**Myoblast Differentiation of Umbilical Cord Blood Derived Stem Cells on Biocompatible Composites Scaffold Meshes Myoblast Differentiation of Umbilical Cord Blood Derived Stem Cells on Biocompatible Composites Scaffold Meshes**

Biswadeep Chaudhuri Additional information is available at the end of the chapter

Biswadeep Chaudhuri

Additional information is available at the end of the chapter

http://dx.doi.org/10.5772/65032

#### **Abstract**

Tissue Engineering (TE) is emerging as an effective way of curing different tissue oriented disorders and new tissue regeneration. Here, it has been attempted to show that biocompatible graphene oxide nanoplatelets (GOnPs)-polymer nanocomposites are novel materials for the fabrication of TE scaffolds for myoblast differentiation of human umbilical cord blood derived mesenchymal stem cells (CB-hMSCs). Addition of GOnPs in bioactive polymers like PCL (poly-caprolactone) and GO-PLGA (poly lactic co-glycolic acid) enhances electrical conductivity and biocompatibility of the electrospun composite scaffolds. CB-hMSCs were used for the direct differentiation to skeletal muscle cells (hSkMCs) on the electrospun GOnPs–PCL and GOnPs-PLGA nanocomposite scaffolds. These scaffolds exhibited admirable myoblast differentiation, proliferation and also promoted self-aligned myotubes formation. Moreover, IGF-1 cell signaling pathway study carried out on GOnPs-PCL composite scaffold meshes also showed their potentiality for excellent myoblast differentiation and proliferation. Structural, mechanical, microstructural and vibration spectroscopic studies were carried out to characterize the scaffold materials. Significantly enhanced values of both conductivity and dielectric constant provided favorable cues for the increase of cellsscaffold interaction and biocompatibility of GOnPs based polymer composite scaffolds. Present study confirmed GOnPs-polymer composite scaffolds as the potential candidates for the myoblast differentiation of CB-hMSCs for skeletal muscle or other tissues repair and regenerations.

**Keywords:** umbilical cord blood, mesenchymal stem cells, myoblast differentiation, graphene oxide, polymer nanocomposite, electrospinning

© 2016 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. © 2017 The Author(s). Licensee InTech. This chapter is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/3.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

## **1. Introduction**

Fabricated artificial scaffolds using suitable biomimetic substrates were found to control the cellular behaviour and deliver appropriate cues for the differentiation and proliferation of different cell types for tissue engineering (TE) applications. Ideal scaffolds directly or indirectly help the growth of cells because they interact with the cells. Excellent biocompatibility, antibacterial property and mechanical stability of the scaffolds are necessary to provide adequate environment for cell growth and proliferation. Over the last decade, various nanomaterials (nanodiamond, graphene and carbon nanotubes) have been used along with suitable polymer to meet the requirements of the desired scaffold materials [1].Among the various techniques used for the fabrications of scaffolds, electrospinning drew worldwide attention as a simple and effective method [2] of preparing scaffold meshes of different biocompatible polymers like poly-caprolactone (PCL), polyvinyl alcohol (PVA) and poly(lactic-co-glycolic acid (PLGA)). These polymers have extensively been used for TE and biomedical applications [3–5] because of their appropriate biocompatibility, biodegradability and good solubility in different organic solvents. However, as the values of both conductivity (*σ* ~ 10−9 S/m) and dielectric permittivity (*ε* < 10) of most of these bioactive polymers are extremely small and also highly biodegradable; these polymers alone are not very suitable for many electronic as well as biomedical applications. Conducting polymers like polyaniline (PANi) and other solid materials blended with insulating polymers (such as PCL and PLGA PVA) were found to enhance biocompatibility, biodegradability and better cell scaffolds constructs [5, 6]. Interestingly, addition of very small amount (~0.3–0.5 wt%) of graphene oxide (GO) nanoplatelets (GOnPs) in different polymers like poly(methyl methacrylate) (PMMA) [7] and polyvinyl alcohol (PVA) [8] were reported to enhance both *ε* and *σ* values of the resulting composites by 2–3 orders of magnitudes along with enhancement of thermomechanical stability [8]. Conductivity enhancement of the scaffolds was also reported to provide important cues for myoblast differentiation [9].

Graphene oxide (GO) or reduced graphene oxide (rGO) are the oxidized forms of graphene. These oxides possess many oxygen-containing functional groups, such as hydroxyl, carboxyl and epoxy groups, and they can also adsorb small molecular weight chemical [10] which favour cell-scaffold interaction and cell viability. Therefore, graphene oxides might be considered as suitable biocompatible filler materials for making polymer composites for different biomedical and clinical applications. Moreover, chemically synthesized GO and rGO have also attracted more interest instead of graphene, in some respects, due to their extraordinary physicochemical properties, ease of synthesis in pure form and fabrication for applications in drug delivery [11], cancer therapies [12] and TE [13, 14].

Other than scaffolds, another important issue of tissue engineering is the availability of constant supply of stem cells which are mostly procured from bone marrow (BM), adipose tissue, etc. It is rather difficult to obtain these stem cells requiring sophisticated surgical procedures. Moreover, there are also problems to find appropriate donors. However, it has already been established that human umbilical cord blood, considered to be a biological waste, is an import and cost effective source of mesenchymal stem cells (MSCs) and there is immense scope of utilizing such MSCs for TE and other clinical applications.

**1. Introduction**

cues for myoblast differentiation [9].

[11], cancer therapies [12] and TE [13, 14].

Fabricated artificial scaffolds using suitable biomimetic substrates were found to control the cellular behaviour and deliver appropriate cues for the differentiation and proliferation of different cell types for tissue engineering (TE) applications. Ideal scaffolds directly or indirectly help the growth of cells because they interact with the cells. Excellent biocompatibility, antibacterial property and mechanical stability of the scaffolds are necessary to provide adequate environment for cell growth and proliferation. Over the last decade, various nanomaterials (nanodiamond, graphene and carbon nanotubes) have been used along with suitable polymer to meet the requirements of the desired scaffold materials [1].Among the various techniques used for the fabrications of scaffolds, electrospinning drew worldwide attention as a simple and effective method [2] of preparing scaffold meshes of different biocompatible polymers like poly-caprolactone (PCL), polyvinyl alcohol (PVA) and poly(lactic-co-glycolic acid (PLGA)). These polymers have extensively been used for TE and biomedical applications [3–5] because of their appropriate biocompatibility, biodegradability and good solubility in different organic solvents. However, as the values of both conductivity (*σ* ~ 10−9 S/m) and dielectric permittivity (*ε* < 10) of most of these bioactive polymers are extremely small and also highly biodegradable; these polymers alone are not very suitable for many electronic as well as biomedical applications. Conducting polymers like polyaniline (PANi) and other solid materials blended with insulating polymers (such as PCL and PLGA PVA) were found to enhance biocompatibility, biodegradability and better cell scaffolds constructs [5, 6]. Interestingly, addition of very small amount (~0.3–0.5 wt%) of graphene oxide (GO) nanoplatelets (GOnPs) in different polymers like poly(methyl methacrylate) (PMMA) [7] and polyvinyl alcohol (PVA) [8] were reported to enhance both *ε* and *σ* values of the resulting composites by 2–3 orders of magnitudes along with enhancement of thermomechanical stability [8]. Conductivity enhancement of the scaffolds was also reported to provide important

222 Umbilical Cord Blood Banking for Clinical Application and Regenerative Medicine

Graphene oxide (GO) or reduced graphene oxide (rGO) are the oxidized forms of graphene. These oxides possess many oxygen-containing functional groups, such as hydroxyl, carboxyl and epoxy groups, and they can also adsorb small molecular weight chemical [10] which favour cell-scaffold interaction and cell viability. Therefore, graphene oxides might be considered as suitable biocompatible filler materials for making polymer composites for different biomedical and clinical applications. Moreover, chemically synthesized GO and rGO have also attracted more interest instead of graphene, in some respects, due to their extraordinary physicochemical properties, ease of synthesis in pure form and fabrication for applications in drug delivery

Other than scaffolds, another important issue of tissue engineering is the availability of constant supply of stem cells which are mostly procured from bone marrow (BM), adipose tissue, etc. It is rather difficult to obtain these stem cells requiring sophisticated surgical procedures. Moreover, there are also problems to find appropriate donors. However, it has already been established that human umbilical cord blood, considered to be a biological waste, In this investigation, GOnPs-impregnated biomimetic PCL and PLGA matrices were prepared with GO concentration within nontoxicity limit for human cells (≤50/ml [15]) forthe fabrication of fibrous scaffolds by electrospinning technique. The widely used PCL and PLGA were chosen as the polymer matrices because of their low percolation threshold limit (*f*c ~ 0.80 wt% GO) forming homogeneous composites with GOnPs as filler [16]. So far, very little work [6, 10] has been done forthe direct differentiation and proliferation of human umbilical cord blood derived mesenchymal stem cells (CB-hMSCs) on pure biopolymer scaffolds or graphene oxide (GO)-polymer based composites scaffolds. The use of such biocompatible GO-PCL or GO-PLGA composite meshes for the myoblast differentiation of CB-hMSCs and oriented myotube formations are novel and challenging with immense future prospects of *in-vivo* tissue regeneration. The confirmation of myoblast differentiation of CB-hMSCs and oriented myotubes formation on GO-polymer electrospun meshes would also be advantageous for the next generation TE applications using such antibacterial biocompatible GO-polymer meshes. Since both PCL and PLGA-graphene oxide nonocomposite meshes exhibited excellent myoblast differentiation potential, in the present chapter, we shall mainly concentrate our discussion on the GOnPs-PCl composite system.
