This study investigated the effects of SIRT1 gene knock-out on osteoarthritis in mice, and the possible roles of SREBP2 protein and the PI3K/AKT signaling pathway in the effects. Mice were randomly divided into a normal group and a SIRT1 gene knock-out group (6 mice in each group). In these groups, one side of the knee anterior cruciate ligament was traversed, and the ipsilateral medial meniscus was cut to establish an osteoarthritis model of knee joint. The countralateral synovial bursa was cut out, serving as controls. The knee joint specimens were then divided into four groups: SIRT1control group (group A, n=6); SIRT1osteoarthritis group (group B, n=6); SIRT1control group (group C, n=6); SIRT1osteoarthritis group (group D, n=6). HE staining, Masson staining, Safranin O-Fast Green staining and Van Gieson staining were used to observe the morphological changes in the articular cartilage of the knee. Immunohistochemical staining was employed to detect the expression of SIRT1, SREBP2, VEGF, AKT, HMGCR and type II collagen proteins. SA-β-gal staining was utilized to evaluate chondrocyte aging. The results showed clear knee joint cartilage destruction and degeneration in the SIRT1osteoarthritis group. The tidal line was twisted and displaced anteriorly. Type II collagen was destroyed and distributed unevenly. Compared with the SIRT1osteoarthritis group and SIRT1control group, SIRT1 protein expression was not obviously changed in the SIRT1osteoarthritis group (P>0.05), while the expression levels of the SREBP2, VEGF and HMGCR proteins were significantly increased (P<0.05) and the levels of AKT and type II collagen proteins were significantly decreased (P<0.05). SIRT1 gene knock-out may aggravate cartilage degeneration in osteoarthritis by activating the SREBP2 protein-mediated PI3K/AKT signalling pathway, suggesting that SIRT1 gene may play a protective role against osteoarthritis.
Animals ; Cartilage ; pathology ; Chondrocytes ; metabolism ; Collagen Type II ; metabolism ; Disease Models, Animal ; Humans ; Knee Joint ; metabolism ; pathology ; Mice ; Mice, Knockout ; Oncogene Protein v-akt ; genetics ; Osteoarthritis ; genetics ; pathology ; Phosphatidylinositol 3-Kinases ; genetics ; Signal Transduction ; genetics ; Sirtuin 1 ; genetics ; Sterol Regulatory Element Binding Protein 2 ; biosynthesis ; genetics ; Vascular Endothelial Growth Factor A ; biosynthesis

OBJECTIVETo study the effect of oral administration of dimethyl dimethoxy biphenyl dicarboxylate (DDB) on adjusting angiogeneic/inflammatory mediators and ameliorating the pathology of bones in rats with collagen-induced arthritis (CIA).

METHODSWistar rat model of CIA was set up using bovine collagen type II. Fifty rats were divided into five groups randomly: normal, CIA model, DDB treatment, methotrexate (MTX) treatment, and combined DDB+MTX treatment. Ankle joints of rats were imaged with digital X-ray machine to show the destruction of joints. Fore and hind paw and knee joints were removed above the ankle joint then processed for haematoxylin and eosin staining. Plasma levels of vascular endothelial growth factor (VEGF), platelet derived growth factor, interleukin-8 (IL-8), IL-4, tumor necrosis factor α (TNF-α), and cyclooxygenase-2 (COX-2) were quantified by enzyme-linked immunosorbent assay. Nitric oxide levels were detected by Griess reagent.

RESULTSCompared with the CIA model group, a remarkable reduction in various angiogenic (VEGF and IL-8) and inflammatory mediators (TNF-α, IL-4 and COX-2) after treatment with DDB either alone or combined with MTX P<0.05 or P<0.01). Histopathological and X-ray findings were confirmatory to the observed DDB anti-arthritic effect. The DDB-treated group showed amelioration in signs of arthritis which appeared essentially similar to normal.

CONCLUSIONOur data shed light on the therapeutic efficacy of DDB in experimental rheumatoid arthritis (RA) compared with a choice drug (MTX) and it may be offered as a second-line drug in the treatment of RA.

Animals ; Arthritis, Experimental ; chemically induced ; diagnostic imaging ; drug therapy ; pathology ; Arthritis, Rheumatoid ; diagnostic imaging ; drug therapy ; pathology ; Collagen ; Cyclooxygenase 2 ; blood ; Dioxoles ; therapeutic use ; Enzyme-Linked Immunosorbent Assay ; Female ; Interleukin-4 ; blood ; Interleukin-8 ; blood ; Methotrexate ; therapeutic use ; Nitric Oxide ; biosynthesis ; Platelet-Derived Growth Factor ; analysis ; Radiography ; Rats ; Rats, Wistar ; Tumor Necrosis Factor-alpha ; blood ; Vascular Endothelial Growth Factor A ; blood

The purpose of this study was to devise an expanded ischemic flap model and to investigate the role of AMD-3100 (Plerixafor, chemokine receptor 4 inhibitor) in this model by confirming its effect on mobilization of stem cells from the bone marrow. Male Sprague-Dawley rats were used as an animal research model. The mobilization of stem cells from the bone marrow was confirmed in the AMD-3100-treated group. The fractions of endothelial progenitor cells (EPC) and the vascular endothelial growth factor receptor (VEGFR) 2+ cells in the peripheral blood were increased in groups treated with AMD-3100. The expression of vascular endothelial growth factor (VEGF) was increased in response to expansion or AMD injection. The expression of stromal cell derived factor (SDF)-1 and VEGFR2 were increased only in unexpanded flap treated with AMD-3100. Treatment with AMD-3100 increased both the number and area of blood vessels. However, there were no statistically significant differences in the survival area or physiologic microcirculation in rats from the other groups. This endogenous neovascularization induced by AMD-3100 may be a result of the increase in both the area and number of vessels, as well as paracrine augmentation of the expression of VEGF and EPCs. However, the presence of a tissue expander under the flap could block the neovascularization between the flap and the recipient regardless of AMD-3100 treatment and expansion.
Animals ; Anti-HIV Agents/pharmacology ; Bone Marrow Cells/cytology ; Chemokine CXCL12/biosynthesis ; Endothelial Progenitor Cells/*cytology ; Hematopoietic Stem Cells/*cytology ; Heterocyclic Compounds/*pharmacology ; Hypoxia-Inducible Factor 1, alpha Subunit/metabolism ; Male ; Neovascularization, Physiologic ; Nitric Oxide Synthase Type III/metabolism ; Rats ; Rats, Sprague-Dawley ; Receptors, CXCR4/antagonists & inhibitors ; Surgical Flaps/*blood supply/surgery ; Tissue Expansion/*methods ; Vascular Endothelial Growth Factor A/biosynthesis ; Vascular Endothelial Growth Factor Receptor-2/biosynthesis/metabolism

In order to investigate the expression of nerve growth factor (NGF) and hypoxia inducible factor-1α (HIF-1α) and its correlation with angiogenesis in non-small cell lung cancer (NSCLC), paraffin-embedded tissue blocks from 20 patients with NSCLC were examined. Twenty corresponding para-cancerous lung tissue specimens were obtained to serve as a control. The expression of NGF, HIF-1α, and vascular endothelial growth factor (VEGF) in the NSCLC tissues was detected by using immunohistochemistry. The microvascular density (MVD) was determined by CD31 staining. The results showed that the expression levels of NGF, HIF-1α and VEGF in the NSCLC tissues were remarkably higher than those in the para-cancerous lung tissues (P<0.05). There was significant difference in the MVD between the NSCLC tissues (9.19±1.43) and para-cancerous lung tissues (2.23±1.19) (P<0.05). There were positive correlations between NGF and VEGF, between HIF-1α and VEGF, and between NGF and HIF-1α in NSCLC tissues, with the spearman correlation coefficient being 0.588, 0.519 and 0.588, respectively. In NSCLC tissues, the MVD had a positive correlation with the three factors (P<0.05). Theses results suggest that NGF and HIF-1α are synergically involved in the angiogenesis of NSCLC.
Adult ; Aged ; Carcinoma, Non-Small-Cell Lung ; blood supply ; metabolism ; Female ; Humans ; Hypoxia-Inducible Factor 1, alpha Subunit ; biosynthesis ; Immunohistochemistry ; Lung ; blood supply ; metabolism ; pathology ; Lung Neoplasms ; blood supply ; metabolism ; Male ; Middle Aged ; Neovascularization, Pathologic ; metabolism ; Nerve Growth Factor ; biosynthesis ; Vascular Endothelial Growth Factor A ; metabolism ; Young Adult

OBJECTIVETo observe the therapeutic angiogenesis effect of naotai recipe (NR) on local ischemia/reperfusion (I/R) injury of rats by observing signaling pathway of hypoxia-inducible factor-lα (HIF-1α) and vascular endothelial growth factor (VEGF).

METHODSTotally 120 Sprague-Dawley (SD) rats were randomly divided into 4 groups, namely, the normal control group (n =12), the sham-operation group (n =12), the I/R model group (n =48), and the NR group (n =48). Cerebral I/R injury models were established using thread suture method. Rats in the I/R model group and the NR group were sub-divided into 4 sub-groups according to the 1st, 3rd, 5th, and 7th I/R day (n =12). The phenomenon of neovasculization was observed by immunofluorescence staining. The protein and mRNA expression levels of HIF-la, VEGF-A, and VEGFR II receptor were detected by RT-PCR.

RESULTSThere were a large amount of labels for neovasculization in the ischemic area of the NR group. Double-immunofluorescence labeling [vWF (red) and BrdU (green)] was observed in the NR group. Compared with the model group, the HIF-1α protein expression was obviously enhanced on the 1 st day of I/R (P <0.01), and the VEGF protein expression started to enhance on the 3rd day in the NR group (P <0.01). The VEGFR protein expression level was the highest in the NR group on the 5th day of I/R (P <0.01). The protein expression of VEGF and HIF-1α started to decrease on the 7th day of I/R.

CONCLUSIONNR could strengthen angiogenesis after I/R by elevating the expression of HIF-lα and activating HIF-lα/VEGF signaling pathway.

Animals ; Brain Ischemia ; metabolism ; Cerebral Infarction ; Hypoxia-Inducible Factor 1, alpha Subunit ; genetics ; metabolism ; Hypoxia-Ischemia, Brain ; metabolism ; Ischemia ; Neovascularization, Pathologic ; Rats, Sprague-Dawley ; Reperfusion Injury ; Signal Transduction ; Vascular Endothelial Growth Factor A ; biosynthesis

OBJECTIVETo investigate the differential expression profile and bioinformatic analysis of microRNA (miRNA) in human dental pulp cells (DPC) during endothelial differentiation.

METHODSDPC were cultured in endothelial induction medium (50 µg/L vascular endothelial growth factor, 10 µg/L basic fibroblast growth factor and 2% fetal calf serum) for 7 days. Meanwhile non-induced DPC were used as control.Quantitative real-time PCR (qRT-PCR) was applied to detect vascular endothelial marker genes [CD31, von Willebrand factor (vWF) and vascular endothelial-cadherin (VE-cadherin)] and in vitro tube formation on matrigel was used to analyze the angiogenic ability of differentiated cells. And then miRNA expression profiles of DPC were examined using miRNA microarray and then the differentially expressed miRNA were validated by qRT-PCR. Furthermore, bioinformatic analysis was employed to predict the target genes of miRNA and to analyze the possible biological functions and signaling pathways that were involved in DPC after induction.

RESULTSThe relative mRNA level of CD31, vWF and VE-cadherin in the control group were (3.48 ± 0.22) ×10(-4), (3.13 ± 0.31) ×10(-4) and (39.60 ± 2.36) ×10(-4), and (19.57 ± 2.20) ×10(-4), (48.13 ± 0.54) ×10(-4) and (228.00 ± 8.89) ×10(-4) in the induced group. The expressions of CD31, vWF and VE-cadherin were increased significantly in endothelial induced DPC compared to the control group (P < 0.05). For in vitro tube formation assay, tubular structures were formed on the matrigel by differentiated DPC. A total of 47 miRNA were differentially expressed, in which 15 miRNA were up-regulated and 32 miRNAs down-regulated in differentiated DPC compared with the control. Of these, 4 miRNA were confirmed by qRT-PCR. The target genes of differential miRNA were predicted to associate with several biological functions, such as the regulation of transcription, cell motion, blood vessel morphogenesis, angiogenesis and cytoskeletal protein, and signaling pathways including the mitogen-activated protein kinase (MAPK) and the Wnt signaling pathway.

CONCLUSIONSThe differential miRNA expression identified in this study may be involved in governing DPC endothelial differentiation, thus contributing to the future research on regulatory mechanisms in dental pulp angiogenesis.

Antigens, CD ; Cadherins ; Cell Differentiation ; Collagen ; Computational Biology ; Dental Pulp ; metabolism ; Drug Combinations ; Fibroblast Growth Factor 2 ; Humans ; Laminin ; MicroRNAs ; Platelet Endothelial Cell Adhesion Molecule-1 ; biosynthesis ; Proteoglycans ; RNA, Messenger ; Real-Time Polymerase Chain Reaction ; Signal Transduction ; Vascular Endothelial Growth Factor A ; Wnt Signaling Pathway ; von Willebrand Factor

Cartilage oligomeric matrix protein-angiopoietin-1 (COMP-Ang1) is an angiogenic factor for vascular angiogenesis. The aim was to investigate the effect of an intracavernosal injection of COMP-Ang1 on cavernosal angiogenesis in a diabetic rat model. Male Otsuka Long-Evans Tokushima Fatty (OLETF) rats made up the experimental group (1 yr old) and Long-Evans Tokushima Otsuka (LETO) rats made up the control group. The experimental group was divided into vehicle only, 10 microg COMP-Ang1, and 20 microg COMP-Ang1. COMP-Ang1 was injected into the corpus cavernosum of the penis. After 4 weeks, the penile tissues of the rats were obtained for immunohistochemistry and Western blot analysis. The immunoreactivity of PECAM-1 and VEGF was increased in the COMP-Ang1 group compared with the vehicle only group. Moreover, the expression of PECAM-1 and VEGF was notably augmented in the 20 microg Comp Ang-1 group. In the immunoblotting study, the expression of PECAM-1 and VEGF protein was significantly less in the OLEFT rats than in the control LETO rats. However, this expression was restored to control level after intracavernosal injection of COMP-Ang1. These results show that an intracavernosal injection of COMP-Ang1 enhances cavernous angiogenesis by structurally reinforcing the cavernosal endothelium.
Angiopoietin-1/genetics/*metabolism ; Animals ; Antigens, CD31/metabolism ; Blood Glucose/analysis ; Blotting, Western ; Body Weight ; Cartilage Oligomeric Matrix Protein/genetics/*metabolism ; Diabetes Mellitus, Experimental/*pathology ; Immunohistochemistry ; Male ; Neovascularization, Physiologic/*drug effects ; Penis/metabolism/pathology ; Rats ; Rats, Long-Evans ; Recombinant Fusion Proteins/biosynthesis/genetics/*pharmacology ; Vascular Endothelial Growth Factor A/metabolism

OBJECTIVE: To determine whether quantitative perfusion parameters of dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) correlate with immunohistochemical markers of angiogenesis in rectal cancer. MATERIALS AND METHODS: Preoperative DCE-MRI was performed in 63 patients with rectal adenocarcinoma. Transendothelial volume transfer (Ktrans) and fractional volume of the extravascular-extracellular space (Ve) were measured by Interactive Data Language software in rectal cancer. After surgery, microvessel density (MVD) and vascular endothelial growth factor (VEGF) expression scores were determined using immunohistochemical staining of rectal cancer specimens. Perfusion parameters (Ktrans, Ve) of DCE-MRI in rectal cancer were found to be correlated with MVD and VEGF expression scores by Spearman's rank coefficient analysis. T stage and N stage (negative or positive) were correlated with perfusion parameters and MVD. RESULTS: Significant correlation was not found between any DCE-MRI perfusion parameters and MVD (rs = -0.056 and p = 0.662 for Ktrans; rs = -0.103 and p = 0.416 for Ve), or between any DCE-MRI perfusion parameters and the VEGF expression score (rs = -0.042, p = 0.741 for Ktrans ; r = 0.086, p = 0.497 for Ve) in rectal cancer. TN stage showed no significant correlation with perfusion parameters or MVD (p > 0.05 for all). CONCLUSION: DCE-MRI perfusion parameters, Ktrans and Ve, correlated poorly with MVD and VEGF expression scores in rectal cancer, suggesting that these parameters do not simply denote static histological vascular properties.
Adult ; Aged ; Aged, 80 and over ; Contrast Media/*diagnostic use ; Female ; Follow-Up Studies ; Humans ; Immunohistochemistry ; Magnetic Resonance Imaging/*methods ; Male ; Middle Aged ; Neoplasm Staging ; Neovascularization, Pathologic/diagnosis/metabolism ; Rectal Neoplasms/blood supply/*diagnosis/metabolism ; Retrospective Studies ; Tumor Markers, Biological/biosynthesis ; Vascular Endothelial Growth Factor A/*biosynthesis

BACKGROUNDPomegranate (punica granatum) belongs to the family Punicaceae, and its peel has been used as a traditional Chinese medicine because of its efficacy in restraining intestine, promoting hemostasis, and killing parasites. Pomegranate peel has been reported to possess wound-healing properties which are mainly attributed to its polyphenol extracts. The purpose of this study was to investigate the effect of pomegranate peel polyphenols (PPP) gel on cutaneous wound healing in diabetic rats.

METHODSAlloxan-induced diabetic rats were given incisional wounds on each side of the mid-back and then treated daily with PPP gel (polyphenol mass fraction = 30%) post-wounding. Rats were sacrificed on days 4, 7, 14, and 21 post-wounding to assess the rates of wound closure, histological characteristics; and to detect the contents of hydroxyproline, production of nitric oxide (NO), and activities of NO synthase (NOS), as well as the expressions of transforming growth factor-β1 (TGF-β1), vascular endothelial growth factor (VEGF), and epidermal growth factor (EGF) in wound tissue.

RESULTSWound closure was significantly shortened when PPP gel was applied to the wounds of diabetic rats. Histological examination showed the ability of PPP gel to increase fibroblast infiltration, collagen regeneration, vascularization, and epithelialization in the wound area of diabetic rats. In addition, PPP gel-treated diabetic rats showed increased contents of hydroxyproline, production of NO, and activities of NOS and increased expressions of TGF-β1, VEGF, and EGF in wound tissues.

CONCLUSIONPPP gel may be a beneficial method for treating wound disorders associated with diabetes.

Alloxan ; Animals ; Diabetes Mellitus, Experimental ; pathology ; physiopathology ; Female ; Gels ; Hydroxyproline ; analysis ; Male ; Nitric Oxide ; biosynthesis ; Polyphenols ; pharmacology ; Punicaceae ; Rats ; Rats, Wistar ; Transforming Growth Factor beta1 ; physiology ; Vascular Endothelial Growth Factor A ; physiology ; Wound Healing ; drug effects

Impaired tumor necrosis factor receptor-1 (TNFR-1) signaling has been found in some malignant tumors with poor prognosis. However, the exact role of TNFR-1 signaling in fibrosarcoma remains unclear. Here, we explored the question by comparing the growth of TNFR-1 deficient (Tnfr1 (-)) and TNFR-1 competent (Tnfr1 (+)) fibrosarcoma FB61 cells (FB61-m and FB61-R1) in mice. TNFR-1 expression on fibrosarcoma cells delayed their growth in vivo but not in vitro. Moreover, reduced FB61-R1 tumor growth was also obtained in TNFR-1 knockout mice. The mechanism relies mainly on the TNFR-1-mediated downregulation of vascular endothelial growth factor (VEGF) production by tumor cells. Importantly, treatment of FB61-m tumors with melphalan resulted in a short delay of tumor growth, followed by a quick remission. However, when FB61-R1 tumors were treated with melphalan, tumor growth was similarly delayed at first and then completely rejected. Our results reveal evidence for TNFR-1 on tumor cells as a prerequisite in chemotherapy for fibrosarcoma, and provide novel insight into the therapeutic approach against some types of tumors using TNFR-1 angonist.
Animals ; Down-Regulation ; drug effects ; Fibrosarcoma ; drug therapy ; genetics ; pathology ; Gene Expression Regulation, Neoplastic ; drug effects ; Humans ; Melphalan ; administration & dosage ; Mice ; Molecular Targeted Therapy ; Receptors, Tumor Necrosis Factor, Type I ; genetics ; Signal Transduction ; drug effects ; Vascular Endothelial Growth Factor A ; biosynthesis