Anuradha Subramanian

Development of biomaterial scaffold for nerve tissue engineering: Biomaterial mediated neural regeneration

Neural tissue repair and regeneration strategies have received a great deal of attention because it directly affects the quality of the patients life. There are many scientific challenges to regenerate nerve while using conventional autologous nerve grafts and from the newly developed therapeutic strategies for the reconstruction of damaged nerves. Recent advancements in nerve regeneration have involved the application of tissue engineering principles and this has evolved a new perspective to neural therapy. The success of neural tissue engineering is mainly based on the regulation of cell behavior and tissue progression through the development of a synthetic scaffold that is analogous to the natural extracellular matrix and can support three-dimensional cell cultures. As the natural extracellular matrix provides an ideal environment for topographical, electrical and chemical cues to the adhesion and proliferation of neural cells, there exists a need to develop a synthetic scaffold that would be biocompatible, immunologically inert, conducting, biodegradable, and infection-resistant biomaterial to support neurite outgrowth. This review outlines the rationale for effective neural tissue engineering through the use of suitable biomaterials and scaffolding techniques for fabrication of a construct that would allow the neurons to adhere, proliferate and eventually form nerves.

Fabrication of uniaxially aligned 3D electrospun scaffolds for neural regeneration

Nanofibrous scaffolds are very promising physical guidance substrates for regenerating nerves to traverse larger nerve gaps. In this study, we have attempted to develop 2D random and 3D longitudinally oriented nanofibers of poly(lactide-co-glycolide) (PLGA) by the modified electrospinning process and characterized the surface morphology, mechanical properties, porosity, degradation and wettability. The orientation of aligned fibers was optimized by varying the speed of the rotating mandrel in the electrospinning process. The mean diameter of random PLGA nanofibers was 197 ± 72 nm, whereas that of the aligned PLGA fiber was 187 ± 121 nm. The pore size of aligned PLGA nanofibers (3.5 ± 1.1 µm) was significantly lower than their respective random nanofibers (8.0 ± 2.0 µm) (p < 0.05). However, the percentage porosity of both scaffolds was comparable (p > 0.05). The mass loss percentage and molecular weight loss percentage due to degradation was higher in random PLGA fibers when compared to aligned PLGA after 5 weeks (p < 0.05). The tensile strength and Youngs modulus of random PLGA fibers were significantly higher than those of the aligned PLGA nanofibers (p < 0.05). Both random and longitudinally aligned scaffolds were used for the in vitro culture of Schwann cells. Morphology and cell proliferation results demonstrated that the aligned fibers assist the direction of Schwann cells and a better proliferation rate than their random fibers. The results confirmed that aligned nanofibers have better deformability, slow degradation, comparable porosity and orientation cues than random nanofibers. Hence the longitudinally aligned nanofibers may be ideal scaffolds for nerve regeneration.

Role of biomaterials, therapeutic molecules and cells for hepatic tissue engineering.

Current liver transplantation strategies face severe shortcomings owing to scarcity of donors, immunogenicity, prohibitive costs and poor survival rates. Due to the lengthy list of patients requiring transplant, high mortality rates are observed during the endless waiting period. Tissue engineering could be an alternative strategy to regenerate the damaged liver and improve the survival and quality of life of the patient. The development of an ideal scaffold for liver tissue engineering depends on the nature of the scaffold, its architecture and the presence of growth factors and recognition motifs. Biomimetic scaffolds can simulate the native extracellular matrix for the culture of hepatocytes to enable them to exhibit their functionality both in vitro and in vivo. This review highlights the physiology and pathophysiology of liver, the current treatment strategies, use of various scaffolds, incorporation of adhesion motifs, growth factors and stem cells that can stabilize and maintain hepatocyte cultures for a long period.

Multi-functional nanoparticles as theranostic agents for the treatment & imaging of pancreatic cancer.

Theranostics has received considerable attention since both therapy and imaging modalities can be integrated into a single nanocarrier. In this study, fluorescent iron oxide (FIO) nanoparticles and gemcitabine (G) encapsulated poly(lactide-co-glycolide) (PLGA) nanospheres (PGFIO) conjugated with human epidermal growth factor receptor 2, (HER-2) antibody (HER-PGFIO) were prepared and characterized. HER-PGFIO showed the magnetic moment of 10emu/g, relaxivity (r2) of 773mM-1s-1 and specific absorption rate (SAR) of 183W/g. HER-PGFIO showed a sustained release of gemcitabine for 11days in PBS (pH 7.4). In vitro cytotoxicity evaluation of HER-PGFIO in 3D MIAPaCa-2 cultures showed 50% inhibitory concentration (IC50) of 0.11mg/mL. Subcutaneous tumor xenografts of MIAPaCa-2 in SCID mice were developed and the tumor regression study at the end of 30days showed significant tumor regression (86±3%) in the HER-PGFIO with magnetic hyperthermia (MHT) treatment group compared to control group. In vivo MRI imaging showed the enhanced contrast in HER-PGFIO+MHT treated group compared to control. HER-PGFIO showed significant tumor regression and enhanced MRI in treatment groups, which could be an effective nanocarrier system for the treatment of pancreatic cancer. STATEMENT OF SIGNIFICANCE Combination therapies are best suitable to treat pancreatic cancer. Theranostics are the next generation therapeutics with both imaging and treatment agents encapsulated in a single nanocarrier. The novelty of the present work is the development of targeted nanocarrier that provides chemotherapy, thermotherapy and MRI imaging properties. The present work is the next step in developing the nanocarriers for pancreatic cancer treatment. Different treatment modalities embedding into a single nanocarrier is the biggest challenge that was achieved without compromising the functionality of each other. The surface modification of polymeric nanocarriers for antibody binding and their multifunctional abilities will appeal to wider audience.

In Vivo Biocompatibility of PLGA-Polyhexylthiophene Nanofiber Scaffolds in a Rat Model

Electroactive polymers have applications in tissue engineering as a physical template for cell adhesion and carry electrical signals to improve tissue regeneration. Present study demonstrated the biocompatibility and biodegradability of poly(lactide-co-glycolide)-poly(3-hexylthiophene) (PLGA-PHT) blend electrospun scaffolds in a subcutaneous rat model. The biocompatibility of PLGA-undoped PHT, PLGA-doped PHT, and aligned PLGA-doped PHT nanofibers was evaluated and compared with random PLGA fibers. The animals were sacrificed at 2, 4, and 8 weeks; the surrounding tissue along with the implant was removed to evaluate biocompatibility and biodegradability by histologic analysis and GPC, respectively. Histology results demonstrated that all scaffolds except PLGA-undoped PHT showed decrease in inflammation over time. It was observed that the aligned PLGA-doped PHT fibers elicited moderate response at 2 weeks, which further reduced to a mild response over time with well-organized tissue structure and collagen deposition. The degradation of aligned nanofibers was found to be very slow when compared to random fibers. Further, there was no reduction in the molecular weight of undoped form of PHT throughout the study. These experiments revealed the biocompatibility and biodegradability of PLGA-PHT nanofibers that potentiate it to be used as a biomaterial for various applications.

Nanoarchitecture of scaffolds and endothelial cells in engineering small diameter vascular grafts

Regeneration of functional small diameter blood vessels still remains a challenge, as the synthetic vascular grafts fail to mimic the complex structural architecture and dynamic functions of blood vessels and also lack with the lack of non‐thrombogenicity. Although, the existence of nanofibrous extracellular matrix components in the native tissue promotes many physical and molecular signals to the endothelial cells for the regulation of morphogenesis, homeostasis, and cellular functions in vascular tissue, poor understanding of the structural architecture on the functional activation of appropriate genes limits the development of successful vascular graft design. Hence, the present review outlines the functional contributions of various nanofibrous extracellular matrix components in native blood vessels. Further, the review focuses on the role of nanofiber topography of biomaterial scaffolds in endothelial cell fate processes such as adhesion, proliferation, migration, and infiltration with the expression of vasculature specific genes; thereby allowing the reader to envisage the communication between the nano‐architecture of scaffolds and endothelial cells in engineering small diameter vascular grafts.

Injectable glycosaminoglycan–protein nano-complex in semi-interpenetrating networks: A biphasic hydrogel for hyaline cartilage regeneration

Articular hyaline cartilage regeneration remains challenging due to its less intrinsic reparability. The study develops injectable biphasic semi-interpenetrating polymer networks (SIPN) hydrogel impregnated with chondroitin sulfate (ChS) nanoparticles for functional cartilage restoration. ChS loaded zein nanoparticles (∼150nm) prepared by polyelectrolyte-protein complexation were interspersed into injectable SIPNs developed by blending alginate with poly(vinyl alcohol) and calcium crosslinking. The hydrogel exhibited interconnected porous microstructure (39.9±5.8μm pore diameter, 57.7±5.9% porosity), 92% swellability and >350Pa elastic modulus. Primary chondrocytes compatibility, chondrocyte-matrix interaction with cell-cell clustering and spheroidal morphology was demonstrated in ChS loaded hydrogel and long-term (42days) proliferation was also determined. Higher fold expression of cartilage-specific genes sox9, aggrecan and collagen-II was observed in ChS loaded hydrogel while exhibiting poor expression of collagen-I. Immunoblotting of aggregan and collagen II demonstrate favorable positive influence of ChS on chondrocytes. Thus, the injectable biphasic SIPNs could be promising composition-mimetic substitute for cartilage restoration at irregular defects.

Development of nanotheranostics against metastatic breast cancer — A focus on the biology & mechanistic approaches

Treatment for metastatic breast cancer still remains to be a challenge since the currently available diagnostic and treatment strategies fail to detect the micro-metastasis resulting in higher mortality rate. Moreover, the lack of specificity to target circulating tumor cells is also a factor. In addition, currently available imaging modalities to identify the secondaries vary with respect to various metastatic anatomic areas and size of the tumor. The drawbacks associated with the existing clinical management of the metastatic breast cancer demands the requirement of multifunctional nanotheranostics, which could diagnose at macro- and microscopic level, target the solid as well as circulating tumor cells and control further progression with the simultaneous evaluation of treatment response in a single platform. However, without the understanding of the biology as well as preferential homing ability of circulating tumor cells at distant organs, it is quite impossible to address the existing challenges in the present diagnostics and therapeutics against the breast cancer metastasis. Hence this review outlines the severity of the problem, basic biology and organ specificity with the sequential steps for the secondary progression of disease followed by the various mechanistic approaches in diagnosis and therapy at different stages.

Self-assembling peptide nanostructures on aligned poly(lactide-co-glycolide) nanofibers for the functional regeneration of sciatic nerve

AIM Regeneration of functional peripheral nerve tissue at critical-sized defect requires extracellular matrix analogs impregnated with appropriate biosignals to regulate the cell fate process and subsequent tissue progression. The aim of the study was to develop electrospun aligned nanofibers as architectural analogs integrated with RADA16-I-BMHP1 as biofunctional peptides. MATERIALS & METHODS Aligned poly(lactide-co-glycolide) (PLGA)-RADA16-I-BMHP1 nanofibers were fabricated and characterized for their in vitro potential using rat Schwann cell line and in vivo potential using a 10 mm sciatic nerve transection rat model. RESULTS PLGA-peptide scaffolds significantly promoted higher expression of genotypic markers and bipolar extension of Schwann cells. Further, PLGA-peptide treated animals promoted the native collagen organization, remyelination and showed significantly higher recovery of sensorimotor and motor function than PLGA-treated groups (p < 0.05). CONCLUSION Our results demonstrate that self-assembling peptide nanostructures on aligned PLGA nanofibers provided better cell-matrix communication with significant functional restoration of the sciatic nerve.

Phase-induced porous composite microspheres sintered scaffold with protein–mineral interface for bone tissue engineering

Scaffolds for orthopedic reconstruction should best mimic the microenvironment of the bone to instil regenerative potential. In this study, a three-dimensional porous microsphere sintered scaffold that closely resembles bone micro-architecture has been fabricated. Synthesized nanohydroxyapatite (∼30–90 nm) using wet chemical precipitation and poly(hydroxybutyrate)/poly(e-caprolactone) were blended to develop composite porous microspheres via emulsion induced phase separation. Brunauer–Emmett–Teller analysis reveals presence of cylindrical mesopores in microspheres with surface area 10.64 m2 g−1, which on solvent/non-solvent sintering to scaffold, resembles trabecular section of rat sternum. The scaffold demonstrates desirable compressive strength (1.16 ± 0.17 MPa) and compressive modulus (6.8 ± 1.3 MPa) with slow degradation and microstructural stability for four weeks. Further, incorporation of bovine serum albumin (BSA) in the scaffold establishes protein–mineral interface and exhibits two-phase protein release mechanism (diffusion and polymer degradation). Pore interconnectivity and functional protein loading of scaffold have been evidenced by protein distribution analysis and secondary structural stability. The transverse section of MG63 cells cultured scaffolds with and without protein demonstrates cell infiltration and extension across the pores, while significantly higher proliferation is observed in protein scaffolds at day 7 (p < 0.05). Hence, the biomimetic scaffold with protein–mineral interface could be an ideal substitute for bone regeneration as it establish cell–matrix interaction.