Sanjeev Kumar Mahto

Assessment of cytocompatibility of surface-modified CdSe/ZnSe quantum dots for BALB/3T3 fibroblast cells

With the widespread use of quantum dots (QDs), the likelihood of exposure to QDs has been assumed to have increased substantially. Recently, QDs have been employed in numerous biological and medical applications. However, there is a lack of toxicological data pertaining to QDs. In this study, we aimed to investigate the cytocompatibility of surface-modified CdSe/ZnSe QDs for BALB/3T3 fibroblast cells. The ligands used for surface modification are mercaptopropionic acid (MPA) and Gum arabic (GA)/tri-n-octylphosphine oxide (TOPO). Cells were exposed to different concentrations of QDs followed by illustrative cytotoxicity analyses. Furthermore, we used a confocal microscope to assess intracellular uptake of QDs. Confocal images showed that MPA-coated QDs were distributed inside the cytoplasmic region of cells. In contrast, GA/TOPO-coated QDs were not found inside cells. MPA-coated QDs were highly cytocompatible, whereas GA/TOPO-coated QDs were toxic to the cells. Cells treated with GA/TOPO-coated QDs showed altered morphology, decreased viability, significant concentrations of intracellular free cadmium, detectable reactive oxygen species (ROS) formation, depolymerized cytoskeleton, and irregular-shaped nuclei. This study suggests that surface modification by ligands plays a significant role in the prevention of cytotoxicity of QDs.

A new perspective on in vitro assessment method for evaluating quantum dot toxicity by using microfluidics technology.

In this study, we demonstrate a new perspective on in vitro assessment method for evaluating quantum dot (QD) toxicity by using microfluidics technology. A new biomimetic approach, based on the flow exposure condition, was applied in order to characterize the cytotoxic potential of QD. In addition, the outcomes obtained from the flow exposure condition were compared to those of the static exposure condition. An in vitro cell array system was established that used an integrated multicompartmented microfluidic device to develop a sensitive flow exposure condition. QDs modified with cetyltrimethyl ammonium bromide∕trioctylphosphine oxide were used for the cytotoxicity assessment. The results suggested noticeable differences in the number of detached and deformed cells and the viability percentages between two different exposure conditions. The intracellular production of reactive oxygen species and release of cadmium were found to be the possible causes of QD-induced cytotoxicity, irrespective of the types of exposure condition. In contrast to the static exposure, the flow exposure apparently avoided the gravitational settling of particles and probably assisted in the homogeneous distribution of nanoparticles in the culture medium during exposure time. Moreover, the flow exposure condition resembled in vivo physiological conditions very closely, and thus, the flow exposure condition can offer potential advantages for nanotoxicity research.

Microfluidic platforms for advanced risk assessments of nanomaterials

Abstract In the past few years, promising efforts to utilize microfabrication-based technologies have laid the foundation for developing advanced, and importantly more physiologically-realistic, microfluidic methods for risk assessment of engineered nanomaterials (ENMs). In the present review, we discuss the wave of recent developments using microfluidic-based in vitro models and platforms for nanotoxicological assays, such as determination of cell viability, cellular dose, oxidative stress and nuclear damage. Here, we specifically highlight the tangible advantages of microfluidic devices in providing promising tools to tackle many of the current and ongoing challenges faced with traditional toxicology assays. Most importantly, microfluidic technology not only allows to recreate physiologically-relevant in vitro models for nanotoxicity examinations, but also provides platforms that deliver an attractive strategy towards improved control over applied ENM doses. In a final step, we present examples of state-of-the-art microfluidic platforms for in vitro assessment of potential adverse ENM effects.

Pharmacokinetics, biodistribution, in vitro cytotoxicity and biocompatibility of Vitamin E TPGS coated trans resveratrol liposomes

The clinical application of trans resveratrol (RSV) in glioma treatment is largely limited because of its rapid metabolism, fast elimination from systemic circulation and low biological half life. Therefore, the objectives of this study were to enhance the circulation time, biological half life and passive brain targeting of RSV using d-α-tocopheryl polyethylene glycol 1000 succinate (TPGS) coated liposomes (RSV-TPGS-Lipo). In addition to basic liposomal characterizations, in vitro anticancer potential against C6 glioma cell lines and cellular internalization of liposomes were carried out by MTT assay and confocal laser scanning microscopy (CLSM), respectively. Pharmacokinetics and tissue distribution studies were also carried out after intravenous administration in Charles Foster rats. RSV-TPGS-Lipo 2 showed significantly higher cytotoxicity than RSV-Lipo (uncoated liposomes) and RSV. Both uncoated and TPGS coated liposomes showed excellent cellular uptake. RSV, RSV-Lipo and RSV-TPGS-Lipo 2 were found to be haemocompatible and safe after i.v. administration. Area under the curve (AUC) and plasma half life (t1/2) after i.v. administration of RSV-TPGS-Lipo 2 was found to be approximately 5.73 and 6.72 times higher than that of RSV-Lipo as well as 29.94 and 29.66 times higher than that of RSV, respectively. Thus, the outcome indicates that RSV-TPGS-Lipo 2 is a promising carrier for glioma treatment with improved pharmacokinetic parameters. Moreover, brain accumulation of RSV-Lipo and RSV-TPGS-Lipo 2 was found to be significantly higher than that of RSV (P<0.05). Results are suggesting that both RSV-Lipo and RSV-TPGS-Lipo 2 are the promising tools of RSV for the treatment of brain cancer.

Multicompartmented microfluidic device for characterization of dose-dependent cadmium cytotoxicity in BALB/3T3 fibroblast cells

This paper describes the development of a miniaturized multicompartmented microfluidic device for high-throughput cell cytotoxicity assays and its applicability to the investigation of cadmium-induced cytotoxicity. A steady gradient of cadmium was generated inside the compartments to study the effects of cadmium ion on BALB/3T3 fibroblast cells in a dose-dependent fashion. The device allowed the performance of multiplexed assays to probe the dosage effect of cadmium, morphological alterations of live cells, regulation of proliferation and viability of cells, determination of reactive oxygen species, mechanisms of cell death, i.e. apoptosis and/or necrosis, and immunocytochemical staining of cells in parallel and/or serially, or on a single population simultaneously. The outcomes of all the microfluidic assays were compared to conventional plates-based cytotoxicity assays. The results indicated that the cells cultured in this device were morphologically healthy with greater than 90% viability. They further suggested that the basic mode of cell death behind cadmium-induced cytotoxicity was apoptosis, which was regulated by intracellular oxidative stress via cytoskeleton disorganization and nuclear condensation. Such microenvironments resemble the in vivo physiological conditions very closely and thus offer a unique platform for more accurate observations of cytotoxicity assays and more precise estimation of the IC50 value in comparison to conventional analytical assays.

Respiratory Physiology on a Chip

Our current understanding of respiratory physiology and pathophysiological mechanisms of lung diseases is often limited by challenges in developing in vitro models faithful to the respiratory environment, both in cellular structure and physiological function. The recent establishment and adaptation of microfluidic-based in vitro devices (μFIVDs) of lung airways have enabled a wide range of developments in modern respiratory physiology. In this paper, we address recent efforts over the past decade aimed at advancing in vitro models of lung structure and airways using microfluidic technology and discuss their applications. We specifically focus on μFIVDs covering four major areas of respiratory physiology, namely, artificial lungs (AL), the air-liquid interface (ALI), liquid plugs and cellular injury, and the alveolar-capillary barrier (ACB).

Intravenous administration of trans-resveratrol-loaded TPGS-coated solid lipid nanoparticles for prolonged systemic circulation, passive brain targeting and improved in vitro cytotoxicity against C6 glioma cell lines

trans-Resveratrol (RSV), a natural molecule isolated from red wine, is widely known for several therapeutic potentials. RSV is proved for cardioprotective, vasodilation, anti-inflammatory, and anticancer effects. Recently, anticancer potential against glioma cells has also been reported. However, the clinical application of RSV in glioma treatment is largely limited because of its rapid metabolism and elimination from systemic circulation thereby exhibiting low biological half-life and poor brain distribution as well. Therefore, the main objective of this study was to enhance the circulation time, biological half-life and passive brain targeting of RSV using D-α-tocopheryl polyethylene glycol 1000 succinate (TPGS)-coated solid lipid nanoparticles (RSV-TPGS-SLN). RSV-TPGS-SLN formulations were prepared by a solvent emulsification evaporation method and evaluated for several nanoparticulate characteristics. In vitro anticancer potential and cellular internalization of nanoparticles were also investigated in C6 glioma cell lines. Pharmacokinetics and biodistribution studies were carried out following intravenous administration in healthy Charles Foster rats. RSV-TPGS-SLN showed significantly higher in vitro cytotoxicity against C6 glioma cell lines and excellent cellular internalization. RSV-TPGS-SLN showed 11.12 and 9.37 times higher area under the curve and plasma half-life than RSV solution, respectively. Moreover, brain distribution of RSV-TPGS-SLN was found to be 9.23 times higher in comparison to that of RSV alone. Thus, we anticipate that the RSV-TPGS-SLN formulation can be applied as a potential tool for improving circulation time, biological half-life and passive brain targeting of RSV, thereby being immensely useful in the treatment of glioma.

Chemical modification of poly(vinyl chloride) for blood and cellular biocompatibility

Poly(vinyl chloride) (PVC) was modified with three different ionomers including thiosulphate, thiourea and sulphite for improving the biocompatibility of the polymer. All ionomers were prepared by nucleophilic substitution using a phase transfer catalyst method. The modified forms of PVC were characterized using ultraviolet-visible (UV-Vis) spectroscopy, Fourier Transform Infrared (FTIR) spectroscopy, scanning electron microscopy (SEM) and thermal gravimetric analysis (TGA). They were found to be less stable thermally compared to the untreated polymer. The biocompatibility of the polymers was evaluated by assessing their wettability via contact angle measurements and by performing hemolysis and thrombogenicity assays. Their cellular biocompatibility was evaluated by assessing their adhesion and proliferation, and by carrying out cytotoxicity assays and nuclear staining. The results reveal that modification of the polymer with the specified ionomers significantly enhances the bio- and blood-compatibility properties.

Microfluidic shear stress-regulated surfactant secretion in alveolar epithelial type II cells in vitro

We investigated the role of flow-induced shear stress on the mechanisms regulating surfactant secretion in type II alveolar epithelial cells (ATII) using microfluidic models. Following flow stimulation spanning a range of wall shear stress (WSS) magnitudes, monolayers of ATII (MLE-12 and A549) cells were examined for surfactant secretion by evaluating essential steps of the process, including relative changes in the number of fusion events of lamellar bodies (LBs) with the plasma membrane (PM) and intracellular redistribution of LBs. F-actin cytoskeleton and calcium levels were analyzed in A549 cells subjected to WSS spanning 4-20 dyn/cm(2). Results reveal an enhancement in LB fusion events with the PM in MLE-12 cells upon flow stimulation, whereas A549 cells exhibit no foreseeable changes in the monitored number of fusion events for WSS levels ranging up to a threshold of ∼8 dyn/cm(2); above this threshold, we witness instead a decrease in LB fusion events in A549 cells. However, patterns of LB redistribution suggest that WSS can potentially serve as a stimulus for A549 cells to trigger the intracellular transport of LBs toward the cell periphery. This observation is accompanied by a fragmentation of F-actin, indicating that disorganization of the F-actin cytoskeleton might act as a limiting factor for LB fusion events. Moreover, we note a rise in cytosolic calcium ([Ca(2+)]c) levels following stimulation of A549 cells with WSS magnitudes ranging near or above the experimental threshold. Overall, WSS stimulation can influence key components of molecular machinery for regulated surfactant secretion in ATII cells in vitro.

Tailored Chemical Properties of 4-Arm Star Shaped Poly(d,l-lactide) as Cell Adhesive Three-Dimensional Scaffolds

Biodegradable poly(lactic acid) (PLA) is widely used to fabricate 3D scaffolds for tissue regeneration. However, PLA lacks cell adhering functional moieties, which limit its successful application in tissue engineering. Herein, we have tailored the cell adhesive properties of star shaped poly(d,l-lactide) (ss-PDLLA) by grafting gelatin to their 4 arms. Grafting of gelatin on PDLLA backbone was confirmed by 1H NMR and FTIR. The synthesized star shaped poly(d,l-lactide)-b-gelatin (ss-pLG) exhibited enhanced wettability and protein adsorption. The modification also facilitated better cell adhesion and proliferation on their respective polymer coated 2D substrates, compared to their respective unmodified ss-PDLLA. Further, 3D scaffolds were fabricated from gelatin grafted and unmodified polymers. The fabricated scaffolds were shown to be cytocompatible to 3T3-L1 cells and hemocompatible to red blood cells (RBCs). Cell proliferation was increased up to 2.5-fold in ss-pLG scaffolds compared to ss-PDLLA scaffolds. Furthermore, a significant increase in cell number reveals a high degree of infiltration of cells into the scaffolds, forming a viable and healthy 3D interconnected cell community. In addition to that, burst release of docetaxal (DTX) was observed from ss-pLG scaffolds. Hence, this new system of grafting polymers followed by fabricating 3D scaffolds could be utilized as a successful approach in a variety of applications where cell-containing depots are used.