Polyglycerol sebacate-based elastomeric materials for arterial regeneration.

Synthetic vascular grafts are commonly used in patients with severe occlusive arterial disease when autologous grafts are not an option. Commercially available synthetic grafts are confronted with challenging outcomes: they have a lower patency rate than autologous grafts and are currently unable to promote arterial regeneration. Polyglycerol sebacate (PGS), a non-toxic polymer with a tunable degradation profile, has shown promising results as a small-diameter vascular graft component that can support the formation of neoarteries. In this review, we first present an overview of the synthesis and modification of PGS followed by an examination of its mechanical properties. We then report on the performance, degradation, regeneration, and remodeling of PGS-based small-diameter vascular grafts, with a focus on efforts to reduce thrombosis, prevent dilation, and promote cellular residency and extracellular matrix regeneration that resembles the native artery in spatial distribution and organization. We also highlight recent advances in the incorporation of novel in situ cell sources for arterial regeneration and their potential application in PGS-based vascular grafts. Finally, we compare vascular grafts fabricated using PGS-based materials with other elastomeric alternatives.

Tropoelastin modulates systemic and local tissue responses to enhance wound healing.

Wound healing is facilitated by biomaterials-based grafts and substantially impacted by orchestrated inflammatory responses that are essential to the normal repair process. Tropoelastin (TE) based materials are known to shorten the period for wound repair but the mechanism of anti-inflammatory performance is not known. To explore this, we compared the performance of the gold standard Integra Dermal Regeneration Template (Integra), polyglycerol sebacate (PGS), and TE blended with PGS, in a murine full-thickness cutaneous wound healing study. Systemically, blending with TE favorably increased the F4/80+ macrophage population by day 7 in the spleen and contemporaneously induced elevated plasma levels of anti-inflammatory IL-10. In contrast, the PGS graft without TE prompted prolonged inflammation, as evidenced by splenomegaly and greater splenic granulocyte and monocyte fractions at day 14. Locally, the inclusion of TE in the graft led to increased anti-inflammatory M2 macrophages and CD4+T cells at the wound site, and a rise in Foxp3+ regulatory T cells in the wound bed by day 7. We conclude that the TE-incorporated skin graft delivers a pro-healing environment by modulating systemic and local tissue responses. STATEMENT OF SIGNIFICANCE: Tropoelastin (TE) has shown significant benefits in promoting the repair and regeneration of damaged human tissues. In this study, we show that TE promotes an anti-inflammatory environment that facilitates cutaneous wound healing. In a mouse model, we find that inserting a TE-containing material into a full-thickness wound results in defined, pro-healing local and systemic tissue responses. These findings advance our understanding of TE's restorative value in tissue engineering and regenerative medicine, and pave the way for clinical applications.

Universal CRISPR/Cas12a-associated aptasensor suitable for rapid detection of small proteins with a plate reader.

With the discovery of the collateral cleavage activity, CRISPR/Cas12a has recently been identified as a key enabling approach in novel DNA biosensor development. Despite its remarkable success in nucleic acid detection, realizing a universal CRISPR/Cas biosensing system for non-nucleic acid targets remains challenging, particularly at extremely high sensitivity ranges for analyte concentrations lower than the pM level. DNA aptamers can be designed to bind to a range of specific target molecules, such as proteins, small molecules, and cells, with high affinity and specificity through configuration changes. Here, by harnessing its diverse analyte-binding ability and also redirecting the specific DNA-cutting activity of Cas12a to selected aptamers, a simple, sensitive, and universal biosensing platform has been established, termed CRISPR/Cas and aptamer-mediated extra-sensitive assay (CAMERA). With simple modifications to the aptamer and guiding RNA of Cas12a RNP, CAMERA demonstrated 100 fM sensitivity for targeting small proteins, such as IFN-γ and insulin, with less than 1.5-h detection time. Compared with the gold-standard ELISA, CAMERA achieved higher sensitivity and a shorter detection time while retaining ELISA's simple setup. By replacing the antibody with an aptamer, CAMERA also achieved improved thermal stability, allowing to eliminate the requirement for cold storage. CAMERA shows potential to be used as a replacement for conventional ELISA for a variety of diagnostics but with no significant changes for the experimental setup.

Development of tropoelastin-functionalized anisotropic PCL scaffolds for musculoskeletal tissue engineering.

The highly organized extracellular matrix (ECM) of musculoskeletal tissues, encompassing tendons, ligaments and muscles, is structurally anisotropic, hierarchical and multi-compartmental. These features collectively contribute to their unique function. Previous studies have investigated the effect of tissue-engineered scaffold anisotropy on cell morphology and organization for musculoskeletal tissue repair and regeneration, but the hierarchical arrangement of ECM and compartmentalization are not typically replicated. Here, we present a method for multi-compartmental scaffold design that allows for physical mimicry of the spatial architecture of musculoskeletal tissue in regenerative medicine. This design is based on an ECM-inspired macromolecule scaffold. Polycaprolactone (PCL) scaffolds were fabricated with aligned fibers by electrospinning and mechanical stretching, and then surface-functionalized with the cell-supporting ECM protein molecule, tropoelastin (TE). TE was attached using two alternative methods that allowed for either physisorption or covalent attachment, where the latter was achieved by plasma ion immersion implantation (PIII). Aligned fibers stimulated cell elongation and improved cell alignment, in contrast to randomly oriented fibers. TE coatings bound by physisorption or covalently following 200 s PIII treatment promoted fibroblast proliferation. This represents the first cytocompatibility assessment of novel PIII-treated TE-coated PCL scaffolds. To demonstrate their versatility, these 2D anisotropic PCL scaffolds were assembled into 3D hierarchical constructs with an internally compartmentalized structure to mimic the structure of musculoskeletal tissue.

Rapid Regeneration of a Neoartery with Elastic Lamellae.

Native arteries contain a distinctive intima-media composed of organized elastin and an adventitia containing mature collagen fibrils. In contrast, implanted biodegradable small-diameter vascular grafts do not present spatially regenerated, organized elastin. The elastin-containing structures within the intima-media region encompass the elastic lamellae (EL) and internal elastic lamina (IEL) and are crucial for normal arterial function. Here, the development of a novel electrospun small-diameter vascular graft that facilitates de novo formation of a structurally appropriate elastin-containing intima-media region following implantation is described. The graft comprises a non-porous microstructure characterized by tropoelastin fibers that are embedded in a PGS matrix. After implantation in mouse abdominal aorta, the graft develops distinct cell and extracellular matrix profiles that approximate the native adventitia and intima-media by 8 weeks. Within the newly formed intima-media region there are circumferentially aligned smooth muscle cell layers that alternate with multiple EL similar to that found in the arterial wall. By 8 months, the developed adventitia region contains mature collagen fibrils and the neoartery presents a distinct IEL with thickness comparable to that in mouse abdominal aorta. It is proposed that this new class of material can generate the critically required, organized elastin needed for arterial regeneration.

Fabricating Organized Elastin in Vascular Grafts.

Surgically bypassing or replacing a severely damaged artery using a biodegradable synthetic vascular graft is a promising treatment that allows for the remodeling and regeneration of the graft to form a neoartery. Elastin-based structures, such as elastic fibers, elastic lamellae, and laminae, are key functional components in the arterial extracellular matrix. In this review, we identify the lack of elastin in vascular grafts as a key factor that prevents their long-term success. We further summarize advances in vascular tissue engineering that are focused on either de novo production of organized elastin or incorporation of elastin-based biomaterials within vascular grafts to mitigate failure and enhance enduring in vivo performance.

Applications of Engineering Techniques in Microvasculature Design.

Achieving successful microcirculation in tissue engineered constructs in vitro and in vivo remains a challenge. Engineered tissue must be vascularized in vitro for successful inosculation post-implantation to allow instantaneous perfusion. To achieve this, most engineering techniques rely on engineering channels or pores for guiding angiogenesis and capillary tube formation. However, the chosen materials should also exhibit properties resembling the native extracellular matrix (ECM) in providing mechanical and molecular cues for endothelial cells. This review addresses techniques that can be used in conjunction with matrix-mimicking materials to further advance microvasculature design. These include electrospinning, micropatterning and bioprinting. Other techniques implemented for vascularizing organoids are also considered for their potential to expand on these approaches.

CRISPR/Cas Multiplexed Biosensing: A Challenge or an Insurmountable Obstacle?

Performing multiplex detection is still an elusive goal for molecular diagnostics. CRISPR/Cas-based biosensing has demonstrated potential for multiplex detection. Instead of being an insurmountable obstacle, CRISPR/Cas multiplexed biosensing is a realistic challenge with some recent successful applications. Strategic considerations are required to fully explore its potential in multiplex diagnostics.

Signal amplification strategies for paper-based analytical devices.

Paper-based analytical devices (PADs) are very popular for point-of-care diagnostics, which provide a fast, cost-effective and possible multiplexed detection of a spectrum of molecules. They have been matching forward proudly to contribute to the modern analytical science and life science. Accompanying with their advantages and huge potentials, low detection sensitivity is continuing to challenge the application of PADs from bench to bedside. In order to improve the sensitivity and enhance the signal readout, variable signal amplification strategies have been investigated and applied for PADs. In this review, we have firstly classified formats of PADs according to the engineering design. Advances for improving sensitivity of PADs in recent five years are then summarised according to three popular types of signal amplification strategies (nanomaterial based, nucleic acid based, and engineering of PADs based). Pros and cons of each signal amplification approach have been discussed accordingly. Finally, the future perspectives of PADs are proposed.

Turn-On Fluorescence Aptasensor on Magnetic Nanobeads for Aflatoxin M1 Detection Based on an Exonuclease III-Assisted Signal Amplification Strategy.

In order to satisfy the need for sensitive detection of Aflatoxin M1 (AFM1), we constructed a simple and signal-on fluorescence aptasensor based on an autocatalytic Exonuclease III (Exo III)-assisted signal amplification strategy. In this sensor, the DNA hybridization on magnetic nanobeads could be triggered by the target AFM1, resulting in the release of a single-stranded DNA to induce an Exo III-assisted signal amplification, in which numerous G-quadruplex structures would be produced and then associated with the fluorescent dye to generate significantly amplified fluorescence signals resulting in the increased sensitivity. Under the optimized conditions, this aptasensor was able to detect AFM1 with a practical detection limit of 9.73 ng kg-1 in milk samples. Furthermore, the prepared sensor was successfully used for detection of AFM1 in the commercially available milk samples with the recovery percentages ranging from 80.13% to 108.67%. Also, the sensor performance was evaluated by the commercial immunoassay kit with satisfactory results.

Advances in biosensors for the detection of ochratoxin A: Bio-receptors, nanomaterials, and their applications.

Ochratoxin A (OTA) is a class of mycotoxin mainly produced by the genera Aspergillus and Penicillium. OTA can cause various forms of kidney, liver and brain diseases in both humans and animals although trace amount of OTA is normally present in food. Therefore, development of fast and sensitive detection technique is essential for accurate diagnosis of OTA. Currently, the most commonly used detection methods are enzyme-linked immune sorbent assays (ELISA) and chromatographic techniques. These techniques are sensitive but time consuming, and require expensive equipment, highly trained operators, as well as extensive preparation steps. These drawbacks limit their wide application in OTA detection. On the contrary, biosensors hold a great potential for OTA detection at for both research and industry because they are less expensive, rapid, sensitive, specific, simple and portable. This paper aims to provide an extensive overview on biosensors for OTA detection by highlighting the main biosensing recognition elements for OTA, the most commonly used nanomaterials for fabricating the sensing interface, and their applications in different read-out types of biosensors. Current challenges and future perspectives are discussed as well.