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Objective:To explore the effects of long noncoding RNA adenosine diphosphate-dependent glucokinase antisense RNA 1(ADPGK-AS1) on the proliferation, migration and invasion of human retinoblastoma (RB) Y-79 cells and its regulatory effect on microRNA-623 (miR-623).Methods:The peritumoral tissue and RB specimens were collected from 39 eyes of 39 patients with RB during surgery in The First Affiliated Hospital of Zhengzhou University and Zhumadian Central Hospital from February 2017 to November 2018.Real-time fluorescence quantitative PCR was employed to detect the expression of ADPGK-AS1 and miR-623 in the specimens.Human RB line Y-79 cells were cultured in vitro and divided into small interfering RNA-normal control (siRNA-NC) group, siRNA-ADPGK-AS1 group, microRNA (miR)-NC group, miR-623 group, siRNA-ADPGK-AS1+ anti-miR-NC group and siRNA-ADPGK-AS1+ anti-miR-623 group.The cell proliferation rate was detected by MTT method.Transwell cell experiment was performed to detect the number of migrating and invading cells.The dual luciferase reporter experiment was used to evaluate the targeting relationship between ADPGK-AS1 and miR-623.The expression of Ki-67, matrix metalloproteinases (MMP)-2, and MMP-9 in the cells was detected by Western blot assay.Written informed consent was obtained from each patient prior to any medical examination and treatment.This study protocol adhered to the Declaration of Helsinki.The use of the human specimens was approved by an Ethics Committee of The First Affiliated Hospital of Zhengzhou University (No.2017-KY-73). Results:Compared with the peritumoral tissue, the relative expression level of ADPGK-AS1 in the RB tissue was significantly increased, and the relative expression level of miR-623 was significantly reduced ( t=40.522, 48.497; both at P<0.01). Compared with the siRNA-NC group, both the relative expression level of Ki-67 protein and the proliferation A value of RB Y-79 cells were significantly reduced in the siRNA-ADPGK-AS1 group ( t=26.833, 18.522; both at P<0.01). The relative expression levels of MMP-2 and MMP-9 proteins in the siRNA-ADPGK-AS1 group were significantly lower than those in the siRNA-NC group ( t=22.123, 26.183; both at P<0.01). The number of migrating and invading cells in the siRNA-ADPGK-AS1 group was significantly less than that in the siRNA-NC group ( t=12.385, 19.201; both at P<0.01). The dual luciferase report experiment confirmed that ADPGK-AS1 targeted miR-623.The protein expression levels of the Ki-67, MMP-2 and MMP-9 in the miR-623 group were significantly lower than those in the miR-NC group ( t=22.137, 22.200, 21.094; all at P<0.01). Compared with the miR-NC group, the proliferation A value of Y-79 cells in the miR-623 group was significantly lower, and the number of migrating and invadoing cells was significantly less ( t=16.398, 11.400, 17.846; all at P<0.01). The relative expressions levels of Ki-67, MMP-2 and MMP-9 proteins in the siRNA-ADPGK-AS1+ anti-miR-623 group were significantly higher than those in the siRNA-ADPGK-AS1+ anti-miR-NC group ( t=20.795, 17.493, 23.479; all at P<0.01). Compared with the siRNA-ADPGK-AS1+ anti-miR-NC group, the proliferation A value of Y-79 cells in the siRNA-ADPGK-AS1+ anti-miR-623 group was significantly increased ( t=15.600, P<0.01), and the number of migrating and invading cells was obviously elevated ( t=14.495, 17.855; both at P<0.01). Conclusions:Knockdown of ADPGK-AS1 gene can inhibit the proliferation, migration and invasion of Y-79 cells by up-regulating the expression of miR-623.
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Objective:To investigate the epidemiological and molecular biological characteristics of heterogeneous vancomycin-intermediate Staphylococcus aureus (hVISA) in blood culture. Methods:hVISA was detected using Mueller-Hinton agar containing 5 μg/ml of teicoplanin (MHA5T) and Populats profiles/area under the curve (PAP/AUC). Staphylococcal cassette chromosome mec ( SCCmec), Staphylococcus aureus protein A ( spa) and accessory gene regulator ( agr) typing and multilocus-sequence typing (MLST) were analyzed using PCR. Difference in autolysis between hVISA and vancomycin-sensitive Staphylococcus aureus (VSSA) isolates were evaluated with Triton X-100-inducd autolysis. Expression of vraR, mgrA, icaA, icaR, pbp4 and agr genes in hVISA and VSSA strains were detected by real-time PCR. Results:The positive detection rate of methicillin-resistant Staphylococcus aureus (MRSA) in blood culture was 39.5% (136/344) in our hospital. Among the MRSA strains, there were 31 strains of hVISA (22.8%). The minimum inhibitory concentrations (MIC) of vancomycin were mainly 1.5 μg/ml (54.8%) and 2 μg/ml(25.8%)against hVISA isolates, and 0.5 μg/ml (46.7%) and 0.75 μg/ml (39.0%) against VSSA isolates. The predominant clone of hVISA was ST239- SCCmecⅢ-t030- agrⅠ accounting for 71.0% (22/31). The autolysis of hVISA isolates decreased significantly as compared with that of VSSA isolates ( χ2=13.583, P=0.032). Compared with VSSA strains, the expression of vraR, mgrA and icaA genes in hVISA strains increased by 1.58, 1.53 and 1.06 times ( P<0.01), while the expression of icaR, agr and pbp4 genes decreased by 0.85, 0.61 and 1.03 times ( P<0.05). Conclusions:The prevalence rate of hVISA in our hospital reached 22.8% and the main epidemic clone was ST239- SCCmecⅢ-t030- agrⅠ, which should be paid great attention to clinically. Rational use of antibiotics, strengthening the prevention and control of nosocomial infection, and avoiding the spread of hVISA strains and the emergence of VISA and VRSA (vancomycin-resistance Staphylococcus aureus) were also necessary.
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Objective To explore the inhibitory effect of 24 pseudomonas aeruginosa(PA) on pathogenic fungi ,such as candida albicans ,candida tropicalis ,candida glabrata ,candida parapsilosis ,candida krusei ,mucous spore bacterium (MSB) etc .Methods 24 PA isolates were collected from clinical specimens and identified by Gram′s stain ,oxidase production and the API 20NE system(bi-oMerieux ,France) .Cross-streaking method and sterilizing filter paper-disk method and co-cultured method were applied to observe the inhibitory effect of PA .Sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE) analyzed the difference of bacte-rial proteins of PA .Results The results showed that some strains of 24 PA had strong inhibitory effect against pathogenic fungi , some strain had partial effect and others had no effect .Co-cultured test showed that PA could inhibit the growth of fungal hyphae . SDS-PAGE displayed the significant difference in secretive proteins between the PA strains which had strong effect and no effect . Conclusion PA have inhibitory effect upon common pathogenic fungi and and this might be related to inhibit fungal hyphae forma-tion ,various protein secretion and inhibit the growth of fungi .
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Objective To investigate antibiotic resistance and resistant trend of Streptococcus pneumonia. Methods To investigate 753 Streptococcus pneumoniae isolated from Tongji Hospital in recent 10 years from January 1st 2000 to December 31st 2009, most of them were from respiratory tract specimens,followed by blood and cerebrospinal fluid. The MIC to penicillin & cefatriaxone were determined by E-test,and other antimicrobial susceptibility were tested by Kirby-Bauer method. Results For non-cerebrospinal fluid specimen, the total rate of PNSSP was 23.8%( 93/392 ), it was significant different between the rate of PNSSP from children ( 26. 4%, 47/178 ) and adults ( 16. 8%, 36/214, χ2 = 7. 642, P ＜ 0. 01 ). All of 10 strains isolated from cerebrospinal fluid were PRSP. Most isolates were high-susceptive to moxifloxacin and levofloxacin, and the rate of susceptibility were 96. 9% ( 720/743 ) and 90. 5% ( 672/743 )respectively. None of Streptococcus pneumonia was resistant to vancomycin and meropenam. The resistant rate of most tested antibiotics increased in different degree year by year, especially penicillin, erythromycin and clindamycin. The rate of PNSSP was only 19%( 19/99 )in 2006 ,but in 2009 the rate increased to 30%( 35/114 ). The susceptibility rate of erythromycin was 22% ( 28/125 )in 2000, but only 3% ( 3/114 )in 2009 ;and the susceptibility rate of clindamycin decreased from 40% ( 13/32 ) in 2004 to 4% (5/114) in 2009. Conclusions From 2000 to 2009, Streptococcus pneunoniae was more likely resistant to penicillin,erythromycin and clindamycin year by year, especially those isolates recovered from children. It was suggested that antibiotics should be chosen to use according to antimicrobial susceptibility test results.