Nanofilm generated non-pharmacological anabolic bone stimulus.

Stimulus-responsive nanomaterials have mainly been employed to ablate or destroy tissues or to facilitate controlled release of drugs or biologics. Herein, we demonstrate the potential of stimulus-responsive nanomaterials to promote tissue regeneration via a non-pharmacological and noninvasive strategy. Thin nanofilms of an optically-absorbing organic dye or nanoparticle (single-walled graphene nanoribbons [SWOGNR]) were placed over (without touching the skin) a rodent femoral fracture site. A nanosecond pulsed near-infrared laser diode was employed to generate photoacoustic (PA) signals from the nanofilms. X-ray micro-computed tomography (microCT), histology, and mechanical testing results showed that daily PA stimulations of upto 45 min for 6 weeks (complete fracture healing) do not adversely affect bone regeneration and quality. Further, microCT and histological analysis showed 10 min daily stimulation for 2 weeks significantly increases bone quantity at the fracture sites of rats exposed to the nanoparticle-generated PA signals. In these rats, up to threefold increase in bone volume to callus volume ratio and twofold increase in bone mineral density within the callus were noted, compared to rats that were not exposed to the photoacoustic signals. The results taken together indicate that nanofilm-generated photoacoustic signals serve as an anabolic stimulus for bone regeneration. The results, in conjugation with the ability of these nanofilms to serve as PA contrast agents, present opportunities toward the development of integrated noninvasive imaging and noninvasive or invasive treatment strategies for bone loss due to disease or trauma.

Two-dimensional graphene oxide-reinforced porous biodegradable polymeric nanocomposites for bone tissue engineering.

This study investigates the mechanical properties and in vitro cytotoxicity of two-dimensional (2D) graphene oxide nanoribbons and nanoplatelets (GONRs and GONPs) reinforced porous polymeric nanocomposites. Highly porous poly(propylene fumarate) (PPF) nanocomposites were prepared by dispersing 0.2 wt % single- and multiwalled SONRs (SWGONRs and MWGONRs) and GONPs. The mechanical properties of scaffolds were characterized using compression testing and in vitro cytocompatibility was assessed using QuantiFlour assay for cellularity and PrestoBlue assay for cell viability. Immunofluorescence was used to assess collagen-I expression and deposition in the extracellular matrix. Porous PPF scaffolds were used as a baseline control and porous single and multiwalled carbon nanotubes (SWCNTs and MWCNTs) reinforced nanocomposites were used as positive controls. Results show that incorporation of 2D graphene nanomaterials leads to an increase in the mechanical properties of porous PPF nanocomposites with following the trend: MWGONRs > GONPs > SWGONRs > MWCNTs > SWCNTs > PPF control. MWGONRs showed the best enhancement of compressive mechanical properties with increases of up to 26% in compressive modulus (i.e., Young's modulus), ~60% in yield strength, and ~24% in the ultimate compressive strength. Addition of 2D nanomaterials did not alter the cytocompatibility of porous PPF nanocomposites. Furthermore, PPF nanocomposites reinforced with SWGONRs, MWGONRs, and GONPs show an improvement in the adsorption of collagen-I compared to PPF baseline control. The results of this study show that 2D graphene nanomaterial reinforced porous PPF nanocomposites possess superior mechanical properties, cytocompatibility, and increased protein adsorption. The favorable cytocompatibility results opens avenues for in vivo safety and efficacy studies for bone tissue engineering applications. © 2019 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 107A: 1143-1153, 2019.

In Vitro Bioactivity of One- and Two-Dimensional Nanoparticle-Incorporated Bone Tissue Engineering Scaffolds.

This study investigates the effect of incorporation of one- or two-dimensional nanoparticles with distinct composition and morphology on the bioactivity of biodegradable, biocompatible polymer matrices. 0.2 wt% multiwalled carbon nanotubes, multiwalled graphene nanoribbons, graphene oxide nanoplatelets (GONPs), molybdenum disulfide nanoplatelets (MSNPs), or tungsten disulfide nanotubes (WSNTs) were uniformly dispersed in poly(lactic-co-glycolic acid) (PLGA) polymer. PLGA or nanoparticle-incorporated PLGA were then incubated with simulated body fluid (SBF) under physiological conditions for 1, 3, 7, or 14 days. Apatite collection on control and incorporated scaffolds was assessed. All groups showed apatite precipitate on the surface after 1 day of SBF incubation. After 14 days of SBF incubation, scaffolds incorporated with GONPs, MSNPs, or WSNTs showed significantly higher phosphate accumulation compared to PLGA scaffolds. Scaffolds incorporated with GONPs, MSNPs, or WSNTs should be studied in vivo to further investigate potential bioactivity, leading to enhanced integration and tissue repair at the bone-implant interface.

Three-dimensional carbon nanotube scaffolds for long-term maintenance and expansion of human mesenchymal stem cells.

Expansion of mesenchymal stem cells (MSCs) and maintenance of their self-renewal capacity in vitro requires specialized robust cell culture systems. Conventional approaches using animal-derived or artificial matrices and a cocktail of growth factors have limitations such as consistency, scalability, pathogenicity, and loss of MSC phenotype. Herein, we report the use of all-carbon 3-D single- and multiwalled carbon nanotube scaffolds (SWCNTs and MWCNTs) as artificial matrices for long-term maintenance and expansion of human MSCs. Three-dimensional SWCNT and MWCNT scaffolds were fabricated using a novel radical initiated thermal cross-linking method that covalently cross-links CNTs to form 3-D macroporous all-carbon architectures. Adipose-derived human MSCs showed good cell viability, attachment, proliferation, and infiltration in MWCNT and SWCNT scaffolds comparable to poly(lactic-co-glycolic) acid (PLGA) scaffolds (baseline control). ADSCs retained stem cell phenotype after 30 days and satisfied the International Society for Cellular Therapy's (ISCT) minimal criteria for MSCs. Post expansion, (1) ADSCs showed in vitro adherence to tissue culture polystyrene (TCPS); (2) MSC surface antigen expression [CD14(-), CD19(-), CD34(-), CD45(-), CD73(+), CD90(+), CD105(+)]; and (3) trilineage differentiation into osteoblasts, adipocytes, and chondrocytes. Results show that cross-linked 3-D MWCNTs and SWCNTs scaffolds are suitable for ex vivo expansion and maintenance of MSCs for therapeutic applications. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1927-1939, 2017.

Boron nitride nanotubes and nanoplatelets as reinforcing agents of polymeric matrices for bone tissue engineering.

This study investigates the mechanical properties and in vitro cytotoxicity of one- and two-dimensional boron nitride nanomaterials-reinforced biodegradable polymeric nanocomposites. Poly(propylene fumarate) (PPF) nanocomposites were fabricated using crosslinking agent N-vinyl pyrrolidone and inorganic nanomaterials: boron nitride nanotubes (BNNTs) and boron nitride nanoplatelets (BNNPs) dispersed at 0.2 wt % in the polymeric matrix. The incorporation of BNNPs and BNNTs resulted in a ∼38 and ∼15% increase in compressive (Young's) modulus, and ∼31 and ∼6% increase in compressive yield strength compared to PPF control, respectively. The nanocomposites showed a time-dependent increased protein adsorption for collagen I protein. The cytotoxicity evaluation of aqueous BNNT and BNNP dispersions (at 1-100 µg/mL concentrations) using murine MC3T3 preosteoblast cells showed ∼73-99% viability. The cytotoxicity evaluation of media extracts of nanocomposites before crosslinking, after crosslinking, and upon degradation (using 1×-100× dilutions) showed dose-dependent cytotoxicity responses. Crosslinked nanocomposites showed excellent (∼79-100%) cell viability, cellular attachment (∼57-67%), and spreading similar to cells grown on the surface of tissue culture polystyrene control. The media extracts of degradation products showed a dose-dependent cytotoxicity. The favorable cytocompatibility results in combination with improved mechanical properties of BNNT and BNNP nanocomposites opens new avenues for further in vitro and in vivo safety and efficacy studies towards bone tissue engineering applications. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 406-419, 2017.

In Vivo Hard and Soft Tissue Response of Two-Dimensional Nanoparticle Incorporated Biodegradable Polymeric Scaffolds.

Current efforts in the design of bone tissue engineering scaffolds have focused on harnessing the physiochemical properties of two-dimensional organic and inorganic nanoparticles to improve bulk and surface properties of biodegradable polymers. Herein, we investigate the hard and soft tissue in vivo biocompatibility of two such constructs: 90% porous poly(lactic-co-glycolic acid) (PLGA) nanocomposite scaffolds incorporated with 0.2 wt % graphene oxide nanoplatelets (GONPs) or molybdenum disulfide nanoplatelets (MSNPs). Scaffolds were implanted in a noncritical sized monocortical defect in the tibia or subcutaneously on the dorsum of a rat model for 2 or 6 weeks. Hard and soft tissue in vivo biocompatibility of the nanoparticle reinforced scaffolds was comparable to that of the PLGA control. In addition, 2 weeks after implantation, significantly less bone growth (∼35%) was observed for the PLGA group compared to that of the empty defect group; it was not observed for the experimental groups which showed 20% and 15% greater bone growth compared to that of the PLGA group. This may indicate that the nanoparticles do play a role in assisting bone regeneration. Taken together, the results suggest that scaffolds incorporated with GONPs or MSNPs show promise for bone tissue engineering applications.

Gene delivery to mammalian cells using a graphene nanoribbon platform.

We have developed a novel oxidized graphene nanoribbon-based platform (O-GNR) for gene delivery of double-stranded DNA into mammalian cells. O-GNRs, synthesized via longitudinal unzipping of multi-walled carbon nanotubes (MWCNTs), exhibited efficient DNA loading of small dsDNA fragments. Fourier Transform Infrared Spectroscopy identified stretching peaks in the O-P-O and DNA sugar phosphate backbone that were consistent with DNA loading onto O-GNRs. The presence of salts in the loading buffer promoted DNA loading and effective dispersion of O-GNRs. DNA:O-GNR complexes were stable upon treatment with surfactants Tween 20 and Triton-X100. O-GNRs did not impact the viability of mammalian cells. Last, the detection of GFP expression upon transfection of the DNA:O-GNR complex indicated that the cargo DNA is expressed in the nucleus. Taken together, O-GNRs function as a platform for gene delivery to mammalian cells.

Three-dimensional macroporous graphene scaffolds for tissue engineering.

The assembly of carbon nanomaterials into three-dimensional (3D) porous scaffolds is critical to harness their unique physiochemical properties for tissue engineering and regenerative medicine applications. In this study, we report the fabrication, characterization, and in vitro cytocompatibility of true 3D (>1 mm in all three dimensions), macroscopic (3-8 mm in height and 4-6 mm in diameter), chemically cross-linked graphene scaffolds prepared via radical initiated thermal cross-linking of single- and multiwalled graphene oxide nanoribbons (SWGONRs and MWGONRs). SWGONR and MWGONR scaffolds possess tunable porosity (∼65-80%) and interconnected macro-, micro-, and nanoscale pores. Human adipose derived stem cells (ADSCs) and murine MC3T3 preosteoblast cells show good cell viability on SWGONR and MWGONR scaffolds after 1, 3, and 5 days comparable to 3D poly(lactic-co-glycolic) acid (PLGA) scaffolds. Confocal live-cell imaging showed that cells were metabolically active and could spread on SWGONR and MWGONR scaffolds. Immunofluorescence imaging showed the presence of focal adhesion protein vinculin and expression of cell proliferation marker Ki-67 suggesting that cells could attach and proliferate on SWGONR and MWGONR scaffolds. These results indicate that cross-linked SWGONR and MWGONR scaffolds are cytocompatible and opens-avenues toward the development of 3D multifunctional graphene scaffolds for tissue engineering applications. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 73-83, 2017.

Toxicology of graphene-based nanomaterials.

Graphene based nanomaterials possess remarkable physiochemical properties suitable for diverse applications in electronics, telecommunications, energy and healthcare. The human and environmental exposure to graphene-based nanomaterials is increasing due to advancements in the synthesis, characterization and large-scale production of graphene and the subsequent development of graphene based biomedical and consumer products. A large number of in vitro and in vivo toxicological studies have evaluated the interactions of graphene-based nanomaterials with various living systems such as microbes, mammalian cells, and animal models. A significant number of studies have examined the short- and long-term in vivo toxicity and biodistribution of graphene synthesized by variety of methods and starting materials. A key focus of these examinations is to properly associate the biological responses with chemical and morphological properties of graphene. Several studies also report the environmental and genotoxicity response of pristine and functionalized graphene. This review summarizes these in vitro and in vivo studies and critically examines the methodologies used to perform these evaluations. Our overarching goal is to provide a comprehensive overview of the complex interplay of biological responses of graphene as a function of their physiochemical properties.

Two- and Three-Dimensional All-Carbon Nanomaterial Assemblies for Tissue Engineering and Regenerative Medicine.

Carbon nanomaterials such as carbon nanotubes and graphene have gained significant interest in the fields of materials science, electronics and biomedicine due to their interesting physiochemical properties. Typically these carbon nanomaterials have been dispersed in polymeric matrices at low concentrations to improve the functional properties of nanocomposites employed as two-dimensional (2D) substrates or three-dimensional (3D) porous scaffolds for tissue engineering applications. There has been a growing interest in the assembly of these nanomaterials into 2D and 3D architectures without the use of polymeric matrices, surfactants or binders. In this article, we review recent advances in the development of 2D or 3D all-carbon assemblies using carbon nanotubes or graphene as nanoscale building-block biomaterials for tissue engineering and regenerative medicine applications.

Graphene-based platforms for cancer therapeutics.

Graphene is a multifunctional carbon nanomaterial and could be utilized to develop platform technologies for cancer therapies. Its surface can be covalently and noncovalently functionalized with anticancer drugs and functional groups that target cancer cells and tissue to improve treatment efficacies. Furthermore, its physicochemical properties can be harnessed to facilitate stimulus responsive therapeutics and drug delivery. This review article summarizes the recent literature specifically focused on development of graphene technologies to treat cancer. We will focus on advances at the interface of graphene based drug/gene delivery, photothermal/photodynamic therapy and combinations of these techniques. We also discuss the current understanding in cytocompatibility and biocompatibility issues related to graphene formulations and their implications pertinent to clinical cancer management.

Sulfobutyl ether ß-cyclodextrin (Captisol(®) ) and methyl ß-cyclodextrin enhance and stabilize fluorescence of aqueous indocyanine green.

As the only FDA-approved near-infrared fluorophore, indocyanine green (ICG) is commonly used to image vasculature in vivo. ICG degrades rapidly in solution, which limits its usefulness in certain applications, including time-sensitive surgical procedures. We propose formulations that address this shortcoming via complexation with ß-cyclodextrin derivatives (ß-CyD), which are known to create stabilizing inclusion complexes with hydrophobic molecules. Here, we complexed ICG with highly soluble methyl ß-CyD and FDA-approved sulfobutyl ether ß-CyD (Captisol(®) ) in aqueous solution. We measured the fluorescence of the complexes over 24 h. We found that both CyD+ICG complexes exhibit sustained fluorescence increases of >2.0× versus ICG in water and >20.0× in PBS. Using transmission electron microscopy, we found evidence of reduced aggregation in complexes versus ICG alone. We thus conclude that this reduction in aggregation helps mitigate fluorescence autoquenching of CyD+ICG complexes compared in ICG alone. We also found that while ICG complexed with methyl ß-CyD severely reduced the viability of MRC-5 fibroblasts, ICG complexed with sulfobutyl ether ß-CyD had no effect on viability. These results represent an important first step toward enhancing the utility of aqueous ICG by reducing aggregation-dependent fluorescence degradation. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 104B: 1457-1464, 2016.

Interactions of 1D- and 2D-layered inorganic nanoparticles with fibroblasts and human mesenchymal stem cells.

AIM: This study investigates the effects of tungsten disulfide nanotubes (WSNTs) and molybdenum disulfide nanoplatelets (MSNPs) on fibroblasts (NIH-3T3) and mesenchymal stem cells (MSCs) to determine safe dosages for potential biomedical applications. MATERIALS & METHODS: Cytotoxicity of MSNPs and WSNTs (5-300 µg/ml) on NIH-3T3 and MSCs was assessed at 6, 12 or 24 h. MSC differentiation to adipocytes and osteoblasts was assessed following treatment for 24 h. RESULTS: Only NIH-3T3 cells treated with MSNPs showed dose or time dependent increase in cytotoxicity. Differentiation markers of MSCs in treated groups were unaffected compared with untreated controls. CONCLUSION: MSNPs and WSNTs at concentrations less than 50 µg/ml are potentially safe for treatment of fibroblasts or MSCs for up to 24 h.

Fabrication and cytocompatibility of in situ crosslinked carbon nanomaterial films.

Assembly of carbon nanomaterials into two-dimensional (2D) coatings and films that harness their unique physiochemical properties may lead to high impact energy capture/storage, sensors, and biomedical applications. For potential biomedical applications, the suitability of current techniques such as chemical vapor deposition, spray and dip coating, and vacuum filtration, employed to fabricate macroscopic 2D all carbon coatings or films still requires thorough examination. Each of these methods presents challenges with regards to scalability, suitability for a large variety of substrates, mechanical stability of coatings or films, or biocompatibility. Herein we report a coating process that allow for rapid, in situ chemical crosslinking of multi-walled carbon nanotubes (MWCNTs) into macroscopic all carbon coatings. The resultant coatings were found to be continuous, electrically conductive, significantly more robust, and cytocompatible to human adipose derived stem cells. The results lay groundwork for 3D layer-on-layer nanomaterial assemblies (including various forms of graphene) and also opens avenues to further explore the potential of MWCNT films as a novel class of nano-fibrous mats for tissue engineering and regenerative medicine.

Porous three-dimensional carbon nanotube scaffolds for tissue engineering.

Assembly of carbon nanomaterials into three-dimensional (3D) architectures is necessary to harness their unique physiochemical properties for tissue engineering and regenerative medicine applications. Herein, we report the fabrication and comprehensive cytocompatibility assessment of 3D chemically crosslinked macrosized (5-8 mm height and 4-6 mm diameter) porous carbon nanotube (CNT) scaffolds. Scaffolds prepared via radical initiated thermal crosslinking of single- or multiwalled CNTs (SWCNTs and MWCNTs) possess high porosity (>80%), and nano-, micro-, and macroscale interconnected pores. MC3T3 preosteoblast cells on MWCNT and SWCNT scaffolds showed good cell viability comparable to poly(lactic-co-glycolic) acid (PLGA) scaffolds after 5 days. Confocal live cell and immunofluorescence imaging showed that MC3T3 cells were metabolically active and could attach, proliferate, and infiltrate MWCNT and SWCNT scaffolds. SEM imaging corroborated cell attachment and spreading and suggested that cell morphology is governed by scaffold surface roughness. MC3T3 cells were elongated on scaffolds with high surface roughness (MWCNTs) and rounded on scaffolds with low surface roughness (SWCNTs). The surface roughness of scaffolds may be exploited to control cellular morphology and, in turn, govern cell fate. These results indicate that crosslinked MWCNTs and SWCNTs scaffolds are cytocompatible, and open avenues toward development of multifunctional all-carbon scaffolds for tissue engineering applications.

In vitro cytocompatibility of one-dimensional and two-dimensional nanostructure-reinforced biodegradable polymeric nanocomposites.

This study investigates the in vitro cytocompatibility of one-dimensional and two-dimensional (1D and 2D) carbon and inorganic nanomaterial reinforced polymeric nanocomposites fabricated using biodegradable polymer poly (propylene fumarate), crosslinking agent N-vinyl pyrrolidone (NVP) and following nanomaterials: single and multiwalled carbon nanotubes, single and multiwalled graphene oxide nanoribbons, graphene oxide nanoplatelets, molybdenum disulfide nanoplatelets, or tungsten disulfide nanotubes dispersed between 0.02 and 0.2 wt% concentrations in the polymer. The extraction media of unreacted components, crosslinked nanocomposites and their degradation products were examined for effects on viability and attachment using two cell lines: NIH3T3 fibroblasts and MC3T3 preosteoblasts. The extraction media of unreacted PPF/NVP elicited acute dose-dependent cytotoxicity attributed to leaching of unreacted components into cell culture media. However, extraction media of crosslinked nanocomposites showed no dose dependent adverse effects. Further, all crosslinked nanocomposites showed high viability (78-100%), high cellular attachment (40-55%), and spreading that was confirmed by confocal and scanning electron microscopy. Degradation products of nanocomposites showed a mild dose-dependent cytotoxicity possibly due to acidic degradation components of PPF. In general, compared to PPF control, none of the nanocomposites showed significant differences in cellular response to unreacted components, crosslinked nanocomposites and their degradation products. Initial minor cytotoxic response and lower cell attachment numbers were observed only for a few nanocomposite groups; these effects were absent at later time points for all PPF nanocomposites. The favorable cytocompatibility results for all the nanocomposites opens avenues for in vivo safety and efficacy studies for bone tissue engineering applications.

Enzymatic Degradation of Oxidized and Reduced Graphene Nanoribbons by Lignin Peroxidase.

The expanding use of graphene for various industrial and biomedical applications requires efficient remediation strategies during their disposal into waste streams. Additionally, the interactions of graphene with the biota need thorough evaluation. In this study, we investigated the interactions of oxidized and reduced graphene oxide nanoribbons (GONRs and rGONRs) with lignin peroxidase (LiP), a ligninolytic enzyme released from white rot fungus. GONRs and rGONRs were treated with LiP in the presence and absence of veratryl alcohol (VA; an electron transfer mediator and secondary metabolite of white rot fungi). Transmission electron microscopy showed the formation of large defects (holes) in the graphene sheet, which increased in diameter with increased degradation time. Raman spectroscopic analysis indicated that, within 96 hours, in the presence of hydrogen peroxide and VA, the GONRs and rGONRs were completely and partially degraded by LiP, respectively. Comparisons between groups with or without VA showed that degradation of GONRs was accelerated in the presence of VA. These results indicated that LiP could efficiently degrade GONRs and rGONRs in the presence of VA, suggesting that VA may be an essential factor needed to degrade rGONRs via LiP treatment. Thus, the wide presence of white rot fungi, and thereby LiP, in nature, could lead to efficient degradation of graphene present in the environment. Additionally, LiP, which has a higher theoretical redox potential compared to horseradish peroxidases and myeloperoxidases, could be a better candidate for the environmental remediation of graphene.

Synthesis, Characterization, In Vitro Phantom Imaging, and Cytotoxicity of A Novel Graphene-Based Multimodal Magnetic Resonance Imaging - X-Ray Computed Tomography Contrast Agent.

Graphene nanoplatelets (GNPs), synthesized using potassium permanganate-based oxidation and exfoliation followed by reduction with hydroiodic acid (rGNP-HI), have intercalated manganese ions within the graphene sheets, and upon functionalization with iodine, show excellent potential as biomodal contrast agents for magnetic resonance imaging (MRI) and computed tomography (CT). Structural characterization of rGNP-HI nanoparticles with low- and high-resolution transmission electron microscope (TEM) showed disc-shaped nanoparticles (average diameter, 200 nm, average thickness, 3 nm). Energy dispersive X-ray spectroscopy (EDX) analysis confirmed the presence of intercalated manganese. Raman spectroscopy and X-ray diffraction (XRD) analysis of rGNP-HI confirmed the reduction of oxidized GNPs (O-GNPs), absence of molecular and physically adsorbed iodine, and the functionalization of graphene with iodine as polyiodide complexes (I3- and I5-). Manganese and iodine content were quantified as 5.1 ± 0.5 and 10.54 ± 0.87 wt% by inductively-coupled plasma optical emission spectroscopy and ion-selective electrode measurements, respectively. In vitro cytotoxicity analysis, using absorbance (LDH assay) and fluorescence (calcein AM) based assays, performed on NIH3T3 mouse fibroblasts and A498 human kidney epithelial cells, showed CD50 values of rGNP-HI between 179-301 µg/ml, depending on the cell line and the cytotoxicity assay. CT and MRI phantom imaging of rGNP-HI showed high CT (approximately 3200% greater than HI at equimolar iodine concentration) and MRI (approximately 59% greater than equimolar Mn2+ solution) contrast. These results open avenues for further in vivo safety and efficacy studies towards the development of carbon nanostructure-based multimodal MRI-CT contrast agents.

The effects of graphene nanostructures on mesenchymal stem cells.

We report the effects of two-dimensional graphene nanostructures; graphene nano-onions (GNOs), graphene oxide nanoribbons (GONRs), and graphene oxide nanoplatelets (GONPs) on viability, and differentiation of human mesenchymal stem cells (MSCs). Cytotoxicity of GNOs, GONRs, and GONPs dispersed in distearoyl-sn-glycero-3-phosphoethanolamine-N-[amino(polyethylene glycol)] (DSPE-PEG), on adipose derived mesenchymal stem cells (adMSCs), and bone marrow-derived mesenchymal stem cells (bmMSCs) was assessed by AlamarBlue and Calcein AM viability assays at concentrations ranging from 5 to 300 µg/ml for 24 or 72 h. Cytotoxicity of the 2D graphene nanostructures was found to be dose dependent, not time dependent, with concentrations less than 50 µg/ml showing no significant differences compared to untreated controls. Differentiation potential of adMSCs to adipocytes and osteoblasts, - characterized by Oil Red O staining and elution, alkaline phosphatase activity, calcium matrix deposition and Alizarin Red S staining - did not change significantly when treated with the three graphene nanoparticles at a low (10 µg/ml) and high (50 µg/ml) concentration for 24 h. Transmission electron microscopy (TEM) and confocal Raman spectroscopy indicated cellular uptake of only GNOs and GONPs. The results lay the foundation for the use of these nanoparticles at potentially safe doses as ex vivo labels for MSC-based imaging and therapy.

Frontoethmoidal encephalocele: Case report and review on management.

Encephaloceles, especially in the frontoethmoidal region, are a form of neural tube defect affecting patients in Southeast Asia more commonly than those in western countries, where they are more common in the occipital regions. All patients with classical frontoethmoidal encephalocele had swelling over the bridge of nose or inner canthus of eye since birth, with varying degrees of hypertelorism. This paper emphasizes on the clinical features of this pathology and its surgical management, along with, reviewing the evolution of single-stage correction and fine refinements.