Multilevel Assessment of Stent-Induced Inflammation in the Adjacent Vascular Tissue.

A recent U.S. Food and Drug Administration report presented the currently available scientific information related to biological response to metal implants. In this work, a multilevel approach was employed to assess the implant-induced and biocorrosion-related inflammation in the adjacent vascular tissue using a mouse stent implantation model. The implications of biocorrosion on peri-implant tissue were assessed at the macroscopic level via in vivo imaging and histomorphology. Elevated matrix metalloproteinase activity, colocalized with the site of implantation, and histological staining indicated that stent surface condition and implantation time affect the inflammatory response and subsequent formation and extent of neointima. Hematological measurements also demonstrated that accumulated metal particle contamination in blood samples from corroded-stetted mice causes a stronger immune response. At the cellular level, the stent-induced alterations in the nanostructure, cytoskeleton, and mechanical properties of circulating lymphocytes were investigated. It was found that cells from corroded-stented samples exhibited higher stiffness, in terms of Young's modulus values, compared to noncorroded and sham-stented samples. Nanomechanical modifications were also accompanied by cellular remodeling, through alterations in cell morphology and stress (F-actin) fiber characteristics. Our analysis indicates that surface wear and elevated metal particle contamination, prompted by corroded stents, may contribute to the inflammatory response and the multifactorial process of in-stent restenosis. The results also suggest that circulating lymphocytes could be a novel nanomechanical biomarker for peri-implant tissue inflammation and possibly the early stage of in-stent restenosis. Large-scale studies are warranted to further investigate these findings.

Pro-Healing Nanomatrix-Coated Stent Analysis in an In Vitro Vascular Double-Layer System and in a Rabbit Model.

Cardiovascular stent technologies have significantly improved over time. However, their optimal performance remains limited by restenosis, thrombosis, inflammation, and delayed re-endothelialization. Current stent designs primarily target inhibition of neointimal proliferation but do not promote functional arterial healing (pro-healing) in order to restore normal vascular reactivity. The endothelial lining that does develop with current stents appears to have loose intracellular junctions. We have developed a pro-healing nanomatrix coating for stents that enhances healing while limiting neointimal proliferation. This builds on our prior work evaluating the effects of the pro-healing nanomatrix coating on cultures of vascular endothelial cells (ECs), smooth muscle cells (SMCs), monocytes, and platelets. However, when a stent is deployed in an artery, multiple vascular cell types interact, and their interactions affect stent performance. Thus, in our current study, an in vitro vascular double-layer (VDL) system was used to observe stent effects on communication between different vascular cell types. Additionally, we assessed the pro-healing ability and vascular cell interactions after stent deployment in the VDL system and in a rabbit model, evaluating the nanomatrix-coated stent compared to a commercial bare metal stent (BMS) and a drug eluting stent (DES). In vitro results indicated that, in a layered vascular structure, the pro-healing nanomatrix-coated stent could (1) improve endothelialization and endothelial functions, (2) regulate SMC phenotype to reduce SMC proliferation and migration, (3) suppress inflammation through a multifactorial manner, and (4) reduce foam cell formation, extracellular matrix remodeling, and calcification. Consistent with this, in vivo results demonstrated that, compared with commercial BMS and DES, this pro-healing nanomatrix-coated stent enhanced re-endothelialization with negligible restenosis, inflammation, or thrombosis. Thus, these findings indicate the unique pro-healing features of this nanomatrix stent coating with superior efficacy over commercial BMS and DES.

Use of Telepathology to Facilitate COVID-19 Research and Education through an Online COVID-19 Autopsy Biorepository.

INTRODUCTION: The coronavirus disease 2019 (COVID-19) pandemic has increased the use of technology for communication including departmental conferences, working remotely, and distance teaching. Methods to enable these activities should be developed and promulgated. OBJECTIVE: To repurpose a preexisting educational website to enable the development of a COVID-19 autopsy biorepository to support distance teaching and COVID-19 research. METHODS: After consent was obtained, autopsies were performed on patients with a confirmed positive severe acute respiratory syndrome coronavirus-2 reverse-transcriptase-polymerase-chain reaction test. Autopsies were performed according to a COVID-19 protocol, and all patients underwent both gross and microscopic examination. The H and E histology slides were scanned using a Leica Biosystems Aperio CS ScanScope whole slide scanner and the digital slide files were converted to deep zoom images that could be uploaded to the University of Alabama at Birmingham (UAB) Pathology Educational Instructional Resource website where virtual microscopy of the slides is available. RESULTS: A total of 551 autopsy slides from 24 UAB COVID-19 cases, 1 influenza H1N1 case and 1 tuberculosis case were scanned and uploaded. Five separate COVID-19 research teams used the digital slides remotely with or without a pathologist on a Zoom call. The scanned slides were used to produce one published case report and one published research project. The digital COVID-19 autopsy biorepository was routinely used for educational conferences and research meetings locally, nationally and internationally. CONCLUSION: The repurposing of a pre-existing website enabled telepathology consultation for research and education purposes. Combined with other communication technology (Zoom) this achievement highlights what is possible using pre-existing technologies during a global pandemic.

In vivo monitoring of the inflammatory response in a stented mouse aorta model.

The popularity of vascular stents continues to increase for a variety of applications, including coronary, lower limb, renal, carotid, and neurovascular disorders. However, their clinical effectiveness is hindered by numerous postdeployment complications, which may stimulate inflammatory and fibrotic reactions. The purpose of this study was to evaluate the vessel inflammatory response via in vivo imaging in a mouse stent implantation model. Corroded and noncorroded self-expanding miniature nitinol stents were implanted in mice abdominal aortas, and novel in vivo imaging techniques were used to assess trafficking and accumulation of fluorescent donor monocytes as well as cellular proliferation at the implantation site. Monocytes were quantitatively tracked in vivo and found to rapidly clear from circulation within hours after injection. Differences were found between the test groups with respect to the numbers of recruited monocytes and the intensity of the resulting fluorescent signal. Image analysis also revealed a subtle increase in matrix metalloproteinase activity in corroded compared with the normal stented aortas. In conclusion, this study has been successful in developing a murine stent inflammation model and applying novel in vivo imaging tools and methods to monitor the complex biological processes of the host vascular wall response.

Histologic analysis of a covered stent implanted for pseudoaneurysm in a coronary artery.

Covered stents are the standard of care when coronary perforations complicate percutaneous coronary interventions and have also been utilized in the treatment of coronary aneurysms. We present the clinical and histologic features of a patient who developed a coronary perforation and pseudoaneurysm 4 years following deployment of intracoronary stents. Although the covered stent successfully sealed the perforation, subsequent thrombosis led to a fatal acute myocardial infarction.

Stent overlapping and geometric curvature influence the structural integrity and surface characteristics of coronary nitinol stents.

Preliminary studies have revealed that some stents undergo corrosion and fatigue-induced fracture in vivo, with significant release of metallic ions into surrounding tissues. A direct link between corrosion and in-stent restenosis has not been clearly established; nonetheless in vitro studies have shown that relatively high concentrations of heavy metal ions can stimulate both inflammatory and fibrotic reactions, which are the main steps in the process of restenosis. To isolate the mechanical effects from the local biochemical effects, accelerated biomechanical testing was performed on single and overlapping Nickel-Titanium (NiTi) stents subjected to various degrees of curvature. Post testing, stents were evaluated using Scanning Electron Microscopy (SEM) to identify the type of surface alterations. Fretting wear was observed in overlapping cases, in both straight and curved configurations. Stent strut fractures occurred in the presence of geometric curvature. Fretting wear and fatigue fractures observed on stents following mechanical simulation were similar to those from previously reported human stent explants. It has been shown that biomechanical factors such as arterial curvature combined with stent overlapping enhance the incidence and degree of wear and fatigue fracture when compared to single stents in a straight tube configuration.

Smooth muscle cell-specific runx2 deficiency inhibits vascular calcification.

RATIONALE: Vascular calcification is a hallmark of atherosclerosis, a major cause of morbidity and mortality in the United States. We have previously reported that the osteogenic transcription factor Runx2 is an essential and sufficient regulator of calcification of vascular smooth muscle cells (VSMC) in vitro. OBJECTIVE: To determine the contribution of osteogenic differentiation of VSMC to the pathogenesis of vascular calcification and the function of VSMC-derived Runx2 in regulating calcification in vivo. METHODS AND RESULTS: SMC-specific Runx2-deficient mice, generated by breeding SM22α-Cre mice with the Runx2 exon 8 floxed mice, exhibited normal aortic gross anatomy and expression levels of SMC-specific marker genes. Runx2 deficiency did not affect basal SMC markers, but inhibited oxidative stress-reduced expression of SMC markers. High-fat-diet-induced vascular calcification in vivo was markedly inhibited in the Runx2-deficient mice in comparison with their control littermates. Runx2 deficiency inhibited the expression of receptor activator of nuclear factor κB ligand, which was accompanied by decreased macrophage infiltration and formation of osteoclast-like cells in the calcified lesions. Coculture of VSMC with bone marrow-derived macrophages demonstrated that the Runx2-deficient VSMC failed to promote differentiation of macrophages into osteoclast-like cells. CONCLUSIONS: These data have determined the importance of osteogenic differentiation of VSMC in the pathogenesis of vascular calcification in mice and defined the functional role of SMC-derived Runx2 in regulating vascular calcification and promoting infiltration of macrophages into the calcified lesion to form osteoclast-like cells. Our studies suggest that the development of vascular calcification is coupled with the formation of osteoclast-like cells, paralleling the bone remodeling process.

The role of vascular calcification in inducing fatigue and fracture of coronary stents.

Traditional approaches for in-vitro pulsatile and fatigue testing of endovascular stents do not take into consideration the pathologies of the stented vessel and their associated biomechanical effects. One important pathology is calcification, which may be capable of inducing changes in the vessel wall leading to inhomogeneous distribution of stresses combined with wall motion during the cardiac cycle. These local property changes in the region adjacent to stents could directly influence in-vivo stent performance. Seven cases containing a total of 18 stents were obtained from autopsy. Radiographs were evaluated and vessels were sectioned for histology and stent topographical analysis. Stents were retrieved by chemical removal of surrounding tissue and surfaces were evaluated using 3D digital optical and scanning electron microscopy for biomechanical abrasion and fracture features. Pathologic complications such as restenosis and thrombus formation were assessed from histological sections. Direct evidence of fracture was found in 6 of the 7 cases (in 12 out of 18 stents; 9 drug eluting and 3 bare metal). The degree of stent alterations was variable, where separation of segments due to fracture occurred mostly in drug-eluting stents. All fracture surfaces were representative of a high cycle fatigue mechanism. These fractures occurred in complex lesions involving the presence of diffuse calcification alone, or in combination with vessel angulations and multiple overlapping stents. Morphologic analysis of tissue at or near some fracture sites showed evidence of thrombus formation and/or neointimal tissue growth.

Aging and living with HIV/AIDS.

With individuals either living longer with the disease or contracting it at a later stage in life, HIV/AIDS can no longer be regarded as just a young person's disease. In fact, people older than 50 will represent 50% of HIV/AIDS cases by 2015. The intersection of aging and HIV/AIDS is explored in this article through the use of an individual example highlighting typical age-related issues encountered in living with a chronic HIV infection and two of the more common comorbid conditions. Nursing implications for managing these conditions-depression and dyslipidemia-as well as other considerations for providing care to older adults with HIV/AIDS are addressed.

The role of fibrinolytic genes and proteins in the development of allograft vascular disease.

BACKGROUND: We have previously shown that lack of plasminogen activator inhibitor-1 (PAI-1) expression in donor tissue greatly increases intimal proliferation (IP) after allogeneic transplantation. We sought to determine the relative role of PAI-1 and other fibrinolytic proteins in the development of IP. METHODS: We used an abdominal aortic transplant model in mice to investigate IP in 3 groups of 6 recipients. In the isograft group, CBA/J strain mice were donors and recipients, donors for allograft group were C57BL/6J mice, and for the allograft/knockout group, C57BL/6J PAI-1 knockout mice. All groups received weekly injections of anti-CD8/CD4 monoclonal antibodies. IP was calculated at 50 days, and sections were analyzed for fibrinolytic proteins, messenger RNA (mRNA) and PAI-1 activity using immunohistochemistry (IHC), in situ hybridization (ISH), reverse transcription-polymerase chain reaction (RT-PCR), and Western blot analysis. RESULTS: Significantly more IP developed in the allograft/knockout group vs the isograft (p < 0.001) and the allograft groups (p = 0.003). There was marked intimal expression of tissue plasminogen activator (tPA), urokinase PA (uPA), and uPA receptor (uPAR) proteins and mRNA in the allograft and allograft/knockout groups vs the isograft group. Allografts also showed significant intimal staining for PAI-1 protein and mRNA. RT-PCR demonstrated a stepwise increase in profibrinolytic protein mRNA from isograft to allograft to allograft/knockout groups, particularly uPA (p = 0.02) and uPAR (p = 0.016). Western blot data showed complementary findings. PAI-1 activity was persistently present in isograft and allograft animals, only. Intimas in allograft and allograft/knockout groups were primarily smooth muscle cells. CONCLUSIONS: PAI-1 reduces IP by limiting smooth muscle cell activity, with little change in matrix composition likely by modulating profibrinolytic protein expression.

Runx2-upregulated receptor activator of nuclear factor κB ligand in calcifying smooth muscle cells promotes migration and osteoclastic differentiation of macrophages.

OBJECTIVE: Clinical and experimental studies demonstrate the important roles of vascular smooth muscle cells (VSMC) in the pathogenesis of atherosclerosis. We have previously determined that the osteogenic transcription factor Runx2 is essential for VSMC calcification. The present study characterized Runx2-regulated signals and their potential roles in vascular calcification. METHODS AND RESULTS: In vivo studies with atherogenic apolipoprotein E(-/-) mice demonstrated that increased oxidative stress was associated with upregulation of Runx2 and receptor activator of nuclear factor κB ligand (RANKL), which colocalized in the calcified atherosclerotic lesions and were juxtaposed to infiltrated macrophages and osteoclast-like cells that are positively stained for an osteoclast marker, tartrate-resistant acid phosphatase. Mechanistic studies using RNA interference, a luciferase reporter system, chromatin immunoprecipitation, and electrophoretic mobility shift assays indicated that Runx2 regulated the expression of RANKL via a direct binding to the 5'-flanking region of the RANKL. Functional characterization revealed that RANKL did not induce VSMC calcification, nor was RANKL required for oxidative stress-induced VSMC calcification. Using a coculture system, we demonstrated that VSMC-expressed RANKL induced migration as well as differentiation of bone marrow-derived macrophages into multinucleated, tartrate-resistant acid phosphatase-positive osteoclast-like cells. These effects were inhibited by the RANKL antagonist osteoprotegerin and with VSMC deficient in Runx2 or RANKL. CONCLUSION: We demonstrate that Runx2 directly binds to the promoter and controls the expression of RANKL, which mediates the crosstalk between calcifying VSMC and migration and differentiation of macrophages into osteoclast-like cells in the atherosclerotic lesions. Our studies provide novel mechanistic insights into the regulation and function of VSMC-derived RANKL in the pathogenesis of atherosclerosis and vascular calcification.

In-vivo corrosion and local release of metallic ions from vascular stents into surrounding tissue.

OBJECTIVES: To evaluate retrieved bare metal vascular stents and surrounding tissue. BACKGROUND: Limited information is available regarding the condition of stent surfaces and their interaction with vascular tissue following implantation. Corrosion of stents presents two main risks: release of metallic ions into tissue and deterioration of the mechanical properties of stents which may contribute to fracture. Release of heavy metal ions could alter the local tissue environment leading to up-regulation of inflammatory mediators and promote in-stent restenosis. METHODS: Nineteen cases were collected from autopsy, heart explants for transplant, and vascular surgery (23 vessels containing 33 bare metal stents). A method was developed for optimal tissue dissolution and separation of the stent/tissue components without inducing stent corrosion. When available, chemical analysis was performed to assess metallic content in both the control and dissolved tissue solutions. Electron microscopy and digital optical microscopy imaging were used to evaluate stents. RESULTS: Twelve of the 33 stents showed varying degrees of corrosion. Metallic levels in the tissue surrounding the corroded stents were significantly higher (0.5-3.0 mcg/cm² stent) than in control solutions (0-0.30 mcg/cm² stent) and in tissue surrounding stents that did not undergo corrosion (0- 0.20 mcg/cm² stent). CONCLUSIONS: Corrosion of some retrieved stents is described which leads to transfer of heavy metal ions into surrounding tissue. The contribution of this metallic ion release to the mechanisms of in-stent restenosis as well as its effect on the mechanical properties of stents is unknown and requires further investigation.

Clinical device-related article surface characterization of explanted endovascular stents: evidence of in vivo corrosion.

Limited information exists regarding the in vivo stability of endovascular stents. Nine excised human vascular segments with implanted stents (n = 16) manufactured from stainless steel, nickel-titanium, tantalum, and cobalt-based alloys were analyzed. The stent/tissue components were separated using an established tissue dissolution protocol and control and explanted stents were evaluated by digital optical and electron microscopy. Metallic content in surrounding tissues was measured by mass spectroscopy. Surface alterations, consistent with corrosion mediated by electrochemical and mechanical factors, were observed in 9 of the 16 explanted stents and were absent from control stents. Tissue dissolved from around corroded stents corresponded with a higher metallic content. The effect of these changes in the microtopography of stents on their mechanical properties (fatigue strength and fracture limit) in addition to the potential for released metallic debris contributing to the biological mechanisms of in-stent restenosis supports the need for further investigations.

Perinatal tobacco smoke exposure increases vascular oxidative stress and mitochondrial damage in non-human primates.

Epidemiological studies suggest that events occurring during fetal and early childhood development influence disease susceptibility. Similarly, molecular studies in mice have shown that in utero exposure to cardiovascular disease (CVD) risk factors such as environmental tobacco smoke (ETS) increased adult atherogenic susceptibility and mitochondrial damage; however, the molecular effects of similar exposures in primates are not yet known. To determine whether perinatal ETS exposure increased mitochondrial damage, dysfunction and oxidant stress in primates, archived tissues from the non-human primate model Macaca mulatta (M. mulatta) were utilized. M. mulatta were exposed to low levels of ETS (1 mg/m(3) total suspended particulates) from gestation (day 40) to early childhood (1 year), and aortic tissues were assessed for oxidized proteins (protein carbonyls), antioxidant activity (SOD), mitochondrial function (cytochrome oxidase), and mitochondrial damage (mitochondrial DNA damage). Results revealed that perinatal ETS exposure resulted in significantly increased oxidative stress, mitochondrial dysfunction and damage which were accompanied by significantly decreased mitochondrial antioxidant capacity and mitochondrial copy number in vascular tissue. Increased mitochondrial damage was also detected in buffy coat tissues in exposed M. mulatta. These studies suggest that perinatal tobacco smoke exposure increases vascular oxidative stress and mitochondrial damage in primates, potentially increasing adult disease susceptibility.

A nitric oxide releasing, self assembled peptide amphiphile matrix that mimics native endothelium for coating implantable cardiovascular devices.

Cardiovascular disease is the number one cause of death in the United States. Deployment of stents and vascular grafts has been a major therapeutic method for treatment. However, restenosis, incomplete endothelialization, and thrombosis hamper the long term clinical success. As a solution to meet these current challenges, we have developed a native endothelial ECM mimicking self-assembled nanofibrous matrix to serve as a new treatment model. The nanofibrous matrix is formed by self-assembly of peptide amphiphiles (PAs), which contain nitric oxide (NO) donating residues, endothelial cell adhesive ligands composed of YIGSR peptide sequence, and enzyme-mediated degradable sites. NO was successfully released from the nanofibrous matrix rapidly within 48 h, followed by sustained release over period of 30 days. The NO releasing nanofibrous matrix demonstrated a significantly enhanced proliferation of endothelial cells (51+/-3% to 67+/-2%) but reduced proliferation of smooth muscle cells (35+/-2% to 16+/-3%) after 48 h of incubation. There was also a 150-fold decrease in platelet attachment on the NO releasing nanofibrous matrix (470+/-220 platelets/cm(2)) compared to the collagen-I (73+/-22 x 10(3)platelets/cm(2)) coated surface. The nanofibrous matrix has the potential to be applied to various cardiovascular implants as a self-assembled coating, thereby providing a native endothelial extracellular matrix (ECM) mimicking environment.

Stainless steel ions stimulate increased thrombospondin-1-dependent TGF-beta activation by vascular smooth muscle cells: implications for in-stent restenosis.

BACKGROUND/AIMS: Despite advances in stent design, in-stent restenosis (ISR) remains a significant clinical problem. All implant metals exhibit corrosion, which results in release of metal ions. Stainless steel (SS), a metal alloy widely used in stents, releases ions to the vessel wall and induces reactive oxygen species, inflammation and fibroproliferative responses. The molecular mechanisms are unknown. TGF-beta is known to be involved in the fibroproliferative responses of vascular smooth muscle cells (VSMCs) in restenosis, and TGF-beta antagonists attenuate ISR. We hypothesized that SS ions induce the latent TGF-beta activator, thrombospondin-1 (TSP1), through altered oxidative signaling to stimulate increased TGF-beta activation and VSMC phenotype change. METHODS: VSMCs were treated with SS metal ion cocktails, and morphology, TSP1, extracellular matrix production, desmin and TGF-beta activity were assessed by immunoblotting. RESULTS: SS ions stimulate the synthetic phenotype, increased TGF-beta activity, TSP1, increased extracellular matrix and downregulation of desmin in VSMCs. Furthermore, SS ions increase hydrogen peroxide and decrease cGMP-dependent protein kinase (PKG) signaling, a known repressor of TSP1 transcription. Catalase blocks SS ion attenuation of PKG signaling and increased TSP1 expression. CONCLUSIONS: These data suggest that ions from stent alloy corrosion contribute to ISR through stimulation of TSP1-dependent TGF-beta activation.

Severe, diffuse coronary artery spasm after drug-eluting stent placement.

OBJECTIVES: Three cases of severe, diffuse coronary artery spasm after drug-eluting stent placement at our institution prompted this review. BACKGROUND: Drug-eluting stents have gained widespread use due to extraordinarily low rates of restenosis. Despite these generally superior clinical outcomes, the specter of rare idiosyncratic reactions remains a concern. METHODS: We performed searches of Medline and the U.S. FDA Manufacturer And User facility Device Experience database (MAUDE) to identify and describe spasm after coronary stent placement. Searches included drug-eluting and bare-metal stents. Institutional cases are reviewed. Location, time course and outcome of cases are described. RESULTS: Thirteen cases of spasm were identified after stent placement. Seven cases occurred after Cypher drug-eluting stent placement, 2 after Taxus drug-eluting stent placement, 1 after BiodivYsio and 3 after bare-metal stents. Five patients experienced diffuse, multivessel spasm--2 after Cypher, 2 after Taxus, and 1 after a Velocity stent. Of these 5 patients, 2 died. An additional 2 required intra-aortic balloon pump placement for cardiogenic shock. Another had persistent symptomatic, diffuse coronary spasm documented by angiography at 1 year. CONCLUSIONS: We describe coronary spasm after stent placement, particularly after drug-eluting stents. Outcomes associated with diffuse severe spasm after stenting are poor, and the pathophysiology remains poorly understood.