The purpose of this study was to investigate the age-related NADPH oxidase (arNOX) activity in patients with age-related knee osteoarthritis (OA). Serum and cartilage arNOX activities were determined using an oxidized ferricytochrome C reduction assay. Full-thickness knee joint cartilages obtained through total knee replacement surgery were graded according to the Outerbridge (OB) classification. Radiographic severity of OA was determined on Knee X-rays according to the Kellgren-Lawrence (K/L) grading system. Cartilage beta-galactosidase, HIF-1alpha, and GLUT-1 expression levels were evaluated as markers for tissue senescence, hypoxia, and glycolysis. Higher arNOX activities occurred with higher levels of cartilage beta-galactosidase, HIF-1alpha, and GLUT-1 (P = 0.002). arNOX activity in cartilages with surface defects (OB grade II, III) was higher than in those without the defects (OB grade 0, I) (P = 0.012). Cartilage arNOX activity showed a positive correlation with serum arNOX activity (r = -0.577, P = 0.023). Serum arNOX activity was significantly higher in the OA subgroup with bilateral ROA than in the OA with no or unilateral ROA (2.449 +/- 0.81, 2.022 +/- 0.251 nM/mL, respectively, P = 0.019). The results of this study demonstrate that OA itself is not a cause to increase arNOX activities, however, arNOX hyperactivity is related to a high degree of cartilage degradation, and a high grade and extent of ROA in age-related OA.
Biomarkers/metabolism ; Cartilage Diseases/*enzymology ; Cartilage, Articular/*enzymology ; Enzyme Activation ; Female ; Humans ; Male ; Middle Aged ; NADH, NADPH Oxidoreductases ; Osteoarthritis, Knee/*diagnosis/*enzymology ; Osteoporosis/*diagnosis/*enzymology ; Reproducibility of Results ; Sensitivity and Specificity ; Statistics as Topic

OBJECTIVETo investigate the effect of Ermiao Recipe (, EMR) with medicinal guide Angelicae Pubescentis Radix (APR) on the homing of bone marrow stem cells (BMSCs) to focal zone in osteoarthritis (OA) rats.

METHODSForty-eight Sprague-Dawley rats were randomly assigned to the sham-operated, model, EMR, and EMR plus APR groups (12 rats in each group). The OA rat model was induced by anterior cruciate ligament transection and medial meniscus resection. All rats were injected with recombinant human granulocyte colonystimulating factor [rhG-CSF, 30 μg/(kg·d) for continuous 7 days], and rats in the EMR and EMR plus APR groups were treated with EMR or EMR plus APR at 1.6 or 1.9 g/(kg·d) for 3 or 6 weeks, respectively. Cartilage histopathologic changes were observed by hematoxylin and eosin staining. Chondrocytes apoptosis and cartilage matrix components were tested by transferase-mediated deoxyuridine triphosphate-biotin nick end labeling assay and special staining. Interleukin-1β (IL-1 β), tumor necrosis factor α (TNF-α), bone morphogenetic protein 2 (BMP-2), and transforming growth factor beta-1 (TGF-β1) in serum were detected by enzyme-linked immunosorbent assay or radioimmunoassay assay. Matrix metalloproteinase (MMP)-13, tissue inhibitors of metalloproteinase (TIMP)-1, bromodeoxyuridine (BrdU), cluster of differentiation 34 (CD34), and stromal cell derived factor 1 (SDF-1) were measured by immunohistochemistry assay.

RESULTSEMR and EMR plus APR significantly inhibited articular cartilage damage and synovium inflammation in OA rats at 3 or 6 weeks of treatment, the most obvious changes in these parameters were found in the EMR plus APR group. At 6 weeks, compared with EMR treatment, EMR plus APR remarkably inhibited chondrocytes apoptosis and the release of IL-1β and TNF-α, obviously decreased MMP-13 expression, and significantly increased expressions of proteoglycan, collagen, type II collagen and TIMP-1, serum levels of BMP-2 and TGF-β1 as well as expressions of BrdU, CD34 and SDF-1 in cartilage articular (P<0.01 or P<0.05).

CONCLUSIONThe medicinal guide APR improved the therapeutic effects of EMR on OA rats by promoting directional homing of BMSCs to focal zone.

Animals ; Apoptosis ; drug effects ; Bone Marrow Cells ; drug effects ; Bone Morphogenetic Protein 2 ; blood ; Bromodeoxyuridine ; metabolism ; Cartilage, Articular ; drug effects ; enzymology ; pathology ; Chemokine CXCL12 ; metabolism ; Chondrocytes ; drug effects ; pathology ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Granulocyte Colony-Stimulating Factor ; administration & dosage ; pharmacology ; Humans ; Interleukin-1beta ; blood ; Knee Joint ; drug effects ; pathology ; Male ; Matrix Metalloproteinase 13 ; metabolism ; Nitric Oxide ; metabolism ; Nitric Oxide Synthase Type II ; blood ; Osteoarthritis ; blood ; drug therapy ; pathology ; Rats, Sprague-Dawley ; Synovial Membrane ; pathology ; Tissue Inhibitor of Metalloproteinase-1 ; metabolism ; Transforming Growth Factor beta1 ; blood ; Tumor Necrosis Factor-alpha ; blood

OBJECTIVETo observe the metabolic, regulatory and anti-oxidative effects of modified Bushen Huoxue Decoction (BSHXD), a Chinese herbal medicine for kidney (Shen)-reinforcement and blood-activation, on an osteoarthritis (OA) rabbit model.

METHODSA rabbit model for knee joint OA was established by the classic Hulth's method. The OA model rabbits were randomized into 5 groups: the model control group, the positive control group treated with glucosamine sulfate, and the three BSHXD treated groups treated respectively with low, moderate, and high doses of BSHXD. In addition, a normal control group and a sham-operated group were set up. Experimental animals were sacrificed after a 7-week treatment, and pathological changes in cartilaginous tissue were estimated using the Mankin criteria. Hydroxyproline (Hyp) and malonaldehyde (MDA) contents in blood serum and urine, as well as superoxide dismutase (SOD) activity and nitric oxide (NO) content in blood serum and knee joint synovial homogenates were detected.

RESULTSMankin scoring showed insignificant statistical differences between the various treatment groups (P >0.05), but all were better than the model control group (P <0.05). Serum and urinary contents of Hyp and MDA as well as serum and synovial levels of NO were significantly lower, but the SOD activity in blood serum and synovial tissue was higher in the BSHXD treated groups than in the model group P <0.01); the effect of BSHXD was dose-dependent to some extent.

CONCLUSIONThe modified BSHXD shows an effect of improving cartilage metabolism in experimental rabbits with OA, and possesses osteo-chondric protective effects in antagonizing peroxidation injury.

Animals ; Antioxidants ; pharmacology ; therapeutic use ; Cartilage, Articular ; drug effects ; pathology ; Disease Models, Animal ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Hydroxyproline ; blood ; urine ; Male ; Malondialdehyde ; metabolism ; Nitric Oxide ; blood ; Osteoarthritis ; blood ; drug therapy ; metabolism ; pathology ; Rabbits ; Superoxide Dismutase ; blood ; Synovial Membrane ; drug effects ; enzymology ; pathology

OBJECTIVETo investigate the in vitro effect of caffeic acid phenethyl ester (CAPE), a NF-κB inhibitor, on the apoptosis of osteoarthritic (OA) chondrocytes and on the regulation of the gelatinases matrix metalloproteinase 2 (MMP-2) and matrix metalloproteinase 9 (MMP-9).

METHODSAnnexin V-FITC/propidium iodide (PI) labeling and western blotting were used to observe and determine the apoptosis in TNFα-stimulated primary cultured osteoarthritic chondrocytes. Also, gelatin zymography was applied to examine MMP-2 and MMP-9 activities in supernatants.

RESULTSIt was confirmed by both flow cytometry and western blotting that chondrocytes from OA patients have an apoptotic background. Use of CAPE in combination with 10 ng/mL of TNFα for 24 h facilitated the apoptosis. MMP-9 in the supernatant could be autoactivated (from proMMP-9 to active MMP-9), and the physiologic calcium concentration (2.5 mmol/L) could delay the autoactivation of MMP-9. The activities of MMP-2 and MMP-9 in the fresh supernatant increased significantly in response to stimulation by 10 ng/mL of TNFα for 24 h. The stimulatory effect of TNFα just on proMMP-9 was counteracted significantly by CAPE.

CONCLUSIONNF-κB could prevent chondrocytes apoptosis though its activation was attributed to the increase of proMMP-9 activity induced by TNFα (a pro-apoptotic factor). Therefore, therapeutic NF-κB inhibitor was a 'double-edged swords' to the apoptosis of chondrocytes and the secretion of MMP-9.

Aged ; Apoptosis ; drug effects ; Caffeic Acids ; pharmacology ; therapeutic use ; Calcium ; physiology ; Cells, Cultured ; Chondrocytes ; drug effects ; enzymology ; secretion ; Drug Evaluation, Preclinical ; Female ; Humans ; Matrix Metalloproteinase 2 ; metabolism ; Matrix Metalloproteinase 9 ; metabolism ; Middle Aged ; NF-kappa B ; antagonists & inhibitors ; Osteoarthritis ; drug therapy ; enzymology ; Phenylethyl Alcohol ; analogs & derivatives ; pharmacology ; therapeutic use ; Tumor Necrosis Factor-alpha ; pharmacology

Activation of caspase-1 by NALP3 inflammasomes has been shown to be important in initiating acute gouty arthritis. The objectives of this study were to measure the levels of caspase-1 in synovial fluid in gout and various arthritides, and to elucidate the clinical significance of caspase-1 levels in synovial fluid. Caspase-1, IL-1beta, IL-18, and uric acid were measured in synovial fluid from 112 patients with gout and other arthritides, such as rheumatoid arthritis, osteoarthritis, and spondyloarthropathy. Caspase-1 in synovial fluid from patients with crystal-induced arthritis, inflammatory arthritis, osteoarthritis, and spondyloarthropathy was 35.9 +/- 86.7, 49.7 +/- 107.7, 2.1 +/- 7.0, and 152.6 +/- 155.7 pg/mL, respectively. The mean level and the frequency of high levels (> or =125 pg/mL) of caspase-1 in spondyloarthropathy were significantly higher than those in the other arthritides including gout. Caspase-1 was detectible in the synovial fluid of patients with the various arthritides. Contrary to our hypothesis, the caspase-1 level in the synovial fluid of patients with gout was not higher than in that of other arthritides. High levels of caspase-1 may be helpful in differentiating spondyloarthropathy from other arthritides.
Adult ; Aged ; Arthritis, Rheumatoid/enzymology/metabolism/pathology ; Caspase 1/*analysis ; Female ; Gout/*enzymology/metabolism/pathology ; Humans ; Interleukin-18/analysis ; Interleukin-1beta/analysis ; Leukocyte Count ; Male ; Middle Aged ; Osteoarthritis/enzymology/metabolism/pathology ; Spondylarthropathies/*enzymology/metabolism/pathology ; Synovial Fluid/*enzymology/metabolism ; Uric Acid/analysis

The levels of tartrate resistant acid phosphatase (TRAP), matrix metalloproteinase-2 (MMP-2), and tissue inhibitor of matrix metalloproteinase-2 (TIMP-2) in synovial fluid (SF) and serum in cases of canine osteoarthritis (OA) were measured. OA was induced by a surgically-created medial patellar luxation in the left stifle of 24 dogs. SF and blood samples were collected at 1.5- and 3-month intervals, respectively. Every 3 months, one dog was euthanatized to collect tissue samples from both stifles. TRAP levels in SF and serum were measured using a spectrophotometer, and TRAP-positive cells in joint tissues were identified by enzyme histochemistry. MMP-2 and TIMP-2 in SF and serum were detected by Western blotting and ELISA, respectively. TRAP in SF from the stifles and serum was significantly increased (p < 0.05) after 3 months. TIMP-2 in SF and serum was significantly decreased (p < 0.05), whereas MMP-2 in SF was significantly increased (p < 0.05) during the progression of OA. Histochemistry revealed an increased number of TRAP-positive cells in tissues from OA-affected joints. Assays measuring TRAP, MMP-2, and TIMP-2 in SF and serum, and methods that detect increased numbers of TRAP-positive cells in the joint tissues can play an important role in identifying the early phases of degenerative changes in canine joint components.
Acid Phosphatase/analysis/blood ; Animals ; Arthritis, Experimental/enzymology/etiology/veterinary ; Biological Markers/*analysis/*blood ; Blotting, Western/veterinary ; Dislocations/complications/*veterinary ; Dog Diseases/*enzymology/etiology ; Dogs ; Enzyme-Linked Immunosorbent Assay/veterinary ; Female ; Isoenzymes/analysis/blood ; Male ; Matrix Metalloproteinase 2/analysis/blood ; Osteoarthritis/enzymology/etiology/*veterinary ; Spectrophotometry/veterinary ; Stifle/physiopathology ; Synovial Fluid/*enzymology ; Tissue Inhibitor of Metalloproteinase-2/analysis/blood

OBJECTIVETo study the clinical effificacy of electroacupuncture (EA) on treating knee osteoarthritis (KOA) of Shen ()-Sui () insuffificiency (SSI) syndrome type.

METHODSA total of 245 patients (279 knees) of KOA-SSI were randomly assigned to two groups by lottery: 141 knees in the treatment group and 138 knees in the control group. The treatment group was managed with EA at the dominant points of Neixiyan (Ex-LE4) and Waixiyan (Ex-LE5) as well as the conjugate points of Xuanzhong (GB39) and Taixi (KI3) for 30 min, once a day, with 15 days as one course; 2 courses were applied with a 5-day interval in between. The control group was treated with intra-articular injection of 2 mL hyaluronic acid into the affected joint every 7 days for 5 times in total. The clinical effects on the patients in different stages were observed, and their symptom scores of knee and contents of cytokines, including interleukin-1 (IL-1), interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), prostaglandin E(2alpha) (PGE(2alpha)) and matrix metalloproteinases-3 (MMP-3), in the knee joint fluid were measured before and after treatment.

RESULTSThe study was completed in 235 patients (263 knees); four patients (7 knees) in the treatment group and six patients (9 knees) in the control group dropped out. Comparison of therapeutic effects (excellent and effective rates) between the two groups showed insignificant differences (P>0.05). Symptom scores of knee and contents of cytokines in the knee flfluid after treatment were lowered signifificantly in the patients of stage I-III in both groups (P<0.05 or P<0.01). However, the lowering of the total symptom score of knee in the patients of stage III in the treatment group was more signifificant (P<0.05).

CONCLUSIONSEA could effectively alleviate the clinical symptoms in KOA patients of stage III, showing an effect superior to that of hyaluronic acid. EA also shows action in suppressing the secretion of IL-1, IL-6, TNF-alpha, PGE(2alpha) and MMP-3 in the knee flfluid.

Aged ; Cytokines ; metabolism ; Electroacupuncture ; adverse effects ; methods ; Female ; Humans ; Male ; Matrix Metalloproteinase 3 ; metabolism ; Middle Aged ; Osteoarthritis, Knee ; diagnostic imaging ; enzymology ; therapy ; Radiography ; Syndrome ; Synovial Fluid ; enzymology ; Treatment Outcome

BACKGROUNDThere are few reports of a biological role for glycosyltransferases in the infiltration of osteoarthritic synovitis. The aim of this research was to investigate the expression and cellular location of β-1,4-galactosyltransferase I (β-1,4-GalT-I) in a surgically-induced rat model of knee osteoarthritis (OA), and explore the role of β-1,4-GalT-I in the pathogenesis of OA.

METHODSMale Sprague-Dawley rats were randomly divided into three groups: OA group, sham group and normal group. The model of OA was established in the right knees of rats by anterior cruciate ligament transaction (ACLT) with partial medial meniscectomy. Fibroblast-like synoviocytes (FLSs) obtained from normal rat synovial tissue were cultured. The expression of β-1,4-GalT-I mRNA in the synovial tissue, articular cartilage and FLSs treated with tumor necrosis factor-α (TNF-α) were assayed by real-time PCR. Western-blotting and immunohistochemisty were used to observe the expression of β-1,4-GalT-I at the protein level. Double immunofluorescent staining was used to define the location of the β-1,4-GalT-I with macrophage-like synoviocytes, FLSs, neutrophils, and TNF-α in the OA synovium. The alteration of TNF-α in FLSs which were treated with lipopolysaccharide (LPS) and β-1,4-GalT-I-Ab were detected by enzyme-linked immunosorbent assay (ELISA).

RESULTSThe mRNA and protein expression of β-1,4-GalT-I increased in synovial tissue of the OA group compared with the normal and sham groups at two and four weeks after the surgery, however, no significant difference appeared in the articular cartilage. Immunohistochemistry also indicated that the β-1,4-GalT-I expression in OA synovium at four weeks after surgery increased sharply compared with the control group. β-1,4-GalT-I co-localized with macrophage-like synoviocytes, FLSs, neutrophils and TNF-α in rat OA synovitis. Moreover, in vitro β-1,4-GalT-I mRNA in FLSs was affected in a dose- and time-dependent manner in response to TNF-α stimulation. ELISA revealed that the expression of TNF-α was attenuated in FLSs in vitro when treated with anti β-1,4-GalT-I antibody.

CONCLUSIONβ-1,4-GalT-I may play an important role in the inflammation process of rat OA synovial tissue which would provide the foundation for further researching into the concrete mechanism of β-1,4-GalT-I in OA synovitis.

Animals ; Blotting, Western ; Cells, Cultured ; Enzyme-Linked Immunosorbent Assay ; Galactosyltransferases ; genetics ; metabolism ; Immunohistochemistry ; Knee Joint ; enzymology ; pathology ; surgery ; Male ; Osteoarthritis, Knee ; enzymology ; genetics ; pathology ; Polymerase Chain Reaction ; Rats ; Rats, Sprague-Dawley ; Synovial Membrane ; enzymology ; Synovitis ; enzymology ; etiology

The degradation of arthrodial cartilage is a typical characteristic in the pathogenesis of osteoarthritis. matrix metalloproteinases (MMPs) are the primary enzymes involved in extracellular matrix degradation of cartilage. The mechanism of MMPs in extracellular matrix degradation of cartilage is becoming clear with the in-depth study about MMPs, such as activation, activity regulation, related signal transduction pathways and transcription factors. This artice reviewed the activation, expression and regulation of MMPs in the related theory and empirical study of osteoarthritis cartilage.
Cartilage ; enzymology ; Enzyme Activation ; Gene Expression Regulation, Enzymologic ; Humans ; Matrix Metalloproteinases ; genetics ; metabolism ; Multigene Family ; Osteoarthritis ; enzymology ; genetics ; Signal Transduction

Adult ; Aged ; Female ; Humans ; Hyaluronic Acid ; therapeutic use ; Male ; Matrix Metalloproteinase 9 ; blood ; metabolism ; Middle Aged ; Osteoarthritis ; drug therapy ; enzymology ; Synovial Fluid ; drug effects ; enzymology