High-mobility group box 1 accelerates distraction osteogenesis healing via the recruitment of endogenous stem/progenitor cells.

BACKGROUND AIMS: While distraction osteogenesis (DO) achieves substantial bone regeneration, prolonged fixation may lead to infections. Existing stem cell and physical therapies have limitations, requiring the development of novel therapeutic approaches. Here, we evaluated high-mobility group box 1 (HMGB1) as a novel therapeutic target for DO treatment. METHODS: Micro-computed tomography (Micro-CT) analysis and histological staining of samples obtained from tibial DO model mice was performed. Transwell migration, wound healing, and proliferation assays were also performed on cultured human mesenchymal stem cells (hMSCs) and human umbilival vein endothelial cells (HUVECs). Tube formation assay was performed on HUVECs, whereas osteogenic differentiation assay was performed on hMSCs. RESULTS: Micro-CT analysis and histological staining of mouse samples revealed that HMGB1 promotes bone regeneration during DO via the recruitment of PDGFRα and Sca-1 positve (PαS+) cells and endothelial progenitor cells. Furthermore, HMGB1 accelerated angiogenesis during DO, promoted the migration and osteogenic differentiation of hMSCs as well as the proliferation, migration and angiogenesis of HUVECs in vitro. CONCLUSIONS: Our findings suggest that HMGB1 has a positive influence on endogenous stem/progenitor cells, representing a novel therapeutic target for the acceleration of DO-driven bone regeneration.

Comparison study of the Le Fort I osteotomy using 2- and 4-plate fixation.

This study was conducted to evaluate the postsurgical stability of Le Fort I osteotomy using zygomatic buttress internal fixation alone with no piriform aperture internal fixation. Patients with maxillary retrognathia and mandibular prognathism underwent the Le Fort I osteotomy with a bilateral sagittal split ramus osteotomy. In group I, fixation was accomplished using titanium plate and screws placed at the piriform aperture and the zygomatic buttress (4 plates). In group II, fixation was accomplished using titanium plate and screws placed at the zygomatic buttress (2 plates). Lateral cephalometric radiographs were taken preoperatively (T1), immediately after surgery (T2), and at 6 months to 1 year (T3) to evaluate skeletal movement. In total, 32 patients were included in this study. None of the patients had wound infection, dehiscence, bone fragment instability, and long-term malocclusion. Regarding point A and the posterior nasal spine (PNS), vertical and horizontal relapse in groups I and II did not differ significantly. In most hospitals, the maxilla was fixed using four plates (piriform aperture and zygomatic buttress); however, within the limitations of the study, the choice of the number of plates for osteosynthesis following Le Fort I osteotomy and repositioning of the maxilla can be left to the discretion of the surgeon without putting the patients at risk for increased relapse by careful intraoperative management.

Isolation and structural characterization of bioactive glycosaminoglycans from the green-lipped mussel Perna canaliculus.

Chondroitin sulfate (CS) and heparan sulfate (HS) are sulfated glycosaminoglycan (GAG) chains that consist of repeating disaccharide units composed of hexosamine and hexuronic acid. GAG chains exhibit diverse bioactivities in a structure-specific manner. Marine invertebrates are a rich source of highly sulfated and rare structures of GAG chains. Here, we isolated GAGs from the green-lipped mussel Perna canaliculus, an aquaculture species that is produced on a large scale. We separated GAGs based on the degree of negative charges and analyzed their disaccharide compositions. CS and HS both exhibited characteristic compositions of differently sulfated disaccharides. CS chains showed a higher degree of sulfation than HS chains and contained a high percentage of the E unit disaccharide GlcA-GalNAc(4,6-O-disulfate). Furthermore, CS chains rich in the E unit stimulated the neurite outgrowth of primary cultured neurons. The present results indicate the potential of P. canaliculus GAGs as biomaterials to study the structure-function relationships of GAGs.

Conditioned media from mesenchymal stromal cells and periodontal ligament fibroblasts under cyclic stretch stimulation promote bone healing in mouse calvarial defects.

BACKGROUND AIMS: When cells are exposed to stresses such as mechanical stimuli, they release growth factors and adapt to the surrounding environment H ere, we demonstrated that mechanical stimulation during culture affects the production of osteogenic and angiogenic factors. METHODS: Human bone marrow derived mesenchymal stromal cells (hMSCs) and human periodontal ligament fibroblasts (HPLFs ) were cultured under cyclic stretch stimulation for 24 h. Collected of the cells and conditioned media (CM), the gene and protein expression levels of osteogenic and angiogenic factors were evaluated. CM was also evaluated for angiogenic activity and calc ification ability. In in vivo study, CM was administered to a mouse calvarial defect model and histologically and radiologically evaluated. RESULTS: Quantitative real time polymerase chain reaction results showed that the expression of bone morphogenetic pro tein 2, 4 (BMP 2, 4), vascular endothelial growth factor A (VEGF A), and platelet derived growth factor AA (PDGF AA) was upregulated in the cyclic stretch stimulation group in comparison with the non stretch group in each cell type. Enzyme linked immunosor bent assay results revealed that the expression of BMP 2,4, VEGF A was upregulated in the cyclic stretch group in comparison with the non stretch group in each cell type. Only HPLFs showed significant difference in PDGF AA expression between the cyclic str etch and the non stretch group. Tube formation assay and Alizarin Red S staining results showed that angiogenic activity and calcification ability of CM was upregulated in the cyclic stretch stimulation group in comparison with the non stretch group in eac h cell type. CM was administered to the mouse calvarial defect model. Histological and radiological examination showed that the bone healing was promoted by CM from the cyclic stretch culture group. Immunohistological staining revealed that CM from cyclic stretch group have greater angiogenic effect than CM from the non stretch group. CONCLUSIONS: These results indicate that osteogenesis was promoted by CM obtained under cyclic stretch stimulation through the increase of angiogenesis in the mouse calvarial defect model.

Geranylgeraniol Induces PPARγ Expression and Enhances the Biological Effects of a PPARγ Agonist in Adipocyte Lineage Cells.

BACKGROUND: The global incidence of diabetes mellitus (DM) has risen precipitously, even in middle- and low-income countries. Peroxisome proliferator-activated receptor γ (PPARγ) plays an important role in the control of cellular glucose metabolism. Activation of PPARγ beneficially results in increased insulin sensitivity. However, the expression of PPARγ is reduced by obesity and several nutritional factors. Here we examined the effect of geranylgeraniol (GGOH), a bioactive compound found naturally in fruits, vegetables, and grains, on the expression and activation of PPARγ. MATERIALS AND METHODS: C3H10T1/2 mouse embryonic fibroblasts and 3T3-L1 pre-adipocytes were used as in vitro models of adipocyte differentiation and function. Quantitative reverse-transcriptase polymerase chain reaction, western blotting, Oil Red O staining, and luciferase assay were performed to respectively assess mRNA expression, protein levels, lipid droplet formation and transcriptional activity. RESULTS: GGOH increased the expression of PPARγ in adipocyte lineage cells. GGOH also enhanced adipogenesis induced by rosiglitazone, a thiazolidinedione class PPARγ agonist. CONCLUSION: GGOH induces PPARγ expression and enhances the biological effects of a PPARγ agonist in adipocyte lineage cells.

Salmon cartilage proteoglycan attenuates allergic responses in mouse model of papain­induced respiratory inflammation.

Proteoglycan (PG) is a complex glycohydrate, which is widely distributed in the extracellular matrix. It has been reported that daily oral administration of PG (extracted from salmon nasal cartilage) modulates the severity of proinflammatory cytokine responses in mouse experimental colitis, autoimmune encephalomyelitis, collagen­induced arthritis and obesity­induced inflammation. The present study investigated the effect of salmon nasal cartilage PG on allergic responses using a mouse model of papain­induced respiratory inflammation. Low titers of immunoglobulin E were identified in the sera of the PG­administered mice. Oral administration of PG attenuated eosinophil infiltration in the lung. In the acute model of papain­induced allergic inflammation, PG­administered mice exhibited low titers of epithelium­derived and T helper 2­associated cytokines. The results of the present study demonstrated that salmon cartilage PG has an immunomodulatory effect on intranasally delivered papain. These results suggest a potential role for PG as a prophylactic agent which may attenuate allergic respiratory inflammation.

Evaluation of the effect of salmon nasal proteoglycan on biomarkers for cartilage metabolism in individuals with knee joint discomfort: A randomized double-blind placebo-controlled clinical study.

A randomized double-blind placebo-controlled clinical trial was conducted to evaluate the chondroprotective action of salmon nasal cartilage proteoglycan on joint health. The effect of oral administration of proteoglycan (10 mg/day) on cartilage metabolism was evaluated in individuals with knee joint discomfort but without diagnosis of knee osteoarthritis. The average age of patients was 52.6±1.1 years old. The effect of proteoglycan was evaluated by analyzing markers for type II collagen degradation (C1,2C) and synthesis (PIICP), and the ratio of type II collagen degradation to synthesis. The results indicated that the change in C1,2C levels significantly differed in the proteoglycan group compared with the placebo group following 16 weeks intervention among subjects with high levels of knee pain and physical dysfunction (total score of Japan Knee Osteoarthritis Measure ≥41) and subjects with constant knee pain (both P<0.05). There was a greater increase in PIICP levels in the proteoglycan group than the placebo group following intervention, although this difference was not significant in both sets of patients. Thus, the C1,2C/PIICP ratios decreased in the proteoglycan group, whereas they slightly increased in the placebo group following the intervention. Furthermore, no test supplement-related adverse events were observed during the intervention. Therefore, oral administration of salmon nasal cartilage proteoglycan at a dose of 10 mg/day may exert a chondroprotective action in subjects with knee joint discomfort. This effect was achieved by improving cartilage metabolism (reducing type II collagen degradation and enhancing type II collagen synthesis), without causing apparent adverse effects.

Cynaropicrin is dual regulator for both degradation factors and synthesis factors in the cartilage metabolism.

AIMS: The molecular mechanism of osteoarthritis (OA) has never been understood clearly, but it has been suggested that imbalance of degradation and synthesis in cartilage contribute to the underlying mechanisms of OA. In this study, we investigated the effectiveness in the cartilage metabolism of the artichoke extract that includes the compound cynaropicrin. MAIN METHODS: We evaluated the efficacy of the artichoke extract or cynaropicrin in the cartilage metabolism factors and NF-κB signaling activity stimulated by inflammatory cytokine in chondrogenic cell lines, OUMS-27 and SW1353, using qRT-PCR, immunofluorescence and immunoblotting. KEY FINDINGS: We initially found that an artichoke extract and cynaropicrin both inhibited the increase of cartilage degradation factor MMP13 and further decreased the synthesis factor aggrecan induced by TNF-α in OUMS-27. In addition, cynaropicrin suppressed the enhancement of master regulator HIF-2α on cartilage degradation and further reduced the master regulator Sox9 on cartilage synthesis induced by TNF-α. We observed that cynaropicrin suppresses NF-κB signaling, which controls HIF-2α and Sox9. Since, HIF-2α is induced by p65 (RelA), we evaluated the effect of cynaropicrin and observed that it suppressed the nuclear transport of p65 (RelA) by inhibiting phosphorylation of IκBα. Moreover, cynaropicrin not only suppressed TNF-α stimulation, it had a similar effect on IL-1ß stimulation. No significant cytotoxicity with cynaropicrin was observed. SIGNIFICANCE: These finding suggest that cynaropicrin is an effective substance that can improve the balance of cartilage metabolism, by altering the equilibrium of cartilage degradation and synthesis induced by multiple mediators know to contribute to OA.

Effects of Salmon Nasal Cartilage Proteoglycan on Plasma Glucose Concentration and Active Glucose Transport in the Small Intestine.

Recently, proteoglycan was purified from the nasal cartilage of salmon. Although several physiological effects have been reported, the effect of salmon nasal cartilage proteoglycan (salmon PG) on glucose metabolism remains unclear. We studied the effect of salmon PG on rat plasma glucose levels. Oral administration of 1% salmon PG significantly attenuated the increase in portal plasma glucose levels following an oral glucose tolerance test (OGTT). Additionally 1% salmon PG delayed the increase in peripheral glucose concentration induced by the OGTT. Mucosal administration of 1% salmon PG significantly decreased active glucose transport using the everted jejunal sac method. Furthermore, transmural potential difference (ΔPD) measurements using the everted jejunum revealed that 1% salmon PG significantly decreased glucose-dependent and phlorhizin (inhibitor of sodium-glucose co-transporter 1; SGLT1)-sensitive ΔPD. These results suggest that salmon PG decreases glucose absorption via SGLT1 in the jejunum, thereby attenuating the increase in portal and peripheral plasma glucose levels in rats.

Absorption of proteoglycan via clathrin-mediated endocytosis in the small intestine of rats.

The mechanism underlying proteoglycan (PG) absorption in the intestine is not clear. Hence we analyzed the transport of salmon PG in the rat jejunum, ileum, and colon by the everted-sac method. The jejunum showed the largest capacity for PG transport. Jejunal transport of PG was also greater than that of chondroitin A and C. An inhibitor of clathrin-mediated endocytosis reduced jejunal PG transport. We conclude that intestinal PG transport is highest in the jejunum, and is partially dependent on clathrin-mediated endocytosis.

Cynaropicrin from Cynara scolymus L. suppresses photoaging of skin by inhibiting the transcription activity of nuclear factor-kappa B.

Aging of skin is characterized by skin wrinkling, laxity, and pigmentation induced by several environmental stress factors. Histological changes during the photoaging of skin include hyperproliferation of keratinocytes and melanocytes causing skin wrinkles and pigmentation. Nuclear factor kappa B (NF-κB) is one of the representative transcription factors active in conjunction with inflammation. NF-κB is activated by stimulation such as ultraviolet rays and inflammatory cytokines and induces the expression of various genes such as those of basic fibroblast growth factor (bFGF) and matrix metalloprotease-1 (MMP-1). We screened several plant extracts for their possible inhibitory effect on the transcriptional activity of NF-κB. One of them, an extract from Cynara scolymus L., showed a greatest effect on the suppression of NF-κB transactivation. As a result, we found that cynaropicrin, which is a sesquiterpene lactone, inhibited the NF-κB-mediated transactivation of bFGF and MMP-1. Furthermore, it was confirmed that in an in vivo mouse model cynaropicrin prevented skin photoaging processes leading to the hyperproliferation of keratinocytes and melanocytes. These findings taken together indicate that cynaropicrin is an effective antiphotoaging agent that acts by inhibiting NF-κB-mediated transactivation.

Hyuganatsu orange (Citrus tamurana Hort. Ex Tanaka) contains a water soluble substance that suppresses bone loss in ovariectomized rats.

We investigated whether Hyuganatsu orange (Citrus tamurana Hort. ex Tanaka) contains water and acetic-acid soluble substances that increase bone mineral density (BMD) in ovariectomized rats. In in vivo study, femoral BMD can significantly increased. In in vitro study, tartrate-resistant acid phosphatase (TRAP) positive cells significantly decreased. We speculate that Hyuganatsu orange contains biologically active substances other than hesperidin that increase BMD.

Nitrocapsanthin and nitrofucoxanthin, respective products of capsanthin and fucoxanthin reaction with peroxynitrite.

The in vitro reactivity of capsanthin (1) and fucoxanthin (2) with peroxynitrite was investigated, and the reaction products produced by scavenging with peroxynitrite were analyzed. (14'Z)-Nitrocapsanthin (3) and 12-nitrocapsanthin (4) were isolated from the products of the reaction of capsanthin with peroxynitrite. Similarly, (14Z)-15-nitrofucoxanthin (5), (11Z)-11-nitrofucoxanthin (6), and (14Z,9'Z)-15-nitrofucoxanthin (7) were obtained from the reaction of peroxynitrite reaction with fucoxanthin. Capsanthin and fucoxanthin inhibited the nitration of tyrosine by peroxynitrite. Furthermore, nitrocapsanthins (3 and 4) and nitrofucoxanthins (5 and 6) exhibited an inhibitory effect on Epstein-Barr virus early antigen activation in Raji cells and an antiproliferative effect on human pancreatic carcinoma. Moreover, nitrocapsanthins (3 and 4) inhibited carcinogensis of mouse skin tumors initiated by 7,12-dimethylbenz[a]anthracene (DMBN).

Effects of artichoke leaf extract on acute gastric mucosal injury in rats.

The present study was designed to clarify the effects of an ethanol extract of artichoke leaf on acute gastric mucosal injury in rats. Oral administration of artichoke leaf extract dose-dependently prevented absolute ethanol-induced (125-500 mg/kg) or restraint plus water immersion stress-induced gastric mucosal injury (1000-2000 mg/kg). The artichoke leaf extract contains 1% cynaropicrin and 0.8% chlorogenic acid as main components and 70% dextrin as a vehicle. Cynaropicrin at doses of 1/100 of artichoke leaf extract [ethanol-induced mucosal injury: 5 mg/kg, per os (p.o.); stress-induced mucosal injury: 20 mg/kg, p.o.] also prevented gastric mucosal injury in both animal models. However, dextrin and chlorogenic acid at doses contained in the leaf extract were ineffective in both models. When artichoke leaf extract was given orally to normal rats, it (500-2000 mg/kg, p.o.) dose-dependently increased gastric mucus content. In addition, it (125-500 mg/kg, p.o.) dose-dependently prevented the decrease in gastric mucus content by absolute ethanol. When the effects of artichoke leaf extract on basal gastric acid secretion in rats were evaluated, it (500-2000 mg/kg, p.o.) dose-dependently increased the volume of gastric juice in normal rats. However, it was ineffective in decreasing basal gastric acid secretion in normal rats. These results indicate that artichoke leaf extract is effective against acute gastritis and its beneficial effect is due to that of cynaropicrin. The gastric mucus-increasing action of artichoke leaf extract may be, at least in part, related to the anti-gastritic action of the extract.

Preparation of a colored conductive paint electrode for electrochemical inactivation of bacteria.

In this study we describe the preparation of a colored conductive paint electrode containing In(2)O(3), SnO(2), or TiO(2) for the electrochemical inactivation of marine bacteria. When each colored conductive paint electrode was immersed in seawater containing 10(6) cells/mL for 90 min, marine microbe attachment to the TiO(2)/SnO(2)/Sb electrode surface was minimal. Preparation of electrodes coated with 40% particles is shown to be more cost-effective, and because of their more translucent coatings they can be painted over with bright colors. When a potential of 1.0 V was applied for 30 min to the colored conductive paint electrode (40 wt% TiO(2)/SnO(2)/Sb) in sterile seawater, the survival ratio decreased to 55%. When 1.5 V vs. saturated calomel electrode (SCE) was applied, all attached cells were inactivated. Chlorine was not detected below an applied potential of 1.5 V. A change in pH was not observed in the range of 0 to 1.5 V. This method might be effective for preventing bacterial cell accumulation and the formation of biofilms.