Plant-derived extracellular vesicles: a novel nanomedicine approach with advantages and challenges.

BACKGROUND: Many eukaryote cells produce membrane-enclosed extracellular vesicles (EVs) to establish cell-to-cell communication. Plant-derived EVs (P-EVs) contain proteins, RNAs, lipids, and other metabolites that can be isolated from the juice, the flesh, and roots of many species. METHODS: In the present review study, we studied numerous articles over the past two decades published on the role of P-EVs in plant physiology as well as on the application of these vesicles in different diseases. RESULTS: Different types of EVs have been identified in plants that have multiple functions including reorganization of cell structure, development, facilitating crosstalk between plants and fungi, plant immunity, defense against pathogens. Purified from several edible species, these EVs are more biocompatible, biodegradable, and extremely available from many plants, making them useful for cell-free therapy. Emerging evidence of clinical and preclinical studies suggest that P-EVs have numerous benefits over conventional synthetic carriers, opening novel frontiers for the novel drug-delivery system. Exciting new opportunities, including designing drug-loaded P-EVs to improve the drug-delivery systems, are already being examined, however clinical translation of P-EVs-based therapies faces challenges. CONCLUSION: P-EVs hold great promise for clinical application in the treatment of different diseases. In addition, despite enthusiastic results, further scrutiny should focus on unravelling the detailed mechanism behind P-EVs biogenesis and trafficking as well as their therapeutic applications. Video Abstract.

Unraveling the Effect of Breast Cancer Patients' Plasma on the Targeting Ability of Folic Acid-Modified Chitosan Nanoparticles.

The formation of protein corona (PC) around nanoparticles (NPs) has been reported inside biological conditions. This effect can alter delivery capacity toward the targeted tissues. Here, we synthesized folic acid-modified chitosan NPs (FA-CS NPs) using different concentrations of folic acid (5, 10, and 20%). FA-CS NPs were exposed to plasmas of breast cancer patients and healthy donors to evaluate the possibility of PC formation. We also monitored uptake efficiency in in vitro conditions after incubation with human breast cancer cell line MDA-MB-231 and monocyte/macrophage-like Raw264.7 cells. Data showed that the formation of PC around FA-CS NPs can change physicochemical properties coincided with the rise in NP size and negative surface charge. SDS-PAGE electrophoresis revealed differences in the type and content rate of plasma proteins attached to NP surface in a personalized manner. Based on MTT data, the formation of PC around NPs did not exert cytotoxic effects on MDA-MB-231 cells while this phenomenon reduced uptake rate. Fluorescence imaging and flow cytometry analyses revealed reduced cellular internalization rate in NPs exposed to patients' plasma compared to the control group. In contrast to breast MDA-MB-231 cells, Raw264.7 cells efficiently adsorbed the bare and PC-coated NPs from both sources, indicating the involvement of ligand-receptor-dependent and independent cellular engulfment. These data showed that the PC formed on the FA-CS NPs is entirely different in breast cancer patients and healthy counterparts. PC derived from patients' plasma almost abolishes the targeting efficiency of FA-CS NPs even in different mechanisms, while this behavior was not shown in the control group. Surprisingly, Raw264.7 cells strongly adsorbed the PC-coated NPs, especially when these particles were in the presence of patients' sera. It is strongly suggested that the formation of PC around can affect delivering capacity of FA-CS NPs to cancer cells. It seems that the PC-coated FA-CS NPs can be used as an efficient delivery strategy for the transfer of specific biomolecules in immune system disorders.

Fabrication and Optimization of Bilayered Nanoporous Anodic Alumina Structures as Multi-Point Interferometric Sensing Platform.

Herein, we present an innovative strategy for optimizing hierarchical structures of nanoporous anodic alumina (NAA) to advance their optical sensing performance toward multi-analyte biosensing. This approach is based on the fabrication of multilayered NAA and the formation of differential effective medium of their structure by controlling three fabrication parameters (i.e., anodization steps, anodization time, and pore widening time). The rationale of the proposed concept is that interferometric bilayered NAA (BL-NAA), which features two layers of different pore diameters, can provide distinct reflectometric interference spectroscopy (RIfS) signatures for each layer within the NAA structure and can therefore potentially be used for multi-point biosensing. This paper presents the structural fabrication of layered NAA structures, and the optimization and evaluation of their RIfS optical sensing performance through changes in the effective optical thickness (EOT) using quercetin as a model molecule. The bilayered or funnel-like NAA structures were designed with the aim of characterizing the sensitivity of both layers of quercetin molecules using RIfS and exploring the potential of these photonic structures, featuring different pore diameters, for simultaneous size-exclusion and multi-analyte optical biosensing. The sensing performance of the prepared NAA platforms was examined by real-time screening of binding reactions between human serum albumin (HSA)-modified NAA (i.e., sensing element) and quercetin (i.e., analyte). BL-NAAs display a complex optical interference spectrum, which can be resolved by fast Fourier transform (FFT) to monitor the EOT changes, where three distinctive peaks were revealed corresponding to the top, bottom, and total layer within the BL-NAA structures. The spectral shifts of these three characteristic peaks were used as sensing signals to monitor the binding events in each NAA pore in real-time upon exposure to different concentrations of quercetin. The multi-point sensing performance of BL-NAAs was determined for each pore layer, with an average sensitivity and low limit of detection of 600 nm (mg mL-1)-1 and 0.14 mg mL-1, respectively. BL-NAAs photonic structures have the capability to be used as platforms for multi-point RIfS sensing of biomolecules that can be further extended for simultaneous size-exclusion separation and multi-analyte sensing using these bilayered nanostructures.

Assessment of Binding Affinity between Drugs and Human Serum Albumin Using Nanoporous Anodic Alumina Photonic Crystals.

In this study, we report an innovative approach aiming to assess the binding affinity between drug molecules and human serum albumin by combining nanoporous anodic alumina rugate filters (NAA-RFs) modified with human serum albumin (HSA) and reflectometric interference spectroscopy (RIfS). NAA-RFs are photonic crystal structures produced by sinusoidal pulse anodization of aluminum that present two characteristic optical parameters, the characteristic reflection peak (λPeak), and the effective optical thickness of the film (OTeff), which can be readily used as sensing parameters. A design of experiments strategy and an ANOVA analysis are used to establish the effect of the anodization parameters (i.e., anodization period and anodization offset) on the sensitivity of HSA-modified NAA-RFs toward indomethacin, a model drug. To this end, two sensing parameters are used, that is, shifts in the characteristic reflection peak (ΔλPeak) and changes in the effective optical thickness of the film (ΔOTeff). Subsequently, optimized NAA-RFs are used as sensing platforms to determine the binding affinity between a set of drugs (i.e., indomethacin, coumarin, sulfadymethoxine, warfarin, and salicylic acid) and HSA molecules. Our results verify that the combination of HSA-modified NAA-RFs with RIfS can be used as a portable, low-cost, and simple system for establishing the binding affinity between drugs and plasma proteins, which is a critical factor to develop efficient medicines for treating a broad range of diseases and medical conditions.

Label-Free real-time quantification of enzyme levels by interferometric spectroscopy combined with gelatin-modified nanoporous anodic alumina photonic films.

Herein, we present an interferometric sensor based on the combination of chemically functionalized nanoporous anodic alumina photonic films (NAA-PFs) and reflectometric interference spectroscopy (RIfS) aimed to detect trace levels of enzymes by selective digestion of gelatin. The fabrication and sensing performance of the proposed sensor were characterized in real-time by estimating the changes in effective optical thickness (i.e., sensing principle) of gelatin-modified NAA-PFs (i.e., sensing element) during enzymatic digestion. The working range (WR), sensitivity (S), low limit of detection (LLoD), and linearity (R(2)) of this enzymatic sensor were established by a series of experiments with different concentrations of gelatin (i.e., specific chemical sensing element) and trypsin (i.e., analyte), a model protease enzyme with relevant implications as a biomarker in the diagnosis of several diseases. The chemical selectivity of the sensor was demonstrated by comparison of gelatin digestion by other nonspecific enzyme models such as chymotrypsin and horseradish peroxidase. Furthermore, the role of the chemical sensing element (i.e., gelatin) was assessed by using hemoglobin instead of gelatin. Finally, we demonstrated that this sensor can be readily used to establish the kinetic parameters of enzymatic reactions. The obtained results revealed that the presented sensor has a promising potential to be used as a point-of-care system for fast detection of gastrointestinal diseases at early stages.

Discovery of a Novel Dual Targeting Peptide for Human Glioma: From In Silico Simulation to Acting as Targeting Ligand.

Purpose: Receptor-mediated transcytosis (RMT) is a more specific, highly efficient, and reliable approach to crossing the blood-brain-barrier (BBB) and releasing the therapeutic cargos into the brain parenchyma. Methods: Here, we introduced and characterized a human/mouse-specific novel leptin-derived peptide using in silico, in vitro and in vivo experiments. Results: Based on the bioinformatics analysis and molecular dynamics (MD) simulation, a 14 amino acid peptide sequence (LDP 14) was introduced and its interaction with leptin-receptor (ObR) was analyzed in comparison with an well known leptin-derived peptide, Lep 30. MD simulation data revealed a significant stable interaction between ligand binding domains (LBD) of ObR with LDP 14. Analyses demonstrated suitable cellular uptake of LDP 14 alone and its derivatives (LDP 14-modified G4 PAMAM dendrimer and LDP 14-modified G4 PAMAM/pEGFP-N1 plasmid complexes) via ObR, energy and species dependent manner (preferred uptake by human/mouse cell lines compared to rat cell line). Importantly, our findings illustrated that the entry of LDP 14-modified dendrimers in hBCEC-D3 cells not only is not affected by protein corona (PC) formation, as the main reason for diminishing the cellular uptake, but also PC per se can enhance uptake rate. Finally, fluorescein labeled LDP 14-modified G4 PAMAM dendrimers efficiently accumulated in the mice brain with lower biodistribution in other organs, in our in vivo study. Conclusion: LDP 14 introduced as a novel and highly efficient ligand, which can be used for drugs/genes delivery to brain tissue in different central nervous system (CNS) disorders.

Docosahexaenoic Acid Reduced Vascular Endothelial Cell Injury in Diabetic Rats Via the Modulation of Autophagy.

Purpose: Among varied ω-3 polyunsaturated fatty acid types, the therapeutic properties of docosahexaenoic acid (DHA) have been indicated under diabetic conditions in different cell lineages. Here, we investigated the anti-diabetic properties of DHA in rats with type 2 diabetes mellitus (D2M) focusing on autophagy-controlling factors. Methods: D2M was induced in male Wistar rats using a single dose of streptozocin (STZ) and a high-fat diet for 8 weeks. On week 2, diabetic rats received DHA 950 mg/kg/d until the end of the study. After that, rats were euthanized, and aortic and cardiac tissue samples were stained with H&E staining for histological assessment. The expression of adhesion molecules, ICAM-1 and VCAM-1, was measured in heart samples using real-time PCR analysis. Using western blotting, protein levels of BCLN1, LC3, and P62 were measured in D2M rats pre- and post-DHA treatment. Results: Data showed intracellular lipid vacuoles inside the vascular cells, and cardiomyocytes, after induction of D2M and DHA reduced intracellular lipid droplets and in situ inflammatory response. DHA can diminish increased levels of ICAM-1 in diabetic conditions (P Control vs. D2M rats=0.005) and reach near-to-control values (P Control vs. D2M rats=0.28; P D2M rats vs. D2M rats+DHA=0.033). Based on western blotting, D2M slightly increased the BCLN1 and LC3-II/I ratio without affecting P62. DHA promoted the LC3II/I ratio (P=0.303) and reduced P62 (P Control vs. D2M rats+DHA =0.0433; P D2M vs. D2M rats+DHA=0.096), leading to the completion of autophagy flux under diabetic conditions. Conclusion: DHA can reduce lipotoxicity of cardiovascular cells possibly via the activation of adaptive autophagy response in D2D rats.

Exosomes; multifaceted nanoplatform for targeting brain cancers.

At the moment, anaplastic changes within the brain are challenging due to the complexity of neural tissue, leading to the inefficiency of therapeutic protocols. The existence of a cellular interface, namely the blood-brain barrier (BBB), restricts the entry of several macromolecules and therapeutic agents into the brain. To date, several nano-based platforms have been used in laboratory settings and in vivo conditions to overcome the barrier properties of BBB. Exosomes (Exos) are one-of-a-kind of extracellular vesicles with specific cargo to modulate cell bioactivities in a paracrine manner. Regarding unique physicochemical properties and easy access to various biofluids, Exos provide a favorable platform for drug delivery and therapeutic purposes. Emerging data have indicated that Exos enable brain penetration of selective cargos such as bioactive factors and chemotherapeutic compounds. Along with these statements, the application of smart delivery approaches can increase delivery efficiency and thus therapeutic outcomes. Here, we highlighted the recent advances in the application of Exos in the context of brain tumors.

Tumor Cells-derived exosomal CircRNAs: Novel cancer drivers, molecular mechanisms, and clinical opportunities.

Recently, circular RNAs (circRNAs) have appealed to a growing interest due to their abundant expression and potential functions in cancer development. The most biological function of circRNAs may include acting as a sponge for miRNAs and proteins in different physio/pathological conditions. CircRNAs promote cancer progression by regulating several procedures such as growth, invasion, metastasis, angiogenesis, and drug resistance. Emerging evidence has shown that circRNAs frequently have tumor-specific expression, proposing these molecules serve as diagnostic and prognostic cancer biomarkers. Furthermore, circRNAs may be used as a potential target for the treatment of cancers as they can sponge oncogenic miRNAs and proteins. Exosomes, a subtype of extracellular vesicles mediate intercellular communication, contain circRNAs and deliver them to target cells inducing cancer development through different signaling pathways. Exosomal circRNAs may serve as a diagnostic and prognostic biomarker for cancers. Targeting exosomes may represent novel approaches for the treatment of cancers through using them as cell-free therapy and drug-delivery system and inhibiting their biogenesis and distribution. However, research on circRNAs biology is advancing and some concerns such as technical issues in the characterization and analysis of circRNAs along with biological understanding gaps necessary to be considered to transfer this undeveloped field to the vanguard of clinical studies. In this review, we discuss the existing information on the formation of circRNA and its roles in the tumor as a biomarker and treatment target. Furthermore, we describe tumor-derived exosomes enclosed circRNAs and their possible roles in cancer development and their potential as biomarkers and therapeutic approaches.

Recent Progress in Nanotechnology for COVID-19 Prevention, Diagnostics and Treatment.

The COVID-19 pandemic is currently an unprecedented public health threat. The rapid spread of infections has led to calls for alternative approaches to combat the virus. Nanotechnology is taking root against SARS-CoV-2 through prevention, diagnostics and treatment of infections. In light of the escalating demand for managing the pandemic, a comprehensive review that highlights the role of nanomaterials in the response to the pandemic is highly desirable. This review article comprehensively discusses the use of nanotechnology for COVID-19 based on three main categories: prevention, diagnostics and treatment. We first highlight the use of various nanomaterials including metal nanoparticles, carbon-based nanoparticles and magnetic nanoparticles for COVID-19. We critically review the benefits of nanomaterials along with their applications in personal protective equipment, vaccine development, diagnostic device fabrication and therapeutic approaches. The remaining key challenges and future directions of nanomaterials for COVID-19 are briefly discussed. This review is very informative and helpful in providing guidance for developing nanomaterial-based products to fight against COVID-19.

Anticancer Effect of Cisplatin-Loaded Poly (Butylcyanoacrylate) Nanoparticles on A172 Brain Cancer Cells Line

Background: Drug delivery systems have been designed to achieve targeted delivery and control the release rate of the drugs. A serious challenge associated with drug delivery systems is the presence of the blood-brain barrier which limits drugs penetration. In the current study, the effects of cisplatin nanoparticles on A172 brain cancer cell line were investigated. Methods: Cisplatin nanoparticles were produced by miniemulsion polymerization technique and their properties were evaluated. Drug release assay was performed to characterize the nanoparticles' properties. Here, we examined the effects of cisplatin nanoparticles and free form of cisplatin on A172 cancer cell line. MTT assay was performed for different concentrations of the drug. To measure the apoptosis rate in A172 cell line in the presence of cisplatin nanoparticles or its free from, Annexin V staining method was used. Results: Our results indicated that loading type of cisplatin was physical loading and only 4.7% of cisplatin was released after 68 h. Furthermore, MTT assay showed that cisplatin nanoparticles in all concentrations had more cytotoxic effects on the cells comparing with the free form of cisplatin and control groups. We also showed that cisplatin nanoparticles could increase apoptosis in cancer cells more than the drug in the free form by using flow cytometry technique. Conclusion: Overall, these findings proved that cisplatin loaded on poly (Butylcyanoacrylate) nanoparticles, was more efficient than the free form of cisplatin in treating A172 cancer cell line.

Structural Engineering of Nanoporous Anodic Alumina Photonic Crystals by Sawtooth-like Pulse Anodization.

This study presents a sawtooth-like pulse anodization approach aiming to create a new type of photonic crystal structure based on nanoporous anodic alumina. This nanofabrication approach enables the engineering of the effective medium of nanoporous anodic alumina in a sawtooth-like manner with precision. The manipulation of various anodization parameters such as anodization period, anodization amplitude, number of anodization pulses, ramp ratio and pore widening time allows a precise control and fine-tuning of the optical properties (i.e., characteristic transmission peaks and interferometric colors) exhibited by nanoporous anodic alumina photonic crystals (NAA-PCs). The effect of these anodization parameters on the photonic properties of NAA-PCs is systematically evaluated for the establishment of a fabrication methodology toward NAA-PCs with tunable optical properties. The effective medium of the resulting NAA-PCs is demonstrated to be optimal for the development of optical sensing platforms in combination with reflectometric interference spectroscopy (RIfS). This application is demonstrated by monitoring in real-time the formation of monolayers of thiol molecules (11-mercaptoundecanoic acid) on the surface of gold-coated NAA-PCs. The obtained results reveal that the adsorption mechanism between thiol molecules and gold-coated NAA-PCs follows a Langmuir isotherm model, indicating a monolayer sorption mechanism.