Nanofibers and their applications in tissue engineering

Rajesh Vasita and Dhirendra S Katti

Department of Biological Sciences and Bioengineering, Indian Institute of Technology – Kanpur, Kanpur, Uttar Pradesh, India

Correspondence: Dhirendra S Katti Department of Biological Sciences and Bioengineering, Indian Institute of Technology – Kanpur, Kanpur-208016, Uttar Pradesh, India Tel +91 512 259 4028 Fax +91 512 259 4010 Email dsk@iitk.ac.in

Int J Nanomedicine. 2006 March; 1(1): 15–30.

PMCID: PMC2426767

Copyright © 2006 Dove Medical Press Limited. All rights reserved

Developing scaffolds that mimic the architecture of tissue at the nanoscale is one of the major challenges in the field of tissue engineering. The development of nanofibers has greatly enhanced the scope for fabricating scaffolds that can potentially meet this challenge. Currently, there are three techniques available for the synthesis of nanofibers: electrospinning, self-assembly, and phase separation. Of these techniques, electrospinning is the most widely studied technique and has also demonstrated the most promising results in terms of tissue engineering applications. The availability of a wide range of natural and synthetic biomaterials has broadened the scope for development of nanofibrous scaffolds, especially using the electrospinning technique. The three dimensional synthetic biodegradable scaffolds designed using nanofibers serve as an excellent framework for cell adhesion, proliferation, and differentiation. Therefore, nanofibers, irrespective of their method of synthesis, have been used as scaffolds for musculoskeletal tissue engineering (including bone, cartilage, ligament, and skeletal muscle), skin tissue engineering, vascular tissue engineering, neural tissue engineering, and as carriers for the controlled delivery of drugs, proteins, and DNA. This review summarizes the currently available techniques for nanofiber synthesis and discusses the use of nanofibers in tissue engineering and drug delivery applications.

Abstract

Introduction

Methods for nanofiber synthesis

Natural polymeric materials for nanofibers

Synthetic polymeric materials for nanofibers

Applications of nanofibers in tissue engineering

Nanofibers for musculoskeletal tissue engineering

Nanofibers for blood vessel tissue engineering

Nanofibers for controlled drug delivery

Application of carbon nanofibers in tissue engineering

Applications of alumina nanofibers in tissue engineering

Conclusion

References

Self-assembly

Eukaryotic cells can sense their local environment through cell receptors that recognize their corresponding extra-cellular tissue markers such as collagen and fibronectin. Therefore, mimicking the extracellular matrix (ECM) using biomaterials would be a logistic approach for engineering of a variety of tissue types. To mimic the human ECM, Berndt et al synthesized a peptide amphiphile (PA)-based self-assembling system with the goal of designing a simple self-assembly system that allows for the formation of thermally stable protein-like molecular architectures. The authors developed PAs that consisted of a dialkyl chain moiety (hydrophobic component/tail group) attached to an N-alpha amino group of a peptide chain (hydrophilic component/head group), resulting in a “peptide amphiphile” (Berndt et al 1995). The peptide head groups were derived from the ECM collagen ligand sequence. The synthesized PA hydrophilic head group consisted of Gly-Val-Lys-Gly-Asp-Lys-Gly-Asn-Pro-Gly-Trp-Pro-Gly-Ala-Pro [IV-H1], which is similar to the human α1 (IV) 1263-1277 collagen sequence. In another study, Yu et al replaced the dialkyl chains of the PA used in the previous study with monoalkyl chains (Berndt et al 1995; Yu et al 1996, 1998; Fields et al 1998). They demonstrated that with an increase in the length of the monoalkyl chain from C₆ to C₁₆, the thermal stability of the PA increased because of the hydrophobic interaction between alkyl chains (Yu et al 1998). Both the dialkyl and monoalkyl chain-based PAs readily self-assembled to form a stabilized triple-helical conformation in an aqueous solvent at the liquid–air interface (Yu et al 1996, 1999).

In a more recent study, Malker et al determined the bioactivity of the PA self-assemblies by incorporating bioactive sequences within the PA. Their results indicated that the formation of the triple-helix for such a PA (ie, containing a bioactive sequence) produced an ordered structure of the bioactive sequence on the exterior of the triple helix that led to a favorable cell response (ie, cell adhesion, spreading, and proliferation) because of the similarity of the self-assembled triple helix to natural ECM. The results of this study and another previous study by Fields et al indicated that these PA structures have potential to be used as surface coatings for biomaterials to improve biocompatibility (Fields et al 1998; Malkar et al 2003).

Based on prior knowledge of PA self-assembling systems (Berndt et al 1995; Stupp et al 1997; Fields et al 1998; Yu et al 1998; Malkar et al 2003), Stupp et al designed di- and tri-block PAs that self-assembled into a rod-like architecture. By engineering the peptide head group of the PA, the authors developed a new technique for the self-assembly of PAs into nanofibers using pH control (Hartgerink et al 2001).

The synthesis of the PA involved the following salient features (Hartgerink et al 2001).

• Incorporation of phosphoserin residue to enable enhanced hydroxyapatite (HA) mineralization.
• Incorporation of RGD (Arg-Gly-Asp) peptide to increase integrin-mediated cell adhesion.
• Incorporation of four consecutive cystine residues, which form inter-molecular disulfide bonds that polymerize to provide improved structural stability.
• Incorporation of a flexible linker region consisting of three glycine residues to provide flexibility to the head group.

The preparation of nanofibers involved reduction of cystine residues of the PA to free thiol groups using dithiotheritol followed by acidification below pH 4 to cause self-assembly of the PAs into cylindrical micelles/nanofibers. The resulting nanofibers had a hydrophobic core of alkyl residues and a hydrophilic exterior lined by peptide residues. Their results indicated that the nanofibers produced by self-assembly were approximately 5–8 nm in diameter and several microns in length (see Figure 2). Hartgerink et al further investigated the mineralization potential of these nanofibers. The authors observed the formation of HA crystals that were oriented along the length of the nanofibers. This nanoscale orientation resembles the orientation of HA crystals in mineralized ECM and collagen fibers of bone tissue. Since the mineralized, self-assembled nanofibers were similar to the lowest level of the hierarchical structure of bone tissue, the authors believe that the nanofibers show potential to be used as primary building blocks for the engineering of bone or other mineralization tissue (Hartgerink et al 2001).

Figure 2 Transmission electron micrograph (TEM) of nanofibers formed from peptide amphiphile molecules (N terminus – C10H19O and Peptide – CCCCGG GS(PO4)RGD) that self-assembled by drying directly onto a TEM grid without adjusted pH (negatively (more ...)

In another study, Hartgerink et al (2002) investigated the effect of variations in the molecular structure of the PAs on the self-assembled nanofibers. It was observed that modifications in the alkyl chain length of the PA alter the pH sensitivity of nanofibers, which affects self-assembly. Modification of the C- terminal region (ie, the region that is expressed on the surface of the nanofibers after self-assembly) led to changes in length and stiffness of the nanofibers. Replacement of cystine residues by alanine did not affect the self-assembly of the PAs into nanofibers. These results suggested that the self-assembled nanofibers show potential for development as novel biomaterials (Hartgerink et al 2002; Hwang et al 2002). This study also introduced three different methods of forming self-assembled PAs, including pH-controlled self-assembly, drying on surface-induced self-assembly, and divalent-ion-induced self-assembly. The study demonstrated that PAs can be self-assembled reversibly into nanofibers that result in the formation of gels through pH changes. These PA nanofibers can also be reversibly polymerized to improve their stability. The reversibility of these two procedures makes the self-assembly technique attractive as it enables the fabrication of remarkably versatile materials. In addition, this technique produces a good yield of nanofibers with low polydispersity.

Therefore, the self-assembly technique, by virtue of the modifications possible in the structure of the PA, enables a variety of self-assemblies including layered and lamellar structures, and by virtue of the aforementioned reversibilities lends flexibility to the system. Thus, the self-assembly technique shows good potential for further exploration with the goal of designing novel scaffolds for tissue engineering applications.

Reference: http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=2426767