Laser-textured grooves extended to the sidewall edges of endovascular metallic materials to accelerate endothelialization

Abstract

The inventive concept relates to an endovascular stent. The stent includes a metal device comprising a metal or metal alloy, an outer or external surface formed by the metal or metal alloy, and a three-dimensional surface texture modification applied to or formed on the outer or external surface, wherein the surface texture modification extends to the sidewall edge of the metal device. In addition, the inventive concept includes methods of making and implanting the endovascular stent in a human body.

Description

LASER-TEXTURED GROOVES EXTENDED TO THE SIDEWALL EDGES OF ENDOVASCULAR METALLIC MATERIALS TO ACCELERATE ENDOTHELIALIZATIONCROSS REFERENCE TO RELATED APPLICATIONS

[0001] This patent application claims priority to U.S. Provisional Patent Application 63 / 709,663, filed on October 21, 2024, entitled “LASER-TEXTURED GROOVES EXTENDED TO THE SIDEWALL EDGES OF ENDOVASCULAR METALLIC MATERIALS TO ACCELERATE ENDOTHELIALIZATION”, the contents of which are incorporated herein in their entirety.Field of the Invention

[0002] The inventive concept includes a metal or metal alloy endovascular stent, such as a metal coronary stent, having a surface textured modification applied to or formed on an outer or external surface of the endovascular stent. Additionally, the inventive concept includes methods for preparing or conditioning, e.g., treating, the endovascular stent. The surface textured modification extends to the sidewall edges of the endovascular stent, which includes groove edge angles with different slopes that improve cell attachment and proliferation.Background

[0003] Over the past decade, stenting has emerged as a fundamental therapy for coronary artery disease (CAD), which continues to be the leading cause of mortality globally. As reported in the 2016 Heart Disease and Stroke Statistics by the American Heart Association (AHA), 15.5 million Americans aged 20 and above are afflicted with CAD [1], This condition is typified by the narrowing of arteries (stenosis) due to atherosclerosis, which is the accumulation of fatty deposits beneath the endothelial layer of blood vessels. In such severe cases, stent implantation is the preferred intervention to restore normal blood flow [2],

[0004] Coronary stents have become a cornerstone in the treatment of coronary heart disease, yet their long-term efficacy is often compromised by significant complications. Coronary artery stents, typically expandable mesh metallic tubes, are inserted into narrowed arteries to maintain or restore vascular patency. Initially, bare metal stents (BMS) were designed as mechanical supportsto prevent vascular elastic recoil. However, BMS are plagued by high rates of restenosis and poor endothelialization. Then coronary stents evolved to include drug-eluting capabilities to reduce neointimal hyperplasia and adverse immune responses under the name of drug-eluting stents (DES), thereby mitigating the risk of restenosis characterized by excessive tissue in-growth into the arterial lumen [3-7], Although effective in reducing restenosis, DES inhibit endothelialization leading to late thrombosis and ultimately behaving similarly to BMS post-drug release. Despite their extensive use, the long-term effectiveness of stents is frequently compromised by complications such as late in-stent thrombus and restenosis. The BMS are particularly susceptible to high restenosis rates and suboptimal endothelialization, while DES reduce restenosis but leads to incomplete endothelialization [8], leading to late thrombosis and similar issues once the drug is depleted. Notably, there is increasing evidence that stent-related thrombosis is more prevalent in DES than in BMS, especially after the discontinuation of adjunctive antiplatelet therapy [9-12], The established link between DES and delayed healing due to drug-induced hyperplasia inhibition, which hinders healthy endothelium re-establishment and increases thrombotic risk

[0013] , underlines the need for strategies that enhance endothelialization of stent surfaces. The rapid adhesion and proliferation of endothelial cells on the stent surface are critical for early vascular healing following stenting, which can significantly mitigate these major complications.

[0005] Addressing the persistent issues of late in-stent thrombus and restenosis, research has emphasized the importance of surface treatment in stent design; a focus increasingly relevant given the rising incidence of coronary artery diseases requiring stenting interventions. Surface modification of coronary stents has aimed at improving device biocompatibility. Surface properties play a pivotal role in manipulating cell growth, directly influencing the success and longevity of stent implants. Various surface engineering techniques have been employed to enhance the biological response of biomedical devices in vivo. The methods include chemical etching, plasma-based techniques, laser-based techniques, and the like. Chemical etching offers advantages like anti-bacterial properties

[0014] but may leave harmful residues, such as fluoride from hydrofluoric acid, which can damage DNA

[0015] , Whereas plasma techniques, while effective at roughening materials, require complex setups for precise freeform textures such as radiofrequency plasma

[0016] , making laser-based techniques a more promising and precise alternative.

[0006] Laser surface engineering, characterized by its precision, speed, and adaptability, holds promise for stent material treatment. This technique enables the fabrication of diverse 3D textures — such as grooves, pillars, nanowires, and porous structures — while simultaneously altering surface chemistry through processes like nitridation and oxidation. The laser-based modification of biodegradable materials can decelerate degradation rates, thereby improving stent performance by enhancing endothelialization, inhibiting smooth muscle cell proliferation, reducing platelet adhesion, and controlling corrosion and degradation. Laser texturing is an effective technique for creating grooves and complex freeform surface structures, with the unique advantage of simultaneously altering surface structure and chemistry in a rapid, precise manner. Utilizing direct laser processing, laser ablation has been used to fabricate groove-like structures on 316L stents, resulting in a threefold increase in endothelial cell adhesion and an eightfold increase in cell proliferation. Additionally, studies have found that endothelial cell proliferation and migration were significantly influenced by the geometrical dimensions of the grooves, such as width and depth [17, 18],

[0007] The size and shape of laser- induced surface textures are crucial factors influencing endothelialization on coronary stents. Various texture designs, including lines and spot grooves, have been investigated to understand their impact on endothelialization and stent performance [18-20], For instance, it has been reported

[0021] that micrometer-scale line grooves on Nitinol surfaces considerably accelerated endothelial cell migration compared to smooth surfaces, suggesting that such patterns could reduce the time required for endothelialization of vascular stents, ultimately decreasing the risk of in-stent restenosis and late-stage thrombosis

[0021] , However, research has shown that nano- to submicron-scale patterns significantly enhance cellular responses by affecting cell alignment and morphology, thereby improving cell function

[0022] , Studies indicate that the groove dimensions, specifically width and depth, critically determine the pace of endothelial cell proliferation and migration. Larger grooves have been found to improve cell alignment, while a higher concentration of ridges on the grooved surface significantly impacts cell morphology

[0023] , These findings underscore the importance of optimizing both the size and shape of laser-induced grooves to enhance the biocompatibility and performance of coronary stents.

[0008] Accordingly, surface properties play a pivotal role in manipulating cell growth, and directly influencing the success and longevity of stent implants. Despite extensive research, theoptimal surface characteristics for coronary stents remain elusive. Thus, there is a need in the art to design and develop coronary stents, and methods for their preparation and fabrication, that include laser-generated surface micro textures, material chemistry changes, endothelial cell behavior, and combinations thereof. It is important to elucidate how laser- generated microtextures and the consequent changes in material chemistry on metallic, such as cobalt chromium (CoCr), substrates affect endothelial cell behavior. Therefore, the inventive concept provides improved stents that demonstrate superior biocompatibility and clinical outcomes, as well as addressing the critical challenges of restenosis and late thrombosis.SUMMARY OF THE INVENTION

[0009] In one aspect, the inventive concept provides an endovascular stent including a metal device, which includes a metal or metal alloy; an outer or external surface formed by the metal or metal alloy; a three-dimensional, surface texture modification applied to or formed on the outer or external surface, wherein the surface texture modification extends to the sidewall edge of the metal device.

[0010] In certain embodiments, the metal or metal alloy includes cobalt-chromium.

[0011] In certain embodiments, the surface texture modification includes one or more of grooves, ridges, pillars, nanowires, and porous structures. The grooves may extend to the sidewall edge of the metal device. The grooves can have one or more groove edge angles. The groove edge angles can have different slopes. In certain embodiments, one of the slopes may be selected to improve cell attachment and proliferation on the metal device.

[0012] In certain embodiments, the surface texture modification includes a biomimetic surface.

[0013] In certain embodiments, the surface texture modification includes superior hydrophilicity, minimal contact angles, and high surface free energy.

[0014] In certain embodiments, the outer or external surface of the metal device, the chemistry of the metal device, and the sidewall edge of the metal device, are all modified by the surface texture modification.

[0015] In another aspect, the inventive concept provides a method of preparing an endovascular stent. The method includes forming a metal device including a metal or metal alloy; an outer or external surface formed by the metal or metal alloy; and a sidewall edge; applying or forming athree-dimensional, surface texture modification on the outer or external surface; and extending the surface texture modification to the sidewall edge of the metal device.

[0016] In certain embodiments of the method, the metal or metal alloy includes cobaltchromium.

[0017] In certain embodiments, the applying or forming step includes one or more of grooves, ridges, pillars, nanowires, and porous structures.

[0018] In certain embodiments, the extending step includes forming grooves in the outer or external surface that extends to the sidewall edge of the metal device. The grooves can have one or more groove edge angles. The groove edge angles can have different slopes. In certain embodiments, one of the slopes may be selected to improve cell attachment and proliferation on the metal device.BRIEF DESCRIPTION OF THE DRAWINGS

[0019] FIGS. 1A-1F show scanning electron microscope (SEM) images of CoCr patterned surface features alongside corresponding contact angle measurements as follows: (1A): grid, 5 pm, (IB): grid, 10 pm, (1C): lines, 5 pm, (ID) non-patterned substrate, (IE) stent strut with sidewall engraving, and (IF) grid-textured stent strut, in accordance with certain embodiments of the invention.

[0020] FIGS. 2A-2C show SEM images of CoCr patterned surface features that illustrate groove slope angle variations as follows: (2A) depicts grooves with a gentle slope angle of 26.5 ± 0.5°, representing the least inclined configuration; (2B) shows grooves with an intermediate slope of 45 ± 1.2°, and (2C) shows grooves with the steepest slope angle of 63.5 ± 0.25°, with the thinnest edge thickness at the groove boundary, in accordance with certain embodiments of the invention.DETAILED DESCRIPTION

[0021] The inventive concept includes endovascular stents, such as but not limited to, coronary stents in the treatment of coronary heart disease. B are-metal stents (BMS), which are known in the art, demonstrate high rates of restenosis and poor endothelialization. Thus, the inventive concept provides improved coronary stents, as well as methods for their preparation, that demonstrate low rates of restenosis and improved endothelialization.

[0022] The coronary stents according to the inventive concept are composed of metal or metal alloy, which is selected from a variety of suitable metals or metal alloys known in the art. In certain embodiments, the material composition of the coronary stents includes cobalt-chromium (CoCr).

[0023] The inventive concept includes processing, conditioning, and / or treatment of coronary stents. The process or treatment includes applying or forming a micro-hierarchical surface modification or texture, in a grid-like micro-pattern, to the (e.g., CoCr) stents that integrates sidewall edge structuring into the grid patterns on the stent surface. The sidewall structuring provides gateway channels that promote EC adhesion, proliferation, and migration. In addition, the modified stent surface positively influences endothelial cell (EC) interactions, e.g., with foreign material of the coronary stents which exhibits reduced complications typically associated with coronary stenting, without compromising mechanical integrity. Furthermore, the stent surface with modification closely mimics the arterial luminal environment.

[0024] According to the inventive concept, the surface modification includes innovative surface textures and / or patterns that introduce unique microstructures and edge modifications that enhance EC adhesion and proliferation. In certain embodiments, laser-textured surfaces according to the inventive concept exhibit superior hydrophilicity, characterized by minimal contact angles, which correlates with high surface free energy. This property enhances protein adsorption, thus promoting cell attachment. Moreover, the increased surface area provided by the textured surface and extended sidewall edges accommodates a higher number of cells, which likely underlies the observed improvements in cell adhesion and proliferation.

[0025] The surface modifications are applied post-fabrication, such that the textured features are integrated directly into the already fabricated stent mesh. In accordance with the inventive concept, the surface texture modification is applied to or formed on an outer or external surface of the stent including the extended sidewall edges of the endovascular stent. The sidewall edges provide stable anchoring sites and gateway channels for improving EC attachment and selective alignment, while substantially reducing platelet deposition in the grooved regions and maintaining essential mechanical properties. In certain embodiments, the grooves are included on the extended sidewall edges of the stent such that the edges include groove angles with different slopes. The slope is selectable to improve or optimize cell attachment and proliferation.

[0026] In certain embodiments laser fabrication is used to create microgrooves, e.g., from 5-30 pm) with extended sidewall edges that promote rapid EC adhesion and proliferation. According to the inventive concept, stents with grid pattern and sidewall edge structuring on an already fabricated stent enhances EC viability approximately six-fold compared to a non-pattemed control stent.

[0027] In certain embodiments, the surface texture includes one or more, e.g., a plurality of, grooves or ridges applied to an outer or external surface of the coronary stents. The number of grooves or ridges, as well as the depth and / or width of the grooves or ridges, is variable.Without intending to be bound by any particular theory, it is contemplated that the presence of the surface texture (as compared to a smooth surface) on the outer or external surface of the stents (including the sidewall edge), improves adhesion and proliferation of endothelial cells on the stents’ surface, thereby enhancing endothelialization of the stents’ surface, which is critical for early vascular healing following the stenting procedure.

[0028] The surface modification, e.g., texture, may be applied to the coronary stents using a variety of suitable procedures known in the art, such as but not limited to, chemical etching, plasma-based techniques, laser-based techniques, and the like. In certain embodiments, laser surface engineering is employed to apply or form the surface modification. Laser surface engineering is characterized by its precision, speed, and adaptability, and therefore is particularly suitable for stent material treatment. This technique enables the fabrication of diverse 3D textures — such as grooves, pillars, nanowires, and porous structures — while simultaneously altering the surface chemistry through processes, such as, nitridation and oxidation. The laserbased modification of biodegradable materials can decelerate degradation rates, thereby improving stents’ performance by enhancing endothelialization, inhibiting smooth muscle cell proliferation, reducing platelet adhesion, and controlling corrosion and degradation.

[0029] Laser texturing is an effective technique for creating grooves and complex freeform surface structures, with the unique advantage of simultaneously altering surface structure and chemistry in a rapid, precise manner. In certain embodiments, utilizing direct laser processing, laser ablation fabricates groove-like structures on stents, resulting in a threefold increase in endothelial cell adhesion and an eightfold increase in cell proliferation. This technique leverages the well-established principle that substrate surface properties significantly influence cell growth behaviors. As aforementioned, the rapid adhesion and proliferation of endothelial cells on thestent surface are critical for early vascular healing following stenting, which can significantly mitigate significant complications associated with stenting. Additionally, endothelial cell proliferation and migration are significantly influenced by the geometrical dimensions of the grooves, such as width and depth.

[0030] According to the inventive concept, direct laser surface texturing technology is employed to fabricate micro grid patterns on a common stent material, such as but not limited to a CoCr substrate. As aforementioned, the direct laser surface texturing technique simultaneously modifies both the surface structure of the substrate, and the chemistry of the substrate, to favor and promote endothelial cell (EC) adhesion and proliferation. Additionally, according to the inventive concept, edge structuring is developed to further promote rapid EC proliferation.Accordingly, the inventive concept includes processing or treatment of coronary stents with use of direct laser surface texturing technology that extends to the sidewall edges of the stents. Thus, this unique surface conditioning includes both modification of the substrate surface structure and chemistry modification of the substrate, as well as the edge structure of the substrate, to achieve up to a tenfold increase in EC proliferation with minimal cytotoxicity, representing a significant improvement as compared to known BMS and DES techniques.

[0031] As aforementioned, the known BMS and DES techniques result in restenosis, poor endothelialization, and inhibited endothelialization post-drug release. In contrast, the inventive concept provides enhanced performance for both BMS and DES, while preserving the beneficial features of DES and mitigating the drawbacks. Furthermore, platelet aggregation assays demonstrate that the novel grid microstructures effectively reduce platelet aggregation, highlighting the potential for laser-textured stents to improve overall biocompatibility and functionality.

[0032] In certain embodiments, the processing, conditioning or treatment of a stent with use of direct laser surface texturing technology that extends to the sidewall edges of the stent, includes the following procedure. Optionally, prior to laser processing, the stent substrate is subjected to a cleaning process using one of the various suitable cleaning processes that are known in the art. In certain embodiments, the cleaning process includes subjecting the stent to 70% ethanol in an ultrasonic bath for 10 minutes to ensure complete removal of surface contaminants. Following the cleaning process (or absent of the cleaning process), laser patterning is conducted by use of a fiber laser system wherein the stent substrate is securely positioned under a laser head of thefiber laser system for laser patterning. The system provides for precise engraving of the substrate. In certain embodiments, the laser beam, e.g., characterized by a near-Gaussian intensity distribution (e.g., beam quality factor M2 is approximately 1), is focused to a spot diameter, e.g., less than 20 pm, on the CoCr substrate. Using this technique, various groove patterns, including lines, spots, and grid textures, are created on the outer or external surface of the stent surface. Additionally, according to the inventive concept, the edge of the stents is also structured to enhance a biological response. Accordingly, the laser parameters are adjusted to achieve desired groove widths and depths.

[0033] In certain embodiments, grooves with a width of 5 microns are formed in the outer or external surface of the stent utilizing a laser power of 12.6 watts and a feed rate of 46 mm / sec. In contrast, in certain other embodiments, grooves with a width of 10 microns are formed using a laser power or 18.2 watts and a feed rate of 37 mm / sec. In certain embodiments, the parameters are fine-tuned to produce either fixed groove widths with variable depths or fixed groove depths with variable widths. Following laser processing, the samples are typically rapidly cooled to attain finer microstructures, which enhance cell adhesion by providing higher energy anchoring sites within the grooves. The precise control over the laser setting enables the production of well-defined groove patterns tailored to meet stent requirements.

[0034] In certain embodiments, following laser beam engraving, a chromium oxide layer forms on the surface of the CoCr substrate. Further, in certain embodiments, HC1 acid is used to remove the oxide layer. EDS analysis may be utilized to determine the effectiveness of the oxide removal process and how the absence of the oxide layer affects the surface chemistry.

[0035] In certain embodiments, the direct laser surface textured pattern illustrates a grooved surface on the CoCr sample, featuring parallel grooves with clear edges and uniform depth, measuring approximately 5 pm in width and spaced 20 pm apart. Alternately, the direct laser surface textured pattern illustrates grooves of 10 pm in width and spaced 40 pm apart.Optionally, a novel grid pattern is formed by intersecting grooves at right angles, creating a gridlike structure with grooves about 5 pm wide, and highlighting the complexity of the intersecting lines. Optionally, a larger grid-grooved pattern with grooves 10 pm wide and approximately 0.5 pm deep is formed. In further embodiments, a pattern of spot grooves with a diameter of 5 pm and spaced 10 pm apart is formed. All patterns may be compared to a non-patterned CoCr substrate surface that serves as a control, presenting a smooth, featureless appearance.

[0036] In certain embodiments, upon surface modification, there is a significant alteration in hydrophilicity, such as, the laser-textured samples showing a marked reduction in surface contact angles. This suggests that laser treatment is effective in producing hydrophilic surfaces.

[0037] In certain embodiments, the inventive concept includes selecting an edge structure for the stents based on its impact on endothelial cell proliferation. Without intending to be bound by any particular theory, cells exhibit a marked preference for the edges of the grooves at early culture stages; this is primarily due to the tapered structure at the groove edges, which reduces edge thickness and facilitates easier cell proliferation; at these edges, an increased ligand density at the groove ridges further enhances this effect. For example, a pattern with a minimal slope angle, likely results in lower cell attachment whereas, in contrast, patterns with steeper slopes, significantly improve cell attachment and proliferation. The engraving parameters may be adjusted to create grooves with different slopes by altering the engraving intensities. Grooves with a slope angle of 26.5 degrees represents the least inclined configuration, as compared to grooves with a slightly increased slope angle of 45 degrees, and as compared to grooves with steeper slope angle of 63.5 degrees. The grooves with the steepest slope angle of 63.5 is characterized by the thinnest edge thickness at the groove edge.

[0038] Furthermore, with respect to how different edge angles influence cell adhesion and proliferation, the 63-degree edge angle demonstrates a significantly enhanced cell adhesion response, with a 50% increase compared to the grid pattern and a substrate lacking edge grooves. This indicates that specific groove edge angles, particularly 63 degrees, substantially improves cell adhesion.

[0039] Accordingly, based on the interplay between laser-generated surface microtextures, material chemistry changes, and endothelial cell behavior on metallic substrates, e.g., CoCr, the inventive concept provides improved stents including 3D textures applied to, or formed on, an outer or exterior surface of the stents, that exhibit enhanced endothelialization, inhibition of smooth muscle cell proliferation, reduced platelet adhesion, and controlled corrosion and degradation. These improved stents represent next-generation coronary stents with superior biocompatibility and clinical outcomes, addressing the critical challenges of restenosis and late thrombosis.EXAMPLES

[0040] In the following examples, direct laser surface texturing technology was employed to fabricate defined novel micro patterns on cobalt-chromium (CoCr) coupons, a common stent material. This technique simultaneously modified both surface structure and chemistry to favor endothelial cell (EC) adhesion and proliferation, leveraging the well-established principle that substrate surface properties, significantly influence cell growth behaviors [24-26], Additionally, edge structuring was developed to further promote rapid EC proliferation, confirmed through various in vivo cell assays. This unique surface condition achieved up to a tenfold increase in EC proliferation with minimal cytotoxicity, a significant improvement compared to known bare-metal stents (BMS) and drug-eluting stents (DES), which suffer from restenosis, poor endothelialization, and inhibited endothelialization post-drug release, respectively. The inventive concept provides enhanced performance for both BMS and DES, preserving the beneficial features of DES while mitigating their drawbacks. Furthermore, platelet aggregation assays demonstrated that the microstructures effectively reduced platelet aggregation, highlighting the potential for the novel laser-textured stents to improve overall biocompatibility and functionality.Materials and MethodsPatterned cobalt chromium (CoCr) substrates fabrication

[0041] To fabricate the patterned cobalt chromium (CoCr) substrates, CoCr foil (C0120240 / 1) was obtained from Good Fellow. The foil was subsequently cut into testing samples, each with dimensions of 5 x 5 mm2and a thickness of 100 pm. Before laser processing, the samples underwent a cleaning process using 70% ethanol in an ultrasonic bath for 10 minutes to ensure complete removal of surface contaminants. For laser patterning, the samples were securely positioned under the laser head of an IPG Photonics YLR-Series fiber laser system equipped with an IPGCut Micro Cutting Head. This setup, featuring a wavelength of 1070 nm and a pulse duration of 10 nanoseconds, provided precise engraving capabilities. The laser beam, characterized by a near-Gaussian intensity distribution (beam quality factor M2~ 1), was focused to a spot diameter of less than 20 pm on the CoCr substrates. Using this technique, various groove patterns, including lines, spots, and grid textures, were created. Additionally, the edges of the samples were structured to enhance biological response.

[0042] The laser parameters were meticulously adjusted to achieve the desired groove widths and depths. For grooves with a width of 5 microns, a laser power of 12.6 watts and a feed rate of 46 mm / sec were employed. In contrast, for grooves with a width of 10 microns, a laser power of 18.2 watts and a feed rate of 37 mm / sec were utilized. These parameters were fine-tuned to produce either fixed groove widths with variable depths or fixed depths with variable widths. Following laser processing, the samples were rapidly cooled to attain finer microstructures, which enhance cell adhesion by providing higher energy anchoring sites within the grooves

[0027] , This precise control over the laser settings enabled the production of well-defined groove patterns tailored to meet the experimental requirements.

[0043] Additionally, a coronary stent with a strut thickness of 65 pm was prepared for a series of biological testing. The stent as textured with integrated sidewall inline patterns using the same laser processing technique, allowing for precise surface modifications without compromising the structural integrity. These textured stents were then used in biological assays to evaluate endothelial cell adhesion, proliferation, and overall biocompatibility, ensuring the relevance of the micro- structuring approach in a clinically applicable setting.Surface Topography Characterization

[0044] Optical microscopy (Zeiss Imager.Mlm) was initially employed to examine the surface topography of the samples, allowing for detailed observation of the effects of varying power and scanning rates on the widths and depths of features in the CoCr material. This preliminary assessment was crucial for optimizing the fabrication parameters. Subsequently, Scanning Electron Microscopy (SEM) using a FEI Apreo SEM was utilized to obtain high-resolution images of the patterned CoCr surfaces. SEM imaging was performed at an accelerating voltage of 5 kV to ensure detailed surface characterization. Prior to SEM analysis and subsequent cell culture experiments, the substrates were prepared by sputter-coating with a thin layer of gold / palladium. This process, conducted in a 50 m Torr argon environment for 30 secs at a current of 40 mA, improved sample conductivity and imaging quality.

[0045] The surface topography of the patterned CoCr substrates was comprehensively characterized using a surface profiler (Broker DektakXT stylus profilometer), which facilitated the precise measurement of substrate grooves and pattern dimensions. Following this initial profiling, Atomic Force Microscopy (AFM) was employed to achieve high-resolution imaging ofthe surface features. The AFM analysis was conducted with a Bruker Dimension Icon AFM, equipped with a NanoScope VI controller. For these measurements, Bruker AFM tips (SCANSYST-AIR) with a tip curvature radius of 2 nm were utilized in tapping mode. To ensure accurate and reproducible data, the scan rate was set at 0.5 Hz throughout the imaging process.Surface Chemistry Characterization

[0046] After laser beam engraving, a chromium oxide layer forms on the surface of the CoCr samples. This layer's presence necessitates an evaluation of its availability and impact on the rapid adhesion of endothelial cells. To this end, surface chemistry was characterized for normal patterned substrates, and patterned substrates with the oxide layer removed using Hydrochloric acid (HC1).

[0047] An Octane Elite Energy Dispersive Spectroscopy (EDS) system was employed to conduct this analysis. The EDS assessments were carried out to compare the elemental composition across the two types of substrates, focusing on detecting any variations in the oxide layer and understanding its influence on cellular interactions. For patterned substrates, the EDS provided detailed information on the surface's elemental makeup, revealing the presence and distribution of chromium oxide. In the case of patterned substrates treated with HC1 acid to remove the oxide layer, the EDS analysis was crucial in determining the effectiveness of the oxide removal process and how the absence of the oxide layer affected the surface chemistry.Hydrophilicity Characterization

[0048] The hydrophilicity of the various substrate surfaces was assessed through contact angle measurements performed using phosphate-buffered saline (PBS). A contact angle goniometer equipped with a camera setup was utilized to capture the contact angles of different texture designs, including line patterns, grid patterns, and non-patterned surfaces. The captured images were then analyzed digitally to accurately measure the contact angles, providing a comparative evaluation of the hydrophilicity across the different substrate designs.

[0049] The hydrophilicity of the various substrate surfaces was evaluated using the sessile drop method to measure the contact angle. Phosphate- buffered saline (PBS, Sigma- Aldrich, MO) was used as the liquid medium, and droplets with a precise volume of 0.8 pL ±0.1 pL were dispensed onto the substrate surfaces. Based on the droplet volume, assuming a spherical cap shape, thecalculated droplet diameter was approximately 1.17 mm. A contact angle goniometer equipped with a high-resolution camera was employed to capture the images of the droplet profile on different surface designs, including line patterns, grid patterns, and non-pattemed control surfaces. The images were digitally analyzed to determine the contact angles with high precision. This setup enabled a comparative assessment of the wettability across various textures, offering insight into how surface patterning influences hydrophilicity.Mechanical properties characterization

[0050] The mechanical properties of a thin strut CoCr coronary stent structure (Osstemcardio, Korea) were assessed to ensure structural integrity post-modification. Longitudinal bending flexibility testing was performed using a three-point bending test setup based on the ASTM F2606-08 standard for balloon-expandable vascular stents and stent delivery systems. A 5 N load cell (FLC-5P, Starrett. MA) measured applied forces, recording displacement and force data at a 10 Hz sampling rate. Bending stiffness, representing resistance to bending deformation, was calculated as a function of free bending length, deflection, and bending force according to Equation (1) below. Additionally, a radial compression test was conducted to evaluate radial strength after the engraving process, ensuring the stent’s ability to resist inward forces and maintain vessel patency.Bending stiffness (ET)-F.P 148f. Equation (1)In vitro Endothelial Cell Culture

[0051] Rat aortic endothelial cell (RAEC) attachment and proliferation on various cobalt chromium (CoCr) surface textures were comprehensively evaluated. RAECs, sourced from (Cell Biologies), were cultured on both non-pattemed and texture-patterned CoCr coupons. Initially, CoCr samples underwent sterilization via immersion in 70% ethanol for 10 minutes, followed by a 10-minute UV exposure. Each sterilized sample was then placed into an individual well of a 24-well cell culture plate, to which 500 pL of RAEC cultured in complete endothelial cell medium (Cell Biologies) suspension (5.0 x 104cells / mL) was added. The samples were incubated at standard cell culture conditions of 37 °C with 5% CO2 for either 24 or 72 hours,with the cell culture medium being replenished every 24 hours. Post-incubation, the samples were transferred to new cell culture plates and washed three times with phosphate-buffered saline (PBS; Sigma- Aldrich) for subsequent evaluation.

[0052] For Scanning Electron Microscopy (SEM) imaging, the attached cells were fixed by immersing the samples in a 2.5% glutaraldehyde (Sigma- Aldrich) solution for 2 hours. The samples were then sequentially dehydrated using 30%. 50%. 75%, and 100% ethanol, followed by hexamethyldisilazane (Fisher Scientific). After overnight drying in a vacuum, the surfaces of the samples were coated with gold / palladium to facilitate imaging at an accelerating voltage of 5 kV.In vitro Endothelial cell Viability and Proliferation assays

[0053] Cell adhesion assays were initially conducted over a short period, where (RAECs) are seeded at a density of (105cells / 100 pL) in Complete Endothelial Cell Medium onto the patterned Cobalt Chromium (CoCr) samples. The cells were incubated for 4 hours under standard cell culture conditions. Post-incubation, the samples were rinsed three times with (PBS) to remove non-adherent cells, preparing them for cell viability analysis. Additionally, longer cell density assays were performed over 1 and 3 days, wherein RAECs were seeded at a density of (5.0 x 104cells / 100 pL) in Complete Endothelial Cell Medium onto the CoCr samples. The culture medium was refreshed only once, after the first day. At the conclusion of days 1 and 3, non-adherent cells were removed by rinsing the samples three times with PBS to prepare them for cell viability analysis.

[0054] To quantify the number of RAECs on the CoCr samples, the CellTiter 96® AQueous One Solution Cell Proliferation Assay (MTS; Promega) was utilized. Briefly, after culturing the cells on the patterned samples, the supernatant was removed on both day 1 and day 3 with culture medium replenishment after the first day. The samples were then transferred to fresh cell culture plates. Subsequently, 500 pL of the fresh Complete Medium was added onto the samples. Thereafter, 75 pL of CellTiter 96® AQueous One Solution Reagent was pipetted into the wells containing samples with complete medium. Samples with MTS reagent were incubated for 2 hours at the standard conditions before transferring the aliquots from each sample’s well to 96-well plates. Absorbance was recorded at 490 nm using a (Thermo Scientific Multiskan FC) 96-well plate reader to determine the number of RAECs on each sample (n = 3). The cell densities at4 hours, 1 , and 3 days were calculated by referencing the MTS assay's standard curve, which correlates absorbance readings to specific cell densities.

[0055] Live / Dead staining was also performed to assess cell orientation and distribution on the patterned samples and to validate the MTS assay results. Briefly, after incubating the patterned CoCr coupons for 1 and 3 days at the standard conditions, with medium replenished every 24 hours, the samples were transferred to fresh cell culture plates and washed three times with PBS to remove unattached RAECs. Cell viability was determined in situ using the LIVE / DEAD Cell Viability Kit from AAT Bioquest. The stain working solution is prepared by adding 20 pl of CytoCalcein solution and 20 pl of Propidium Iodide into 10 ml of Assay Buffer, followed by a 30-minute incubation at the standard conditions before visualization using fluorescence microscopy. ImageJ software was utilized to enhance the visualization of viable cells, providing improved contrast between the fluorescently stained cells and the underlying groove pattern shapes. This approach ensured accurate assessment of cell distribution and alignment relative to the surface textures.In vitro Platelet aggregation assay

[0056] To evaluate the platelet deposition over the predesigned patterned Cobalt Chromium (CoCr) coupons, whole ovine blood was collected with sodium citrate (0.106 M) via jugular venipuncture and deposited over the samples. The procedures adhered to National Institutes of Health (NIH) guidelines for the care and use of laboratory animals and were approved by the Institutional Animal Care and Use Committee (IACUC) at the University of Pittsburgh. Briefly, non-patterned and various patterned CoCr coupons were prepared as previously described. The samples were sterilized by immersion in 70% ethanol for 10 minutes, followed by exposure to ultraviolet (UV) light for an additional 10 minutes. Each sample was then placed into no-additive (Z) tubes (BD Vacutainers®, Becton, Dickinson and Co., Franklin, NI, USA) containing 5 mL of fresh ovine blood. The samples were incubated for 3 hours at 37 °C with gentle rocking. Postincubation, non-adherent platelets were removed by rinsing the samples five times with PBS.

[0057] The platelets deposition on each sample was analyzed using both scanning electron microscopy (SEM) and a lactate dehydrogenase (LDH) assay. For SEM imaging, the adhered platelets were fixed by immersion in a 2.5% glutaraldehyde solution for 2 hours. Following fixation, the platelets were dehydrated using a graded ethanol series (30%, 50%, 75%, and100%) and treated with hexamethyldisilazane. The samples were then sputter-coated with gold / palladium prior to imaging at 5 kV (n = 3). For quantification via the LDH assay, the adhered platelets were lysed by immersion in 1 mL of 2% Triton X-100 in PBS, followed by stirring for 20 minutes. The lysis solution was centrifuged at 250 g for 10 minutes, and 100 pL of the supernatant was collected and reacted with 100 pL of LDH reagent. The absorbance of the reacted solution was measured at 490 nm, with 650 nm as the reference, using a microplate reader (n = 3). This absorbance data was used to calculate the number of platelets adhered to each sample.Statistics

[0058] For each experimental condition, a minimum of three independent tests were conducted to ensure the reliability of the results. The data obtained from these experiments are presented as mean ± standard deviation (SD). Statistical comparisons between groups were performed using Student’s t-test to determine the significance of the observed differences. A p-value of less than 0.05 was considered indicative of a statistically significant difference.Micro pattern fabrication and characterization

[0059] The surface topographies of the CoCr samples were analyzed using Scanning Electron Microscopy (SEM), revealing distinct morphological features from various laser patterning processes (FIGs. 1A -IF). As shown in FIGS. 1A and IB, the grid patterns illustrate a grooved surface on the CoCr sample. FIG. 1A includes groove widths measuring approximately 5 pm and FIG. IB has grooves measuring approximately 10 pm in width. FIGS. 1A and IB exhibited contact angles of 8.9 ± 0.74° and 12.2 ± 1.04°, respectively, indicating enhanced hydrophilicity with narrower grooves. This improvement could be attributed to increased surface area, more contact points, and capillary action facilitated by intersecting grooves. In contrast, FIG. 1C illustrates linear grooves with a 5 pm width. FIG. 1C showed a higher contact angle of 22.2 ± 0.32°, reflecting reduced wettability due to the absence of intersecting features. FIG. ID illustrates a flat, non-patterned CoCr surface. FIG. ID demonstrated the lowest wettability, with a contact angle of 58.8 ± 0.48°. Thus, FIGS. 1A-1D show a decrease in contact angle with increasing pattern complexity enhancing endothelial cell (EC) adhesion and proliferation . To translate these findings into a clinically relevant model, CoCr coronary stents with strutdimension of 65 pirn were engraved with grid grooves (10-20 pm wide) integrated with sidewall edge texturing, linking the inner and outer strut faces as illustrated in FIG. IE. These complex intersections, clearly visible in FIG. IF, are designed to maximize surface wettability and create gateway channels to support EC attachment and proliferation in subsequent biological assays.

[0060] The topographical characterization of the CoCr patterned surface features showed the precise and uniform nature of the grooves, essential for influencing cell behavior; atomic force microscope (AFM) images confirmed the regularity and consistency of the micro-patterned grid, which is crucial for ensuring reproducibility in cell adhesion and viability studies.

[0061] The mechanical behavior of the grid-textured stent was assessed and compared to the control stent using bending and crushing (radial compression) tests. The grid-textured stent demonstrated higher bending stiffness, lower flexibility, than the control during the first halfway of the test, with the difference diminishing as deflection increased, suggesting comparable flexibility while preserving structural integrity. Crushing test results showed that both stents exhibited similar crushing loads, with the grid-textured stent maintaining sufficient radial strength to endure physiological pressures. Notably, initial deviations in both bending and crushing tests may be attributed to strain hardening induced by the laser engraving process. This effect could be mitigated through post-engraving heat treatment to restore material ductility. Overall, the findings confirm that laser-induced grid microtexturing preserves mechanical robustness while enhancing flexibility, ensuring the stent’s suitability for vascular deployment.

[0062] To complete the surface characterization analysis, it is essential to evaluate the influence of the oxide layer typically formed during laser texturing. A detailed comparison of the surface composition of CoCr substrates was conducted, examining patterned surfaces both with and without the oxide layer. The effect of surface chemistry on behavior showed rat aortic endothelial cell (RAEC) adhesion and viability on CoCr grid patterns using the MTS assay over multiple culture durations. The results revealed a consistent trend: substrates with an oxide layer exhibited slightly higher cell densities compared to those without. Although the oxide layer contributed to a modest enhancement in RAEC adhesion and proliferation across all time points, this improvement was not statistically significant. These findings suggest that the influence of microgroove topographies on cellular responses can be more accurately assessed when the confounding effects of the oxide layer are minimized, allowing for a clearer understanding of the role of surface patterning in promoting endothelialization.Groove Pattern’s geometry integrated with sidewall edges and endothelialization relationship

[0063] Micro-groove parameters, specifically width and depth significantly impacted the adhesion of rat aortic endothelial cells (RAECs) on laser micrometer grid patterns of CoCr surfaces across various culture durations. The effect of micro-groove width (5 pm, 10 pm, 20 pm, and 30 pm) with a constant groove depth of 0.5 pm on RAEC adhesion was tested. Cell adhesion was measured at 4 h, one day, and three days, with minimal differences among the various depths. The data indicated a substantial variation in cell adhesion across different groove widths. Cell adhesion increased as groove size decreased. Specifically, the 5 pm (15337 ± 492 cells / ml) and 10 pm (12998 ± 712 cells / ml) grooves showed the highest adhesion levels at the three-day time point, compared to the 20 pm (12611 + 585 cells / ml) grooves. The groove showed a notable rise in cell adhesion, indicating optimal conditions for cell attachment and proliferation. The impact of micro-groove depth (0.5 pm, 1 pm, and 1.5 pm) on RAEC adhesion with a constant groove width of 10 pm was tested. Cell adhesion was similarly measured at 4 h, one day, and three days, with minimal differences among the various depths. Specifically, at 0.5 pm (13136 ± 636 cells / ml) and 1 pm (12998 + 824 cells / ml), adhesion showed a slight but statistically insignificant increase for the 0.5 pm depth compared to 1 pm.

[0064] The comparative analysis of different groove dimensions revealed that the optimal selection of both width and depth for the interconnected micro-hierarchical groove array significantly enhances surface energy, thus promoting a higher degree of hydrophilicity. This increase in hydrophilicity, attributed to the finer hierarchical structures at the micro-scale, enhanced the surface’s ability to interact with biological fluids. The improved wetting behavior was facilitated by the increased contact area between the surface and PBS, ultimately supporting more favorable conditions for endothelial cell adhesion and proliferation.

[0065] MTS assay results revealed a substantial increase in cell density on laser-patterned micrometer grids compared to various designs, including parallel, spot, and non-pattemed CoCr surfaces, across multiple culture durations (4 h, one day, and three days). Notably, grids with a width of 5 pm and a depth of 0.5 pm demonstrated a cell density of (15,337 ± 492 cells / ml) on the third day, nearly tenfold that of non-patterned surfaces, which recorded (1590 + 361 cells / ml). These grid patterns, characterized by a groove width of 5 pm and a depth of 0.5 pm, consistently outperformed other topographies, with the most significant enhancements observed after one and three days of culture.

[0066] Moreover, it was observed that RAEC viability enhancement due to the grid pattern could be further amplified by carefully tuning the sidewall edge texture angle. FIGS. 2A-2C illustrate the groove slope angle variations through SEM imaging, showing how engraving parameters were adjusted to create grooves with different slopes by altering engraving intensities. FIG. 2A depicts grooves with a gentle slope angle of 26.5 ± 0.5°, representing the least inclined configuration. FIG. 2B shows grooves that were formed with an intermediate slope of 45 ± 1.2°. FIG. 2C shows grooves that were formed with the steepest slope angle of 63.5 + 0.25°, characterized by the thinnest edge thickness at the groove boundary. These variations in sidewall slope influence the available surface area and microenvironment at the groove edges, which likely affects cell attachment and proliferation. Grooves with steeper slopes may offer enhanced focal adhesion points, further promoting RAEC viability and highlighting the importance of microstructural refinement in optimizing surface- guided endothelialization.

[0067] The viability responses from the MTS assay, showcased the influence of groove edge angles on rat aortic endothelial cell (RAEC) adhesion and proliferation over 4 h, 1 day, and 3 days. The results revealed that sharper edge angles significantly enhanced adhesion, with the 63° edge angle demonstrating a 50 % increase in cell density compared to both standard grid patterns and substrates lacking edge grooves. This underscores the critical role of groove geometry in optimizing surface modifications for improved cellular responses. The laser-induced sidewall edge structuring on CoCr substrates markedly impacted endothelial cell (EC) adhesion by generating high-energy regions along the prominent groove edges. These thin, steep sidewalls facilitated rapid endothelialization, as evidenced by the uniform distribution of viable RAECs across various edge angles within the first 4 h of culture. A direct correlation between increasing sidewall angles and enhanced EC adhesion was observed, highlighting the pivotal role of edge geometry in accelerating endothelialization. To further amplify this effect, a continuous sidewall structure with a fixed 63° angle was engineered along the substrate edges, leading to a substantial improvement in EC adhesion; cell density increased significantly from (5689.33 + 452 cells / mL) to (20,957 ± 715 cells / mL) within one day of culture.

[0068] Further, live / dead staining assays provided a visual confirmation of cell viability and distribution across the CoCr-pattemed and non-pattemed substrates; rat aortic endothelial cell (RAEC) adhesion and viability after 4 h, 1 day, and 3 days of culture was evaluated. Grid-patterned surfaces with 5 pm grooves consistently exhibited the highest density of viable cells,particularly at groove intersections, with attachment ratios of —4:1 and 3:1 compared to nonpatterned areas. In contrast, parallel line patterns showed reduced cell adhesion, indicating less favorable conditions. After one day, grids maintained superior RAEC density, significantly outperforming other patterns, while by day three, cells not only covered the grid but also extended into adjacent non-patterned regions, reflecting sustained proliferation. Parallel lines supported moderate cell growth, whereas non-patterned surfaces consistently displayed the lowest viability and the highest number of dead cells. Across all patterned surfaces, cell death remained minimal, underscoring the grid pattern’s effectiveness in promoting early adhesion, viability, and long-term proliferation.

[0069] Live / Dead assay on the 63° inclined groove’s edges revealed that RAECs began early attachment predominantly at the groove locations, indicating these regions as favorable anchoring sites that facilitated initial adhesion and migration. This observation was further validated by examining a fully textured edge where cells displayed enhanced attachment along the structured edge. This strategic adhesion suggests a higher propensity for cells to migrate smoothly and efficiently from the edge, promoting a faster and more uniform coverage of the strut area.

[0070] Extensive investigation using SEM was conducted to provide a full picture of the cell adhesion and proliferation. The grid pattern after one day of culture revealed a uniform distribution of cells adhering to the surface grooves. A long-term viability assessment of the grid pattern after five days showed cells proliferated extensively, forming a dense and interconnected network that integrated well with the micro-patterned substrate. In contrast, a non-pattemed surface after three days highlighted sporadic and uneven cell attachment indicating suboptimal conditions for cell growth.Influence on endothelialization with the micro-textured thin strut stents

[0071] Grid-sidewall edges-integrated microtextures were applied directly onto thin strut CoCr coronary stents. The testing evaluated whether the enhanced endothelial cell adhesion and proliferation observed on flat substrates could be replicated on complex stent geometries, which present additional challenges such as curved surfaces, variable wall thickness and intricate strut architectures. MTS data demonstrated a significant increase in RAEC adhesion on grid-textured stents compared to non-pattemed control stents over time. After three days of culture, the grid-textured stents exhibited an 81 % more increase in cell density, confirming the sustained benefits of microtextured features in promoting endothelialization.

[0072] In the control stent, minimal cell attachment was observed, with sparse RAECs adhering randomly along the strut surface. In contrast, the grid-textured stent showed extensive cell coverage, particularly concentrated along the microgroove intersections and edges. These results underscored the critical role of microtopography in facilitating cell attachment, even on the complex three-dimensional surfaces of stents. Early attachment at 8 hours revealed cells initially favoring intersections and sidewalls, where microgrooves provide mechanical cues for possible anchorage. By 24 hours, cells began to bridge gaps between struts, particularly along intersections that facilitated access to the stent’s inner wall. After three days, more extensive cell coverage was evident, with endothelial cells spreading along the strut surfaces and initiating coverage of the inner luminal areas.

[0073] Live / Dead fluorescence images corresponding to the same time points showed preferential attachment of live cells along groove intersections and sidewalls, while dead cells remained minimal, indicating high cell viability throughout the culture period. By 72 h, RAECs migrated along the strut surfaces and began colonizing the inner walls, suggesting that the microtextured patterns not only promote initial attachment but also facilitate directional cell migration critical for complete stent endothelialization.

[0074] The results indicated that sidewalls of the microtextured patterns could serve as a key possible anchoring points, guiding endothelial cell attachment and facilitating migration along the strut surfaces; the vertical surfaces enhance initial cell adhesion by providing additional contact areas and create a topographical pathway that directs cells toward the inner luminal walls of the stent; this strategic alignment encourages efficient cell coverage across the complex stent geometry, significantly advancing beyond the capabilities observed on flat CoCr coupons.Hemocompatibility: platelet deposition on patterned CoCr substrates

[0075] After laser grooving, platelet deposition assays were performed to evaluate the hemocompatibility of the modified CoCr surfaces. Laser-induced micro structuring can significantly alter surface characteristics, potentially impacting platelet deposition, which is critical in determining the thrombogenic risk of biomaterials . Assessing platelet deposition provides valuable insights into the material’s interaction with blood components following laserexposure. Quantitative and imaging analyses of platelet deposition on CoCr samples were performed. Lactate dehydrogenase (LDH) assay results compared the number of deposited platelets on control, line-patterned, and grid-patterned surfaces. The control samples exhibited the highest level of platelet deposition, while the grid patterns showed the least, since the platelet deposition was decreased on the groove regions. There was dense deposition and aggregation of platelets on the control sample, while the platelet deposition showed a significant reduction on the grooved region of the grid-patterned sample.Discussion

[0076] The impact of micro-hierarchical grid grooving on CoCr surfaces was evaluated. The engineered grooves, ranging from 5 to 10 pm in width, featured precisely angled sidewall edges that enhanced surface energy and optimized interactions with endothelial cells (ECs). These sidewalls, serving as proximal anchoring points to surrounding tissue, established stable adhesion sites, promoting rapid endothelialization. The microtextured channels, with the optimized geometry, appeared to foster enhanced EC proliferation and adhesion. Further in vitro results from the grid-textured stents exhibited substantial endothelial coverage, particularly along the groove sidewalls. These regions facilitated cell migration across the stent surface, suggesting that such surface modifications have the potential to accelerate endothelialization and improve overall stent integration.

[0077] Both the groove’s size and shape are crucial factors influencing endothelialization on coronary stents. The innovative surface patterns developed in this study, along with engineered sidewall edge structure modifications, revealed an ideal micro-scale surface design for enhancing EC adhesion and proliferation.

[0078] The results of the MTS and Live / Dead assays revealed an outstanding enhancement in cell viability facilitated by the innovative interconnected grid design. After a few hours of culture, the Live RAEC images showed a notable adhesion started at the edges, proliferating over the micro-texture locations. This distinct proliferation and attachment could be attributed to the increased surface area and roughness associated with these patterns has led to a higher ligand density, creating more attachment points for cells. Notably, samples with narrower groove widths and higher hydrophilicity demonstrated superior expression of EC -related proteins, effectively supporting EC adhesion, and proliferation. For a longer time of culture (24-h), thegrid design exhibited a cell density of (12,604 ± 502 cells / ml), a remarkable 4-fold increase compared to the non-pattemed control (2775 ± 145 cells / ml). This substantial improvement reflects the effectiveness of the grid’s structural features in promoting early-stage cell adhesion.

[0079] A more significant impact was observed upon optimizing the sidewall structure, where the maximum achievable angle of 63° led to a substantial increase in endothelial cell (EC) adhesion. The cell density reached (20,957 ± 715 cells / ml) after just one day of culture, representing more than a six-fold increase over non-pattemed samples. This pronounced enhancement was attributed to the continuous structuring of the substrate sidewalls, which facilitated consistently elevated rates of RAEC adhesion as early as 4 h into the culture. SEM imaging enabled high-resolution visualization of EC morphology, allowing direct observation of cell spreading, attachment, and interaction with the microstructured surfaces. Complementarity, cell viability assays quantitatively confirmed minimal cytotoxicity, reflecting preserved cell function proliferation.

[0080] In addition to these results, further analysis revealed a marked reduction in platelet deposition, particularly in the number of deposited platelets to the groove locations on the grid. The narrow grooves and reduced interspaces of the grid pattern appeared to play a key role in minimizing platelet aggregation by limiting available adhesion sites.Conclusion

[0081] The foregoing examples studied cardiovascular biomaterials through precision laser-engineered micro-patterns and sidewall edge structuring on CoCr substrates. The interconnected grid design, with optimized groove dimensions and tailored sidewalls, enhanced endothelial cell adhesion and proliferation by providing stable endothelial adhesion points that promoted rapid cell attachment and uniform coverage. These micro-hierarchical features also reduced platelet aggregation, emphasizing the role of groove geometry and surface wettability in minimizing thrombogenic risks. In vitro mechanical testing confirmed that the grid-patterned stents maintained essential radial strength and flexibility, while biological assays demonstrated a sixfold increase in endothelial cell proliferation with minimal cytotoxicity. Enhanced cell attachment was observed along the sidewalls and groove intersections, supporting improved endothelialization.References:[1] Mozaffarian, D., Benjamin, E. J., Go, A. S., Arnett, D. K., Blaha, M. J., Cushman, M., ... & Turner, M. B. (2016). Heart disease and stroke statistics — 2016 update: a report from the American Heart Association, circulation, 133(4), e38-e360.[2] Serruys, P. W., De Jaegere, P., Kiemeneij, F., Macaya, C., Rutsch, W., Heyndrickx, G., ... & Morel, M. A. (1994). A comparison of balloon-expandable- stent implantation with balloon angioplasty in patients with coronary artery disease. New England Journal of Medicine, 331(8), 489-495.[3] Drachman DE, Edelman ER, Seifert P, et al. Neointimal thickening after stent delivery of paclitaxel: change in composition and arrest of growth over six months. J Am Coll Cardiol 2000;36:2325-32.[4] Klugherz BD, Llanos G, Lieuallen W, et al. Twenty-eight-day efficacy and phamacokinetics of the sirolimus-eluting stent. Coron Artery Dis 2002;13:183-8.[5] Suzuki T, Kopia G, Hayashi S, et al. Stent-based delivery of sirolimus reduces neointimal formation in a porcine coronary model. Circulation 2001;104:1188-93.[6] Farb A, Heller PF, Shroff S, et al. Pathological analysis of local delivery of paclitaxel via a polymer-coated stent. Circulation 200E104: 473-9.[7] Finn AV, Kolodgie FD, Hamek J, et al. Differential response of delayed healing and persistent inflammation at sites of overlapping sirolimus- or paclitaxel-eluting stents. Circulation 2005;112:270-8.[8] Pendyala LK, Yin X, Li J, Chen JP, Chronos N, Hou D. The first-generation drugeluting stents and coronary endothelial dysfunction. JACC Cardiovasc Interv 2009;2:1169-1177.[9] Joner M, Finn AV, Farb A, Mont EK, Kolodgie FD, Ladisch E, et al. Pathology of drugeluting stents in humans: delayed healing and late thrombotic risk. J Am Coll Cardiol 2005:48(1): 193-202.

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Claims

CLAIMSWe claim:

1. An endovascular stent, comprising:a metal device, comprising:a metal or metal alloy;an outer or external surface formed by the metal or metal alloy;a three-dimensional, surface texture modification applied to or formed on the outer or external surface,wherein the surface texture modification extends to the sidewall edge of the metal device.

2. The endovascular stent of claim 1, wherein the metal or metal alloy comprises cobaltchromium.

3. The endovascular stent of claim 1, wherein the surface texture modification comprises one or more of grooves, ridges, pillars, nanowires, and porous structures.

4. The endovascular stent of claim 1, wherein the grooves extend to the sidewall edge of the metal device.

5. The endovascular stent of claim 4, wherein the grooves have one or more groove edge angles.

6. The endovascular stent of claim 5, wherein the groove edge angles have different slopes.

7. The endovascular stent of claim 6, wherein the one of the different slopes is selected to improve cell attachment and proliferation on the metal device.

8. The endovascular stent of claim 1, wherein the surface texture modification comprises a biomimetic surface.

9. The endovascular stent of claim 1 , wherein the surface texture modification comprises superior hydrophilicity, minimal contact angles and high surface free energy.

10. The endovascular stent of claim 1, wherein the outer or external surface of the metal device, the chemistry of the metal device, and the sidewall edge of the metal device are all modified by the surface texture modification.

11. A method of preparing an endovascular stent, comprising:forming a metal device comprising:a metal or metal alloy;an outer or external surface formed by the metal or metal alloy; and a sidewall edge;applying or forming a three-dimensional, surface texture modification on the outer or external surface; andextending the surface texture modification to the sidewall edge of the metal device.

12. The method of claim 11, wherein the metal or metal alloy comprises cobalt-chromium.

13. The method of claim 11, wherein the applying or forming step comprises one or more of grooves, ridges, pillars, nanowires, and porous structures.

14. The method of claim 11, wherein the extending step comprises forming grooves in the outer or external surface that extends to the sidewall edge of the metal device.

15. The method of claim 14, wherein the grooves have one or more groove edge angles.

16. The method of claim 15, wherein the groove edge angles have different slopes.

17. The method of claim 16, wherein the one of the different slopes is selected to improve cell attachment and proliferation on the metal device.

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