Structural and functional characterization of proteins adsorbed on hydrophilized polylactide-co-glycolide microfibers

Rajesh Vasita, Dhirendra S KattiDepartment of Biological Sciences and Bioengineering, Indian Institute of Technology, Kanpur, Uttar Pradesh, IndiaBackground: Hydrophobic biopolymers such as polylactide-co-glycolide (PLGA, 85:15) have been extensively explored as scaffolding materials for tissue engineering applications. More recently, electrospun microfiber-based and nanofiber-based scaffolds of PLGA have received increased attention because they act as physical mimics of the fibrillar extracellular matrix. However, the hydrophobicity of the PLGA microfiber surface can limit its use in biomedical applications. Therefore, in a previous study, we fabricated Pluronic® F-108 (PF-108)-blended PLGA microfibrous scaffolds that alleviated the hydrophobicity associated with PLGA by enriching the surface of microfibers with the ethylene oxide units present in PF-108.Methods: In this study, we report the influence of the extent of surface enrichment of PLGA microfibers on their interaction with two model proteins, ie, bovine serum albumin (BSA) and lysozyme. BSA and lysozyme were adsorbed onto PLGA microfiber meshes (unmodified and modified) and studied for the amount, secondary structure conformation, and bioactivity of released protein.Results: Irrespective of the type of protein, PF-108-blended PLGA microfibers showed significantly greater protein adsorption and release than the unblended PLGA samples. However, in comparison with BSA, lysozyme showed a 7–9-fold increase in release. The Fourier transform infrared spectroscopy studies for secondary structure determination demonstrated that irrespective of type of microfiber surface (unblended or blended), adsorbed BSA and lysozyme did not show any significant change in secondary structure (α-helical content) as compared with BSA and/or lysozyme in the free powder state. Further, the bioactivity assay of lysozyme released from blended PLGA microfiber meshes demonstrated 80%–85% bioactivity, indicating that the process of adsorption did not significantly affect biological activity. Therefore, this study demonstrated that the decreased hydrophobicity of blended PLGA microfibrous meshes not only improved the amount of protein adsorbed (lysozyme and BSA) but also maintained the secondary structure and bioactivity of the adsorbed proteins.Conclusion: Modulating the hydrophobicity of PLGA via blending with PF-108 could be a viable strategy to improve its interaction with proteins and subsequent cell interaction in tissue engineering applications.Keywords: microfiber, protein adsorption, electrospinnin

A Synergistic Combination of Niclosamide and Doxorubicin as an Efficacious Therapy for All Clinical Subtypes of Breast Cancer

Drug resistance is one of the major hurdles in the success of cancer chemotherapy. Notably, aberrantly expressed Wnt/β-catenin signaling plays a major role in the initiation and maintenance of oncogenesis along with development of chemoresistance. Therefore, the combinatorial approach of targeting Wnt/β-catenin pathway along with using a chemotherapeutic agent seems to be a promising strategy to improve cancer therapy. In the present study, we evaluated the combination of niclosamide (Nic), an FDA-approved antihelminthic drug repurposed as a Wnt signaling inhibitor, and doxorubicin (Dox), a conventional anticancer agent, in all clinical subtypes of breast cancer viz. triple negative breast cancer, HER2 positive breast cancer, and hormone receptor positive breast cancer. The results demonstrated that the combination induced apoptosis and caused synergistically enhanced death of all breast cancer cell types at multiple combinatorial concentrations using both the sequential and concurrent treatment regimens. Mechanistically, downregulation of Wnt/β-catenin signaling and cell cycle arrest at G0/G1 phase by Nic and increase in reactive oxygen species by both Nic and Dox along with the inherent cytotoxicity of Dox mediated the synergism between the two drugs in both the treatment regimens. Overall, the combination of Nic and Dox holds promise to be developed as an efficient therapeutic option for breast cancer irrespective of its clinical subtype

Pore Alignment in Gelatin Scaffolds Enhances Chondrogenic Differentiation of Infrapatellar Fat Pad Derived Mesenchymal Stromal Cells

One of the major strategies in tissue engineering is the biomimetic scaffold-based approach that aims at providing a near-native-like environment for cells to facilitate the regeneration of damaged/lost tissue. The extracellular matrix in native articular cartilage contains aligned collagen fibrils in the superficial (parallel to the articular surface) and deep zones (perpendicular to articular surface) of the tissue. Therefore, we hypothesized that scaffolds with aligned pore architecture may offer aligned collagen deposition upon cell seeding, and as a result, may enable enhanced chondrogenesis. We tested this hypothesis by comparing gelatin scaffolds with random and aligned pore architecture for their ability to differentiate infrapatellar fat pad derived mesenchymal stromal cells (IFP-MSCs) toward the chondrogenic lineage. The fabricated scaffolds with random and aligned pore architecture were comparable in terms of pore size, degree of cross-linking, equilibrium swelling ratio, and in vitro degradation behavior. However, scaffolds with aligned pore architecture demonstrated higher compressive modulus along with cellular infiltration and alignment in comparison to the scaffolds with random pore architecture. An in vitro chondrogenesis study of IFP-MSCs seeded in the developed scaffold systems revealed that scaffolds with aligned pore architecture supported better chondrogenesis in terms of sGAG and total collagen (histology and biochemical) and cartilage specific matrix deposition (immunofluorescence). Further, scaffolds with aligned pore architecture also supported oriented deposition of cell secreted collagen. Taken together, these results suggest that scaffolds with aligned pore architecture enhance in vitro chondrogenic differentiation of IFP-MSCs as compared to scaffolds with random pore architecture and hence could be a potential design criterion in the development of scaffolds for cartilage regeneration

Scanning electron micrograph of poly(l-lactic acid) (PLLA) nanofibrous foam synthesized from 2

5% (wt/v) PLLA/tetrahydrofuran solution at a gelation temperature of 8°C using the phase separation technique (image 500 ×). Source: Ma PX, Zhang R. 1998. Synthetic nano-scale fibrous extracellular matrix. , 46:60–72. Copyright © 1998 J Wiley. Reprinted with permission of John Wiley&Sons Inc.<p><b>Copyright information:</b></p><p>Taken from "Nanofibers and their applications in tissue engineering"</p><p></p><p>International Journal of Nanomedicine 2006;1(1):15-30.</p><p>Published online Jan 2006</p><p>PMCID:PMC2426767.</p><p>© 2006 Dove Medical Press Limited. All rights reserved</p

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

Photoresist derived electrospun carbon nanofibers with tunable morphology and surface properties

A new precursor, SU-8, which is a negative photoresist, was electrospun to produce ultrafine polymeric fibers with a wide range of morphology and wettability characteristics. Electrospun nanofibers of SU-8 were pyrolyzed at 1173 K in an inert atmosphere to give carbon nanofibers. A set of parameters, including electric potential, distance between source and collector, polymer flow rate, and polymer concentration, was optimized for high-viscosity SU-8 photoresist to synthesize long continuous carbon fibers having diameters in the range of 120-600 nm. However, for the same conditions, medium- and lower-viscosity SU-8 yielded beaded fibers and isolated beads, respectively. The wettability of the carbon web was significantly influenced by its surface morphology, as shown by water contact angle measurements. These SU-8-derived carbon nanostructures with tunable surface properties and morphologies could be especially suitable for integration with photoresist-based carbon-MEMS to produce multiscale hierarchal assemblies and could be of potential use in a broad range of applications