The Telomeric Repeat Amplification Protocol(TRAP)and its modified versions change the size and/or the ratio of the telomerase products in the amplification stage of the assay.Based on TRAP,a useful method was developed for detecting telomerase activity in rice.A special precursor primer and a special reverse primer and conducted two steps of PCR cycles were designed.GENE Genius? Bio-imaging System was applied for this quantitative analysis for exploring telomerase activity and its optimal reaction conditions.The method ensured that the optimal reaction conditions for the telomerase was 19℃,13minutes,at a concentration of 0.28 u g/? l.A quantitative analysis method was established for detecting telomerase activity in rice.With this method,we detected telomerase activity in roots,young leaves and young panicles of six parental lines of hybrid rice.The results show that young panicles have the highest telomerase activity,demonstrating that telomerase activity is closely related to the cell vitality in plants.

Objective To study the rationality and possibility of 64 slice spiral CT in the examination of the temporal bone at low dose.Methods The same CT technique and temporal bone mode as those for clinical CT were used,two cranium specimens(four ears)were scanned with Somatom Sensation 64-slice spiral CT at different mA(380,300,200,160,120,80),and muhi-planar reformation was performed.The CT dose index at different mA groups were measured by 10 cm pencil ionization chamber and head dose phantom.Four anatomic structures on axial images(subarcuate fossa,tendon of tensor tympani, facial recess,etc),four anatomic structures on coronal images(scute,crista transversa,fenestra cochleae, etc)and six anatomic structures on double oblique images(malleus,incus,stirrup bone,upper bony semicircular,etc)were chosen to evaluate and grade the reformation images among different mA groups,and to determine the minimum mA value.Ten ears of five patients were used to test the validity of the minimum mA value.Results CT radiation dose was significantly reduced from(47.8?2.7)to(20.1?2.0)raCy (P

OBJECTIVETo investigate the possible protective effect and mechanism of ginsenoside Rb1 against oxidative damage and renal interstitial fibrosis on rats with unilateral ureteral obstruction (UUO).

METHODSIn total, 80 male rats were randomly divided into 4 groups, 20 in each group: the sham operated group (SOR), UUO group, UUO with ginsenoside Rb1 treatment group (treated with intraperitoneal injection of 50 mg/ kg daily) and UUO with Losartan treatment group (as the positive control, treated with 20 mg/kg by gastrogavage per day). The rats were randomly sacrificed on day 3, 7 and 14 after surgery, respectively. The histopathologic changes of renal interstitial tissues were observed with Masson staining. The mRNA of transforming growth factor beta 1 (TGF-beta 1), collagen I and fibronectin were reversed transcribed and quantified by Real-time PCR. Enzyme-linked immunosorbent assay was used to quantitatively detect TGF-beta 1 and 8-hydroxy-2'-deoxyguanosine (8-OHdG) levels. P47phox protein expression was assessed by immunohistochemistry and Western blot analysis.

RESULTSIn the UUO model, the obstructed kidney showed typical features of progressive renal tubulointerstitial fibrosis, and the levels of TGF-beta1, collagen I and fibronectin increased (P<0.05). As compared with the UUO group, ginsennoside Rb1 significantly inhibited the interstitial fibrosis including tubular injury and collagen deposition, and decreased the levels of TGF-beta1 (P<0.05). Ginsenoside Rb1 also inhibited the heme oxygenase (HO-1) and 8-OHdG, two markers of oxidative stress (P<0.05). Moreover, ginsenoside Rb1 suppressed the expression of p47phox, a subunit of nicotinamide adeninedinucleotide phosphate (NADPH) oxidase (P<0.05).

CONCLUSIONGinsenoside Rb1 can obviously inhibit renal interstitial fibrosis in rats with UUO, its mechanism possibly via against the oxidative damage and suppressing TGF-beta1 expression.

Animals ; Deoxyguanosine ; analogs & derivatives ; urine ; Drug Evaluation, Preclinical ; Fibrosis ; genetics ; metabolism ; prevention & control ; Gene Expression Regulation ; drug effects ; Ginsenosides ; therapeutic use ; Heme Oxygenase (Decyclizing) ; metabolism ; Kidney ; drug effects ; metabolism ; pathology ; Kidney Diseases ; etiology ; genetics ; pathology ; prevention & control ; Male ; Models, Biological ; NADPH Oxidases ; genetics ; metabolism ; Oxidative Stress ; drug effects ; Rats ; Rats, Sprague-Dawley ; Saponins ; therapeutic use ; Transforming Growth Factor beta1 ; genetics ; metabolism ; Ureteral Obstruction ; complications ; drug therapy ; genetics ; metabolism

Objective To construct four recombinant Lactococcus lactis strains exhibiting high β-galactosidase activity in fusion or non-fusion ways, and to study the influence factors for their protein expression and secretion. Methods The gene fragments encoding β-galactosidase from two strains of Lactobacillus bulgaricus, wch9901 isolated from yogurt and 1.1480 purchased from the Chinese Academy of Sciences, were amplified and inserted into lactococcal expression vector pMG36e. For fusion expression, the open reading frame of the β-galactosidase gene was amplified, while for non-fusion expression, the open reading frame of the β-galactosidase gene was amplified with its native Shine-Dalgarno sequence upstream. The start codon of the β-galactosidase gene partially overlapped with the stop codon of vector origin open reading frame. Then, the recombinant plasmids were transformed into Escherichia coli DH5α and Lactococcus lactis subsp, lactis MG1363 and confirmed by determining β-galactosidase activities. Results The non-fusion expression plasmids showed a significantly higher β-galactosidase activity in transformed strains than the fusion expression plasmids. The highest enzyme activity was observed in Lactococcus lactis transformed with the non-fusion expression plasmids which were inserted into the β-galactosidase gene from Lactobacillus bulgaricus wch9901. The β-galactosidase activity was 2.75 times as high as that of the native counterpart. In addition, β-galactosidase expressed by recombinant plasmids in Lactococcus lactis could be secreted into the culture medium. The highest secretion rate (27.1%) was observed when the culture medium contained 20 g/L of lactose. Conclusion Different properties of the native bacteria may have some effects on the protein expression of recombinant plasmids. Non-fusion expression shows a higher enzyme activity in host bacteria. There may be a ost-related weak secretion signal peptide gene within the structure gene of Lb. Bulgaricus β-galactosidase, and its translation product may introduce the enzyme secretion out of cells in special hosts.

OBJECTIVETo investigate the effects of ginsenoside R(g1) on the transdifferentiation of rat renal tubular epethelial cells induced by transforming growth factor-beta1, (TGF-beta1).

METHODCultured normal rat renal tubular epethelial cells (NRK-52E) were divided into control group, TGF-beta1-induced group and treated with ginsenoside R(g1) at different concentration (10, 20, 40 mg x L(-1)) group. The morphology of tubular epithelial-myofibroblast transdifferentiation induced by TGF-beta1 was observed through light microscope. alpha-SMA and E-cadherin protein expression were assessed by immunohistochemistry and western blot analyses. alpha-SMA, collagen I and and fibronectin gene expression were assessed by real-time quantitative chain reaction. Enzyme-linked immunosorbent assay was used to quantitatively detect collagen I and fibronectin in the supernatant.

RESULT10 mg x L(-1) TGF-beta1 could induce the transdifferentiation of tubular epithelial myofibroblast, showing fibroblast-like in morphology, with significantly enhanced expression of alpha-SMA, depressed expression of E-cadherin and increased secretion of fibronectin and collagen I (P < 0.05). Compared to TGF-beta1-induced group, ginsenoside R(g1) partly abrogated the alpha-SMA expression and E-cadherin depression triggered by TGF-beta1 in tubular epithelial cells in a dose-dependent manner (P < 0.05). Meanhile, ginsenoside R(g1) blocked morphologic transformation of tubular epithelial cells and decreased levels of collagen I and fibronectin (P < 0.05).

CONCLUSIONGinsenoside R(g1) could inhibit TGF-beta1 induced the tubular epithelial-myofibroblast transdifferentiation and decreased levels of collagen I and fibronectin in NRK52E.

Animals ; Cadherins ; genetics ; metabolism ; Cell Line ; Cell Transdifferentiation ; drug effects ; Drugs, Chinese Herbal ; pharmacology ; Epithelial Cells ; cytology ; drug effects ; Gene Expression ; drug effects ; Ginsenosides ; pharmacology ; Kidney Tubules ; cytology ; drug effects ; Panax ; chemistry ; Rats ; Transforming Growth Factor beta1 ; pharmacology

Total saponins of Panax notoginseng (PNS) have been shown to ameliorate renal interstitial fibrosis. Ginsenoside Rg1, a panaxatriol saponin, is one of the major active molecules from PNS. The present study was undertaken to investigate the effect of ginsenoside Rg1 on renal fibrosis in rats with unilateral ureteral obstruction (UUO). The rats were randomly divided into 3 groups: sham-operation (n=15), UUO (n=15) and UUO with ginsenoside Rg1 treatment (n=15, 50 mg per kg body weight, intraperitoneally (i.p.) injected). The rats were sacrificed on Days 7 and 14 after the surgery. Histological examination demonstrated that ginsenoside Rg1 significantly inhibited interstitial fibrosis including tubular injury as well as collagen deposition. alpha-smooth muscle actin (alpha-SMA) and E-cadherin are two markers of tubular epithelial-myofibroblast transition (TEMT). Interestingly, ginsenoside Rg1 notably decreased alpha-SMA expression and simultaneously enhanced E-cadherin expression. The messenger RNA (mRNA) of transforming growth factor-beta1 (TGF-beta1), a key mediator to regulate TEMT, in the obstructed kidney increased dramatically, but was found to decrease significantly after administration of ginsenoside Rg1. Further study showed that ginsenoside Rg1 considerably decreased the levels of both active TGF-beta1 and phosphorylated Smad2 (pSmad2). Moreover, ginsenoside Rg1 substantially suppressed the expression of thrombospondin-1 (TSP-1), a cytokine which can promote the transcription of TGF-beta1 mRNA and the activation of latent TGF-beta1. These results suggest that ginsenoside Rg1 inhibits renal interstitial fibrosis in rats with UUO. The mechanism might be partly related to the blocking of TEMT via suppressing the expression of TSP-1.
Actins ; biosynthesis ; Animals ; Cadherins ; biosynthesis ; Collagen Type I ; genetics ; metabolism ; Fibronectins ; genetics ; metabolism ; Ginsenosides ; pharmacology ; Immunohistochemistry ; Male ; Nephritis, Interstitial ; drug therapy ; genetics ; metabolism ; pathology ; Panax notoginseng ; chemistry ; RNA, Messenger ; biosynthesis ; genetics ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Reverse Transcriptase Polymerase Chain Reaction ; Smad2 Protein ; biosynthesis ; Thrombospondin 1 ; biosynthesis ; genetics ; Transforming Growth Factor beta1 ; biosynthesis ; genetics ; Ureteral Obstruction ; metabolism ; pathology

OBJECTIVETo determine the susceptibilities of M. tuberculosis H37Ra and M. chelonei subsp. absecessus to several frequently-used disinfectants and to evaluate the practicability of surrogating M. tuberculosis by the latter.

METHODSA suspension quantitative bactericidal test was set up in accordance with Chinese Technique Standard for Disinfection to evaluate the susceptibility of each mycobacteria strain to each selected disinfectant. Killing log value was used as criterion in comparing the susceptibility to disinfectants between the two strains.

RESULTSM. chelonei subsp. abscessus was more resistant to chlorine disinfectant than M. tuberculosis while the two strains were similarly resistant to iodophor disinfectant, peracetic acid, alcohol and glutaraldehyde disinfectant.

CONCLUSIONM. chelonei subsp. abscessus has the potential to surrogate M. tuberculosis in evaluating mycobactericidal efficacies of disinfectants.

Alcohols ; pharmacology ; Bacteriological Techniques ; Chlorine Compounds ; pharmacology ; Disease Outbreaks ; Disinfectants ; pharmacology ; Drug Resistance, Microbial ; Glutaral ; pharmacology ; Iodophors ; pharmacology ; Microbial Sensitivity Tests ; Mycobacterium Infections ; Mycobacterium chelonae ; drug effects ; Mycobacterium tuberculosis ; drug effects ; Peracetic Acid ; pharmacology ; Time Factors

OBJECTIVETo develop a fast, high-throughput screening method with suspension array technique for simultaneous detection of biothreat bacteria.

METHODS16 S rDNA universal primers for Bacillus anthracis, Francisella tularensis, Yersinia pestis, Brucella spp.and Burkholderia pseudomallei were selected to amplify corresponding regions and the genus-specific or species-specific probes were designed. After amplification of chromosomal DNA by 16 S rDNA primers 341A and 519B, the PCR products were detected by suspension array technique. The sensitivity, specificity, reproducibility and detection power were also analyzed.

RESULTSAfter PCR amplification by 16 S rDNA primers and specific probe hybridization, the target microorganisms could be identified at genus level, cross reaction was recognized in the same genus. The detection sensitivity of the assay was 1.5 pg/microl (Burkholderia pseudomallei), 20 pg/microl (Brucella spp.), 7 pg/microl (Bacillus anthracis), 0.1 pg/microl (Francisella tularensis), and 1.1 pg/microl (Yersinia pestis), respectively. The coefficient of variation for 15 test of different probes was ranged from 5.18% to 17.88%, it showed good reproducibility. The assay could correctly identify Bacillus anthracis and Yersinia pestis strains in simulated white powder samples.

CONCLUSIONThe suspension array technique could be served as an opening screening method for biothreat bacteria rapid detection.

Bacillus anthracis ; isolation & purification ; Bioterrorism ; prevention & control ; DNA Primers ; DNA, Bacterial ; analysis ; Francisella tularensis ; isolation & purification ; Oligonucleotide Array Sequence Analysis ; methods ; Polymerase Chain Reaction ; methods ; RNA, Ribosomal, 16S ; genetics ; Yersinia pestis ; isolation & purification

OBJECTIVETo investigate the association of serum vitamin D [25-(OH)D] level with the severity and treatment in children with Henoch-Schönlein purpura (HSP).

METHODSA total of 50 children with newly-diagnosed HSP between January and December, 2015 were enrolled as HSP group, and 49 healthy children were enrolled as control group. Fasting serum samples were collected, and ELISA was used to measure serum 25-(OH)Dlevel. According to the serum 25-(OH)Dlevel, the HSP group were further divided into normal group (>20 ng/mL) (n=9), insufficiency group (15-20 ng/mL) (n=15), deficiency group (≤15 ng/mL) (n=25), and severe deficiency group (≤5 ng/mL) (n=1). The general data, clinical manifestations, hormone therapy, course of disease before admission, and length of hospital stay were compared between groups.

RESULTSThe HSP group had a significantly lower serum 25-(OH)Dlevel than the control group (16±6 ng/mL vs 29±5 ng/mL; P<0.01). Compared with the normal and insufficiency groups, the deficiency and severe deficiency groups had significant increases in the incidence rate of renal involvement, rate of hormone application, and median length of hospital stay (P<0.05), while there was no significant difference in course of disease before admission (P>0.05).

CONCLUSIONSChildren with HSP have a low serum 25-(OH)Dlevel, and such children may have a high risk of renal involvement, a high rate of hormone application, and a prolonged length of hospital stay. However, further studies are needed to investigate whether vitamin D supplementation is helpful to the treatment of HSP and can shorten the course of disease in children with HSP.

Child ; Female ; Humans ; Length of Stay ; Male ; Purpura, Schoenlein-Henoch ; blood ; complications ; drug therapy ; Severity of Illness Index ; Vitamin D ; analogs & derivatives ; blood

OBJECTIVETo construct four recombinant Lactococcus lactis strains exhibiting high beta-galactosidase activity in fusion or non-fusion ways, and to study the influence factors for their protein expression and secretion.

METHODSThe gene fragments encoding beta-galactosidase from two strains of Lactobacillus bulgaricus, wch9901 isolated from yogurt and 1.1480 purchased from the Chinese Academy of Sciences, were amplified and inserted into lactococcal expression vector pMG36e. For fusion expression, the open reading frame of the beta-galactosidase gene was amplified, while for non-fusion expression, the open reading frame of the beta-galactosidase gene was amplified with its native Shine-Dalgarno sequence upstream. The start codon of the beta-galactosidase gene partially overlapped with the stop codon of vector origin open reading frame. Then, the recombinant plasmids were transformed into Escherichia coli DH5 alpha and Lactococcus lactis subsp. lactis MG1363 and confirmed by determining beta-galactosidase activities.

RESULTSThe non-fusion expression plasmids showed a significantly higher beta-galactosidase activity in transformed strains than the fusion expression plasmids. The highest enzyme activity was observed in Lactococcus lactis transformed with the non-fusion expression plasmids which were inserted into the beta-galactosidase gene from Lactobacillus bulgaricus wch9901. The beta-galactosidase activity was 2.75 times as high as that of the native counterpart. In addition, beta-galactosidase expressed by recombinant plasmids in Lactococcus lactis could be secreted into the culture medium. The highest secretion rate (27.1%) was observed when the culture medium contained 20 g/L of lactose.

CONCLUSIONDifferent properties of the native bacteria may have some effects on the protein expression of recombinant plasmids. Non-fusion expression shows a higher enzyme activity in host bacteria. There may be a host-related weak secretion signal peptide gene within the structure gene of Lb. bulgaricus beta-galactosidase, and its translation product may introduce the enzyme secretion out of cells in special hosts.

Erythromycin ; pharmacology ; Gene Expression Regulation, Bacterial ; Lactobacillus ; drug effects ; enzymology ; genetics ; Lactococcus lactis ; drug effects ; enzymology ; genetics ; Lactose ; metabolism ; pharmacology ; Recombinant Proteins ; genetics ; metabolism ; Time Factors ; beta-Galactosidase ; genetics ; metabolism