Rapid Detection of Fusarium oxysporum Using Insulated Isothermal PCR and a Rapid, Simple DNA Preparation Protocol.

We developed an insulated isothermal PCR (iiPCR) method for the efficient and rapid detection of Fusarium oxysporum (Fo), which is a fungus that infects various hosts and causes severe crop losses. The Fo iiPCR method was sensitive enough to detect up to 100 copies of standard DNA template and 10 fg of Fo genomic DNA. In addition, it could directly detect 1 pg of mycelium and 10 spores of Fo without DNA extraction. Our study compared the performance of Fo iiPCR to that of three published in planta molecular detection methods-conventional PCR, SYBR green-based real-time PCR, and hydrolysis probe-based real-time PCR-in field detection of Fo. All diseased field samples yielded positive detection results with high reproducibility when subjected to an Fo iiPCR test combined with a rapid DNA extraction protocol compared to Fo iiPCR with an automated magnetic bead-based DNA extraction protocol. Intraday and interday assays were performed to ensure the stability of this new rapid detection method. The results of detection of Fo in diseased banana pseudostem samples demonstrated that this new rapid detection method was suitable for field diagnosis of Fusarium wilt and had high F1 scores for detection (the harmonic mean of precision and recall of detection) for all asymptomatic and symptomatic Fo-infected banana samples. In addition, banana samples at four growth stages (seedling, vegetative, flowering and fruiting, and harvesting) with mild symptoms also showed positive detection results. These results indicate that this new rapid detection method is a potentially efficient procedure for on-site detection of Fo.

E-cadherin gene methylation in lung cancer.

E-cadherin, a tumor repressor gene, has been shown to play an important role in maintaining the polarity and structural integrity of epithelial and is closely associated with tumorigenesis, invasion, and metastasis. The current study aimed to investigate the effects of E-cadherin methylation on lung cancer (LC) quantitatively through a meta-analysis. We searched electronic databases to identify eligible studies from their inception through September 30, 2013. Pooled odds ratio (OR) with 95 % confidence interval (CI) was used to assess the relationship between E-cadherin gene methylation and LC risk. A hazard ratio (HR) with 95 % CI was used to assess the impact of E-cadherin gene methylation on overall survival (OS) of LC patients. Seventeen studies comprising 983 LC cases and 669 controls met the inclusion criteria. Summary results revealed that hypermethylation frequencies in LC tissues were significantly higher than those in normal control tissues (OR = 4.11, 95 % CI 2.78-6.07, P < 0.001). Subgroup analysis indicated that higher methylation frequencies were observed in Asian population. Interestingly, we found that hypermethylation of E-cadherin was associated with significantly better survival with HR of 0.47 (95 % CI 0.31-0.71). This meta-analysis revealed that E-cadherin gene promoter methylation was associated with an increased risk of LC, especially in Asian population, and methylated E-cadherin predicted long survival in patients with LC. However, further studies with large numbers of patients will be needed to confirm the findings.

Expression of survivin and patients survival in non-small cell lung cancer: a meta-analysis of the published studies.

Among new biological markers that could become useful prognostic factors for non-small cell lung cancer (NSCLC). Survivin is one of the most commonly over-expressed oncogenes, however, its role in NSCLC remains controversial. We performed a systematic review of the literature with meta-analysis to clarify this issue. Electronic databases were used to identify published studies before August 2011. Pooled hazard ratio (HR) with 95 % confidence interval (95 % CI) was used to estimate the strength of the association of survivin expression with survival of NSCLC patients. Heterogeneity and publication bias were also assessed. Overall 29 relevant published studies including 2,517 lung cancer patients were identified from electronic databases. We found that overexpression of survivin in NSCLC patients might be a poor prognostic factor for survival 1.95 (95 % CI: 1.65-2.29; P < 0.001). Heterogeneity testing indicated that there was heterogeneity among studies. When stratified by histology types, the heterogeneity was absent. We should point out that the publication bias may partly account for the result, but the conclusion might not be affected deeply by the publication bias. When we accounted for publication bias using the trim and fill method, the results remained significant (HR = 1.71, 95 % CI: 1.44-2.02, P < 0.001), suggesting the stability of our results. Therefore, our study suggested that survivin overexpression had a poor prognosis value in patients with NSCLC.

[Ultrastructure of the digestive system in Dermatophagoides farinae (Acariformes:Pyroglyphidae)].

Fifty living mites (Dermatophagoides farinae) were fixed in 2.5% glutaraldehyde, postfixed in 1% osmium tetroxide, dehydrated in a graded ethanol series, embedded in embedding medium. The ultrastructure of the digestive tract in D. farinae was observed by serial ultrathin sections with a transmission electron microscope. The alimentary canal of D. farinae consists of the cuticle-lined foregut and hindgut separated by a microvilli-lined midgut (anterior midgut, posterior midgut). There are different types of epithelial cells in the anterior midgut The microvilli of epithelial cells in posterior midgut are longer than that of the anterior midgut In posterior midgut, the food bolus is surrounded by the peritrophic membrane. The midgut is the main site of digestion and absorption.

Low molecular weight heparin in treating patients with lung cancer received chemotherapy: A meta-analysis.

OBJECTIVE: To systematically review the efficacy and safety of low molecular weight heparin (LMWH) in treating patients with lung cancer received chemotherapy. MATERIALS AND METHODS: Databases including PubMed, The Cochrane Library, Excerpt Medica Database, Chinese Biomedical Literature Database, China National Knowledge Infrastructure, VIP, and Wanfang Data were searched for the randomized controlled trials (RCTs) about LMWH in treating patients with lung cancer received chemotherapy from the establishment to May 31, 2015. According to the inclusion and exclusion criteria, two reviewers independently screened literature, extracted data, and assessed quality of the included studies. Meta-analysis was then performed by using Review Manager 5.3 (Cochrane Collaboration, Oxford, UK) software. RESULTS: A total of eight RCTs involving 952 patients were finally included. Meta-analysis showed that compared with the control group, LMWH significantly improved the 1- and 2-year overall survival (OS) rates of the patients with lung cancer received chemotherapy (risk ratio [RR] =1.65, 95% confidence interval [95% CI] [1.20-2.26], P = 0.002; RR = 2.63, 95% CI [1.40-4.94], P = 0.003, respectively), and significantly reduced the incidence of venous thromboembolism (VTE) (RR = 0.40, 95% CI [0.23-0.69], P = 0.001), not significantly increased the incidence of major bleeding events and thrombocytopenia (RR = 1.29, 95% CI [0.57-2.96], P = 0.54; RR = 0.86, 95% CI [0.69-1.07], P = 0.18, respectively), and not significantly improved the overall response rate (RR = 1.24, 95% CI [0.98-1.57], P = 0.07). CONCLUSION: LMWH improves the 1- and 2-year OS rates of the patients with lung cancer received chemotherapy and reduces the incidence of VTE, not increase the incidence of major bleeding events and thrombocytopenia. These show that there is a certain effect of LMWH, and the security is good.

Testin is a tumor suppressor in non-small cell lung cancer.

The Testin gene was previously identified in the fragile chromosomal region FRA7G at 7q31.2. It has been implicated in several types of cancers including prostate, ovarian, breast and gastric cancer. In the present study, we investigated the function of the candidate tumor-suppressor Testin gene in non-small cell lung cancer (NSCLC). In NSCLC cell lines, we observed lower expression of Testin compared to that noted in normal human bronchial epithelial cells. MTT assays, flow cytometry, clonogenic assay and invasion assay showed that the overexpression of the Testin gene inhibited cancer cell proliferation, invasion and colony formation. In tumor xenograft models, Testin markedly inhibited lung cancer cell xenograft formation and growth in athymic nude mice. Taken together, these data suggest that Testin plays an important role in the development and progression of NSCLC. Testin may be an effective novel target in NSCLC prevention and treatment.

HIST1H3D: A promising therapeutic target for lung cancer.

HIST1H3D gene encodes histone H3.1 and is involved in gene-silencing and heterochromatin formation. HIST1H3D expression is upregulated in primary gastric cancer tissue. In this study, we explored the effects of HIST1H3D expression on lung cancer, and its mechanisms. HIST1H3D expression was measured by immunohistochemistry and RT-PCR in lung cancer tissues and human lung cancer cell lines. Cell proliferation was assessed by MTT assay. Flow cytometric analysis was used to determine cell cycle distribution and apoptosis. Levels of related proteins were detected by western blotting. Bioinformatics analysis was performed to investigate related signaling pathways. cDNA microarray analysis was performed to identify differentially expressed genes following HIST1H3D knockdown. HIST1H3D expression was upregulated in lung cancer tissue samples and the H1299 human lung cancer cell line (P<0.01). Regulation of HIST1H3D expression in nucleus of cells in lung cancer tissues was significant associated with tumor stage (P=0.02) and lymph node metastases (P=0.04). Downregulation of HIST1H3D expression led to suppression of proliferation and colony forming ability, cell cycle arrest at the G0/G1 phase, and promotion of cell apoptosis. The microarray data revealed 522 genes that were differentially expressed after HIST1H3D knockdown in H1299 cells. These genes were shown to be linked to numerous pathways, including the cell cycle, p53 signaling, and MCM. Western blot analysis confirmed upregulated expression of the THBS1 and TP53I3 genes, and downregulated expression of the CDK6, CDKN1 and CCNE2 genes. In conclusion, our results suggest that HIST1H3D is highly expressed in lung cancer cell lines and tissues. Furthermore, HIST1H3D may be important in cell proliferation, apoptosis and cell cycle progression, and is implicated as a potential therapeutic target for lung cancer.

[Molecular mechanism of anti-apoptotic action of survivin in NCI-H446 lung cancer cells].

OBJECTIVE: To investigate cell apoptosis induced by survivin ASODN and clarify the precise mechanism of anti-apoptotic action of survivin. METHODS: Cells of lung cancer cell line NCI-H446 were treated with survivin ASODN at different concentrations. The changes of survivin mRNA and protein expression were assessed by RT-PCR and Western blot assay. The apoptosis index (AI) and proliferation index (PI) were determined by flow cytometry (FCM). After 500 mmol/L survivin ASODN treatment, cells were stained with Rh123 to detect changes of mitochondrial membrane potential (deltapsim) by FCM. The concentration of cytoplasmic cytochrome c (cyt-c) was continuously determined by ELISA. Relative activities of caspase-9 and caspase-3 were assessed by colorimetric assay. The expression of caspase-8 protein was measured by Western blot assay. The apoptotic rates of lung cancer cells induced by survivin ASODN with or without mitochondrial permeability transition pole (MPTP) inhibitor CsA treatment were assessed by FCM. RESULTS: Down-regulated survivin mRNA was shown to be in dose-dependent and time-dependent manners. Its maximal effect was achieved at a concentration of 500 nmol/L for 72 h, at which mRNA was down-regulated by 62.7%, the expression of survivin protein in NCI-H446 cells was also obviously decreased. After treatment with survivin ASODN at concentration of 500 mmol/L for 72 h, AI was 48.35%, higher than that of control, lipofectin, NSODN, survivin ASODN 100 mmol/L and 300 mmol/L groups (3.75%, 3.41%, 4.69%, 19.85% and 34.39%, respectively). PI was 24.38%, lower than that of control, lipofectin, NSODN, survivin ASODN100 and 300 mmol/L groups (75.74%, 73.12%, 71.76%, 51.03% and 38.94%, respectively). Deltapsim was decreased in 9.54% of NCI-H446 cells treated with survivin ASODN for 3 h and 97.06% for 24 h. Following it, release of cyt-c from mitochondria to cytosol and activation of caspase-9 and caspase-3 increased significantly. The above mentioned indicators changed with a time-dependent and time diversity relationship. In the presence of CsA, the apoptotic rate of lung cancer cells induced by survivin ASODN was decreased significantly. No up-regrulation and activation in caspase-8 protein was observed. CONCLUSION: Survivin inhibits apoptosis via regulation of mitochondrial-dependent pathway. survivin ASODN can not only induce apoptosis but also inhibit cell proliferation through blocking the expression of survivin mRNA and protein.

Prognosis value of HIF-1α expression in patients with non-small cell lung cancer.

BACKGROUND: Hypoxia-inducible factor 1 (HIF-1) is a transcription factor that plays a critical role in the development and progression of tumors. Various studies evaluating the prognostic value of HIF-1α in patients with lung cancer (LC) remain controversial. To comprehensively and quantitatively summarize the evidence on the effect of HIF-1α expression on the survival of patients with LC, a meta-analysis was carried out. MATERIAL AND METHODS: Electronic databases were used to identify published studies before August 31st, 2013. Studies were assessed for quality using REMARK. Data were collected comparing overall survival in patients with high HIF-1α expression with those with low expression. RESULTS: Totally, 13 papers including 1420 patients were subjected to final analysis. The combined hazard ratio (HR) was 1.60 (95% CI: 1.14-2.25, P=0.007), suggesting that high expression of HIF-1α was an indicator of poor prognosis. Further, when stratified by LC histological type (SCLC and NSCLC), study region (Asia and Europe), cut-off values (10%), tumor stage (I-III and I-IV), antibody for IHC (H1α67 and ESEE 122), and HR estimated method (univariate/multivariate analysis), most of the results were statistically significant. CONCLUSIONS: Taken together, this meta-analysis revealed that HIF-1α overexpression might be a predicative factor of poor prognosis for NSCLC particularly in Asia.

Meta-analysis for cyclin E in lung cancer survival.

BACKGROUND: To assess the prognosis value of cyclin E expression in survival of patients with lung cancer (LC), we performed a systematic review of the literature with meta-analysis. METHODS: Electronic databases were used to identify published studies before August 2011. Pooled hazard ratio (HR) with 95% confidence interval (95% CI) was used to estimate the strength of the association of cyclin E expression with survival of LC patients. Heterogeneity and publication bias were also assessed. RESULTS: Fourteen studies (2606 cases) were eligible and subjected to analysis. Cyclin E over-expression was found to be a strong predictor of poor prognosis in LC patients (HR: 1.38, 95% CI: 1.07-1.79; P=0.014). When only non-small cell lung cancer (NSCLC) was considered, the combined HR was 1.53 (95% CI: 1.19-1.97, P=0.001). A significant association was also evident when the analysis was limited to studies involving adenocarcinoma (AD), but not squamous cell carcinoma (SQ). Publication bias was absent. Sensitivity analyses suggested that the summary statistics obtained should approximate the actual average.