The purpose of the present study was to evaluate effects of VEGF on osteosclerosis with poor blood supply. Teflon cylinders, of which bottoms were covered with e-PTFE membranes, were embedded in two anterior and two posterior regions one of the rabbit skulls. The cylinders were packed with collagen sponge containing 0.2 µg VEGF or sterile physiological saline. Four weeks later, the membranes were removed. Histological analysis was performed before removing the membrane (week0), one week (week1), two weeks (week2), three weeks (week3), and four weeks (week4) after removing the membrane. At week4, newly-formed bone was analyzed with µCT and image analysis software. Compared to the anterior region, blood supply was anatomically favorable in the posterior region. At week0, favorable neovascularization was seen in the posterior cylinder with VEGF. However, slightly inferior neovascularization was found in the anterior cylinder with VEGF. After week2, the most extensive osteogenesis was observed in the posterior cylinder with VEGF. At week4, newly-formed bone thickened and matured. The amount of newly-formed bone in the anterior cylinder with physiological saline was the smallest. The results suggest that VEGF can facilitate neovascularization and osteogenesis in osteosclerosis.
We prepared a three-dimensional cell culture system using atelocollagen gel derived from chum salmon sources. To examine whether collagen gel derived from salmon is suitable as a scaffold for ES-D3 cell differentiation, we compared the degrees of differentiation of cells cultured in a salmon collagen (FC), a conventional collagen gel (CG), and a collagen sponge derived from porcine sources (CS). No significant differences in cell differentiation were noted between cells cultured with the collagen gel derived from salmon and the control cells, the differentiation of which was inhibited by adding mLIF. With FC, efficient differentiation of ES-D3 cells was not observed, whereas both the collagen gel supported cell differentiation and the collagen sponge derived from the porcine sources. The results of studies using ES cells have proven useful in studies of ES cell differentiation factors. Moreover, these study results may yield important clues as to cell differentiation factors for adult stem cells. To that end, the development of scaffolds that facilitate the in vitro culturing of ES cells for long periods is desirable.
Type I collagen hydrogel treated by an ascorbate-copper ion crosslinking system is highly biocompatible and highly degradable in the body. The purpose of this study was to examine the effect of collagen hydrogel on periodontal wound healing in beagles. Sixty-four periodontal dehiscence type defects were created on the buccal roots of four beagles. Subsequently, collagen hydrogel was implanted in each defect in the experimental group and sutured, and, in the control group, the defect was sutured without any application of collagen hydrogel. The percentage of the length of new bone and new cementum in the experimental group was significantly greater than that in the control group at weeks 4 (p<0.01) and 8 (p<0.05). Significantly more junctional epithelium was observed in the control group than in the experimental group at weeks 2 (p<0.01) and 4 (p<0.05). These findings thus indicate that periodontal regeneration was stimulated by collagen hydrogel implantation in beagles.
In the present study, we prepared lipid-coated basic-fibroblast growth factor (b-FGF) and evaluated its tissue response. The lipid, didodecyl N-D-glucono-L-glutamate (2C12-Glu) was prepared from didodecyl L-glutamate and D-glucono-1,5-lactone, and the lipid-coated b-FGF was prepared from the mixing reaction of 2C12-Glu in dioxane solution and b-FGF in phosphate-buffered saline solution. After centrifugation and freeze drying, liplid-coated b-FGF was obtained as a white substance. The lipid-coated b-FGF was insoluble in water or any aqueous buffer solution, but freely soluble in most organic solvent. Ultraviolet spectra measurement of lipid-coated b-FGF between 250-300 nm indicated that 15% of b-FGF was coated with lipid. Circular dichroism spectroscopy measurement confirmed that the conformation of b-FGF was maintained in organic solvent after lipid-coating. The lipid-coated b-FGF or lipid (2C12-Glu) alone was implanted in the tibia of rats. After 2 weeks of implantation, histological observation revealed that the lipid-coated b-FGF induced the new bone formation faster than lipid (2C12-Glu) alone. In conclusion, lipid-coated b-FGF will be useful for bone regeneration in drug delivery system or tissue engineering.
Objectives; To establish a method for nondestructive observation of bone in peri-implant tissue, we compare radiographs with soft X-ray, microcomputed tomography (micro-CT) and confocal laser scanning microscopy (CLSM). Materials and Methods; After extraction of the 3rd incisors in the maxillary bone of beagle dogs (n=3), six titanium implants were placed in the bone. Two fluorescent dyes, calcein and alizarin red, were administered to the animals at different weeks. At 4, 8 and 12 weeks after implantation, the animals were sacrificed and temporal patterns of new bone formation at the interface between implant and host bone were observed in undecalcified materials using soft X-ray, micro-CT and CLSM. Results; Bone density was analyzed with dental X-ray and soft X-ray. Micro-CT enables three-dimensional analysis of the bone in peri-implant tissue. CLSM had the advantage of enabling observation of the process of calcification at high magnification. In addition, the analysis of CLSM showed that a little newly formed bone appeared close to the host bone at 4 weeks after implantation, after which porous bone formation tended to increase with time up to 12 weeks, and filled the screw grooves of the implant. It was concluded that these three techniques are useful in assessing the relation of bone formation to implant design and surgical methods.
The history of animal use in research laboratories around the world is outlined with particular attention given to the concept of alternatives (the three Rs of Replace-ment, Reduction and Refinement). Using data from the UK and the USA, the paper demonstrates how far the "alternatives" concept has advanced in the last thirty years and how much progress has been made over that period.