RESUMO
One strategy to correct alveolar bone defects is use of bioactive bone substitutes to maintain the structure of defect site and facilitate cells and vessels' ingrowth. This study aimed to fabricate and characterize the freeze-dried bone regeneration scaffolds composed of polymeric Type I collagen, nano Beta-tricalcium phosphate (ß-TCP), and gelatin. The stable structures of scaffolds were obtained by thermal crosslinking and EDC/NHS ((1-ethyl-3-(3-dimethylaminopropyl) carbodiimide)/(N-hydroxysuccinimide)) chemical crosslinking processes. Subsequently, the physicochemical and biological properties of the scaffolds were characterized and assessed. The results indicated the bioactive composite scaffolds containing 10% and 20% (w/v) nano ß-TCP exhibited suitable porosity (84.45 ± 25.43 nm, and 94.51 ± 14.69 nm respectively), a rapid swelling property (reaching the maximum swelling rate at 1 h), excellent degradation resistance (residual mass percentage of scaffolds higher than 80% on day 90 in PBS and Type I collagenase solution respectively), and sustained calcium release capabilities. Moreover, they displayed outstanding biological properties, including superior cell viability, cell adhesion, and cell proliferation. Additionally, the scaffolds containing 10% and 20% (w/v) nano ß-TCP could promote the osteogenic differentiation of MC3T3-E1. Therefore, the bioactive composite scaffolds containing 10% and 20% (w/v) nano ß-TCP could be further studied for being used to treat alveolar bone defects in vivo.
Assuntos
Gelatina , Osteogênese , Gelatina/farmacologia , Alicerces Teciduais/química , Regeneração Óssea , Colágeno/química , Fosfatos de Cálcio/farmacologia , Fosfatos de Cálcio/química , Polímeros , Engenharia Tecidual/métodosRESUMO
The natural healing process of extraction socket and traditional socket plug material could not prevent buccal bone wall resorption and down growth of epithelium from the socket orifice. A multiphase bioactive socket plug (BP) is designed to overcome the natural healing process by maintaining the three-dimensional (3D) volume of extraction sockets, particularly in sockets with wall defects, and later provide sufficient alveolar bone volume for implant placement. The study aimed to fabricate and evaluate the physical, chemical, and biological performance of BPin vitro. The BP was fabricated through freeze-drying and layer-by-layer assembly, comprised of a base serving as a scaffold, a central portion for promoting bone regeneration, an upper buccal portion for maintaining alveolar socket dimension with a covering collagen membrane (Memb) on the top and upper buccal surface to prevent soft tissue infiltration. The BP as the experimental group and a pure collagen plug (CP) as the control group were investigated and compared. Radiograph, scanning electron microscopy, and energy-dispersive spectroscopy mapping confirmed that the four-part BP was successfully assembled and fabricated. Swelling rate analysis indicated that BP, CP, and Memb reached swelling equilibrium within 1 hour. BP exhibited a high remaining weight percentage in collagenase solution (68.81 ± 2.21% on day 90) and sustained calcium ion release, reaching the maximum 0.13 ± 0.04 mmol l-1on day 14. In biological assays, BP exhibited excellent cell proliferation (The OD value increased from 0.02 on day 1 to 0.23 on day 21.). The BP group exhibited higher alkaline phosphatase activity and osteocalcin content than the CP group within 21 days. Memb and BP exhibited outstanding barrier function, as evidenced by Hematoxylin and eosin staining. In summary, the multiphase bioactive socket plug represents a promising scaffold for alveolar ridge preservation application.
Assuntos
Colágeno , Alicerces Teciduais , Alvéolo Dental , Alvéolo Dental/cirurgia , Animais , Colágeno/química , Alicerces Teciduais/química , Materiais Biocompatíveis/química , Regeneração Óssea/efeitos dos fármacos , Processo Alveolar , Proliferação de Células , Microscopia Eletrônica de Varredura , Humanos , Teste de Materiais , Aumento do Rebordo Alveolar/métodos , Extração Dentária , Osteoblastos/citologia , Substitutos Ósseos/química , Substitutos Ósseos/farmacologia , Cicatrização , Cálcio/metabolismo , Cálcio/química , Osteocalcina/metabolismoRESUMO
Currently two factors hinder the use of collagen as building block of regenerative devices: the limited mechanical strength in aqueous environment, and potential antigenicity. Polymeric collagen is naturally found in the cross-linked state and is mechanically tougher than the monomeric, acid-soluble collagen ex vivo. The antigenicity of collagen, on the other hand, is mainly ascribed to inter-species variations in amino acid sequences of the non-helical terminal telopeptides. These telopeptides can be removed through enzymatic treatment to produce atelocollagen, although the effect of this cleavage on triple helix organization, amino acidic composition and thermal properties is often disregarded. Here, we compare the structural, chemical and physical properties of polymeric and monomeric type I collagen with and without telopeptides, in an effort to elucidate the influence of either mature covalent crosslinks or telopeptides. Circular dichroism (CD) was used to examine the triple helical conformation and quantify the denaturation temperature (Td) of both monomeric collagen (36.5°C) and monomeric atelocollagen (35.5°C). CD measurements were combined with differential scanning calorimetry (DSC) in order to gain insight into the triple helix-to-coil thermal transition and shrinkage temperature (Ts) of polymeric atelo collagen (44.8°C), polymeric collagen (62.7°C), monomeric atelo collagen (51.4°C) and monomeric collagen (66.5°C). Structural and thermal analysis was combined with high pressure liquid chromatography (HPLC) to determine the content of specific collagen amino acidic residues used as markers for the presence of telopeptides and mature crosslinks. Hydroxylamine was used as the marker for polymeric collagen, and had a total content of 9.66% for both polymeric and polymeric atelo collagen; tyrosine was used as the marker for telopeptide cleavage, was expressed as 0.526% of the content of polymeric collagen and the partially-reduced content of 0.39% for atelocollagen.
Assuntos
Colágeno Tipo I/química , Sequência de Aminoácidos , Aminoácidos , Dicroísmo Circular , Colágeno , PolímerosRESUMO
Currently one factor hindering the development of collagen hydrogel constructs for tissue engineering is the mismatch between initial cellularity and mechanical strength. The main advantage of collagen hydrogel tissue constructs is their ability to support interstitially seeded cells. However, cells are sensitive to their environment, in particular, substrate stiffness, which cannot easily be replicated within hydrogels without cytotoxic cross-linking treatment. In this study, pre-crosslinked polymeric collagen fibrils are introduced as a starting material, thereby avoiding artificial cross-linking. Shear aggregation of this material in solution results in fibril alignment, but cell addition is only possible when polymeric collagen is blended with its monomeric counterparts to slow the aggregation of collagen fibrils. The hydrogel can then be brought to physiological collagen density by plastic compression. Interstitially seeded fibroblasts were supported for 14days. Although compression of blended gels resulted in some cell death due to increased rate of fluid expulsion, not normally seen in conventional collagen hydrogels, the surviving cell population recovers during subsequent culture. Importantly, the compression process can be controlled and customized to limit cell damage. This is the first report of native polymeric collagen used in a tissue engineering context, for the rapid production of a stiff collagen-cell constructs.