BACKGROUND: Congenital scoliosis (CS) is a complex spinal malformation of unknown etiology with abnormal bone metabolism. Fibroblast growth factor 23 (FGF23), secreted by osteoblasts and osteocytes, can inhibit bone formation and mineralization. This research aims to investigate the relationship between CS and FGF23. METHODS: We collected peripheral blood from two pairs of identical twins for methylation sequencing of the target region. FGF23 mRNA levels in the peripheral blood of CS patients and age-matched controls were measured. Receiver operator characteristic (ROC) curve analyses were conducted to evaluate the specificity and sensitivity of FGF23. The expression levels of FGF23 and its downstream factors fibroblast growth factor receptor 3 (FGFr3)/tissue non-specific alkaline phosphatase (TNAP)/osteopontin (OPN) in primary osteoblasts from CS patients (CS-Ob) and controls (CT-Ob) were detected. In addition, the osteogenic abilities of FGF23-knockdown or FGF23-overexpressing Ob were examined. RESULTS: DNA methylation of the FGF23 gene in CS patients was decreased compared to that of their identical twins, accompanied by increased mRNA levels. CS patients had increased peripheral blood FGF23 mRNA levels and decreased computed tomography (CT) values compared with controls. The FGF23 mRNA levels were negatively correlated with the CT value of the spine, and ROCs of FGF23 mRNA levels showed high sensitivity and specificity for CS. Additionally, significantly increased levels of FGF23, FGFr3, OPN, impaired osteogenic mineralization and lower TNAP levels were observed in CS-Ob. Moreover, FGF23 overexpression in CT-Ob increased FGFr3 and OPN levels and decreased TNAP levels, while FGF23 knockdown induced downregulation of FGFr3 and OPN but upregulation of TNAP in CS-Ob. Mineralization of CS-Ob was rescued after FGF23 knockdown. CONCLUSIONS Our results suggested increased peripheral blood FGF23 levels, decreased bone mineral density in CS patients, and a good predictive ability of CS by peripheral blood FGF23 levels. FGF23 may contribute to osteopenia in CS patients through FGFr3/TNAP / OPN pathway.
Humans ; Osteopontin/genetics* ; Alkaline Phosphatase/metabolism* ; Receptor, Fibroblast Growth Factor, Type 3/metabolism* ; Scoliosis/genetics* ; Osteoblasts/metabolism* ; Calcinosis ; RNA, Messenger/metabolism* ; Bone Diseases, Metabolic/metabolism* ; Fibroblast Growth Factors/genetics*

Amelogenin (AMG) is a cell adhesion molecule that has an important role in the mineralization of enamel and regulates events during dental development and root formation. The purpose of the present study was to investigate the effects of recombinant human AMG (rhAMG) on mineralized tissue-associated genes in cementoblasts. Immortalized mouse cementoblasts (OCCM-30) were treated with different concentrations (0.1, 1, 10, 100, 1000, 10,000, 100,000 ng · mL) of recombinant human AMG (rhAMG) and analyzed for proliferation, mineralization and mRNA expression of bone sialoprotein (BSP), osteocalcin (OCN), collagen type I (COL I), osteopontin (OPN), runt-related transcription factor 2 (Runx2), cementum attachment protein (CAP), and alkaline phosphatase (ALP) genes using quantitative RT-PCR. The dose response of rhAMG was evaluated using a real-time cell analyzer. Total RNA was isolated on day 3, and cell mineralization was assessed using von Kossa staining on day 8. COL I, OPN and lysosomal-associated membrane protein-1 (LAMP-1), which is a cell surface binding site for amelogenin, were evaluated using immunocytochemistry. F-actin bundles were imaged using confocal microscopy. rhAMG at a concentration of 100,000 ng · mL increased cell proliferation after 72 h compared to the other concentrations and the untreated control group. rhAMG (100,000 ng · mL) upregulated BSP and OCN mRNA expression levels eightfold and fivefold, respectively. rhAMG at a concentration of 100,000 ng · mL remarkably enhanced LAMP-1 staining in cementoblasts. Increased numbers of mineralized nodules were observed at concentrations of 10,000 and 100,000 ng · mL rhAMG. The present data suggest that rhAMG is a potent regulator of gene expression in cementoblasts and support the potential application of rhAMG in therapies aimed at fast regeneration of damaged periodontal tissue.
Alkaline Phosphatase ; metabolism ; Amelogenin ; physiology ; Animals ; Biomarkers ; metabolism ; Calcification, Physiologic ; Cell Adhesion Molecules ; metabolism ; Cell Proliferation ; Cementogenesis ; physiology ; Collagen Type I ; metabolism ; Core Binding Factor Alpha 1 Subunit ; metabolism ; Gene Expression Regulation ; In Vitro Techniques ; Integrin-Binding Sialoprotein ; metabolism ; Mice ; Microscopy, Confocal ; Osteocalcin ; metabolism ; Osteopontin ; metabolism ; Real-Time Polymerase Chain Reaction

OBJECTIVETo explore the effect of total flavonoids of Herba Epimedium (FHE) on BMP-2/RunX2/OSX signaling pathway in promoting osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs).

METHODSPassage 3 BMSCs were randomly divided into the control group, the experimental group, and the inhibitor group. BMSCs in the control group were cultured in 0.2% dimethyl sulfoxide + Osteogenuxic Supplement (OS) fluid + DMEM/F12 culture media. BMSCs in the experimental group were intervened by 20 microg/mL FHE. BMSCs in the inhibitor group were intervened by 20 microg/mL FHE and 1 microg/mL NOGGIN recombinant protein. At day 9 alkaline phosphatase (ALP) activity was measured. Calcium nodules were stained by alizarin red staining and the density was observed. The transcription expression of osteogenic differentiation-related proteins (type I collagen, osteocalcin, and osteopontin) and related factors of BMP-2/RunX2/OSX signaling pathway was assayed by RT-PCR.

RESULTSCompared with the control group, ALP activities were enhanced and the density of calcium nodules significantly increased; type I collagen, osteocalcin, and osteopontin expression levels were increased in the experimental group. The expression of osteogenesis-related transcription factor was also increased in the experimental group. Noggin recombinant protein inhibited FHE promoting BMSCs osteogenesis in the inhibitor group. Compared with the experimental group, ALP activity decreased (P < 0.05), the density of calcium nodules was lowered, expression levels of type I collagen, osteocalcin, osteopontin significantly decreased (P < 0.05) in the inhibitor group.

CONCLUSION20 microg/mL FHE promoted osteogenic differentiation process of BMSCs by BMP-2/RunX2/OSX signaling pathway.

Bone Morphogenetic Protein 2 ; metabolism ; Cell Differentiation ; drug effects ; Cells, Cultured ; Collagen Type I ; metabolism ; Core Binding Factor Alpha 1 Subunit ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Epimedium ; chemistry ; Flavonoids ; pharmacology ; Humans ; Mesenchymal Stromal Cells ; cytology ; drug effects ; Osteocalcin ; metabolism ; Osteogenesis ; drug effects ; Osteopontin ; metabolism ; Signal Transduction ; Sp7 Transcription Factor ; Transcription Factors ; metabolism

OBJECTIVETo compare the osteogenic differentiation potential and osteoclast capacity between stem cells from human exfoliated deciduous teeth (SHED) in the physiological root resorption period and dental pulp stem cells (DPSCs).

METHODSSHED and DPSCs were isolated, purified and cultured in vitro. The two stem cells were examined with ALP staining at 14 days and with alizarin red staining at 21 days of osteogenic induction, and the expressions of the genes associated with osteogenesis and osteoclastogenesis were detected using real-time PCR.

RESULTSThe isolated SHED and DPSCs both showed an elongate spindle-shaped morphology. After osteogenic induction of the cells, Alizarin red staining visualized a greater number of mineralized nodules in SHED than in DPSCs (P<0.05), and SHED also exhibited a stronger ALP activity than DPSCs (P<0.05). RT-PCR test results showed that the two stem cells expressed RANKL,OCN, ALP, OPG and Runx2 mRNA after osteogenic induction, but the expression levels of Runx2, OCN and ALP were lower in DPSCs than in SHED (P<0.05), and the ratio of RANKL/OPG was significantly higher in SHED (P<0.05).

CONCLUSIONSCompared with DPSCs, SHED has not only the ability of osteogenic differentiation but also an osteoclast capacity, which sheds light on the regulatory role of SHED in physiological root resorption bone remodeling.

Alkaline Phosphatase ; metabolism ; Cell Differentiation ; Cell Proliferation ; Cells, Cultured ; Core Binding Factor Alpha 1 Subunit ; metabolism ; Dental Pulp ; cytology ; Humans ; Osteoclasts ; cytology ; Osteogenesis ; Osteopontin ; metabolism ; RANK Ligand ; metabolism ; Real-Time Polymerase Chain Reaction ; Stem Cells ; cytology ; Tooth, Deciduous ; cytology

OBJECTIVETo investigate the effects and mechanisms of high glucose on the phenotype transformation of rat vascular smooth muscle cells (VSMCs).

METHODSVSMCs ere isolated from rat thoracic aorta and the 3rd-5th VSMCs were incubated with normal glucose (5.5 mmol/L), high glucose (25 mmol/L), or high glucose (25 mmol/L) + P38 inhibitor (25 mmol/L +SB203580) for another 24 hours. Then the gene expression of osteopontin (OPN), alpha smooth-actin (alpha-SMA), matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9(MMP-9) were assayed by real time RT-PCR, the protein expression of P38 were assayed by Western blot.

RESULTS(1) High glucose promoted the phenotype transformation of VSMCs and up-regulated the expression of MMP-2 and MMP-9. (2) High glucose promoted the phosphorylation of P38. (3) SB203580, the inhibitor of P38/MAPK signal pathway, inhibited the effects of high glucose on phenotype transformation and expression of MMP-2 and MMP-9.

CONCLUSIONHigh glucose may promote phenotype transformation of VSMCs via the signal pathway of P38/MAPK.

Actins ; metabolism ; Animals ; Aorta, Thoracic ; cytology ; Blotting, Western ; Cells, Cultured ; Glucose ; pharmacology ; Imidazoles ; pharmacology ; MAP Kinase Signaling System ; Matrix Metalloproteinase 2 ; Matrix Metalloproteinase 9 ; metabolism ; Muscle, Smooth, Vascular ; cytology ; Myocytes, Smooth Muscle ; cytology ; drug effects ; Osteopontin ; metabolism ; Phenotype ; Pyridines ; pharmacology ; Rats ; p38 Mitogen-Activated Protein Kinases ; metabolism

In the study, the inhibitory effect of epigallocatechin gallate (EGCG) on Calcium oxalate nephrolithiasis and its possible mechanism were investigated. The rat Calcium oxalate nephrolithiasis model was induced through the combined oral administration of ethylene glycol and ammonium chloride, which was intervened with EGCG. Rat blood samples were collected to detect blood creatinine (Cr), blood urea nitrogen (BUN) and blood calcium. Rat urine samples were collected to observe and compare 24-hour urine volume, oxalic acid (Ox) and calcium in urine. Renal samples were collected to prepare tissue slices and observe the pathological changes in Calcium oxalate nephrolithiasis. The expression of osteopontin (OPN) in renal tissues was evaluated by Real-time PCR and Western blot. According to the results, compared with normal rats, rats in the nephrolithiasis model showed significant increases in Cr, BUN, urine Calcium, urine Ox and renal OPN expression (P < 0.05), but obvious decrease in 24-hour urine volume (P < 0.05); Compared with rats with nephrolithiasis, those processed with EGCG revealed remarkable declines in Cr, BUN, urine Calcium and urine Ox (P < 0.05), with significant rise in 24-hour urine volume (P < 0.05) in a concentration-dependent manner. Additionally, compared with the control group, nephrolithiasis rats showed significant pathological changes in Calcium oxalate calculus. After ECCG treatment, the renal pathological changes and OPN expression attenuated significantly in a concentration-dependent manner. The results showed that EGCG inhibits the formation of calcium oxalate nephrolithiasis in rats and shows a notable protective effect on renal functions.
Animals ; Blood Urea Nitrogen ; Calcium ; blood ; Calcium Oxalate ; metabolism ; Catechin ; administration & dosage ; analogs & derivatives ; Creatinine ; blood ; Disease Models, Animal ; Humans ; Kidney ; drug effects ; metabolism ; Male ; Nephrolithiasis ; blood ; drug therapy ; genetics ; Osteopontin ; genetics ; metabolism ; Rats ; Rats, Wistar

OBJECTIVETo investigate the expression of osteopontin (OPN) splice variant mRNA, including the three isoforms OPN-A, OPN-B, and OPN-C, to explore its correlation with clinicopathologic features in gastric cancer, and to elucidate their role in tumor invasion and distant metastasis of gastric cancer.

METHODSThe expression of OPN-A, OPN-B and OPN-C mRNA were detected by real-time reverse transcriptase-polymerase chain reaction in 66 gastric cancer tissues. The relationship between the expression of OPN-A, OPN-B and OPN-C mRNA and clinicopathologic features of gastric cancer was analyzed.

RESULTSThe expression of OPN-C mRNA in the gastric cancer tissue was 3.21-fold higher than that in peritumoral mucosal tissue, showing a significant difference between them (P < 0.001). OPN-C mRNA expression was correlated with the depth of tumor invasion, tumor diameter, lymph node meatastasis, distant meatastasis and had no correlation with differentiation grades. The low expression of OPN-C mRNA was correlated with long survival benefit (P = 0.03). The expression of OPN-A and OPN-B mRNA had no significant relationship with clinicopathologic features of gastric cancer.

CONCLUSIONSOne of the isoform of osteopontin (OPN) OPN-C mRNA is overexpressed in gastric cancer. The overexpression of OPN-C mRNA may reflect the progression and is associated with the prognosis of gastric cancer. OPN-C mRNA may have value as a prognostic biomarker in gastric cancer. However, the expression of OPN-A and OPN-B are not correlated with the progression and metastasis of gastric cancer.

Disease Progression ; Gastric Mucosa ; metabolism ; Humans ; Lymph Nodes ; Lymphatic Metastasis ; Neoplasm Invasiveness ; Neoplasm Proteins ; genetics ; Osteopontin ; genetics ; Prognosis ; Protein Isoforms ; genetics ; RNA, Messenger ; metabolism ; Real-Time Polymerase Chain Reaction ; Stomach Neoplasms ; genetics ; mortality ; pathology

OBJECTIVE: To investigate osteopontin (OPN) expression in plasma and tissue of patients with layngeal squamous cell carcinoma and analyze its role in invasion, metastasis, and clinical significance in laryngeal quamous cell carcinoma. METHOD: Enzyme-linked immunosorbent assay (ELISA) and immunohistochemistry were used to detect expression of OPN in plasma and tissue of 60 cases of laryngeal squamous cell carcinoma, 20 cases of adjacent normal laryngeal tissue and 20 cases of plasma from healthy subjects. RESULT: The expression of plasma OPN was closely correlated with clinical stage and cervical lymphatic metastasis in laryngeal squamous cell carcinoma (P < 0.05), but no significant correlation with the tumor location, pathological grade, gender and age (P > 0.05). The expression of OPN increased in plasma during cancer development: laryngeal squamous cell carcinoma (38.089 ± 9.225) ng/ml, healthy subjects (18.563 ± 9.308) ng/ml. There was a significant difference between the groups (P < 0.05). The expression of OPN in tissue was closely correlated with clinical stage (P < 0.05), pathological grade (P < 0.05) and cervical lymphatic metastasis (P < 0.05) in laryngeal squamous cell carcinoma adjacent atypical hyperplastic epithelium and carcinoma. The expression of OPN increased in tissue during cancer development: laryngeal squamous cell carcinoma (56.67%), adjacent normal laryngeal tissue (15.00%). There was a significant difference between the groups (P < 0.05). Elevated expression of plasma OPN is positively correlated with the expression of OPN in tissue in laryngeal squamous cell carcinoma patients (r = 0. 871, P < 0.05). CONCLUSION OPN plays an important role in the infiltration, metastasis and carcinogenesis in laryngeal squamous cell carcinoma. Combination of serum OPN, tissue OPN detection can be used as diagnostic and surveillance indicators for laryngeal squamous cell carcinoma infiltration and metastasis.
Carcinoma, Squamous Cell ; metabolism ; pathology ; Case-Control Studies ; Enzyme-Linked Immunosorbent Assay ; Head and Neck Neoplasms ; metabolism ; pathology ; Humans ; Hyperplasia ; pathology ; Immunohistochemistry ; Laryngeal Neoplasms ; metabolism ; pathology ; Larynx ; pathology ; Lymphatic Metastasis ; Neck ; Osteopontin ; metabolism ; Squamous Cell Carcinoma of Head and Neck

OBJECTIVETo study the correlation between airway inflammation and osteopontin (OPN) level in the lung tissue, and to study the effect of dexamethasone (DXM) on OPN expression.

METHODSFifty mice were randomly divided into 5 groups: normal control, ovalbumin (OVA)-challenged asthma groups (OVA inhalation for 1 week or 2 weeks) and DXM-treated asthma groups (DXM treatment for 1 week or 2 weeks). The mice were sensitized and challenged with OVA to prepare mouse model of acute asthma. Alterations of airway inflammation were observed by haematoxylin-eosin staining. Serum level of OVA-sIgE was evaluated using ELISA. OPN expression in the lung tissue was located and measured by immunohistochemistry and Western blot respectively. OPN mRNA level in the lung tissue was detected by real-time PCR.

RESULTSThe asthma groups showed more pathological changes in the airway than the normal control and the DXM-treated groups. Compared with the OVA-challenged 1 week group, the pathological alterations increased in the OVA-challenged 2 weeks group. The level of OVA-sIgE in serum increased in the asthma groups compared with the control and the DXM groups (P<0.01). Serum OVA-sIgE sevel increased more significantly in the OVA-challenged 2 weeks group compared with the OVA-challenged 1 week group (P<0.01). OPN protein and mRNA levels were significantly raised in the asthma groups compared with the normal control and the DXM groups (P<0.01), and both levels increased more significantly in the OVA-challenged 2 weeks group compared with the OVA-challenged 1 week group (P<0.01).

CONCLUSIONSThe increased OPN expression in the lung tissue is associated with more severe airway inflammation in asthmatic mice, suggesting that OPN may play an important role in the pathogenesis of asthma. DXM can alleviate airway inflammation possibly by inhibiting OPN production.

Animals ; Asthma ; drug therapy ; metabolism ; pathology ; Dexamethasone ; therapeutic use ; Enzyme-Linked Immunosorbent Assay ; Female ; Immunoglobulin E ; blood ; Lung ; metabolism ; pathology ; Mice ; Mice, Inbred BALB C ; Osteopontin ; analysis ; genetics ; physiology ; Ovalbumin ; immunology

OBJECTIVETo investigate the composition, function, and regulatory mechanisms of the secreted phosphoprotein 1 (SPP1) gene in metastatic prostate cancer.

METHODSWe obtained the data about the whole genomic expression profiles on prostate cancer metastasis from the GEO database, and performed data-mining and bioinformatic analysis using BRB-Array Tools and such softwares as Protparam, MotifScan, SignalP 4.0, TMHMM, NetPhos2.0, PredictProtein, GO, KEGG, and STRING.

RESULTSTotally, 73 co-expressed differential genes in prostate cancer metastasis were identified, 21 up-regulated and 52 down-regulated (P <0.01). Bioinformatic analysis indicated that the highly expressed SPP1 gene encoded 314 amino acids and contained 2 N-glycosylation sites, 8 casein kinase II phosphorylation sites and 3 protein kinase C phosphorylation sites, playing essential roles in extracellular matrix (ECM) binding, ossification, osteoblast differentiation, cell adhesion, PI3K-Akt signaling pathway, focal adhesion, Toll-like receptor signaling pathway, and ECM-receptor interaction.

CONCLUSIONThe bioinformatic method showed a high efficiency in analyzing microarray data and revealing internal biological information. SPP1 may play an important role in prostate cancer metastasis and become a novel biomarker for the diagnosis of prostate cancer metastasis and a new target for its treatment.

Computational Biology ; Data Mining ; Down-Regulation ; Humans ; Male ; Microarray Analysis ; Osteopontin ; chemistry ; genetics ; secretion ; Phosphatidylinositol 3-Kinases ; metabolism ; Prostatic Neoplasms ; genetics ; metabolism ; pathology ; Signal Transduction ; Toll-Like Receptors ; metabolism