Objective To investigate the effect of alteration of CRELD1 gene expression on the related genes in the endocardial cushion development.Methods Over-expression and silence of CRELD1 gene were realized by the construction of lentiviral vector.Afterwards,the lentiviral vectors were used to infect the human fetal lung fibroblasts (HFL)-I.All of the cells were divided into the following 5 groups:the blank control group,the negative control group of interference,the interference group,the negative control group of over-expression,and the over-expression group.Western blot and real-time fluorescent quantitative polymerase chain reaction were applied to examine the mRNA and protein expression of CRELD1,Sox9,Aggrecan,Scleraxis and Tenascin-C.Results The DNA sequences of 2 recombinant plasmids pLV3-shRNA-CRELD1 and pLV4-CRELD1 matched very well with those which were designed according to the DNA sequence analysis.HFL-I was successfully infected with lentiviral vectors and displayed fluorescent green light under inverted fluorescence microscope.The results of real-time PCR detection and Western blot test were consistent:expressions of Sox9 and Aggrecan in the interference group were significantly higher than those in the negative control group of interference,while the expressions of the 2 genes in the over-expression group were significantly lower than those in the negative control group of over-expression.Expressions of Scleraxis increased in both the interference group and the over-expression group when compared with the negative control groups respectively.Compared to the corresponding negative control groups,Tenascin-C expression decreased markedly in the interference group,whereas it increased significantly in over-expression group.Conclusions CRELD1 gene has negative effect on the expression of the related genes Sox9 and Aggrecan in the endocardial cushion development,whereas it has positive effect on the Tenascin-C expression.It serves as a theoretical framework to illustrate the effect of CRELD1 gene on the atrioventricular septal defect.

Purpose To explore the immunohistochemical ( IHC) expression of ALK antibodies with different clones in anaplastic lym-phoma kinase ( ALK) gene fusion non-small cell lung cancer ( NSCLC) . Methods ALK expression in 60 NSCLCs were detected by IHC including autostainer (D5F3, Ventana+BenchMark) and manual staining using 4 different antibodies of D5F3 (Ventana), D5F3 (Cell Signaling), 1A4/1H7 (OriGene), 5A4 (Abcam), and all cases were verified with ALK FISH. Their expressions with 4 anti-bodies were compared with those by D5F3 (Ventana+BenchMark). Results 32 ALK gene rearrangement NSCLCs and 28 negative cases were identified by FISH and D5F3 (Ventana+BenchMark). The sensitivity of D5F3 (Ventana), D5F3 (Cell Signaling), 1A4/1H7 (OriGene), 5A4 (Abcam) was 93. 8%, 84. 4%, 93. 8%, 56. 3%, and all the speciticity was 100%. The consistency with D5F3 (Ventana+BenchMark) was 96. 7%, 91. 7%, 96. 7% and 76. 7%, respectively. The validity of immunohistochemical staining in surgical resection specimens was better than in small biopsies. Conclusion Effective routine manual immunohistochemistry with high-affinity antibody clone may provide a more economic and widespread pre-screening technique.

OBJECTIVETo investigate the role of nitric oxide in the inhibitory effect of somatic afferent input on the pressor response caused by electrical stimulation of the paraventricular nucleus of the hypothalamus (PVN).

METHODSSD rats anesthetized by urethane were used in the study. Bipolar stainless stimulating electrode was inserted into PVN for electrical stimulation. Multi-barreled micropipettes were used for microinjection of L-NAME or normal saline into the lateral ventricle or amygdala. Deep peroneal nerve (DPN) was stimulated with electrical current pulses of 0.4 mA with duration of 0.5 ms at 4 Hz for 5 min. PVN was stimulated by electrical current pulses of 0.3 mA with duration of 0.5 ms at 80 Hz for 10 sec.

RESULTElectrical stimulation of PVN increased mean arterial pressure. Stimulation of DPN significantly inhibited the pressor response induced by stimulation of PVN (P<0.01), with the inhibitory percentage of 43.27%. Microinjection of L-NAME (0.5 mol/L,10 microl) into the lateral ventricle of brain attenuated the inhibitory effect of DPN. The inhibitory percentage decreased from 47.73% to 12.49% (P<0.05). Microinjection of L-NAME (2 mol/L,100 nl) into amygdala reduced the inhibitory effect of DPN. The inhibitory percentage of stimulating DPN on the pressor response decreased from 50.71% to 25.30% (P<0.05).

CONCLUSIONNitric oxide in the brain and amygdala are involved in the inhibitory effect of somatic afferent input on central pressor response.

Afferent Pathways ; drug effects ; physiology ; Amygdala ; drug effects ; physiology ; Animals ; Blood Pressure ; physiology ; Electric Stimulation ; Enzyme Inhibitors ; pharmacology ; NG-Nitroarginine Methyl Ester ; pharmacology ; Nitric Oxide ; antagonists & inhibitors ; physiology ; Nitric Oxide Synthase ; antagonists & inhibitors ; Paraventricular Hypothalamic Nucleus ; drug effects ; physiology ; Peroneal Nerve ; physiology ; Pressoreceptors ; drug effects ; physiology ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo study the anti-inflammation effect of volatile oil of Centipeda minima and the mechanism of action.

METHODThe animal model was induced by the Car injection into intrapleural of rats, to observe the effect of VOCM on acute inflammation.

RESULTVOCM was able to inhibit the increase of NO, CRP and proinflammatory cytokines such as TNFalpha in the acute inflammation of the rat body.

CONCLUSIONVOCM has a protective effect on acute pleural effusion in rats induced by an intrapleural injection of Car.

Animals ; Asteraceae ; chemistry ; C-Reactive Protein ; metabolism ; Carrageenan ; Dinoprostone ; metabolism ; Interleukin-8 ; blood ; Leukocyte Count ; Male ; Nitric Oxide ; metabolism ; Oils, Volatile ; isolation & purification ; pharmacology ; Plants, Medicinal ; chemistry ; Pleural Effusion ; chemically induced ; metabolism ; pathology ; Rats ; Rats, Sprague-Dawley ; Tumor Necrosis Factor-alpha ; blood

Objective: To compare amplification refractory mutation system(ARMS) and droplet digital PCR (ddPCR) in the detection of epidermal growth factor receptor (EGFR) gene mutations in patients with non-small cell lung cancer (NSCLC), and to investigate the clinical value of ddPCR. Methods: A total of 79 specimens of NSCLC, including 22 cases of cell block, 18 cases of surgical specimens, 12 cases of biopsy specimens and 27 cases of plasma samples, were analyzed for the mutation status of EGFR gene by ARMS and droplet digital PCR method. Results: In 18 cases of surgical specimens and 12 cases of biopsy specimens, the detection results by the two methods were identical with positive rates of 9/18 and 5/12, respectively. In 22 cases of effusion cell blocks, ARMS detected 19-del and L858R of EGFR gene in two cases, in which droplet digital PCR detected 19-del+ T790M mutations in one case and L858R+ T790M mutation in another. L858R mutation was detected by droplet digital PCR in one case but ARMS assay was negative. The remaining 19 cases were consistent by the two methods. In blood samples, the positive rate was 33.3%(9/27) by ARMS and 37.0%(10/27) by droplet digital PCR. Two cases showed L858R and 19-del+ T790M mutation by droplet digital PCR but ARMS assay detected only 19-del. The remaining 25 cases were consistent by the two methods. Conclusion Droplet digital PCR method is more sensitive and accurate than ARMS for the detection of EGFR mutations in pleural fluid and blood samples, can be used in clinical test.

OBJECTIVETo reconfirm the association of KPNB3 with schizophrenia in Chinese population.

METHODSTwo single nucleotide polymorphisms (SNPs), rs2588014 and rs626716 at the KPNB3 locus, were genotyped in 304 Chinese Han family trios consisting of fathers, mothers, and affected offsprings with schizophrenia. These 2 SNPs were detected by PCR-based restriction fragment length polymorphism (RFLP) analysis. The Hardy-Weinberg equilibrium for genotypic distributions was estimated by the goodness-of-fit test. The UNPHASED program was used to perform transmission disequilibrium test (TDT), haplotype analysis, and pair-wise measure of linkage disequilibrium (LD) between these 2 SNPs.

RESULTSThe genotypic distributions of both rs2588014 and rs626716 were in the Hardy-Weinberg equilibrium (P > 0.05). The TDT revealed allelic association with rs626716 (chi2 = 9.31, P = 0.0023) but not with rs2588014 (chi2 = 3.44, P = 0.064). The global P-value was 0.0099 for 100 permutations. The haplotype analysis also showed a disease association (chi2 = 25.97, df = 3, P = 0.0000097).

CONCLUSIONThe present study provides further evidence in support of the KPNB3 association with schizophrenia in Chinese population.

Adult ; China ; epidemiology ; Female ; Genetic Predisposition to Disease ; Genotype ; Humans ; Male ; Polymorphism, Single Nucleotide ; Schizophrenia ; epidemiology ; genetics ; beta Karyopherins ; genetics

Objective To investigate the feasibihty and superiority of transthoracic echocardiography(TTE)guided transcatheter occlusion for patent ductus arteriosus (PDA).Methods Totally 58 patients with simple funnel shape(Type A) PDA were studied retrospectively,and they were divided into angiography group(n =21) and echocardiography group(n =37).The angiography group received traditional transcatheter occlusion technique,and the echocardiography group received TTE guided transcatheter occlusion of PDA.An observation for the effect was taken post operatively,as well as for the comparison between the 2 groups in the aspects of the operation duration,the time of X-ray exposure,the radiation dose,the length of hospital stay and the expense of medication.Results Successful occlusion was performed in all patients.Both of the TTE and angiogram group showed good position of the occlusion devices.Moreover,the velocities of blood flow in left pulmonary artery and the descending aorta were in a normal range.Compared with angiography group,echocardiography group experienced shorter operation duration (P ＜ 0.05),shorter X-ray exposure time (P ＜ 0.05),lower radiation dose,fewer days of hospital stay (P ＜ 0.05) and less expense of the medication (P ＜ 0.05).Conclusions TTE-guided transcatheter occlusion for simple funnel shape PDA (Type A) is a simplified occlusion method,which is effective and preferential treatment to the method of traditional angiography guidance in clinical trials.

BACKGROUNDMyocardial tissue-level perfusion failure is associated with adverse outcomes following ST-elevation myocardial infarction (STEMI) despite successful epicardial recanalization. We have developed a new quantitative index-thrombolysis in myocardial infarction (TIMI) myocardial perfusion frame count (TMPFC)--for assessing myocardial tissue level perfusion. However, factors affecting this novel index of myocardial perfusion are currently unknown.

METHODSA total of 255 consecutive STEMI patients undergoing primary angioplasty were enrolled. Myocardial tissue level perfusion was assessed by TMPFC, which measures the filling and clearance of contrast in the myocardium using cine-angiographic frame counting. We differentiate three groups with two cut off values for TMPFC: a TMPFC of 90 frames was the upper boundary of the 95% confidence interval (CI) for the TMPFC observed in normal arteries, and a TMPFC of 130 was the 75th percentile of TMPFC.

RESULTSSTEMI patients with TMPFC > 130 frames (68 patients, 26.7%) had higher clinical and angiographic risk factor profiles as well as a higher 30-day MACE rate compared with those with TMPFC ≤ 90 frames and those with TMPFC > 90 and ≤ 130 frames. Multivariable analysis identified that the independent predictors of TMPFC > 130 frames were age ≥ 75 years (OR 2.08, 95%CI 1.21 to 3.58, P = 0.007), diabetes (OR 1.37, 95%CI 1.01 to 1.86, P = 0.042), Killip class ≥ 2 (OR 1.52, 95%CI 1.05 to 2.21, P = 0.027), and prolonged pain-to-balloon time (OR 1.73, 95%CI 1.07 to 2.79, P = 0.013). TMPFC > 130 frames was identified as the strongest independent predictor of 30-day major adverse cardiac event (MACE) (OR 2.77, 95%CI 1.21 to 6.31, P = 0.008), along with age ≥ 75 years (OR 2.19, 95%CI 1.11 to 4.33, P = 0.016), female gender (OR 1.67, 95%CI 1.03 to 2.70, P = 0.038), and Killip class ≥ 2 (OR 1.83, 95%CI 1.07 to 3.14, P = 0.021).

CONCLUSIONSSTEMI patients with poor myocardial perfusion assessed by TMPFC had higher risk factor profiles. Advanced age, diabetes, higher Killip class, and longer ischemia time were independent predictors of impaired TMPFC after primary percutaneous coronary intervention. These results emphasize that particular attention should be paid on myocardial microvascular reperfusion in STEMI patients with these risk factors.

Aged ; Angioplasty, Balloon, Coronary ; Coronary Angiography ; Female ; Humans ; Male ; Middle Aged ; Myocardial Infarction ; diagnostic imaging ; pathology ; therapy ; Myocardial Reperfusion ; Myocardium ; metabolism ; pathology

OBJECTIVETo study the clinicopathologic features, immunophenotype and genetic changes of perivascular epithelioid cell neoplasms (PEComa).

METHODSA total of 25 cases of PEComa located in various anatomic sites were selected for immunohistochemical staining (SP or EnVision method). TFE3 fluorescence in-situ hybridization was also performed to determine the TFE3 gene status.

RESULTSThe age of patient ranged from 21 to 61 years (mean = 43 years). The male-to-female ratio was 1: 1.3. Histologically, 22 cases represented conventional angiomyolipomas, composed of a mixture of adipose tissue, spindle element, epithelioid smooth muscle cells and abnormal thick-walled blood vessels in various proportions. Three cases involving lung, soft tissue and broad ligament had subtle but distinctive morphologic features. Nested or sheet-like architecture with epithelioid or spindle cells was observed. Immunohistochemical study showed that HMB 45, melan A, smooth muscle actin and cathepsin K were expressed in 80% (20/25), 88% (22/25), 88% (22/25) and 100% (25/25) of PEComa, respectively. Within positive cases, the average proportion of positive tumor cells was 36%, 41%, 35% and 90% respectively for HMB 45, melan A, smooth muscle actin and cathepsin K. TFE3 was negative in all of the 22 renal and hepatic PEComa studied, while it was positive in the 3 cases of extra-hepatorenal PEComa. None of the 25 cases exhibited evidence of TFE3 gene fusion or amplification.

CONCLUSIONSExtra-hepatorenal PEComa have distinctive morphologic features and are associated with TFE3 overexpression. Cathepsin K immunostaining demonstrates high sensitivity and specificity in PEComa, better than other commonly employed immunomarkers. This marker is thus useful in diagnosis of PEComa and distinction with other neoplasms.

Actins ; metabolism ; Adult ; Angiomyolipoma ; metabolism ; pathology ; Basic Helix-Loop-Helix Leucine Zipper Transcription Factors ; genetics ; metabolism ; Cathepsin K ; metabolism ; Female ; Humans ; Immunohistochemistry ; Kidney Neoplasms ; metabolism ; pathology ; Liver Neoplasms ; metabolism ; pathology ; MART-1 Antigen ; metabolism ; Male ; Melanoma-Specific Antigens ; metabolism ; Middle Aged ; Perivascular Epithelioid Cell Neoplasms ; metabolism ; pathology ; Young Adult

BACKGROUNDStreptococcus pneumoniae (S. pneumoniae) is a major causative agent of severe infections, including sepsis, pneumonia, meningitis, and otitis media, and has become a major public health concern. We report the pneumococcal serotype and sequence type (ST) distribution, and antimicrobial resistance of 39 S. pneumoniae strains from seven hospitals in China.

METHODSBlood/cerebrospinal fluid (CSF) and sputum isolates from patients were analyzed to determine S. pneumoniae serotypes by polymerase chain reaction (PCR) and the Neufeld Quellung reaction, the multilocus sequence types (MLST) by PCR and sequencing, and susceptibility to antimicrobial agents by the VITEK Gram Positive Susceptibility Card.

RESULTSA total of 39 isolates were collected including 21 blood/CSF and 18 sputum isolates. Conventional serotyping by the Quellung reaction required 749 reactions. In contrast, PCR based typing needed only 106 PCR reactions. The most frequent serotypes from the blood/CSF isolates were 14 (38.1%), 19A (14.3%), 23F (9.5%), and 18C (9.5%). In the sputum isolates the most frequent serotypes were 19F (33.3%), 23F (16.7%), 19A (11.1%), and 3 (11.1%). The incidence of penicillin resistance in the blood/CSF and sputum isolates was 66.7% and 55.6%, respectively. Statistical analysis showed that patients = 5 years old had a higher resistance to penicillin when they compared with the patients = 65 years old (P = 0.011). Serotypes 14, 19A and 19F were significantly associated with penicillin resistance (P < 0.001). ST320, ST271, and ST876 isolates showed high resistant rates to several antibiotics including penicillin (P = 0.006). All of the isolates of serotype 19A were resistant to both penicillin and erythromycin, and they were all multi-drug resistant (MDR) isolates.

CONCLUSIONSThe specificity and sensitivity of multiplex-PCR are good, and this method represents a substantial savings of time and money, and can be widely used in the laboratory and clinical practice. Data from this research showed an extremely high prevalence of penicillin resistance and an increasing prevalence of multi-drug resistant (MDR) rate in S. pneumoniae. A distinctive emergence of serotype 19A was observed which was also associated with the increasing prevalence of antimicrobial resistance. Therefore, nationwide surveillance of pneumococcal resistance and serotypes is strongly warranted.

Adolescent ; Adult ; Aged ; Child ; Child, Preschool ; Drug Resistance, Multiple, Bacterial ; Humans ; Infant ; Microbial Sensitivity Tests ; Middle Aged ; Molecular Typing ; methods ; Multilocus Sequence Typing ; methods ; Pneumococcal Infections ; microbiology ; Serotyping ; Streptococcus pneumoniae ; classification ; drug effects