Monitoring Cell Cytoskeleton Variations upon Piezoelectric Stimulation: Implications for the Immune System.

Piezoelectric stimulation can have a significant impact on different cellular functions with possible applications in several fields, such as regenerative medicine, cancer therapy, and immunoregulation. For example, piezoelectric stimulation has been shown to modulate cytoskeleton variations: the implications of this effect range from the regulation of migration and invasion of cancer cells to the activation of pro- or anti-inflammatory phenotypes in immune cells. In this chapter, we will present different methodologies to evaluate cytoskeleton variations, focusing on modifications on f-/g-actin ratio and on the migration and invasion ability of tumor cells.

Microglia Polarization and Antiglioma Effects Fostered by Dual Cell Membrane-Coated Doxorubicin-Loaded Hexagonal Boron Nitride Nanoflakes.

Microglial cells play a critical role in glioblastoma multiforme (GBM) progression, which is considered a highly malignant brain cancer. The activation of microglia can either promote or inhibit GBM growth depending on the stage of the tumor development and on the microenvironment conditions. The current treatments for GBM have limited efficacy; therefore, there is an urgent need to develop novel and efficient strategies for drug delivery and targeting: in this context, a promising strategy consists of using nanoplatforms. This study investigates the microglial response and the therapeutic efficacy of dual-cell membrane-coated and doxorubicin-loaded hexagonal boron nitride nanoflakes tested on human microglia and GBM cells. Obtained results show promising therapeutic effects on glioma cells and an M2 microglia polarization, which refers to a specific phenotype or activation state that is associated with anti-inflammatory and tissue repair functions, highlighted through proteomic analysis.

Cell-Membrane-Coated and Cell-Penetrating Peptide-Conjugated Trimagnetic Nanoparticles for Targeted Magnetic Hyperthermia of Prostate Cancer Cells.

Prostate malignancy represents the second leading cause of cancer-specific death among the male population worldwide. Herein, enhanced intracellular magnetic fluid hyperthermia is applied in vitro to treat prostate cancer (PCa) cells with minimum invasiveness and toxicity and highly specific targeting. We designed and optimized novel shape-anisotropic magnetic core-shell-shell nanoparticles (i.e., trimagnetic nanoparticles - TMNPs) with significant magnetothermal conversion following an exchange coupling effect to an external alternating magnetic field (AMF). The functional properties of the best candidate in terms of heating efficiency (i.e., Fe3O4@Mn0.5Zn0.5Fe2O4@CoFe2O4) were exploited following surface decoration with PCa cell membranes (CM) and/or LN1 cell-penetrating peptide (CPP). We demonstrated that the combination of biomimetic dual CM-CPP targeting and AMF responsiveness significantly induces caspase 9-mediated apoptosis of PCa cells. Furthermore, a downregulation of the cell cycle progression markers and a decrease of the migration rate in surviving cells were observed in response to the TMNP-assisted magnetic hyperthermia, suggesting a reduction in cancer cell aggressiveness.

Serum Asprosin Level as a New Biomarker in Differentiating Familial Mediterranean Fever Attacks.

Introduction Familial Mediterranean fever (FMF) is a recessively inherited disease characterized by recurrent attacks of fever and sterile polyserositis. Recently, some proteins originating from adipose tissue have been demonstrated to play a critical role in the inflammatory process. Asprosin is a new adipokine secreted by adipose tissue, and proinflammatory cytokines have been determined to increase with the decrease of circulating asprosin. This study was designed to evaluate the level of asprosin in the acute attack and attack-free period in FMF patients. Materials and methods A total of 65 FMF patients were evaluated for this cross-sectional case-control study. Those who were obese and had concomitant diabetes mellitus, hypertension, heart failure, and rheumatological disease were excluded from the study. The patients were divided into two groups: attack-free period and attack period. Fifteen healthy individuals who were not obese and had no additional disease were included as the control group. Demographic data, gene analyses, laboratory findings, and symptoms were recorded at the time of diagnosis. Serum asprosin level was studied by enzyme-linked immunosorbent assay test in the outpatient clinic controls of the patients. Asprosin levels and other laboratory findings were compared between the attack, attack-free, and control groups. Results Of the patients included in the study, 50% were in the attack period, and 50% were in the free-attack period. The mean age of the FMF patients was 34±10 years. Asprosin level in the control [median (interquartile range (IQR))=30.4 (21.5-57.7) ng/mL] group was significantly higher than the attack [median (IQR)=21.5 (17.5-28) ng/mL] and attack-free [median (IQR)=19(18.7-23) ng/mL] groups (p=0.001). C-reactive protein and sedimentation levels were significantly higher in the attack group compared to the other two groups (p<0.001). There was a moderate correlation between C-reactive protein and asprosin levels (Ro=-0.314, p=0.01). The cut-off value of serum asprosin level was determined as 21.6 ng/mL; sensitivity was 78%, and specificity was 77% (p<0.001). Conclusion The study demonstrated that the serum asprosin levels of FMF patients with acute attack were lower than those in the attack-free periods and healthy controls. Asprosin is likely to have a role in the anti-inflammatory cascade.

Sumac (Rhus coriaria) Extract-Loaded Polymeric Nanosheets Efficiently Protect Human Dermal Fibroblasts from Oxidative Stress.

Under healthy physiological conditions, living organisms possess a variety of antioxidant mechanisms to scavenge overproduced reactive oxygen species (ROS). However, under pathological circumstances, endogenous antioxidant systems may not be adequate to eliminate the excessive amount of oxidants, and thus, a continuous exogenous antioxidant income is required. In this regard, sumac (Rhus coriaria) extract is a good candidate for therapeutic applications, because of its high content of antioxidant polyphenolic compounds. In this work, sumac extract-loaded nanosheets (sumac-nanosheet) have been exploited for loading and controlled release of sumac extract, envisioning topical drug delivery applications. Sumac extract has been obtained through the solvent extraction method, and polymeric nanosheets have been thereafter prepared through the spin coating-assisted layer-by-layer deposition of polycaprolactone (PCL), sumac extract, and poly(d,l-lactic acid) (PDLLA). The collected data show a rich content of the sumac extract in terms of polyphenolic compounds, as well as its strong antioxidant properties. Moreover, for the first time in the literature, we demonstrated the possibility of efficiently loading such extract in polymeric nanosheets and the suitability of this nanoplatform as a reactive oxygen species scavenger in human dermal fibroblasts treated with a pro-oxidant insult.

Hazelnut extract-loaded nanostructured lipid carriers and evaluation of their antioxidant properties.

Reactive oxygen species (ROS) are a common hallmark of many degenerative diseases, developing in all those cases where a failure of physiological antioxidant mechanisms occurs (in particular, antioxidant enzymes and the glutathione system), or in case of exposure to an extremely high level of oxidants. In this regard, antioxidant natural extracts are promising compounds as preventive or therapeutic agents against ROS-dependent degenerations. In this study, a deep investigation of hazelnut (Corylus avellana) extract has been performed in terms of mass spectroscopy, evaluation of phenolic content, and antioxidant capacity. Then, nanostructured lipid carriers (NLCs) have been exploited for encapsulation of the hazelnut extracts in order to achieve prolonged bioactivity, increased stability, and targeting through a sustainable delivery approach. The hazelnut extract-loaded NLCs (NE_NLCs) have been deeply characterized for their stability, production yield, and encapsulation efficiency. Moreover, NE_NLCs showed optimal cytocompatibility on human dermal fibroblast (HDF) cells, as well as excellent antioxidant activity, upon pro-oxidant stimulus on HDF cells.

3D Fiber Reinforced Hydrogel Scaffolds by Melt Electrowriting and Gel Casting as a Hybrid Design for Wound Healing.

Emerging biomanufacturing technologies have revolutionized the field of tissue engineering by offering unprecedented possibilities. Over the past few years, new opportunities arose by combining traditional and novel fabrication techniques, shaping the hybrid designs in biofabrication. One of the potential application fields is skin tissue engineering, in which a combination of traditional principles of wound dressing with advanced biofabrication methods could yield more efficient therapies. In this study, a hybrid design of fiber-reinforced scaffolds combined with gel casting is developed and the efficiency for in vivo wound healing applications is assessed. For this purpose, 3D fiber meshes produced by melt electrowriting are selectively filled with photocrosslinkable gelatin hydrogel matrices loaded with different growth factor carrier microspheres. Additionally, the influence of the inclusion of inorganic bioactive glass particles within the composite fibrous mesh is evaluated. Qualitative evaluation of secondary wound healing criteria and histological analysis shows that hybrid scaffolds containing growth factors and bioactive glass enhances the healing process significantly, compared to the designs merely providing a fiber-reinforced bioactive hydrogel matrix as the wound dressing. This study aims to explore a new application area for melt electrowriting as a powerful tool in fabricating hybrid therapeutic designs for skin tissue engineering.

Modulation of anti-angiogenic activity using ultrasound-activated nutlin-loaded piezoelectric nanovectors.

Angiogenesis plays a fundamental role in tumor development, as it is crucial for tumor progression, metastasis development, and invasion. In this view, anti-angiogenic therapy has received considerable attention in several cancer types in order to inhibit tumor vascularization, and the progress of nanotechnology offers opportunities to target and release anti-angiogenic agents in specific diseased areas. In this work, we showed that the angiogenic behavior of human cerebral microvascular endothelial cells can be inhibited by using nutlin-3a-loaded ApoE-functionalized polymeric piezoelectric nanoparticles, which can remotely respond to ultrasound stimulation. The anti-angiogenic effect, derived from the use of chemotherapy and chronic piezoelectric stimulation, leads to disruption of tubular vessel formation, decreased cell migration and invasion, and inhibition of angiogenic growth factors in the presence of migratory cues released by the tumor cells. Overall, the proposed use of remotely activated piezoelectric nanoparticles could provide a promising approach to hinder tumor-induced angiogenesis.

Ultrasound-responsive nutlin-loaded nanoparticles for combined chemotherapy and piezoelectric treatment of glioblastoma cells.

Glioblastoma multiforme (GBM), also known as grade IV astrocytoma, represents the most aggressive primary brain tumor. The complex genetic heterogeneity, the acquired drug resistance, and the presence of the blood-brain barrier (BBB) limit the efficacy of the current therapies, with effectiveness demonstrated only in a small subset of patients. To overcome these issues, here we propose an anticancer approach based on ultrasound-responsive drug-loaded organic piezoelectric nanoparticles. This anticancer nanoplatform consists of nutlin-3a-loaded ApoE-functionalized P(VDF-TrFE) nanoparticles, that can be remotely activated with ultrasound-based mechanical stimulations to induce drug release and to locally deliver anticancer electric cues. The combination of chemotherapy treatment with chronic piezoelectric stimulation resulted in activation of cell apoptosis and anti-proliferation pathways, induction of cell necrosis, inhibition of cancer migration, and reduction of cell invasiveness in drug-resistant GBM cells. Obtained results pave the way for the use of innovative multifunctional nanomaterials in less invasive and more focused anticancer treatments, able to reduce drug resistance in GBM. STATEMENT OF SIGNIFICANCE: Piezoelectric hybrid lipid-polymeric nanoparticles, efficiently encapsulating a non-genotoxic drug (nutlin-3a) and functionalized with a peptide (ApoE) that enhances their passage through the BBB, are proposed. Upon ultrasound stimulation, nanovectors resulted able to reduce cell migration, actin polymerization, and invasion ability of glioma cells, while fostering apoptotic and necrotic events. This wireless activation of anticancer action paves the way to a less invasive, more focused and efficient therapeutic strategy.

Effect of anesthesia induction on cerebral tissue oxygen saturation in hypertensive patients: an observational study.

OBJECTIVE: In hypertensive patients, the autoregulation curve shifts rightward, making these patients more sensitive than normotensive individuals to hypotension. Hypotension following the induction of anesthesia has been studied in normotensive patients to determine its effects on brain tissue oxygenation, but not enough studies have examined the effect of hypotension on brain oxygenation in hypertensive patients. The current study aimed to use near-infrared spectroscopy to evaluate brain tissue oxygen saturation after the induction of anesthesia in hypertensive patients, who may have impaired brain tissue oxygen saturation. METHODS: The study included a total of 200 patients aged > 18 years old with ASA I-III. Measurements were taken while the patient was breathing room air, after the induction of anesthesia, when the lash reflex had disappeared following the induction of anesthesia, after intubation, and in the 5th, 10th, and 15th minutes of surgery. The patients were divided into nonhypertensive and hypertensive groups. RESULTS: There was a significant difference in age between the groups (p = 0.000). No correlation was found between cerebral tissue oxygen saturation and age (r = 0.015, p = 0.596). Anesthesia induction was observed to decrease mean arterial blood pressure in both groups (p = 0.000). Given these changes, there was no significant difference in brain tissue oxygen saturation between the nonhypertensive and hypertensive groups (p > 0.05). CONCLUSION: There was no difference between hypertensive and normotensive groups in terms of the change rates in cSO2 values. However, there was a difference between the groups in terms of cSO2 values.

Synthesis and characterization of silver nanoparticles decorated polydopamine coated hexagonal boron nitride and its effect on wound healing.

BACKGROUND: Wound healing is an essential physiological process involving many cell types and their products acting in a marvellous harmony to repair damaged tissues. During the healing process, cellular proliferation and extracellular matrix remodelling stages could be interrupted by undesired factors including microorganisms and altered metabolic activities. In such a case, the process requires some external stimulants to accelerate or remediate the healing stages. METHODS: In this study, we report a multifunctional wound healing stimulating agent. In this context, hexagonal boron nitride (hBN) nanoparticles, silver nanoparticles (AgNPs) and polydopamine(pdopa) were used through mussel-inspired chemistry of dopamine to obtain pdopa coated hBN (hBN@pdopa) and AgNPs decorated hBN@pdopa (hBN@pdopa-AgNPs). These two nanostructures were investigated to observe stages of healing. RESULTS: AgNPs were chosen for inflammation reduction and hBN for induced cell proliferation and migration. In in vitro experiments, firstly, high cellular uptake capacity and biocompatibility of hBN@pdopa and hBN@pdopa-AgNPs were evaluated. They were also tested for their reaction against increased concentration of reactive oxygen species (ROS) in injured cells. Finally, their effect on cellular migration, intracellular tube formation and F-actin organization were monitored by light and confocal microscopy, respectively. CONCLUSION: The results clearly indicate that the hBN@pdopa-AgNPs significantly decrease ROS production, promote wound closure, and reorganize tube formation in cells.

Nanotechnology-Based Strategies to Evaluate and Counteract Cancer Metastasis and Neoangiogenesis.

Cancer metastasis is the major cause of cancer-related morbidity and mortality. It represents one of the greatest challenges in cancer therapy, both because of the ability of metastatic cells to spread into different organs, and because of the consequent heterogeneity that characterizes primary and metastatic tumors. Nanomaterials can potentially be used as targeting or detection agents owing to unique chemical and physical features that allow tailored and tunable theranostic functions. This review highlights nanomaterial-based approaches in the detection and treatment of cancer metastasis, with a special focus on the evaluation of nanostructure effects on cell migration, invasion, and angiogenesis in the tumor microenvironment.

3D printing of silver-doped polycaprolactone-poly(propylene succinate) composite scaffolds for skin tissue engineering.

Scaffold-based tissue engineering approaches have been commonly used for skin regeneration or wound healings caused by diseases or trauma. For an ideal complete healing process, scaffold structures need to meet the criteria of biocompatibility, biodegradability, and antimicrobial properties, as well as to provide geometrical necessities for the regeneration of damaged tissue. In this study, design, synthesis and characterization of a three dimensional (3D) printable copolymer based on polycaprolactone-block-poly(1,3-propylene succinate) (PCL-PPSu) including anti-microbial silver particles is presented. 3D printing of PCL-PPSu copolymers provided a lower processing temperature compared to neat PCL, hence, inclusion of temperature-sensitive bioactive reagents into the developed copolymer could be realized. In addition, 3D printed block copolymer showed an enhanced hydrolytic and enzymatic degradation behavior. Cell viability and cytotoxicity of the developed copolymer were evaluated by using human dermal fibroblast (HDF) cells. The addition of silver nitrate within the polymer matrix resulted in a significant decrease in the adhesion of different types of microorganisms on the scaffold without inducing any cytotoxicity on HDF cells in vitro. The results suggested that 3D printed PCL-PPSu scaffolds containing anti-microbial silver particles could be considered as a promising biomaterial for emerging skin regenerative therapies, in the light of its adaptability to 3D printing technology, low-processing temperature, enhanced degradation behavior and antimicrobial properties.

Hexagonal boron nitrides reduce the oxidative stress on cells.

The molecular stress caused by a drug administered to treat a disorder on healthy cells appears as a side effect. In this study, we aim to understand the potential of hexagonal boron nitrides (hBNs) as a therapeutic agent to relieve the cellular stress exerted by drugs. First, the cytotoxicity of hBNs and their possible degradation product, boric acid (BA), on the embryonic mouse hippocampal cell line mHippo E-14 was assessed in a wide concentration range (4.4-440 µg ml-1) of boron including hBNs and BA for 24 and 72 h exposure. Then, cell cycle, reactive oxygen species generation, cell death mechanism and apoptotic body formation in nuclei with hBN and BA exposure were evaluated at increased concentrations and incubation times. Finally, the cells, exposed to doxorubicin (DOX), an anti-cancer chemotherapy drug, to exert oxidative stress, were treated with hBNs and BA. The results indicate that hBNs decrease the oxidative stress at the concentrations that are nontoxic to cells. The study suggests that hBNs can open new venues for their investigation to reduce or eliminate the adverse effects of toxic drugs used in the treatment of several fatal diseases including neurological disorders and cancer with their slow degradation feature.

Electrospun Nanofibers With pH-Responsive Coatings for Control of Release Kinetics.

Functional and stimuli-responsive nanofibers with an enhanced surface area/volume ratio provide controlled and triggered drug release with higher efficacy. In this study, chemotherapeutic agent Rose Bengal (RB) (4,5,6,7-tetrachloro-2', 4',5',7'-tetraiodofluoresceindisodium)-loaded water-soluble polyvinyl alcohol (PVA) nanofibers were synthesized by using the electrospinning method. A thin layer of poly(4-vinylpyridine-co-ethylene glycol dimethacrylate) p(4VP-co-EGDMA) was deposited on the RB-loaded nanofibers (PVA-RB) via initiated chemical vapor deposition (iCVD), coating the fiber surfaces to provide controllable solubility and pH response to the nanofibers. The uncoated and [p(4VP-co-EGDMA)-PVA] coated PVA-RB nanofiber mats were studied at different pH values to analyze their degradation and drug release profiles. The coated nanofibers demonstrated high stability at neutral and basic pH values for long incubation durations of 72 h, whereas the uncoated nanofibers dissolved in <2 h. The drug release studies showed that the RB release from coated PVA-RB nanofibers was higher at neutral and basic pH values, and proportional to the pH of the solution, whereas the degradation and RB release rates from the uncoated PVA-RB nanofibers were significantly higher and did not depend on the pH of environment. Further analysis of the release kinetics using the Peppas model showed that while polymer swelling and dissolution were the dominant mechanisms for the uncoated nanofibers, for the coated nanofibers, Fickian diffusion was the dominant release mechanism. The biocompatibility and therapeutic efficiency of the coated PVA-RB nanofibers against brain cancer was investigated on glioblastoma multiforme cancer cells (U87MG). The coated PVA nanofibers were observed to be highly biocompatible, and they significantly stimulated the ROS production in cells, increasing apoptosis. These promising results confirmed the therapeutic activity of the coated PVA-RB nanofibers on brain cancer cells, and encouraged their further evaluation as drug carrier structures in brain cancer treatment.

Synthesis, Functionalization, and Bioapplications of Two-Dimensional Boron Nitride Nanomaterials.

Two-dimensional boron nitride nanostructures (2D-BNNs) have been increasingly investigated for their applications in several scientific and technological areas. This considerable interest is due to their unique physicochemical properties, which include high hydrophobicity, heat and electrical insulation, resistance to oxidation, antioxidation capacity, thermal conductivity, high chemical stability, mechanical strength, and hydrogen storage capacity. They are also used as fillers, antibacterial agents, protective coating agents, lubricants, boron neutron capture therapy agents, nanocarriers for drug delivery, and for the receptor phase in chemosensors. The investigations for their use in medicine and biomedicine are very promising, including cancer therapy and wound healing. In this review, 2D-BNNs synthesis and their surface modification strategies, biocompatibility, and bioapplication studies are discussed. Finally, a perspective for the future use of these novel nanomaterials in the biomedical field is provided.

Stimulatory Effect of Hexagonal Boron Nitrides in Wound Healing.

There is an increased interest in the use of hexagonal boron nitrides (hBNs) in medicine due to their unique properties that include low toxicity, absorption of UV light and neutrons, high chemical stability, and mechanical strength. In this study, hBNs were investigated for wound healing therapy as a potential therapeutic agent. Boron compounds (especially boric acid) are increasingly used in wound healing therapy; however, their short half-life is a bottleneck and limits their use. Because hBNs slowly degrade and one of the degradation products is boric acid (BA), hBNs and BA were comparatively evaluated in vitro to assess their influence on wound healing. First, antimicrobial activity and cytotoxicity of synthesized hBNs were evaluated for dose determination. Then, the results of scratch assay indicated that hBNs accelerated the wound closure at low concentrations, showing also enhanced angiogenic activity compared to BA. The cell cycle analysis showed that hBNs did not arrest the cells in the G2/M phase but induced the cells to go into S phase, whereas BA had almost no effect on the cell proliferation. It was also found that the hBNs decreased the reactive oxygen species level more than BA. Apoptosis/necrosis assay showed that the hBNs rescued the cells from apoptosis, whereas BA had almost no effect on the cell death mechanism. Beside, hBNs did not cause damage to the mitochondria at low concentration and did not perturb the F-actin conformation at all tested concentrations. These findings suggest that hBNs might be a potential therapeutic agent in wound healing therapy.

One-Step Synthesis of Hexagonal Boron Nitrides, Their Crystallinity and Biodegradation.

Hexagonal boron nitrides (hBNs) have recently been investigated for several novel applications due to their unique properties such as biocompatibility, superhydrophobicity, electrical insulation, and thermal and chemical stability. In addition, their biodegradation products have recently reported to have therapeutic effect on certain cancer types. hBNs are easily synthesized from boron and nitrogen precursors at moderately low temperatures. However, crystallinity and yield vary depending on the type of precursor, reaction temperature, and duration. In this study, a simple one-step hBNs synthesis method is reported without a catalyst, which might be an undesired contaminant for biomedical applications. The influence of boron precursors (boric acid, colemanite, or boron trioxide) on hBNs crystallinity, stability, and biodegradation in suspensions containing oxidative and hydrolytic degradation agents is investigated with the aim of their possible application in biomedicine. We found that the choice of boron precursor is a critically important parameter controlling the hBNs crystallinity and dependently influencing the biodegradation rate.

Boron nitride nanotubes for gene silencing.

BACKGROUND: Non-viral gene delivery is increasingly investigated as an alternative to viral vectors due to low toxicity and immunogenicity, easy preparation, tissue specificity, and ability to transfer larger sizes of genes. METHODS: In this study, boron nitride nanotubes (BNNTs) are functionalized with oligonucleotides (oligo-BNNTs). The morpholinos complementary to the oligonucleotides attached to the BNNTs (morpholino/oligo-BNNTs) are hybridized to silence the luciferase gene. The morpholino/oligo-BNNTs conjugates are administered to luciferase-expressing cells (MDA-MB-231-luc2) and the luciferase activity is monitored. RESULTS: The luciferase activity is decreased when MDA-MB-231-luc2 cells were treated with morpholino/oligo-BNNTs. CONCLUSIONS: The study suggests that BNNTs can be used as a potential vector to transfect cells. GENERAL SIGNIFICANCE: BNNTs are potential new nanocarriers for gene delivery applications.

Evaluation of boron nitride nanotubes and hexagonal boron nitrides as nanocarriers for cancer drugs.

AIM: Boron nitride nanotubes (BNNTs) and hexagonal boron nitrides (hBNs) are novel nanostructures with high mechanical strengths, large surface areas and excellent biocompatibilities. Here, the potential use of BNNTs and hBNs as nanocarriers was comparatively investigated for use with cancer drugs. MATERIALS & METHODS: Doxorubicin (Dox) and folate are used as model drugs and targeting agents, respectively. RESULTS & DISCUSSION: The obtained results indicate that BNNTs have about a threefold higher Dox loading capacity than hBNs. It was also found that cellular uptake of folate-Dox-BNNTs was much higher when compared with Dox-BNNTs for HeLa cells, due to the presence of folate receptors on the cell surface, leading to increased cancer cell death. In summary, folate and Dox conjugated BNNTs are promising agents in nanomedicine and may have potential drug delivery applications.