OBJECTIVES: To investigate the role of apelin in regulating proliferation, migration and angiogenesis of bladder cancer cells and the possible regulatory mechanism. METHODS: GEO database was used to screen the differentially expressed genes in bladder cancer tissues and cells. Bladder cancer and paired adjacent tissues were collected from 60 patients for analysis of apelin expressions in relation to clinicopathological parameters. In cultured bladder cancer J82 cells and human umbilical vein endothelial cells (HUVECs), the effects of transfection with an apelin-overexpressing plasmid or specific siRNAs targeting apelin, fibroblast growth factor 2 (FGF2) and fibroblast growth factor receptor 1 (FGFR1) on proliferation and migration of J82 cells and tube formation in HUVECs were examined using plate cloning assay, Transwell assay, and angiogenesis assay; the changes in FGF2 expression and FGFR1 phosphorylation were detected using Western blotting. RESULTS: The expression level of apelin was significantly higher in bladder cancer tissues than adjacent tissues, and bladder cancer cell lines (T24 and J82) also expressed higher mRNA and protein levels of apelin than SV-HUC-1 cells. Apelin expression level in bladder cancer tissues was correlated with tumor invasion, distant metastasis and advanced TNM stages. Apelin knockdown significantly suppressed proliferation and migration of J82 cells and decreased the total angiogenic length of HUVECs. In contrast, apelin overexpression significantly promoted proliferation and migration and enhanced FGFR1 phosphorylation in J82 cells, and increased the total angiogenesis length in HUVECs, but this effects were effectively mitigated by transfection of the cells with FGF2 siRNA or FGFR1 siRNA. CONCLUSIONS High expression of apelin promotes J82 cell proliferation and migration and HUVEC angiogenesis by promoting activation of the FGF2/FGFR1 pathway.
Humans ; Urinary Bladder Neoplasms/blood supply* ; Receptor, Fibroblast Growth Factor, Type 1/metabolism* ; Cell Proliferation ; Cell Movement ; Fibroblast Growth Factor 2/metabolism* ; Neovascularization, Pathologic ; Human Umbilical Vein Endothelial Cells ; Cell Line, Tumor ; Signal Transduction ; Apelin ; Intercellular Signaling Peptides and Proteins/genetics* ; Female ; Male ; Angiogenesis

Platelet-enriched plasma (PRP) contains high concentration of platelets and abundant growth factors, which is made by centrifuging of blood and separating of blood elements. PRP promotes tendon repair by releasing various cytokines to enhance cell proliferation, tenogenic differentiation, formation and secretion of matrix; meantime, it can reduce pain by inhibiting the expression of pain-associated molecules. A number of clinical studies demonstrated that PRP was effective in treatment of tendinopathy, including patellar tendinopathy, lateral epicondylitis and plantar fasciopathy. However, some studies did not support this conclusion, because of disparity of PRP types, therapeutic courses and injections protocols in clinical application. Based on its safety, PRP can be a choice of treatment for tendinopathy, in case other non-surgical therapies are of no effect.
Blood Platelets ; cytology ; Cytokines ; metabolism ; Humans ; Intercellular Signaling Peptides and Proteins ; metabolism ; Platelet-Rich Plasma ; Tendinopathy ; therapy

PURPOSE: Dickkopf-1 (DKK-1) is a Wnt/beta-catenin signaling pathway inhibitor. We investigated whether DKK-1 is related to progression in hepatocellular carcinoma (HCC) cells and HCC patients. MATERIALS AND METHODS: In vitro reverse-transcription polymerase chain reaction (RT-PCR), wound healing assays, invasion assays, and ELISAs of patient serum samples were employed. The diagnostic accuracy of the serum DKK-1 ELISA was assessed using receiver operating characteristic (ROC) curves and area under ROC (AUC) analyses. RESULTS: RT-PCR showed high DKK-1 expression in Hep3B and low in 293 cells. Similarly, the secreted DKK-1 concentration in the culture media was high in Hep3B and low in 293 cells. Wound healing and invasion assays using 293, Huh7, and Hep3B cells showed that DKK-1 overexpression promoted cell migration and invasion, whereas DKK-1 knock-down inhibited them. When serum DKK-1 levels were assessed in 370 participants (217 with HCC and 153 without), it was significantly higher in HCC patients than in control groups (median 1.48 ng/mL vs. 0.90 ng/mL, p<0.001). The optimum DKK-1 cutoff level was 1.01 ng/mL (AUC=0.829; sensitivity 90.7%; specificity 62.0%). Although DKK-1 had a higher AUC than alpha-fetoprotein (AFP) and des-gamma-carboxy prothrombin (DCP) (AUC=0.829 vs. 0.794 and 0.815, respectively), they were statistically similar (all p>0.05). When three biomarkers were combined (DKK-1 plus AFP plus DCP), they showed significantly higher AUC (AUC=0.952) than single marker, DKK-1 plus AFP, or DKK-1 plus DCP (all p<0.001). CONCLUSION: DKK-1 might be a key regulator in HCC progression and a potential therapeutic target in HCC. Serum DKK-1 could complement the diagnostic accuracy of AFP and DCP.
Area Under Curve ; Biomarkers/blood/metabolism ; Biomarkers, Tumor/blood ; Carcinoma, Hepatocellular/blood/*diagnosis ; Enzyme-Linked Immunosorbent Assay ; Female ; Humans ; Intercellular Signaling Peptides and Proteins/*blood/*metabolism ; Liver Neoplasms/blood/*diagnosis ; Male ; Middle Aged ; Protein Precursors/blood/metabolism ; Prothrombin/metabolism ; ROC Curve ; Reverse Transcriptase Polymerase Chain Reaction/*methods ; Sensitivity and Specificity ; alpha-Fetoproteins/analysis/metabolism

Recent studies have shown that Er-Zhi-Wan (EZW), a traditional Chinese medicine consisting of Herba Ecliptae (HE) and Fructus Ligustri Lucidi (FLL), had a definite antiosteoporotic effect on osteoporotic femur, but its effect on osteoporosis of alveolar bone remains unknown. In the present study, we investigated the effects of Er-Zhi-Wan (EZW) on the microarchitecture and the regulation of Wnt/β-catenin signaling pathway in the alveolar bone of ovariectomized rats. Thirty Sprague-Dawley rats were randomly divided into three groups: sham operation group (sham, n=10), ovariectomy (OVX) group (n=10), and OVX with EZW treatment group (EZW group, n=10). From one week after ovariectomy, EZW (100 mg/mL) or vehicle (distilled water) was fed (1 mL/100 g) once per day for 12 weeks until the sacrifice of the rats. The body weights were measured weekly. After sacrifice, the sera and mandible were collected and routinely prepared for the measurement of alveolar trabecular microarchitecture, serum levels of E2, bone-specific alkaline phosphatase (BALP) and tartrate-resistant acid phosphatase 5b (TRAP5b), as well as mandibular mRNA expression of Wnt/β-catenin signaling pathway molecules wnt3a, low-density lipoprotein receptor-related protein 5 (LRP5), β-catenin and dickkopf homolog 1 (DKK1). The results showed that EZW treatment significantly prevented the body weight gain, degradation of alveolar trabecular microarchitecture and alveolar bone loss in the OVX rats. Furthermore, we observed that EZW could increase the serum levels of E2 and BALP, and decrease levels of serum TRAP5b in EZW group compared with vehicle group. In addition, RT-PCR results revealed that EZW upregulated the expression levels of wnt3a, LRP5 and β-catenin, and reduced the expression of DKK1 in OVX rats. Taken together, our results suggested that EZW may have potential anti-osteoporotic effects on osteoporotic alveolar bone by stimulating Wnt/LRP5/β-catenin signaling pathway.
Acid Phosphatase ; blood ; Alkaline Phosphatase ; blood ; Alveolar Process ; drug effects ; metabolism ; Animals ; Body Weight ; drug effects ; Drugs, Chinese Herbal ; pharmacology ; Estradiol ; blood ; Female ; Gene Expression ; drug effects ; Intercellular Signaling Peptides and Proteins ; genetics ; Isoenzymes ; blood ; Low Density Lipoprotein Receptor-Related Protein-5 ; genetics ; Mandible ; drug effects ; metabolism ; Medicine, Chinese Traditional ; methods ; Organ Size ; drug effects ; Ovariectomy ; Rats, Sprague-Dawley ; Reverse Transcriptase Polymerase Chain Reaction ; Tartrate-Resistant Acid Phosphatase ; Time Factors ; Up-Regulation ; drug effects ; Uterus ; drug effects ; growth & development ; Wnt Signaling Pathway ; drug effects ; genetics ; Wnt3A Protein ; genetics ; beta Catenin ; genetics

OBJECTIVETo investigate the association of serum chemerin with degenerative aortic valve disease (DAVD) in patients with metabolic syndrome.

METHODSFrom July, 2012 to July, 2013, 48 patients with metabolic syndrome (mean age 56.33∓6.14 years, including 25 male and 23 female patients), 48 patients with metabolic syndrome and DAVD (mean age 60.16∓6.72 years, 24 males and 21 females), and 48 adult healthy volunteers (mean age 52.94∓8.28 years, 23 males and 25 females) were examined for triglyceride, total cholesterol, low-density lipoprotein-cholesterol, high-density lipoprotein, fasting glucose, C-reactive protein and other biochemical indexes. Serum chemerin levels were detected using ELISA for all the subjects.

RESULTSPatients with metabolic syndrome had higher levels of serum chemerin than the healthy subjects, and patients with DAVD had higher chemerin levels than those with DAVD. Multivariate logistic regression analysis showed that increased serum chemerin level is a predictor of aortic valve degeneration in patients with metabolic syndrome. Univariate linear regression analysis showed that serum chemerin levels, body mass index, systolic blood pressure, total triglyceride and C-reactive protein were associated with metabolic syndrome. Stepwise multiple linear regression analysis identified correlations of body mass index and C-reactive protein with serum chemerin level.

CONCLUSIONElevated serum chemerin level can be a predictor for DAVD in patients with metabolic syndrome.

Adult ; Aged ; Aortic Valve ; Blood Pressure ; Body Mass Index ; C-Reactive Protein ; metabolism ; Chemokines ; blood ; Cholesterol, LDL ; blood ; Female ; Heart Defects, Congenital ; complications ; Heart Valve Diseases ; complications ; Humans ; Intercellular Signaling Peptides and Proteins ; blood ; Lipoproteins, HDL ; blood ; Male ; Metabolic Syndrome ; blood ; complications ; Middle Aged ; Triglycerides ; blood

OBJECTIVETo explore the effect of puerarin combined with felodipine on the mRNA and protein expression of apelin and APJ in renal tissue of renovascular hypertensive rat.

METHODSixty-two Sprague-Dawley rats were used, of which 8 rats were randomly chosen as sham-operation group. The remaining rats were made for the rat model with renovascular hypertension. The renovascular hypertensive rats were randomly divided into 5 groups as follows: 4 groups which were treated with felodipine (0.8 mg x kg(-1) x d(-1)), puerarin (50 mg x kg(-1) x d(-1)), puerarin combined with felodipine (puerarin 25 mg x kg(-1) x d(-1) + felodipine 0.4 mg x kg(-1) x d(-1)) or captopril combined with felodipine (captopril 15 mg x kg(-1) x d(-1) x felodipine 0.4 mg x kg(-1) x d(-1)), and 1 group which was treated with distilled water. Drugs or distilled water were administered for 8 weeks. The expression of apelin and APJ mRNA and protein in ischemic and non-ischemic kidneys was assessed by RT-PCR or Western blot.

RESULTCompared with sham-operation group, the expression of apelin mRNA and protein in ischemic and non-ischemic kidneys in model group was increased significantly (P < 0.01); the expression of APJ mRNA and protein in ischemic kidneys had no significance, while that in non-ischemic kidneys was decreased (P < 0. 01). Compared with model group, the expression of apelin mRNA and protein in ischemic and non-ischemic kidneys was decreased significantly in all drug-treated groups (P < 0.01); while that of APJ mRNA and protein in non-ischemic kidneys was upregulated (P < 0.01). Compared with felodipine group, the expression of apelin mRNA and protein in ischemic and non-ischemic kidneys was decreased (P < 0.01 or P < 0.05) in the group treated with both puerarin and felodipine; and the expression of APJ mRNA and protein in ischemic kidneys did not reach significant level, however, that was upregulated in non-ischemic kidneys (P < 0.01 or P < 0.05).

CONCLUSIONPuerarin downregulates the expression of apelin mRNA and protein in ischemic and non-ischemic kidneys, and upregulates that of APJ mRNA and protein in non-ischemic kidneys. Combination of puerarin and felodipine enhances the above-mentioned effects and shows no significant difference versus the combination of felodipine and captopril. The results suggest that puerarin regulates blood pressure and protects target organ through apelin/APJ pathway and that puerarin has synergetic effects with CCB.

Animals ; Antihypertensive Agents ; pharmacology ; Apelin ; Apelin Receptors ; Blotting, Western ; Captopril ; pharmacology ; Drug Synergism ; Felodipine ; pharmacology ; Gene Expression ; drug effects ; Hypertension, Renovascular ; genetics ; metabolism ; Intercellular Signaling Peptides and Proteins ; genetics ; metabolism ; Ischemia ; Isoflavones ; pharmacology ; Kidney ; blood supply ; drug effects ; metabolism ; Male ; RNA, Messenger ; genetics ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Receptors, G-Protein-Coupled ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Vasodilator Agents ; pharmacology

OBJECTIVETo investigate the biological behavior including survival and proliferation of CD34 + CD38--Lin--cells when they are cultured at single cell level.

METHODSPurified umbilical cord blood CD34 + CD38--Lin--cells were separated at single cell level in 96-well plates using flow cytometry for four groups: control group (CD34 + CD38--Lin--cell plus stem cell medium) , Shh group (CD34 + CD38--Lin--cell plus stem cell medium and Shh), BMP-4 group (CD34 + CD38--Lin--cell plus stem cell medium and BMP-4), Jagged-1 group (CD34 + CD38--Lin--cell plus stem cell medium and Jagged-1). Methylcellulose medium was used in the colony-forming experiment which was also in four groups as previously. The number of cells and colony-forming units in each well for the four groups was evaluated at different time points (day 1, 3, 7) with fluorescence microscopy counting method.

RESULTSDivision of single cell was observed to be amplified in all of these groups from day 3. And meanwhile, after 1-week culture, the survival rates for the treated groups were all higher than the control group (Jagged-1 group > BMP-4 group > Shh group > control), while the cell number in each well was also highest in the Jagged-1 group (Jagged-1 group > BMP-4 group > control). The number of wells with a cell number of zero was significantly fewer in all treated groups (especially the Jagged-1 group) than in the control group; meanwhile, the number of wells with a cell number higher than 17 was evidently higher in all the treated groups (especially the BMP-4 group) more than controls. Colony-forming units for erythroid (BFU-E), granulocyte (CFU-G), macrophage (CFU-M), and granulocyte macrophage (CFU-GM) were observed for all of these experimental groups, and there was no significant difference between the four experimental groups.

CONCLUSIONSCD34 + CD38 - Lin - cell can achieve the survival, self-renewal and proliferation when cultured at single cell level, and the adding of Shh, BMP-4, and Jagged-1 can enhance such capabilities. However, CD34 + CD38 - Lin - cell can only maintain cell totipotency in its niche.

ADP-ribosyl Cyclase 1 ; metabolism ; Antigens, CD34 ; metabolism ; Bone Morphogenetic Protein 4 ; chemistry ; Calcium-Binding Proteins ; chemistry ; Cell Culture Techniques ; Cell Proliferation ; Cell Survival ; Cells, Cultured ; Colony-Forming Units Assay ; Culture Media ; Fetal Blood ; cytology ; Hedgehog Proteins ; chemistry ; Hematopoietic Stem Cells ; cytology ; Humans ; Intercellular Signaling Peptides and Proteins ; chemistry ; Jagged-1 Protein ; Membrane Proteins ; chemistry ; Serrate-Jagged Proteins

BACKGROUNDAs two novel adipocytokines, chemerin and apelin play a key role in the pathological process of insulin resistance (IR), glucose metabolism and obesity, researchers have found that the levels of chemerin and apelin changed significantly in type 2 diabetic patients with obesity, however, the underlying mechanism involved remains unclear. The aim of this study was to investigate whether chemerin and apelin play an important role in the pathophysiologic proceeding of diabetes.

METHODSThis study enrolled 81 newly diagnosed obese type 2 diabetes mellitus (T2DM) patients (T2DM group, n = 81). All the patients were randomly assigned to DM1 group treated with metformin (n = 41) and DM2 group treated with pioglitazone (n = 40). After hypoglycemic agents treatment, patients under better blood glucose control were chosen to be given antioxidant treatment. Another 79 subjects without T2DM were recruited as normal control group (NC group), including 40 subjects (NC1 group) with normal body mass index (BMI) and 39 obese subjects (NC2 group). Levels of chemerin, apelin, BMI, tumor necrosis factor-α (TNF-α), homeostasis model assessment of IR (HOMA-IR) and 8-isoprotaglandim F2α (8-iso-PGF2α) were examined at baseline and post-treatment. The relationship between chemerin, apelin and BMI, TNF-α, HOMA-IR, 8-iso-PGF2α was analyzed.

RESULTSThe baseline levels of chemerin, apelin, TNF-α, HOMA-IR and 8-iso-PGF2α in T2DM group were significantly higher than normal control group (P < 0.001). All indices mentioned above were significantly decreased after treatment (P < 0.05). In T2DM patients treated with pioglitazone, indices mentioned above except for HOMA-IR, were decreased significantly compared with patients treated with metformin (P < 0.05). After antioxidant treatment using lipoic acid, levels of chemerin, apelin, TNF-α and 8-iso-PGF2α were further significantly decreased (P < 0.05). Correlation analysis showed that the levels of chemerin and apelin correlated positively with BMI, TNF-α, HOMA-IR and 8-iso-PGF2α before and after treatment with hypoglycemic agents (P < 0.01). The levels of chemerin and apelin also had positive correlation with TNF-α and 8-iso-PGF2α after antioxidant treatment (P < 0.05).

CONCLUSIONSThe levels of chemerin and apelin in obese T2DM patients are closely related to IR. The increased levels may be a result of compensatory response to IR, and also may be the causative factor of IR. The levels of chemerin and apelin correlate closely with oxidative stress and inflammation. The two adipokines may be inflammatory factors playing important roles in the initiation and development of obese T2DM. Chemerin and apelin are related to the pathophysiology of IR, oxidative stress and inflammation.

Apelin ; Blood Glucose ; metabolism ; Body Mass Index ; Chemokines ; metabolism ; Diabetes Mellitus, Type 2 ; drug therapy ; immunology ; metabolism ; Dinoprost ; analogs & derivatives ; metabolism ; Humans ; Hypoglycemic Agents ; therapeutic use ; Inflammation ; metabolism ; Intercellular Signaling Peptides and Proteins ; metabolism ; Metformin ; therapeutic use ; Thiazolidinediones ; therapeutic use ; Tumor Necrosis Factor-alpha ; metabolism

BACKGROUND: The objective of this study was to explore whether individual variations in the concentration of growth factors (GFs) influence the biologic effects of platelet-rich plasma (PRP) on human mesenchymal stem cells (HMSCs). METHODS: The concentrations of 7 representative GFs in activated PRP (aPRP) were measured using ELISA. The effects of PRP on the proliferation and alkaline phosphatase (ALP) activity of HMSCs were examined using several concentrations of aPRP from 3 donors; the relationships between the GF levels and these biologic effects were then evaluated using 10% aPRP from 5 subgroups derived from 39 total donors. HMSCs were cultured in DMEM with the addition of aPRP for 4 or 12 days; then, DNA content and ALP activity were measured. RESULTS: The quantity of DNA increased significantly at a 10% concentration of aPRP, but the ALP activity was suppressed at this concentration of aPRP. The GF concentrations varied among donors, and 5 subgroups of characteristic GF release patterns were identified via cluster analysis. DNA levels differed significantly between groups and tended to be higher in groups with higher concentrations of transforming growth factor-beta1 (TGF-beta1) and platelet-derived growth factors (PDGFs). DNA quantity was positively correlated with TGF-beta1 concentration, and was negatively correlated with donor age. ALP activity was negatively correlated with PDGF-BB concentration. CONCLUSIONS: The varying GF concentrations may result in different biologic effects; thus, individual differences in GF levels should be considered for reliable interpretation of the biologic functions and standardized application of PRP.
Alkaline Phosphatase/metabolism ; Blood Donors ; Cell Differentiation ; Cells, Cultured ; Culture Media/chemistry ; DNA/analysis ; Humans ; Intercellular Signaling Peptides and Proteins/*pharmacology ; Mesenchymal Stem Cells/*cytology/drug effects ; Platelet-Derived Growth Factor/pharmacology ; Platelet-Rich Plasma/*metabolism ; Transforming Growth Factor beta1/pharmacology

OBJECTIVE: To determine the mechanism of protective effect of noninvasive limb ischemic preconditioning (NIPC) on myocardium of patients with rheumatic heart disease undergoing heart valve surgery under cardiopulmonary bypass (CPB). METHODS: A total of 32 patients with rheumatic heart disease undergoing heart valve surgeries under CPB were randomly divided into 2 groups: a control group(n=16)and an NIPC group(n=16).Tourniguet was used for each patient in the NIPC group around both the upper extremities in turn, inflated for 8 min and deflated for 5 min for 3 cycles. After the anesthesia, the remaining procedures were the same as in the control group. Blood samples were collected from the central vein after the induction of anesthesia (T(1)), 5 min before aortic clamp (T(2)),30 min after aortic opening (T(3)), 6 h after the operation (T(4)), and 24 h after the operation (T(5)) to measure the concentration of cardiac troponin I and creatine kinase MB in the plasma and CGRP and ET-1 in the serum. Pathologic change of the right auricle of the heart tissue during the superior vena cave intubation and extubation was detected. RESULTS: The content of cardiac troponin I and creatine kinase MB at T(4) and T(5) in the 2 groups was higher than that of other time points in the same group, and it reached the peak at T(5). Comparison of the content of cardiac troponin I and creatine kinase MB at T(4) and T(5) in the 2 groups showed significant difference, and that of the NIPC group was lower than the control group(P<0.05).CGRP and ET-1 contents reached the peak at T(2) in the NIPC group and at T(3) in the control group, but the peak of CGRP in the NIPC group was higher than that in the control group(P<0.01).The peak of ET-1 content in the NIPC group was lower than that in the control group(P<0.01). After the CPB, myocardial and mitochondrion impairment was lighter in the NIPC group than in the control group. CONCLUSION Noninvasive limb ischemic preconditioning can protect the myocardium through increasing CGRP, inhibiting ET-1, and advancing the peak of CGRP and ET-1.
Adult ; Arm ; blood supply ; Calcitonin Gene-Related Peptide ; metabolism ; Cardiopulmonary Bypass ; Female ; Granulins ; Heart Valve Diseases ; surgery ; Heart Valve Prosthesis Implantation ; methods ; Humans ; Intercellular Signaling Peptides and Proteins ; metabolism ; Ischemic Preconditioning ; methods ; Male ; Middle Aged ; Mitral Valve ; surgery ; Myocardial Reperfusion Injury ; prevention & control ; Rheumatic Heart Disease ; surgery