To establish a molecular identification method for Bletillae Rhizoma, this paper extracted genome DNA from Bletillae Rhizoma and its adulterants. The sequences of rDNA ITS2 were sequenced after amplifying. Then multiple alignments of ITS2 were constructed phylogenetic tree with Neighbor Joining by MEGA 5. 1 and found out SNPs loci. The result showed that rDNA ITS2 region could identify Bletillae Rhizoma and its adulterants. There existed the SNPs loci, which could identify Bletilla striata and B. ochracea. Furthermore, we designed specific primers against the SNPs loci of B. striata and B. ochracea, then screened primers and optimized the PCR amplification conditions. Finally, the DNA of B. striata and B. ochracea were specifically amplified by BJ59-412F, BJ59-412R and HHBJ-225R. The length of amplification products were respectively about 350 bp and 520 bp that were effectively identified of B. striata and B. ochracea. While, the adulterants of Bletillae Rhizoma were no-reaction occurring. To sum up, the amplification conditions of the primers can identify B. striata, B. ochracea and their adulterants successfully at the same time. This method was easy, time-saving, and reliable, which can be used as a rapid method for molecular identification of Bletillae Rhizoma.
Base Sequence ; DNA Primers ; genetics ; DNA, Intergenic ; genetics ; DNA, Plant ; genetics ; Drug Contamination ; prevention & control ; Molecular Sequence Data ; Orchidaceae ; classification ; genetics ; Phylogeny ; Polymorphism, Single Nucleotide ; Rhizome ; classification ; genetics

In order to investigate the mechanisms of phenylhexyl isothiocyanate (PHI) inhibiting the proliferation of multiple myeloma cell RPMI8226 in vitro, the RPMI8226 cells were co-cultured with PHI of various concentrations. The inhibition of proliferation was measured by MTT test and the cell apoptosis was assayed by DAPI staining. The changes of Notch1, Jagged2, BCL-2 and p-Akt proteins in the PHI-treated cells were detected by Western blot. The results showed that PHI inhibited RPMI8226 cell proliferation in certain concentration range and induced their apoptosis. The inhibiting effect caused by PHI showed a concentration-and time-dependent manner. The PHI decreased expressions of Notch1 and Jagged2 proteins in a concentration-and time-dependent manners, the levels of BCL-2 and p-Akt declined at the same time. It is concluded that PHI can inhibit proliferation of RPMI8226 cells, and induce their apoptosis. The cell apoptosis is associated with the inhibition of Notch signaling and downstream targets BCL-2 and p-Akt proteins of RPMI8226 cells, PHI may be a new Notch signaling inhibitor and a promising therapeutic drug for multiple myeloma.
Cell Line, Tumor ; Humans ; Intercellular Signaling Peptides and Proteins ; metabolism ; Isothiocyanates ; pharmacology ; Jagged-2 Protein ; Membrane Proteins ; metabolism ; Multiple Myeloma ; metabolism ; Proto-Oncogene Proteins c-akt ; metabolism ; Proto-Oncogene Proteins c-bcl-2 ; metabolism ; Receptor, Notch1 ; metabolism ; Signal Transduction ; drug effects

Objective To prepare a capillary electrophoresis sieving medium and apply it in GA118-16A genetic analyzer. Methods The white solid polyacrylamide (LPA) was prepared by polymerization and lyophilized. Through the swelling of the sol buffer, the sieving medium was obtained. The sieving medium was evaluated by 1) characterizing the parameters, including molecular weight, structure and viscosity, 2) applying in the GA118-16A genetic analyzer, including the spatial calibration, the spectral calibration and the STR analysis.. Results The prepared sieving medium Mw 1.8 x 105Da, Mn 1.2 x 105 Da, is of correct structure and high purity. The polydispersity was 1.5The spatial calibration and spectral calibration files can be established successfully in GA118-16A genetic analyzer, and the sieving medium can effectively separate the DNA fragments with 1bp difference. The STR profile is of sharp peaks, no impurity peaks, no tail, and no peak loss. Conclusion The sieving medium prepared by the method can be applied to domestic genetic analyzer such as GA118-16A.

BACKGROUNDCorneal neovascular leakage can lead to edema and secondary scarring. Previous studies have shown that pericytes play a key role in maturation of angiogenesis. The present studies investigate the relationship between vascular permeability and pericyte coverage of endothelial cells in rat corneal neovascular induced by alkali burns.

METHODSCorneal neovascular vessels induced by alkali burns was performed in Sprague-Dawley rats. Corneas were excised on 1, 2, 3, 5, 7 and 10 days after cauterization. The vascular permeability rate was measured by the Evans blue method. The microvessel pericyte coverage index (MPI) was applied to quantify the pericyte coverage through double immunofluorescent staining of frozen sections of corneas with CD31 as the endothelial and alpha-smooth muscle actin (alpha-SMA) as the pericyte markers. The correlation between permeability rate and MPI was analyzed. Pericyte coverage was confirmed ultrastructually using transmission electron microscopy.

RESULTSThe vascular permeability rate was (1.14 +/- 0.17), (0.24 +/- 0.08), (0.29 +/- 0.16), (0.14 +/- 0.10), (0.09 +/- 0.06) and (0.05 +/- 0.04) microg x ml(-1) x mm(-2) respectively on 1, 2, 3, 5, 7 and 10 days after cauterization. The MPI was 0, 16.07%, 11.95%, 43.84%, 73.97% and 86.21% respectively at the above mentioned time points. The correlation coefficient between MPI and the permeability rate was -0.943 (P = 0.005).

CONCLUSIONSPericyte recruitment was significantly correlated with the permeability of corneal neovascularization induced by alkali burns in rats. Therapeutic strategies aiming at anti-leakage should be most effective if they promote pericytes proliferation in the course of corneal neovascularization.

Alkalies ; Animals ; Burns, Chemical ; physiopathology ; Capillary Permeability ; Cell Movement ; Cornea ; blood supply ; ultrastructure ; Corneal Neovascularization ; physiopathology ; Eye Burns ; chemically induced ; physiopathology ; Female ; Fluorescent Antibody Technique ; Pericytes ; physiology ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo investigate the epidemiological changes of erectile dysfunction (ED) patients in the past five years.

METHODSIn 2003 and 2008, we conducted two questionnaire investigations on the epidemiological changes of ED outpatients in 11 Chinese cities in such aspects as age, disease course, ED severity, smoking and drinking habits, accompanying hypertension, diabetes and coronary heart disease (CHD), and sexual intercourse satisfaction.

RESULTSAccording to the valid copies of the questionnaire collected (808 in 2003 and 858 in 2008), the age pattern of the ED patients hardly changed in the past five years, over 60% aged 30 - 50 years. Compared with the results obtained in 2003, the second investigation showed obvious increases in the following numbers of the ED patients: by 13% in those with longer disease courses (5 - 10 yr), from 24.1 to 42.9% in those with moderate ED, from 20.4 to 29.9% in those with severe ED, by at least 10% in those with smoking and drinking habits, from 11.5 to 16.2% in those with hypertension, from 9.4 to 13.5% in those with diabetes, and from 57.6 to 73.3% in those without sexual satisfaction, while the number of those with CHD did not change significantly.

CONCLUSIONIncreased unhealthy living habits and erectile function impairing diseases have added to the incidence and severity of ED. There is still much work to be done in the prevention and early treatment of ED.

Adult ; China ; epidemiology ; Erectile Dysfunction ; epidemiology ; Humans ; Incidence ; Male ; Middle Aged ; Outpatients ; Penile Erection ; Surveys and Questionnaires ; Urban Population

OBJECTIVETo detect the expression of recombinant plasmid PcDNA3.1-sTNFRI in vitro and evaluate the bioactivity of expressed sTNFRI.

METHODSCHO cells were transfected with recombinant plasmid PcDNA3.1-sTNFRI by liposome. sTNFRI in cell culture supernatant was detected by ELISA and sTNFRI blockage of TNF-alpha cytotoxicity in L929 cells evaluated by MTT assay.

RESULTSThe expression of sTNFRI in transfected cell culture supernatant was higher than control groups (P < 0.001). The expressed sTNFRI could significantly neutralize TNF-alpha cytotoxicity in L929 cells.

CONCLUSIONSThese results showed that the recombinant plasmid PcDNA3.1-sTNFRI can be expressed in mammalian cells and the recombinant sTNFRI has biological function.

Animals ; Antigens, Surface ; biosynthesis ; CHO Cells ; Cloning, Molecular ; Cricetinae ; DNA, Complementary ; biosynthesis ; genetics ; Eukaryotic Cells ; metabolism ; Genetic Vectors ; drug effects ; Humans ; Plasmids ; biosynthesis ; genetics ; Receptors, Tumor Necrosis Factor, Type I ; biosynthesis ; genetics ; Recombinant Fusion Proteins ; biosynthesis ; genetics ; pharmacology ; Transfection

BACKGROUNDIodine staining during endoscopy has been successfully used to detect early carcinomatous and precancerous lesions in the esophagus, cervix, and oral cavity. The objective of this study was to determine the diagnostic accuracy of fiberoptic ductoscopy (FDS) plus in vivo iodine staining for intraductal proliferative lesions of the breast.

METHODSWe performed periodic acid-Schiff (PAS) and in vitro iodine staining on 52 and 64 specimens of benign mammary hyperplasia, respectively, and 57 and 53 specimens of ductal carcinoma in situ (DCIS), respectively. Next, FDS was performed on 177 recurrent nipple discharge patients who were randomly divided into two groups. One group was iodine-staining group in which 92 patients were randomly selected to undergo iodine staining during FDS, and the remaining 85 were assigned to the control group. Biopsy specimens of suspicious lesions were obtained and subjected to histopathological examination.

RESULTSFollowing PAS staining, benign mammary hyperplasia lesions were positively stained, while negligible PAS positivity was observed in the DCIS lesions (P < 0.05). Following in vitro iodine staining, benign mammary hyperplasia specimens appeared dark brown, whereas DCIS samples appeared significantly lighter or unstained. Compared with the pathological examination results, FDS with iodine staining showed an agreement rate in the diagnosis of ductal intraepithelial neoplasia (DIN), sensitivity, specificity, positive likelihood ratio, negative likelihood ratio, and Youden index of 97.82%, 98.83%, 83.33%, 5.93, 0.014, and 0.8216, respectively; the corresponding values for FDS without iodine staining were 88.24%, 89.16%, 50.00%, 1.78, 0.217, and 0.3916, respectively.

CONCLUSIONFDS with iodine staining was superior to conventional FDS for the diagnosis of DIN and is valuable for breast cancer prevention.

Adult ; Aged ; Breast ; pathology ; Breast Neoplasms ; diagnosis ; pathology ; Carcinoma, Ductal, Breast ; diagnosis ; pathology ; Carcinoma, Intraductal, Noninfiltrating ; diagnosis ; pathology ; Female ; Fiber Optic Technology ; Humans ; Hyperplasia ; Iodine ; Middle Aged ; Periodic Acid-Schiff Reaction ; Staining and Labeling

AIMTo study the effects of hepatocyte growth factor (HGF) and vascular endothelial growth factor (VEGF) on proliferation and migration of bovine coronary artery endothelial cells (BCAEC) in vitro.

METHODSBCAECs were isolated and cultured in vitro, and divided into control group, VEGF group and HGF group. BCACEs proliferation were measured using MTT, and their migration was observed using reverse microscope.

RESULTSThe OD value of control, VEGF and HGF group were 0.22 +/- 0.01, 0.40 +/- 0.14, 0.44 +/- 0.15 respectively. The proliferation ratio of BCAECs in VEGF and HGF group was 81.8% +/- 16.9%, 100.0% +/- 21.1% respectively. There was no migration in control group, but significant migration in VEGF and HGF group.

CONCLUSIONBoth VEGF and HGF can promote proliferation and migration of BCAECs, the effect of HGF is stronger than VEGF.

Animals ; Cattle ; Cell Division ; drug effects ; Cell Movement ; drug effects ; Cell Proliferation ; drug effects ; Cells, Cultured ; Coronary Vessels ; cytology ; Culture Media, Conditioned ; Endothelial Cells ; drug effects ; Endothelium, Vascular ; cytology ; Hepatocyte Growth Factor ; pharmacology ; Vascular Endothelial Growth Factor A ; pharmacology

OBJECTIVETo investigate the changes of proportion and suppression function of CD-4+ CD-25+ regulatory T cells in the peripheral blood of patients with aggressive periodontitis.

METHODSFlow cytometric analysis was used to detect the frequency of CD-4+ CD-25+ regulatory T cells in the peripheral blood of 16 patients with generalized aggressive periodontitis and 17 patients with chronic periodontitis, as well as 17 periodontal healthy controls. Furthermore, CD-4+ CD-25+ regulatory T cells and CD-4+ CD-25- T cells were separated from peripheral blood of each enrolling subject using magnetic cell sorting technology. The direct suppression effect of CD-4+ CD-25+ regulatory T cells on CD-4+ CD-25- T lymphocytes proliferation was performed by the mixed lymphocytes reaction and measured by 3H-thymidine radioactive assay.

RESULTSThe patients with generalized aggressive periodontitis had a lower frequency of CD4+ CD-25+ regulatory T cells (9.71 +/- 4.01)% in the peripheral blood than periodontal healthy controls [(14.72 +/- 3.51)%] and chronic periodontitis patients [(17.01 +/- 5.16 )%], P < 0.05. A significant decrease was found in the suppression function of CD-4+ CD-25+ regulatory T cells from peripheral blood of patients with generalized aggressive periodontitis when co-cultured with CD-4+ CD-25- T lymphocytes in the proportion of 2 : 1, 1 : 1 and 1 : 2 as compared with chronic periodontitis patients and periodontal healthy controls (P < 0.05).

CONCLUSIONSDiminished numbers and decreased suppression function of CD-4+ CD-25+ regulatory T cells were found in patients with generalized aggressive periodontitis.

Adult ; Aggressive Periodontitis ; blood ; Case-Control Studies ; Female ; Flow Cytometry ; Humans ; Male ; Middle Aged ; T-Lymphocytes, Regulatory ; cytology

OBJECTIVETo gain a deeper insight into the benefits of oral tadalafil for patients with erectile dysfunction (ED).

METHODSFrom January 2008 to June 2009, we conducted a nationwide survey on the quality of male erectile function among the outpatients under the direction of the Chinese Association of Andrology. A total of 205 ED patients were prescribed oral tadalafil and accomplished a questionnaire investigation after 4 weeks of medication. We compared various parameters of the patients before and after the treatment.

RESULTSFour weeks of oral tadalafil medication achieved a total rate of effectiveness of 85.9% (176/205). The proportion of those with moderate to severe ED was decreased from 67.8% (139/205) before medication to 16.6% (34/205) after it. Those who enjoyed sexual life were increased from 21.5% (44/205) before medication to 84.9% (174/205) after it. Only 1.0% (2/205) of the patients could achieve grade 4 penile hardness before the treatment, as compared with 60.5% (124/205) after it. And the frequency of sexual intercourse was significantly increased, over 4 times in 90.2% (159/205) of the patients.

CONCLUSIONOral tadalafil, with its sure effectiveness on ED, can bring great benefits to the sexual life of ED patients.

Adult ; Carbolines ; therapeutic use ; Erectile Dysfunction ; drug therapy ; Humans ; Male ; Middle Aged ; Penile Erection ; Phosphodiesterase Inhibitors ; therapeutic use ; Surveys and Questionnaires ; Tadalafil ; Treatment Outcome