Breaking of the Phosphodiester Bond: A Key Factor That Induces Hemolysis

In-depth understanding the toxicity of nanomaterials in red blood cells (RBCs) is of great interest, because of the importance of RBCs in transporting oxygen in blood circulation. Although the toxic effects of nanoparticles in RBCs have been revealed, the conclusions from the literature are conflicting, and in particular, the toxic mechanism is still at the infant stage. Herein, we investigated the size-dependent toxicity of well-known CdTe semiconductor quantum dots (QDs) and revealed the exact toxic mechanism at the molecular level by confocal microscopy and Fourier transform infrared (FT-IR) spectroscopy techniques. We found that smaller mercaptosuccinic acid-capped CdTe QDs (MSA-QDs) with the green-emitting color could cause hemagglutination whereas the middle-size yellow-emitting MSA-QDs induced the formation of stomatocytes and echinocytes and the bigger size red-emitting MSA-QDs induced heavy hemolysis and the formation of lots of ghost cells. The FT-IR data proved that all the MSA-QDs were likely to bond to the RBCs membranes and caused the structural changes of lipid and protein in RBCs. But only the red-emitting MSA-QDs caused the breakage of the phosphodiester bond, which might cause the heavy hemolysis. To some extent, this is the first example that reveals the hemolysis mechanism at the molecular level

Size-Dependent Stability of Water-Solubilized CdTe Quantum Dots and Their Uptake Mechanism by Live HeLa Cells

Water-solubilized quantum dots have led to a promising application in cellular labeling and biological imaging. The physicochemical properties of water-solubilized quantum dots, particularly in a physiological environment, are strongly dependent on their size. In this paper, we systematically studied the stability of mercaptosuccinic acid-coated CdTe quantum dots (MSA-QDs) of about 2.3 and 5.4 nm diameters in various buffers with different pH values and under laser irradiation by fluorescence spectroscopy. It was found that larger MSA-QDs showed better stability. Size-dependent uptake of MSA-QDs by living HeLa cells was further investigated by confocal microscopy. In phosphate buffer solution, the larger MSA-QDs entered the cells mainly by endocytosis, and part of the smaller ones entered the cells by passive penetration. In cell culture medium, their uptake pathways could be changed due to the changes of their surface properties. The cytotoxicity of smaller and larger MSA-QDs was significantly decreased due to the adsorption of some biological components in the cell culture medium on the nanoparticles surface

A Facile One-Pot Synthesis of Copper Sulfide-Decorated Reduced Graphene Oxide Composites for Enhanced Detecting of H₂O₂ in Biological Environments

The high levels of H₂O₂ are closely associated with cancer and progressive neurodegenerative diseases, such as Parkinson’s disease. In this study, we developed a novel CuS nanoparticle-decorated reduced graphene oxide-based electrochemical biosensor for the reliable detection of H₂O₂. The new electrocatalyst, CuS/RGO composites was successfully prepared by heating the mixture of CuCl₂ and Na₂S aqueous solutions in the presence of PVP-protected graphene oxide at 180 °C. A potential application of CuS/RGO composite-modified electrode as a biosensor to monitor H₂O₂ has been investigated. The steady-state current response increases linearly with H₂O₂ concentration from 5 to 1500 μM with a fast response time of less than 2 s. The detection limit (3σ) for determination of H₂O₂ has been estimated to be 0.27 μM, which was lower than certain enzymes and noble metal nanomaterial-based biosensors. In addition, the study of storage time on the amperometric response of the sensor indicates super stability. Due to these remarkable analytical advantages, the as-made sensor was applied to determine the H₂O₂ levels in human serum and urine samples and H₂O₂ released from human cervical cancer cells with satisfactory results. These results demonstrate that this new nanocomposite with the high surface area and electrocatalytic activity is a promising candidate for use as an enhanced electrochemical sensing platform in the design of nonenzymatic biosensors

Monitoring of the Enzymatic Degradation of Protein Corona and Evaluating the Accompanying Cytotoxicity of Nanoparticles

Established nanobio interactions face the challenge that the formation of nanoparticle–protein corona complexes shields the inherent properties of the nanoparticles and alters the manner of the interactions between nanoparticles and biological systems. Therefore, many studies have focused on protein corona-mediated nanoparticle binding, internalization, and intracellular transportation. However, there are a few studies to pay attention to if the corona encounters degradation after internalization and how the degradation of the protein corona affects cytotoxicity. To fill this gap, we prepared three types of off/on complexes based on gold nanoparticles (Au NPs) and dye-labeled serum proteins and studied the extracellular and intracellular proteolytic processes of protein coronas as well as their accompanying effects on cytotoxicity through multiple evaluation mechanisms, including cell viability, adenosine triphosphate (ATP) content, mitochondrial membrane potential (MMP), and reactive oxygen species (ROS). The proteolytic process was confirmed by recovery of the fluorescence of the dye-labeled protein molecules that was initially quenched by Au NPs. Our results indicate that the degradation rate of protein corona is dependent on the type of the protein based on systematical evaluation of the extracellular and intracellular degradation processes of the protein coronas formed by human serum albumin (HSA), γ-globulin (HGG), and serum fibrinogen (HSF). Degradation is the fastest for HSA corona and the slowest for HSF corona. Notably, we also find that the Au NP-HSA corona complex induces lower cell viability, slower ATP production, lower MMP, and higher ROS levels. The cytotoxicity of the nanoparticle-protein corona complex may be associated with the protein corona degradation process. All of these results will enrich the database of cytotoxicity induced by nanomaterial-protein corona complexes

Influence of the Molecular Structure of Carboxyl-Terminated Self-Assembled Monolayer on the Electron Transfer of Cytochrome c Adsorbed on an Au Electrode: In Situ Observation by Surface-Enhanced Infrared Absorption Spectroscopy

Surface-enhanced infrared adsorption spectroscopy (SEIRAS) was employed for the in situ observation of structural changes that occur in a self-assembled monolayer (SAM) of 11-mercaptoundecanoic acid (MUA) bound on a gold surface. The observed SEIRA spectra reveal a deprotonation of the carboxyl head group of the MUA-SAM layer after adsorption. An analysis of the vibrational spectra suggests that the deprotonation process occurs when the adsorbed MUA molecules reach a critical mutual distance. MUA-SAMs promote direct electron transfer between the metal electrode and cytochrome c, the electron mediator between the integral membrane protein complexes of the respiratory chain. The results show that the coverage of cytochrome c increases with the coverage of deprotonated MUA on the surface. On the other hand, the electron transfer of cytochrome c is optimized only when a moderate amount of the carboxyl head group is deprotonated. The electron transfer of cytochrome c is suppressed with a further increase of the deprotonated MUA. The relationship between the surface structure of the MUA layer and the electron transfer of cytochrome c is discussed on the basis of the spectroscopic data

Revealing the Nature of Interaction between Graphene Oxide and Lipid Membrane by Surface-Enhanced Infrared Absorption Spectroscopy

Revealing the nature of interaction between graphene oxide (GO) and lipid membrane is a crucial issue but still remains challenging. Here, we describe our recent effort toward this direction by studying the GO-induced vibrational changes of interfacial water and lipid membrane with surface-enhanced infrared absorption (SEIRA) spectroscopy. The experimental results provide evidence that overcoming the electrostatic repulsion of phosphate group, its hydrogen bonding attraction as well as the electrostatic and hydrophobic interaction of choline group are the driving forces for the effective adsorption of GO on lipid membrane. This work will open exciting new avenues to explore the use of SEIRA spectroscopy technique in probing nanobio interface

Molecular Impact of the Membrane Potential on the Regulatory Mechanism of Proton Transfer in Sensory Rhodopsin II

Metabolism establishes a potential difference across the cell membrane of every living cell which drives and regulates secondary ion and solute transfer across membrane proteins. Unraveling the effect of the membrane potential on the level of single molecular groups of the membrane protein was long hampered by the lack of appropriate analytical techniques. We have developed Surface Enhanced Infrared Difference Absorption Spectroscopy (SEIDAS), a highly sensitive vibrational technique for surface analysis, for the study of solid-supported monolayers of orientated membrane proteins. Here, we present spectroscopic data on vibrational changes of sensory rhodopsin II from Natronomonas pharaonis (NpSR II). The application of the electrode potential provides a voltage drop across the NpSR II monolayer through the Helmholtz double layer that mimics the cellular membrane potential. IR difference spectra indicated a shift of the photostationary equilibrium from an M and O mixture toward an M dominant equilibrium. The shift of positive to negative potential exhibited similar effects on the light-induced SEIDA spectra as the increase in pH. This effect is explained in terms of local pH change raised by the compensation of excess charge from the electrode. As we have shown earlier (Jiang, et al. Proc. Natl. Acad. Sci. U.S.A. 2008, 105 (34), 12113−12117), the application of an electric field opposite to the physiological proton transfer from the retinal Schiff base to its counterion Asp75 leads to the selective halt of the latter. However, when the solution pH is much higher than 5.8, that is, when the proton releasing group at the extracellular side is ionized, proton transfer of Asp75 becomes insensitive to the electric field exerted by the electrode. We infer that the deprotonation of the proton release group creates a local polar environment surrounding Asp75 as a consequence of hydrogen-bonding rearrangements that exceeds the energy of the external dipole. Our results reveal a molecular model for the physiological regulation of the photocycle of NpSR II by the potential drop across the membrane which came about by the interplay between the change in local pH at the membrane surface and the external electric field

Impact of Shape and Pore Size of Mesoporous Silica Nanoparticles on Serum Protein Adsorption and RBCs Hemolysis

With the rapid development of nanotechnology, mesoporous silica nanoparticles (MSNs) with numerous forms and structures have been synthesized and extensively applied in biomedicine in the past decades. However, our knowledge about the biocompatibility of the developed MSNs has not matched their development. Therefore, in this work, we have synthesized sphere-shaped MSNs with different pore scales (<i>s</i>-SPs and <i>l</i>-SPs) and rod-shape (RPs-3) MSNs to evaluate the influence of the morphology and pore size on their interaction with serum proteins and red blood cells (RBCs). The adsorption of human albumin (HSA), globulin (HGG), and fibrinogen (HSF) onto different kinds of MSNs has been analyzed by pseudo second-order kinetic model, and the conformational changes of the adsorbed proteins have been studied by FTIR spectroscopy. We find that the conformation of absorbed HSA and HSF, while not HGG, will be affected by the pore size and morphology of the MSNs. The conformational changes of the adsorbed proteins will further affect their saturated adsorption capacity. However, the initial adsorption rate is only determined by the property of MSNs and proteins. Additional hemolysis assay shows that the pore size and morphology of the MSNs will also affect their hemolytic activity in RBCs which will be extremely depressed by the formation of protein corona. These systematic studies will provide an overall understanding in the blood compatibility of MSNs as well as useful guidelines for fabrication of blood-compatible nanomaterials