Objective To observe the expressions of transforming growth factor-?1(TGF-?1) and plasminogen activator inhibitor-1(PAI-1) mRNA in tubulointerstitial fibrosis( TIF ) rats,and explore the effects of TGF-?1 and PAI-1 on TIF and the interaction between TGF-?1 and PAI-1.Methods Sixty male SD rats were randomly divided into sham operation group (SOR group,n=30) and unilateral urethral obstruction group(UUO group,n=30).TIF model was established via UUO.At d 7,d 14,d 21 after operation,10 rats of every group were killed to obtain renal samples.Histological changes were observed in tubulointerstitium injury under microscope;mRNA and proteins of TGF-?1 and PAI-1 were detected by real-time PCR assays and Western blotting respectively at d 7,d 14,d 21 after experiment onset.SPSS 10.0 software was used to analyze data.Results 1.HE,Masson staining of renal tissues of all groups indicated that there were no fibrosis in SOR rats,there were vacuole degeneration in tubular epithelial cells and inflammatory cell infiltration in tubulointerstitial in UUO rats at d 7,fibrosis aggravated gradually and became severe fibrosis at d 21.2.The expression locations of TGF-?1 and PAI-1 were renal sites of fibrosis by immunohistochemical staining.The mRNA and proteins of TGF-?1 in rats from SOR groups were lower than those of UUO groups at d 7,d 14,d 21(P

Objective To investigate the radioresistant effects of pprI gene of Deinococcus radiodurans on BALB/c mice.Methods Male BALB/c mice in SPF level were applied for this work.The pEGFP-c1 plasmid and pEGFP-c1-pprI gene recombinant plasmid were transferred into anterolateral muscle of mice with in vivo electroporation technology.The mice were irradiated by 6 Gy 60Co γ-rays in whole body and the mortality of mice was observed within 30 days after irradiation.In addition,the mouse were irradiated with 4 Gy γ-rays and then the peripheral blood cell number,apoptosis rates of thymocyte cells,spleen cells and bone marrow cells were observed in the days of 1,7,14,28 and 35 after irradiation while the histopathological changes of lung and testis were observed in the days 7 and 28 after γ-ray irradiation.Results The highest gene transfection efficiency of muscle cells was obtained in a Plasmid injection amount of 50 μg/50 μl and electric field strength of 200 V/cm.The acute radiation mortality of pEGFP-c1-pprI gene recombinant plasmid transfer group was 30％,lower than that of irradiation group (60.0％) and pEGFP-c1 plasmid transfer group (63.3％) after 6 Gy γ-ray irradiation (x2 =4.90,6.24,P ＜ 0.05).Compared with the irradiation group and pEGFP-c1 plasmid transfer group,the WBC count of pEGFP-c1-pprI gene recombinant plasmid group in peripheral blood of mice was significantly higher in the days of 1,7,14 and 28 (F =16.26,8.10,6.37,10.74,P ＜0.05),PLT count was significantly higher in days of 7 and 14 (F =7.36,5.71,P ＜ 0.05),meanwhile the lymphocyte percentage was increased significantly on the 7th day (F =18.43,P ＜ 0.05) after irradiation.On the other hand,the apoptosis rates of thymocyte cells and bone marrow cells were significantly decreased in the days of 1,7,14,28 and 35 (F =3.88,14.91,14.14,39.86,5.65,P ＜0.05 and F=53.70,11.75,21.78,41.40,4.54,P ＜0.05) while the apoptosis rate of spleen cells was significantly decreased in the days of 1,7,14 and 28 (F =97.95,56.61,33.55,14.71,P ＜0.05) after irradiation.Finally,the radiation histopathological changes of lung and testis of the pEGFP-c1-pprI gene recombinant plasmid group were slight and easy to recover.Conclusions Transfection of pprI gene of Deinococcus radiodurans by in vivo electroporation has significant protective effect on the acute radiation injury in BALB/c mice,which may have important clinical applications.

Objective To construct the eukaryotic expression vector of pprI gene from Deinococcus radiodurans R1 and investigate its radioresistant effects in eukaryotic cells.Methods A recombinant vector pEGFP-c1-pprI was constructed by DNA recombinant technique.The empty vector pEGFP-c1 and the pEGFP-c1-pprI were transferred into human lung epithelial cells Beas-2B by LipofectamineTM 2000,respectively.Then the infected cells were screened in order to develop a cell line with stable expression of pprI gene.Cell survival rate was tested by clone-forming assay.Cell cycle distribution and apoptosis were detected by a flow cytometry.The fluorescence intensity of reactive oxygen species (ROS) was observed by a fluorescent microscope.γ-H2AX foci in the irradiated cell was detected by immunofluorescence.Results The eukaryotic expression plasmid of pprI prokaryotic gene was constructed and PprI fusion protein was expressed in human lung epithelial cells successfully,and the cell line (2BG) with a stable pprI gene expression was established.After irradiation,the cell survival fraction of 2BG cells was significantly higher than Beas-2B cells so that the value of D0 、Dq and N of the survival curve were increased.Moreover,the fluorescence intensity of ROS and the number of γ-H2AX foci in 2BG cells were also lower than those of B eas-2B cells(F =16.73,19.47,6.94,P ＜ 0.05).Between these two cell lines,the apoptosis rate and cell cycle G2 arrest also had significant difference (F =139.73,237.92,P ＜ 0.05).Conclusions The pprI gene from Deinococcus radiodurans RI can be stably expressed in the eukaryotic cells and it allows the transferred cells to have a radioresistant function.

(3S)-Linalool synthase (LIS) is a key enzyme involved in the monoterpene biosynthetic pathway. Based on our previous transcriptome study, the expression level of LIS gene was exceedingly related to glycyrrhizic acid (GA) biosynthesis. Therefore, we used hairy root culturing to further investigate the effect of LIS on the GA biosynthesis. A LIS gene (GenBank accession number: MZ169552) was cloned from Glycyrrhiza uralensis. The plant binary overexpression vector pCA-LIS was constructed by gene fusion. G. uralensis hairy roots overexpressing LIS were induced by the Agrobacterium rhizogenes ATCC15834. The expression levels of LIS were analyzed by real-time quantitative PCR (RT-qPCR) and the contents of GA in hairy root lines were determined by UPLC. It was found that in the hairy root lines overexpressing LIS, the expression levels of LIS were significantly higher than that in the wild type, while the contents of GA were remarkably lower than those in the wild type and negative control. These findings indicate that the expression level of LIS is negatively correlated with the accumulation of GA. In this study, LIS was cloned from G. uralensis for the first time and the negative regulatory effect of LIS on GA biosynthesis was confirmed by reverse genetics. This work provides support for further improvement of the molecular regulatory network of GA biosynthesis in G. uralensis.

Aristolochic Acids ; adverse effects ; Child ; Drugs, Chinese Herbal ; adverse effects ; Female ; Humans ; Male ; Nephrosis ; chemically induced

Plants in Ainsliaea genus, belongs to Compositae family, are traditional Chinese medicine and widely used in folk. These plants contain various types of chemical components, and main components are sesquiterpene lactone and its glycosides. In addition, there are triterpenoids, flavonoids, steroids, phenolic acid, long chain fatty acid and volatile oils. Recently, much attention has been payed to varlous research of A. fragrans. This paper reviewed and summarized the chemical components to provide the theoretical basis for the use of Ainsliaea.
Asteraceae ; chemistry ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; Molecular Structure

Background Some drugs with inhibitory effect on the proliferation of lens epithelial cells have a limiting application in clinic because of their adverse response.To screen the effective and less side-effect drug for supressing LECs growth is very inportant for the prevention and treatment of after cataract.Objective This study was to explore the effects of docetaxel on LECs growth and compare its role with epirubicin hydrochloride,pirarubicin hydrochloTide and rahitrexed.Methotis Immortalized human LECs line (SRA01/04) were cultured and passaged.Different concentrations of docetaxel,epirubicin hydrochloride,pirarubicin hydrochloride and rahitrexed were added into the medium respectively for 24.48 and 72 hours.The proliferation of LECs was detect by M1Yr.Flow cytometry analysis Was used to analyze the influence of different concentrations of docetaxel on cellular cycle at 48 hours after addition of docetaxel,and Annexin V-FITC/PI marking method was used to assesse the apoptosis of LECs under the action of docetaxel.Expression of bcl-2 protein in LECs Was evaluated by Westeru blot. Result The growth rate of LECs Wag 100%in 8-519 pmol/L doeetaxel groups with the normal cell shape.Majority of abnormal cells and low growth rate were found in 66 nmoVL docetaxel group at 48 and 72 hours.The IC50 of docetaxel was lowest in 48 and 72 hours in docetaxel group in comparison to epirubicin hydrochloride and pirarubicin hydrochloride. However,no evident inhibition on LECs growth in 23.22-523.56 μmol/L of raltitrexed.At 48 hours,the percentage of LECs in G2/M phase increased as the asccnte of concentration of docetaxel,showing a significant difference among 4 groups(F=2633.05,P＜0.01).The percentage of early apoptotic cells increased to 22.4%(χ2=20.00,P＜0.01) and 27.9%(χ2=42.68,P＜0.01)from normal control 3.1% at 48 hours after LECs exposed to 8.3 nmol/L and 266 nmol/L docetaxe.The expression of bcl-2 protein in LECs was obviously weakened after addition of docetaxel,especially 8.3 nmol/L docetaxel group. Conclusion Docetaxel,epirubicin hydrochloride and pirarubicin hydrochloride can inhibit the proliferation of human LECs in vitro.But there is no supression on LECs growth inraltitrexed.Docetaxel is proved to have a strongly arrested effect on the proliferation of LECs in comparison with epirubicin hydrochloride and pirarubicin hydrochloride and play its role at concentration-and time-dependent manner.

Objective To investigate the expression of proliferating cell nuclear antigen(PCNA) in renal tissues of children with primary nephrotic syndrome(NS),and elucidate the relationship between PCNA expression and cell proliferation in renal tissues from the children with primary NS.Methods Paraffin-embedded renal biopsy tissue sections from 39 patients with primary NS were examined by immunohistochemical staining with anti-PCNA monoclonal antibody,normal renal tissue sections from 6 nephrectomized patients with nephroma were selected as control. Possible correlation between the percentage of PCNA positive cells and the pathologic type , histopathological score, clinical indices (serum albumin ,serum cholesterol ,serum creatinine and 24 hours urine protein ) before renal biopsy of NS were evaluated separately .Results The percentage of PCNA positive cells in glomeruli and tubulom terstitium of NS patients was significantly higher than that of the control (P

To look for a more stable and convenient way of constructing short hairpin RNA expression vectors targeting the latent membrane protein-1(LMP-1) encoded by Epstein-Barr virus(pshLMP1), and to study the inhibition function of pshLMP1 expression vectors in HNE1 cells, we designed the pshLMP1 expression cassette and pshLMP1 expression vectors by both the annealing method and PCR method and then co-transfected with pEGFP-N1-1158 into HNE1 cells to observe the mRNA and protein levels of LMP-1 genes by green fluorescence analysis, RT-PCR and western blot. pshLMP1 expression vectors were successfully obtained by both methods but better cloning efficiency was achieved and fewer deletions and mutations of nucleotides were achieved with the PCR method. Furthermore, the mRNA and protein levels of LMP-1 genes were down-regulated by pshLMP1 expression vectors. According to our research, we found that the PCR method provides a more efficient way to construct pshLMP1 expression vectors which have the ability to inhibit the function of LMP-1 genes expressed in HNE1 cells, and also provides a novel application of RNA interference technology against-EBV.

Objective To investigate the pathogen distribution and susceptibility profile of fungal isolates from bloodstream infections,and valuate the clinical utility of G test in diagnosis of fungal infections for the purpose to improve antifungal therapy.Methods A retrospective analysis was carried out to analyze the fungal pathogens isolated from bloodstream infections in the First Affiliated Hospital of Nanjing Medical University during the period from January 2013 through December 2015 and their antimicrobial susceptibility.Results A total of 114 fungal strains were isolated from bloodstream infections during the 3-year period,most of which were Candida (99/114,86.8％),especially Candida albicans (30.7％).About 41.2％ (47/114) of the fungal strains were isolated from Department of Thoracic Surgery (10,5 and 4 strains in 2013,2014 and 2015),Hematology (11 strains in 2014),and ICU (7 strains in 2014).Antimicrobial susceptibility testing showed that all the fungal strains (100％) were susceptible to amphotericin B,but 83.5％ susceptible to itraconazole (the lowest).G test was positive before the result of blood culture in 13 of the 54 patients who received G test.Conclusions Candida was the most common fungus in fungal bloodstream infection.Amphotericin B is the most active antifungal agent in vitro.Blood culture combined with serological test can provide clinicians an earlier and reliable diagnosis.