Cooperative regulation of adherens junction expansion through epidermal growth factor receptor activation.

The mechanisms controlling the dynamics of expansion of adherens junctions are significantly less understood than those controlling their static properties. Here, we report that for suspended cell aggregates, the time to form a new junction between two cells speeds up with the number of junctions that the cells are already engaged in. Upon junction formation, the activation of epidermal growth factor receptor (EGFR) distally affects the actin turnover dynamics of the free cortex of the cells. The 'primed' actin cortex results in a faster expansion of the subsequent new junctions. In such aggregates, we show that this mechanism results in a cooperative acceleration of the junction expansion dynamics (kinetype) but does not alter the cell contractility, and hence the final junction size (phenotype). This article has an associated First Person interview with the first author of the paper.

Co-culture of infrapatellar fat pad-derived mesenchymal stromal cells and articular chondrocytes in plasma clot for cartilage tissue engineering.

BACKGROUND: Cell source plays a deterministic role in defining the outcome of a cell-based cartilage regenerative therapy and its clinical translational ability. Recent efforts in the direction of co-culture of two or more cell types attempt to combine the advantages of constituent cell types and negate their demerits. METHODS: We examined the potential of co-culture of infrapatellar fat pad-derived mesenchymal stromal cells (IFP MSCs) and articular chondrocytes (ACs) in plasma clots in terms of their ratios and culture formats for cartilage tissue engineering. RESULTS AND DISCUSSION: It was observed that IFP MSCs and ACs interact positively to produce a better quality hyaline cartilage-like matrix. While a supra-additive deposition of sulfated Glycosaminoglycans (sGAG), collagen type II, aggrecan and link protein was observed, deposition of collagen type I and X was sub-additive. (Immuno)-histologically similar cartilage was generated in vitro in IFP MSC:AC ratio of 50:50 and pure AC groups thus yielding a hyaline cartilage with 50% reduced requirement of ACs. Subsequently, we investigated if this response could be improved further by enabling better cell-cell interactions using scaffold-free systems such as self-assembled cartilage or by encapsulating cellular micro-aggregates in plasma clot. However, it was inferred that while self-assembly may have enabled better cell-cell interaction, poor cell survival negated its overall beneficial role, whereas the micro-aggregate group demonstrated highly heterogeneous matrix deposition within the construct, thus diminishing its translational utility. Overall, it was concluded that co-culture of IFP MSCs and ACs at a ratio of 50:50 within plasma clots demonstrated potential for cell-based cartilage regenerative therapy.

Learning Enriched Features for Fast Image Restoration and Enhancement.

Given a degraded input image, image restoration aims to recover the missing high-quality image content. Numerous applications demand effective image restoration, e.g., computational photography, surveillance, autonomous vehicles, and remote sensing. Significant advances in image restoration have been made in recent years, dominated by convolutional neural networks (CNNs). The widely-used CNN-based methods typically operate either on full-resolution or on progressively low-resolution representations. In the former case, spatial details are preserved but the contextual information cannot be precisely encoded. In the latter case, generated outputs are semantically reliable but spatially less accurate. This paper presents a new architecture with a holistic goal of maintaining spatially-precise high-resolution representations through the entire network, and receiving complementary contextual information from the low-resolution representations. The core of our approach is a multi-scale residual block containing the following key elements: (a) parallel multi-resolution convolution streams for extracting multi-scale features, (b) information exchange across the multi-resolution streams, (c) non-local attention mechanism for capturing contextual information, and (d) attention based multi-scale feature aggregation. Our approach learns an enriched set of features that combines contextual information from multiple scales, while simultaneously preserving the high-resolution spatial details. Extensive experiments on six real image benchmark datasets demonstrate that our method, named as MIRNet-v2, achieves state-of-the-art results for a variety of image processing tasks, including defocus deblurring, image denoising, super-resolution, and image enhancement. The source code and pre-trained models are available at https://github.com/swz30/MIRNetv2.

Isolation, Expansion, and Differentiation of Mesenchymal Stem Cells from the Infrapatellar Fat Pad of the Goat Stifle Joint.

The IFP, present in the knee joint, serves as a promising source of MSCs. The IFP is an easily accessible tissue as it is routinely resected and discarded during arthroscopic procedures and knee replacement surgeries. Additionally, its removal is associated with minimal donor site morbidity. Recent studies have demonstrated that IFP-MSCs do not lose their proliferation capacity during in vitro expansion and have age-independent osteogenic differentiation potential. IFP-MSCs possess superior chondrogenic differentiation potential compared to bone marrow-derived MSCs (BMSCs) and adipose-derived stem cells (ADSCs). Although these cells are easily obtainable from aged and diseased patients, their effectiveness is limited. Hence, using IFP-MSCs from healthy donors is important to determine their efficacy in biomedical applications. As access to a healthy human donor is challenging, animal models could be a better alternative to enable fundamental understanding. Large animals such as dogs, horses, sheep, and goats play a crucial role in translational research. Amongst these, the goat could be a preferred model since the stifle joint of the goat has the closest anatomy to the human knee joint. Moreover, goat-IFP can fulfill the higher MSC numbers needed for tissue regeneration applications. Furthermore, low cost, availability, and compliance with the 3R principles for animal research make them an attractive model. This study demonstrates a simple protocol for isolating IFP-MSCs from the stifle joint of goats and in vitro culture conditions for their expansion and differentiation. The aseptically isolated IFP from the goat was washed, minced, and digested enzymatically. After filtration and centrifugation, the collected cells were cultured. These cells were adherent, had MSCs-like morphology, and demonstrated remarkable clonogenic ability. Further, they differentiated into adipogenic, chondrogenic, and osteogenic lineages, demonstrating their multipotency. In conclusion, the study demonstrates the isolation and expansion of MSCs, which show potential in tissue engineering and regenerative medicine applications.

In vivo cartilage regeneration in a multi-layered articular cartilage architecture mimicking scaffold.

AIMS: Extracellular matrix (ECM) and its architecture have a vital role in articular cartilage (AC) structure and function. We hypothesized that a multi-layered chitosan-gelatin (CG) scaffold that resembles ECM, as well as native collagen architecture of AC, will achieve superior chondrogenesis and AC regeneration. We also compared its in vitro and in vivo outcomes with randomly aligned CG scaffold. METHODS: Rabbit bone marrow mesenchymal stem cells (MSCs) were differentiated into the chondrogenic lineage on scaffolds. Quality of in vitro regenerated cartilage was assessed by cell viability, growth, matrix synthesis, and differentiation. Bilateral osteochondral defects were created in 15 four-month-old male New Zealand white rabbits and segregated into three treatment groups with five in each. The groups were: 1) untreated and allogeneic chondrocytes; 2) multi-layered scaffold with and without cells; and 3) randomly aligned scaffold with and without cells. After four months of follow-up, the outcome was assessed using histology and immunostaining. RESULTS: In vitro testing showed that the secreted ECM oriented itself along the fibre in multi-layered scaffolds. Both types of CG scaffolds supported cell viability, growth, and matrix synthesis. In vitro chondrogenesis on scaffold showed an around 400-fold increase in collagen type 2 (COL2A1) expression in both CG scaffolds, but the total glycosaminoglycan (GAG)/DNA deposition was 1.39-fold higher in the multi-layered scaffold than the randomly aligned scaffold. In vivo cartilage formation occurred in both multi-layered and randomly aligned scaffolds treated with and without cells, and was shown to be of hyaline phenotype on immunostaining. The defects treated with multi-layered + cells, however, showed significantly thicker cartilage formation than the randomly aligned scaffold. CONCLUSION: We demonstrated that MSCs loaded CG scaffold with multi-layered zonal architecture promoted superior hyaline AC regeneration.Cite this article: Bone Joint Res 2020;9(9):601-612.

Ageing and the natural history of dry eye disease: A prospective registry-based cross-sectional study.

PURPOSE: To investigate the impact of ageing on ocular surface parameters, and empirically determine optimal prognostic cut-off ages for clinical markers of dry eye disease, aqueous tear deficiency, and meibomian gland dysfunction. METHODS: A total of 1331 community residents (785 females, 546 males; mean ± SD age, 38 ± 19 years) were recruited in a prospective registry-based cross-sectional study. Dry eye symptomology, ocular surface characteristics, and tear film quality were evaluated for each participant within a single clinical session, in accordance with the global consensus recommendations of the TFOS DEWS II reports. RESULTS: Multivariate regression analysis demonstrated positive associations between ageing and clinical markers of dry eye disease (all p ≤ 0.001). The Youden-optimal prognostic cut-off ages for signs of meibomian gland dysfunction occurred during the third decade of life (24-29 years); the optimal predictive ages for lid wiper epitheliopathy, tear film instability, hyperosmolarity, and dry eye symptoms occurred during the fourth decade of life (31-38 years); while the optimal prognostic thresholds for signs of aqueous tear deficiency and ocular surface staining occurred in the fifth and sixth decades of life (46-52 years). CONCLUSIONS: Advancing age is a significant risk factor for dry eye disease, which represents a growing public health concern with the ageing population worldwide. Signs of meibomian gland dysfunction appeared earlier in the natural history of disease progression, and the brief delay prior to the development of other clinical dry eye signs might represent a window of opportunity for preventative interventions in the young adult age group.

Two high-yield complementary methods to sort cell populations by their 2D or 3D migration speed.

The potential to migrate is one of the most fundamental functions for various epithelial, mesenchymal, and immune cells. Image analysis of motile cell populations, both primary and cultured, typically reveals an intercellular variability in migration speeds. However, cell migration chromatography, the sorting of large populations of cells based on their migratory characteristics, cannot be easily performed. The lack of such methods has hindered our understanding of the direct correlation between the capacity to migrate and other cellular properties. Here, we report two novel, easily implementable and readily scalable methods to sort millions of live migratory cancer and immune cells based on their spontaneous migration in two-dimensional and three-dimensional microenvironments, respectively. Correlative downstream transcriptomic, molecular and functional tests reveal marked differences between the fast and slow subpopulations in patient-derived cancer cells. We further employ our method to reveal that sorting the most migratory cytotoxic T lymphocytes yields a pool of cells with enhanced cytotoxicity against cancer cells. This phenotypic assay opens new avenues for the precise characterization of the mechanisms underlying hither to unexplained heterogeneities in migratory phenotypes within a cell population, and for the targeted enrichment of the most potent migratory leukocytes in immunotherapies.

Single-cell analysis of EphA clustering phenotypes to probe cancer cell heterogeneity.

The Eph family of receptor tyrosine kinases is crucial for assembly and maintenance of healthy tissues. Dysfunction in Eph signaling is causally associated with cancer progression. In breast cancer cells, dysregulated Eph signaling has been linked to alterations in receptor clustering abilities. Here, we implemented a single-cell assay and a scoring scheme to systematically probe the spatial organization of activated EphA receptors in multiple carcinoma cells. We show that cancer cells retain EphA clustering phenotype over several generations, and the degree of clustering reported for migration potential both at population and single-cell levels. Finally, using patient-derived cancer lines, we probed the evolution of EphA signalling in cell populations that underwent metastatic transformation and acquisition of drug resistance. Taken together, our scalable approach provides a reliable scoring scheme for EphA clustering that is consistent over multiple carcinomas and can assay heterogeneity of cancer cell populations in a cost- and time-effective manner.

Author Correction: Single-cell analysis of EphA clustering phenotypes to probe cancer cell heterogeneity.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Author Correction: Single-cell analysis of EphA clustering phenotypes to probe cancer cell heterogeneity.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Procurement, storage and utilization trends of eye banks in India.

Purpose: To study the trends in collection, storage and utilization of donor corneas in eye banks in India. Methods: The data was collected from 12 eye banks in India that collected more than 1000 corneas per year. The retrospective analysis of the parameters like characteristics of the donor and the host, storage media used, number of eyes collected, number of eyes utilized, causes of non-utilization of the tissue and the procedures performed was done. Results: A total of 20,564 eyes were collected by the 12 eye banks during the year 2013-2014. Voluntary eye donation (VED), and hospital cornea retrieval program (HCRP) contributed to 59.6% and 40.4% of tissue procurement respectively. Whole globe enucleation (52.3%) was more commonly performed as compared to in-situ excision of the donor corneas. The most commonly used storage media at all eye banks was McCarey-Kaufman (MK) media (83.3%). The utilization rate of the donor eyes was 50.5%. The most frequent indication for corneal transplantation was infection (active infection - 33.13%, healed infection - 10.78%) followed by Pseudophakic bullous keratopathy (PBK) (13.57%). Full thickness keratoplasty (optical penetrating keratoplasty - 47.23%, therapeutic penetrating keratoplasty - 31.74%) was performed most often followed by endothelial keratoplasty (12.41%) in the developing country. Conclusion: VED still contributes to majority of the donor tissue retrieval in India. The majority of the eye banks still utilize whole globe enucleation technique and store tissues in MK media. Trends from previous years showed a change towards HCRP, in-situ excision technique and preservation in the long-term storage media.

Sulfated polysaccharide mediated TGF-ß1 presentation in pre-formed injectable scaffolds for cartilage tissue engineering.

In this work, a plant-derived polysaccharide carboxymethylcellulose (CMC) was chemically modified to incorporate sulfate groups to facilitate binding of cationic growth factors. The sulfated CMC (heparin mimic) was then used with CMC (glycosaminoglycan mimic) and gelatin (collagen mimic) to fabricate injectable pre-formed, macroporous scaffolds for cartilage tissue engineering. These scaffolds demonstrated high resilience and shape memory, thereby making them injectable through a 14G needle for up to 4-6 aspiration and injection cycles. Further, the scaffolds could sequester cationic proteins and growth factors (TGF-ß1) through affinity-based interactions. When seeded with infrapatellar fat pad derived MSCs, the scaffolds demonstrated enhanced chondrogenesis after 28 days of in vitro culture when compared to controls. Taken together; these results demonstrate a polysaccharide-based minimally-invasive and translatable pre-formed injectable scaffold-based cell and growth factor delivery system for cartilage regeneration.

TGF-ß1 presenting enzymatically cross-linked injectable hydrogels for improved chondrogenesis.

In this work, we developed a novel enzymatically cross-linked injectable hydrogel composed of carboxymethyl cellulose (CMC), sulfated carboxymethyl cellulose (sCMC) and gelatin for delivery of infrapatellar fat pad derived MSCs and articular chondrocytes to a cartilage defect site while enabling TGF-ß1 mediated chondrogenesis. The sCMC component in the hydrogel served the purpose of mimicking heparan sulfate and thus enabled strong binding with TGF-ß1 and its consequential long term presentation to the encapsulated cells. We demonstrated that amongst CMC/sCMC/gelatin hydrogels cross-linked with 1 and 2mM H2O2, the latter demonstrated significantly higher compressive modulus and supported better in vitro cartilage formation. Thereafter, we explored the utility of this system to present TGF-ß1 to encapsulated cells for prolonged time period. It was observed that these hydrogels could sequester >90% of encapsulated TGF-ß1 for at least 4 weeks. The encapsulated TGF-ß1 was shown to be bioactive and supported significantly better cell survival over control hydrogels. Further, the TGF loaded hydrogels demonstrated good sulfated GAG and collagen deposition which was higher than control hydrogels and comparable to those treated with soluble TGF-ß1 through media. Interestingly, incorporation of TGF-ß1 in hydrogels not only enhanced the expression and deposition of hyaline cartilage markers, but it also significantly reduced the deposition of fibrocartilage and hypertrophy markers. Overall, it was concluded that TGF-ß1 immobilized CMC/sCMC/gelatin injectable hydrogels encapsulated with IFP MSCs and ACs present a promising, cost effective and easily translatable strategy for cartilage tissue engineering.

Understanding the influence of phosphorylation and polysialylation of gelatin on mineralization and osteogenic differentiation.

Post-translational modifications such as phosphorylation and sialylation impart crucial functions such as mineral deposition and osteogenic differentiation to non-collagenous bone matrix proteins. In this work, the influence of phosphorylation and polysialylation of gelatin on mineralization in simulated body fluid (SBF) and on osteogenic differentiation of mesenchymal stem cells (MSC) was studied. It was observed that increase in phosphorylation could be directly correlated with the mineralization ability of phosphorylated gelatin in SBF. The total calcium and phosphate deposited increased with increase in degree of phosphorylation and was >3 fold higher on the highest degree of phosphorylation. Whereas, polysialylation did not have any significant influence on mineral deposition in SBF. On the other hand, when MSCs were cultured on polysialylated surfaces they showed relatively higher cell elongation with 1.5 fold higher cell aspect ratio, higher alkaline phosphatase activity and 3 fold higher mineral deposition when compared to control and phosphorylated gelatin surfaces. In conclusion, phosphorylation and polysialylation of gelatin show a significant influence on mineralization and osteogenic differentiation respectively which can be advantageously used for bone tissue engineering.

Fabrication and characterization of Pluronic modified poly(hydroxybutyrate) fibers for potential wound dressing applications.

Electrospun poly(hydroxybutyrate) (PHB) fiber meshes have shown some success in wound dressing applications, however, their use is limited by their high hydrophobicity and brittle nature. In this study we investigated the effect of hydrophilization of electrospun PHB fibers by blending with Pluronic F-108 (PF) for use as a wound dressing material. Blending of PHB with different concentrations of PF (0.5%PF-PHB and 1.0% PF-PHB) before electrospinning led to a significant increase in the water wettability and swelling properties of fibers as compared to pristine PHB fibers. Further, it was observed that though the tensile moduli of PF blended PHB fibers were relatively lower as compared to PHB fibers, they show higher resistance to failure measured in terms of strain to failure and energy to failure. Moreover, PF blending significantly improved the in vitro blood clotting rate on PHB fibers when compared to control PHB fibers. Furthermore, the fabricated fiber systems were found to be cytocompatible and supported adhesion of fibroblasts in vitro. Finally, it was demonstrated that the PF blended fiber systems were suitable for the encapsulation of an antibiotic (doxycycline) to render them with antibacterial properties. Taken together, this study demonstrates that PF blending can be used to significantly improve properties of PHB fibers for wound dressing applications.

Pericellular plasma clot negates the influence of scaffold stiffness on chondrogenic differentiation.

Matrix stiffness is known to play a pivotal role in cellular differentiation. Studies have shown that soft scaffolds (<2-3kPa) promote cellular aggregation and chondrogenesis, whereas, stiffer ones (>10kPa) show poor chondrogenesis in vitro. In this work we investigated if fibrin matrix from clotted blood can act as a soft surrogate which nullifies the influence of the underlying stiff scaffold, thus promoting chondrogenesis irrespective of bulk scale scaffold stiffness. For this we performed in vitro chondrogenesis on soft (∼1.5kPa) and stiff (∼40kPa) gelatin scaffolds in the presence and absence of pericellular plasma clot. Our results demonstrated that in absence of pericellular plasma clot, chondrocytes showed efficient condensation and cartilaginous matrix secretion only on soft scaffolds, whereas, in presence of pericellular plasma clot, cell rounding and cartilaginous matrix secretion was observed in both soft and stiff scaffolds. More specifically, significantly higher collagen II, chondroitin sulfate and aggrecan deposition was observed in soft scaffolds, and soft and stiff scaffolds with pericellular plasma clot as compared to stiff scaffolds without pericellular plasma clot. Moreover, collagen type I, a fibrocartilage/bone marker was significantly higher only in stiff scaffolds without plasma clot. Therefore, it can be concluded that chondrocytes surrounded by a soft fibrin network were unable to sense the stiffness of the underlying scaffold/substrate and hence facilitate chondrogenesis even on stiff scaffolds. This understanding can have significant implications in the design of scaffolds for cartilage tissue engineering. STATEMENT OF SIGNIFICANCE: Cell fate is influenced by the mechanical properties of cell culture substrates. Outside the body, cartilage progenitor cells express significant amounts of cartilage-specific markers on soft scaffolds but not on stiff scaffolds. However, when implanted in joints, stiff scaffolds show equivalent expression of markers as seen in soft scaffolds. This disparity in existing literature prompted our study. Our results suggest that encapsulation of cells in a soft plasma clot, present in any surgical intervention, prevents their perception of stiffness of the underlying scaffold, and hence the ability to distinguish between soft and stiff scaffolds vanishes. This finding would aid the design of new scaffolds that elicit cartilage-like biochemical properties while simultaneously being mechanically comparable to cartilage tissue.

Pore orientation mediated control of mechanical behavior of scaffolds and its application in cartilage-mimetic scaffold design.

Scaffolds with aligned pores are being explored in musculoskeletal tissue engineering due to their inherent structural anisotropy. However, influence of their structure on mechanical behavior remains poorly understood. In this work, we elucidate this dependence using chitosan-gelatin based random and aligned scaffolds. For this, scaffolds with horizontally or vertically aligned pores were fabricated using unidirectional freezing technique. Random, horizontal and vertical scaffolds were characterized for their mechanical behavior under compressive, tensile and shear loading regimes. The results revealed conserved trends in compressive, tensile and shear moduli, with horizontal scaffolds showing the least moduli, vertical showing the highest and random showing intermediate. Further, these scaffolds demonstrated a highly viscoelastic behavior under cyclic compressive loading, with a pore orientation dependent relative energy dissipation. These results established that mechanical behavior of porous scaffolds can be modulated by varying pore orientation alone. This finding paved the way to recreate the structural and consequent mechanical anisotropy of articular cartilage tissue using zonally varied pore orientation in scaffolds. To this end, monolithic multizonal scaffolds were fabricated using a novel sequential unidirectional freezing technique. The superficial zone of this scaffold had horizontally aligned pores while the deep zone consisted of vertically aligned pores, with a transition zone between the two having randomly oriented pores. This depth-dependent pore architecture closely mimicked the collagen alignment of native articular cartilage which translated into similar depth-dependent mechanical anisotropy as well. A facile fabrication technique, biomimetic pore architecture and associated mechanical anisotropy make this multizonal scaffold a promising candidate for cartilage tissue engineering.

Pullulan-based composite scaffolds for bone tissue engineering: Improved osteoconductivity by pore wall mineralization.

Porous hydrogels have been explored for bone tissue engineering; however their poor mechanical properties make them less suitable as bone graft substitutes. Since incorporation of fillers is a well-accepted method for improving mechanical properties of hydrogels, in this work pullulan hydrogels were reinforced with nano-crystalline hydroxyapatite (nHAp) (5 wt% nHAp in hydrogel) and poly(3-hydroxybutyrate) (PHB) fibers (3 wt% fibers in hydrogel) containing nHAp (3 wt% nHAp in fibers). Addition of these fillers to pullulan hydrogel improved compressive modulus of the scaffold by 10 fold. However, the hydrophilicity of pullulan did not support adhesion and spreading of cells. To overcome this limitation, porous composite scaffolds were modified using a double diffusion method that enabled deposition of hydroxyapatite on pore walls. This method resulted in rapid and uniform coating of HAp throughout the three-dimensional scaffolds which not only rendered them osteoconductive in vitro but also led to an improvement in their compressive modulus. These results demonstrate the potential of mineralized pullulan-based composite scaffolds in non-load bearing bone tissue engineering.

Surface hydrophilicity of PLGA fibers governs in vitro mineralization and osteogenic differentiation.

Interfacial properties of biomaterials play an important role in governing their interaction with biological microenvironments. This work investigates the role of surface hydrophilicity of electrospun poly(lactide-co-glycolide) (PLGA) fibers in determining their biological response. For this, PLGA is blended with varying amounts of Pluronic®F-108 and electrospun to fabricate microfibers with varying surface hydrophilicity. The results of mineralization study in simulated body fluid (SBF) demonstrate a significant enhancement in mineralization with an increase in surface hydrophilicity. While presence of serum proteins in SBF reduces absolute mineral content, mineralization continues to be higher on samples with higher surface hydrophilicity. The results from in vitro cell culture studies demonstrate a marked improvement in mesenchymal stem cell-adhesion, elongation, proliferation, infiltration, osteogenic differentiation and matrix mineralization on hydrophilized fibers. Therefore, hydrophilized PLGA fibers are advantageous both in terms of mineralization and elicitation of favorable cell response. Since most of the polymeric materials being used in orthopedics are hydrophobic in nature, the results from this study have strong implications in the future design of interfaces of such hydrophobic materials. In addition, the work proposes a facile method for the modification of electrospun fibers of hydrophobic polymers by blending with a poloxamer for improved bone tissue regeneration.

Combination of single walled carbon nanotubes/graphene oxide with paclitaxel: a reactive oxygen species mediated synergism for treatment of lung cancer.

Heterogeneity in tumors has led to the development of combination therapies that enable enhanced cell death. Previously explored combination therapies mostly involved the use of bioactive molecules. In this work, we explored a non-conventional strategy of using carbon nanostructures (CNs) [single walled carbon nanotube (SWNT) and graphene oxide (GO)] for potentiating the efficacy of a bioactive molecule [paclitaxel (Tx)] for the treatment of lung cancer. The results demonstrated enhanced cell death following combination treatment of SWNT/GO and Tx indicating a synergistic effect. In addition, synergism was abrogated in the presence of an anti-oxidant, N-acetyl cysteine (NAC), and was therefore shown to be reactive oxygen species (ROS) dependent. It was further demonstrated using bromodeoxyuridine (BrdU) incorporation assay that treatment with CNs was associated with enhanced mitogen associated protein kinase (MAPK) activation that was ROS mediated. Hence, these results for the first time demonstrated the potential of SWNT/GO as co-therapeutic agents with Tx for the treatment of lung cancer.