BACKGROUND: Tumor necrosis factor-associated protein 1 (TRAP1) is a heat-shock protein 90-related mitochondrial chaperone. Accumulative evidence has demonstrated that TRAP1 overexpression is closely related to carcinogenesis. However, the exact function and mechanism of TRAP1 in the occurrence of laryngeal carcinoma remains unclear. OBJECTIVE: To investigate whether RNA interference can inhibit TRAP1 overexpression and to explore its effects on growth and apoptosis of CD133+CD44+ laryngeal carcinoma stem cells. METHODS: CD133+CD44+ laryngeal carcinoma stem cells were sorted from human laryngeal carcinoma Hep-2 cellsusing immunomagnetic beads. The shRNA sequence of TRAP1 was designed and synthesized and CD133+CD44+ laryngeal carcinoma stem cells were transfected with LipofectamineTM 2000. Cell counting kit-8 assay, colony formation assay and flow cytometry were used to investigate the effects of interference of TRAP1 expression on growth and apoptosis of CD133+CD44+ laryngeal carcinoma stem cells. Spectrophotometric method was used to detect the activity of caspase-3, -8 and -9. RESULTS AND CONCLUSION: TRAP1 mRNA and protein expression levels were significantly decreased in TRAP1 shRNA-transfected CD133+CD44+ laryngeal carcinoma stem cells (P < 0.01). Compared with the blank control and negative control groups, the growth and colony formation of CD133+CD44+ laryngeal carcinoma stem cells were significantly inhibited in the TRAP1 shRNA-transfected group (P < 0.05). Apoptosis of CD133+CD44+ laryngeal carcinoma stem cells was significantly inhibited in the TRAP1 shRNA-transfected group as compared with the blank control and negative control groups (P < 0.05). TRAP1 shRNA-mediated cell apoptosis was associated with the activation of caspase-3, -8 and -9. These results suggest that RNA interference targeting inhibition of TRAP1 suppresses cell growth but promotes apoptosis in CD133+CD44+ aryngeal carcinoma stem cells. TRAP1 is likely to be a gene target for treatment of laryngeal carcinoma.

Objective: To establish a UPLC-MS/MS method for the determination of the residual quantity of potential genetoxic impurity 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC) in ledipasvir.Methods: The chromatographic conditions were as follows: an Agilent ZORBAX SB-C18 (75 mm×4.6 mm, 3.5 μm) column was used with methanol-0.1% formic acid (50∶50) as the mobile phase.The flow rate was 0.3 ml·min-1.The column temperature was 30 ℃ and the injection volume was 5 μl.The mass spectrometry conditions were as follows: electrospray ionization source (ESI), multistage reaction monitoring (MRM), positive ion scanning mode, ion spray voltage of 2 500 V, ion source temperature of 500 ℃, atomization gas of 379.0 kPa, auxiliary gas of 275.6 kPa, curtain gas of 137.8 kPa, collision gas of 41.3 kPa, ion collision energy of 15 V and scan time of 100 ms.The MRM ion pair for quantitative analysis was m/z→156.2/86.1.Results: The linear range of EDC was 0.03-2.25 μg·ml-1 (r=0.999 0).The limit of detection was 0.03 ng and the limit of quantitation was 0.08 ng.The average recovery was 98.3%(RSD=5.7%, n=9).Conclusion: The method is simple, rapid and accurate, and can be used for the quality control of EDC in ledipasvir.

Objective To analyze the clinical characteristics and prognosis of disseminated cryptococcosis in children.Methods The data of disseminated cryptococcosis inpatients were reviewed retrospectively at Beijing Children's Hospital,Capital Medical University,from January 2002 to September 2014.The demographic data,clinical manifestations,laboratory findings,imaging,antifungal treatments and outcomes of all the patients were analyzed.Results Overall 25 children with disseminated cryptococcosis were enrolled including 17 boys(68.0％).The average age was 7 years old.Four cases (16.0％) had underlying conditions,among them 1 case had human immunodeficiency virus(HIV)-positive.The median time to diagnosis was 32 (23-47) days,44.0％ of the patients (11 cases) were misdiagnosed,and 8 cases misdiagnosed as tuberculosis.All patients had fever.Other common clinical manifestations included cough (16 cases),headache (10 cases),vomiting (10 cases),altered mental status (6 cases) and stomachache (6 cases).Respiratory system involvement was seen in all cases,central nervous system was involved in 18 cases,other organ involvement included lymph nodes,spleen,liver,kidney,skin,skeleton and costicartilage.Amphotericin B (AmB) + Fluconazole (Flu) ± 5-flucytosine (5-FC) was the most common therapy (15 cases),Flu ± 5-FC for 6 cases,AmB ± 5-FC for 2 cases,Voricanazole (VOR) for 1 case.Sixteen cases (66.7％) got recovery/improved on discharge,8 cases (33.3％) rejected to the advise and discharged with treatment failure,and the HIV-infected patient transferred to a special hospital.Fifteen patients (60.0％) were followed up,and 13 cases (87.0％) showed recovery,but 2 died in the long-term prognosis.Conclusions Disseminated cryptococcosis in children is more common in school-age boys.Most patients are without underlying conditions.Disseminated cryptococcosis can cause multiple organ damage.Patients with prolonged fever,cough,headache,with or without underlying diseases,can be suspected as disseminated cryptococcosis.Blood and cerebrospinal fluid culture for fungus,cryptococcal antigen should be performed for early diagnosis and treatment.

OBJECTIVETo correlate the abnormal expression of anapastic lymphoma kinase (ALK) protein with the genetic and epigenetic changes of ALK, and to analyze its clinical application in pediatric neuroblastoma.

METHODSThree neuroblastoma (NB) cell lines (two ALK positive: SH-SY5Y and SK-N-SH, one ALK negative: SK-N-AS) and 43 paraffin-embedded NB tissues were included in the study. In both cell lines and clinical cases, immunohistochemistry was used to detect ALK protein expression; PCR and Sanger sequencing were used to detect ALK point mutation; fluorescence in situ hybridization (FISH) was used to detect ALK abnormality and bisulfite sequencing PCR (BSP) was used to detect methylation of CpG island in the promoter area of ALK.

RESULTSThe cell lines SH-SY5Y and SK-N-SH were positive for ALK expression (cytoplasm), while the SK-N-AS was negative; among the 43 cases of NB, 26 (60.5%, 26/43) were positive for ALK protein (membrane and cytoplasm), and the rest were negative. Survival analysis showed ALK protein expression was related to survival time, with ALK positive cases having shorter survival time than ALK negative cases (P = 0.020). But ALK protein expression had no association with tumor differentiation (P = 0.503), tumor sites (P = 1.000) and age of patients (P = 0.063). FISH showed ALK amplification in two cases (4.6%, 2/43), ALK gain was found in 30 cases (69.7%, 30/43), and the remaining cases had normal ALK copy (25.6%, 11/43). The presence of extra copies (amplification and gain) of ALK was associated with ALK positive protein expression (P = 0.020), but there was no association with tumor differentiation (P = 1.000), tumor sites (P = 0.775) and age of patients (P = 0.328). No point mutation was found in all three cell lines. Of the 43 NB cases, only one case (2.3%, 1/43) showed point mutation in exon 23, and was a synonymous mutation [A1200A (G4552C)]. The case was ALK negative, but the patient died two months after diagnosis. BSP analysis showed that CpG island in ALK promoter region were all unmethylated in three cell lines and 6 NB cases (including 3 ALK positive, 3 ALK negative).

CONCLUSIONSALK protein is expressed in most NB, and the expression indicates poor outcome. ALK expression is associated with extra copies of ALK, but there is no association with the methylation status of CpG island of ALK; the presence of extra copies of ALK is the most common genetic aberration in NB. Point mutation of ALK is rare, and may predict poor prognosis in pediatric NB.

Adolescent ; Cell Line, Tumor ; Child ; Exons ; Humans ; Immunohistochemistry ; In Situ Hybridization, Fluorescence ; Neuroblastoma ; enzymology ; genetics ; Receptor Protein-Tyrosine Kinases ; genetics ; metabolism

Objective To study the patients' clinical characteristics and prognosis when only C3 deposition exists in endocapillary proliferative glomerulonephritis and try to understand deeply the role of C3 in kidney damage deeply. Methods The patients who were diagnosed with endocapillary proliferative glomerulonephritis but only had C3 deposited in immunofluorescence(to avoid false positive,C3≥2 ﹢ was included)were selected from Beijing Children's Hospital Affiliated to Capital Medical University during November 2010 to October 2014. Their clinical manifestations,la-boratory examinations,treatments,prognosis,and pathological changes were analyzed,and literature review was performed. Their clinical characteristics and prognosis were summarized. Results There were 11 patients diagnosed with endocapil-lary proliferative glomerulonephritis which had only C3 deposition(≥2 ﹢ ). Nine of them had onset with acute nephritis syndrome(81. 8% ),and 2 cases presented recurrent paroxysmal gross hematuria(18. 2% ). Seven cases were diagnosed with acute post streptococcal glomerulonephritis(63. 6% ). Eleven cases' clinical manifestations were relatively severe, and the complement C3 was significantly lower than the normal(100. 0% ). Their light microscope showed capillary proli-ferative glomerulonephritis,and the electron microscope showed the immune complexes were deposited in the endothelium,the epithelium or the mesangial area. The patients received corresponding treatment respectively,and all the patients had good prognosis during following up of 7 months up to 39 months. Conclusions Streptococcus infection is a common cause in endocapillary proliferative glomerulonephritis with only C3 deposition. The clinical manifestations of some children are similar to post streptococcal glomerulonephritis but relatively severe. Only deposition of C3 without IgG may be involved in another complement activation mechanism.

Objective To investigate the molecular genetic changes of anaplastic lymphoma kinase (ALK) gene and c-myc gene in anaplastic large cell lymphoma (ALCL). Methods The structural aberrations and changes of copy numbers in ALK and c-myc genes in 72 paraffin-embedded ALCL specimens were detected by interphase fluorescence in situ hybridization (FISH). Results Among 72 ALCL specimens, ALK protein was expressed in 42, ALK gene translocation was detected in 40 specimens in which extra copies of ALK gene were detected in 17. ALK gene translocation was not found in all 30 ALK negative specimens, but extra copies of ALK gene were detected in 14 cases. The difference of incidence rates of extra copies in ALK gene between ALK positive and ALK negative specimens was not significant (P＞0.05). c-myc gene translocation was not found in any of 72 ALCL specimens, but extra copies were detected in 24 cases.Conclusion Most (75.0%) ALCL have ALK gene aberration, in which ALK gene translocations are most common (55.6%), and the extra copies of ALK gene are relatively common genetic changes (43.1%). The ALK gene aberration is only detected in ALK positive ALCL and the gene translocations are in either ALK positive and negative ALCL. There is no or rare c-myc gene translocation in ALCL, but extra copies of c-myc gene are relatively common (33.3%).

Objective:To investigate the effect of serum cystatin C level on the occurrence and its long-term prognosis of contrast agent-induced acute kidney injury(CI-AKI)after percutaneous coronary intervention(PCI)in elderly patients.Methods:A total of 848 elderly patients(≥60 years)undergoing PCI in our department between Mar 2015 and Dec 2017 were enrolled in a prospective cohort.The CI-AKI was defined as the increase of serum creatinine ≥44.2 μmol/L within 48-72 h after using iodine contrast agent or more than 25 % higher than base level within 48-72 h after PCI.A receiver operating characteristic curve was used to analyze the optimal cut-off value of Cystatin C for predicting CI-AKI after PCI.Patients were divided into 2 groups based on the optimal cut-off value of Cystatin C: the high Cystatin C group(Cystatin C ≥1.3 mg/L, n=178)and the control group(Cystatin C<1.3 mg/L, n=670). The differences in the incidence of CI-AKI after PCI and major adverse cardiac events(MACE)at 1 year follow-up were compared between the two groups.The Cox regression model was further used to analyze the predictors of the long-term prognosis after PCI.Results:Of 848 patients receiving PCI, the incidence of CI-AKI was 9.4%.The incidence of MACE at 1 year after PCI was higher in the high Cystatin C group than in the control group(15.7% vs.9.3%, χ2=6.524, P=0.011). Cox regression analysis confirmed that the high baseline level of Cystatin C was the most independent predictive factor for MACE at 1 year of follow-up( HR=16.244, P<0.001). Conclusions:The high baseline level of Cystatin C(≥1.3 mg/L)is an independent risk factor for CI-AKI and is also the most important predictor for the occurrence of long-term MACE in elderly patients undergoing PCI.

Objective To explore the impact of Enalapril-folic acid tablet on development of contrast-induced nephropathy (CIN) after percutaneous coronary intervention (PCI) in elderly patients with coronary heart diseases(CHD) complicated with mild renal insufficiency. Methods A total of 935 old patients (≥60 years) undergoing selective PCI at our hospital from January 2013 to June 2016 were enrolled in this retrospective study.Based on treatments during the perioperative phase ,the 935 patients were divided into Enalapril-folic acid tablet intervention group (n= 296 ,31.7％ ) ,and other RAS blocker control group with Angiotensin eonverting-enzyme inhibitor (ACEI) or Angiotensin Ⅱreceptor blocker (ARB) (n=639 ,68.3％ ). The levels of serum creatinine for 72 hours after operation were monitored and evaluated.Baseline data and CIN incidence were compared between two groups.The clinical events within 1 month after contrast media application were recorded.Multivariate Logistic regression analysis was used to analyze independent risk factors for CIN after PCI. Results The incidence of CIN after PCI was lower in the intervention group than in the control group with no statistical significance[7.1％ (21/296) vs.11.1％ (71/639) ,χ2 = 3.679 ,P= 0.059].Multivariate regression analysis showed that age (OR=1.103 ,P=0.001) ,hypertension (OR=3.362 ,P=0.017) , and hyper-homocysteinemia (OR=3.528 ,P=0.003) were independent risk factors ,but the treatment with Enalapril-folic acid tablet might be a protective factor for development of CIN after PCI (OR=0.443 ,P=0.042).During the 1-month follow-up ,the rate of recurrent angina pectoris was lower in the intervention group than in the control group ( P < 0.05 ) ,while there were no statistically significant differences between two groups in the rates of rehospitalization due to worsening renal dysfunction ,dialysis/hemofiltration ,acute heart failure ,and new myocardial infarction (all P>0.05). Conclusions The treatment with enalapril-folic acid tablet may prevent the occurrence of CIN after PCI in elderly patients with coronary heart disease and mild renal insufficiency.

Transcription factor is a key trans-acting factor to mediate stress response by regulating gene expression. Plants have developed a series of mechanisms to modulate development, stress response, signaling and disease resistance at transcription level. DNA binding with one finger (DOF), containing one C₂-C₂ zinc finger domain, is a special plant transcription factor. Specifically, the conserved domain at N-terminus of DOF has multiple functions, including interacting with DNA and protein, which could be involved in plant development and stress response. Although many DOF family genes are characterized in plant stress response, it is not clear if DOF genes have functions in cereal plants. In the present paper, the role of DOF family genes on cereal plants were discussed based on a comprehensive phylogenetic relationship analysis, expression profiles in different tissues and various environmental conditions. The results obtained here will provide an important reference for further understanding the mechanism of gramineous crops in stress resistance.
DNA-Binding Proteins ; metabolism ; Gene Expression Regulation, Plant ; Phylogeny ; Plant Proteins ; metabolism ; Plants ; genetics ; Transcription Factors ; metabolism ; Zinc Fingers