\u3cem\u3eIn vitro\u3c/em\u3e Effect of Graphene Structures as an Osteoinductive Factor in Bone Tissue Engineering: A Systematic Review

Graphene and its derivatives have been well‐known as influential factors in differentiating stem/progenitor cells toward the osteoblastic lineage. However, there have been many controversies in the literature regarding the parameters effect on bone regeneration, including graphene concentration, size, type, dimension, hydrophilicity, functionalization, and composition. This study attempts to produce a comprehensive review regarding the given parameters and their effects on stimulating cell behaviors such as proliferation, viability, attachment and osteogenic differentiation. In this study, a systematic search of MEDLINE database was conducted for in vitro studies on the use of graphene and its derivatives for bone tissue engineering from January 2000 to February 2018, organized according to the PRISMA statement. According to reviewed articles, different graphene derivative, including graphene, graphene oxide (GO) and reduced graphene oxide (RGO) with mass ratio ≤1.5 wt % for all and concentration up to 50 μg/mL for graphene and GO, and 60 μg/mL for RGO, are considered to be safe for most cell types. However, these concentrations highly depend on the types of cells. It was discovered that graphene with lateral size less than 5 µm, along with GO and RGO with lateral dimension less than 1 µm decrease cell viability. In addition, the three‐dimensional structure of graphene can promote cell‐cell interaction, migration and proliferation. When graphene and its derivatives are incorporated with metals, polymers, and minerals, they frequently show promoted mechanical properties and bioactivity. Last, graphene and its derivatives have been found to increase the surface roughness and porosity, which can highly enhance cell adhesion and differentiation

Graphite/Gold Nanoparticles Electrode for Direct Protein Attachment: Characterization and Gas Sensing Application

In this work, graphite/gold nanoparticles (G/AuNPs) were synthesized through a facile chemical method, and its potential application for direct protein attachment for electrochemical detection of carbon monoxide (CO) was investigated. The preparation of G/AuNPs electrodes was optimized by synthesizing the nanoparticles in different concentration of HAuCl4.3H2O at various temperatures. The G/AuNPs electrode was subsequently modified by four types of mercaptopropionic acid, including 1-mercaptopropionic, 3-mercaptopropionic, 6-mercaptopropionic, and 11-mercaptopropionic acid, to achieve the best structure for protein attachment. Visible absorption and electrochemical studies showed that 3-mercaptopropionic acid possesses the best performance regarding the electrical conductivity between electrode and protein redox center. The cyclic voltammetry results revealed that the modified electrode has an appropriate performance for CO detection at very low concentrations while keeping a linear response. The limit of detection for the modified electrode was calculated to be about 0.2 ppb. Finally, the interactions of cytochrome C and carbon monoxides were simulated using molecular dynamics (MD), and the effect of protein conformation changes on the electrochemical signal was thoroughly examined. The simulation results suggested that the proposed electrochemical sensor has an acceptable performance for the detection of CO due to less fluctuation of amino acids near the protein chain in the presence of CO molecules

A Glassy Carbon Electrode Modified with Reduced Graphene Oxide and Gold Nanoparticles for Electrochemical Aptasensing Of Lipopolysaccharides from Escherichia Coli Bacteria

An electrochemical aptasensor is described for the voltammetric determination of lipopolysaccharide (LPS) from Escherichia coli 055:B5. Aptamer chains were immobilized on the surface of a glassy carbon electrode (GCE) via reduced graphene oxide and gold nanoparticles (RGO/AuNPs). Fast Fourier transform infrared, X-ray diffraction and transmission electron microscopy were used to characterize the nanomaterials. Cyclic voltammetry, square wave voltammetry and electrochemical impedance spectroscopy were used to characterize the modified GCE. The results show that the modified electrode has a good selectivity for LPS over other biomolecules. The hexacyanoferrate redox system, typically operated at around 0.3 V (vs. Ag/AgCl) is used as an electrochemical probe. The detection limit is 30 fg·mL−1. To decrease the electrochemical potential for detection of LPS, Mg/ carbon quantum dots were used as redox active media. They decrease the detection potentialto 0 V and the detection of limit (LOD) to 1 fg·mL−1. The electrode was successfully used to analyze serum of patients and healthy persons

Layer-By-Layer Assembly of Graphene Oxide on Thermosensitive Liposomes for Photo-Chemotherapy

Stimuli responsive polyelectrolyte nanoparticles have been developed for chemo-photothermal destruction of breast cancer cells. This novel system, called layer by layer Lipo-graph (LBL Lipo-graph), is composed of alternate layers of graphene oxide (GO) and graphene oxide conjugated poly (l-lysine) (GO-PLL) deposited on cationic liposomesencapsulating doxorubicin. Various concentrations of GO and GO-PLL were examined and the optimal LBL Lipo-graph was found to have a particle size of 267.9 ± 13 nm, zeta potentialof +43.9 ± 6.9 mV and encapsulation efficiency of 86.4 ± 4.7%. The morphology of LBL Lipo-graph was examined by cryogenic-transmission electron microscopy (Cryo-TEM), atomic force microcopy (AFM) and scanning electron microscopy (SEM). The buildup of LBL Lipo-graph was confirmed via ultraviolet-visible (UV–Vis) spectrophotometry, thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC) analysis. Infra-red (IR) response suggests that four layers are sufficient to induce a gel-to-liquid phase transition in response to near infra-red (NIR) laser irradiation. Light-matter interaction of LBL Lipo-graph was studied by calculating the absorption cross section in the frequency domain by utilizing Fourier analysis. Drug release assay indicates that the LBL Lipo-graph releases much faster in an acidic environment than a liposome control. A cytotoxicity assay was conducted to prove the efficacy of LBL Lipo-graph to destroy MD-MB-231 cells in response to NIR laser emission. Also, image stream flow cytometry and two photon microcopy provide supportive data for the potential application of LBL Lipo-graph for photothermal therapy. Study results suggest the novel dual-sensitive nanoparticles allow intracellular doxorubin delivery and respond to either acidic environments or NIR excitation. Statement of Significance Stimuli sensitive hybrid nanoparticles have been synthesized using a layer-by-layer technique and demonstrated for dual chemo-photothermal destruction of breast cancer cells. The hybrid nanoparticles are composed of alternating layers of graphene oxide and graphene oxide conjugated poly-l-lysine coating the surface of a thermosensitive cationic liposome containing doxorubicin as a core. Data suggests that the hybrid nanoparticles may offer many advantages for chemo-photothermal therapy. Advantages include a decrease of the initial burst release which may result in the reduction in systemic toxicity, increase in pH responsivity around the tumor environment and improved NIR light absorption

Electrochemical nanobiosensor based on reduced graphene oxide and gold nanoparticles for ultrasensitive detection of microRNA-128

Acute lymphoblastic leukemia (ALL) is one of the most prevalent cancers in children and microRNA-128 is amongst the most useful biomarkers not only for diagnosis of ALL, but also for discriminating ALL from acute myeloid leukemia (AML). In this study, a novel electrochemical nanobiosensor based on reduced graphene oxide (RGO) and gold nanoparticles (AuNPs) has been fabricated to detect miRNA-128. Cyclic Voltametery (CV), Square Wave Voltametery (SWV) and Electrochemical Impedance Spectroscopy (EIS) have been applied to characterize the nanobiosensor. Hexacyanoferrate as a label-free and methylene blue as a labeling material were used in the design of the nanobiosensors. It was found that the modified electrode has excellent selectivity and sensitivity to miR-128, with a limit of detection of 0.08761 fM in label-free and 0.00956 fM in labeling assay. Additionally, the examination of real serum samples of ALL and AML patients and control cases confirms that the designed nanobiosensor has the potential to detect and discriminate these two cancers and the control samples.Comunidad de Madri

The CRISPR growth spurt: from bench to clinic on versatile small RNAs

Clustered Regulatory Interspaced Short Palindromic Repeats (CRISPR) in association with CRISPR associated protein (Cas) is an adaptive immune system, playing a pivotal role in the defense of bacteria and archaea. Ease of handling and cost effectiveness make CRISPR-Cas system an ideal programmable nuclease tool. Recent advances in understanding the CRISPR-Cas system have tremendously improved its efficiency. For instance, it is possible to recapitulate the chronicle CRISPR-Cas from its infancy and inaugurate a developed version by generating novel variants of Cas proteins, subduing off-target effects and optimization of innovative strategies. In summary, CRISPR-Cas system could be employed in a number of applications including providing model systems, rectification of detrimental mutations, and antiviral therapies

Microfluidic synthesis of PLGA/carbon quantum dot microspheres for vascular endothelial growth factor delivery

In this study, vascular endothelial growth factor (VEGF) loaded poly(D,L-lactide-co-glycolide) (PLGA) – carbon quantum dot microspheres were produced using microfluidic platforms. The microcapsules were fabricated in flow-focusing geometry with a biphasic flow to generate solid/oil/water (s–o–w) droplets. To avoid any damage to protein functional and structural stability during the encapsulation process, the VEGF was PEGylated. The produced microspheres were intact and highly monodisperse in size (CV \u3c 5%). Furthermore, microspheres in a size range of 16–36 µm were achieved by adjusting the flow ratio parameter. The encapsulation efficiency, release profile, and bioactivity of the produced microparticles were also studied. The loading efficiency of PEGylated VEGF in the microparticles was varied from 51–69% and more than 90% of PEGylated VEGF was released within 28 days. Furthermore, the release of VEGF was indirectly monitored by carbon quantum dots. The present monodisperse and controllable VEGF loaded microspheres with reproducible manner could be widely used in tissue engineering and therapeutic applications

Microfluidic-Assisted Fabrication of Reverse Micelle/PLGA Hybrid Microspheres for Sustained Vascular Endothelial Growth Factor Delivery

In this study, poly (d, l-lactide-co-glycolide) (PLGA) composite microspheres containing anhydrous reverse micelle (R.M.) dipalmitoylphosphatidylcholine (DPPC) nanoparticles loaded vascular endothelial growth factor (VEGF) were produced using microfluidic platforms. The VEGF-loaded R.M. nanoparticles (VRM) were achieved by initial self-assembly and subsequent lipid inversion of the DPPC vesicles. The fabricated VRMs were encapsulated into the PLGA matrix by flow-focusing geometry microfluidic platforms. The encapsulation efficiency, in vitro release profile, and the bioactivity of the produced composite microspheres were investigated. The release study showed that VEGF was slowly released from the PLGA composite microspheres over 28 days with a reduced initial burst (18  ±  4.17% in the first 24 H). The VEGF stability during encapsulation and release period was also investigated, and the results indicated that encapsulated VEGF was well preserved. Also, the bioactivity assay of the PLGA composite microspheres on human umbilical vein endothelial cells was confirmed that the encapsulated VEGF was utterly active. The present monodisperse and controllable VEGF-loaded microspheres with reproducible manner could be widely used in tissue engineering and therapeutic applications

Protein-based nanobiosensor for direct detection of hydrogen sulfide

The chemically modified cytochrome c from equine heart, EC (232-700-9), was immobilized onto gold nanoparticles in order to develop a specific biosensing system for monitoring hydrogen sulfide down to the micromolar level, by means of a localized surface plasmon resonance spectroscopy. The sensing mechanism is based on the cytochrome-c conformational changes in the presence of H2S which alter the dielectric properties of the gold nanoparticles and the surface plasmon resonance peak undergoes a redshift. According to the experiments, it is revealed that H2S can be detected at a concentration of $4.0\ \mu \text{M}\ (1.3\ \text{ppb})$ by the fabricated biosensor. This simple, quantitative and sensitive sensing platform provides a rapid and convenient detection for H2S at concentrations far below the hazardous limit

Cancer cell detection using electrochemical nanobiosensor based on graphene / gold nanoparticle

Introduction: Nowadays, early cancer detection and effective treatment is crucial for improved prognosis and cancer management. In particular, the accurate qualitative detection of cancer cells represents a critical step in cancer diagnosis. The aim of this study was to examine Cancer cell detection using electrochemical nanobiosensor based on graphene / gold nanoparticle. Materials and Methods: Modified graphene oxide/gold nanoparticle electrodes were employed to increase the sensitivity of human breast cancer MCF-7 cells detection, using CD44 biomarker. Frist the electrodes were modified with graphene, then gold nanoparticles were sediment on graphene-modified electrode. Then CD44 monoclonal antibody conjugated on the surface of gold nanoparticles, on graphene-modified electrode. Finally, the performance of the fabricated biosensors were investigated by using a common reference electrode composed of silver-silver chloride and a common platinum counter electrode at different antigen concentrations with the buffer and serum. Results: The proposed electrochemical cytosensor delivered a high sensitivity with the average of 1.12 μA / cells ml-1, and a low detection limit of 6 cells. Conclusion: These results indicate that the cytosensor has great potential in diagnosis of cancer