AIM:To investigate the effect of cyclopamine on Hedgehog (HH) signaling, phenotypic transfor-mation and matrix accumulation induced by aristolochic acid (AA) in renal tubular epithelial cell NRK-52E.METHODS:NRK-52E cells were randomly divided into control group (treated with solvent only), AA group (treated with AA at con-centrations of 1, 5, 10 mg/L) and cyclopamine group (treated with AA at concentration of 10 mg/L plus cyclopamine at concentrations of 1, 5, 10μmol/L).After cultured for 24 h, the mRNA expression of Ptch1, Smo,α-SMA, E-cadherin, ZO-1, BMP-7, type I collagen and type III collagen was quantified by real-time PCR.The protein levels of Shh and TGF-β1 were detected by ELISA .Immunofluorescence staining was used to evaluate the expression of Ptch 1, Smo,α-SMA, E-cadherin and type III collagen in the NRK-52E cells.RESULTS: AA increased the expression of TGF-β1, α-SMA and type III collagen, decreased the expression of E-cadherin and ZO-1 protein, and down-regulated the expression of Ptch1, Shh and Smo mRNA in the NRK-52E cells, indicating that AA activated HH signaling , and phenotypic transformation and matrix accumulation occurred in AA-treated NRK-52E cells.Treatment with cyclopamine inhibited HH signaling by decrea-sing Smo expression and increasing Ptch 1 expression.Moreover, cyclopamine also down-regulated the expression of TGF-β1,α-SMA, type I collagen and III collagen , and up-regulated the expression of BMP-7, ZO-1 and E-cadherin.CON-CLUSION:AA induces phenotypic transformation and matrix accumulation in renal tubular epithelial cells , which can be inhibited by cyclopamine treatment .The possible mechanism is that cyclopamine suppresses the activation of HH signaling , resulting in the reduction of epithelial-to-mesenchymal transition and matrix deposition .

Cell Membrane Permeability ; Crystallization ; Dosage Forms ; Nephelometry and Turbidimetry ; Pharmaceutical Preparations ; chemistry ; Pharmacokinetics ; Quantitative Structure-Activity Relationship ; Solubility ; Technology, Pharmaceutical ; methods ; trends

AIMTo investigate the combined effect of cosolvent and cyclodextrin (CD) on solubilization of insoluble drugs.

METHODSPhase-solubility method was applied to determine solubilization of two diterpenoids in cosolvent / cyclodextrin combinations. The combined effect was evaluated and explained with an established mathematical model, and the model parameters were calculated by means of nonlinear regression analysis.

RESULTSThe strong agreement between the predicted and the observed solubility data supports the validity of the proposed model, with the determination coefficients of two regression models were 0.993 and 0.992, separately.

CONCLUSIONThe validated mathematical model can be used to explain and predict the combined solubilization of the two insoluble drugs in different cosolvent systems.

2-Hydroxypropyl-beta-cyclodextrin ; Algorithms ; Diterpenes ; chemistry ; Models, Chemical ; Solubility ; Solvents ; chemistry ; Water ; chemistry ; beta-Cyclodextrins ; chemistry

Objective To compare the differences between two statistical methods for evaluating non-sensitivity of pathogenic bacteria to antimicrobial agents,and explore effect of non-consideration of clinical background on evalua-ting extent of bacterial resistance.Methods Data of Staphylococcus aureus and Acinetobacter spp .in a hospital in the first half year of 2008,2010 and 2013 were collected and conducted statistical analysis with two methods (me-thod 1 :based on all clinically isolated bacteria;method 2 :based on infection-related non-repetitive bacteria),two methods for evaluating bacterial non-sensitive rates to antimicrobial agents were compared.Results The non-sensi-tive rates of Acinetobacter spp .to various antimicrobial agents :statistical results by using method 1 were generally higher than those using method 2,absolute difference between two statistical methods was 10.46%-33.77%;the non-sensitive rates of Staphylococcus aureus to various antimicrobial agents :except compound sulfamethoxazole in 2010 and 2013(difference were 6.17% and 10.21 % respectively),penicillin G (difference was 3.86%),erythromy-cin (difference was 2.71 %),and azithromycin in 2013 (difference was 2.43%),statistical results by using method 1 were generally higher than those using method 2,absolute difference between two statistical methods was 0-18.04%.Conclusion There are deviation in the non-sensitive rates of bacterial strains to antimicrobial agents by using two different statistical methods,deviation is larger in Acinetobacter spp ..The resistance level might be incorrectly higher when evaluating the resistance status without considering clinical background of bacteria.

Objective To investigate the expression of monocyte-macrophage-related factors and interstitial fibrosis in kidney tissues of rats with ureter obstruction and recanalization .Methods Forty-eight male Spragur-Dawley rats were divided randomly into the obstructive group:sham (n=6), unilateral ureteral obstruction(UUO)3 days (n=6), UUO 7 days (n=6), and UUO 14 days (n=6) and recanalization group:bilateral ureteral obstruction(RBUO)0 day (n=6), 3 days after RBUO (n=6), 7 days after RBUO (n=6), and 14 days after RBUO (n=6).The kidneys were excised on day 3, 7, and 14, and the deposition of collagen fibers in kidney was detected with HE and Masson staining . Immunohistochemical analysis was performed to evaluate the protein expressions of monocyte chemoattractant protein -1 (MCP-1), macrophage colony-stimulating factor (M-CSF) and activated-macrophage marker CD68.Real-time PCR was used to detect the mRNA expressions of MCP-1 and M-CSF.TGF-β1 levels were determined by ELISA .Results Fibrosis observed with HE and Masson staining was obviously increased in kidney tissue of UUO rats , and aggravated as time prolonged, but alleviated in rats with recanalization .TGF-β1 levels were increased obviously in the UUO group , but decreased in rats with recanalization compared with those in BUO rats .In UUO rats, mRNA and protein expression levels of MCP-1 and M-CSF were increased .MCP-1 and M-CSF expression was gradually decreased in rats with recanalization compared with those in BUO rats .The dynamic change in expression of MCP-1 and M-CSF in both UUO rats and recanalization rats was consistent with the change in expression of CD 68. Conclusion Dynamic change in expression of MCP-1 and M-CSF in kidney tissues reflects change of activated and accumulated monocyte -macrophages , which may be one of the major mechanisms contributing to fibrosis induced by ureter obstruction .Renal fibrosis is alleviated by down-regulated expression of monocyte-macrophages factors with recanalization operation .

Objective To probe into the clinical curative effect of laparoscopic high ligation of spermatic vein in the treatment of critical varicocele.Methods The clinical data of the 26 eases of laparoscopic high ligation of apermatic vein in the treatment of critical varicocde were reviewed and analyzed in the last two years in this hospi- tal.Results All the 26 cases had been conducted smoothly with the operation time 25～50min,an average of(28?3)min.After 3～24 months being followed-up,all the symptoms and signs disappeared with no relapse and testicle a- trophy.Eight wives of the patients who had been operated on got pregnant.Conclusion With soon recovery and small wound,it was safe to adopt laparoscopic high ligation of spermatic vein in the treatment of critical varicocele, especially for critical two-sided varicocde.

Animals ; Epithelial-Mesenchymal Transition ; Fibrosis ; Kidney ; pathology ; Kidney Tubules ; cytology ; Male ; Rats ; Rats, Sprague-Dawley ; Ureteral Obstruction ; pathology

Objective To investigate the efficiency of Trichostatin A (TSA) in inducing cell apoptosis and altering the Notch pathway genes expression in PANC-1 cells line.Methods The survival rate and apoptosis of PANC-1 cells were measured by MTT assay and Hoechst 33258 staining,respectively.mRNA expression levels of the genes,numb,gcn512,dll3,hes6,eaf2,cytohesins,in PANC-1 cells were assessed by real-time quantitive PCR.Western blot was used to measure the expression of bcl-2,bax,actived caspase-3 and NICD protein which was the biologically active form of Notch-1.Results After culturing with 0.1,0.2,and 0.4 μmol/L TSA for 24 hours,the cellular survival rate of PANC-1 cells significantly decreased to 72％,58％ and 39％,respectively.The survival rate of PANC-1 was negatively correlated to time length of culture with TSA.Increased apoptosis of PANC-1 cells after 12,24 and 36 h culture with TSA was detected by Hoechst 33258 staining.Western blotting showed that the expression of bax,actived caspase-3 and NICD protein increased while the bcl-2 protein decreased after culture with TSA.In real time quantitive PCR assessment,the mRNA expression of numb and hes6 in PANC-1 cells were upregulated by TSA (P ＜ 0.05),while the mRNA expression of gcn512 and dll3 were down-regulated by TSA (P ＜ 0.05).While mRNA expressions of eaf2 and cytohesin1,2,3,4 were not affected by TSA.Conclusions TSA induces apoptosis of pancreatic cancer cell line PANC-1.The Notch signal pathway may be involved in inducing cellular apoptosis of PANC-1 when cultured with TSA.

Blood-eye barrier (BEB) is one of the most important structures of organism to maintain homeostasis of the eye. However, it is the major constraint for the medication of intraocular diseases. Traditional Chinese medicines have distinctive advantages for the treatment of intraocular diseases, which can be used to regulate the physiological function of human body with low toxicity. In this article, we have briefly summarized the feature of BEB, with the domestic and foreign literatures combined, and mainly reviewed current progress in the field of study on the permeability of traditional Chinese medicines and effective components in BEB and promoting methods.

Previous methods used for nuclear transplantation were further investigated to develop a method that was both easy to carryout and did not require any special apparatus, such as Piezoimpact or Spindle-View. Following the puncture of zona pellucida with two holes by injection pipette that contained donor nuclei or cells, the injection pipette was pulled back to the perivitelline space while the negative pressure was increased in the holding pipette until the polar body and karyoplasm were wiped off completely. Then a reconstructed embryo was completed by the direct injection of the donor nucleus or cell without pulling out the injection pipette. 200 oocytes were manipulated using this method and it cost about 40 seconds with nucleus injection and about 30 seconds with cell injection to complete a reconstructed embryo. The success rates were 62.6% and 86. 0%, respectively, and enucleation rate was about 73.3% validated by Hoechst 33342. Using this method, the nucleus was completely eliminated and another was injected using the microscope and micromanipulator. Moreover, the efficiency of nuclear transplantation and survival rate of reconstructed embryos were greatly improved. Furthermore, it is very easy to manipulate and popularize in practice.
Animals ; Cell Culture Techniques ; methods ; Cell Nucleus ; metabolism ; Cells, Cultured ; Cloning, Organism ; methods ; Embryo, Mammalian ; cytology ; metabolism ; Embryonic Development ; Female ; Male ; Mice ; Mice, Inbred C57BL ; Mice, Inbred DBA ; Mice, Inbred Strains ; Nuclear Transfer Techniques ; Oocytes ; cytology ; metabolism ; Zona Pellucida ; metabolism