Developmental heterogeneity of vascular cells: Insights into cellular plasticity in atherosclerosis?

Smooth muscle cells, endothelial cells and macrophages display remarkable heterogeneity within the healthy vasculature and under pathological conditions. During development, these cells arise from numerous embryological origins, which confound with different microenvironments to generate postnatal vascular cell diversity. In the atherosclerotic plaque milieu, all these cell types exhibit astonishing plasticity, generating a variety of plaque burdening or plaque stabilizing phenotypes. And yet how developmental origin influences intraplaque cell plasticity remains largely unexplored despite evidence suggesting this may be the case. Uncovering the diversity and plasticity of vascular cells is being revolutionized by unbiased single cell whole transcriptome analysis techniques that will likely continue to pave the way for therapeutic research. Cellular plasticity is only just emerging as a target for future therapeutics, and uncovering how intraplaque plasticity differs across vascular beds may provide key insights into why different plaques behave differently and may confer different risks of subsequent cardiovascular events.

Colchicine promotes atherosclerotic plaque stability independently of inflammation.

Atherosclerosis is a chronic inflammatory disease which is driven in part by the aberrant trans -differentiation of vascular smooth muscle cells (SMCs). No therapeutic drug has been shown to reverse detrimental SMC-derived cell phenotypes into protective phenotypes, a hypothesized enabler of plaque regression and improved patient outcome. Herein, we describe a novel function of colchicine in the beneficial modulation of SMC-derived cell phenotype, independent of its conventional anti-inflammatory effects. Using SMC fate mapping in an advanced atherosclerotic lesion model, colchicine induced plaque regression by converting pathogenic SMC-derived macrophage-like and osteoblast-like cells into protective myofibroblast-like cells which thickened, and thereby stabilized, the fibrous cap. This was dependent on Notch3 signaling in SMC-derived plaque cells. These findings may help explain the success of colchicine in clinical trials relative to other anti-inflammatory drugs. Thus, we demonstrate the potential of regulating SMC phenotype in advanced plaque regression through Notch3 signaling, in addition to the canonical anti-inflammatory actions of drugs to treat atherosclerosis.

Sustainable Nanomaterials for Biomedical Applications.

Significant progress in nanotechnology has enormously contributed to the design and development of innovative products that have transformed societal challenges related to energy, information technology, the environment, and health. A large portion of the nanomaterials developed for such applications is currently highly dependent on energy-intensive manufacturing processes and non-renewable resources. In addition, there is a considerable lag between the rapid growth in the innovation/discovery of such unsustainable nanomaterials and their effects on the environment, human health, and climate in the long term. Therefore, there is an urgent need to design nanomaterials sustainably using renewable and natural resources with minimal impact on society. Integrating sustainability with nanotechnology can support the manufacturing of sustainable nanomaterials with optimized performance. This short review discusses challenges and a framework for designing high-performance sustainable nanomaterials. We briefly summarize the recent advances in producing sustainable nanomaterials from sustainable and natural resources and their use for various biomedical applications such as biosensing, bioimaging, drug delivery, and tissue engineering. Additionally, we provide future perspectives into the design guidelines for fabricating high-performance sustainable nanomaterials for medical applications.

Tribological and Rheological Properties of Poly(vinyl alcohol)-Gellan Gum Composite Hydrogels.

Polymeric poly(vinyl alcohol) (PVA)-based composite hydrogels are promising materials with various biomedical applications. However, their mechanical and tribological properties should be tailored for such applications. In this study, we report the fabrication of PVA-gellan gum (GG) composite hydrogels and determine the effect of GG content on their rheological and tribological properties. The rheology tests revealed an enhanced storage (elastic) modulus with increased gellan gum (GG) concentration. The results showed up to 89% enhancement of the elastic modulus of PVA by adding 0.5 wt% gellan gum. This elastic modulus (12.1 ± 0.8 kPa) was very close to that of chondrocyte and its surrounding pericellular matrix (12 ± 1 kPa), rendering them ideal for cartilage regeneration applications. Furthermore, the friction coefficient was reduced by up to 80% by adding GG to PVA, demonstrating the increased elastic modulus improved chance of survival under mechanical shear stresses. Examining PVA/GG at different concentrations of 0.1, 0.3, and 0.5 wt% of GG, we demonstrate that at a load of 5 N, the friction coefficient decreases by increasing the GG concentration. However, at higher loads of 10 and 15 N, a 0.3 wt% concentration was sufficient to significantly reduce the friction coefficient. For PVA and PVA/GG composites, we observed a reduction in friction coefficient by increasing the load from 5 to 15 N. We also found the friction to be independent of the sliding velocity. Possible mechanisms of achieving a reduced friction coefficient are discussed.

Recent Advancements in the Fabrication of Functional Nanoporous Materials and Their Biomedical Applications.

Functional nanoporous materials are categorized as an important class of nanostructured materials because of their tunable porosity and pore geometry (size, shape, and distribution) and their unique chemical and physical properties as compared with other nanostructures and bulk counterparts. Progress in developing a broad spectrum of nanoporous materials has accelerated their use for extensive applications in catalysis, sensing, separation, and environmental, energy, and biomedical areas. The purpose of this review is to provide recent advances in synthesis strategies for designing ordered or hierarchical nanoporous materials of tunable porosity and complex architectures. Furthermore, we briefly highlight working principles, potential pitfalls, experimental challenges, and limitations associated with nanoporous material fabrication strategies. Finally, we give a forward look at how digitally controlled additive manufacturing may overcome existing obstacles to guide the design and development of next-generation nanoporous materials with predefined properties for industrial manufacturing and applications.

Stereolithographic Visible-Light Printing of Poly(l-glutamic acid) Hydrogel Scaffolds.

Bioprinting is a promising fabrication technique aimed at developing biologically functional, tissue-like constructs for various biomedical applications. Among the different bioprinting approaches, vat polymerization-based techniques offer the highest feature resolution compared to more commonly used extrusion-based methods and therefore have greater potential to be utilized for printing complex hierarchical tissue architectures. Although significant efforts have been directed toward harnessing digital light processing techniques for high-resolution bioprinting, the use of stereolithography (SLA) setups for producing distinct hydrogel filaments smaller than 20 µm has received less attention. Improving the bioprinting resolution is still a technical challenge that must consider both the practical limitations of the bioprinter apparatus and the formulation of the cytocompatible bioresin. In this study, we developed a novel bioresin compatible with SLA and capable of printing high-resolution features. This resin, composed of a biosynthetic polypeptide poly(l-glutamic acid) functionalized with tyramine moieties (PLGA-Tyr), was crosslinked using a visible-light photoinitiator system. Varying concentrations of PLGA-Tyr and the co-photoinitiator were evaluated for the hydrogel system's gelation ability, swelling characteristics, degradation profiles, mechanical properties, and cell viability post-encapsulation. This study introduces a custom-built, cost-effective, visible-light SLA bioprinting system named the "MicroNC". Using the newly developed visible-light bioresin, we demonstrated for the first time the ability to fabricate hydrogel scaffolds with well-resolved filaments (less than 8 µm in width) capable of supporting cell viability and proliferation and directing cellular morphology at the single-cell level for up to 14 days. Overall, these experiments have underscored the exciting potential of using the visible-light-photoinitiated PLGA-Tyr material system for developing physiologically relevant in vitro hydrogel scaffolds with feature resolutions comparable to the dimensions of individual human cells for a wide range of biomedical applications.

Modified N-linked glycosylation status predicts trafficking defective human Piezo1 channel mutations.

Mechanosensitive channels are integral membrane proteins that sense mechanical stimuli. Like most plasma membrane ion channel proteins they must pass through biosynthetic quality control in the endoplasmic reticulum that results in them reaching their destination at the plasma membrane. Here we show that N-linked glycosylation of two highly conserved asparagine residues in the 'cap' region of mechanosensitive Piezo1 channels are necessary for the mature protein to reach the plasma membrane. Both mutation of these asparagines (N2294Q/N2331Q) and treatment with an enzyme that hydrolyses N-linked oligosaccharides (PNGaseF) eliminates the fully glycosylated mature Piezo1 protein. The N-glycans in the cap are a pre-requisite for N-glycosylation in the 'propeller' regions, which are present in loops that are essential for mechanotransduction. Importantly, trafficking-defective Piezo1 variants linked to generalized lymphatic dysplasia and bicuspid aortic valve display reduced fully N-glycosylated Piezo1 protein. Thus the N-linked glycosylation status in vitro correlates with efficient membrane trafficking and will aid in determining the functional impact of Piezo1 variants of unknown significance.

Mural Cells: Potential Therapeutic Targets to Bridge Cardiovascular Disease and Neurodegeneration.

Mural cells collectively refer to the smooth muscle cells and pericytes of the vasculature. This heterogenous population of cells play a crucial role in the regulation of blood pressure, distribution, and the structural integrity of the vascular wall. As such, dysfunction of mural cells can lead to the pathogenesis and progression of a number of diseases pertaining to the vascular system. Cardiovascular diseases, particularly atherosclerosis, are perhaps the most well-described mural cell-centric case. For instance, atherosclerotic plaques are most often described as being composed of a proliferative smooth muscle cap accompanied by a necrotic core. More recently, the role of dysfunctional mural cells in neurodegenerative diseases, such as Alzheimer's and Parkinson's disease, is being recognized. In this review, we begin with an exploration of the mechanisms underlying atherosclerosis and neurodegenerative diseases, such as mural cell plasticity. Next, we highlight a selection of signaling pathways (PDGF, Notch and inflammatory signaling) that are conserved across both diseases. We propose that conserved mural cell signaling mechanisms can be exploited for the identification or development of dual-pronged therapeutics that impart both cardio- and neuroprotective qualities.

Tuneable manganese oxide nanoparticle based theranostic agents for potential diagnosis and drug delivery.

Among various magnetic nanoparticles, manganese oxide nanoparticles are considered as established T 1 magnetic resonance imaging (MRI) contrast agents for preclinical research. The implications of their degradation properties and use as therapeutic carriers in drug delivery systems have not been explored. In addition, how the chemical composition and size of manganese oxide nanoparticles, as well as the surrounding environment, influence their degradation and MRI contrast properties (T 1 vs. T 2) have not been studied in great detail. A fundamental understanding of their characteristic properties, such as degradation, is highly desirable for developing simultaneous diagnosis and therapeutic solutions. Here, we demonstrate how the precursor type and reaction environment affect the size and chemical composition of manganese oxide nanoparticles and evaluate their influence on the nanoparticle degradability and release of the drug l-3,4-dihydroxyphenylalanine (l-dopa). The results show that the degradation rate (and the associated release of drug l-dopa molecules) of manganese oxide nanoparticles depends on their size, composition and the surrounding environment (aqueous or biometric fluid). The dependence of MRI relaxivities of manganese oxide nanoparticles on the size, chemical composition and nanoparticle degradation in water is also established. A preliminary cell viability study reveals the cytocompatible properties of l-dopa functionalized manganese oxide nanoparticles. Overall, this work provides new insights into smartly designed manganese oxide nanoparticles with multitasking capabilities to target bioimaging and therapeutic applications.

Nature-inspired topographies on hydroxyapatite surfaces regulate stem cells behaviour.

Surface topography is one of the key factors in regulating interactions between materials and cells. While topographies presented to cells in vivo are non-symmetrical and in complex shapes, current fabrication techniques are limited to replicate these complex geometries. In this study, we developed a microcasting technique and successfully produced imprinted hydroxyapatite (HAp) surfaces with nature-inspired (honeycomb, pillars, and isolated islands) topographies. The in vitro biological performance of the developed non-symmetrical topographies was evaluated using adipose-derived stem cells (ADSCs). We demonstrated that ADSCs cultured on all HAp surfaces, except honeycomb patterns, presented well-defined stress fibers and expressed focal adhesion protein (paxillin) molecules. Isolated islands topographies significantly promoted osteogenic differentiation of ADSCs with increased alkaline phosphatase activity and upregulation of key osteogenic markers, compared to the other topographies and the control unmodified (flat) HAp surface. In contrast, honeycomb topographies hampered the ability of the ADSCs to proliferate and differentiate to the osteogenic lineage. This work presents a facile technique to imprint nature-derived topographies on the surface of bioceramics which opens up opportunities for the development of bioresponsive interfaces in tissue engineering and regenerative medicine.

Baghdadite Ceramics Prevent Senescence in Human Osteoblasts and Promote Bone Regeneration in Aged Rats.

Bone fractures and critical-sized bone defects present significant health threats for the elderly who have limited capacity for regeneration due to the presence of functionally compromised senescent cells. A wide range of synthetic materials has been developed to promote the regeneration of critical-sized bone defects, but it is largely unknown if a synthetic biomaterial (scaffold) can modulate cellular senescence and improve bone regeneration in aged scenarios. The current study investigates the interaction of Baghdadite (Ca3ZrSi2O9) ceramic scaffolds with senescent human primary osteoblast-like cells (HOBs) and its bone regeneration capacity in aged rats. A senescent HOB model was established by repeatedly passaging HOBs till passage 7 (P7). Compared to the clinically used hydroxyapatite/tricalcium phosphate (HA/TCP), Baghdadite prevented senescence induction in P7 HOBs and markedly negated the paracrine effect of P7 HOB secretomes that mediated the up-regulations of cellular senescence-associated gene expression levels in P2 HOBs. We further demonstrated that conditioned media extracted from Baghdadite corrected the dysfunctional mitochondria in P7 HOBs. In vivo, the bone regeneration capacity was enhanced when 3D printed Baghdadite scaffolds were implanted in a calvaria critical-sized bone defect model in both young and aged rats compared to HA/TCP scaffolds, but a better effect was observed in aged rats than in young rats. This study suggests that Baghdadite ceramic represents a novel and promising biomaterial approach to promote bone regeneration capacity in the elderly by providing an anti-senescent microenvironment.

Visible light mediated PVA-tyramine hydrogels for covalent incorporation and tailorable release of functional growth factors.

The translation of growth factors (GFs) into clinical applications is limited by their low stability in physiological environments. Controlled GF delivery through biomaterial vehicles provides protection from proteases, targeted delivery, and longer term release profiles. However, current methods used to incorporate GFs into biomaterials still present limitations. While direct adsorption and encapsulation result in burst release, covalent incorporation provides a tailorable release profile but generally requires more complicated processes and chemical modification of the GFs. Bioaffinity methods provide long-term release profiles but fail in their specificity, resulting in GF-dependent applicability and release profiles. In the present study, we introduce tyraminated poly-vinyl-alcohol (PVA-Tyr) as a GF-delivery vehicle that can covalently incorporate native GFs through a photo-initiated cross-linking process via formation of bi-phenol bonds. Mass loss and release studies revealed that protein-loaded PVA-Tyr hydrogels had highly tailorable degradation times from 7 to 92 days, during which the covalently incorporated proteins were released in a linear fashion. The incorporation of bovine serum albumin (BSA), vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), or brain-derived growth factor (BDNF) resulted in similar incorporation efficiencies and release profiles, demonstrating the low specificity and versatility of the system. Furthermore, functional studies demonstrated that VEGF, bFGF and BDNF released from the PVA-Tyr hydrogels retained the ability to increase the metabolic activity, migration, and 3D vessel formation of endothelial cells and mesenchymal stem cells. Taken together, this demonstrates that PVA-Tyr shows high potential as a highly tailorable GF delivery tool for a range of different regenerative medicine applications.

Mechanically stressed cancer microenvironment: Role in pancreatic cancer progression.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal solid malignancies in the world due to its insensitivity to current therapies and its propensity to metastases from the primary tumor mass. This is largely attributed to its complex microenvironment composed of unique stromal cell populations and extracellular matrix (ECM). The recruitment and activation of these cell populations cause an increase in deposition of ECM components, which highly influences the behavior of malignant cells through disrupted forms of signaling. As PDAC progresses from premalignant lesion to invasive carcinoma, this dynamic landscape shields the mass from immune defenses and cytotoxic intervention. This microenvironment influences an invasive cell phenotype through altered forms of mechanical signaling, capable of enacting biochemical changes within cells through activated mechanotransduction pathways. The effects of altered mechanical cues on malignant cell mechanotransduction have long remained enigmatic, particularly in PDAC, whose microenvironment significantly changes over time. A more complete and thorough understanding of PDAC's physical surroundings (microenvironment), mechanosensing proteins, and mechanical properties may help in identifying novel mechanisms that influence disease progression, and thus, provide new potential therapeutic targets.

Small Molecule KRAS Inhibitors: The Future for Targeted Pancreatic Cancer Therapy?

Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest solid tumors in the world. Currently, there are no approved targeted therapies for PDAC. Mutations in Kirsten rat sarcoma viral oncogene homologue (KRAS) are known to be a major driver of PDAC progression, but it was considered an undruggable target until recently. Moreover, PDAC also suffers from drug delivery issues due to the highly fibrotic tumor microenvironment. In this perspective, we provide an overview of recent developments in targeting mutant KRAS and strategies to overcome drug delivery issues (e.g., nanoparticle delivery). Overall, we propose that the antitumor effects from novel KRAS inhibitors along with strategies to overcome drug delivery issues could be a new therapeutic way forward in PDAC.

Two-Photon Dual-Emissive Carbon Dot-Based Probe: Deep-Tissue Imaging and Ultrasensitive Sensing of Intracellular Ferric Ions.

Carbon dots (CDs)-based nanoparticles have been extensively explored for biological applications in sensing and bioimaging. However, the major translational barriers to CDs for imaging and sensing applications include synthetic strategies to obtain monodisperse CDs with tunable structural, electronic, and optical properties in order to achieve high-resolution deep-tissue imaging, intracellular detection, and sensing of metal ions with high sensitivity down to nanomolar levels. Herein, we report a novel strategy to synthesize and develop a multifunctional nitrogen-doped CDs probe of different sizes using a new combination of carbon and nitrogen sources. Our results show that the structural characteristics (i.e., the surface density of emissive traps and bandgaps levels) depend on the size of the CDs, which ultimately influences their optical properties. This work also demonstrates the development of a two-photon dual-emissive fluorescent multifunctional probes (3-FCDs) by conjugating fluorescein isothiocyanate on the surface of nitrogen-doped CDs. 3-FCDs show excellent near-infrared two-photon excitation ability, single-wavelength excitation, high photostability, biocompatibility, low cytotoxicity, and good cell permeability. Using two-photon fluorescence imaging, our multifunctional probe shows excellent deep-tissue high-resolution imaging capabilities with penetration depth up to 3000 and 280 µm in hydrogel scaffold and pigskin tissue, respectively. The designed probe exhibits ultrasensitivity and specificity toward Fe3+ ions with a remarkable detection limit of 2.21 nM using two-photon excitation. In addition, we also demonstrate the use of multifunctional CDs probe for ultrasensitive exogenous and real-time endogenous sensing of Fe3+ ions and imaging in live fibroblasts with rapid response times for intracellular ferric ion detection.

Reprogramming of human fibroblasts into osteoblasts by insulin-like growth factor-binding protein 7.

The induced pluripotent stem cell (iPSC) is a promising cell source for tissue regeneration. However, the therapeutic value of iPSC technology is limited due to the complexity of induction protocols and potential risks of teratoma formation. A trans-differentiation approach employing natural factors may allow better control over reprogramming and improved safety. We report here a novel approach to drive trans-differentiation of human fibroblasts into functional osteoblasts using insulin-like growth factor binding protein 7 (IGFBP7). We initially determined that media conditioned by human osteoblasts can induce reprogramming of human fibroblasts to functional osteoblasts. Proteomic analysis identified IGFBP7 as being significantly elevated in media conditioned with osteoblasts compared with those with fibroblasts. Recombinant IGFBP7 induced a phenotypic switch from fibroblasts to osteoblasts. The switch was associated with senescence and dependent on autocrine IL-6 signaling. Our study supports a novel strategy for regenerating bone by using IGFBP7 to trans-differentiate fibroblasts to osteoblasts.

High-Strength Fiber-Reinforced Composite Hydrogel Scaffolds as Biosynthetic Tendon Graft Material.

The development of suitable synthetic scaffolds for use as human tendon grafts to repair tendon ruptures remains a significant engineering challenge. Previous synthetic tendon grafts have demonstrated suboptimal tissue ingrowth and synovitis due to wear particles from fiber-to-fiber abrasion. In this study, we present a novel fiber-reinforced hydrogel (FRH) that mimics the hierarchical structure of the native human tendon for synthetic tendon graft material. Ultrahigh molecular weight polyethylene (UHMWPE) fibers were impregnated with either biosynthetic polyvinyl alcohol/gelatin hydrogel (FRH-PG) or with polyvinyl alcohol/gelatin + strontium-hardystonite (Sr-Ca2ZnSi2O7, Sr-HT) composite hydrogel (FRH-PGS). The scaffolds were fabricated and assessed to evaluate their suitability for tendon graft applications. The microstructure of both FRH-PG and FRH-PGS showed successful impregnation of the hydrogel component, and the tendon scaffolds exhibited equilibrium water content of ∼70 wt %, similar to the values reported for native human tendon, compared to ∼50 wt % water content retained in unmodified UHMWPE fibers. The tensile strength of FRH-PG and FRH-PGS (77.0-81.8 MPa) matched the range of human Achilles' tendon tensile strengths reported in the literature. In vitro culture of rat tendon stem cells showed cell and tissue infiltration into both FRH-PG and FRH-PGS after 2 weeks, and the presence of Sr-HT ceramic particles influenced the expression of tenogenic markers. On the other hand, FRH-PG supported the proliferation of murine C2C12 myoblasts, whereas FRH-PGS seemingly did not support it under static culture conditions. In vivo implantation of FRH-PG and FRH-PGS scaffolds into full-thickness rat patellar tendon defects showed good collagenous tissue ingrowth into these scaffolds after 6 weeks. This study demonstrates the potential viability for our FRH-PG and FRH-PGS scaffolds to be used for off-the-shelf biosynthetic tendon graft material.

Role of Biomaterials and Controlled Architecture on Tendon/Ligament Repair and Regeneration.

Engineering synthetic scaffolds to repair and regenerate ruptured native tendon and ligament (T/L) tissues is a significant engineering challenge due to the need to satisfy both the unique biological and biomechanical properties of these tissues. Long-term clinical outcomes of synthetic scaffolds relying solely on high uniaxial tensile strength are poor with high rates of implant rupture and synovitis. Ideal biomaterials for T/L repair and regeneration need to possess the appropriate biological and biomechanical properties necessary for the successful repair and regeneration of ruptured tendon and ligament tissues.

Novel injectable strontium-hardystonite phosphate cement for cancellous bone filling applications.

Injectable bone cement (IBC) such as those based on methacrylates and hydraulic calcium phosphate and calcium sulfate-based cements have been used extensively for filling bone defects with acceptable clinical outcomes. There is a need however for novel IBC materials that can address some of the inherent limitations of currently available formulations to widen the clinical application of IBC. In this study, we characterized a novel hydraulic IBC formulation consisting of bioactive strontium-doped hardystonite (Sr-HT) ceramic microparticles and sodium dihydrogen phosphate, herein named Sr-HT phosphate cement (SPC). The resultant cement is comprised of two distinct amorphous phases with embedded partially reacted crystalline reactants. The novel SPC formulation possesses a unique combination of physicochemical properties suitable for use as an IBC, and demonstrates in vitro cytocompatibility when seeded with primary human osteoblasts. In vivo injection of SPC into rabbit sinus defects show minor new bone formation at the SPC periphery, similar to those exhibited in sinus defects filled with a clinically available calcium phosphate cement. The current SPC formulation presented in this paper shows promise as a clinically applicable IBC which can be further enhanced with additives.

Promoting Cell Survival and Proliferation in Degradable Poly(vinyl alcohol)-Tyramine Hydrogels.

A photopolymerizable-tyraminated poly(vinyl alcohol) (PVA-Tyr) system that has the ability to covalently bind proteins in their native state was evaluated as a platform for cell encapsulation. However, a key hurdle to this system is the radicals generated during the cross-linking that can cause oxidative stress to the cells. This research hypothesized that incorporation of anti-oxidative proteins (sericin and gelatin) into PVA-Tyr gels would mitigate any toxicity caused by the radicals. The results showed that although incorporation of 1 wt% sericin promoted survival of the fibroblasts, both sericin and gelatin acted synergistically to facilitate long-term 3D cell function. The encapsulated cells formed clusters with deposition of laminin and collagen, as well as remaining metabolically active after 21 d.