Platelet derived growth factor releasing chitosan sponge for periodontal bone regeneration

Abstract

With an aim of improving bone regeneration, chitosan sponge containing platelet-derived growth factor-BB (PDGF-BB) were developed. For fabrication of chitosan sponge, chitosan solution was freeze-dried, crosslinked and freeze-dried again. PDGF-BB was incorporated into the chitosan sponge by soaking chitosan sponge into the PDGF-BB solution. Release kinetics of PDGF-BB, cell attachment, proliferation capacity and bony regenerative potentials of PDGF-BB-loaded chitosan sponge were investigated. Prepared chitosan sponge retained porous structure with 100 μm pore diameter that was suitable for cellular migration and growth. Release rate of PDGF-BB could be controlled by varying initial loading content of PDGF-BB to obtain optimal therapeutic efficacy. PDGF-BB-loaded chitosan sponge induced significantly high cell attachment and proliferation level, which indicated good cellular adaptability. PDGF-BB-loaded chitosan sponge demonstrated marked increase in new bone formation and rapid calcification. Degradation of the chitosan sponge was proceeded at defect site and subsequently replaced with new bone. Histomorphometric analysis confirmed that PDGF-BB-loaded chitosan sponge significantly induced new bone formation. These results suggested that chitosan sponge and PDGF-BB-loaded chitosan sponge may be beneficial to enhance periodontal bone regeneration.

Introduction

Periodontal regenerative therapy with bone-substituting materials has gained favorable clinical efficacy by enhancing osseous regeneration in periodontal bony defects [1], [2], [3], [4], [5], [6], [7], [8], [9]. As bone-substituting materials, bone powder [1], [2], calcium phosphate ceramic [3], [4], modified forms of hydroxyapatite (Osteogen^®) [5], [6], and hard tissue replacement polymer (HTR polymer^®) [7], [8], [9] have demonstrated their periodontal bony regenerative potency. Bone-substituting materials should fulfill several requirements such as biocompatibility, osteogenecity, malleability, biodegradability [1], [2], [3], [7]. However, these materials revealed some drawbacks including bone resorption, immune response, disease transmission, low biodegradability, poor adaptation. Several attempts have been made to overcome these problems; use of biodegradable polymers and combination of ceramics with bioactive polymers such as collagen and polylactides [10], [11], [12]. These attempts have been aimed at developing new bone substitutes that resemble bone more closely than other materials. However, these techniques have some limitations in inducing bone regeneration within whole therapeutic period. These materials act only as physical scaffolds for bone tissue. Bone formation after grafting these materials occurred over a period of several months or years [1], [4], [6], [7], [10]. Additional function to these materials should be imparted to shorten bone forming period and enhancing bone forming efficacy. As an effective approach, drug incorporation within these materials might be suggested to obtain this purpose.

Early healing concept of bone formation by some bioactive agents such as flurbiprofen and tetracycline has been recently suggested in periodontal regeneration [13], [14]. Especially, growth factors including platelet-derived growth factor-BB (PDGF-BB), insulin-like growth factor (IGF), transforming growth factor-β (TGF) function as modulators of chemotaxis, proliferation, and differentiation of pluripotent cells concerning bone regeneration [15], [16]. Overall healing periods might be significantly shortened by using these agents. It would thus be more advantageous for the substituting material to have release-controlling capacity of bioactive agents. This combination of controlled drug delivery concept with bone substitute technique can be highly beneficial for bone regeneration.

In this study, chitosan sponge were developed as an osteoconductive material which induces or stimulates bone formation. Chitosan is a biodegradable cationic polysaccharide composed of N-acetylglucosamine residues which is known to accelerate wound healing and bone formation [17]. Many previous reports corroborated enhanced wound healing and hemostatic effect of chitosan [18], [19], [20]. In addition to these biomedical applicability, chitosan can regulate release of bioactive agents [21], [22], [23]. Platelet-derived growth factor-BB (PDGF-BB) was used in this study. PDGF-BB is reported as a potential mediator of bone regeneration [24]. It stimulates proliferation and differentiation of mesenchymally derived cells including fibroblasts, smooth muscle cells, ligament cells and osteoblasts [24], [25], [26]. Also, it has revealed significant neoosteogenic effect in beagle dogs and monkeys [27], [28], [29]. However, as yet, extremely high dose of PDGF-BB (above 10 μg/ml) has been applied in clinical trials owing to rapid clearance of PDGF-BB in vivo and inability to maintain therapeutic concentration during implantation period [25], [26], [27], [28]. Therefore, controlled PDGF-BB delivery from the sponge may be highly beneficial for the treatment of periodontal disease.

The objective of this study is to develop growth factor releasing chitosan sponge matrices as a osteoconductive materials for bone regeneration. This paper reports on the fabrication of chitosan sponge, release kinetics of growth factor, osteoblast attachment and proliferation and bone regeneration capacity of growth-factor-loaded chitosan sponge.