Reactive oxygen species (ROS) are involved in the initiation and progression of atherosclerosis. ROS-derived hydroperoxides, as an indicator of ROS production, have been measured by using the diacron reactive oxygen metabolites (d-ROMs) test, which requires iron-containing transferrin in the reaction mixture. In this study we developed a modified d-ROMs test, termed the Fe-ROMs test, where iron ions were exogenously added to the reaction mixture. This modification is expected to exclude the assay variation that comes from different blood iron levels in individuals. In addition, this Fe-ROMs test was helpful for determining the class of plasma lipoproteins that are hydroperoxidized. Low-density lipoprotein/very low-density lipoprotein (LDL/VLDL) and high-density lipoprotein (HDL) were purified by use of an LDL/VLDL purification kit and the dextran sulfate-Mg2+ precipitation method, respectively; their hydroperoxide contents were assessed by performing the Fe-ROMs test. The majority of the hydroperoxides were detected only in the HDL fraction, not in the LDL/VLDL. Further detailed analysis of HDLs by size-exclusion high-performance liquid chromatography revealed that the hydroperoxide-containing molecules were small-sized HDLs. Because HDL was shown to be the principal vehicle for the plasma hydroperoxides, this Fe-ROMs test is a beneficial method for the assessment of oxidized-HDL levels. Indeed, Fe-ROMs levels were strongly associated with the levels of oxidized HDL, which were determined by performing the malondialdehyde-modified HDL enzyme immunoassay. In conclusion, the Fe-ROMs test using plasma itself or the HDL fraction after dextran sulfate-Mg2+ precipitation is useful to assess the functionality of HDL, because the oxidation of HDL impairs its antiatherogenic capacity.

Kawasaki disease (KD) is a systemic vasculitis in infants that develops predominantly in the coronary arteries. Despite the clinically transient nature of active inflammation in childhood albeit rare complications (e.g., coronary artery aneurysm), KD has recently been suggested to increase the incidence of ischemic heart diseases in young adulthood. However, little is known about the histopathology of the coronary artery long after development of the acute KD vasculitis. To address this, we conducted histological studies of rabbit coronary arteries in adolescent phase after induction of the KD-like vasculitis induced by horse serum administration. After a transmural infiltration of inflammatory cells in acute phase at day 7, the artery exhibited a gradual decrease in the number of inflammatory cells and thickening of the intima during the chronic phase up to day 90, where proteoglycans were distinctly accumulated in the intima with abundant involvement of α-smooth muscle actin (α-SMA)-positive cells, most of which accompanied expression of VCAM-1 and NF-κB. Distinct from classical atherosclerosis, inflammatory cells, e.g., macrophages, were barely detected during the chronic phase. These observations indicate that the KD-like coronary arteritis is followed by intimal thickening via accumulation of proteoglycans and proliferation of α-SMA-positive cells, reflecting aberrant coronary artery remodeling.

Apelin is a selective endogenous ligand of the APJ receptor, which genetically has closest identity to the angiotensin II type 1 receptor (AT-1). The effects of the apelin/APJ system on renal fibrosis still remain unclear.

In this study, we developed porous medium cross-linked recombinant collagen peptide (mRCP) with two different ranges of interconnected pore sizes, Small-mRCP (S-mRCP) with a range of 100-300 μm and Large-mRCP (L-mRCP) with a range of 200-500 μm, to compare the effect of pore size on bone regeneration in a calvarial bone defect.

We recently demonstrated the bone-forming potential of medium-cross-linked recombinant collagen peptide (mRCP) in animal models of bone defects. However, these studies were limited to a 4-week observation period; therefore, in the present study, we aimed to further evaluate mRCP as a suitable bone graft material for the alveolar cleft by analyzing its bone-forming potential, osteogenic-inducing ability, and biodegradation over an extended period of 12 weeks, using a rat critical-size calvarial defect model.

Guided bone regeneration (GBR) is an accepted method in dental practice that can successfully increase the bone volume of the host at sites chosen for implant placement; however, existing GBR membranes exhibit rapid absorption and lack of adequate space maintenance capabilities. We aimed to compare the effectiveness of a newly developed resorbable bilayer membrane composed of poly (L-lactic acid) and poly (-caprolactone) (PLACL) with that of a collagen membrane in a rat GBR model.

Lipid-free fibroblast-like cells, known as dedifferentiated fat (DFAT) cells, can be generated from mature adipocytes with a large single lipid droplet. DFAT cells can re-establish their active proliferation ability and can transdifferentiate into various cell types under appropriate culture conditions. The first objective of this study was to compare the multilineage differentiation potential of DFAT cells with that of adipose-derived stem cells (ASCs) on mesenchymal stem cells. We obtained DFAT cells and ASCs from inbred rats and found that rat DFAT cells possess higher osteogenic differentiation potential than rat ASCs. On the other hand, DFAT cells show similar adipogenic differentiation, and chondrogenic differentiation potential in comparison with ASCs. The second objective of this study was to assess the regenerative potential of DFAT cells combined with novel solid scaffolds composed of PLGA (Poly d, l-lactic-co-glycolic acid) on periodontal tissue, and to compare this with the regenerative potential of ASCs combined with PLGA scaffolds. Cultured DFAT cells and ASCs were seeded onto PLGA scaffolds (DFAT/PLGA and ASCs/PLGA) and transplanted into periodontal fenestration defects in rat mandible. Micro computed tomography analysis revealed a significantly higher amount of bone regeneration in the DFAT/PLGA group compared with that of ASCs/PLGA and PLGA-alone groups at 2, 3, and 5 weeks after transplantation. Similarly, histomorphometric analysis showed that DFAT/PLGA groups had significantly greater width of cementum, periodontal ligament and alveolar bone than ASCs/PLGA and PLGA-alone groups. In addition, transplanted fluorescent-labeled DFAT cells were observed in the periodontal ligament beside the newly formed bone and cementum. These findings suggest that DFAT cells have a greater potential for enhancing periodontal tissue regeneration than ASCs. Therefore, DFAT cells are a promising cell source for periodontium regeneration.

The effect of NiCl2 on oral squamous cell carcinoma-derived cell line HSC3 was examined. Incubation with 1 mM NiCl2 significantly reduced the expression of MMPs at mRNA and protein levels. The in vivo orthotopic implantation model was established by injecting highly metastatic subcell line HSC3-M3 to nude mouse tongue. After 1 week of injection, mice were fed with or without 1 mM NiCl2-containing water for two to three weeks. Immunohistochamical examination revealed that MMP9 expression was drastically reduced in NiCl2-fed mice. By CT images, cancer mass was observed as a translucent area in control mice. In NiCl2-fed mice, much highly translucent area was observed within the translucent area. Histologically, this area corresponded to the necrotic area in the tumor mass. Real-time PCR analysis revealed the reduced expression of angiogenic factors such as IL-8 and VEGF mRNA in NiCl2-fed mice. To further examine the effect of NiCl2 on metastasis, human β-globin gene expression in regional lymphnodes was compared. The β-globin gene was totaly absent in NiCl2-fed mice. Moreover, various cancer metastasis-related genes were inhibited in NiCl2-fed mice by PCR array analysis. The results indicated that NiCl2 might be a promising new anti-cancer therapeutics for the oral cancer treatment.

This study aimed to examine the optimal cross-link density of recombinant peptide (RCP) particles, based on human collagen type I, for bone reconstruction in human alveolar cleft. Low- (group 1), medium- (group 2), and high- (group 3) cross-linked RCP particles were prepared by altering the duration of the heat-dependent dehydration reaction. Rat palatine fissures (n = 45), analogous to human congenital bone defects, were examined to evaluate the potential of bone formation by the three different RCP particles. Microcomputed tomography images were obtained to measure bone volume and bone mineral density at 4, 8, 12, and 16 weeks post grafting. Specimens were obtained for histological analysis at 16 weeks after grafting. Additionally, alkaline phosphatase and tartrate acid phosphatase staining were performed to visualize the presence of osteoblasts and osteoclasts. At 16 weeks, bone volume, bone mineral density, and new bone area measurements in group 2 were significantly higher than in any other group. In addition, the number of osteoblasts and osteoclasts on the new bone surface in group 2 was significantly higher than in any other group. Our results demonstrated that medium cross-linking was more suitable for bone formation-and could be useful in human alveolar cleft repairs as well.

Mesenchymal stem cells (MSCs) are known to have different differentiation potential depending on the tissue of origin. Dedifferentiated fat cells (DFATs) are MSC-like multipotent cells that can be prepared from mature adipocytes by ceiling culture method. It is still unknown whether DFATs derived from adipocytes in different tissue showed different phenotype and functional properties. In the present study, we prepared bone marrow (BM)-derived DFATs (BM-DFATs), BM-MSCs, subcutaneous (SC) adipose tissue-derived DFATs (SC-DFATs), and adipose tissue-derived stem cells (ASCs) from donor-matched tissue samples. Then, we compared their phenotypes and multilineage differentiation potential in vitro. We also evaluated in vivo bone regeneration ability of these cells using a mouse femoral fracture model.

Kawasaki Disease (KD) involves a diffuse and systemic vasculitis of unknown etiology that mainly affects infants and children. Although a considerable number of analyses of the clinical, histopathological and molecular biological details underlying the mechanism responsible for the development of coronary arterial lesions, it is still poorly understood.The purpose of this study was to analyze the state of angiogenesis, vasculogenesis and the distribution of blood vessels using an animal model of KD like vasculitis. We investigated the involvement of the vasa vasorum from the adventitia in the vascular involvement and the development of the disease state by performing sequential histopathology, scanning electron microscopy (SEM) and micro computed tomography (CT) studies using a murine model of vasculitis induced by the Candida albicans water-soluble fraction (CAWS).

The methyltransferase G9a was originally isolated as a histone methyltransferase that catalyzes the methylation of histone 3 lysine 9 (H3K9) to a dimethylated state (H3K9me2). Recent studies have revealed that G9a has multiple functions in various cells, including osteoblasts. Here, we investigated G9a function during cranial bone formation. Crossing Sox9-cre with G9aflox/flox (fl/fl) mice generated conditional knockout mice lacking G9a expression in Sox9-positive neural crest-derived bone cells. Sox9-Cre/G9afl/fl mice showed severe hypo-mineralization of cranial vault bones, including defects in nasal, frontal, and parietal bones with opened fontanelles. Cell proliferation was inhibited in G9a-deleted calvarial bone tissues. Expression levels of bone marker genes, i.e., alkaline phosphatase and osteocalcin, were suppressed, whereas Runx2 expression was not significantly decreased in those tissues. In vitro experiments using G9a-deleted calvarial osteoblasts showed decreased cell proliferation after G9a deletion. In G9a-deleted osteoblasts, expression levels of fibroblast growth factor receptors and several cyclins were suppressed. Moreover, the expression of bone marker genes was decreased, whereas Runx2 expression was not altered by G9a deletion in vitro. G9a enhanced the transcriptional activity of Runx2, whereas siRNA targeting G9a inhibited the transcriptional activity of Runx2 in C3H10T1/2 mesenchymal cells. We confirmed the direct association of endogenous Runx2 with G9a. Chromatin immunoprecipitation experiments showed that G9a bound to Runx2-target regions in promoters in primary osteoblasts. Furthermore, Runx2 binding to the osteocalcin promoter was abrogated in G9-deleted osteoblasts. These results suggest that G9a regulates proliferation and differentiation of cranial bone cells through binding to and activating Runx2.