A bi-phasic scaffold consisting of a columnar formaldehyde-acetalized polyvinyl alcohol (PVF) sponge and a cylindrical porous hydroxyapatite (HA) with a hollow center was devised. Rat bone marrow cells (rBMCs) were seeded into the sponge placed in the hollow center of the cylindrical porous HA. The bi-phasic scaffold, a cylindrical porous HA and a PVF sponge separated from a bi-phasic scaffold after rBMC seeding, and a PVF sponge without rBMCs as a negative control, were implanted for 6 weeks into rat dorsal subcutaneous tissue. In each construct, bone formation was examined histologically and osteocalcin was measured immunochemically. Bone formation was observed in the bi-phasic scaffold and also in the cylindrical porous HA isolated from the bi-phasic scaffold. A significant difference in the quantity of osteocalcin was observed between the bi-phasic scaffold and the isolated cylindrical porous HA. No bone formation was found in the isolated PVF sponge. The bi-phasic scaffold as an outer layer of the scaffold seemed to inhibit the outflow of rBMCs from the PVF sponge. This type of bi-phasic scaffold may have two specific characteristics: Attachment of cells both in PVF sponge and cylindrical porous HA.

Because of the three-dimensional structure of bone or hard tissue such as a tooth, a scaffold is necessary for its regeneration by cellular engineering. Commonly, for in vivo examination, hydroxyapatite (HA) has been used as such a scaffold. Cylindrical HA with a hollow center, which included a columnar formalin-treated polyvinyl alcohol sponge, was used in this examination as a scaffold. The sponge had been coated with L-tryptophan or L-lysine before insertion into the hollow center of the HA. Rat bone marrow cells (rBMCs) derived from the femur were seeded in the sponge before insertion into the hollow center of HA. The number of rBMCs seeded in each sponge was 1.5 × 10 6 . These scaffolds were implanted subcutaneously into the backs of Fischer 344 rats for 6 weeks. In the amino-acid-coated sponge in HA, osteogenesis was found histologically. An osteocalcin level of approximately 10 µg was measured in the scaffolds with L-tryptophan-coated formalized polyvinyl alcohol sponge containing rBMCs, 4 µg on average in the scaffolds with L-lysine-coated sponge containing the cells and about 2 µg in each scaffold with uncoated sponge containing the cells. The structure of the scaffolds used in this study was thought to be suitable for osteogenesis by rBMCs. It was concluded that tryptophan, as a factor for bone formation by stem cells, functioned by promoting cell adhesion and the differentiation of stem cells into osteoblasts.

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Enterococcus faecalis is an etiological agent of endodontic infections. The present study was performed to investigate the gene profiles of E. faecalis induced by type I collagen stimulation. E. faecalis ATCC 19433 was cultivated with [collagen (+)] or without type I collagen [collagen (−)], and transcriptome analysis was performed using high-throughput sequencing technology. A total of 3.6 gb of information was obtained by sequence analysis and 77 differentially expressed genes (DEGs) between the two culture conditions were identified. Among the 77 DEGs, 35 genes were upregulated in collagen (+) E. faecalis, whereas 42 genes were downregulated. Gene Ontology (GO) enrichment analysis was performed and 11 GO terms, including metalloendopeptidase activity (GO:0004222) and two related GO terms (GO:0031012, GO:0044421), were significantly enriched in the set of upregulated genes. We focused on an upregulated DEG belonging to the matrixin metalloprotease gene family, and matrix metalloprotease (MMP) activities of the bacterial cell were examined. The generic MMP, MMP-8, and MMP-9 activities of collagen (+) E. faecalis were significantly higher than those of collagen (−) E. faecalis. These results suggested that contact with type I collagen may alter the gene expression profile of E. faecalis, and upregulation of metalloprotease genes may result in enhanced MMP activities in E. faecalis.

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