Single-cell and bulk RNA sequencing reveal heterogeneity and diagnostic markers in papillary thyroid carcinoma lymph-node metastasis.

PURPOSE: Papillary thyroid carcinoma (PTC) is characterized by lymph-node metastasis (LNM), which affects recurrence and prognosis. This study analyzed PTC LNM by single-cell RNA sequencing (scRNA-seq) data and bulk RNA sequencing (RNA-seq) to find diagnostic markers and therapeutic targets. METHODS: ScRNA-seq data were clustered and malignant cells were identified. Differentially expressed genes (DEGs) were identified in malignant cells of scRNA-seq and bulk RNA-seq, respectively. PTC LNM diagnostic model was constructed based on intersecting DEGs using glmnet package. Next, PTC samples from 66 patients were used to validate the two most significant genes in the diagnostic model, S100A2 and type 2 deiodinase (DIO2) by quantitative reverse transcription-polymerase chain reaction (RT-qPCR) and immunohistochemical (IHC). Further, the inhibitory effect of DIO2 on PTC cells was verified by cell biology behavior, western blot, cell cycle analysis, 5-ethynyl-2'-deoxyuridine (EdU) assay, and xenograft tumors. RESULTS: Heterogeneity of PTC LNM was demonstrated by Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Ontology (GO) analysis. A total of 19 differential genes were used to construct the diagnostic model. S100A2 and DIO2 differ significantly at the RNA (p < 0.01) and protein level in LNM patient tissues (p < 0.001). And differed in PTC tissues with different pathologic typing (p < 0.001). Further, EdU (p < 0.001) and cell biology behavior revealed that PTC cells overexpressed DIO2 had reduced proliferative capacity. Cell cycle proteins were reduced and cells are more likely to be stuck in G2/M phase (p < 0.001). CONCLUSIONS: This study explored the heterogeneity of PTC LNM using scRNA-seq. By combining with bulk RNA-seq data, diagnostic markers were explored and the model was established. Clinical diagnostic efficacy of S100A2 and DIO2 was validated and the treatment potential of DIO2 was discovered.

High-throughput miRNA sequencing and identification of a novel ICE1-targeting miRNA in response to low temperature stress in Eucalyptus camaldulensis.

MicroRNAs (miRNAs) play a crucial role in the growth, development, morphogenesis, signal transduction, and stress response in plants. The ICE (Inducer of CBF expression)-CBF (C-repeat binding factor)-COR (Cold-regulated gene) regulatory cascade is an important signalling pathway in plant response to low temperature stress, and it remains unknown whether this pathway is regulated by miRNAs. In this study, high-throughput sequencing was employed for predicting and identifying the miRNAs that were likely to target the ICE-CBF-COR pathway in Eucalyptus camaldulensis. A novel ICE1-targeting miRNA, eca-novel-miR-259-5p (nov-miR259), was further analysed. A total of 392 conserved miRNAs and 97 novel miRNAs were predicted, including 80 differentially expressed miRNAs. Of these, 30 miRNAs were predicted to be associated with the ICE-CBF-COR pathway. The full-length of mature nov-miR259 was 22 bp and its precursor gene was 60 bp in length, with a typical hairpin structure. The RNA ligase-mediated 5' amplification of cDNA ends (5'-RLM-RACE) and Agrobacterium-mediated tobacco transient expression assays demonstrated that nov-miR259 could cleave EcaICE1 in vivo. Moreover, qRT-PCR and Pearson's correlation analysis further revealed that the expression levels of nov-miR259 were almost significantly negatively correlated with those of its target gene, EcaICE1, and the other genes in the ICE-CBF-COR pathway. We first identified the nov-miR259 as a novel ICE1-targeting miRNA, and the nov-miR259-ICE1 module may be involved in regulating the cold stress response in E. camaldulensis.

[Specification of statistical graphics in medical research].

Statistical graphics has a long history and is an important mean to present study design, analysis results and conclusions of medical research. A survey of statistical graphs of recent publications in Chinese and English academic journals shows that scientific and technical specifications of statistical graphics are still lacking. Based on the requirements of statistical graphics in prestigious medical journals (NEJM, Lancet, JAMA and The BMJ), this paper summarizes the technical requirements, key points of design of statistical graphs and practical conditions of common statistical graphs to provide reference for clinical researchers.

ZNF295-AS1 inhibits autophagy via the ZNF295-AS1/miR-508-5p/ATG7 axis in AS.

OBJECTIVE: As a result of gene-environment interactions, the incidence of atherosclerosis (AS) is rapidly increasing worldwide. Autophagy in endothelial cells is a key process of AS and is difficult to control when it becomes excessive in the end stage of AS. MATERIALS AND METHODS: In this study, we found increased expression levels of ZNF295-AS1 in the serum of AS patients, as well as in ox-LDL-treated HUVECs. The autophagy level was also upregulated in both samples. We demonstrated that ZNF295-AS1 may interact directly with miR-508-5p to act as a miR-508-5p sponge. The negative relationship between ZNF295-AS1 and miR-508-5p indicated that ZNF295-AS1 may be an upstream suppressor of miR-508-5p. RESULTS: ATG7 plays a critical role in autophagy and was predicted to be a target of miR-508-5p. Therefore, we overexpressed miR-508-5p, which reduced the expression level of ATG7, enhanced cell proliferation and prevented autophagy. These data indicated that the ZNF295-AS1/miR-508-5p/ATG7 axis may participate in autophagy regulation in ox-LDL-treated HUVECs. The subsequent rescue experiments revealed the specificity of the ZNF295-AS1/miR-508-5p/ATG7 axis in the contribution of ZNF295-AS1 to autophagy. CONCLUSIONS: Overall, our findings demonstrate a novel mechanism by which ZNF295-AS1 silencing regulates ATG7 reduction and inhibits autophagy, which may delay the progression of AS. The ZNF295-AS1/miR-508-5p/ATG7 axis may be of therapeutic significance in AS.

[Analysis of influencing factors of ureteral anastomotic stenosis following urinary diversion after laparoscopic radical cystectomy].

Objective: To investigate the influencing factors of ureteroenteric strictures (UES) in patients undergoing laparoscopic radical cystectomy plus urinary diversion (UD). Method: A total of 412 patients who underwent UD after radical prostatectomy from January 2008 to December 2016 were retrospectively included in this study. Age, gender, body mass index (BMI), diversion type, time to diagnosis of UES, duration of ureteral stent, postoperative complications, including urinary tract infections, ureteroenteric leakage and UES were collected. Kaplan-Meier curves were used to describe time to developing UES. Prognostic factors of UES were analyzed using COX proportional hazard regression model. Result: Median follow-up time was 37 (IQR 17-120) months. A total of 59 patients (70 sides) developed UES, including 34 cases on the left side, 14 cases on the right side and 11 cases on both sides, following UD after radical cystectomy. The median time to diagnosis of UES was 7 (IQR 4-11) months. The total incidence of UES was 14.3%. The incidence of UES was 10.9%, 13.3% and 14.1% at 1, 3 and 5 years after UD, respectively. Cox proportional hazard regression model analysis demonstrated that BMI≥25kg·m(-2) (P=0.008), ureteroenteric leakage (P=0.001) and urinary tract infections (P=0.037) were the independent risk factors associated with UES following UD after radical cystectomy. Conclusion: The incidence rate of UES following UD after radical cystectomy was relatively high, which occurs more common on the left side. Obese patients, combined with ureteroenteric leakage, urinary tract infection after UD, are more likely to develop into UES.

[Surgical technique and outcomes of uteri retrieval from brain-dead multi-organ donors: a preclinical research of human uterine transplantation].

Objective: To evaluate the surgical technique and outcomes of uteri retrieval from brain-dead multi-organ donors.This study is a preclinical research of human living uterine transplantation. Methods: From May, 2015 to May, 2017, four uteri retrieval procedures, characterized with radical hysterectomy and uterine vascular pedicles dissection, were performed in multi-organ brain-dead donors.The uterus was the third authorized organ after the kidney and liver retrieval procedures in the first two cases.The uterine pedicles included the uterus, ovaries, fallopian tubes, the upper one-third of the vagina and internal iliac vessels or external iliac vessels.The perfusion of the uterus was conducted after the retrieval for evaluating the availability, followed by histopathological examination of the uterine issues per 30 minutes. Results: Since the uterine vein was quite difficult to identify and dissect in the first two case, which result in the rupture of triple uterine veins.Therefore, the uterine venous vessels including uterine vein connected with internal iliac vein and internal iliac arteries were selected as vascular grafts and dissected successfully in the last two cases, which could be perfused with the mixture of 4 ℃ heparinized physiological saline through each artery because of shortening the surgical time and arranging the uterine procurement as the first authorized organ procedure.Mean (SD) operative time was 152.5±39.0 min (115-215 min, n=4). Conclusion: Our preliminary experience indicated that the uterus could be retrieved from the brain-dead multi-organ donors and transplanted to the recipient.The attempt of skeletonizing the uterine veins should be replaced by dissection of internal iliac vein.

[Relationship between c-kit mRNA expression and prognosis in patients with rectal carcinoma].

Objective: To investigate the correlation between c-kit mRNA expression and prognosis in patients with rectal carcinoma. Methods: The expression of c-kit mRNA in rectal carcinoma tissues(n=66) was detected by multiplex branched-DNA liquid chip method. According to the expression level, the patients were classified into the c-kit mRNA high expression group and the low group. We analyzed the relationship between the c-kit mRNA expression and the clinicopathological characteristics of patients, as well as the factors affecting patients'prognosis. Results: Of the 66 rectal carcinoma patients, 18(27.3%)cases were c-kit mRNA high expression. No significant correlation was found between the c-kit mRNA expression and gender, age, preoperative carcinoembryonic antigen, preoperative hemoglobin, distance to verge, lymph node metastasis, tumor thrombus, T stage, TNM stage and tumor differentiation (P>0.05). In follow-up, 34 patients died, 32 patients and 36 patients were recurrence or metastasis. The 1-, 3-, 5-year overall survival(OS) of c-kit mRNA high expression group were 100.0%, 77.8%, 77.8%, respectively, while those of the low one were 93.8%, 56.3%, 45.8%, respectively. The difference was statistically significant(P=0.025). Lymph node metastasis, T stage and TNM stage were also significant associated with OS(P<0.05). The 1-, 3-, 5-year disease free rate (DFS)of the c-kit mRNA high expression group were 100.0%, 77.8% and 77.8%, respectively, while those of the low one were 77.1%、43.8% and 41.7%, respectively, and the difference between the two groups was significant (P=0.044). As a reslut, c-kit mRNA expression (P=0.038) and TNM stage (P=0.039) were the independent prognostic factors affecting the OS in rectal cancer patients. Conclusions: Low expression of c-kit was associated with poor prognosis of rectal carcinoma. And the mechanism underlying this phenomenon deserves further exploration.

MiR-613 promotes cell proliferation and invasion in cervical cancer via targeting PTPN9.

OBJECTIVE: To investigate the potential effects of miR-613 on the development of cervical cancer (CC) and the relevant mechanism. PATIENTS AND METHODS: The expression level of miR-613 was detected in CC tissues and cells (siHa) by comparing with corresponding adjacent normal tissues and normal human embryonic kidney cells (293T). Luciferase assay was performed to evaluate the interaction between miR-613 and PTPN9. The effects of the miR-613 on siHa cells were determined by subsequent experiments including cell proliferation, invasion and migration. RESULTS: In our study, miR-613 was found up-regulated in CC tissues and the same result was found at cellular level. The potential target of miR-613 was analyzed by three public databases. We found that tyrosine-protein phosphatase non-receptor type 9 (PTPN9) was a direct target of miR-613, and Luciferase assays confirmed our hypothesis. The subsequent experiments showed that decreased expression of PTPN9 resulting from up-regulation of miR-613 could promote the cell proliferation, invasion and migration of CC cells. CONCLUSIONS: We showed the promotion function of miR-613 on CC by targeting PTPN9 and revealed that miR-613/PTPN9 axis might be a potential therapeutic target for the treatment of CC.

[Effects of urethral fibrosis on urinary control after laparoscopic radical prostatectomy].

Objective: To determine the influence of urethral fibrosis on the recovery of urinary continence after laparoscopic radical prostatectomy. Method: A retrospective study of 203 patients from January 2010 to January 2014 who were underwent laparoscopic radical prostatectomy for prostate cancer in the First Affiliated Hospital of Fujian Medical University. The patients were divided into 2 groups according to preoperative T2-weighted magnetic resonance imaging of fibrosis status of the urethral wall and periurethral tissue. One hundred and forty-four(≤2 grade) and 59 (≥3 grade) were classified into the no/mild and severe urethral fibrosis groups respectively. Urinary continence at 1, 3, 6, 12 months after operation were compared between this two groups respectively. Result: There was no significant difference in the two groups with respect to age, body mass index (BMI), Charlson comorbidity index (CCI), international prostate symptom score (IPSS), prostate volume, preoperative prostate-specific antigen value, nerve-sparing procedure, postoperative Gleason score and pathological stage. The operation was completed successfully in all cases. With a median follow-up time of 15 months (ranged from 12 to 24 months), there was no statistical difference between the two groups in urinary continence at 1 month after operation (P>0.05). The incidences of continence in patients with no/mild fibrosis were significantly higher at 3, 6, 12 months after operation than those with severe fibrosis. (In the no/mild fibrosis group and severe fibrosis group, the continue rate at 3 mouths was 50.0% vs 28.8% P=0.005; at 6 mouths was 91.0% vs 59.3% P<0.001; at 12 mouths was 98.6% vs 88.1% P=0.003). Conclusion: Preoperative urethral fibrosis could be a significant predictor of recovery of the long-term urinary continence status after laparoscopic radical prostatectomy. Compared with no/mild fibrosis, severe fibrosis had worse long-term continence status.

Histidine-tryptophan-ketoglutarate solution decreases mortality and morbidity in high-risk patients with severe pulmonary arterial hypertension associated with complex congenital heart disease: an 11-year experience from a single institution.

Cardioplegic reperfusion during a long term ischemic period interrupts cardiac surgery and also increases cellular edema due to repeated solution administration. We reviewed the clinical experiences on myocardial protection of a single perfusion with histidine-tryptophan-ketoglutarate (HTK) for high-risk patients with severe pulmonary arterial hypertension associated with complex congenital heart disease. This retrospective study included 101 high-risk patients undergoing arterial switch operation between March 2001 and July 2012. We divided the cohort into two groups: HTK group, myocardial protection was carried out with one single perfusion with HTK solution; and St group, myocardial protection with conventional St. Thomas' crystalloid cardioplegic solution. The duration of cardiopulmonary bypass did not differ between the two groups. The mortality, morbidity, ICU stay, post-operative hospitalization time, and number of transfusions in HTK group were lower than those in St group (P<0.05). Univariate and multivariate analysis showed that HTK is a statistically significant independent predictor of decreased early mortality and morbidity (P<0.05). In conclusion, HTK solution seems to be an effective and safe alternative to St. Thomas' solution for cardioplegic reperfusion in high-risk patients with complex congenital heart disease.

Histidine-tryptophan-ketoglutarate solution decreases mortality and morbidity in high-risk patients with severe pulmonary arterial hypertension associated with complex congenital heart disease: an 11-year experience from a single institution

Cardioplegic reperfusion during a long term ischemic period interrupts cardiac surgery and also increases cellular edema due to repeated solution administration. We reviewed the clinical experiences on myocardial protection of a single perfusion with histidine-tryptophan-ketoglutarate (HTK) for high-risk patients with severe pulmonary arterial hypertension associated with complex congenital heart disease. This retrospective study included 101 high-risk patients undergoing arterial switch operation between March 2001 and July 2012. We divided the cohort into two groups: HTK group, myocardial protection was carried out with one single perfusion with HTK solution; and St group, myocardial protection with conventional St. Thomas' crystalloid cardioplegic solution. The duration of cardiopulmonary bypass did not differ between the two groups. The mortality, morbidity, ICU stay, post-operative hospitalization time, and number of transfusions in HTK group were lower than those in St group (P<0.05). Univariate and multivariate analysis showed that HTK is a statistically significant independent predictor of decreased early mortality and morbidity (P<0.05). In conclusion, HTK solution seems to be an effective and safe alternative to St. Thomas' solution for cardioplegic reperfusion in high-risk patients with complex congenital heart disease.

Head mirror versus headlight: illumination, visual identification and visual acuity for otolaryngological examination.

OBJECTIVE: To investigate and compare the performance of head mirrors and headlights during otolaryngological examination. METHODS: The illuminance and illumination field of each device were measured and compared. Visual identification and visual acuity were also measured, in 13 medical students and 10 otolaryngology specialists. RESULTS: The illuminance (mean ± standard deviation) of the LumiView, Kimscope 1 W and Kimscope 3 W headlights and a standard head mirror were 352.3 ± 9, 92.3 ± 4.5, 438 ± 15.7 and 68.3 ± 1.2 lux, respectively. The illumination field of the head mirror (mean ± standard deviation) was 348 ± 29.8 grids, significantly greater than that of the Kimscope 3 W headlight (183 ± 9.2 grids) (p = 0.0017). The student group showed no statistically significant difference between visual identification with the best headlight and the head mirror (score means ± standard deviations: 56.2 ± 9 and 53.3 ± 14.1, respectively; p = 0.3). The expert group scored significantly higher for visual identification with head mirrors versus headlights (59.7 ± 3.3 vs 55.2 ± 5.8, respectively; p = 0.0035), but showed no difference for visual acuity. CONCLUSION: Despite the advantages of headlight illumination, head mirrors provided better, shadow-free illumination. Despite no differences amongst students, head mirrors performed better than headlights in experienced hands.

Clinicopathological characteristics and computed tomography features of mucinous gastric carcinoma.

This study investigated the clinicopathological characteristics of mucinous gastric carcinoma (MGC) and assessed whether multidetector-row computed tomography (MDCT) could differentiate MGC from non-mucinous gastric carcinoma (NGC). Clinicopathological data from 542 patients with gastric carcinoma (23 MGC, 519 NGC), who underwent pre-operative MDCT examination and curative or palliative gastrectomy, were analysed. Only seven of the 23 patients with MGC were correctly diagnosed pre-operatively by endoscopic biopsy. The MGC patients had larger tumours, a higher frequency of lymph node metastases, were more likely to have tumours of tumour, node, metastasis stages III and IV, and were less likely to have a curative resection than NGC patients. In addition, five MGC patients had calcifications in the thickened gastric wall. In conclusion, MGC is rare and is detected mostly at an advanced stage. The diagnostic sensitivity of MGC by endoscopic biopsy was relatively low, whereas MDCT was helpful in distinguishing MGC from NGC.

Size of the largest lymph node visualized on multi-detector-row computed tomography (MDCT) is useful in predicting metastatic lymph node status of gastric cancer.

This study was designed to investigate whether the size of the largest lymph node (long-axis diameter [LAD] and short-axis diameter [SAD]) visualized using multi-detector-row computed tomography (MDCT) was useful for predicting the metastatic lymph node (MLN) status of gastric cancer. A retrospective analysis of 305 gastric cancer patients who underwent pre-operative MDCT was performed, followed by a prospective study in 61 gastric cancer patients to determine the diagnostic effectiveness of LAD and SAD. In the retrospective study, the accuracy of LAD and SAD for predicting the MLN status of gastric cancer was 51.1% and 45.9%, respectively. In the prospective study, the accuracy of LAD and SAD measurement and the traditional MDCT method of counting MLNs was 52.5%, 49.2% and 57.4%, respectively; the differences were not significant. In conclusion, the size of the largest lymph node in terms of LAD and SAD visualized on MDCT was useful for predicting the MLN status of gastric cancer, with accuracy comparable to the traditional MDCT method of counting the total number of MLNs detected.

Bifidobacterium as an oral delivery carrier of oxyntomodulin for obesity therapy: inhibitory effects on food intake and body weight in overweight mice.

INTRODUCTION: Oxyntomodulin (OXM) is a gut hormone released from intestinal L cell. Synthetic OXM and its analog reduce food intake and body weight in both rodents and human beings by being administered intravenously. However, people find intravenous administration difficult because of its side effects and inconvenience. The aim of this study is to develop a novel oral delivery system for OXM and its analog using genetically engineered Bifidobacterium as the carrier. METHODS: An OXM gene expression vector pBBADs-OXM for the Bifidobacterium genus was constructed. Human OXM sequence was fused with extracellular exo-xylanase (XynF) signal peptide (Xs) from Bifidobacterium longum under the control of the pBAD promoter. B. longum NCC2705 was transformed with the recombinant plasmid pBBADs-OXM by electroporation, and the transformed B. longum was selected using MRS plates containing 60 microg ml(-1) ampicillin. The OXM expression in vitro was identified by western blot and enzyme-linked immunosorbent assay (ELISA) assay after L-arabinose induction. Overweight BALB/c mice were treated with B. longum transformed with OXM after 0.2% L-arabinose induction every day for 4 weeks to investigate the effects of OXM-transformed B. longum on food intake and body weight by oral administration. The B. longum transformed with the green fluorescent protein (GFP) gene was used as negative control; orlistat, a gastrointestinal lipase inhibitor, was used as positive control; Normal saline (NS, 0.9% saline) was used as blank control. The food intakes of each group were measured every day, and body weights were measured once a week. Normal BALB/c (2 months old) mice were treated with OXM-transformed B. longum after induction by intragastric administration every day for 6 days to reveal the mechanism of transformed B. longum, with OXM exerting its biological function by oral administration. Plasma OXM, plasma ghrelin and the OXM of intestinal contents were detected by the ELISA method. Plasma glucose and triglyceride levels were analyzed using the Automatic Biochemistry Analyzer. RESULTS: Transformed B. longum with OXM was selected and identified without biological and morphological alteration. An approximately 4-5 kDa OXM peptide was detected in both the supernatant and the cell pellet of transformed B. longum after L-arabinose induction in vitro. The food intake, body weight and blood triglyceride level of overweight mice treated with OXM-transformed B. longum were all significantly reduced compared with that of the GFP negative control group and NS control group (P<0.01). Interestingly, the plasma triglyceride level of the GFP group was significantly decreased compared with that of the NS control group (P<0.01). The OXM level in the intestinal contents of the OXM group was significantly increased compared with that of the GFP negative control group and the NS group (P<0.05). The plasma ghrelin level of the OXM group was significantly decreased compared with that of the GFP and NS groups (P<0.01). Unexpectedly, the ghrelin level of the GFP group was significantly increased compared with that of the NS control group (P<0.01). CONCLUSION: A novel oral delivery system of Bifidobacterium for human OXM has been successfully established. The expression of recombinant OXM can be detected in the supernatant and cell pellet of transformed B. longum. OXM-transformed B. longum reduces food intake, body weight and plasma lipid level in overweight mice by oral administration.

Ecological stabilization of thickened wastewater sludge from CAST process.

Wastewater sludge ecological stabilization (WWSES) pilot scale experiments were conducted for thickening treatment and disposal of sludge which came from Cyclic Activated Sludge Technology (CAST) process. The study was performed over the periods from June to November 2005 and from May to November 2006, on a bed of 80 m2. The sludge loadings were stopped for the winter from December 2005 and resumed in May 2006. The results shows that dried sludge layer has higher permeation coefficients of 0.15-1.3 m/h. It is suggested that the percolate did not filtrate downwards evenly, part of percolate filtrates downwards along stems, roots and cracks existing in dried sludge which have lower flow resistance. The relationship of dried sludge thickness and operation time is in accord with quadratic equation under fluctuating sludge loadings. Linear regression equation can indicate dried sludge thickness variation under fixed sludge loading. In comparison with natural ones, coarse protein content of Phragmites australis roots in the system is twice as high, coarse fiber content of roots, coarse fat content of stems and leaf are obviously higher; and coarse protein content of Typha augustifolia in the system are obviously higher, while coarse fat and coarse fiber contents have no significant difference.

Characterization of resistance gene analogs with a nucleotide binding site isolated from a triploid white poplar.

The majority of cloned plant disease resistance genes (R genes) encode a nucleotide binding site (NBS) and a leucine-rich repeat (LRR) domain. In this study, to better understand the R genes in white poplar, 59 resistance gene analogues (RGAs) were identified from a triploid white poplar [(Populus tomentosa x Populus bolleana) x P. tomentosa], based on conserved NBS regions. The 59 RGAs were phylogenetically classified into 10 subfamilies, and 54 RGAs with open-reading frames (ORFs) were further grouped into two classes, toll and interleukin-1 receptor (TIR) and non-TIR. BLAST searches with reference to the genomic sequence of Populus trichocarpa found 96 highly homologous regions distributed in 37 loci, suggesting the abundance and divergence of NBS-encoding genes in the triploid poplar genome. Within subfamilies 1-3, the average non-synonymous/synonymous substitution (omega) rates were < 1, indicating purifying selection on these RGAs, but some sites were clearly under diversifying selection with omega > 1. Many intergenic exchanges were also detected among these RGAs, indicating a probable role in homogenising NBS domains. Quantitative real-time PCR analysis revealed dramatic variations in the transcript level of 18 RGAs in the mature leaves, bark and roots of the triploid poplar, and identified two RGAs that had significantly higher level of transcripts in bark, four RGAs in mature leaves, and 14 in the above-ground portion of poplars, suggesting their probable roles in resistance against diseases attacking the organs. Our results shed light on genetic resources of poplar resistance and will be useful for further resistance gene isolation and exploitation.

Effect of NF-kappaB constitutive activation on proliferation and apoptosis of gastric cancer cell lines.

OBJECTIVE: To observe whether there is constitutive activation of nuclear transcription factor kappaB (NF-kappaB) and its effect on proliferation and apoptosis of human gastric cancer cell lines. METHODS: Nuclear/cytoplasmic protein expression of NF-kappaB was analyzed by Western blot in four different gastric cancer cell lines. Trans AM(TM) NF-kappaB p65 Kit was used for detecting the difference of p65 activity. The effect of PDTC (pyrrolidine dithiocarbamate), a specific inhibitor of NF-kappaB on the proliferation of gastric cancer cells, was measured by MTT (3-[4,5-dimethythiazol-2-yl]-2,5-diphenyltetrazolium bromide) method. The apoptotic rates of AGS and SGC-7901 gastric cancer cell lines were measured with flow cytometer (FCM) after treatment by PDTC. RESULTS: The constitutive activations of NF-kappaB were identified in four gastric cancer cell lines. The expression of activated subunit of p50 was lower in AGS cell line, and higher in MKN28, MKN45 and SGC-7901 cell lines. The expression of activated subunit of p65 was lower in MKN28 and MKN45 cell lines, and higher in AGS and SGC-7901 cell lines. Both the activity of NF-kappaB and the cell proliferation were significantly inhibited in experimental group treated by PDTC, compared with control groups (p<0.01). An increased apoptotic rate and a decreased proliferating activity were observed after the gastric cancer cells were exposed to PDTC for 24 h. CONCLUSIONS: These results suggested that the constitutive activation and the protein expression of NF-kappaB are different in gastric cancer cell lines. PDTC can inhibit NF-kappaB activity and cell proliferation, which related to an increased cell apoptosis. The results disclosed that NF-kappaB could be a potential therapeutic target for solid tumor therapy.