The fate of an endothelium layer after preconditioning

BackgroundA strategy in minimizing thrombotic events of vascular constructs is to seed the luminal surface with autologous endothelial cells (ECs). The task of seeding ECs can be achieved via bioreactors, which induce mechanical forces (shear stress, strain, pressure) onto the ECs. Although bioreactors can achieve a confluent layer of ECs in vitro, their acute response to blood remains unclear. Moreover, the necessary mechanical conditions that will increase EC adhesion and function remain unclear. We hypothesize that preconditioning seeded endothelium under physiological flow will enhance their retention and function.ObjectiveTo determine the role of varying preconditioning protocols on seeded ECs in vitro and in vivo.MethodsScaffolds derived from decelluarized arteries seeded with autologous ECs were preconditioned for 9 days. Three specific protocols, low steady shear stress (SS), high SS, and cyclic SS were investigated. After preconditioning, the seeded grafts were exposed to 15 minutes of blood via an ex vivo arteriovenous shunt model or alternately an in vivo arteriovenous bypass graft model.ResultsThe shunt model demonstrated ECs remained intact for all conditions. In the arteriovenous bypass model, only the cyclic preconditioned grafts remained intact, maintained morphology, and resisted the attachment of circulating blood elements such as platelets, red blood cells, and leukocytes. Western blotting analysis demonstrated an increase in the protein expression of eNOS and prostaglandin I synthase for the cyclic high shear stress-conditioned cells relative to cells conditioned with high shear stress alone.ConclusionCyclic preconditioning has been shown here to increase the ECs ability to resist blood flow-induced shear stress and the attachment of circulating blood elements, key attributes in minimizing thrombotic events. These studies may ultimately establish protocols for the formation of a more durable endothelial monolayer that may be useful in the context of small vessel arterial reconstruction.Clinical RelevanceThe importance of ECs toward patency has been demonstrated by the superior performance of endothelialized vein compared with prosthetic vascular graft materials. This article evaluates conditioning protocols for bioengineered vascular conduits to improve endothelial retention. This study describes approaches to improve bioengineered vessels as a potential alternative to conventional prosthetic vascular grafts

The Pathology of Neoatherosclerosis in Human Coronary Implants Bare-Metal and Drug-Eluting Stents

ObjectivesHuman coronary bare-metal stents (BMS) and drug-eluting stents (DES) from autopsy cases with implant duration >30 days were examined for the presence of neointimal atherosclerotic disease.BackgroundNeointimal atherosclerotic change (neoatherosclerosis) after BMS implantation is rarely reported and usually occurs beyond 5 years. The incidence of neoatherosclerosis after DES implantation has not been reported.MethodsAll available cases from the CVPath stent registry (n = 299 autopsies), which includes a total of 406 lesions—197 BMS, 209 DES (103 sirolimus-eluting stents [SES] and 106 paclitaxel-eluting stents [PES])—with implant duration >30 days were examined. Neoatherosclerosis was recognized as clusters of lipid-laden foamy macrophages within the neointima with or without necrotic core formation.ResultsThe incidence of neoatherosclerosis was significantly greater in DES lesions (31%) than BMS lesions (16%; p < 0.001). The median stent duration with neoatherosclerosis was shorter in DES than BMS (DES, 420 days [interquartile range [IQR]: 361 to 683 days]; BMS, 2,160 days [IQR: 1,800 to 2,880 days], p < 0.001). Unstable lesions characterized as thin-cap fibroatheromas or plaque rupture were more frequent in BMS (n = 7, 4%) than in DES (n = 3, 1%; p = 0.17), with relatively shorter implant durations for DES (1.5 ± 0.4 years) compared to BMS (6.1 ± 1.5 years). Independent determinants of neoatherosclerosis identified by multiple logistic regression included younger age (p < 0.001), longer implant durations (p < 0.001), SES usage (p < 0.001), PES usage (p = 0.001), and underlying unstable plaques (p = 0.004).ConclusionsNeoatherosclerosis is a frequent finding in DES and occurs earlier than in BMS. Unstable features of neoatherosclerosis are identified for both BMS and DES with shorter implant durations for the latter. The development of neoatherosclerosis may be yet another rare contributing factor to late thrombotic events

In vitro and in vivo Assessment of Keratose as a Novel Excipient of Paclitaxel Coated Balloons

Purpose: Drug coated balloons (DCB) are continually improving due to advances in coating techniques and more effective excipients. Paclitaxel, the current drug choice of DCB, is a microtubule-stabilizing chemotherapeutic agent that inhibits smooth muscle cell proliferation. Excipients work to promote coating stability and facilitate paclitaxel transfer and retention at the target lesion, although current excipients lack sustained, long-term paclitaxel retention. Keratose, a naturally derived protein, has exhibited unique properties allowing for tuned release of various therapeutic agents. However, little is known regarding its ability to support delivery of anti-proliferative agents such as paclitaxel. The goal of this project was to thus demonstrate the feasibility of keratose as a DCB-coating excipient to promote the release and delivery of paclitaxel.Methods: Keratose was combined with paclitaxel in vitro and the release kinetics of paclitaxel and keratose were evaluated through high performance liquid chromatograph-mass spectroscopy (HPLC-MS) and spectrophotometry, respectively. A custom coating method was developed to deposit keratose and paclitaxel on commercially available angioplasty balloons via an air spraying method. Coatings were then visualized under scanning electron microscopy and drug load quantified by HPLC-MS. Acute arterial transfer of paclitaxel at 1 h was assessed using a novel ex vivo model and further evaluated in vivo in a porcine ilio-femoral injury model.Results: Keratose demonstrated tunable release of paclitaxel as a function of keratose concentration in vitro. DCB coated via air spraying yielded consistent drug loading of 4.0 ± 0.70 μg/mm2. Under scanning electron microscopy, the keratose-paclitaxel DCB showed uniform coverage with a consistent, textured appearance. The acute drug transfer of the keratose-paclitaxel DCB was 43.60 ± 14.8 ng/mg at 1 h ex vivo. These measurements were further confirmed in vivo as the acute 1 h arterial paclitaxel levels were 56.60 ± 66.4 ng/mg.Conclusion: The keratose-paclitaxel coated DCB exhibited paclitaxel uptake and achieved acute therapeutic arterial tissue levels, confirming the feasibility of keratose as a novel excipient for DCB

Acute Thrombogenicity of a Durable Polymer Everolimus-Eluting Stent Relative to Contemporary Drug-Eluting Stents With Biodegradable Polymer Coatings Assessed Ex Vivo in a Swine Shunt Model

AbstractObjectivesThis study sought to evaluate whether the permanent fluoropolymer-coated Xience Xpedition everolimus-eluting stent (Xience-EES) exhibits lower acute thrombogenicity compared with contemporary drug-eluting stents (DES) with biodegradable polymer coatings in an acute swine shunt model.BackgroundPrevious pre-clinical and clinical experience suggests that several factors may influence the predisposition for acute thrombus formation of polymer-coated DES, including stent design and the polymer coating technology. It remains unclear whether relevant differences exist with respect to acute thrombogenicity, particularly between current commercial stent designs using permanent polymers and those using biodegradable polymers.MethodsAn ex vivo carotid to jugular arteriovenous porcine shunt model involving a test circuit of 3 in-line stents, was used to test acute thrombogenicity, where Xience-EES (n = 24) was compared with 4 CE-marked DES with biodegradable polymer coatings (BioMatrix Flex, Synergy, Nobori, and Orsiro [n = 6 each]). After 1 h of circulation, platelet aggregation in whole mount stents was evaluated by confocal microscopy with immunofluorescent staining against dual platelet markers (CD61/CD42b) along with scanning electron microscopy.ResultsXience-EES showed the least percentage of thrombus-occupied area as compared with the biodegradable polymer-coated DES, with a significant difference compared with BioMatrix Flex and Synergy (mean differences: [BioMatrix Flex: 15.54, 95% confidence interval [CI]: 11.34 to 19.75, p < 0.001; Synergy: 8.64, 95% CI: 4.43 to 12.84, p < 0.001; Nobori: 4.22, 95% CI: -0.06 to 8.49, p = 0.055; Orsiro: 2.95, 95% CI: -1.26 to 7.15, p = 0.286). The number of cell nuclei on strut surfaces was also the least in Xience-EES, with a significant difference relative to BioMatrix Flex, Nobori, and Orsiro (mean ratios: BioMatrix Flex: 4.73, 95% CI: 2.46 to 9.08, p < 0.001; Synergy: 1.44, 95% CI: 0.75 to 2.76, p = 0.51; Nobori: 5.97, 95% CI: 3.11 to 11.44, p < 0.001; Orsiro: 5.16, 95% CI: 2.69 to 9.91, p < 0.001).ConclusionsXience-EES’s overall design confers acute thromboresistance relative to contemporary DES with biodegradable coatings, with less platelet aggregation versus BioMatrix Flex and Synergy, and less inflammatory cell attachment versus BioMatrix Flex, Nobori, and Orsiro, in an ex vivo swine shunt model, which lends support to reported clinical findings of lower early stent thrombosis

Microvascular Obstruction Underlying Pathophysiology and Clinical Diagnosis

Successful restoration of epicardial coronary artery patency after prolonged occlusion might result in microvascular obstruction (MVO) and is observed both experimentally as well as clinically. In reperfused myocardium, myocytes appear edematous and swollen from osmotic overload. Endothelial cell changes usually accompany the alterations seen in myocytes but lag behind myocardial cell injury. Endothelial cells become voluminous, with large intraluminal endothelial protrusions into the vascular lumen, and together with swollen surrounding myocytes occlude capillaries. The infiltration and activation of neutrophils and platelets and the deposition of fibrin also play an important role in reperfusion-induced microvascular damage and obstruction. In addition to these ischemia-reperfusion-related events, coronary microembolization of atherosclerotic debris after percutaneous coronary intervention is responsible for a substantial part of clinically observed MVO. Microvascular flow after reperfusion is spatially and temporally complex. Regions of hyperemia, impaired vasodilatory flow reserve and very low flow coexist and these perfusion patterns vary over time as a result of reperfusion injury. The MVO first appears centrally in the infarct core extending toward the epicardium over time. Accurate detection of MVO is crucial, because it is independently associated with adverse ventricular remodeling and patient prognosis. Several techniques (coronary angiography, myocardial contrast echocardiography, cardiovascular magnetic resonance imaging, electrocardiography) measuring slightly different biological and functional parameters are used clinically and experimentally. Currently there is no consensus as to how and when MVO should be evaluated after acute myocardial infarction