Effects of fibrin-binding oligopeptide on osteopromotion in rabbit calvarial defects

Purpose: Fibronectin (FN) has been shown to stimulate bone regeneration in animal models. The aim of this study was to evaluate the capacity of bovine bone mineral coated with synthetic oligopeptides to enhance bone regeneration in rabbit calvarial defects. Methods: Oligopeptides including fibrin-binding sequences of FN repeats were synthesized on the basis of primary and tertiary human plasma FN structures. Peptide coated and uncoated bone minerals were implanted into 10 mm calvarial defects in New Zealand white rabbits, and the animals were sacrificed at 4 or 8 weeks after surgery. After specimens were prepared, histologic examination and histomorphometric analysis were performed. Results: At 4 weeks after surgery, the uncoated groups showed a limited amount of osteoid formation at the periphery of the defect and the oligopeptide coated groups showed more osteoid formation and new bone formation in the center of the defect as well as at the periphery. At 8 weeks, both sites showed increased new bone formation. However, the difference between the two sites had reduced. Conclusions: Fibrin-binding synthetic oligopeptide derived from FN on deproteinized bovine bone enhanced new bone formation in rabbit calvarial defects at the early healing stage. This result suggests that these oligopeptides can be beneficial in reconstructing oral and maxillofacial deformities or in regenerating osseous bone defects. ⓒ 2010 Korean Academy of Periodontology.

Nontoxic membrane translocation peptide from protamine, low molecular weight protamine (LMWP), for enhanced intracellular protein delivery: in vitro and in vivo study

Naturally derived, nontoxic peptides from protamine by the authors, termed low molecular weight protamines (LMWPs), possess high arginine content and carry significant sequence similarity to that of TAT, by far the most potent protein transduction domain peptide. Therefore, it was hypothesized that these LMWPs would also inherit the similar translocation activity across the cell membrane, which enables any impermeable species to be transduced into the cells. LMWPs were prepared by enzymatic digestion of protamine, examined their capability of transducing an impermeable protein toxin into the tumor cells by chemical conjugation, and determined cytotoxicity of transduced protein toxin (e.g., gelonin) against cancer cell lines and a tumorâ bearing mouse. In vitro results showed that LMWPs could indeed translocate themselves into several mammalian cell lines as efficiently as TAT, thereby transducing impermeable gelonin into the cells by chemical conjugation. In vivo studies further confirmed that LMWP could carry an impermeable gelonin across the tumor mass and subsequently inhibit the tumor growth. In conclusion, the presence of equivalent cell translocation potency, absence of toxicity of peptide itself, and the suitability for lowâ cost production by simple enzymatic digestion could expand the range of clinical applications of LMWPs, including medical imaging and gene/protein therapies.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/154356/1/fsb2fj042322fje-sup-0125.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/154356/2/fsb2fj042322fje.pd

Initial adhesion of bone marrow stromal cells to various bone graft substitutes

Purpose: The aim of this study is to determine whether certain biomaterials have the potential to support cell attachment. After seeding bone marrow stromal cells onto the biomaterials, we investigated their responses to each material in vitro. Methods: Rat bone marrow derived stromal cells were used. The biomaterials were deproteinized bovine bone mineral (DBBM), DBBM coated with fibronectin (FN), synthetic hydroxyapatite (HA), HA coated with FN, HA coated with beta-tricalcium phosphate (TCP), and pure beta-TCP. With confocal laser scanning microscopy, actin filaments and vinculin were observed after 6, 12, and 24 hours of cell seeding. The morphological features of cells on each biomaterial were observed using scanning electron microscopy at day 1 and 7. Results: The cells on HA/FN and HA spread widely and showed better defined actin cytoskeletons than those on the other biomaterials. At the initial phase, FN seemed to have a favorable effect on cell adhesion. In DBBM, very few cells adhered to the surface. Conclusions: Within the limitations of this study, we can conclude that in contrast with DBBM not supporting cell attachment, HA provided a more favorable environment with respect to cell attachment. (C) 2011 Korean Academy of Periodontology.This work was supported by a National Research Foundation of Korea Grant funded by the Government of the Republic of Korea (2008-E00580)

Effect of immobilized cell-binding peptides on chitosan membranes for osteoblastic differentiation of mesenchymal stem cells

Two cell-binding domains from FGF-2 (fibroblast growth factor-2) were shown to increase cell attachment and osteoblastic differentiation. Two synthetic peptides derived from FGF-2, namely residues 36-41 (F36; PDGRVD) and 77-83 (F77; KEDGRLL), were prepared and their N-termini further modified for ease of surface immobilization. Chitosan membranes were used in the present study as mechanical supportive biomaterials for peptide immobilization. Peptides could be stably immobilized on to the surface of chitosan membranes. The adhesion of mesenchymal stem cells to the peptide (F36 and F77)-immobilized chitosan membrane was increased in a dose-dependent manner and completely inhibited by soluble RGD (Arg-Gly-Asp) and anti-integrin antibody, indicating the existence of an interaction between F36/F77 and integrin. Peptide-immobilized chitosan supported human bone-marrow-derived mesenchymal-stem-cell differentiation into osteoblastic cells, as demonstrated by alkaline phosphate expression and mineralization. Taken together, the identified peptide-immobilized chitosan membranes were able to support cell adhesion and osteoblastic differentiation; thus these peptides might be useful as bioactive agents for osteoblastic differentiation and surface-modification tools in bone regenerative therapy.This study was supported in part by the KOSEF (Korea Science and Technology Foundation) Nanobiotechnology Development Program [no. 2007-00952] entitled Regenomics (innovative surface activation of regenerative biomaterials), in part by the Korea Research Foundation [grant no. D00192] and in part by an Engineering Research Center grant from KOSEF through the IBEC (Intelligent Biointerface Engineering Center) [grant no. R11-2000-084-09001-0]

Engineering new bone tissue in vitro on highly porous poly(Α-hydroxyl acids)/hydroxyapatite composite scaffolds

Engineering new bone tissue with cells and a synthetic extracellular matrix (scaffolding) represents a new approach for the regeneration of mineralized tissues compared with the transplantation of bone (autografts or allografts). In the present work, highly porous poly( L -lactic acid) (PLLA) and PLLA/hydroxyapatite (HAP) composite scaffolds were prepared with a thermally induced phase separation technique. The scaffolds were seeded with osteoblastic cells and cultured in vitro . In the pure PLLA scaffolds, the osteoblasts attached primarily on the outer surface of the polymer. In contrast, the osteoblasts penetrated deep into the PLLA/HAP scaffolds and were uniformly distributed. The osteoblast survival percentage in the PLLA/HAP scaffolds was superior to that in the PLLA scaffolds. The osteoblasts proliferated in both types of the scaffolds, but the cell number was always higher in the PLLA/HAP composite scaffolds during 6 weeks of in vitro cultivation. Bone-specific markers (mRNAs encoding bone sialoprotein and osteocalcin) were expressed more abundantly in the PLLA/HAP composite scaffolds than in the PLLA scaffolds. The new tissue increased continuously in the PLLA/HAP composite scaffolds, whereas new tissue formed only near the surface of pure PLLA scaffolds. These results demonstrate that HAP imparts osteoconductivity and the highly porous PLLA/HAP composite scaffolds are superior to pure PLLA scaffolds for bone tissue engineering. © 2000 John Wiley & Sons, Inc. J Biomed Mater Res 54: 284–293, 2001Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/34419/1/16_ftp.pd

Comparison of high molecular coating type dentine hypersensitivity desensitizers

Purpose : The aim of this study was to evaluate the clinical efficacy of two commercially available desensitizing agents over 3 weeks on patients with dentin hypersensitivity. Materials and Methods : An oxalate-based solution, Sensblok ( NIBEC Inc. Seoul, Korea) & Superseal (Pheonix Inc. Michigan, USA), and a distilled water placebo were compared in a clinical setting. Seven volunteers exhibiting three or more teeth that were sensitive to tactile & air were enrolled in the study (35 teeth total). A visual analog scale (VAS) was used for recording each patients level of sensitivity to tactile & air stimuli. A Florida probe was rubbed across the exposed dentin three times with a constant pressure 20 grams to measure the tactile stimulus. The air stimulus was generated by dental unit air syringe for 1-second blast from 1/2 inch distance. The desensitizing agents were applied according manufactures guideline. VAS scores were recorded at baseline, 1 minute after treatment, at 1 week, 2weeks, 3weeks. VAS score were analyzed using a repeated measures ANOVA (α=0.05). Results : The Sensblok, Superseal, and distilled water placebo all decreased dentin sensitivity. A statistically significant difference existed between Sensblok and placebo when tactile stimulus was applied (P<0.05), but no significant difference was found among other desensitizing agents and tactile and air stimuli

The effect of the surface modification of titanium using a recombinant fragment of fibronectin and vitronectin on cell behavior

The surface of titanium implants is in direct contact with host tissue and plays a critical role in determining biocompatibility. Fibronectin (FN) and vitronectin (VN) are major cell adhesive proteins found in the extracellular matrix (ECM) of various tissues, and in circulating blood. The aim of this study was to evaluate the engineered biomimetic surface of titanium by using recombinant fragment of FN8–10 and VNNTD that contains the binding site for integrins. MC3T3-E1 cells seeded upon the FN8−10-coated titanium showed a marked increase in cell adhesion, proliferation, and differentiation over VNNTD-coated titanium. In addition, we confirmed that the surface properties of titanium prefer for FN8–10 over VNNTD (p<0.05) in protein adhesion. These results suggest that the FN8–10-modified titanium surface can be used to improve the osseointegration of titanium implants by enhancing bone formation

Controlled release of platelet-derived growth factor from porous poly(L-lactide) membranes for guided tissue regeneration

Platelet-derived growth factor-BB (PDGF-BB) was incorporated into porous poly (L-lactide) (PLLA) membranes with an aim of improving early bone healing in guided tissue regeneration (GTR) therapy. Porous PDGF-BB loaded membranes were fabricated by coating PDGF-BB-dissolved PLLA methylene chloride–ethyl acetate solutions on polyglycolic acid (PGA) meshes. Release kinetics of PDGF-BB, biologic activity, degradability and guided tissue regenerative potentials of the membranes were investigated. Release of PDGF-BB could be controlled by adding bovine serum albumin that may provide porous diffusion channels for PDGF-BB release and by varying initial loading content of PDGF-BB. Biologic activity of PDGF-BB in the membranes was ascertained by fibroblast chemotaxis. PDGF-BB loaded membranes maintained proper degradation property for periodontal application. PDGF-BB loaded membrane markedly increased new bone formation in rat calvarial defects, and completed bony reunion after 2 weeks of implantation period. These results suggested that PDGF-BB loaded PLLA membrane might potentially enhance guided tissue regenerative efficacy.The authors are thankful to Sam Yang Group R and D Center for providing materials. This work was supported by the grant of Korea Science and Engineering Foundation (KOSEF) #94-0403-05-02-3, Korea

Enhanced fibronectin-mediated cell adhesion of human osteoblast by fibroblast growth factor, FGF-2

Using specific recombinant human fibronectin peptide (hFNIII9-10) that contains the binding site for integrin, we found that the fibroblast growth factor, FGF-2, enhances fibronectin-mediated adhesion in human osteoblast-like MG63 cells. The mechanism of the synergistic adhesion was due to the activation of extracellular-regulated kinase (ERK)-type MAPK upon interaction of integrin to hFNIII9-10 and its downstream activation of signaling pathways