reperfusion Injury and Subsequent Analysis of Macrophage Differentiation

Janine Struve

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Department of Orthopedic Surgery

Medical College of Wisconsin

Vasodilation of Isolated Vessels and the Isolation of the Extracellular Matrix of Tight-skin Mice

Non-thermal Infrared Light Treatment of Ischemia/Reperfusion Injury and Subsequent Analysis of Macrophage Differentiation

Department of Orthopedic Surgery