Elsevier

Carbohydrate Polymers

Volume 98, Issue 1, 15 October 2013, Pages 574-580
Carbohydrate Polymers

Cell culture and characterization of cross-linked poly(vinyl alcohol)-g-starch 3D scaffold for tissue engineering

https://doi.org/10.1016/j.carbpol.2013.06.020Get rights and content

Highlights

  • PVA and starch can be chemically cross-linked to form a PVA-g-starch 3D scaffold-grafted polymer.

  • The absorbency of PVA-g-starch 3D scaffold is up to 800%.

  • The strength of the 3D scaffold strength reaches 4 × 10−2 MPa.

  • The 3D scaffold was degraded by various enzymes at a rate of up to approximately 30–60% in 28 days.

  • In vitro experiments revealed that cells proliferate and grow in the 3D scaffold material.

Abstract

The research goal of this experiment is chemically to cross-link poly(vinyl alcohol) (PVA) and starch to form a 3D scaffold that is effective water absorbent, has a stable structure, and supports cell growth. PVA and starch can be chemically cross-linked to form a PVA-g-starch 3D scaffold polymer, as observed by Fourier transform infrared spectroscopy (FTIR), with an absorbency of up to 800%. Tensile testing reveals that, as the amount of starch increases, the strength of the 3D scaffold strength reaches 4 × 10−2 MPa. Scanning electron microscope (SEM) observations of the material reveal that the 3D scaffold is highly porous formed using a homogenizer at 500 rpm. In an enzymatic degradation, the 3D scaffold was degraded by various enzymes at a rate of up to approximately 30–60% in 28 days. In vitro tests revealed that cells proliferate and grow in the 3D scaffold material. Energy dispersive spectrometer (EDS) analysis further verified that the bio-compatibility of this scaffold.

Introduction

Biodegradable polymers have been extensively utilized in medicine for a long time. Biodegradable polymer are bio-compatible, bio-absorbent and do not induce an immune reaction or inflammation can be used in medical materials in, for example, sutures, cover coatings, fracture fixing materials and other applications (Hsieh et al., 2011, Jiang et al., 2004, Kuo et al., 2012, Li et al., 2005, Wang et al., 2004). In recent years, the use of biodegradable polymer in combination with the cultivation of live cells to form new cartilage tissue has opened up the new research field of tissue engineering (Chen et al., 2000, Lahiji et al., 2000, Lanza et al., 1999, Yoshimoto et al., 2003). The aim of using these special biopolymers to construct 3D scaffolds in which implanted cells can proliferate and grow, is to combine the advantages of the transplantation of human tissue with those of synthetic repair materials, and so establish principles for tissue engineering. Therefore, the use of biodegradable polymers in applications of biomaterials seems more important than ever.

A 3D scaffold can be used to grow and cultivate cells only under the following conditions, 1. Biocompatible scaffold must be non-toxic to implanted cells or tissues and to promote their growth and adhesion. 2. Bio-absorbable or bio-decomposable a 3D scaffold is an auxiliary tool, whose ultimate purpose is to be able to degradation and then be absorbed or excreted by the body after the cells or tissues have grown in a period suitable for cultivation. 3. Exhibit highly linked pores – when cells are being cultivated using a 3D scaffold, they must be able to adopt the designed shape in the scaffold. As the cells grow in the culture liquid, nutrients must be freely input to, and the waste material should be excreted from, the structure. Accordingly, porosity is a key factor in cell cultivation. 4. Exhibit excellent mechanical strength and flexibility – the cells will not be unable to maintain their original shapes during the period of cultivation because of changes in the culture liquid or degradation. Hence, 3D scaffold must have a mechanical strength to support the attachment of cells.

This investigation concerns an inexpensive polymer-polyvinyl alcohol (PVA) which is an extensively used water-soluble polymer. PVA contains numerous polar alcohol groups and may form hydrogen bonds with water. It therefore dissolves easily in water. PVA is also a biodegradable material has that has been used in tissue engineering, and production methods include electrospining to form PVA nanofibrous scaffolds (Asran et al., 2010, Liao et al., 2011); chemical synthesis (Thomas et al., 2009, Mansur and Costa, 2008), freeze-drying (Jiang et al., 2011, Mohan and Nair, 2008, Mohan et al., 2010, Poursamar et al., 2011), and melt-molding particulate-leaching (Oh, Kang, Kim, Cho, & Lee, 2003). Starch, a natural polymer, is a material that is also commonly used in tissue engineering. In most relevant studies starch is blended with other matter to form a porous scaffold (Castillejo et al., 2012, Duarte et al., 2010, Ghosh et al., 2008, Gomes et al., 2002, Santos et al., 2010). Other methods of producing a porous scaffold, such as chemical synthesis (Sundaram et al., 2008, Xiao and Yang, 2006), wet spinning (Pashkuleva et al., 2010, Rodrigues et al., 2012), and use of a blowing agent (Salgado et al., 2002) have also been reported upon in these literature. The cross-linking method that is proposed in this investigation does not seem to have been described before is utilized herein for the first time.

The goal is to exploit an existing, inexpensive, biodegradable material to form a 3D scaffold material for use in regenerative medicine. Since PVA is inexpensive, easy to obtain, highly biodegradable and biocompatibility, PVA and natural polymer-starch are considered herein. No complex or expensive equipment is required: simple chemical cross-linking method, to form 3D scaffold material and to improve its mechanical properties and formation. Not only are the structural changes of the material examined: its gel and swelling, formation, mechanical properties, biodegradability, porosity and other characteristics are elucidated. Finally, NIH3T3 cells are transplanted and cultivated to 3D scaffold, and observe the effectiveness of the material for tissue engineering.

Section snippets

Formation of 3D scaffold

PVA and soluble starch were obtained from the Nitto Chemical Pharmaceutical Companies. Formaldehyde was obtained from Aldrich Chemical Company. Sulfuric acid (H2SO4) was purchased from The First Chemical Company. All materials and chemicals were used as acquired without any processing.

The starch powder (0.5, 0.75, and 1 g) was separately dissolved in RO water (10 mL) at room temperature. The 1 g of PVA was dissolved in RO water (10 mL) at 90 °C. The starch solution was slowly added into the PVA

Morphology of 3D scaffold

SEM was utilized to observe the morphological changes of the chemical cross-linked PVA-g-starch 3D scaffold. Fig. 1 presents SEM image of the PVA-g-starch 3D scaffold. The pores of the 3D material were formed by a homogenizer at 500 rpm. The photographs display the distribution of the pores of the 3D material that was cross-linked by stirring within the material. The physical properties of these materials will be investigated, and the material will be used for cell cultivation.

FT-IR analysis of cross-linked PVA-g-starch

To confirm whether

Conclusions

The cross-linked PVA-g-starch 3D scaffold reveals that is highly porous and easily formed using a homogenizer at 500 rpm. Formaldehyde (aldehyde group) can react with PVA and starch (hydroxyl groups) under the presence of an acid catalyst, and forming rings structure of 1,3-dioxane. The FT-IR analysis demonstrates that the PVA-g-starch had been successfully prepared. The produced 3D scaffolds have highly porosity; can keep a large number of water. The absorption of water of 3D scaffold increased

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