OBJECTIVE: To observe the effects of Taohong Siwu Decoction(, THSWD) on the mesenchymal stem cells(MSCs) migration, homing number and cytokine expression in callus during the early process of fracture healing, and to explore the mechanism of THSWD on accelerationg fracture healing by regulating the homing of MSCs in rats. METHODS: A rat model of right femoral shaft open fracture was established. Thirty-two 5-week-old male Sprague-Dawley rats, weighting 110 to 130 g, were divided into control group, low-dose group, medium-dose group and high-dose group by using random number table. Distilled water was given to the control group, and the other groups were given Taohong Siwu Decoction. The rats were gavaged twice a day for 5 consecutive days after surgery. Bone volume/tissue volume(BV/TV) and bone mineral density(BMD) were observed using micro-computed tomography (micro-CT) at 21 days after surgery. At 5 days post-fracture, peripheral blood MSCs from THSWD treated and untreated rats were cultured in vitro. Subsequently, the migration ability of MSCs was observed by cell migration assay. The number of MSCs homing to the callus at the early stage of fracture (5 d) was detected by Immunohistochemistry (IHC). Protein chip was used to detect the expression of cytokines in callus. RESULTS: Micro-CT results showed that BV/TV was higher in the high-dose group than in the medium-dose group (P=0.032), and higher in the medium-dose group than in the low-dose group(P=0.041), with no difference between the control and low-dose group (P=0.651). In addition, there was no difference in BMD between low-dose group and the model group (P=0.671), and lower in the low-dose group than in the medium-dose group(P=0.018), and the medium-dose group was lower than the high-dose group(P=0.008). Cell migration assay showed that THSWD promotes enhanced the migration ability of peripheral blood MSCs. IHC assay revealed that CD45^-, CD90⁺, CD29⁺ MSCs significantly increased in bone callus after THSWD intervention compared with the control group. Protein chip showed that THSWD promoted the upregulation of CINC-1(×2.91), CINC-3(×1.59), LIX(×1.5), Thymus Chemokine (×2.55), VEGF (×1.22) and the down-regulation of TIMP-1 (×2.98). CONCLUSION THSWD, a representative formula of "promoting blood circulation and removing blood stasis", can significantly accelerate fracture healing, and its mechanism may be related to enhancing the migration ability of peripheral blood MSCs and up-regulating CINC-1, CINC-3, LIX, Thymus Chemokine, VEGF and down-regulating TIMP-1 in bone callus, which promotes the peripheral blood MSCs homing in the early stage of fracture.
Animals ; Drugs, Chinese Herbal ; Fracture Healing ; Fractures, Bone/drug therapy* ; Humans ; Male ; Mesenchymal Stem Cells ; Rats ; Rats, Sprague-Dawley ; Tissue Inhibitor of Metalloproteinase-1/pharmacology* ; Vascular Endothelial Growth Factor A ; X-Ray Microtomography

The objective of this study was to assess the angiogenic potential expressed as a quotient of vascular endothelial growth factor A (VEGF-A), as an indicator of proangiogenic activity, and the circulating receptors (soluble VEGF receptor protein R1 (sVEGFR-1) and sVEGFR-2), as indicators of the effect of angiogenic inhibition, depending on the concentrations of matrix metalloproteinase 2 (MMP-2) and MMP-9 and their tissue inhibitor 1 (TIMP-1) and TIMP-2 in the plasma of patients with lower extremity artery disease (LEAD). These blood parameters in patients with intermittent claudication (IC) and critical limb ischemia (CLI) were compared for select clinical and biochemical features. Stimulation of angiogenesis in the plasma of individuals with LEAD was evident as indicated by the significant increase in VEGF-A concentration along with reduced inhibition depending on circulating receptors sVEGFR-1 and sVEGFR-2. Critical ischemia was associated with higher VEGF-A, MMP-9, TIMP-1, and TIMP-2 concentrations than in the case of IC.
Aged ; Angiogenesis Inhibitors/pharmacology* ; Female ; Gene Expression Regulation ; Humans ; Intermittent Claudication/drug therapy* ; Ischemia/drug therapy* ; Lower Extremity/blood supply* ; Male ; Matrix Metalloproteinase 9/blood* ; Middle Aged ; Neovascularization, Pathologic ; Tissue Inhibitor of Metalloproteinase-1/blood* ; Tissue Inhibitor of Metalloproteinase-2/blood* ; Vascular Endothelial Growth Factor A/blood* ; Vascular Endothelial Growth Factor Receptor-1/blood* ; Vascular Endothelial Growth Factor Receptor-2/blood*

OBJECTIVETo investigate the pathogenic mechanisms of airway inflammation and recurrent wheezing induced by recurrent respiratory virus infection after respiratory syncytial virus (RSV) infection.

METHODSSixty-four female BALB/c mice (aged 6-8 weeks) were randomly divided into four groups: control, RSV, Poly(I:C), and RSV+Poly(I:C) (n=16 each). The bronchoalveolar lavage fluid (BALF) was collected on the 3rd day after Poly(I:C) administration, and the total cell number and differential counts in BALF were determined. Hematoxylin-eosin staining was used to observe pulmonary pathological changes. The airway responsiveness was detected. ELISA was used to measure the levels of interferon-γ (IFN-γ), interleukin-4 (IL-4), interleukin-13 (IL-13), matrix metallopeptidase-9 (MMP-9), and tissue inhibitor of metalloproteinase-1 (TIMP-1) in BALF.

RESULTSCompared with the other three groups, the RSV+Poly(I:C) group had significant increases in the total number of inflammatory infiltrating cells in the airway, airway responsiveness, and MMP-9 level in BALF (P<0.05). The RSV+Poly(I:C) group showed more severe pulmonary tissue injuries compared with the control and RSV groups (P<0.01). Compared with the RSV group, the RSV+Poly(I:C) group showed significant reductions in the levels of IL-4 and TIMP-1 in BALF (P<0.01).

CONCLUSIONSViral re-infection in the late stage of RSV infection may cause an imbalance of MMP-9/TIMP-1 expression and thus contribute to aggravated airway inflammation.

Animals ; Asthma ; etiology ; Bronchoalveolar Lavage Fluid ; chemistry ; Female ; Lung ; pathology ; Matrix Metalloproteinase 9 ; analysis ; Mice ; Mice, Inbred BALB C ; Poly I-C ; pharmacology ; Respiratory Syncytial Virus Infections ; complications ; Tissue Inhibitor of Metalloproteinase-1 ; analysis

PURPOSE: The degradation of the extracellular matrix has been shown to play an important role in the treatment of hepatic cirrhosis. In this study, the effect of thalidomide on the degradation of extracellular matrix was evaluated in a rat model of hepatic cirrhosis. MATERIALS AND METHODS: Cirrhosis was induced in Wistar rats by intraperitoneal injection of carbon tetrachloride (CCl4) three times weekly for 8 weeks. Then CCl4 was discontinued and thalidomide (100 mg/kg) or its vehicle was administered daily by gavage for 6 weeks. Serum hyaluronic acid, laminin, procollagen type III, and collagen type IV were examined by using a radioimmunoassay. Matrix metalloproteinase-13 (MMP-13), tissue inhibitor of metalloproteinase-1 (TIMP-1), and alpha-smooth muscle actin (alpha-SMA) protein in the liver, transforming growth factor beta1 (TGF-beta1) protein in cytoplasm by using immunohistochemistry and Western blot analysis, and MMP-13, TIMP-1, and TGF-beta1 mRNA levels in the liver were studied using reverse transcriptase polymerase chain reaction. RESULTS: Liver histopathology was significantly better in rats given thalidomide than in the untreated model group. The levels of TIMP-1 and TGF-beta1 mRNA and protein expressions were decreased significantly and MMP-13 mRNA and protein in the liver were significantly elevated in the thalidomide-treated group. CONCLUSION: Thalidomide may exert its effects on the regulation of MMP-13 and TIMP-1 via inhibition of the TGF-beta1 signaling pathway, which enhances the degradation of extracellular matrix and accelerates the regression of hepatic cirrhosis in rats.
Actins ; Animals ; Carbon Tetrachloride/toxicity ; Collagen Type III/metabolism ; Down-Regulation ; Extracellular Matrix/metabolism ; Immunohistochemistry ; Immunosuppressive Agents/*pharmacology ; Liver Cirrhosis, Experimental/chemically induced/*metabolism/pathology/*prevention & control ; Male ; RNA, Messenger/analysis/metabolism ; Rats ; Rats, Wistar ; Thalidomide/*pharmacology ; Tissue Inhibitor of Metalloproteinase-1/biosynthesis/*drug effects ; Transcription Factor RelA/biosynthesis/drug effects ; Transforming Growth Factor beta1/biosynthesis/*drug effects ; Transforming Growth Factors/metabolism

OBJECTIVETo explore the effect of the Hsp90 inhibitor anacardic acid on cell proliferation, invasion and migration of breast cancer MDA-MB-231 cells.

METHODSThe inhibitory effect of anacardic acid on Hsp90 was assessed with in vitro ATPase inhibition assay and ATP-sepharose binding assay. MTT assay was used to detect the growth inhibition induced by anacardic acid in MDA-MB-231 cells. Transwell assays were used to evaluate MDA-MB-231 cell invasion and migration. Western blotting was performed to assess the effect of anacardic acid in triggering the degradation of MMP-9, TIMP-1, Hsp90, and Hsp70.

RESULTSAnacardic acid exhibited a modest activity of ATPase inhibition with an IC50 value of 82.5 µmol/L. Anacardic acid significantly suppressed the proliferation of MDA-MB-231 cells in a dose-dependent manner (IC50 value of 29.3 µmol/L). Treatment with 12.5, 25, and 50 µmol/L anacardic acid for 36 h caused inhibition of cell invasion by 23.6%, 56.6%, and 67.0% in MDA-MB-231 cells, respectively (P<0.05), and anacardic acid treatment for 24 h inhibited the cell migration by 30.0%, 45.5%, and 77.5%, respectively (P<0.05). Anacardic acid dose-dependently induced MMP-9 degradation, but did not obviously affect Hsp90 or Hsp70 expressions.

CONCLUSIONAnacardic acid can significantly inhibit the proliferation, invasion, and migration of MDA-MB-231 cells, the mechanism of which may involve the inhibition of Hsp90 ATPse activity and down-regulation of MMP-9 expression.

Anacardic Acids ; pharmacology ; Breast Neoplasms ; pathology ; Cell Line, Tumor ; drug effects ; Cell Movement ; Cell Proliferation ; Down-Regulation ; HSP90 Heat-Shock Proteins ; antagonists & inhibitors ; metabolism ; Humans ; Matrix Metalloproteinase 9 ; metabolism ; Tissue Inhibitor of Metalloproteinase-1 ; metabolism

Osteoarthritis is recognised to be an interactive pathological process involving the cartilage, subchondral bone and synovium. The signals from the synovium play an important role in cartilage metabolism, but little is known regarding the influence of the signalling from bone. Additionally, the collagenases and stromelysin-1 are involved in cartilage catabolism through mitogen-activated protein kinase (MAPK) signalling, but the role of the gelatinases has not been elucidated. Here, we studied the influence of osteoclastic signals on chondrocytes by characterising the expression of interleukin-1β (IL-1β)-induced gelatinases through MAPK signalling. We found that osteoclast-conditioned media attenuated the gelatinase activity in chondrocytes. However, IL-1β induced increased levels of gelatinase activity in the conditioned media group relative to the mono-cultured chondrocyte group. More specifically, IL-1β restored high levels of gelatinase activity in c-Jun N-terminal kinase inhibitor-pretreated chondrocytes in the conditioned media group and led to lower levels of gelatinase activity in extracellular signal-regulated kinase or p38 inhibitor-pretreated chondrocytes. Gene expression generally correlated with protein expression. Taken together, these results show for the first time that signals from osteoclasts can influence gelatinase activity in chondrocytes. Furthermore, these data show that IL-1β restores gelatinase activity through MAPK inhibitors; this information can help to increase the understanding of the gelatinase modulation in articular cartilage.
3T3 Cells ; Animals ; Cartilage, Articular ; cytology ; Cell Survival ; physiology ; Cells, Cultured ; Chondrocytes ; drug effects ; enzymology ; Coculture Techniques ; Culture Media, Conditioned ; Gelatinases ; drug effects ; Interleukin-1beta ; pharmacology ; JNK Mitogen-Activated Protein Kinases ; antagonists & inhibitors ; MAP Kinase Signaling System ; physiology ; Matrix Metalloproteinase 2 ; drug effects ; Matrix Metalloproteinase 9 ; drug effects ; Mice ; Mitogen-Activated Protein Kinases ; antagonists & inhibitors ; drug effects ; Monocytes ; cytology ; NF-kappa B ; antagonists & inhibitors ; Osteoclasts ; physiology ; Protease Inhibitors ; analysis ; Tissue Inhibitor of Metalloproteinase-1 ; drug effects ; Tissue Inhibitor of Metalloproteinase-2 ; drug effects ; p38 Mitogen-Activated Protein Kinases ; antagonists & inhibitors

OBJECTIVETo investigate the effects of Biejia Ruangan Tablet ([symbol in text], BRT)-containing serum on the expression of matrix metalloproteinase (MMP-9) and tissue inhibitor of metalloproteinase (TIMP-1) in cultured renal interstitial fibroblasts.

METHODSDifferent BRT-containing sera were prepared by gastric gavages to rats with the high-dose (7 g/kg), mid-dose (3.5 g/kg), and low-dose (1.75 g/kg) BRT respectively. The expression of extracellular matrix in NRK-49F cells was induced by treatment with human transforming growth factor-β1 (recombined human TGF-β1), and BRT-containing serum. Western blotting and Northern blotting were used to measure type I and III procollagen, MMP-9, and TIMP-1.

RESULTSThe high dose BRT-containing serum could decrease the type I and III procollagen gene expression which boosted by TGF-β1, at the same time cut down TIMP-1 protein and gene expression which increased by TGF-β1 (P <0.05). Treatment of cells with recombined human TGF-β1 had no significant effect on MMP-9 expression and BRT-containing serum also had no effect on MMP-9 expression.

CONCLUSIONSHigh dose BRT has anti-fibrosis effects in NRK-49F cells, as indicated by its inhibition of type I and III procollagen and TIMP-1 expression.

Animals ; Cells, Cultured ; Collagen Type I ; genetics ; metabolism ; Collagen Type III ; genetics ; metabolism ; Drugs, Chinese Herbal ; pharmacology ; Fibroblasts ; drug effects ; metabolism ; Gene Expression Regulation ; drug effects ; Humans ; Kidney ; cytology ; Male ; Matrix Metalloproteinase 9 ; genetics ; metabolism ; RNA, Messenger ; genetics ; metabolism ; Rats, Sprague-Dawley ; Serum ; metabolism ; Tablets ; Tissue Inhibitor of Metalloproteinase-1 ; genetics ; metabolism ; Transforming Growth Factor beta1 ; pharmacology

OBJECTIVE: To study the effect of quercetin, an inhibitor of matrix metalloproteinases 9 (MMP-9), on the growth and metastasis of lung cancer cells and the underlying mechanisms.
 METHODS: We evaluated the inhibitory effect and the inhibitory kinetics of quercetin on MMP-9 by ELISA and enzyme inhibition kinetics, and the inhibitory effect of quercetin on the growth of lung cancer cell (A549) by MTT. The effect of quercetin on levels of MMP-9 (mRNA and protein) and TGF-β1 (protein) in A549 were measured by RT-PCR and Western blot, respectively. The synergistic inhibition effect of quercetin plus TIMP-1 on the growth of lung cancer cell A549 was discussed.
 RESULTS: Quercetin induced the apoptosis of A549. It was a reversible competitive inhibitor of MMP-9 (half inhibition rate IC50 of 5.25 μmol/L, inhibition constant Ki was 2.18 μmol/L). With the increase in quercetin concentration, the levels of MMP-9 (mRNA and protein) and TGF-β1 (protein) were decreased, and the number of tumor cells on wear filter membrane was reduced. The combination of quercetin (at low concentrations) with TIMP-1 showed synergistic inhibitory effect on the growth of A549 cells. 
 CONCLUSION Quercetin is a competitive inhibitor of MMP-9 and could downregulate the expression of MMP-9 and TGF-β1, which plays an important role in A549 apoptosis.
Apoptosis ; Cell Line, Tumor ; drug effects ; Down-Regulation ; Humans ; Lung Neoplasms ; pathology ; Matrix Metalloproteinase 9 ; metabolism ; Matrix Metalloproteinase Inhibitors ; pharmacology ; Neoplasm Metastasis ; Quercetin ; pharmacology ; RNA, Messenger ; metabolism ; Tissue Inhibitor of Metalloproteinase-1 ; pharmacology ; Transforming Growth Factor beta1 ; metabolism

OBJECTIVETo observe effects of Fengbaisan (, FBS) on the expression of matrix metalloproteinase-9 (MMP-9) and tissue inhibitor of metalloproteinase-1 (TIMP-1) in lung tissue of rats with chronic obstructive pulmonary disease (COPD) and to investigate the preventive and therapeutic mechanisms of FBS.

METHODSThe COPD rat model was established by cigarette smoke exposure and lipopolysaccharide (LPS) intra-tracheal dripping. The histopathological changes of lung tissue was observed via hematoxylin/eosin staining. The expression of MMP-9 and TIMP-1 in lung tissue was measured by reverse transcription polymerase chain reaction (RT-PCR) and immunohistochemistry.

RESULTSThe typical histopathological changes of COPD were displayed in the model group, Ambroxol Hydrochloride group and FBS group, and the pathological lesions in the FBS group were less than those in the model group. The expression of MMP-9 and TIMP-1 in the model group increased significantly compared with those in the normal group (P<0.05). After treatment for successive 28 days, the expression of MMP-9 and TIMP-1 in the FBS group decreased remarkably as compared with the model group (P<0.05).

CONCLUSIONSFBS can regulate MMP-9/TIMP-1 imbalance to prevent airway and lung parenchyma remodeling process via reducing the expression of MMP-9 and TIMP-1 in the lung tissue of COPD rats, and this may be a possible therapeutic mechanism of FBS on COPD.

Animals ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Gene Expression Regulation, Enzymologic ; drug effects ; Immunohistochemistry ; Lung ; drug effects ; enzymology ; pathology ; Male ; Matrix Metalloproteinase 9 ; genetics ; metabolism ; Pulmonary Disease, Chronic Obstructive ; drug therapy ; enzymology ; RNA, Messenger ; genetics ; metabolism ; Rats ; Rats, Wistar ; Tissue Inhibitor of Metalloproteinase-1 ; genetics ; metabolism

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