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OBJECTIVE@#To explore potential therapeutic targets other than androgen-deprivation treatment for prostate cancer by screening the proteins induced by androgen at palmitoylation modification level in LNCaP cells.@*METHODS@#The LNCaP cells were treated with androgen (Methyltrienolone, R1881, 5 nmol/L) or dimethyl sulfoxide (DMSO) for 24 h, and then labeled with alkynyl palmitic acid Alk-C16 (100 μmol/L). After that, the cells were collected, lysed, the total protein was extracted, agarose beads labeled with azide (1 mmol/L) were added, and the click-chemistry reaction was carried out at room temperature for 1 h. The covalent bond formed by click-chemistry reaction of azide and alkynyl group was used to enrich the palmitoylated proteins on agarose beads. Label-free quantitation (LFQ) was used to compare the protein palmitoylation level of R1881 treated and untreated cells to screen the proteins induced by androgen at palmitoylation modification level.@*RESULTS@#In this experiment, 907 potential palmitoylated proteins (mascot score>2, P<0.05) were identified, among which 430 proteins had LFQ values not zero at least twice. Among the 430 proteins, the palmitoylation levels of 92 candidates were increased by androgen treatment, and their LFQ values were significantly upregulated (>1.5-fold, P<0.05) in ≥2 samples of androgen-treated vs. untreated LNCaP cells. We also used the software of cytoscape to classify the 92 proteins, and found that the known functional proteins of them could be divided into three categories: metabolism related, protein folding related and translation initiation related. Among them, metabolism related proteins included lipid metabolism (6), glucose metabolism (7) and respiratory electron transport chain (8), and a small amount of amino acid metabolism (2) and other metabolism related proteins (2). Notably, the ratio of LFQ of cytochrome b-c1 complex subunit 2 (UQCRC2) was significantly (>3-fold, P<0.05) higher in androgen-treated cells compared with untreated cells, indicating that the palmitoylation level of UQCRC2 was enhanced by androgen most significantly than that of others. The second was long-chain acyl CoA dehydrogenase (ACADVL) related to lipid metabolism and glucose 6-phosphate dehydrogenase (PGD) related to glucose metabolism, but the LFQ ratio of them was less than 3-fold.@*CONCLUSION@#The research on palmitoylation mechanism of metabolism, especially the proteins related to respiratory electron transport chain, will provide a new guidance for the diagnosis and treatment of prostate cancer and the development of targeted drugs.
Subject(s)
Humans , Male , Androgen Antagonists , Androgens , Lipoylation , Prostatic NeoplasmsABSTRACT
@#AIM:To study the clinical efficacy of phase Ⅱ minimally invasive vitreous surgery in different stages in treatment of open ocular trauma with retinal detachment.<p>METHODS: A retrospective analysis was performed on 41 patients(41 eyes)with open eye trauma combined with retinal detachment from December 2013 to June 2018 in the Ophthalmology Department of Xiaolan People's Hospital of Zhongshan. According to the opportunity of phase Ⅱ vitrectomy, 41 eyes were divided into two groups: 24 eyes in the early group(6d after injury)and 17 eyes in the conventional group(7-14d after injury). Postoperative follow-up with 6mo as the time point, the retinal reattachment rate, TPVR incidence, visual acuity and complications were compared between the two groups.<p>RESULTS: The retinal reduction rate was 92% in the early group and 76% in the conventional group, and the difference was not statistically significant, there was no statistical difference(<i>P</i>=0.692)in retinal reduction rate between the two groups. The incidence of TPVR in the early group was lower than that in the conventional group(<i>P</i>=0.014). The improvement of postoperative visual acuity in early group was better than the conventional group(<i>U</i>=119.5,<i> P</i>=0.0018). There was no significant difference in complications between the two groups.<p>CONCLUSION: Open ocular trauma patients with retinal detachment have better prognosis after phase Ⅱ vitrectomy within 6d after injury than 7-14d.
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[Objective]To evaluate the practicality of non-mydriatic digital fundus camera in the remote screen of diabetic retinopathy for community residents.[Methods]Ninety-two patients(184 eyes)with type 2 diabetes mellitus have been taken 1-field and 5-field non-mydriatic fundus photography and examination in mydriatic fundus by pre-placed-mirror ophthalmoscopy by a ophthalmologist,the results were sent to the hospital with a computer programs. A specialist evaluates the consistency of detectable rate of diabetic retinophathy(DR)among the 3 methods.The time of tak-ing 1-field and 5-field non-mydriatic fundus photography is compared.[Results]All the three methods show good consis-tency in detectable rate of DR compare with each other,the κ value is 0.89 for 1-field non-mydriatic fundus photography and examination in mydriatic fundus by preplaced-mirror ophthalmoscopy,0.95 for 1-field and 5-field non-mydriatic fundus photography and 0.95 for 5-field non-mydriatic fundus photography and examination in mydriatic fundus by pre-placed-mirror ophthalmoscopy,respectively. The 1-field non-mydriatic fundus photography spent less time compares with 5-field non-mydriatic fundus photography(55.4±5.8 vs 405.9±68.5 s,P<0.01).[Conclusion]The remote screen for diabetic retinopathy in community based on non-mydriatic digital fundus camera is worth promoting,we suggest 1-field non-mydriatic fundus photography as a screen method.
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The expression of fibroblast growth factor 9 (FGF9) recombinant fusion protein in Carthamus tinctorius was used to identify its effect on hair regrowth and wound repair system in mice, providing a basis for C. tinctorius as a plant bioreactor, and establishing a foundation for commercial applications of FGF9 fusion protein in hair regrowth and wound repair. The identified pOTBar-oleosin-rhFGF9 plasmid was transformed into Agrobacterium tumefaciens EHA105 by freeze-thaw method, and the oleosin-rhFGF9 gene was transformed into safflower leaves by A. tumefaciens mediated method. Transgenic safflower seedlings were then obtained by tissue culture. After basta screening, transgenic T₃ safflower seeds were obtained by grafting method, PCR verification and propagation. The expression of oleosin-rhFGF9 was detected by Western blot, and the content of oleosin-rhFGF9 fusion protein was 0.09% by using ELISA quantitative method. It was observed that 60 μg·L⁻¹ transgenic safflower oil had better effect on promoting NIH/3T3 cells proliferation in a certain dose-dependent manner. Sixty C57BL/6 mice were used to establish alopecia model and wound model respectively, and then were randomly divided into control group (treated with PBS or saline), negative group (treated with wild type safflower seed oil bodies, 60 g·L⁻¹), positive group (treated with FGF9, 0.054 g·L⁻¹), low dose group (treated with transgenic safflower oil bodies, 10 g·L⁻¹) and high dose group (treated with transgenic safflower oil bodies, 60 g·L⁻¹). The skin of all above-mentioned mice models were coated with soft adhesive manner every other day, 100 μL/time. After 15 days, the mice skin was cut and embedded for histological analysis. The hair regrowth experimental results showed that the hair of mice grew well, and the mice in high dose group had bushy hair, with significant effect on regeneration hair number as compared with the positive group. The healing was obvious in wound experiment, with significant healing effect in positive group, high dose group and low dose group as compared to blank control group. Furthermore, high dose group remarkably showed a better and higher healing effect than the positive group at day 5. Oleosin-rhFGF9 was successfully transformed into safflower, and T₃ transgenic safflower oil bodies expressed oleosin-rhFGF9 fusion protein were obtained, with the role of promoting hair regeneration and wound repair in mice.
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The aim of this study is to investigate antibiotic resistance and molecular epidemiology characteristics of methicillin-resistant Staphylococcus aureus (MRSA) in Shenzhen area.We collected 428 Staphylococcus aureus isolates from eight hospitals in Shenzhen in 2012.According to the results of minimum inhibitory concentration (MIC) to cefoxitin,26.2％ of Staphylococcus aureus isolates (112/428) were identified as MRSA.The MIC of 10 antimicrobial agents was determined by agar dilution method.Panton-Valentine leucocidin(PVL) was detected by polymerase chain reaction(PCR).Positive strains of PVL were detected by multilocus sequence typing(MLST).Among the 112 strains,the resistance rates to trimethoprim-sulfamethoxazole,moxifloxacin,ciprofloxacin,gentamicin,erythromycin,clindamycin and tetracycline were 4.46％,12.50％,16.96％,19.64％,46.42％,25.00％ and 26.79％ respectively.No isolates resistant to vancomycin,linezolid and teicoplanin were found.Among the 112 strains,there were 13 (11.61％) strains carried PVL gene.There were no significant differences in the resistance rates of PVL positive strains and negative strains to 10 kinds of antimicrobial agents.Among the 13 strains carried PVL gene,7 kinds of old sequences and 1 kind of new sequence type were found by MLST.ST338 and ST25 were the most common type.All the data indicate that surveillance of MRSA in Shenzhen has a distinct genetic background from other regions.
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To compare the main nucleosides in Cordyceps genus herbs (C. sinensis, C. millitaris, Hirsutella sinensis and C. sobolifera), an HPLC method for simultaneous determination of uridine, inosine, guanosine, adenosine and cordycepine in Cordyceps genus herbs was developed. The sample was extracted with 0.5% phosphoric acid solution to prepare test solution. The separation was performed on a Zorbax SB-Aq (4.6 mm×150 mm, 5 μm) column with gradient elution by 0.04 mol•L⁻¹ potassium dihydrogen phosphate solution and acetonitrile, column temperature 30 ℃,flow rate 0.8 mL•min⁻¹,and detection wavelength 260 nm. The content of nucleosides in four Cordyceps genus herbs was evaluated by fingerprint analysis and hierarchical cluster analysis (HCA). The calibration curves of five nucleosides showed good linear regression (r>0.99) and the average recoveries were between 95.0% and 105.0%. The contents of the five nucleosides in the four Cordyceps genus herbs were different and could be obviously distinguished by HCA. The fingerprint analysis result showed that the similarity between C. sinensis and the others was less than 0.9. The method was accurate and reliable, which can be used for quality control of Cordyceps genus herbs.
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Objective: To develop a rapid high-performance liquid chromatography coupled with diode array detection (HPLC-DAD) method for the simultaneous determination of six polar compounds in Ophiocordyceps sinensis. Methods: A poroshell SB Aq column (50 mm × 4.6 mm, 2.7 μm) and gradient elution were used; The detection wavelength of compounds was set at 260 nm. The chromatographic peaks of the six investigated compounds in sample were identified by comparing their retention times with reference compounds. Results: All calibration curves showed good linearity (r > 0.999) within the tested ranges. The intra- and inter-day precisions of the six analytes were less than 0.8% and 2.1%, respectively, and the recoveries of the six analytes were between 95% and 103%. The validated method was successfully applied to the determination of six polar compounds in O. sinensis samples. Conclusion: The poroshell SB Aq column is suitable for the rapid analysis of polar components in Chinese materia medica on conventional HPLC system and the developed HPLC method is also helpful to the quality control of O. sinensis. © 2014 Tianjin Press of Chinese Herbal Medicines.
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<p><b>OBJECTIVE</b>To explore the relationship between the polymorphisms of Toll-like receptor 2 (TLR2) and TLR9 and the susceptibility to gastric cancer.</p><p><b>METHODS</b>A population-based case-control study was conducted at Linqu county, Shandong province, China, including a total of 248 cases of gastric cancer. Another total of 496 age and sex-matched controls were randomly selected from the same cohorts. TLR2 rs3804099 and TLR9 rs187084 were detected by polymerase chain reaction-restriction fragment length polymorphism method. Odds ratios (ORs) and 95% confidence interval (CI) were computed from logistic regression models after adjusting for age, sex, Helicobacter pylori (H. pylori) infection and smoking status.</p><p><b>RESULTS</b>The frequencies of TT, TC and CC genotype on TLR2 rs3804099 in control group were 43.5% (216/496), 46.6% (231/496) and 9.9% (49/496), respectively; whereas those in case group were 53.2% (132/248), 39.9% (99/248) and 6.9% (17/248), respectively. Significant differences in the frequencies of TLR2 rs3804099 were found between case and control groups (χ(2) = 6.665, P = 0.036). It was found that compared with the TT genotype, TC + CC genotype carriers obviously less susceptible to gastric cancer (OR = 0.68, 95%CI: 0.50 - 0.93). Joint effects analysis indicated that the TLR2 rs3804099 TT genotype carriers and H.pylori infectors had higher susceptibility to gastric cancer(OR = 3.42, 95%CI: 2.16 - 5.42), compared with TC + CC genotype carriers and non-H.pylori infection group. The frequencies of TT, TC and CC genotype on TLR9 rs187084 in control group were 33.3% (165/496), 49.0% (243/496) and 17.7% (88/496), respectively; whereas those in case group were 35.9% (89/248), 50.0% (124/248) and 14.1% (35/248), respectively. No significant association with gastric cancer was observed for TLR9 rs187084 polymorphism (χ(2) = 1.684, P = 0.431).</p><p><b>CONCLUSION</b>Our findings indicate that TLR2 rs3804099 is closely associated with susceptibility to gastric cancer.</p>
Subject(s)
Female , Humans , Male , Middle Aged , Case-Control Studies , China , Epidemiology , Genetic Predisposition to Disease , Polymorphism, Genetic , Stomach Neoplasms , Epidemiology , Genetics , Toll-Like Receptor 2 , Genetics , Toll-Like Receptor 9 , GeneticsABSTRACT
<p><b>OBJECTIVE</b>To setup a quantitative assay for detection of cyclooxygenase-2 (COX-2) methylation in human gastric mucosa samples.</p><p><b>METHODS</b>A standard analysis system was established by denaturing high performance liquid chromatography (DHPLC) under the condition of 55 degrees C oven temperature and a linear acetonitrile gradient (4.0/min). While, a total of 10 cases of gastric biopsy samples were detected for methylation status of COX-2.</p><p><b>RESULTS</b>The complete methylated human promyelocytic leukemia cells (HL-60) and unmethylated gastric cancer cell line (MGC803) were used as positive and negative control. The proportion of the methylated copies of COX-2 was calculated according to the peak heights of methylated (M) and unmethylated (U) COX-2 in same PCR amplicon. The formula was Y = 1.0608 x M/(M + U), R(2) = 0.9894. Among 10 biopsy samples, the proportions of methylated copies of COX-2 in 2 cases of dysplasia were higher than superficial gastritis and chronic atrophy gastritis (24.5%, 18.4% vs 7.6%, 9.6%).</p><p><b>CONCLUSION</b>The methylation of COX-2 promoter CpG islands can be detected in human gastric mucosa samples by quantitative DHPLC assay, which could be used in the population-based study of precancerous gastric lesions.</p>
Subject(s)
Humans , Cell Line, Tumor , Chromatography, High Pressure Liquid , Methods , CpG Islands , Cyclooxygenase 2 , Genetics , DNA Methylation , Gastric Mucosa , Metabolism , Pathology , Promoter Regions, Genetic , Stomach Diseases , Genetics , Metabolism , PathologyABSTRACT
<p><b>OBJECTIVE</b>To investigate the association between the heat shock protein 70-2 gene polymorphism and ankylosing spondylitis (AS).</p><p><b>METHODS</b>The polymorphisms of HSP70-2 gene Pst I 1267 site were analysed in 176 Chinese Han AS patients and 127 healthy controls by PCR and restriction fragment length polymorphisms(RFLP) methods.</p><p><b>RESULTS</b>In AS patients HSP70-2 genotypes AA, AG and GG were 46.6%, 46.0% and 7.4% respectively, frequencies of A and G were 69.6%(A) and 30.4%(G). In healthy controls HSP70-2 genotypes AA, AG and GG were 44.1%, 48.8% and 6.9% respectively, frequencies of A and G were 68.5%(A)and 31.5%(G). No significant differences were found in the distribution of HSP70-2 genotypes and allele frequencies between AS patients and controls.</p><p><b>CONCLUSION</b>Our results indicate that the there may be no association of the HSP70-2 gene polymorphism with AS in Chinese Han population.</p>
Subject(s)
Adolescent , Adult , Aged , Child , Female , Humans , Male , Middle Aged , Asian People , Genetics , China , Ethnology , Ethnicity , Genetics , Gene Frequency , Genetic Predisposition to Disease , Genotype , HSP70 Heat-Shock Proteins , Genetics , Polymorphism, Single Nucleotide , Spondylitis, Ankylosing , GeneticsABSTRACT
<p><b>OBJECTIVE</b>To investigate the associations of serum gastrin-17 (G-17) concentration with helicobacter pylori (Hp) infection.</p><p><b>METHODS</b>A (13)C-urea breath and ELISA test to determine the Helicobacter pylori status and to detect the serum gastrin concentration was conducted in 242 villagers in Linqu of Shandong Province, a high gastric cancer prevalence area in China.</p><p><b>RESULTS</b>Of 242 subjects, 65 of 111 were found Hp-positive in males (58.56%), compared with 65 of 131 in females (49.62%) (chi(2) = 1.932, P = 0.165). The statistical difference was not observed among different age groups (chi(2) = 4.185, P = 0.123). The average level of G-17 among 242 subjects was (24.43 +/- 25.46) pmol/L and it was statistically higher in females (29.87 +/- 28.18) pmol/L than that in males (18.01 +/- 20.11) pmol/L (Z = -3.618, P < 0.001). However, there was no statistical difference found among age groups (chi(2) = 1.948, P = 0.378). The G-17 level in Hp-negative group (35.50 +/- 30.92) pmol/L was observed significantly higher than in Hp-positive group (14.90 +/- 13.79) pmol/L (Z = 5.368, P = 0.0001).</p><p><b>CONCLUSION</b>The G-17 concentration was found higher in Hp-negative subjects than in Hp-positive subjects, and higher in female than in male, but no difference was found among age groups.</p>