Suppressive activity of RGX-365 on HMGB1-mediated septic responses

Abstract RGX-365 is the main fraction of black ginseng conmprising protopanaxatriol (PPT)-type rare ginsenosides (ginsenosides Rg4, Rg6, Rh4, Rh1, and Rg2). No studies on the antiseptic activity of RGX-365 have been reported. High mobility group box 1 (HMGB1) is recognized as a late mediator of sepsis, and the inhibition of HMGB1 release and recovery of vascular barrier integrity have emerged as attractive therapeutic strategies for the management of sepsis. In this study, we examined the effects of RGX-365 on HMGB1-mediated septic responses and survival rate in a mouse sepsis model. RGX-365 was administered to the mice after HMGB1 challenge. The antiseptic activity of RGX-365 was assessed based on the production of HMGB1, measurement of permeability, and septic mouse mortality using a cecal ligation and puncture (CLP)-induced sepsis mouse model and HMGB1-activated human umbilical vein endothelial cells (HUVECs). We found that RGX-365 significantly reduced HMGB1 release from LPS- activated HUVECs and CLP-induced release of HMGB1 in mice. RGX-365 also restored HMGB1-mediated vascular disruption and inhibited hyperpermeability in the mice. In addition, treatment with RGX-365 reduced sepsis-related mortality in vivo. Our results suggest that RGX- 365 reduces HMGB1 release and septic mortality in vivo, indicating that it is useful in the treatment of sepsis.

Impacts of Drug Interactions on Pharmacokinetics and the Brain Transporters: A Recent Review of Natural Compound-Drug Interactions in Brain Disorders.

Natural compounds such as herbal medicines and/or phyto-compounds from foods, have frequently been used to exert synergistic therapeutic effects with anti-brain disorder drugs, supplement the effects of nutrients, and boost the immune system. However, co-administration of natural compounds with the drugs can cause synergistic toxicity or impeditive drug interactions due to changes in pharmacokinetic properties (e.g., absorption, metabolism, and excretion) and various drug transporters, particularly brain transporters. In this review, natural compound-drug interactions (NDIs), which can occur during the treatment of brain disorders, are emphasized from the perspective of pharmacokinetics and cellular transport. In addition, the challenges emanating from NDIs and recent approaches are discussed.

Fermented Oyster Extract Promotes Insulin-Like Growth Factor-1-Mediated Osteogenesis and Growth Rate.

Fermented oyster (Crassostrea gigas) extract (FO) prevents ovariectomy-induced osteoporosis by inhibiting osteoclastogenesis and activating osteogenesis. However, the molecular mechanisms underlying FO-mediated bone formation and growth rate are unclear. In the current study, we found that FO significantly upregulated the expression of growth-promoting genes in zebrafish larvae including insulin-like growth factor 1 (zigf-1), insulin-like growth factor binding protein 3 (zigfbp-3), growth hormone-1 (zgh-1), growth hormone receptor-1 (zghr-1), growth hormone receptor alpha (zghra), glucokinase (zgck), and cholecystokinin (zccka). In addition, zebrafish larvae treated with 100 µg/mL FO increased in total body length (3.89 ± 0.13 mm) at 12 days post fertilization (dpf) compared to untreated larvae (3.69 ± 0.02 mm); this effect was comparable to that of the ß-glycerophosphate-treated zebrafish larvae (4.00 ± 0.02 mm). Furthermore, FO time- and dose-dependently increased the extracellular release of IGF-1 from preosteoblast MC3T3-E1 cells, which was accompanied by high expression of IGF-1. Pharmacological inhibition of IGF-1 receptor (IGF-1R) using picropodophyllin (PPP) significantly reduced FO-mediated vertebrae formation (from 9.19 ± 0.31 to 5.53 ± 0.35) and growth performance (from 3.91 ± 0.02 to 3.69 ± 0.01 mm) in zebrafish larvae at 9 dpf. Similarly, PPP significantly decreased FO-induced calcium deposition in MC3T3-E1 cells by inhibiting GSK-3ß phosphorylation at Ser9. Additionally, DOI hydrochloride, a potent stabilizer of GSK-3ß, reduced FO-induced nuclear translocation of RUNX2. Transient knockdown of IGF-1Rα/ß using specific silencing RNA also resulted in a significant decrease in calcium deposition and reduction in GSK-3ß phosphorylation at Ser9 in MC3T3-E1 cells. Altogether, these results indicate that FO increased phosphorylated GSK-3ß at Ser9 by activating the autocrine IGF-1-mediated IGF-1R signaling pathway, thereby promoting osteogenesis and growth performance. Therefore, FO is a potential nutritional supplement for bone formation and growth.

Inhibitory effect of oolonghomobisflavan B on osteoclastogenesis by suppressing p38 MAPK activation.

Suppression of differentiation and/or function of osteoclasts is considered an effective therapeutic strategy for osteolytic bone diseases such as periodontitis and osteoporosis. Evidence regarding the health benefits of oolong tea consumption is accumulating, and tea polyphenols have various pharmacological properties such as anti-cancer and anti-diabetes effects. In this study, we investigated the effect of oolonghomobisflavan B (OFB), a polyphenolic compound in oolong tea, on osteoclast differentiation. OFB suppressed receptor activator of nuclear factor-κB (RANKL)-induced formation of tartate-resistant acid phosphatase-positive multinuclear cells without cytotoxicity. OFB also significantly attenuated p38 phosphorylation, which is essential for RANKL-induced osteoclastogenesis, and inhibited the expressions of nuclear factor of activated T cells, cytoplasmic 1 (NFATc1) and osteoclast-specific target genes, including dendritic cell-specific transmembrane protein and cathepsin K. Our findings suggest that OFB exhibits an anti-osteoclastogenic activity by inhibiting RANKL-mediated p38 activation, which is useful for the prevention and treatment of osteolytic bone diseases.

Inhibitory functions of maslinic acid, a natural triterpene, on HMGB1-mediated septic responses.

BACKGROUND: Maslinic acid (MA), a natural triterpenoid from Olea europaea, prevents oxidative stress and pro-inflammatory cytokine generation. High mobility group box 1 (HMGB1) has been recognized as a late mediator of sepsis, and the inhibition of the release of HMGB1 and the recovery of vascular barrier integrity have emerged as attractive therapeutic strategies for the management of sepsis. METHODS: We tested the hypothesis that MA induces sirtuin 1 and heme oxygenase-1, which inhibit the release of HMGB1 in lipopolysaccharide (LPS)-stimulated cells, thus inhibiting HMGB1-induced hyperpermeability and increasing the survival of septic mice. MA was administered after LPS or HMGB1 challenge, and the antiseptic activity of MA was determined based on permeability, the activation of pro-inflammatory proteins, and the production of markers for tissue injury in HMGB1-activated human umbilical vein endothelial cells (HUVECs) and a cecal ligation and puncture (CLP)-induced sepsis mouse model. RESULTS: MA significantly reduced the release of HMGB1 in LPS-activated HUVECs and attenuated the CLP-induced release of HMGB1. Additionally, MA alleviated HMGB1-mediated vascular disruption and inhibited hyperpermeability in mice, and in vivo analysis revealed that MA reduced sepsis-related mortality and tissue injury. CONCLUSION: Taken together, the present results suggest that MA reduced HMGB1 release and septic mortality and thus may be useful in the treatment of sepsis.

Fermented Oyster Extract Promotes Osteoblast Differentiation by Activating the Wnt/ß-Catenin Signaling Pathway, Leading to Bone Formation.

The Pacific oyster, Crassostrea gigas, is well-known as a nutritious food. Recently, we revealed that fermented extract of C. gigas (FO) inhibited ovariectomy-induced osteoporosis, resulting from suppression of osteoclastogenesis. However, since the beneficial effect of FO on osteogenesis is poorly understood, it was examined in mouse preosteoblast MC3T3-E1 cells, human osteosarcoma MG-63 osteoblast-like cells, and zebrafish larvae in this study. We found that FO increased mitochondrial activity from days 1 to 7; however, total cell number of MC3T3-E1 cells gradually decreased without any change in cell viability, which suggests that FO stimulates the differentiation of MC3T3-E1 cells. FO also promoted the expression of osteoblast marker genes, including runt-related transcription factor 2 (mRUNX2), alkaline phosphatase (mALP), collagen type I α1 (mCol1α1), osteocalcin (mOCN), osterix (mOSX), bone morphogenetic protein 2 (mBMP2), and mBMP4 in MC3T3-E1 cells accompanied by a significant increase in ALP activity. FO also increased nuclear translocation of RUNX2 and OSX transcription factors, ALP activity, and calcification in vitro along with the upregulated expression of osteoblast-specific marker proteins such as RUNX2, ALP, Col1α1, OCN, OSX, and BMP4. Additionally, FO enhanced bone mineralization (calcein intensity) in zebrafish larvae at 9 days post-fertilization comparable to that in the ß-glycerophosphate (GP)-treated group. All the tested osteoblast marker genes, including zRUNX2a, zRUNX2b, zALP, zCol1a1, zOCN, zBMP2, and zBMP4, were also remarkably upregulated in the zebrafish larvae in response to FO. It also promoted tail fin regeneration in adult zebrafish as same as the GP-treated groups. Furthermore, not only FO positively regulate ß-catenin expression and Wnt/ß-catenin luciferase activity, but pretreatment with a Wnt/ß-catenin inhibitor (FH535) also significantly decreased FO-mediated bone mineralization in zebrafish larvae, which indicates that FO-induced osteogenesis depends on the Wnt/ß-catenin pathway. Altogether, the current study suggests that the supplemental intake of FO has a beneficial effect on osteogenesis.

Inhibitory effects of compounds isolated from Dioscorea batatas Decne peel on particulate matter-induced pulmonary injury in mice.

Particulate matter 2.5 (PM2.5), with an aerodynamic diameter of ≤2.5 µm, is the primary air pollutant that plays a key role associated with lung injury produced by loss of vascular barrier integrity. Dioscorea batatas Decne (Chinese yam), a perennial plant belonging to Dioscoreaceae family, is widely cultivated in tropical and subtropical regions across Asia. Both aerial parts and root of D. batatas are consumed for nutritional and medicinal purposes. The aim of this study was to (1) identify the bioactive compounds present in D. batatas peel which may be responsible for inhibition of PM2.5-induced pulmonary inflammation in mice and (2) examine in vitro mechanisms underlying the observed effects of these compounds on mouse lung microvascular endothelial cells. The measured parameters include permeability, leukocyte migration, proinflammatory protein activation, reactive oxygen species (ROS) generation, and histology. Two phenanthrene compounds, 2,7-dihydroxy-4,6-dimethoxyphenanthrene (1) and 6,7-dihydroxy-2,4-dimethoxyphenanthrene (2) were isolated from D. batatas peels. Both these phenanthrene compounds exhibited significant scavenging activity against PM2.5-induced ROS and inhibited ROS-induced activation of p38 mitogen-activated protein kinase. In addition, enhancement of Akt pathway, involved in the maintenance of endothelial integrity, was noted. These phenanthrene compounds also reduced vascular protein leakage, leukocyte infiltration, and proinflammatory cytokine release in the bronchoalveolar lavage fluid obtained from PM2.5-induced lung tissues. Evidence thus indicates that phenanthrene compounds derived from D. batatas may exhibit protective effects against PM2.5-induced inflammatory lung injury and vascular hyperpermeability in mice.

2-O-digalloyl-1,3,4,6-tetra-O-galloyl-ß-D-glucose isolated from Galla Rhois suppresses osteoclast differentiation and function by inhibiting NF-κB signaling.

Natural compounds isolated from medicinal herbs and plants have immense significance in maintaining bone health. Hydrolysable tannins have been shown to possess a variety of medicinal properties including antiviral, anticancer, and anti-osteoclastogenic activities. As a part of a study on the discovery of alternative agent against skeletal diseases, we isolated a hydrolysable tannin, 2-O-digalloyl-1,3,4,6-tetra-Ogalloyl- ß-D-glucose (DTOGG), from Galla Rhois and examined the effect on osteoclast formation and function. We found that DTOGG significantly inhibited receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast differentiation by downregulating the expression of the key regulator in osteoclastogenesis as well as osteoclast-related genes. Analysis of RANKL/RANK signaling revealed that DTOGG impaired activation of IκBα and p65 in the nuclear factor kappa-lightchain- enhancer of activated B cells (NF-κB) signaling pathway. Furthermore, DTOGG reduced bone resorbing activity of osteoclasts, compared to the vehicle-treated control. These results suggest that DTOGG could be a useful natural compound to manage osteoclast-mediated skeletal diseases. [BMB Reports 2019; 52(6): 409-414].

Aloin Reduces HMGB1-Mediated Septic Responses and Improves Survival in Septic Mice by Activation of the SIRT1 and PI3K/Nrf2/HO-1 Signaling Axis.

High mobility group box 1 (HMGB1) is recognized as a late mediator of sepsis, and the inhibition of HMGB1 release and recovery of vascular barrier integrity have emerged as attractive therapeutic strategies for the management of sepsis. We tested the hypothesis that aloin induces sirtuin 1 (SIRT1) and heme oxygenase (HO)-1, which inhibit HMGB1 release in lipopolysaccharide (LPS)-stimulated cells, thereby inhibiting HMGB1-induced hyperpermeability and increasing the survival of septic mice. Aloin was administered after LPS or HMGB1 challenge, and the antiseptic activity of aloin was determined from measurements of permeability, activation of pro-inflammatory proteins and production of markers for tissue injury in HMGB1-activated human umbilical vein endothelial cells (HUVECs) and a cecal ligation and puncture (CLP)-induced sepsis mouse model. Aloin significantly reduced HMGB1 release in LPS-activated HUVECs via SIRT1-mediated HMGB1 deacetylation and the PI3K/Nrf2/heme oxygenase (HO)-1 signaling axis. Aloin also suppressed the production of tumor necrosis factor (TNF)- α and interleukin (IL)-6, as well as the activation of nuclear factor (NF)- κ B and extracellular signal-regulated kinase 1/2 (ERK 1/2) by HMGB1. Moreover, aloin restored HMGB1-mediated vascular disruption and inhibited vascular hyperpermeability in mice. In addition, treatment with aloin reduced the CLP-induced release of HMGB1, sepsis-related mortality and tissue injury in vivo. Our results suggest that aloin reduces HMGB1 release and sepsis-related mortality by activating SIRT1 and PI3K/Nrf2/HO-1 signals, indicating that aloin has potential for the treatment of sepsis.

Aloin reduces inflammatory gene iNOS via inhibition activity and p-STAT-1 and NF-κB.

Aloin is the major anthraquinone glycoside obtained from the Aloe species and exhibits anti-inflammatory and anti-oxidative activities. Here, we aimed to determine the effects of aloin on heme oxygenase-1 (HO-1) induction and on the expressions of inducible nitric oxide synthase (iNOS) and cyclooxygenase (COX) 2 in lipopolysaccharide (LPS)-activated human umbilical vein endothelial cells (HUVECs). To the end, aloin was tested whether aloin reduces iNOS protein expression and inflammatory markers (interleukin (IL)-1ß and tumor necrosis factor (TNF)-α) in LPS-treated mice lung tissue. The results indicated that aloin affected HO-1 induction and reduced LPS-activated NF-κB-luciferase activity showed to preferential inhibition of iNOS/NO and COX-2/PGE2 that was partly related to inhibition of STAT-1 phosphorylation. In particular, aloin induced translocation of Nrf2 from cytosol into the nucleus by an increased Nrf2-ARE binding activity, and reduced IL-1ß production in LPS-activated HUVECs. The reduced expression of iNOS/NO by aloin was reversed by siHO-1RNA-transfection. In LPS-treated mice, aloin significantly reduced iNOS protein in lung tissues, and TNF-α levels in the BALF. We concluded that aloin may be beneficial for treatment of lung injury.

JH-4 reduces HMGB1-mediated septic responses and improves survival rate in septic mice.

Inhibition of high mobility group box 1 (HMGB1) and restoration of endothelial integrity are emerging as attractive therapeutic strategies for the management of severe vascular inflammatory diseases. Recently, we found that JH-4, a synthesized decursin derivative, exhibited a strong anti-Hutchinson-Gilford progeria syndrome by efficiently blocking progerin-lamin A/C binding. In this study, we examined the effects of JH-4 on HMGB1-mediated septic responses and the survival rate in a mouse sepsis model. The anti-inflammatory activities of JH-4 were monitored based on its effects on lipopolysaccharide- or cecal ligation and puncture (CLP)-mediated release of HMGB1. The antiseptic activities of JH-4 were determined by measuring permeability, leukocyte adhesion, migration, and the activation of proinflammatory proteins in HMGB1-activated human umbilical vein endothelial cells and mice. JH-4 inhibited the release of HMGB1 and downregulated HMGB1-dependent inflammatory responses in human endothelial cells. JH-4 also inhibited HMGB1-mediated hyperpermeability and leukocyte migration in mice. In addition, treatment with JH-4 reduced CLP-induced release of HMGB1, sepsis-related mortality, and pulmonary injury in vivo. Our results indicate that JH-4 is a possible therapeutic agent to treat various severe vascular inflammatory diseases via the inhibition of the HMGB1 signaling pathway.

Sargassum serratifolium Extract Attenuates Interleukin-1ß-Induced Oxidative Stress and Inflammatory Response in Chondrocytes by Suppressing the Activation of NF-κB, p38 MAPK, and PI3K/Akt.

Osteoarthritis (OA) is a degenerative joint disease that is characterized by irreversible articular cartilage destruction by inflammatory reaction. Among inflammatory stimuli, interleukin-1ß (IL-1ß) is known to play a crucial role in OA pathogenesis by stimulating several mediators that contribute to cartilage degradation. Recently, the marine brown alga Sargassum serratifolium has been reported to exhibit antioxidant and anti-inflammatory effects in microglial and human umbilical vein endothelial cell models using lipopolysaccharide and tumor necrosis factor-α, but its beneficial effects on OA have not been investigated. This study aimed to evaluate the anti-osteoarthritic effects of ethanol extract of S. serratifolium (EESS) in SW1353 human chondrocytes and, in parallel, primary rat articular chondrocytes. Our results showed that EESS effectively blocked the generation of reactive oxygen species in IL-1ß-treated SW1353 and rat primary chondrocytes, indicating that EESS has a potent antioxidant activity. EESS also attenuated IL-1ß-induced production of nitric oxide (NO) and prostaglandin E2, major inflammatory mediators in these cells, which was associated with the inhibition of inducible NO synthase and cyclooxygenase-2 expression. Moreover, EESS downregulated the level of gene expression of matrix metalloproteinase (MMP)-1, -3 and -13 in SW1353 chondrocytes treated with IL-1ß, resulting in their extracellular secretion reduction. In addition, the IL-1ß-induced activation of nuclear factor-kappa B (NF-κB) was restored by EESS. Furthermore, EESS reduced the activation of p38 mitogen-activated protein kinase (MAPK) and phosphatidylinositol-3-kinase (PI3K)/Akt signaling pathways upon IL-1ß stimulation. These results indicate that EESS has the potential to exhibit antioxidant and anti-inflammatory effects through inactivation of the NF-κB, p38 MAPK, and PI3K/Akt signaling pathways. Collectively, these findings demonstrate that EESS may have the potential for chondroprotection, and extracts of S. serratifolium could potentially be used in the prevention and treatment of OA.

Sargassum serratifolium attenuates RANKL-induced osteoclast differentiation and oxidative stress through inhibition of NF-κB and activation of the Nrf2/HO-1 signaling pathway.

Sargassum serratifolium C. Agardh is a marine brown alga that has long been used as an ingredient for food and medicine by many people living along Asian coastlines. Recently, various beneficial effects of extracts or compounds isolated from S. serratifolium have been reported, but their efficacies against bone destruction are unclear. Therefore, in this study, we investigated the inhibitory property of an ethanol extract of S. serratifolium (EESS) on osteoclast differentiation by focusing on the receptor activator of nuclear factor-κB ligand (RANKL)-stimulated osteoclastogenesis model using RAW 264.7 macrophages. Our results demonstrated that EESS reduced RANKL-induced osteoclast differentiation in RAW 264.7 cells, by inhibiting tartrate-resistant acid phosphatase (TRAP) activity and destroying the F-actin ring formation. EESS also attenuated RANKL-induced expressions of key osteoclast-specific genes, such as nuclear factor of activated T cells cytoplasmic 1 (NFATC1), TRAP, cathepsin K and matrix metalloproteinase-9. These effects were mediated by impaired nuclear translocation of nuclear factor (NF)-κB and suppression of IκB-α degradation. In addition, EESS effectively inhibited the production of reactive oxygen species (ROS) by RANKL, which was associated with enhanced expression of nuclear translocation of nuclear factor-erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1). Overall, our findings provide evidence that EESS suppresses RANKL-induced osteoclastogenesis and oxidative stress through suppression of NF-κB and activation of Nrf2/HO-1 signaling pathway, indicating that S. serratifolium has a potential application the prevention and treatment of osteoclastogenic bone disease.

Activation of the Nrf2/HO-1 Signaling Pathway Contributes to the Protective Effects of Sargassum serratifolium Extract against Oxidative Stress-Induced DNA Damage and Apoptosis in SW1353 Human Chondrocytes.

Oxidative stress in chondrocytes plays a critical role in the pathogenesis of osteoarthritis as an important cause of articular cartilage degradation. Sargassum serratifolium C. Agardh, a marine brown algae, is known to have potent antioxidant activity. Nevertheless, no study has been conducted yet on the protective efficacy against oxidative stress in chondrocytes. Therefore, the aim of the current study is to investigate the mechanism of the antioxidative effect of ethanol extract of S. serratifolium (EESS) on DNA damage and apoptosis induced by hydrogen peroxide (H2O2) in SW1353 human chondrocytes. For this purpose, SW1353 cells exposed to H2O2 in the presence or absence of EESS were applied to cell viability assay, comet assay, immunoblotting and flow cytometry analyses. Our results showed that EESS effectively attenuated H2O2-induced cytotoxicity and DNA damage associated with the inhibition of reactive oxygen species (ROS) accumulation. EESS also weakened the mitochondria membrane permeabilization by H2O2, and recovered H2O2-induced decreased expression of anti-apoptotic Bcl-2 and pro-caspase-3, and degradation of poly (ADP-ribose) polymerase. In addition, EESS increased not only expression, but also phosphorylation of nuclear factor-erythroid 2 related factor 2 (Nrf2), and promoted the expression of heme oxygenase-1 (HO-1), a critical target enzyme of Nrf2, but decreased the expression of kelch-like ECH-associated protein-1; however, the inhibition of HO-1 activity by zinc protoporphyrin abolished the antioxidant potential induced by EESS against H2O2-mediated oxidative stress. Therefore, the results of this study suggest that the antioxidant efficacy of EESS in chondrocytes is at least involved in the Nrf2/HO-1 signaling pathway-dependent mechanism.

Rhus javanica Gall Extract Inhibits the Differentiation of Bone Marrow-Derived Osteoclasts and Ovariectomy-Induced Bone Loss.

Inhibition of osteoclast differentiation and bone resorption is a therapeutic strategy for the management of postmenopausal bone loss. This study investigated the effects of Rhus javanica (R. javanica) extracts on bone marrow cultures to develop agents from natural sources that may prevent osteoclastogenesis. Extracts of R. javanica (eGr) cocoons spun by Rhus javanica (Bell.) Baker inhibited the osteoclast differentiation and bone resorption. The effects of aqueous extract (aeGr) or 100% ethanolic extract (eeGr) on ovariectomy- (OVX-) induced bone loss were investigated by various biochemical assays. Furthermore, microcomputed tomography (µCT) was performed to study bone remodeling. Oral administration of eGr (30 mg or 100 mg/kg/day for 6 weeks) augmented the inhibition of femoral bone mineral density (BMD), bone mineral content (BMC), and other factors involved in bone remodeling when compared to OVX controls. Additionally, eGr slightly decreased bone turnover markers that were increased by OVX. Therefore, it may be suggested that the protective effects of eGr could have originated from the suppression of OVX-induced increase in bone turnover. Collectively, the findings of this study indicate that eGr has potential to activate bone remodeling by inhibiting osteoclast differentiation and bone loss.

Proliferation and osteoblastic differentiation of bone marrow stem cells: comparison of vertebral body and iliac crest.

Bone marrow stem cells (BMSCs) can be obtained from the vertebral body (VB) and iliac crest (IC) for augmenting spinal arthrodesis. However, it is still not evaluated, which of the two sites would have a better BMSCs potential on Proliferation and osteoblastic differentiation is still not evaluated. Fourteen patients (10 men and 4 women) undergoing posterolateral lumbar arthrodesis and pedicle screw instrumentation were involved. The mean age was 54.7 years (range 31-75 years). Bone marrow aspirates were obtained from the vertebral body through the bilateral pedicle and were quantified relative to matched, bilateral aspirates from the iliac crest that were obtained from the same patient and at the same time. The mononuclear cell count and concentration of BMSCs were calculated and compared. Proliferation and osteoblastic differentiation of each of the BMSCs were characterized using biochemical and molecular biology techniques. Concentration (cells/mL) of BMSCs from VB and IC were 3.73 × 10(3) and 3.19 × 10(3), respectively (P > 0.05). VB and IC exhibited similar proliferation pattern at 3, 5 and 7 days, but BMSCs from the VB exhibited an increased mineralization staining with Alizarin Red S at 14 days. BMSCs from both anatomic sites expressed comparable levels of CD29, CD34, CD44, CD90 and CD105. VB and IC displayed similar levels of expression of ALP, type I collagen and osterix, but VB expressed higher level of osteocalcin and Runx-2, especially at 14 and 21 days. Our studies show that BMSCs from VB have osteogenic differentiation potential similar to IC. Based on these findings, we suggest that BMSCs from VB would be comparable candidates for osseous graft supplementation especially in spinal fusion procedures.

Association of a RUNX2 promoter polymorphism with bone mineral density in postmenopausal Korean women.

Osteoporosis is characterized by impaired osteoblastogenesis. Bone mineral density (BMD) is a major determinant of bone strength. RUNX2 is an osteoblast-specific transcription factor involved in osteoblast differentiation and ossification. To determine whether RUNX2 is associated with BMD in an ethnically distinct population, we investigated SNPs within the two RUNX2 promoters (P1 and P2) using the Illuminar GoldenGate system in 729 postmenopausal Korean women. Subjects bearing the minor homozygote genotype (CC) at the RUNX2 -1025 T > C SNP (rs7771980) located in P2 showed a significant association with reduced lumbar spine BMD (p = 0.02) and BMDs at proximal femur sites (trochanter, p = 0.05; total femur, p = 0.04) compared with subjects carrying the major homozygote genotype (TT) or the heterozygote genotype (TC), respectively. These results present an interesting genotype association complementary to the previously reported association of BMD with the RUNX2 -1025 T > C P2 SNP in Spanish and Australian cohorts. Therefore, we suggest that the RUNX2 P2 polymorphism (-1025 T > C) may be a useful genetic marker for bone metabolism and may play an important role in BMD in postmenopausal Korean women.

Zinc-deficient diet decreases fetal long bone growth through decreased bone matrix formation in mice.

This study evaluated the effects of zinc on skeletal development during fetal development in pregnant ICR mice fed a zinc-deficient (3 mg/kg) or zinc-adequate (30 mg/kg) diet. We also included a group pair-fed with the zinc-deficient group to control for decreased appetite due to zinc deficiency. Developing fetuses at embryonic day 18.5 were removed by cesarean section, and the skeletal development was evaluated by histological analysis as well as by body weight and longitudinal growth measurement. Reduced maternal food intake in the zinc-deficient and pair-fed groups resulted in a marked and significant (P < .05) decrease in fetal weight compared to that of the zinc-adequate group. However, fetal length retardation in the pair-fed group was less marked than in the zinc-deficient group, suggesting that reduced supply of zinc from maternal circulation may play a role in longitudinal growth through skeletal development. The fetal developing tibia of the zinc-deficient group showed marked shortening of diaphysis and a mild narrowing of the hypertrophic chondrocyte zone width with increased osteoclast number, but there was no influence on the mineralization of bone matrix. This may be the result of reduced activation of osteoblasts and maturation of chondrocytes with increased osteoclastic activity, suggesting that zinc deficiency during the fetal development has a greater impact on the matrix formation of bone than the mineralization of bone matrix.