Objective To investigate the expression pattern of microRNA (miRNA) in the kidneys of unilateral ureteral obstruction (UUO) rats and to identify specific miRNA related to renal interstitial fibrosis (RIF).Methods Forty-eight male SD rats were divided into two groups:UUO group and sham-operated (Sham) group.Rats were sacrificed at 3,7 and 14 days after operation.Histologic changes were examined by Masson staining.Forty-eight selected miRNAs were examined by stem-loop real-time qPCR.Results At the 3rd day after operation,obstructed kidneys from operation rats showed mild edema in the interstitium and mononuclear cell infiltration.At the 7th day after operation,focal interstitial fibrosis was observed.At the 14th day after operation,fibrosis became more severe.The Sham kidneys showed no pathological changes.At the 3th day after operation,25 miRNAs were differentially expressed.At the 7th day after operation,24 miRNAs were aberrantly expressed,whereas 21 miRNAs were differentially expressed at the 14th day after operation (P＜0.05).Among these miRNAs,miR-132,miR-192,miR-194,miR-29c and miR-203 were consistently up-regulated or down-regulated in a time-dependent manner after operation.There were significantly correlations between the expression of five miRNAs and severity of tubulointerstitial injury (P＜0.05).Conclusions There are at least 20 miRNAs differentially expressed in the process of RIF induced by UUO.There are significantly correlations between the expression of miR-132,miR-192,miR-194,miR-29c and miR-203 and the severity of tubulointerstitial injury.They may be closely related to RIF.A further study is needed.

Objective To discuss the efficacy of application combination Of minimal immunosuppressive drugs in chronic allograft nephropathy after renal transplantation. Methods Data were drawn from the First Hospital of Tsinghua University.From September 1,2004 to July 1,2006,31 cadaver kidney transplantations were performed using triple immunosuppression with tacrolimus(n=31)and MMF plus steroids before using new strategy.The new strategy is Rapamycin+tacrolimus+MMF+Prednisone.The serum ereatinine,GFR(ml/min/1.73 m2)and 24-hours urine protein before and after 12 months of using lOW dose combination of calcineurin inhibitors,MMF,Rapamune,Predsone and Q80 were recorded.During this time,the concentration of tacrolimus,rapamune were monitored as well. Results After 12 months follow-up,the serum creatinine of 28 patients were decreased from(300±21)μmol/L to(215±38)μmol/L.GFR(ml/min/1.73m2)was elevated from 42.54±2.95 to 49.98±3.05.Three patients whose serum creatinine was 416-464μmol/L had to take hemodialysis.The 24-hours urine protein(g)of 31 patients below 0.8 g did not increase urine protein during follow-up.One patient's 24-hours urine protein(g)increased from 0.95 to 1.29.The patient and graft survival rate was 100%(31/31),90.3%(28/31)respectively.The rapamune main side effect was hyperlipidemia. Conclusions Rapamune and low dose Tacrolimus+Myeophenolate Mofetil+Corticosteroid could be a safe treatment.It may improve renal function in chronic allograft nephropathy.

OBJECTIVETo observe the effect of combined therapy of Neiyi pill (NP) and Neiyi enema (NE) on endometriosis and its effect on serum levels of endometrial antibody (EMAB) and carcinoembryonic antigen 125 (CA125).

METHODSFifty-eight cases with endometriosis were divided into 3 groups randomly, group A (n = 16) treated by NP, group B (n = 24) treated by NP and NE, and group C (n = 18) treated by danazol, all for 3 menstrual cycle with single blind method. The effect was observed and the serum levels of EMAB and CA125 were detected before and after treatment.

RESULTSComparison of the efficacy between the 3 groups showed: there was no remarkable difference between group A and B (P >0.05), both of group A and group B were superior to that of group C (P<0.05). The levels of EMAB and CA125 had no significant changes in all the three groups after treatment.

CONCLUSIONCombined therapy of NP and NE could improve the curative effect on endometriosis, and without obvious effects on serum levels of EMAB and CA125.

Administration, Rectal ; Adult ; Autoantibodies ; blood ; CA-125 Antigen ; blood ; Drugs, Chinese Herbal ; administration & dosage ; Endometriosis ; blood ; drug therapy ; Endometrium ; immunology ; Enema ; Female ; Humans ; Phytotherapy ; Single-Blind Method ; Tablets

Objective To analyze the expression profile of circular RNA (circRNA) in LPS-treated murine peritoneal macrophages (MPMs) and to investigate the effects of mmu_circ_0000790 (circ790) on the secretion of pro-inflammatory cytokines in MPMs following LPS stimulation.Methods MPMs were isolated from C57BL/6 male mice and then stimulated with (LPS treatment group) or without (blank control group) 1 mg/L of LPS for 12 hours.Supernatants of cell culture and cell pellets were collected from each group.Enzyme linked immunosorbent assay (ELISA) was used to measure the changes in the secretion of IL-1β,IL-6 and TNF-α in the supernatants.Expression profile of circRNA in macrophages from the two groups (n=3) was analyze by circRNA microarray.Some differentially expressed circRNAs were validated by real-time PCR (RT-PCR).Moreover,RT-PCR was also performed to detect the expression of circ790 in MPMs after LPS stimulation.Small interfering RNA (siRNA) oligo specific for circ790 was designed and synthesized.Then the synthesized siRNA oligo and normal control (NC) oligo were transfected into MPMs by LipofectamineTM2000.The transfected MPMs were treated with LPS.ELISA was used to detect the levels of IL-1β,IL-6 and TNF-α in the supernatants.Arraystar's home-made microRNA target prediction software was used to predict circ790/microRNA.Results The concentrations of IL-1β,IL-6 and TNF-α in the LPS treatment group were significantly higher than those in the blank control group (P<0.01).These results indicated that the inflammatory model of MPMs was successfully constructed.Statistical differences in 34 differentially expressed circRNAs were found between the two groups (P<0.05).Among them,12 circRNAs were up-regulated and the other 22 circRNAs were down-regulated in the LPS treatment group.RT-PCR results were generally consistent with the microarray data.The expression of circ790 was increased by (1.94±0.15),(3.18±0.13) and (4.21±0.22) folds as compared with that of the blank control group at 3,6 and 12 h after LPS stimulation (P<0.05).After transfecting the MPMs with circ790-siRNA,the secretion of IL-6 was significantly decreased (P<0.05) without notable influence on the secretion of IL-1β and TNF-α.circ790 could function as a microRNA sponge to regulate the gene expression.Conclusion These data show a significantly altered circRNA expression profile in the LPS-treated MPMs.circ790 may be involved in the regulation of IL-6 secreted by macrophages.

[ ABSTRACT ] AIM: To observe the effect of rapamycin on the apoptosis of mouse astrocytes in vitro.ME-THODS:The astrocytes from C57BL/6J newborn mouse pups were isolated and primarily cultured.The effect of rapamycin on the viability of astrocytes was assessed by MTT assay.The mean fluorescence intensity of SYTOX?Green stain in the astrocytes was detected by fluorescence microplate reader in order to analyze the effects of rapamycin on the cell death in-duced by H2 O2 , ionomycin and/or deferorxamin.DiOC6 (3) staining was used to analyze the mitochondrial membrane po-tential of the astrocytes induced by H2 O2 .Flow cytometry analysis was used to determine the production of ROS in the as-trocytes and mitochondria by staining with H2 DCFDA and MitoSOXTM Red reagent, respectively.RESULTS: Rapamycin at concentration of 0.5 μmol/L protected the astrocytes against cell death induced by H2 O2 or deferoxamine plus ionomy-cin.Rapamycin protected the mitochondrial membrane potential of astrocytes from the injury of H2 O2 .It also reduced the production of ROS in the astrocytes and decreased the level of ROS in the mitochondria.CONCLUSION:Rapamycin re-duces the ROS overload in the mitochondria, keeps mitochondrial membrane potential safety and protects the astrocytes a-gainst apoptosis in vitro.

This study is to establish an UPLC fingerprint of Resina Draconis from different manufacturers, which can provide a comprehensive evaluation for its quality control. The analysis was performed on a Phenomenex Kinetex 2.6 μ C18 100A column by agradientelution program with acetonitrile-water as mobile phase at a flow rate of 1.7 mL x min(-1). The column temperature was 40 degrees C and the detection wavelengthwas 280 nm. The fingerprints of 18 batches of Draconis Resina were further evaluated by chemometrics methods including similarity analysis (SA), hierarchical clustering analysis (HCA) and principal component analysis (PCA). As a result, there were 15 common peaks, 13 of which had been identified by LC-Q-TOF MS, and the similarity degrees of 15 batches of the samples was more than 0.9, and the samples were divided into 4 clusters by their quality difference. The method is reproducible, simple and reliablethat it can be used for quality control and evaluation of Resina Draconis from different manufacturers.
Chromatography, High Pressure Liquid ; methods ; Dracaena ; chemistry ; Drugs, Chinese Herbal ; analysis ; Principal Component Analysis ; Quality Control

OBJECTIVETo investigate the effect of HS3ST3B1 on hepatitis B virus (HBV) replication.

METHODSHepG2 cells were classified into 7 groups according to the plasmids transfected: (1) Blank group, no plasmid transfected; 2. Positive control, transfected with pCH9-HBV which permits HBV replication; (3) Negative control, transfected with pCH9-HBV + pcDNA3.1 + pTZU6+1; (4) Treatment A, transfected with pCH9-HBV + pCDNA3.1-HS3ST3B1 + pTZU6+1; (5) Interference A, transfected with pCH9-HBV + pCDNA3.1-HS3ST3B1 + psh1126 (a plasmid to interfere HS3ST3B1 expression); (6) Treatment B, transfected with pCH9-HBV + pTZU6+1; (7) Interference B, transfected with pCH9-HBV + psh1126. The levels of HBV DNA were detected in the above groups by Southern blotting. HBV total RNA of Negative control, Treatment A and Interference A were quantified by Real-time PCR to determine the influence of HS3ST3B1 over-expression on the HBV RNA transcription. The activity of the four HBV promoters [core promoter (cp), x promoter(xp), surface antigen promoter1(sp1), surface antigen promoter2 (sp2)] were assayed by Dual-Luciferase Reporter Assay System. The data was analyzed using one way ANOVA, with P < 0.05 indicating statistically meaningful difference.

RESULTSouthern blot data revealed the level of HBV DNA in Treatment A and Interference A accounted for 10% +/- 2% and 31% +/- 4% of that in control. Compared with control, a statistical difference existed between Treatment A and Control, with F value equalling to 20.8 and P value equalling to 0.034 respectively. A statistical difference also existed between Interfere A and Treatment A, with F value equalling to 24.9 and P value equalling to 0.021 respectively. The level of HBV DNA in Experiment B was raised by 130% +/- 11% as compared to that in Interference B, and the levels of HBV DNA showed a dose-dependent decrease when H7 cells were transfected with 0.5, 1.0, 1.5 microg pCDNA3.1-HS3ST3B1 respectively. Statistical differences existed between control and H7 transfected with different dose of pCDNA3.1-HS3ST3B1, with F values equalling to 22.7, 20.3, 26.5 and P values equalling to 0.029, 0.041 and 0.015 respectively. Real-time PCR revealed that the HBV total RNA in Treatment A accounted for 17.0% +/- 2.7% of that in control and there was a statistical difference between Treatment A and control, with F value equalling to 25.6 and P value equalling to 0.018. In addition, HBV DNA in Interference A was restored to 74.0% +/- 3.9% of that in control, and there was also a statistical difference between Treatment A and Interference A, with F value equalling to 21.3 and P value equalling to 0.032. However, the down regulation of HBV total RNA had nothing to do with HBV promoters activity.

CONCLUSIONHS3ST3B1 can inhibit HBV replication and reduce the level of HBV total RNA, but the downregulation of HBV total RNA may not be the result of direct interaction of HS3ST3B1 and HBV promoters.

DNA Replication ; DNA, Viral ; biosynthesis ; Hep G2 Cells ; Hepatitis B virus ; genetics ; physiology ; Humans ; Plasmids ; Sulfotransferases ; genetics ; Transfection ; Virus Replication

Depression and insomnia are intimately related. Depressed patients usually manifest sleep discontinuity and early awakening, reduced or no slow wave sleep (SWS) and shortened latency of rapid eye movement (REM) sleep. These sleep abnormalities are very similar to those caused by over activated hypothalamic-pituitary-adrenal (HPA) axis with stress. Therefore, the animal models developed by post-traumatic stress disorder or chronic unpredictable mild stress could be used to evaluate drugs which have effects of both anti-depression and improvement of sleep quality, and to provide a more reliable platform for further studis on the mechanisms of depression and accompanied insomnia. This review mainly focuses on the typical features of sleep disturbance of depression, possible pathophysiological mechanisms, establishment of animal stress models and analysis of their abnormal sleep characteristics.
Animals ; Chronic Disease ; Depression ; physiopathology ; Depressive Disorder ; physiopathology ; Disease Models, Animal ; Humans ; Hypothalamo-Hypophyseal System ; physiopathology ; Pituitary-Adrenal System ; physiopathology ; Sleep ; physiology ; Sleep Initiation and Maintenance Disorders ; physiopathology ; Sleep, REM ; Stress Disorders, Post-Traumatic ; physiopathology ; Stress, Psychological ; physiopathology

OBJECTIVETo investigate the major cardiovascular risk factors affecting small arterial elasticity and the effect of combined multiple risk factors on it.

METHODSArterial elasticity indexes (C(1)-large artery and C(2)-small artery) were measured with CVProfilor DO-2020. The status of insulin resistance was evaluated with HOMA (homeostasis model assessment). Subjects were categorized into abnormal C(2) group and control group according to the level of C(2). The former group was further divided into four subgroups (0 to 3) based on the number of risk factors.

RESULTS(1) The levels of age, total cholesterol (TC), low density lipoprotein- cholesterol (LDL-C), fasting blood glucose (FBG), systolic blood pressure (SBP) and diastolic blood pressure (DBP) in abnormal C(2) group were higher than those in control group, whereas C(2) itself was lower than that in control group (P all < 0.05). Age, TC, LDL-C, FBG, SBP and DBP were significantly inversely correlated with C(2). (2) With the clusters of risk factors increasing, C(2) was decreasing (6.5 +/- 2.6 vs 5.4 +/- 2.3 vs 4.7 +/- 2.7 vs 3.1 +/- 1.6, P < 0.001). C(2) decreased significantly in subjects with multiple risk factors (subgroup 3). (3) Fasting plasma insulin and HOMA-IR (insulin resistance index) were significantly higher in subgroup 3 than in the other subgroups (P < 0.05, P < 0.001 respectively).

CONCLUSIONSThe elevations of age, TC, LDL-C, FBG, SBP and DBP were the major cardiovascular risk factors on the reduction of C(2), and the effects on it were continuously. With their concurrent effects, multiple risk factors could decrease small arterial elasticity much more significantly. Insulin resistance seems to be closely related to the clusters of multiple risk factors.

Adult ; Aged ; Arterioles ; physiopathology ; Blood Pressure ; Cardiovascular Diseases ; blood ; etiology ; physiopathology ; Cholesterol ; blood ; Cholesterol, HDL ; blood ; Cholesterol, LDL ; blood ; Elasticity ; Female ; Humans ; Insulin Resistance ; Male ; Middle Aged ; Risk Factors

AIMTo study the mechanism of inhibition of basic fibroblast growth factor (bFGF) related signal transduction by lidamycin in cancer cells.

METHODSMTT assay was used to determine the growth inhibitory effect of lidamycin (LDM) and adriamycin (ADR) in several cancer cell lines. The inhibition of bFGF bound to its receptor by LDM was measured with [125I]-bFGF binding assay. Intracellular Ca2+ stimulated by bFGF was measured by Fura-3. The formation of bFGF-receptor immune complex and the inhibitory effect of LDM on the activity of PKC isoenzymes induced by bFGF in cancer cells were identified by Western blotting analysis.

RESULTSLDM displayed extremely potent growth inhibitory effect on several cancer cell lines in a dose-dependent manner. A comparison of the IC50 values showed that the effect of LDM was 1000-fold more potent than that of ADR. LDM blocked the specific binding of [125I]-bFGF to rat lung membranes with an IC50 value of 2.0 x 10(-4) nmol x L(-1). As detected by anti-FGFR specific antibody, LDM inhibited the formation of bFGF-receptor immune complex. bFGF induced cytosolic Ca2+ response was obstructed by pretreatment with 10 nmol x L(-1) LDM. Immunoblotting demonstrated that LDM inhibited the activity of PKC isoenzymes in cancer cells stimulated with bFGF.

CONCLUSIONThe blocking of bFGF receptors in the signal transduction pathway may be involved in the effect of LDM on cancer cells.

Aminoglycosides ; administration & dosage ; pharmacology ; Animals ; Antibiotics, Antineoplastic ; administration & dosage ; pharmacology ; Breast Neoplasms ; pathology ; Calcium ; metabolism ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Dose-Response Relationship, Drug ; Doxorubicin ; pharmacology ; Enediynes ; administration & dosage ; pharmacology ; Female ; Fibroblast Growth Factor 2 ; pharmacology ; HT29 Cells ; Humans ; Membrane Proteins ; metabolism ; Protein Binding ; Protein Kinase C ; metabolism ; Rats ; Receptors, Fibroblast Growth Factor ; metabolism ; Signal Transduction